Untitled - Child and Family Research Institute

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Welcome to the 13th annual CFRI Trainee Research Forum. Each year we
host this event to showcase our trainees’ unique and innovative research.
In partnership with the teaching hospitals on this site, CFRI provides our
trainees with a true ‘bench to bedside’ opportunity for research. We train
undergraduates, medical students, residents, sub-specialty fellows,
masters and doctoral students, and post-doctoral fellows. The diversity of
the research here is remarkable, and the Trainee Research Forum provides
a wonderful preview of the interdisciplinary work our trainees will pursue
as they progress into their own careers as scientific investigators.
We are proud of the trainees here at CFRI, and we are pleased to be able
to help them to become leaders in their respective fields.
Sandra Elias
Manager, Research Education
Child & Family Research Institute
2013 TRAINEE RESEARCH FORUM
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TABLE OF CONTENTS
Session 1
Mikaela Barker............................................................................................................................. 11
Joshua Brown.............................................................................................................................. 12
Samuel Chow.............................................................................................................................. 13
Andrew Fam................................................................................................................................ 14
Lisa Kozicky................................................................................................................................. 15
Jesse Olson................................................................................................................................. 16
David Schuberth.......................................................................................................................... 17
Nancy Vertel................................................................................................................................ 18
Session 2
Rika Aleliunas.............................................................................................................................. 21
James Cairns............................................................................................................................... 22
Stephanie Campbell.................................................................................................................... 23
Kaia Hookenson.......................................................................................................................... 24
Michelle Muller............................................................................................................................ 25
Magdalene Payne........................................................................................................................ 26
Alexis Shih .................................................................................................................................. 27
Session 3
Abeer Aljaadi............................................................................................................................... 31
Dayna-Lynn Dymianiw................................................................................................................. 32
Jason Ken-Shun Hung................................................................................................................. 33
Alexandre Lussier........................................................................................................................ 34
Oksana Nemirovsky..................................................................................................................... 35
Cameron Rogers.......................................................................................................................... 36
Arjun Trivedi................................................................................................................................ 37
Sarah White................................................................................................................................. 38
Grace Goh .................................................................................................................................. 39
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TABLE OF CONTENTS
Session 4
Maria Aristizabal.......................................................................................................................... 43
Helen Chen.................................................................................................................................. 44
Jianjia Fan.................................................................................................................................... 45
Yu-Hsuan Huang.......................................................................................................................... 46
Safia Ladha.................................................................................................................................. 47
Sarah Munro................................................................................................................................ 48
Terri Petkau.................................................................................................................................. 49
Magda Price................................................................................................................................ 50
Dominik Sommerfeld................................................................................................................... 51
Yu Yao ......................................................................................................................................... 52
Session 5
Ana Cohen................................................................................................................................... 55
Parastoo Kheirkhah Dehkordi...................................................................................................... 56
Jennifer Grants............................................................................................................................ 57
Grace Leung................................................................................................................................ 58
Elizabeth Marchant...................................................................................................................... 59
Bo Peng....................................................................................................................................... 60
Veronica Schiariti......................................................................................................................... 61
Kevin Tsai..................................................................................................................................... 62
Joanna Yeung ............................................................................................................................. 63
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TABLE OF CONTENTS
Session 6
Chansonette Badduke................................................................................................................. 67
Rebecca De Souza....................................................................................................................... 68
Jonathan Han.............................................................................................................................. 69
Sara Khosravi............................................................................................................................... 70
Chris Laugen................................................................................................................................ 71
Paul Sabatini................................................................................................................................ 72
Ashish Sharma............................................................................................................................. 73
Sophie Stukas.............................................................................................................................. 74
Bryan Tennant.............................................................................................................................. 75
Allison Watts
Session 7
Guilaine Boyce............................................................................................................................ 79
Dominika Nackiewicz................................................................................................................... 80
Wai Hang Cheng......................................................................................................................... 81
Nicole Krentz............................................................................................................................... 82
Phoebe Lu................................................................................................................................... 83
Ashish Marwaha........................................................................................................................... 84
Cecil Ming Yeung Chau............................................................................................................... 85
Tuan-Anh Nguyen........................................................................................................................ 86
Anna Poon .................................................................................................................................. 87
Session 8
Jiadi Wen..................................................................................................................................... 91
Matthias Görges.......................................................................................................................... 92
Jacqueline Lai.............................................................................................................................. 93
Guobin Sun.................................................................................................................................. 94
Alice Wang.................................................................................................................................. 95
Hong Yang................................................................................................................................... 96
Maryam Zarepour ....................................................................................................................... 97
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TABLE OF CONTENTS
Session 9
Bat-Chen Friedman..................................................................................................................... 101
Anita Coté................................................................................................................................... 102
Judith de Niet............................................................................................................................. 103
Shan-Yu Fung.............................................................................................................................. 104
Klaske van Heusden.................................................................................................................... 105
Whitney Weikum ........................................................................................................................ 106
Session 10
Mercedes Chan........................................................................................................................... 109
Heng Gan.................................................................................................................................... 110
Anas Manouzi.............................................................................................................................. 111
Lana Shaiba................................................................................................................................. 112
Tracy Tucker................................................................................................................................. 113
Yuk Joseph Ting ........................................................................................................................ 114
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POSTER SESSION ONE
masters students
MODERATOR:
Liisa Holsti
PARTICIPANTS:
Mikaela Barker
Joshua Brown
Samuel Chow
Andrew Fam
Lisa Kozicky
Jesse Olson
David Schuberth
Nancy Vertel
MASTERS STUDENTS | POSTER SESSION ONE
Trainee: Mikaela Barker
Board #: 1
Session #: 1
Supervisor: Angela Devlin
Department:Pediatrics
Title: Dietary energy intakes in children with mental health conditions treated with second
generation antipsychotics
Author(s): Mikaela Barker, Angela Devlin, Constadina Panagiotopoulos
ABSTRACT:
Background: Mental Health Conditions (MHC) affect approximately 10-20% of Canadian children each
year, with approximately 15% of children in British Columbia diagnosed with a MHC.
MHCs include: anxiety disorder, attention deficit hyperactivity disorder, depressive
disorder, bipolar disorder, and psychotic disorder. One class of medications being
increasingly used to treat MHCs are second-generation antipsychotics (SGA). Recent
data has shown that SGAs can cause severe and rapid weight gain, increased measures
of adiposity, and cardiometabolic dysfunction in a higher proportion of SGA-treated
compared to SGA-naïve children. The mechanisms responsible for these adverse
side-effects in the pediatric SGA-treated population are not fully understood. Studies in
adults have reported an increased susceptibility to hunger that was two times higher for
those on SGAs compared to a reference group. However, very little is known about the
diets of SGA-treated children. The goal of this study is to determine the role for altered
dietary energy intake on adiposity and cardiometabolic dysfunction in SGA-treated
children compared to SGA-naïve children.
Method: This is a cross-sectional study of male and female children (aged 6-19 years) diagnosed
with MHCs and recruited through BC Children’s Hospital inpatient Psychiatric unit.
Children will be categorized as SGA-treated (n=25) if they have been taking SGAs for
greater than 4 wks and SGA-naïve (n=25) if they are not currently or ever been treated
with SGAs. Demographics, global assessment of functioning (GAF) scores, and
medications (dose and duration) were obtained from medical charts. Three 24-hour
dietary recalls were conducted and analyzed by ESHA food processor software to
calculate dietary energy intakes. Anthropometric measurements, blood pressure, and
fasting plasma lipids, insulin, and glucose were also determined.
Result: To date, 7 SGA-treated and 4 SGA-naïve children have been recruited and assessed.
The mean 3-day total energy intake is 2021.71 kcal/day for the SGA-treated children and
1911.71 kcal/day for the SGA-naïve children. The SGA-treated children have higher BMI
z-scores (mean=1.130) and waist circumference (mean=79.21cm) compared to the
SGA-naïve children (BMI z-score = -0.292; waist circumference=67.27 cm).
Conclusion: These are preliminary findings of an ongoing study. My goal is to have all 50 participants
recruited and analyzed by May 2014.
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CELEBRATING EXCELLENCE
Trainee: Joshua Brown
Board #: 2
Session #: 1
Supervisor: Michael Kobor
Department: Medical Genetics
Title: Function and recruitment of the DNA damage response protein Rtt107 in budding yeast
Author(s): Joshua A Brown, Michael S Kobor
ABSTRACT:
DNA replication and environmental stress lead to DNA damage, and repairing this damage is required
for the survival and function of cells. The genomic stability of budding yeast is maintained by a network
of interacting proteins in multiple DNA damage-repair pathways. The chromatin landscape surrounding
sites of DNA damage controls access of repair factors to DNA, and changes to this environment, such as
histone modifications, occur in response to DNA damage. Histone H2A is phosphorylated in response to
damage, and this leads to assembly of repair proteins and chromatin modifiers.
Yeast strains lacking the protein Rtt107 are sensitive to DNA damaging agents and exhibit a delay in
restarting DNA replication after damage repair. Rtt107 contains six BRCT (BRCA1 C-terminal) domains,
which are often found in proteins that act as scaffolds for the assembly of protein complexes in response
to DNA damage.
Rtt107 is known to be involved in the recruitment of other factors involved in the DNA-damage
response, but the details of the function of Rtt107 have not been fully explored. Rtt107 is recruited to
double-strand breaks (DSBs) in DNA, and this recruitment is dependent upon the damage-induced
phosphorylation of H2A. In strains with a non-phosphorylatable H2A S129A allele the DNA damage
sensitivity of Rtt107 knockout stains is suppressed.
As yeast possess multiple repair pathways, pathways can compensate for deficiencies in other pathways
to maintain genome stability. The suppression of Rtt107 deletion phenotypes seen with H2A S129A may
be the result of activation of alternative repair pathways.
Examining sensitivity to these agents of triple mutants with H2A S129A and deletions of Rtt107 and
genes required for alternative pathways will determine which, if any, of these pathways are responsible
for this suppression.
Other phenotypes of Rtt107 knockouts with H2A S129A have not been examined, and a quantitative
measure of DNA replication and cell cycle progression will be obtained to determine if suppression
occurs.
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Trainee: Samuel Chow
Board #: 3
Session #: 1
Supervisor: Jan Ehses
Department:Surgery
Title: Gp130 receptor signaling as a mediator of alpha cell dysfunction in type 2 diabetes
Author(s): Samuel Chow, Madeleine Speck, Piriya Yoganathan, Pedro Herrera, Werner Muller,
Jan Ehses
ABSTRACT:
Dysregulated alpha-cell glucagon secretion contributes to post-meal hyperglycemia in pre-diabetes and
to hyperglycemia in type 2 diabetes (T2D). Islets in T2D are characterized by chronic inflammation, and
increased IL-6 family cytokine expression. We recently discovered that IL-6 increases glucagon secretion
from islets. However, chronically increased IL-6 may drive alpha-cell reprogramming to generate GLP-1.
Cytokines of the IL-6 family all require the gp130 receptor to signal. Therefore, we were interested in
elucidating the role of the alpha-cell gp130 receptor during T2D. Initially, we profiled IL-6 cytokine
family expression in islets in the type 2 diabetic GK rat. GK rat islets overexpressed Il6, Lif, Clcf1, and
Osm mRNA compared to Wistar controls. To study the role of the gp130 receptor in alpha-cell function,
we generated alpha-cell specific gp130 knock out (agp130KO) mice. Primary alpha-cells from agp130KO
mice showed a 50% reduction in pSTAT3 activation by IL-6 and LIF, in parallel with a 50% reduction in
IL-6 mediated glucagon secretion with no effect on GLP-1 secretion. In vivo, agp130KO mice have
normal glucose tolerance, glucose-mediated glucagon suppression, and circulating glucagon at 16-18
wks of age. To model islet inflammation in a setting of insulin resistance, we subjected our agp130KO
mice to STZ followed by HFD for 6 weeks. This model is characterized by increased body weight,
increased fasting blood glucose, glucose intolerance, impaired glucose-stimulated insulin secretion,
hyperinsulinemia, and hyperglucagonemia. Increased Il27 and Il6 mRNA, and increased IL-6 and LIF
protein secretion characterize islet inflammation post-STZ in this model. When subjected to STZ/HFD,
agp130KO mice had reduced fasting glycemia, improved glucose tolerance, and improved glucosemediated glucagon suppression. Fasting insulin was decreased likely due to reduced glycemia in
agp130KO mice, but no differences in glucose-stimulated insulin secretion or insulin sensitivity were
detected. Finally, no differences in pancreatic hormone levels of insulin, glucagon, or GLP-1 were
observed between genotypes in these mice. Our data strongly suggest that reduced glycemia and
improved glucose tolerance in our agp130KO mice is due to improved alpha-cell function. Thus, further
work is needed to understand the mechanisms of gp130 signaling and modification of signaling via
gp130 may be useful for the treatment of T2D.
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Trainee: Andrew Fam
Board #: 4
Session #: 1
Supervisor: Cornelius Boerkoel
Title: Mitochondrial localization of Tdp1 protects the mitochondrial genome from oxidative
stress
Author(s): Andrew Fam, Lauren Fougner, Miraj Chowdhury, Cheryl Walton, Kunho Choi,
Cornelius F Boerkoel
ABSTRACT:
Tyrosyl-DNA phosphodiesterase I (Tdp1) participates in the removal of topoisomerase I-DNA
intermediates by cleavage of the 3’ phosphotyrosine bond. In studying rhabdomyosarcoma (RMS) cell
lines, we have found that overexpression of Tdp1 makes cells resistant to the anticancer drug
camptothecin (CPT), a drug that interferes with DNA repair by stabilizing the topoisomerase I-DNA
intermediate. Although classically considered a nuclear DNA repair enzyme, RMS cells expressed Tdp1
in the mitochondria and had a CPT sensitivity inversely proportionate to Tdp1 expression in the
mitochondria. Within human skeletal muscle, Tdp1 was exclusively expressed in intermyofibrillar
mitochondria but not in subsarcolemnal mitochondria or the nucleus. In contrast, cultured human dermal
fibroblasts and mouse embryonic fibroblasts express Tdp1 exclusively in the nucleus until challenged
with oxidizing agents such as H2O2 and menadione sodium bisulfite. Following such a challenge Tdp1
relocates from the nucleus to the cytoplasm and mitochondria. Affirming a role for Tdp1 in mitochondrial
DNA repair, tissues devoid of functional Tdp1 accumulate, mitochondrial DNA lesions under prolonged
exposure to oxidative stress. In summary, we hypothesize that Tdp1 translocates between the nucleus
and the mitochondria to facilitate DNA repair and that it processes oxidatively damaged mitochondrial
DNA.
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Trainee: Lisa Kozicky
Board #: 5
Session #: 1
Supervisor: Laura Sly
Title: Regulatory macrophages for the treatment of Inflammatory Bowel Disease
Author(s): Lisa Kozicky, Bonnie Cheung, Laura Sly
ABSTRACT:
Inflammatory Bowel Disease (IBD) is a chronic inflammatory disease characterized by inflammation along
the intestinal tract. Current treatment for IBD relies on non-specific immune suppression but up to 40%
of people are, or will become, refractory to current therapies. Macrophages are key mediators of
inflammation initiating the innate immune response. However, macrophages can be skewed to a
regulatory phenotype (Mregs) that plays an equally important role in turning off the inflammatory
response.
My overarching hypothesis is that Mregs can be used to reduce intestinal inflammation, like that which
characterizes IBD. To address this hypothesis, I propose three specific aims:
Aim 1. To compare the anti-inflammatory properties of Mregs primed with serum or IgG
Aim 2. To determine whether Mregs can reduce intestinal inflammation in vivo.
Method: Macrophages were derived from mouse bone marrow aspirates. Macrophages were
primed to create an Mreg phenotype with either normal mouse serum or high dose IgG.
Inflammatory responses of Mregs were measured in vitro and the potential of Mregs
to block inflammatory responses was assessed in vivo using the dextran sodium sulfate
(DSS)-induced mouse model of intestinal inflammation.
Result: Mregs stimulated with high doses of IgG have low pro-inflammatory IL-12/23p40
production and produce high levels of anti-inflammatory IL-10 in response to
inflammatory stimuli. Serum-induced Mregs more potently inhibit LPS-induced
IL-12/23p40 production and also produce high levels of IL-10. Importantly, adoptive
transfer of in vitro-derived Mregs reduces inflammation and clinical disease activity in
mice during DSS-induced colitis.
Conclusion: These studies demonstrate that Mregs have potent anti-inflammatory activity that can be
used to reduce intestinal inflammation in vivo. Skewing macrophages to an Mreg
phenotype in vivo or adoptive transfer of in vitro-derived Mregs may provide a novel
immunotherapeutic strategy to treat intestinal inflammation. In future studies, we will
investigate the mechanism(s) by which macrophages change their phenotype to become
Mregs to determine whether macrophages can be skewed in situ to dampen down
inflammation in people with IBD.
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Trainee: Jesse Olson
Board #: 6
Session #: 1
Department: Pathology & Laboratory Medicine
Title: Programming of glucose homeostasis in female mice by developmental exposure to folic acid and vitamin B12 imbalance
Author(s): Jesse Olson, Rika Aleiunas, Melissa Glier, Abeer Aljaadi, Tim Green, Angela Devlin
ABSTRACT:
Developmental exposure to imbalanced maternal nutrient status may program risk for type 2
diabetes. This phenomenon of developmental programming may be mediated by epigenetic
mechanisms. Population studies have reported that children from mothers with high folate and low
B12 status during pregnancy have greater adiposity and insulin resistance, however the mechanisms
behind these observations remain unknown. The objective of this study is to investigate the molecular
mechanisms underlying programming of adiposity and glucose homeostasis by maternal folate and B12
imbalance. Female C57BL/6J mice were fed a high folic acid (FA) with B12 (HFA+B12), high FA without
B12 (HFA-B12), or a control diet during pregnancy and lactation. At weaning, female offspring were fed
a western or control diet until 30 weeks of age. Offspring mice fed the post weaning western diet had
greater body weight (P<0.001), visceral (P<0.001) and subcutaneous (P<0.001) fat, and glucose
intolerance (P<0.05) than offspring fed the post weaning control diet. Offspring from female mice fed
the HFA-B12 diet had greater subcutaneous fat (P<0.05) than offspring from female mice fed the control
or HFA+B12 diets. This was accompanied by lower (P<0.05) pancreatic islet MafA mRNA, however this
was only observed in offspring fed the post weaning control diet. Offspring mice fed the post weaning
western diet that were from females fed the HFA-B12 diet had lower (P<0.05) fasting glucose and insulin
concentrations compared to offspring from females fed the control diet. These findings provide evidence
of programming of glucose homeostasis by developmental exposure to FA and B12 imbalance.
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Trainee: David Schuberth
Board #: 7
Session #: 1
Supervisor: Robert McMahon
Department:Psychology
Title: Is there a reciprocal relationship between callous-unemotional traits and emotion
recognition in preschool?
Author(s): David A Schuberth, Dave S Pasalich, Robert J McMahon, Joseph J Palamar,
Esther J Calzada, Laurie Miller Brotman
ABSTRACT:
Among risk factors for the development of severe behavioral problems across the lifespan, callousunemotional (CU) traits, characterized by a lack of empathy, have been identified as particularly
significant. Further, developmental research posits the accurate recognition of distress emotions in
others as paramount in the successful establishment of empathy. Some research has suggested that CU
traits and deficits in negative emotion recognition may share common neurological underpinnings, yet
this relationship has yet to be explored in early childhood. The current study investigated the
potentially reciprocal relationship between CU traits and the recognition of fear and sadness across
preschool and kindergarten. Participants include 489 preschoolers from the control group of a larger
randomized controlled trial, with three data collection points spanning an 18-month period. Preschool
and kindergarten teachers were surveyed to obtain children’s reported levels of CU traits and aggression,
while children were assessed directly to obtain verbal ability and emotion recognition skills. Emotion
recognition assessment included emotion knowledge and identification across contexts such as matching
games, spoken vignettes, and puppet shows. It was hypothesized that CU traits and the recognition of
fear and sadness would negatively predict one another at the subsequent time point after controlling for
aggression and verbal ability. Analyses included correlations within and mixed-model linear regressions
across time points. Results partially supported the initial hypotheses: Higher reported levels of CU traits
at the beginning of preschool predicted less accurate recognition of sadness at the end of preschool,
whereas less accurate recognition of fear at the end of preschool predicted higher reported levels of CU
traits at the end of kindergarten. These findings suggest that addressing deficits in the recognition of
distress cues may be important in deterring the further development of CU traits in children who show
these features in early childhood.
17
Trainee: Nancy Vertel
Board #: 8
Session #: 1
Supervisor: Karen Campbell
Department: Oral Health Science
Title: Barriers to access to dental services for children with special needs
Author(s): Nancy Vertel, Karen Campbell, Rosamund Harrison
ABSTRACT:
Objectives of the pilot project:
1. Identify challenges and enabling factors encountered by caregivers in obtaining dental care for their
Children with Special Health Care Needs (CSHCN);
2. Determine the level of understanding and awareness of the oral status of CSHN by their caregivers
Method: Ethical approval from UBC C&W BREB was granted in October 2011. A sample of
CSHCN caregivers (n=51) were interviewed either in person, telephone or electronically
using a pre-tested interview instrument. Closed-ended questions included race/ethnicity,
language spoken at home, caregiver education level, family structure and income range.
Open-ended questions were used to report caregiver’s perception of their child’s oral
health status and enabling factors/barriers when accessing dental services. Demographic
information, medical diagnosis, oral health status, chief dental complaint, referral date/
source, dental benefit type, anticipated dental treatment summary were all extracted
from their child’s dental record. Quantitative data was analyzed using univariate
statistical tests. Qualitative data will be understood by thematic analysis.
Result: Commonly reported medical diagnoses for sample CSHCN included developmental
delay, seizure disorder, autism and cerebral palsy. Of the sample, 50% were referred to
DD-BCCH by a medical specialist and 60% did not have a regular dental provider. About
50% of caregivers reported experiencing refusal of dental care. Caregivers in upper
income levels represented 33% of the sample. CSHCN’s dental services were funded
through private dental insurance (41%), government-based funding (41%), and
out-of-pocket funds (26%).
Conclusion: The present sample of caregivers reported an awareness of their child’s unmet dental
needs, yet consulted a dental provider only on an emergency or as-needed basis. Also,
due to medical complexity and/or behavioral limitations of their child’s condition,
caregivers perceived the importance of dental care as secondary to that of medical care.
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POSTER SESSION TWO
masters students
MODERATOR:
PARTICIPANTS:
Mariana Brussoni
Rika Aleliunas
James Cairns
Stephanie Campbell
Kaia Hookenson
Michelle Muller
Magdalene Payne
Alexis Shih
MASTERS STUDENTS | POSTER SESSION TWO
Trainee: Rika Aleliunas
Board #: 9
Session #: 2
Title: Maternal folic acid and vitamin B12 intake programs offspring body composition and
fasting insulin levels
Author(s): Rika Aleliunas, Jesse Olson, Melissa Glier, Abeer Aljaadi, Tim Green, Angela Devlin
ABSTRACT:
Gestational nutrient status may affect cardiometabolic risk in adulthood. Since the implementation of
folic acid fortification in Canada, circulating levels of unmetabolized folic acid are higher in the
population and the metabolic consequences are unknown. Folic acid and vitamin B12 are essential for
the generation of methyl groups for methylation of cellular components via the methionine cycle. Excess
folic acid intake creates an imbalance in these vitamins if vitamin B12 status is marginal and affects the
methylation capacity of the methionine cycle. A 2008 study from India reported that children born to
women with high folate and low vitamin B12 status had greater adiposity and insulin resistance at age 6.
The majority of Canadians have high folate status but approximately 5% are vitamin B12 deficient and
as many as 1 in 20 women may be vitamin B12 deficient in the early stages of pregnancy. Therefore, the
goal of our study was to investigate the effects of gestational exposure to high folic acid, with or without
vitamin B12 supplementation, on measures of metabolic health in C57BL/6J mice. We fed female
mice diets high in folic acid, with (HFA+B12) and without (HFA-B12) adequate vitamin B12, before
conception, throughout pregnancy, and during lactation. At weaning, offspring mice were fed a western
diet (45% fat and 35% carbohydrate) or a control diet for 20 weeks. At 20 weeks, western-fed male mice
had greater body weight, larger fat pads, and decreased glucose tolerance (P<0.01) relative to
control-fed mice. Maternal folic acid and vitamin B12 intake affected overall weight gain (P<0.05) and
the size of the subcutaneous fat depot (P<0.01) in all mice. We observed an interaction between
maternal diet and weaning diet on retroperitoneal fat pad size. Western-fed mice exposed in utero to
high folic acid (with and without vitamin B12 supplementation) had smaller retroperitoneal fat depots
than maternal controls (P<0.05). At 30 weeks control-fed offspring with in utero exposure to HFA-B12
diet had higher fasting insulin levels than maternal control and HFA+B12 groups (P<0.05). Our work
suggests that maternal folic acid and vitamin B12 intake may program insulin homeostasis and adiposity
in offspring.
21
Trainee: James Cairns
Board #: 10
Session #: 2
Supervisor: Daniel Goldowitz
Title: Systems neurobiology of autism using mouse models
Author(s): James Cairns, Daniel Goldowitz
ABSTRACT:
Autism and Autism Spectrum Disorders (ASD) are a heterogeneous group of disorders, with the
severity of symptoms varying widely between individuals. Autism can be characterized by symptoms
such as impaired and disordered language, decreased social interactions, extreme fixation on certain
objects or activities and impaired fine and gross motor movements. There is an increasing body of
evidence that suggests that structural and functional abnormalities in the cerebellum and its connections
to other brain regions are a key component in the pathogenesis of autism. Examples of abnormalities
include: decreased numbers of neurons in the cerebellum, altered migration patterns of neurons, and
abnormal synaptogenesis between neurons. There are connections between the cerebellar nuclei and
the prefrontal and parietal cortices, which suggests that the cerebellum plays an important role in
modulating a wider variety of neural activity than previously thought. Since the cerebellum can
influence motor, sensory and cognitive neural activities and because it has a long developmental
maturation period like the prefrontal cortex, altered synaptic connections between the cerebellum and
cortex could be a contributing factor in the pathogenesis of autism.
To investigate this question further, we explored differences in neural activity between wildtype mice
(C57BL6/J mice) and autistic-like phenotype mice using two mutant mouse models including the FMR1
knockout mouse and the Lurcher mutant mouse. To induce action potential firing in different brain
regions animals underwent a series of behavioural tests including rotarod activation. Following behavioural testing, cFos positive cells were used as markers of neuronal activity and we evaluated differences
in neuronal activation in a subset of brain regions that have previously been implicated in the pathogenesis of autism and ASD. Preliminary data has shown increased cFos activation in the cerebellum of wildtype animals as compared to Lurcher mutants and increased cFos activation in the somato-motor cortex
of FMR1 wildtype animals as compared to FMR1 knockout animals.
22
Trainee: Stephanie Campbell
Board #: 11
Session #: 2
Supervisor: Brad Hoffman
Department:Surgery
Title: The role of chromatin-modifying proteins in reprogramming of hepatocytes into
pancreatic β-cells
Author(s): Stephanie Campbell, Cheryl Whiting, Brad Hoffman
ABSTRACT:
Islet transplantation is an effective treatment for type 1 diabetes; however, this procedure is limited by
organ donor numbers, and graft failure inevitably occurs. A search for alternative sources of β-cells has
led to the development of reprogramming protocols that attempt to generate “β-like-cells” from
alternative cell types. Although, in vivo such protocols have shown promise, in vitro protocols have had
limited success. Our preliminary data indicates that this may be due to the inability of these protocols to
overcome specific epigenetic barriers. For example, in hepatocytes, which like the pancreas are derived
from the endoderm, cis-regulatory regions critical for β-cell gene expression often either lack activating histone modifications (H3K4me1), or are enriched for repressive marks (H3K27me3). I will determine
whether the manipulation of chromatin-modifying proteins can be used to help overcome these
epigenetic barriers, and improve the reprograming of mouse hepatocytes into “β-like-cells”.
The β-cell phenotype will be induced in vitro with a tdTomato lentivirus expressing the ?-cell-specific
transcription factors: Pdx1, MafA, and Ngn3. We will isolate hepatocytes from adult mice expressing GFP
under control of the insulin promoter. Cells will be co-transduced with a CFP lentivirus overexpressing or
knocking down chromatin-modifying proteins of interest. These include components of the SWI/SNF
nucleosome remodeling complex (Brg1); proteins involved in H3K4 methyltransferase activity (Setd7,
Wdr5); H3K4 demethylases (Jmjd3, Utx); and components of H3K27 methyltransferase complexes (Ring1
and Eed). The reprogramming efficiency will be determined by monitoring the number of triple-positive
(GFP/tdTomato/CFP) cells relative to the number of double-positive (tdTomato/CFP) cells by
fluorescence microscopy. We will examine reprogramming efficacy of the best candidate factors by
immunofluorescent staining for β-cell-specific proteins and other islet cell hormones. Generated “β-likecells” will be selected for further functional tests, including glucose-responsive insulin-secretion, and
their ability to reverse diabetes in mouse models. These studies will interrogate the role of chromatin
structure as a barrier to cellular reprogramming, and in parallel, shed light on the role of chromatinmodifying proteins in β-cell development, and may help us move closer to making cell replacement
therapy a widely applicable treatment for diabetes.
23
Trainee: Kaia Hookenson
Board #: 13
Session #: 2
Supervisor: Tim Oberlander & Angela Devlin
Title: MTHFR C677T genotype and HPA stress regulation at 6 years of age
Author(s): Kaia Hookenson, Joanne Weinberg, Ursula Brain, Angela M Devlin, Tim F Oberlander
ABSTRACT:
Background:
Major depression is often characterized by elevated daily cortisol concentrations
reflecting altered hypothalamic pituitary adrenal (HPA) regulation. Maternal mood and
serotonin reuptake inhibitor (SRI) antidepressants used during pregnancy have been
recognized as key early life experiences that affect developing HPA stress systems.
Thus, prenatal exposure to a dysregulated maternal HPA axis may exert early
programming effects on fetal HPA activity. The methylenetetrahydrofolate reductase
(MTHFR) gene, an enzyme required for folate metabolism, has a variant (C677T)
associated with depression. It remains unknown whether the C677T variant moderates
the effect of prenatal exposure to maternal depression and SRI antidepressants in
shaping HPA function during early childhood.
Method: To study if an association between antenatal maternal mood and offspring HPA
regulation may be moderated by MTHFR C677T genotype, cortisol was quantified in
salivary samples obtained from children (n=55) over four days [early morning awakening
(mean 7:42 ± 0.91 h), 15 minutes later (8:02 ± 0.96 h), and evening (18:40 ± 2.35 h)].
Result: Using a linear regression model, prenatal SRI antidepressant exposure and maternal
mood (Hamilton Depression Rating Scale) at 33 weeks and 6 years, did not predict cortisol
concentrations. However, children with 677TT genotype (n=9) had higher evening cortisol
concentrations (?=-0.384, P=0.004), and a smaller change from early morning to evening
(?=0.292, P=0.037), compared to children with 677CC and CT genotype.
Conclusion: These data suggest that children with a MTHFR 677TT genotype have both elevated and
flattened diurnal cortisol patterns, possibly indicating HPA axis dysregulation. The outcomes of this work offer insight into the links between one-carbon metabolism and risk for
stress-related mood disorders.
24
Trainee: Michelle Muller
Board #: 14
Session #: 2
Supervisor: Blair Leavitt
Title: The role of iron in Huntington’s Disease
Author(s): Michelle Muller, Ge Lu, Blair Leavitt
ABSTRACT:
It has been shown with imaging and post-mortem studies that specific brain areas of neurodegeneration
also accumulate iron. In Huntington’s Disease, it is thought that iron accumulates in the caudate/putamen which is the specific region of neurodegeneration. I am looking at the iron content in the aged HD
mouse model, YAC128, as well as post-mortem analysis via atomic absorption spectroscopy.
25
Trainee: Magdalene Payne
Board #: 15
Session #: 2
Supervisor: Rajavel Elango
Title: Quantification of lysine requirements in healthy pregnant women using stable isotope
tracer techniques
Author(s): Magdalene Payne, Trina Stephens, Ronald O Ball, Paul B Pencharz, Rajavel Elango
ABSTRACT:
Background:
Current protein and amino acid recommendations during pregnancy are extrapolated
from nonpregnant adult data by the Dietary Reference Intakes (DRI). Additionally, these
recommendations remain constant throughout pregnancy and do not take into account
the metabolic adaptations that occur during different stages of pregnancy. This study
aims to define a quantitative requirement for lysine (an essential amino acid) during early
and late stages of gestation.
Method: Two phases of healthy pregnancy (12-19 weeks and 33-38 weeks) were studied using the
stable isotope based indicator amino acid oxidation (IAAO) technique. 20 women (age
range 22-36y) were given a test diet containing a random lysine intake (range = 6 - 88
mg/kg/d) along with a crystalline amino acid mixture. Diets were provided as eight isonitrogenous meals with caloric intake maintained at 1.7 x resting energy expenditure and
protein maintained at 1.5g/kg/d. Baseline and isotopic steady state breath samples were
taken. Lysine requirements were determined by measuring oxidation of L-1- 13C-Phenylalanine to 13CO2 (F13CO2). Linear regression crossover analysis was used to determine
a breakpoint (indicating a mean lysine requirement) in F13CO2 data.
Result: Preliminary results for late gestation (n=35 completed of an expected 40) indicate a mean
lysine requirement of ~53 mg/kg/d, which is higher than current recommendations made
by the DRI of 41 mg/kg/d. Preliminary results for early gestation could not be made (n=17
completed of an expected 40).
Conclusion: These data are the first for lysine requirements during healthy human pregnancy. Whether
lysine requirements differ during different stages of gestation remains to be determined.
26
Trainee: Alexis Shih
Board #: 16
Session #: 2
Supervisor: Dan Luciani
Department:Surgery
Title: Additional roles of the pro-survival factor Bcl-xL in beta-cell mitochondrial physiology
and oxidative stress
Author(s): Alexis Shih, Lerly Luo, Dan S Luciani
ABSTRACT:
A diet high in glucose and fat increases metabolic load and promotes oxidative stress in pancreatic
beta-cells. Chronic stress leads to beta-cell dysfunction and death, resulting in the loss of glucose
tolerance and eventually diabetes. Proteins in the B-cell lymphoma 2 (Bcl-2) family control the execution
of cell death by apoptosis. Moreover, we recently demonstrated that anti-apoptotic Bcl-2 and Bcl-xL
suppress glucose signalling in beta-cells, and studies on non-beta-cells suggest Bcl-xL regulate
mitochondrial morphology. We hypothesize that Bcl-xL protects beta-cells from nutrient-induced
oxidative stress by regulating beta-cell mitochondrial morphology and metabolic flux.
To test our hypothesis we use complementary approaches of acute pharmacological inhibition,
conditional Bcl-x gene deletion and transient Bcl-xL overexpression. We found that prolonged treatment
with a small molecule Bcl-2/Bcl-xL antagonist triggered MIN6 beta-cell death that can be prevented by
addition of the antioxidant N-acetyl-L-cysteine (NAC). We examined the effect of chronic glucolipotoxicity on the death of pancreatic islet cells from beta-cell-specific Bcl-xL knockout (KO) and littermate wild
type (WT) mice. After 48 hours culture of high glucose and free fatty acids, Bcl-xL WT and KO islet cells
show significantly increased cell death. Surprisingly, the glucolipotoxicity-induced death of both Bcl-xL
WT and KO islet cells was potentiated by addition of NAC, suggesting pro-survival roles for reactive
oxygen species (ROS). Next we examined putative role of Bcl-xL in beta-cell mitochondrial morphology by transiently overexpressing YFP-tagged Bcl-xL in MIN6 cells. Using fluorescence microscopy we
observed that Bcl-xL:YFP locates exclusively to the mitochondria and promotes marked mitochondrial
aggregation. Preliminary experiments suggest the observed mitochondrial hyper-fusion does not
signifacntly affect glucose-induced cytosolic calcium responses.
Our results demonstrate that acute Bcl-2/Bcl-xL inhibition triggers ROS-dependent beta-cell death,
suggesting Bcl proteins may constitutively suppress beta-cell ROS formation. Our preliminary data
further implicate Bcl-xL may control beta-cell mitochondrial fusion-fission; processes suggested to
interrelate with ROS signalling and affect beta-cell susceptibility to nutrient-induced stress. These
findings and data from other labs, suggest that Bcl-xL acts as a point of molecular integration between
beta-cell function, stress signalling and survival. Clarifying these roles of Bcl-xL will improve our
understanding of beta-cell physiology in the pathogenesis of type 2 diabetes.
27
POSTER SESSION THREE
masters students
MODERATOR:
PARTICIPANTS:
Rajavel Elango
Abeer Aljaadi
Dayna-Lynn Dymianiw
Jason Ken-Shun Hung
Alexandre Lussier
Oksana Nemirovsky
Cameron Rogers
Arjun Trivedi
Sarah White
Grace Goh
MASTERS STUDENTS | POSTER SESSION THREE
Trainee: Abeer Aljaadi
Board #: 17
Session #: 3
Title: Programming of liver gene expression by maternal exposure to folic acid/vitamin B12
imbalance
Author(s): Abeer M Aljaadi, Rika E Aleliunas, Jesse D Olson, Dian C Sulistyoningrum,
Timothy J Green, Angela M Devlin
ABSTRACT:
Background:
Developmental programming suggests that changes in environment during development can program metabolic response later in life and increase risk for chronic disease.
Food fortification with folic acid has increased folate intakes of Canadians, however there
is concern regarding vitamin B12 status. Around 5% of Canadian women are vitamin B12
deficient. This is concerning because an association between gestational exposure to
high maternal folate and low vitamin B12 status with greater adiposity and insulin
resistance in children has been reported. We examined the effect of gestational
exposure to maternal folic acid/vitamin B12 imbalance on programming of liver gene
expression.
Method: Female C57BL/6 mice were fed a high folic acid/adequate vitamin B12 (HFA+B12),
high folic acid/no vitamin B12 (HFA-B12), or control (adequate folic acid/vitamin B12)
diet 6 weeks prior to mating and through pregnancy and lactation (n=6-8 mice/group).
At weaning, the offspring mice were randomly assigned to receive the control diet or
a Western diet (45% fat, 35% carbohydrate) for 20 weeks (n=5-6 mice/group). Serum
vitamin B12 concentrations were quantified by a microbiological assay. We quantified
mRNA expression by real-time PCR of key enzymes in methyl nutrient metabolism in liver
from adult male offspring: Mtr (encodes methionine synthase), Mat1a (encodes methionine adenosyltransferase), Cbs (encodes cystathionine ?-synthase), and Mthfr (encodes
methylenetetrahydrofolate). As well, we quantified gene expression of Nr3c1 and
Ppara as they are regulated by DNA methylation and involved in carbohydrate and fat
metabolism.
Result: Dams fed the HFA-B12 diet had lower serum B12 concentrations (P<0.01) compared
to the control-fed dams after 12 weeks on diet. Offspring mice dams fed the HFA-B12
diet had lower (P<0.05) Cbs and Mthfr mRNA expression and this was unaffected by
postweaning diet. Offspring mice fed the Western diet had higher Mtr mRNA expression
(P<0.05) compared to control-fed offspring mice, regardless of maternal diet. No effects
of maternal and postweaning diets were observed for Mat1a, Nr3c1, and Ppara
expression.
Conclusion: Our findings suggest that gestational exposure to maternal folic acid/vitamin B12
imbalance programs Cbs and Mthfr mRNA expression in liver of adult male offspring.
31
Trainee: Dayna-Lynn Dymianiw
Board #: 18
Session #: 3
Supervisor: Jehannine Austin
Title: The impact of prenatal screening on maternal mood: A longitudinal study
Author(s): Dayna-Lynn Dymianiw, Catriona Hippman, Angela Inglis, Jehannine Austin
ABSTRACT:
Studies have shown that shortly after receiving results that indicate a pregnancy is at increased risk for
fetal aneuploidy, women often experience symptoms of depression/anxiety. However, it remains unclear
as to whether these symptoms persist. In our ongoing study, our goal is to compare the trajectories of
depression symptoms through pregnancy and the postpartum between two groups: those who receive
‘abnormal’ results that indicate an increased risk for fetal aneuploidy, and those who receive ‘normal’
results.
From enrollment, women complete the Edinburgh Postnatal Depression Scale (EPDS) at 4-week intervals
until 3 months postpartum.
Of the first 111 participants (goal n=200), 59 have ‘normal’ results, and 52 have ‘abnormal’ results. As
compared to those with ‘normal’ results, women with ‘abnormal’ results: a) have higher EPDS scores
(U=932.5, Z= -3.564, p <0.0001) and b) are more likely to score above cut-off for depression on the
EPDS (?2(1, 111) = 6.631, p =0.01).
Women who receive ‘abnormal’ results are more likely to experience depression during their pregnancy.
This information provides a rationale for healthcare providers to monitor the mood of women who
receive ‘abnormal’ results. Referring women who are depressed for appropriate assessment is important
to avoid negative consequences for maternal/infant health.
32
Trainee: Jason Ken-Shun Hung
Board #: 19
Session #: 3
Supervisor: Rusung Tan
Title: Novel liposome therapy prevents type I diabetes in non-obese diabetic mice
Author(s): Jason Ken-Shun Hung, I-Fang Lee, Nicole Leung, Omar Duramad, Rusung Tan
ABSTRACT:
Type I diabetes (T1D) is an autoimmune disease that affects roughly 300,000 Canadians with the
economic burden to Canada reaching $1.2 billion; both are steadily rising. T1D is caused by a selective
breakdown of immunological tolerance towards the insulin-secreting β cells in the pancreas, leading to
uncontrolled regulation of blood glucose levels, which is fatal if left untreated. It is believed that T1D is
mainly caused by a dysregulation of immune cells such as natural killer T (NKT) cell, regulatory T cells
(Treg) and dendritic cells (DC), which are normally involved in maintaining tolerance to self-antigens and
modulating immune responses. In order to restore regulatory function to these cells we have developed
a novel liposome that incorporates NKT agonist alpha-galactosylceramide (?GalCer) and insulin, which is
thought to be a key antigen involved in T1D and potent activator of Tregs. We hypothesize that simultaneous activation of DC, Treg and NKT cells through antigen presenting cells using this novel agent
prevents T1D in NOD mice. We show that the liposomal therapy prevents T1D in NOD mice compared
to untreated mice (p=0.044) but that route of injection was paramount to the efficacy of the therapy; IV
injections provided no protection, IP injections only delayed T1D, while SQ injections fully protected
mice from T1D. However, all three routes of injections showed significantly lower levels of IGRP-specific
T cells during early time points. Furthermore, all three routes show significantly higher levels of DCs in
the spleen, a key organ in immune regulation. We therefore conclude that dependent on the route of
injection, liposomal therapy containing ?GalCer and insulin can prevent the development of T1D in NOD
mice. We believe that the liposome as well as the route of injection alters the pharmokinetics and pharmodynamics of the therapy, possibly by either increasing the efficacy time, stability or by creating drug
reservoirs in different organs. Further studies are currently being done to determine the exact mechanisms, distribution and cytotoxicity of the liposomal therapy. We believe that this type of combinational
liposomal therapy offers a new direction of T1D and other autoimmune disease treatment in the future.
33
Trainee: Alexandre Lussier
Board #: 20
Session #: 3
Title: Acute ethanol exposure alters the histone modification profile in the mouse brain
Author(s): Alexandre Lussier, Kristin Hamre, Daniel Goldowitz, Michael S Kobor
ABSTRACT:
Exposure to alcohol during the early stages of life disrupts a number of key molecular pathways, often
resulting in abnormal brain development. While the exact mechanisms through which ethanol acts are
still unknown, alterations in epigenetic programming may mediate its effects. Subtle changes in the
pattern of repressive or active histone marks could lead to lasting changes in cell and brain function.
Here, we provide evidence for global changes in histone acetylation and methylation in the brain
following acute ethanol exposure by assessing differences in the levels modifications on histones 3
and 4.
C57BL/6J mice were exposed to alcohol on postnatal day 7 via 2 subcutaneous injections of 2.5 g/kg
of ethanol 2 hours apart with controls given isovolumetric saline. Brain tissue (cerebellum, hippocampus and cerebral cortex) was collected and flash frozen 7 hours after the initial injection. Histones were
extracted from the tissue and the changes in various chromatin marks were assayed by western blot
analysis.
The observed shifts in histone modifications suggest changes in transcriptional activity. The ethanol treated samples showed altered levels of H3K4 and H3K27 trimethylation, as well as H4 acetylation. These
differences were region specific and were also marked by differences between male and female mice.
These results provide evidence that changes in histone acetylation and methylation may be an important
means by which ethanol alters gene expression. They also offer a potential therapeutic avenue for
treating and preventing the defects caused by developmental exposure to alcohol.
34
Trainee: Oksana Nemirovsky
Board #: 21
Session #: 3
Supervisor: Christopher Maxwell
Title: The role of BRCA1 in mammary epithelial polarization through the study of mitotic
spindle positioning in health and disease
Author(s): Oksana Nemirovsky, Nagarajan Kannan, Connie Eaves, Chris Maxwell
ABSTRACT:
Women who carry germline mutations in BRCA1 (Breast cancer 1) have an elevated risk of developing
proliferative and difficult to treat tumours, that do not express hormone receptors and resemble
mammary basal epithelia by gene expression profiling. This resemblance to breast stem/progenitor
cells suggests that loss of BRCA1 function disrupts stem cell homeostasis. We have shown that germline
variation in a mitotic spindle assembly factor modifies the risk of developing breast cancer in carriers of
BRCA1 mutations. Directional mitotic spindle assembly manages the balance between self-renewal and
differentiation during stem cell division, through the asymmetric segregation of transcription
factors. We hypothesize that BRCA1 is necessary for directional cell division in primitive cells and
disruption of its function is sufficient to destroy the balance, favoring proliferation. Our objective is to
characterize spindle positioning as a targetable mechanism to control primitive cell turnover.
Design:
We established the matrigel culture system (3D) to measure the spindle orientation
relative to apical surface of spheroid acini, formed from single mammary cell in matrigel,
using a non-malignant cell-line, MCF10A, and normal primary mammary tissue sample
from reduction mammoplastiy. To model the loss of BRCA1 function, cells will be
transduced with lentiviral shRNA. We used matrigel system to study spindle orientation
in a malignant, BRCA1 mutated cell line, HCC1937, along with a BRCA1 vector control
and a BRCA1 reconstituted HCC1937 line. This was done to determine whether BRCA1
gain of function would restore spindle orientation bias.
Result: Normal mammary cells form polarized acini after 7-8 days culture in matrigel. Acinar
polarization was evident from apical positioning of the centrosomes and Golgi
apparatus, as well as basal deposition of CD49f (integrin). Mitotic spindle orientation
relative to the center of the acinar lumen was assessed in anaphase and telophase
cells to exclude the possibility of spindle rotations in metaphase. Mitotic spindles were
positioned predominantly planar to the lumen in MCF10A and both luminal progenitors
(LP) and basal progenitors derived (BP) acini. BRCA1 was reconstituted in HCC1937. To
date, I have not found significant differences between the 3D growth and polarity of the
HCC1937 sub-lines.
35
Trainee: Cameron Rogers
Board #: 22
Session #: 3
Supervisor: Catherine Pallen
Title: Reversible phosphorylation of Protein Tyrosine Phosphatase α (PTPα ) in integrin
signaling and cell migration
Author(s): Cameron Rogers, Min Chen, Catherine J Pallen
ABSTRACT:
Cancer metastasis is a major cause of cancer mortality. It is a pathophysiological process dependent on
tumor cell motile properties of migration and invasion. The integrins are transmembrane receptors that
contact the extracellular matrix (ECM) to critically regulate cell migration. Integrin engagement triggers a
complex array of signaling events to promote the formation of multi-protein complexes called focal
adhesions (FAs). FAs are linked to the cytoskeleton and to integrin-ECM contact points. Signaling
pathways originating at the FA determine the dynamic assembly and disassembly of FAs, as well as
cytoskeleton rearrangements. Understanding the complex molecular mechanisms that regulate integrin
signaling, cell shape and motility is important to identify and develop molecular therapeutics to target
aberrant migration and invasion by tumor cells and inhibit metastasis.
Protein tyrosine phosphatase α (PTPα) plays key roles in integrin-mediated cell migration. Our lab has
shown that integrin stimulation induces the phosphorylation of PTPα at Tyr789 and that this is important
to recruit other proteins to FAs and promote cell migration. PTPα phosphorylation is reversible, as the
termination of integrin signaling via cell detachment induces PTPα dephosphorylation by an unknown
phosphatase. Analogously, in the trailing edge of a migrating cell, the local termination of integrin
signaling and FA disassembly must occur to permit forward cell movement, suggesting that PTPα
dephosphorylation is a part of this process. Our study aims to elucidate the regulation of PTPα
dephosphorylation in this process and identify the responsible phosphatase.
Using mouse embryo fibroblasts (MEFs), we have determined the kinetics of PTPα dephosphorylation
upon termination of integrin signaling by placing cells in suspension, with ~50% of PTPα being dephosphorylated in 30 min. Using MEFs from genetically modified mice deficient in various PTPs, we have
shown that several candidate PTPs implicated in integrin signaling, including PTP1B, TC-PTP, PTP-PEST,
PTEN and SHP2, are not required for PTPα dephosphorylation. We are investigating other PTPs as
candidate regulators of PTPα phosphorylation status, and have recently found that a PTP known as STEP
(Striatal-Enriched Protein Tyrosine Phosphatase) complexes with PTPα in MEFs. Our ongoing studies will
establish whether STEP is an integrin signaling-linked regulator of PTPα-phosphoTyr789.
36
Trainee: Arjun Trivedi
Board #: 23
Session #: 3
Supervisor: Jean-Pierre Chanoine
Title: Insights into the role of ghrelin and growth hormone in regulating maternal glucose
homeostasis in mice
Author(s): Arjun Trivedi, William Gibson, Sandra Babich, Jean-Pierre Chanoine
ABSTRACT:
Background:
Hypothesis: Method: Result: Conclusion: Ghrelin is secreted primarily by the stomach and circulates as acylated (AG) and
unacylated ghrelin (UAG). Acylation is mediated by the enzyme ghrelin O-acyltransferase
(GOAT). AG protects against hypoglycemia in non-pregnant mice by facilitating pituitary growth hormone (GH) release. In humans during the prenatal period, pituitary GH
is replaced by placental GH yet is not well-characterized in mice. In addition, maternal
plasma AG concentrations are markedly low and rise post-partum suggesting a possible
role in the mother during pregnancy. In essence, AG’s influence on GH concentrations in
pregnant mice remains unknown. Furthermore, pregnancy poses a high-risk period for
hypoglycemia during malnutrition to maintain maternal euglycemia whilst meeting fetal
energy demands.
1) Calorie-restriction (CR) causes a more severe hypoglycemia in pregnant mice
2) Upregulation of GOAT/ghrelin, increased plasma AG and GH concentrations are part
of the hormonal response to this hypoglycemia during CR
Wild-type (WT) and GOAT-KO, pregnant and non-pregnant mice were time-mated and
were subjected to a freely-fed (FF) diet or 50% CR for 7 days. CR occurred between days
11 to 18 of pregnancy where they were sacrificed (~day 20=delivery). Blood glucose
(BG) was measured by a OneTouch glucometer, plasma AG, UAG and GH were
measured by EIA and GOAT/ghrelin expression by RT-PCR.
1) In WT mice, CR did not affect BG in non-pregnant mice (6.1±1.8mmol L;p>0.05;n=20)
but caused hypoglycemia in pregnant mice (4.3±0.9mmol/L;p<0.005;n=9). The latter
was associated with higher circulating AG and GH concentrations as well as lower
stomach GOAT and ghrelin mRNA (p<0.05).
2) In GOAT-KO mice, where plasma AG is by definition absent, CR caused severe
hypoglycemia (2.9mmol/L;n=1) in pregnant mice characterized by morbidity and
systemic P termination and hypoglycemia in NP mice (5.3±0.8mmol/L;p<0.001;n=19).
The latter was associated with higher circulating UAG, undetectable GH depletion
and lower stomach ghrelin mRNA (p<0.05).
1) GOAT-KO mice are more prone to hypoglycemia, suggesting a role for AG that is
independent from GH secretion;
2) The AG increase during CR not due to increased stomach production but likely
decreased degradation.
3) AG is not necessary for placental GH production in pregnant mice.
37
Trainee: Sarah White
Board #: 24
Session #: 3
Supervisor: Dan Luciani
Department:Surgery
Title: Combined and individual roles of pro-apoptotic Bax and Bak in beta-cell stress signalling
and death
Author(s): Sarah A White, James D Johnson, Dan S Luciani
ABSTRACT:
Functional failure and loss of pancreatic β-cells is a critical event in the pathogenesis of diabetes and
there is mounting evidence that suggests chronic endoplasmic reticulum (ER) stress contributes
significantly to β-cell dysfunction and apoptotic death. Two core pro-apoptosis proteins, Bax and Bak,
mediate the execution of mitochondrial apoptosis and have also been suggested to regulate aspects
of ER physiology and stress signalling. In this study we set out to determine the relative contributions of
Bax and Bak in the execution of β-cell death and examine their putative roles in β-cell ER-stress
signalling under diabetogenic conditions. We generated mice in which the single or combined knockout
of Bax and Bak could be induced in the pancreatic β-cell. Physiological islet function assessed both in
vivo and in vitro was not affected by the knockout of Bax and/or Bak. However, Bax and Bak knockout
provided protection towards β-cell survival under stress conditions. Single knockout, double knockout,
and wild-type cells were assayed for ER-stress and cell death following treatment with staurosporine,
thapsigargin, and glucolipotoxic conditions. Time-course kinetic cell death analysis demonstrated the
expected protection of single and double knockout cells from staurosporine, and further revealed that
Bax-Bak double knockout was required for significant protection from glucolipotoxic death. ER-stress
signalling was evaluated by quantitative PCR for BAX, XBP1s and CHOP. BAX expression was significantly upregulated after 48 hours of thapsigargin-induced ER-stress. Interestingly, spliced XBP1 expression
was augmented in Bax-Bak double knockout islets under early ER-stress signalling compared to
wild-type controls. CHOP expression increased in a time-dependent manner but was not significantly
different between Bax-Bak double knockout and WT islets. These results suggest that Bax and Bak
regulate a specific arm of the ER-stress response upstream of apoptosis by suppressing maximal XBP1
splicing. Together these data suggest that Bax and Bak have both individual and combined contributions
to β-cell death under various stress conditions and have alternate roles regulating early ER-stress
signalling in the β-cell.
38
Trainee: Grace Goh
Board #: 25
Session #: 3
Supervisor: Stefan Taubert
Title: The Mediator subunit MDT-15 is required for the oxidative stress response
Author(s): Grace Y S Goh, Kulveer S Parhar, Ada W L Kwong, Marcus A Wong, Stefan Taubert
ABSTRACT:
Reactive oxygen species (ROS) can damage cellular components, yet they are also required for a multitude of physiological processes. Thus, ROS levels must be tightly controlled. One critical level of control
is through transcriptional regulation of ROS detoxification genes. Here we identify MDT-15, a subunit of
the Mediator complex, as a key player in ROS detoxification. MDT-15 acts as a transcriptional coregulator and is required to express many detoxification genes; as such, it is required for worms to survive on
both the metalloid sodium arsenite and the organic peroxide t-BOOH. This requirement is independent
of MDT-15’s previously characterized role in lipid metabolism, as knockdown or mutation of the fatty acid
desaturase fat-6, a known target of MDT-15, did not reduce survival on t-BOOH, and supplementation
with polyunsaturated fatty acids did not rescue the hypersensitive phenotype of mdt-15 mutants. The
conserved transcription factor SKN-1/Nrf is required for arsenite-dependent gene inductions, and we
find evidence for functional and physical interaction between MDT-15 and SKN-1. Specifically, we show
that in a yeast two-hybrid system, MDT-15 physically interacts with SKN-1 via a previously uncharacterized region of MDT-15. MDT-15 is also required for the induction of SKN-1 target genes, making it a
novel transcriptional coregulator of SKN-1 in the context of oxidative stress. Furthermore, others have
shown that the t-BOOH response is largely SKN-1 independent, suggesting that other transcription
factors must cooperate with MDT-15 in this context. Our preliminary data suggest that Nuclear Hormone
Receptors may cooperate with MDT-15 to regulate the response to t-BOOH. In sum, we propose that
MDT-15 is broadly required for the transcriptional response to oxidative stress by acting in concert with
multiple transcription factors.
39
POSTER SESSION FOUR
doctoral students
MODERATOR:
PARTICIPANTS:
Tobias Kollmann
Maria Aristizabal
Helen Chen
Jianjia Fan
Yu-Hsuan Huang
Safia Ladha
Sarah Munro
Terri Petkau
Magda Price
Dominik Sommerfeld
Yu Yao
DOCTORAL STUDENTS | POSTER SESSION FOUR
Trainee: Maria Aristizabal
Board #: 26
Session #: 4
Title: High-throughput genetic and gene expression analysis of the RNAPII-CTD reveals
unexpected connections to SRB10/CDK8
Author(s): Maria J Aristizabal, Gian Luca Negri, Joris J Benschop, Frank C P Holstege,
Nevan J Krogan, Michael S Kobor
ABSTRACT:
The C-terminal domain (CTD) of RNA polymerase II (RNAPII) is composed of heptapeptide repeats,
which play a key regulatory role in gene expression. Using genetic interaction, chromatin immunoprecipitation followed by microarrays and mRNA expression analysis we found that truncating the CTD resulted
in distinct changes to cellular function. Truncating the CTD altered RNAPII occupancy to not only cause
decreases, but also increases in mRNA levels. The latter were largely mediated by promoter elements,
and in part were linked to the transcription factor Rpn4. The mediator subunit Cdk8 was enriched at
promoters of these genes, and its removal not only restored normal mRNA and RNAPII occupancy levels,
but also reduced the abnormally high cellular amounts of Rpn4. This suggested a positive role of Cdk8
in relationship to RNAPII, which contrasted with the observed negative role at the activated INO1 gene.
Here, loss of CDK8 suppressed the reduced mRNA expression and RNAPII occupancy levels of CTD
truncation mutants.
43
Trainee: Helen Chen
Board #: 27
Session #: 4
Supervisor: Christopher Maxwell
Title: RHAMM acts near chromosomes to promote mitotic spindle assembly by regulating the
location and abundance of TPX2 and the downstream activation of Aurora A
Author(s): Helen Chen, C James Lim, Christopher A Maxwell
ABSTRACT:
Enhanced cell proliferation is a hallmark of cancer. Thus, molecular pathways underlying mitosis are
important targets for therapy. To form a bipolar mitotic spindle, the mitotic cell must assemble microtubules at centrosomes [i.e. the spindle poles] as well as non-centrosome sites [i.e. kinetochores]. The
spindle then captures and aligns the duplicate chromosomes. Correct attachment satisfies the spindle
assembly checkpoint (SAC) and allows mitotic progression.
Aurora kinase A is a key regulator of spindle assembly. When bound by its activator, TPX2, the kinase
mediates spindle assembly through phosphorylation of downstream substrates. Since TPX2 is necessary
for optimal Aurora A function, its availability is tightly controlled.
The receptor for hyaluronan-mediated motility (RHAMM) is a microtubule associated protein demonstrated to partner with TPX2 to influence Aurora A activity and participate in spindle assembly in both human
and Xenopus cells. We hypothesize that RHAMM interacts with TPX2 to determine TPX2 location and
abundance, which determines Aurora A kinase activity and spindle assembly.
To date, we have followed the location of RHAMM through the early stages of mitosis, its interacting proteins, as well as the consequences of its loss. We tracked the location of RHAMM by IF and GFP-RHAMM
and found it at the kinetochores, an as-yet undescribed localization. We colocalized RHAMM with key
kinetochore proteins, BUBR1 and NDC80, during microtubule nucleation and confirmed physical associations by IP. For insights into its functions, we followed mitotic progression in real-time with HeLa cells.
We quantified a significant increase in the number of cells exhibiting aberrant spindle morphology and
mitotic failures with siRNA mediated RHAMM depletion. We also followed the kinetics of mitosis based
upon duration of spindle assembly, chromosome congression and cytokinesis. Live cell investigations reveal that RHAMM depletion delays mitotic progression at two key events: 1) cells are unable to construct
a proper bipolar spindle; 2) cells are able to construct a phenotypically “normal” mitotic spindle, but fail
to satisfy the SAC. Thus, the protein’s location, physical interactions, and the consequences of its depletion all point to a critical role at the kinetochore.
Recently, we have identified that RHAMM depletion results in TPX2 mislocalization and reduced TPX2
protein abundance at the spindle poles. In addition, we have shown reduced protein abundance of
both native and active forms of Aurora A. These results suggest that RHAMM may be targeting TPX2 to
spindle assembly sites, and protecting TPX2 from proteolytic degradation. TPX2 then influences downstream Aurora A activation.
We are currently investigating which region of RHAMM is of functional importance, in regulating spindle
assembly and TPX2 interactions. Further insights into this pathway may improve treatments for cancers
linked to these proteins, such as paediatric sarcomas and BRCA-1 associated early onset breast cancer.
44
Trainee: Jianjia Fan
Board #: 28
Session #: 4
Supervisor: Cheryl Wellington
Title: Liver-X-Receptor and progesterone receptor activation promote apolipoprotein E
secretion from CCF-STG1 astrocytoma cells
Author(s): Jianjia Fan, Yoko Shimizu, Jeniffer Chan, Anna Wilkinson, Ayaka Ito, Peter Tontonoz,
Tom Pfeifer, Cheryl L Wellington
ABSTRACT:
Apolipoprotein E is the major lipid carrier in the central nervous system. As apoE plays a major role in
the pathogenesis of Alzheimer’s Disease (AD) and also modulates repair pathways after several forms of
acute brain injury, modulating the expression, secretion or function of apoE may provide potential therapeutic approaches for several neurological disorders. Here we report that progesterone and a synthetic
progestin, lynestrenol, significantly induce apoE secretion from human CCF-STTG1 astrocytoma cells.
Intriguingly, lynestrenol increases expression of the cholesterol transporter ABCA1 via a liver-X-receptor
(LXR) dependent pathway in CCF-STTG1 cells. ApoA-I- and apoE3-mediated cholesterol efflux is also
enhanced by lynestrenol in these cells. The progesterone receptor (PR) inhibitor RU486 attenuates the effect of lynestrenol and progesterone on apoE expression in human astrocytoma cells but has no effect on
ABCA1 expression, suggesting that the PR participates in apoE regulation but does not affect ABCA1.
These results demonstrate the selectivity of certain reproductive hormones to regulate apoE secretion by
activating both LXR-dependent and PR-dependent pathways in glial cells.
45
Trainee: Yu-Hsuan Huang
Board #: 29
Session #: 4
Title: SAP is required for CD8 T cells differentiation and function upon B cells stimulation
Author(s): Yu-Hsuan Huang, Kevin Tsai, Rusung Tan, John J Priatel
ABSTRACT:
X-linked lymphoproliferative disease (XLP) is a congenital immunodeficiency and humans with XLP often
develop fatal complications and more severely, lead to B lymphoma upon exposure to Epstein-Barr virus
(EBV). Most people with XLP have mutations in SAP (SLAM-associated protein), a small SH2-containing
adaptor transmits signaling via SLAM (signaling lymphocytic activation molecule) family receptors which
express in hematopoietic cells. The SLAM-SAP signaling, through lymphocytes interaction, has been
shown to be critical for development and function of multiple immune cell types. However, the precise
mechanism of how loss of SAP contributes to susceptibility to EBV, a B lymphotropic virus, and immune
dysfunction remains unclear. Here, we have investigated the role of SAP signaling in regulating the differentiation and function of naïve CD8 T cells upon B cells activation. In vitro experiments revealed that
OT-1 CD8 T cells, recognize ovalbumin peptide specifically, from SAP deficient mice (SAP-/-) exhibit
a greatly diminished potential in proliferation and cytokine secretion as compared to wild-type T cells
upon OVA coated B cells or OVA presented B lymphoma stimulation. More interestingly, decreased differentiation ability are found in SAP-/- OT-1 CD8 T cells in vivo after adoptive transfer to mice inoculated
with OVA presented B lymphoma. Together, our results suggest SAP is required for antigen priming of
CD8 T cells when B cells act as primary APC.
46
Trainee: Safia Ladha
Board #: 30
Session #: 4
Supervisor: Michael Hayden
Title: Examining conditional caspase-6 deficiency as a therapeutic in YAC128
Author(s): Safia Ladha, Bibiana K Y Wong, Michael R Hayden
ABSTRACT:
Huntington disease (HD) is an autosomal dominant neurodegenerative disorder characterized by
motor, cognitive, and psychiatric symptoms, followed by death 15 to 20 years after onset. HD is caused
by a CAG repeat expansion in the huntingtin (HTT) gene resulting in the production of mutant huntingtin (mHTT). Caspase-6 (C6) is a cysteine aspartyl protease that plays a central role in apoptosis and has
recently been implicated in several neurodegenerative diseases including HD. Increased C6 activation is
observed in early grade human HD brain samples and in mouse models of HD, and genetic or peptide
inhibition of C6 activity in vitro protects neurons from degeneration. C6 is one of several proteases that
cleaves mHTT, subsequently releasing toxic fragments. Our lab has previously demonstrated that
preventing C6-mediated cleavage of mHTT at the 586 amino acid site using mice expressing C6-resistant mHTT (C6R) protects against enhanced extrasynaptic NMDAR activity, preserves striatal volume and
rescues cognitive and motor deficits in the YAC128 mouse model of HD. These data point to an essential
role for C6 in the pathogenesis of HD and provide a rationale for the inhibition of C6 as a therapeutic
approach.
The goal of this study is to investigate the effect of adult C6 inhibition on the YAC128 phenotype. This
will be done by crossing a C6flox/Cre-ERT2 mouse with the YAC128 mouse and inducing excision of C6
by treatment with tamoxifen. These mice will be treated at different ages and examined for behavioral,
neuropathological and biochemical endpoints. The use of an inducible C6 knockout mouse allows for
regulated adult C6 knockdown, an approach that is a more accurate model of pharmacological C6
inhibition than a constitutive gene knockout and more closely reflects the effects of a therapy given in
human adulthood. Results from this study will provide insight into the effect of adult C6 inhibition on HD
pathogenesis and will further validate C6 as a therapeutic target in HD.
47
Trainee: Sarah Munro
Board #: 31
Session #: 4
Supervisor: Patricia Janssen
Department: School of Population & Public Health
Title: Choice, autonomy, and consumer demand: The construction of “Cesarean Delivery on
Maternal Request”
Author(s): Sarah Munro, Patricia Janssen ABSTRACT:
Introduction/ In the late 1990s, amid concerns about rising caesarean rates, medical journal editorials
Objective: and letters to the editor began to discuss the subject of “cesarean delivery on maternal
request” (CDMR) – caesarean performed without medical indications at the mother’s
request. I explore how authors persuaded the obstetric community to view the emerging
concept of CDMR as a real phenomenon and as an appropriate/inappropriate mode of
delivery.
Method: The study sample (n=42) consisted of medical journal editorials and letters to the editor
published 1998-2012. Data collection and analysis were guided by genre theory and
social constructionist grounded theory. Sample texts were read in their entirety and
coded for recurring and interesting concepts and generic elements that constituted
rhetorical action. Codes were refined into categories through iterative analytic memoing
and “situational mapping” – an exercise illustrating the elements of the situation of
inquiry and their interrelationships. Results were written into an explanatory narrative and
discussed within the context of the prevalence of CDMR and changing North American
obstetric policy.
Result: My poster will focus on one central theme emerging from analysis: respect for patient
autonomy. Proponents of CDMR constructed patients as autonomous consumers
managing their own reproductive choices, while critics argued that patient autonomy
was circumscribed by medico-legal, financial, and sociocultural factors. Neither side of
the debate explicated the features of informed decision-making that might support and
enact patient autonomy. Early authors (1998-2004) were largely supportive of CDMR
while those who published more recently (2004-2012) were critical of the subject,
reflecting temporal changes in policy and practice guidelines. The prevalence of CDMR
was overstated in early publications, while later articles, responding to emerging data on
the low prevalence of CDMR in Canada and the United States (0.34%-2.8%), questioned
whether CDMR was a real phenomenon.
Discussion: CDMR did not contribute to the rising caesarean rate; rather leaders in the obstetric
community constructed the phenomenon and argued it was an appropriate mode of
delivery based on the principle of respect for patient autonomy. Physician values and
attitudes towards consumer-driven health care influenced health policy regarding CDMR.
48
Trainee: Terri Petkau
Board #: 32
Session #: 4
Title: Sensitivity to neurotoxic stress is not increased in progranulin-deficient mice
Author(s): Terri L Petkau, Shanshan Zhu, Ge Lu, Sarah Fernando, Pam Wagner, Max Cynader,
Blair R Leavitt
ABSTRACT:
Loss-of-function mutations in the progranulin (GRN) gene are a common cause of autosomal dominant
frontotemporal lobar degeneration (FTLD), a fatal and progressive neurodegenerative disorder common
in people under 65. In the brain, progranulin is expressed in multiple regions at varying levels, and has
thus been hypothesized to play a neuroprotective or neurotrophic role. We used in vivo injections of four
neurotoxic agents in constitutive progranulin-knockout (Grn-/-) mice and their wild-type (Grn+/+)
counterparts to assess neuronal sensitivity to toxic stress. Administration of 3-nitropropionic acid,
quinolinic acid, kainic acid, and pilocarpine induced robust and measurable neuronal cell death in
affected brain regions, but no differential cell death was observed between Grn+/+ and Grn-/- mice.
We conclude that the acute chemical models of neuronal injury used in the current study do not cause
increased neuronal cell death in constitutive progranulin knockout mice.
49
Trainee: Magda Price
Board #: 33
Session #: 4
Supervisor: Wendy Robinson
Title: Additional annotation enhances biologically relevant sub classification of the Illumina
HumanMethylation450 BeadChip array
Author(s): E Magda Price, Allison M Cotton, Lucia L Lam, Pau Farre, Eldon Emberly,
Carolyn J Brown, Wendy P Robinson, Michael S Kobor ABSTRACT:
Measurement of genome-wide DNA methylation (DNAm) has become an important avenue for
investigating potential physiologically-relevant epigenetic changes. Illumina Infinium is a commercially
available microarray suite used to measure DNAm at many sites throughout the genome. However, it has
been suggested that a subset of array probes may give misleading results due to issues related to probe
design. To facilitate biologically significant data interpretation, we set out to enhance probe
annotation of the newest Infinium array, the HumanMethylation450 BeadChip (450k), with >485,000
probes covering 99% of RefSeq genes. Annotation that was added or expanded on includes: 1)
documented SNPs in the probe target, 2) probe binding specificity, 3) CpG classification of target sites
and 4) gene feature classification of target sites. Probes with documented SNPs at the target CpG (4.3%
of probes) were associated with increased within tissue variation in DNAm. An example of a probe with a
SNP at the target CpG demonstrated how sample genotype can confound the measurement of DNAm.
Additionally, 8.6% of probes mapped to multiple locations in silico. Measurements from these
non-specific probes likely represent a combination of DNAm from multiple genomic sites. The expanded
biological annotation demonstrated that based on DNAm, grouping probes by an alternative high-density and intermediate-density CpG island classification provided a distinctive pattern of DNAm. Finally,
variable enrichment for differentially methylated probes was noted across CpG classes and gene feature
groups, dependent on the tissues that were compared. DNAm arrays offer a high-throughput approach
for which careful consideration of probe content should be utilized to better understand the biological processes affected. Probes containing SNPs and non-specific probes may affect the assessment of
DNAm using the 450k array. Additionally, probe classification by CpG enrichment classes and to a lesser
extent gene feature groups resulted in distinct patterns of DNAm. Thus, we recommend that compromised probes be removed from analyses and that the genomic context of DNAm is considered in studies
deciphering the biological meaning of Illumina 450k array data.
50
Trainee: Dominik Sommerfeld
Board #: 34
Session #: 4
Title: The role of specific Protein Phosphatase 2A (PP2A) holoenzymes in orchestrating the cell
cycle symphony
Author(s): Dominik D Sommerfeld, Catherine J Pallen
ABSTRACT:
Protein phosphatase 2A (PP2A), a highly abundant, ubiquitous and conserved Ser/Thr-specific
phosphatase, has been shown to regulate a plethora of important cellular processes, including the cell
cycle progression. PP2A is a heterotrimeric holoenzyme complex composed of a catalytic subunit, a
scaffolding subunit, and a variable, regulatory subunit. Genetic alterations in the scaffolding subunit and
genomic deletions or loss of heterozygosity (LOH) of regulatory subunits have been identified in various
cancers, including breast, ovarian and cervical carcinomas. In particular, recurrent genomic deletions of
the PPP2R2A (B55alpha) and PPP2R3B (PR70/48) subunits have been found in specific subtypes of
luminal breast and endometrial carcinoma.
Therefore, it is likely that dysregulation or dysfunction of PP2A holoenzymes containing these specific
subunits plays an important role in these cancers. Many potential downstream targets of PP2A
dysregulation have been reported, including cell cycle regulators. Specifically, the B55alpha regulatory
subunit has been implicated in the regulation of mitotic entry and exit, while the PR70/48 subunit is
believed to be involved in regulation of DNA replication. Our aim is to investigate the functional
consequences of loss of the B55alpha and PR70/48 subunits on cell cycle control, and to elucidate
underlying molecular pathways, so as to gain insight into the roles of PP2A genetic alterations in cancer.
51
Trainee: Yu Yao
Board #: 35
Session #: 4
Supervisor: Megan Levings
Department:Surgery
Title: Treg cells mediate suppression of IL-1β production
Author(s): Yu Yao, Jens Vent-Schmidt, Gijs Hardenberg, Casey Chan, Theodore Steiner,
Megan Levings
ABSTRACT:
Regulatory T cells (Tregs) are indispensable for regulating adaptive immunity as well as inflammation
mediated by the innate immune system. Dysregulation of innate immunity can lead to uncontrolled
activation of the inflammasome and subsequent maturation and release of pro-inflammatory IL 1 family
cytokines, resulting in excess inflammation and tissue destruction. Delivery of Tregs can dampen mucosal
inflammation and cure colitis in experimental models, but whether or not they can suppress inflammasome activation is unknown. To test this, we co-cultured human Tregs or conventional T cells (Tconv) with
allogeneic macrophages and stimulated the inflammasome by addition of LPS and ATP. We found that
Tregs but not Tconvs potently suppressed the transcription and release of mature IL 1β. Similar
experiments confirmed this finding in mice. To investigate the mechanism(s) by which Tregs suppress
IL-1β production, mouse Treg-conditioned media was added to macrophages stimulated with LPS and
ATP, and found to similarly block IL-1β in a dose-dependent manner. Preliminary data suggest that IL-10,
but not TGF β, in the Treg-conditioned media mediates this effect. These data represent the first evidence for Treg-control of IL-1β production and have implications for the application of Treg cell therapy
in inflammatory bowel disease.
52
POSTER SESSION FIVE
doctoral students
MODERATOR:
PARTICIPANTS:
Cheryl Wellington
Ana Cohen Parastoo Kheirkhah Dehkordi
Jennifer Grants
Grace Leung
Elizabeth Marchant
Bo Peng
Veronica Schiariti
Kevin Tsai
Joanna Yeung
DOCTORAL STUDENTS | POSTER SESSION FIVE
Trainee: Ana Cohen
Board #: 36
Session #: 5
Supervisor: William Gibson
Title: Mutational analysis of EZH2 in patients with Weaver and Weaver-like syndromes
Author(s): Ana S A Cohen, Jieqing Xu, Xiaohua Han, Joanne Denny, Katelin Townsend,
Damian B Yap, Samuel A J R Aparício, Colin J D Ross, William T Gibson
ABSTRACT:
In late 2011, our lab found that constitutional mutations in the epigenetic regulator EZH2 (enhancer of
zeste homolog 2) cause Weaver Syndrome (WS). WS is characterized by overgrowth, increased height,
large head, intellectual disability and susceptibility to various cancers. Having established a diagnostic
test, we are now investigating the link between these mutations, WS, and cancer development.
All coding exons of EZH2 were sequenced in 27 individuals referred to us from all over the world with
Weaver-like phenotypes. To date, one novel mutations as well as two reoccurring mutation were
identified in a subset of these individuals. We are looking at phenotype/genotype correlations, with
particular emphasis in determining characteristics that would help us predict the likelihood of these
patients developing cancer. This would allow for early screening to detect both solid tumours and
haematological malignancies in these patients, which should hopefully increase the patients’ chances of
survival.
In collaboration with the Aparicio lab at the BC Cancer Agency, we have investigated the effects of
WS-associated mutations on protein function using in vitro assays. Normally, EZH2 acts as a histone
methyltransferase in the Polycomb repressive complex 2 (PRC2), and silences transcription through
trimethylation of histone H3 lysine 27 (H3K27me3). In contrast to the enhanced trimethylation activity
seen in some somatic mutations that cause leukemia, the WS-associated EZH2 mutants show reduced
methylation activity. Although strategies to reduce EZH2 activity are currently being explored in common
cancers such as leukemias, prostate cancer and breast cancer, our data suggest that these strategies may
not be effective in rare cancers associated with Weaver syndrome.
In our cohort of families with cancer syndromes similar to Weaver syndrome who do not have mutations
in EZH2, trio-based exome sequencing will be applied to find additional cancer genes, and their
oncogenic potential will be investigated through functional studies at the BC Cancer Agency.
55
Trainee: Parastoo Kheirkhah Dehkordi
Board #: 37
Session #: 5
Supervisor: Guy Dumont
Department: Electrical & Computer Engineering
Title: Estimation of heart rate variability from Photoplethysmography (PPG) signal in children
with and without sleep disordered breathing
Author(s): Parastoo Dehkordi, Ainara Garde, Walter Karlen, J Mark Ansermino, Guy A Dumont
ABSTRACT:
Heart Rate Variability (HRV), the variation in the time interval between heartbeats is one of the most
promising and widely used quantitative markers of autonomic activity. HRV can be used as a
discriminable index for identifying individuals with sleep disordered breathing (SDB).Traditionally, HRV is
measured as the series of instantaneous cycle intervals of ECG signal. In this study, we investigated the
estimation of variation in heart rate from a photoplethysmography (PPG) signal. The pulsatile feature of
the PPG waveform is synchronized with each heartbeat and its fundamental frequency depends on heart
rate. We obtained pulse to pulse variability, called pulse rate variability (PRV) from peak to peak time
intervals of PPG signal and assessed its accuracy as an estimate of HRV in children with and without
sleep disordered breathing (SDB). The PPG was recorded from 63 children undergoing sleep study by
an oximeter sensor connected to a mobile phone simultaneously with ECG and other signals in a full
channel PSG. We used correlation and Bland Altman analysis for comparing the parameters of HRV and
PRV in both groups of children. Significant (p <0.05) correlations (0.90 < r < 1) and good agreement
were found between HRV and PRV for mean of intervals, SDNN and RMSSD parameters. However Bland
Altman analysis showed the large divergence for LF/HF ratio parameter. In conclusion, PRV can be used
instead of HRV but the LF/HF ration should be used by more consideration.
56
Trainee: Jennifer Grants
Board #: 38
Session #: 5
Supervisor: Stefan Taubert
Title: The Mediator subunit CDK-8 is a dual negative/positive regulator of EGFR-Ras-MAPK
signaling
Author(s): Jennifer M Grants, Lisa T L Ying, Stefan Taubert
ABSTRACT:
The epidermal growth factor receptor-Ras-mitogen activated protein kinase signaling pathway (EGFR
signaling) controls fundamental cellular processes, such as proliferation and differentiation, which are
required for organismal development and implicated in cancer. Thus, EGFR signaling must be tightly
regulated. The conserved transcriptional coregulator complex, Mediator, may regulate EGFR signaling.
Here, we identify Mediator subunit cyclin dependent kinase 8 (CDK-8) as a novel regulator of the EGFR
signaling pathway in Caenorhabditis elegans. Using C. elegans vulval induction as a measure of EGFR
signaling activity, we demonstrate that CDK-8 negatively impacts this pathway.
Specifically, cdk-8 null mutation increases EGFR signaling activity, which is significantly enhanced by
mutations in the so-called synthetic multivulva (synMuv) genes, well-characterized negative regulators of
EGFR signaling. SynMuv genes primarily regulate EGFR signaling by repressing transcription of the EGF
ligand, lin-3; in line with cdk-8’s genetic interactions with synMuv genes, we find that CDK-8 also
represses lin-3 transcription. Genetic epistasis analysis also places cdk-8 downstream of the receptor
let-23/EGFR. In line with this position, we find evidence that CDK-8 may act as a corepressor for a
transcription factor downstream of the EGFR signaling pathway, LIN-1, as loss of cdk-8 enhances the
increase in EGFR signaling activity caused by lin-1 RNAi knockdown. Unexpectedly, CDK-8 also takes on
a positive regulatory role in the EGFR signaling pathway in the absence of another transcription factor
downstream of this pathway, LIN-31: we observe that cdk-8 is required for the increase in EGFR
signaling activity seen in lin-31 null mutants. Thus, cdk-8 acts downstream or in parallel to lin-31 in this
positive role, suggesting that CDK-8 may be an important coactivator for another transcription factor
downstream of the EGFR signaling pathway. Overall, this work identifies both positive and negative
regulatory roles for CDK-8 within an important developmental and oncogenic signaling cascade.
57
Trainee: Grace Leung
Board #: 39
Session #: 5
Title: Functional characterization of the BRCT-domain containing protein Rtt107 in the DNA
damage response
Author(s): Grace P Leung, Michael S Kobor
ABSTRACT:
Maintenance of genome integrity is critical for proper cellular function, with perturbations potentially
having serious ramifications for health and disease. Cells are constantly exposed to DNA damage,
and therefore have a multifaceted network of pathways that regulate the DNA damage response. One
characteristic of the DNA damage response is the assembly of large protein complexes, often mediated
by scaffolding proteins, which frequently contain BRCT (BRCA1 C-terminal) domains. In Saccharomyces
cerevisiae, the BRCT-domain containing protein Rtt107 is important for the response to DNA damage
during S-phase. It is phosphorylated by the checkpoint kinase Mec1 in response to DNA damage, and
interacts with a number of protein complexes. In this study we sought to further examine the role of
Rtt107 as a scaffolding protein in the DNA damage response. Upon induction of a specific double strand
break (DSB) in the genome, Rtt107 was recruited to the site of the DNA lesion.
Moreover, this recruitment was dependent on the presence of H2A phosphorylation, and was mediated
by the C-terminal BRCT domains of Rtt107. Rtt107 in turn was required for the recruitment of multiple
proteins to DSBs, including the SMC5/6 complex, a highly conserved complex involved in multiple
genome maintenance activities, and Slx4, the interaction partner of Rtt107. The regulation of Rtt107
recruitment and its effects were also studied in the context of a protein-bound nick. The use of a
different type of DNA lesion revealed some differences in Rtt107 function, supporting the idea that the
cell utilizes specific responses to different types of DNA lesions. Together, this work provides new insight
into the role of Rtt107 at DNA damage sites and its interactions with other DNA damage response
factors.
58
Trainee: Elizabeth Marchant
Board #: 40
Session #: 5
Supervisor: Pascal Lavoie
Title: Impaired toll-like receptor responses in very premature neonates during the neonatal
period
Author(s): Elizabeth A Marchant, Mihoko Ladd, Ashish Arunkumar Sharma, Alice van Zanten,
Nicole Farrell, Pascal M Lavoie
ABSTRACT:
Background:
Over the last few decades, the morbidity and mortality from infections has increased in
premature neonates despite improvements in neonatal care. We and others have shown
previously that cord blood cells from very premature neonates exhibit poor inflammatory
responses to Toll-like receptor (TLR) immune stimulation. On the other hand, the
developmental maturation of the innate immune responses during the neonatal period
has not been studied in these infants.
Objective: To determine whether the deficits in immune response previously reported in cord blood
from very premature neonates persists during the neonatal period, at a time when they
are most susceptible to infection.
Design/
Multiple cytokine responses (IL-1-beta, IL-6, IL-10, TNF-alpha, and IL-12p40) to TLR4
Method: (LPS) and TLR 7/8 (R848) stimulation (24h) using whole blood in vitro between very
premature neonates (born < 30 weeks; 6-23 days of age; peripheral blood; n=14),
preterm (cord blood; n=8), term neonates (cord blood; n=20) and healthy adults (n=18),
using rigorous Standard Experimental Procedures. Neonates with suspected or proven
sepsis prior to or at the time of sampling were excluded.
Results: Stimulated pro-inflammatory responses (IL-1-beta, IL-6, IL-10, TNF-alpha, and IL-12/IL23p40) to LPS remained profoundly attenuated in very premature neonates, compared
to term neonates or healthy adults; whereas responses to R848 appeared more mature.
IL-6 and IL-10 responses to LPS were highest in term neonates, and were comparable
between neonates and adults.
Conclusion: This is the first study reporting TLR-mediated immune responses in very premature
neonates during the neonatal period. Our data confirm that the level of innate immune
attenuation previously demonstrated from cord blood persists after birth; these
observations constitute an essential premise to support a major role for impaired innate
immune responses in increasing susceptibility to infections early in life in very premature
neonates.
59
Trainee: Bo Peng
Board #: 41
Session #: 5
Supervisor: Peter Leung
Department: Obstetrics & Gynaecology
Title: GnRH induces RUNX2 expression via ERK and AKT signalling: Possible involvement in
trophoblast invasion and capillary-like network formation?
Author(s): Bo Peng, Hua Zhu, Peter C K Leung
ABSTRACT:
Background:
Gonadotropin-releasing hormone (GnRH) exerts pro-invasive effects on extravillous
trophoblast (EVT)cells in vitro. RUNX2 has been reported to regulate matrix
metalloproteinase MMP2/9 expression and is often associated with invasive and
angiogenic phenotypes. The aim of our study was to investigate the roles of RUNX2 and
MMP2/9 in basal and GnRH-induced trophoblast cell invasion and endovascular
differentiation.
Method: The localization of RUNX2 and MMP2/9 in first-trimester placenta was examined by
immunohistochemistry. Immortalized EVT cells (HTR-8/SVneo) were treated with GnRH,
GnRH antagonist Antide, MEK inhibitor PD98095, PI3K inhibitor LY294002, MMP2/9
inhibitor or siRNA targeting RUNX2 or MMP2/9, alone or in combination. Protein and
mRNA levels were measured by Western blot and RT-qPCR. Invasiveness and vascular
mimicry were evaluated by Matrigel transwell invasion and capillary-like network
formation assays.
Results: GnRH treatment increased capillary-like network formation by HTR-8/SVneo cells,
suggesting a possible role in endovascular differentiation. Moreover, GnRH treatment
increased RUNX2 levels and this effect was attenuated by Antide. GnRH treatment
elevated phospho-ERK and phospho-Akt levels and pre-treatment with PD98095 or
LY294002 attenuated the effects of GnRH on RUNX2. Down-regulation of RUNX2
reduced GnRH-induced invasion and basal MMP2 and MMP9 expression. Moreover,
basal HTR-8/SVneo cell invasion and capillary-like network formation were reduced
following knockdown and/or inhibition of RUNX2, MMP2 or MMP9.
Conclusion: Our results suggest that GnRH acts via its receptor to induce ERK and Akt
phosphorylation which contributes to elevated RUNX2 expression. Moreover,
GnRH-induced increases in trophoblast invasion, and possibly capillary-like network
formation, are mediated by RUNX2, likely in concert with MMP2/9.
60
Trainee: Veronica Schiariti
Board #: 42
Session #: 5
Supervisor: Louise Mâsse
Title: “He does not see himself as being different”: A qualitative analysis of children and
caregivers reports on relevant areas of functioning in Cerebral Palsy using the
International Classification of Functioning coding
Author(s): Schiariti V, Mâsse LC, Sauve K, Klassen A, Cieza, A, O’Donnell M
ABSTRACT:
Background:
Participation in a variety of activities improves children’s health and self-esteem and promotes social integration in their community. Gaining a better understanding of the kind
of activities children with Cerebral Palsy (CP) are able to do and what factors facilitate
their physical abilities and overall functioning will guide professionals planning services
that improve social participation. In the context of the development of an International
Classification of Functioning (ICF) based tool called “ICF Core Sets for children with CP”,
we explored what children with CP and caregivers identify as relevant areas of functioning.
Method: We conducted semi-structured individual interviews with youth diagnosed with CP, as
well as parents of children with CP. The study population represented the full distribution
of CP severity. Each interview covered all ICF-CY components. Interviews qualitatively
assessed the children and caregivers’ perspectives on relevant areas of functioning.
Audio-recorded interviews were transcribed verbatim in Nvivo. Themes were linked to
the ICF-CY by two trained professionals.
Results: 10 youth and 22 parents were interviewed (10 parent-child dyads). Mean age of youth
participants was 11.7 years (range 8-15yr), 74% male. For parent interviews, mean child
age was 9.4 years (range 4–16yr). Children were Gross Motor Function Classification System levels I-V. Overall; we identified 589 themes that were linked to 146 ICF-CY categories. Most themes were linked to the components activity and participation and environmental factors. Children described areas related to mobility, recreation and leisure and
self-care. In contrast, parents discussed more concerns about physical limitations and
environmental factors including barrier/facilitators of everyday activities. Overall, parental perceptions of relevant areas of functioning were discordant from those of children
with CP, both in general and in specific child-parent dyads.
Conclusion: Interpretation of relevant areas of functioning differed between youth with CP and
caregivers. Youth with CP highlighted abilities and facilitators of activities of daily living.
In contrast, caregivers provided a more comprehensive perspective in terms of the ICFCY components, including areas of impairments in body structure/functions, limitations
in activity of daily living and the role of the environment showing the need to consider
both perspectives when assessing functional abilities.
61
Trainee: Kevin Tsai
Board #: 43
Session #: 5
Title: Characterization and regulation of autoreative CD8 T cells
Author(s): Kevin Tsai, Yu-Hsuan Huang, Rusung Tan, John J Priatel
ABSTRACT:
Negative selection via apoptosis is the primary mechanism of central tolerance by which auto-reactive
T cells are eliminated should their T cell receptors (TCR) interact too strongly with the peptide MHC
complexes expressed on thymic epithelial cells. Although negative selection is efficient in eliminating
auto-reactive T cells, some do escape and could potentially cause autoimmunity; to prevent this
development, these T cells could be programmed to enter an antigen-unresponsive state, also known as
the anergy.
Current mouse models employed for the study of T cell anergy are flawed because transgenic TCRs
impede the development of normal T cell compartments. As the result, this raises the question of
whether or not the anergic T cells observed in these mice were the result of altered T cell compartments
or T cell anergic programming. The use of the Vβ5 transgenic mouse strain allows for the preservation of
TCR specificity to OVA while allowing for normal T cell compartments to develop through the
endogenous rearrangement of the TCR Vβ chain. By crossing the Vβ5 strain with RIP-mOVA strain, we
created a more physiologically relevant mouse model to study auto-reactive T cells that have escaped
negative selection. We hypothesize that anergic T cells could be found in this more physically relevant
transgenic strain.
We find that the Vβ5 transgenic mice have near normal T cell compartments, and although present in
very low numbers, OVA specific T cells are detectable in the peripheries of RIP-mOVA Vβ5 mice. Flow
cytometry analysis revealed that these cells are antigen experienced (CD44 high), acutely activated
(CD127 low) and have elevated expression of anergic T cell marker BTLA.
62
Trainee: Joanna Yeung
Board #: 44
Session #: 5
Supervisor: Dan Goldowitz
Title: Disruption of cerebellar nuclear neurons and unipolar brush cells in the Pax6-null Small
Eye mutant
Author(s): J Yeung, T Ha, D J Swanson, M Larouche, D Goldowitz
ABSTRACT:
It is clear that Pax6 is important in cerebellar granule cell development, yet its role in the development of
other rhombic lip (RL) derivatives in the cerebellum is still undefined. The large, glutamatergic
cerebellar nuclear (CN) neurons and unipolar brush cells (UBCs) are derived from the Math1-positive
cerebellar RL. Little is known about the molecular control of the generation of CN neurons and UBCs
during early cerebellar development, other than they require Math1. These neurons transiently express
the transcription factor Pax6 during development.
To elucidate the role of Pax6 in the generation of CN neurons, we studied the Pax6-deficient Small Eye
mutant mouse (Sey/Sey). The developmental, transcriptional profile of wildtype and Pax6-null cerebella
revealed that in the absence of Pax6, the expression of markers of CN neurons and UBCs, such as Tbr1
and Tbr2, respectively, is significantly decreased. Immunohistochemical time-course study of the normal
development of recently postmitotic CN neurons show that they express Pax6, Lmx1a and Tbr1 along
their migratory route from the RL, through the subpial stream, and into the nuclear transitory zone (NTZ).
The loss of Pax6 in the Sey/Sey cerebellum results in a dramatic and previously unheralded absence
of Tbr1-positive and/or Lmx1-positive cells in the NTZ. During later developmental stages, when Tbr1
labels the medial CN in wildtype cerebellum, the Sey/Sey cerebellum lacks Tbr1-positive cells in the
region that should be colonized by CN neurons. To further characterize the changes, we examined the
cytoarchitecture of the NTZ and CN in the Sey mutant and wildtype cerebella. Analysis of cresyl violet
stained material revealed a loss of recognizable organization of the NTZ in the Sey/Sey cerebellum that
coincides with the absence of Tbr1-positive cells. These results indicate a disruption of the CN and UBC
population in the absence of Pax6.
Our present study identifies Pax6 as a critical and necessary transcription factor controlling the
generation of cerebellar nuclear neurons and unipolar brush cells in the mouse cerebellum.
63
POSTER SESSION SIX
doctoral students
MODERATOR:
PARTICIPANTS:
Laura Sly
Chansonette Badduke
Rebecca De Souza
Jonathan Han
Sara Khosravi
Chris Laugen
Paul Sabatini
Ashish Sharma
Sophie Stukas
Bryan Tennant
Allison Watts
DOCTORAL STUDENTS | POSTER SESSION SIX
Trainee: Chansonette Badduke
Board #: 45
Session #: 6
Supervisor: Evica Rajcan-Separovic
Title: Contribution of exome sequencing to understanding the 1q21.1 CNV
Author(s): Chansonette Badduke, Flamingo Tang, Hani Bagheri, Sally Martell, Laura Dumas,
Veronica Searles, James Sikela, Sarah Hamilton, Sandra Marles, Barbara McGillivary,
Suzanne Lewis, Evica Rajcan-Separovic
ABSTRACT:
Copy number variations (CNVs) of 1q21.1 are associated with variable physical abnormalities and levels
of learning difficulties. We used exome sequencing to look for variants in the 1q21.1 region and
genome wide to explain the phenotypic variability seen in two families with 1q21.1 CNVs (3 deletions,
and 2 duplications, and 3 unaffected individuals). Sequence variants for follow-up were selected based
on minor allele frequency (<1%) in common variant databases (NHLBI 6500 and 1000 exomes), high
conservation and pathogenicity scores, and presence in genes with abnormal expression in patient
lymphoblasts derived from previous whole genome expression analysis (Harvard et al., Orphanet Journal
of Rare Diseases, 6:54, 2010). One indel leading to a premature stop codon in an endoplasmic reticulum
(ER) stress response gene in a father with ADHD and his affected son with developmental delay, both
carrying the 1q21.1 duplication, fulfilled all criteria. We used Sanger sequencing to confirm the presence
of the variant and qPCR to show that both individuals with the variant have reduced gene expression.
We then screened our whole genome expression data for abnormal expression in additional ER stress
response genes. Surprisingly, a number of genes from this pathway have reduced expression not only in
the two 1q21.1 duplication subjects but also in carriers of the 1q21.1 deletion from the second family,
who did not have a detectable pathogenic sequence variation.
Combined whole exome sequencing and whole genome expression findings implicate the ER stress
response pathway in carriers of 1q21.1 CNV through pathogenic variants in the stress response
pathway and/or perturbations in the function in one of the genes from 1q21.1 CNV affecting the ER
stress response pathway. This could lead to phenotypic variability dependent on the level of stress each
carrier was exposed to.
67
Trainee: Rebecca De Souza
Board #: 46
Session #: 6
Title: Identification & characterization of regulatory regions of the Huntingtin Gene
Author(s): Rebecca De Souza, Natalia Kosior, Anthony Mathelier, Wyeth Wasserman, Blair Leavitt
ABSTRACT:
Huntington’s Disease (HD) is a neurodegenerative disorder caused by a trinucleotide repeat expansion
in exon 1 of the huntingtin gene (HTT). Despite ongoing research, the function of wild-type HTT remains
largely unknown. In particular, we know very little about how HTT is regulated at the transcriptional level.
By identifying regions of the genome that regulate HTT transcription, and the pathways which activate
these regions, we can better understand wildtype HTT function and identify new therapeutic targets.
Previous studies have focused on short stretches of the promoter region upstream of the transcriptional
start for the HTT gene. While these studies have uncovered a few important regulatory regions and
associated transcription factors (TFs), distant regulatory regions, regions within the HTT gene body and
regions within the 3’ untranslated region have been overlooked. To address this gap in our knowledge
we conducted a bioinfomatic assessment to identify transcriptional regulatory regions using regulation
relative tracks on the UCSC genome browser. Using this method we identified 11 regions of interest
that contain genomic markers indicative of regulatory regions. As screening these regions for putative
transcription factor binding sites (TFBS) generated a lengthy list of TFs. We have subsequently applied
several layers of additional selection criteria, such as ChIP-seq data and expression in relevant tissues, to
shorten and further validate our list of TFs. We are currently validating our top TF hits using EMSA assays
to show that these putative TFBS are functional, and assessing expression in a HTT promoter-luciferase
assay to identify how these TFs could regulate HTT promoter function.
68
Trainee: Jonathan Han
Board #: 47
Session #: 6
Supervisor: Megan Levings
Department:Surgery
Title: Insulin impairs regulatory T cell function: Implications for obesity
Author(s): Jonathan M Han, Scott J Patterson, Jan A Ehses, Megan K Levings
ABSTRACT:
Chronic inflammation is known to drive metabolic dysregulation in obesity and type 2 diabetes.
Although the precise origin of the unchecked inflammatory responses in obesity is unclear, it is known
that over-production of pro-inflammatory cytokines such as TNF-α by innate immune cells has a key role
in the development of metabolic dysfunction. One key hallmark of obesity is high levels of the pancreatic
hormone insulin, and we hypothesized that there may be an unknown link between hyperinsulinemia and
chronic inflammation. Here we show that high levels of insulin impair the ability of regulatory T cells to
suppress inflammatory responses via effects on the AKT/mTOR signaling pathway. Insulin strongly
activates AKT/mTOR signalling in regulatory T cells, leading to specific inhibition of the production of
the anti-inflammatory cytokine IL-10. As a result, insulin hinders the ability of regulatory T cells to
suppress the production of TNF-α by macrophages. Regulatory T cells from the visceral adipose tissue of
hyperinsulinemic, obese mice also have a decrease in IL-10 production and a parallel increase in
production of IFN-γ. These data suggest that the hyperinsulinemia associated with obesity may
contribute to the development of obesity-associated inflammation via a previously unknown effect on
regulatory T cells function.
69
Trainee: Sara Khosravi
Board #: 48
Session #: 6
Supervisor: Mark Ansermino & Guy Dumont
Department: Anesthesia, Pharmacology & Therapeutics/Electrical & Computer Engineering
Title: Robust model predictive control of hypnosis in adults: A simulation study
Author(s): Sara Khosravi, Klaske van Heusden, Guy A Dumont, J Mark Ansermino
ABSTRACT:
Background:
The main difficulty in the design of controllers for closed-loop control of anesthesia is the
significant intra- and inter-patient variability in response to a standard dose of drug. To
attain acceptance for a closed-loop drug delivery from clinicians and regulatory authorities, the control system will require a certification procedure that includes stability and
robust performance criteria. The goal of this work was to design a model predictive
controller (MPC) that provides an adequate propofol infusion rate for a given population
while being robust to the inter-patient variability and the resulting mismatch between
the prediction model and the patient response. An attraction of MPC is that it provides
the ability to control both hypnosis and analgesia in an uncomplicated manner, with
constraints on drug infusion rates and system states. The novel component of this design
is the systematic controller tuning and the inclusion of robustness and performance
analysis in the controller design.
Method: MPC requires a process model to predict the future output values. A nominal model
was constructed from the individual frequency responses of 44 adult subjects previously
reported. The uncertainty with respect to this nominal model was quantified and robustness and performance measures were included in the controller design. A 60-minute surgical procedure was simulated for the complete set of 44 models assuming an infusion
of 10 mg/ml propofol. The WAVCNS index provided by the NeuroSENSE Monitor (NeuroWave Systems, Cleveland Heights, USA) was considered as the clinical endpoint. The
target depth of hypnosis was set to a WAVcns of 50 for the duration of the procedure.
For all patients, a hypothetical surgical stimulus was fixed to start 20min after the beginning of the simulation. The controller tuning was based on the robust stability and robust
nominal performance measures as well as the predicted time domain performance.
Result: Typical performance measures including rise time and overshoot are calculated according
to the definitions from a previous study. Comparing the results of the MPC controller and
a robustly tuned PID controller, reveals that the MPC controller has less overall oscillations for the same population. The MPC controller attains this, while outperforming the
PID controller in terms of rise time 5.4 min and 7.87min and overshoot 1.8 % and 3.3%,
respectively.
Conclusion: The proposed MPC controller can provide adequate and stable depth of hypnosis during
simulated induction and maintenance of anesthesia and achieves robustness against patient uncertainty. Future work includes extending the proposed method to a constrained
robust MPC controller as well as multivariable control of hypnosis and analgesia.
70
Trainee: Chris Laugen
Board #: 49
Session #: 6
Supervisor: Patricia Janssen
Department: School of Population & Public Health
Title: Social support and exclusive breast feeding among Canadian women
Author(s): Chris Laugen, Patricia Janssen
ABSTRACT:
Evidence suggests that exclusive breastfeeding is the most effective feeding choice to prevent infant
morbidity and mortality. The WHO recommendation for exclusive breastfeeding for the first six months
of life has now been adopted by Health Canada, the Canadian Pediatric Society, Dietitians of Canada,
and the Breastfeeding Committee for Canada. While 87% of mothers in Canada initiate breastfeeding,
less than 14% exclusively breastfeed to six months.
This study examines whether or not greater social support may help Canadian mothers to achieve
exclusive breastfeeding for six months. Data was obtained from the 2009-2010 Canadian Community
Health Survey (CCHS). The analytic sample was limited to mothers who gave birth in the five years
prior to the survey, responded when asked whether or not exclusive breastfeeding for six months was
achieved, and answered the module on social support (n=2141). Social support measures included
four scales based on the Medical Outcomes Study. Bivariable and multivariable logistic models were
constructed for exclusive breastfeeding and each social support scale, adjusting for age, marital status,
education, household income, and employment. Odds ratios are reported with 95% confidence intervals.
Approximately 26% of surveyed mothers responded as having exclusively breastfed for 6 months. In
the multivariable adjusted models, all four social support scales resulted in increased odds for exclusive
breastfeeding. The greatest odds of meeting guidelines for exclusive breastfeeding were among women
with higher scores on the affection support scale (OR 1.22, 95% CI 0.84 – 1.79) and the emotional and
information support scale (OR 1.29, 95% CI 0.76 – 2.17). Married women with more education also had
higher odds of exclusively breastfeeding according to the guidelines. Many studies have examined why
women fail to exclusively breastfeed, but fewer studies identify how women achieve this goal. The
effects of all four types of social support measured in the current analysis are positive, though modest,
for achieving exclusive breastfeeding for six months. Future research can look more closely at what kind
of support women receive who successfully achieve the Canadian guidelines for exclusive breastfeeding.
71
Trainee: Paul Sabatini
Board #: 50
Session #: 6
Supervisor: Francis Lynn
Department:Surgery
Title: Npas4 is a novel, activity regulated, cytoprotective factor in pancreatic beta cells
Author(s): Paul Sabatini, Francis Lynn
ABSTRACT:
Diabetes is a disease characterized by chronically elevated blood glucose levels and is caused by the
failure of pancreatic beta cells. In order to design novel therapeutics, a better understanding of factors
that prevent beta cell death is needed. Npas4 is a basic helix-loop-helix, PAS domain transcription factor found in pancreatic beta cells which, to date has only been studied in neurons, it is our objective to
examine how Npas4 is regulated in the beta cell as well as what effects it has on beta cell function and
survival. Within Beta cells, Npas4 is expression is highly dynamic, being rapidly and dramatically induced
at the message and protein level following exposure to high glucose in isolated mouse and human islets
as well as in rat islets, in vivo. To study the effect of increased Npas4 on beta cell function we made
use of an adenovirus which strongly induces npas4 expression at the message and protein level. When
Npas4 is overexpressed in either MIN6 cells or islets, there is a dramatic decrease in the expression of
both insulin 1 and insulin 2. This inhibition is due to decreases in Pdx-1 and Neurod1 protein expression
as well as Npas4 directly binding to the insulin promoters. Hypothesizing that this decrease in insulin
expression may ease endoplasmic reticulum burden and allow beta cells to better respond to ER stress,
we examined the ability of Npas4 to mitigate ER stress. Interestingly, Npas4 expression is significantly
induced in response to classical ER stressors thapsigargin and palmitate. If overexpressed in either MIN6
cells or whole mouse islets, Npas4 significantly reduces the induction of Ddit3 caused by both thapsigargin and palmitate (CHOP) as well as thapsigargin mediated cell death. Our future studies will examine
the protective functions of Npas4 in vivo.
72
Trainee: Ashish Sharma
Board #: 51
Session #: 6
Title: Genetics of IL-1β responses in humans
Author(s): Ashish Arunkumar Sharma, Kelsey Lee, Bernard Kan, Pascal M Lavoie
ABSTRACT:
Interleukin 1β (IL-1β) is a key modulator of inflammation in humans. An excess of IL-1β has been
implicated in several human autoimmune and inflammatory diseases such as type 1 diabetes,
inflammatory bowel diseases and rheumatoid arthritis. On the other hand, a lack IL-1β underlies a
greatest risk of infection. Due to its critical role in regulating inflammation, IL-1β responses require a tight
regulation by a dual mechanism: Initially, stimulation of pathogen recognition receptors (PRRs, e.g. TLR4)
by pathogen associated molecular patterns (PAMPs, e.g. LPS) results in transcription of the IL1B gene
and translation into its precursor protein form (pro-IL-1β). Pro-IL-1β is cleaved/activated by the NLRP3/
caspase-1 inflammasome, upon the recognition of danger associated molecular patterns (DAMPs, e.g.
ATP). Susceptibility to infection or inflammatory diseases in humans has been attributed to genetic
variations within the IL-1β pathway. However, the contribution of common genetic variants to individuals’
responses in the general population is poorly understood. We hypothesize that common genetic variants
modulate IL-1β responses in humans.
Our lab has developed robust, standardized methods to quantify the true biological variability within
rate-limiting steps along the IL-1β secretion pathway. Using these assays, we analyzed the IL-1β response
to LPS and ATP in 24 healthy adults twice within a 1-month period, in order to estimate the degree of
experimental versus biological variability. Expression of responses along the IL-1β pathway is highly
variable, yet they are similar over time in a given individual. The amount of IL-1β secreted is primarily
determined by the proportion of monocytes and by the extent of caspase-1 activation within peripheral
blood mononuclear cells. Moreover, we detect significantly greater variability within women, whereas
men are more stable over time. Our immediate plans is to expand this cohort to conduct a
whole-genome association study to identify single nucleotide polymorphisms associated with ratelimiting components of the IL-1β responses and to confirm that these SNPs significantly impact gene
expression. We expect that our study will provide a first understanding of the importance of SNPs in
determining susceptibility to IL-1β-mediated diseases in humans.
73
Trainee: Sophie Stukas
Board #: 52
Session #: 6
Title: Intravenously injected human apoA-I rapidly enters the central nervous system in mice
Author(s): Sophie Stukas, Michael Lee, Michael Carr, Nicole DeValle, Kaistyn Lemke, Jianjia Fan,
Dhananjay Namjoshi, Michael Oda, Cheryl Wellington
ABSTRACT:
Epidemiological evidence strongly suggests that mid-life “vascular risk factors” such as cardiovascular
disease, type II diabetes, hypertension, and hypercholesterolemia increase the risk for dementia, stroke,
and cerebral haemorrhage later in life. As many of these risk factors are also characterized by alterations
in plasma high-density lipoprotein (HDL; “good cholesterol”) metabolism, we hypothesize that HDL is a
physiological link to the central nervous system (CNS). Apolipoprotein A-I (apoA-I), the primary protein
component of plasma HDL, has potent anti-atherogenic, anti-inflammatory and anti-oxidant properties
and also protects the vascular endothelium.
Apo-I is made in only the liver and intestine and is not synthesized within the CNS. Nevertheless, apoA-I
is abundant in both cerebrospinal fluid (CSF) and brain tissue. Both in vitro and in vivo evidence suggest
that HDL can maintain cerebrovascular health, attenuate neuroinflammation and preserve cognitive function. However, very little is known about how apoA-I enters the CNS. Here we show that human apoA-I
(hapoA-I) delivered via intravenous injection, rapidly enters the CNS of mice in both a time and concentration dependent manner. Imaging studies show that hapoA-I is rapidly localized to cerebral ventricles
and to a lesser extent the luminal side of cerebral blood vessels. Confocal microscopy confirms uptake
of hApoA-I by epithelial cells of the choroid plexus. CSF and brain levels of hApoA-I directly correlate to
circulating plasma hapoA-I levels, ranging from 0.05 – 0.2% of plasma levels or ~0.2-0.8 ug/mL. Over the
course of 4h, hApoA-I levels in plasma decrease significantly but remain relatively constant in CSF and
brain lysates, suggesting decreased catabolism in the CNS. These results support the hypothesis that
apoA-I is able to cross the blood-CSF and possibly also the blood-brain-barrier and may aid in maintaining cerebral vascular endothelial health, thereby preserving neuronal function.
74
Trainee: Bryan Tennant
Board #: 53
Session #: 6
Supervisor: Brad Hoffman
Department:Surgery
Title: The transcription factor Myt3 acts as a pro-survival factor in β-cells
Author(s): Bryan R Tennant, Ratib Islam, Marabeth M Kramer, Yulia Merkulova, Roger L Kiang,
Cheryl J Whiting, Brad G Hoffman
ABSTRACT:
The increasing prevalence of diabetes worldwide, the paucity of islet donors and the obvious short-comings in current treatment options lead to a necessity to find alternate sources of functional β-cells. The
identification of novel factors that are expressed specifically in the pancreas, and a greater understanding
of the transcriptional networks regulating β-cell function and survival is vital to the improvement of current,
and the development of novel, therapeutic options. To this end we previously identified the transcription
factor Myt3 as specifically expressed in pancreas. Here, we sought to determine the expression and regulation of Myt3 in islets and to determine its significance in regulating islet function and survival. We show
that Myt3 is the most abundant MYT family member in adult islets and that Myt3 is expressed in all the major endocrine cell types in the pancreas after E18.5. Further, we determine that Myt3 expression is sensitive
to both glucose and cytokine exposure. Of specific interest, suppressing Myt3 expression reduces insulin
content and increases β-cell apoptosis, at least in part, due to reduced Pdx1, Mafa, Il-6, Bcl-xl, and c-Iap2
levels, while over-expression of Myt3 protects islets from cytokine induced apoptosis. These data are an
important step in clarifying the regulatory networks responsible for β-cell survival, and point to Myt3 as a
potential therapeutic target for improving islet graft survival and functional β-cell mass.
75
Trainee: Allison Watts
Board #: 54
Session #: 6
Title: Patterns of dietary intake among overweight and obese adolescents and their parents
Author(s): Allison Watts, Louise Mâsse, Chris Lovato, Susan Barr, Rhona Hanning
ABSTRACT:
Purpose:
Few studies have compared parent-child dietary intake among adolescents who are
overweight or obese. The purpose of this study was to determine the relationship
between parent and adolescent intake of vegetables and fruits, fat, and sugar-sweetened beverages.
Method: Complete dietary data was collected from 173 parent and adolescent (11-16yrs) pairs
who presented for a lifestyle behaviour change intervention. Baseline dietary intake
was assessed from 3 web-based 24h diet recalls. Parent and adolescent intake of three
dietary components were examined: servings of vegetables & fruits (VF), percent energy
from fat (% fat), and servings of sugar-sweetened beverages (SSB). Multivariable
regression models were used to identify associations between parent-child dietary
variables. Analyses were adjusted for parent BMI, age, sex, education, and child age
and sex.
Result: Adolescents consumed 3.3 servings of VF per day, 33% of their energy was from fat,
and 54% reported consuming SSB, with an average of about 1 serving per day. Parents
consumed slightly more VF (3.7) and fat (35%), but less SSB (31%) than did adolescents.
Parent intake of VF (?=0.17, p=.02), % fat (?=0.25, p=.003) and SSB (OR=3.24, 95% CI
1.56-6.73) were positively associated with their adolescent’s intake of these same dietary
components. The relationship between parent-adolescent fat intake was stronger among
families with less parental education.
Conclusion: Parent’s intake of several dietary components important for good health predicted
adolescent’s intake. Targeting parents, by helping them improve their own diet, may
promote improvements in their adolescent’s diet.
76
POSTER SESSION SEVEN
doctoral students
MODERATOR:
PARTICIPANTS: Gregor Reid
Guilaine Boyce
Dominika Nackiewicz
Wai Hang Cheng
Nicole Krentz
Phoebe Lu
Ashish Marwaha
Cecil Ming Yeung Chau
Tuan-Anh Nguyen
Anna Poon
DOCTORAL STUDENTS | POSTER SESSION SEVEN
Trainee: Guilaine Boyce
Board #: 55
Session #: 7
Title: Development of Th17 immunity in human newborns
Author(s): Guilaine K Boyce, Ashish A Sharma, Pascal M Lavoie
ABSTRACT:
Prematurity and infection account for over two thirds of neonatal deaths worldwide. Neonates possess
an immature immune system, making them more susceptible to infection. This applies particularly to
preterm infants. The immune system provides targeted antimicrobial protection through the
differentiation of T helper (Th) lymphocytes into subsets optimized to combat specific types of
pathogens. The Th17 responses play an important role in mucosal immunity against fungal and
extracellular bacterial pathogens, the main causes of leading neonatal infections, especially in
prematurely born infants. We previously reported a marked attenuation of Toll-like receptor-induced
interleukin (IL)-1beta, IL-6 and IL-23 production in newborns of decreasing gestational age; these
cytokines are critical for antigen-driven differentiation of Th17 responses. According to recent data cord
blood T cells are able to differentiate into IL-17 producing (Th17) cells when exposed to these cytokines.
We hypothesize that the developmental lack of Th17-differentiation in neonates is primarily due to a
poor Th17-inductive ability of neonatal mononuclear cells (MCs). According to our experiments, cord
blood CD4+/CD8+ T cells lack most signature Th17 cytokines (IL-17A, IL-17F, IL-21 and IL-22)
following PMA/ionomycin stimulation, which is consistent with a naïve undifferentiated state. Interestingly, we observed a predominance of IL-17F production within CD4-CD8- cells. Preliminary data show that
IL-17A production in cord blood CD3+ cells is largely detected among γσ T cells in contrast to adults in
which conventional CD4+ T cells appear as the main sources of Th17 cytokines. Collectively, our findings
suggest that Th17 responses in newborns may be driven by immune cells other than conventional γσ
T cells. Further investigation to determine the contribution of γσ T cells and the functional capacity of
neonatal immune cells to drive a Th17 response will provide valuable insight into the mechanisms
regulating human mucosal immunity in early life.
79
Trainee: Dominika Nackiewicz
Board #: 56
Session #: 7
Supervisor: Jan Ehses
Department:Surgery
Title: Islet cell type specific regulation of inflammation and beta cell function by TLR2/6 and
TLR4 ligands
Author(s): Dominika Nackiewicz, Meixia Dan, Madeleine Speck, Anisa Salmi, Amanda Cunningham,
Nico van Rooijen, Beatrice Guardiola, Carole Schuster-Klein, Jan A Ehses
ABSTRACT:
Chronic inflammation contributes to pancreatic beta cell dysfunction in type 2 diabetes. Both toll-like
receptor (TLR)-2 and -4 ligands are increased in the circulation of recently diagnosed type 2 diabetes
patients, and both TLR2 and TLR4 deficient mice are protected from the metabolic consequences of a
high fat diet. Thus, we analyzed murine wild type islets, TLR2-/- islets, TLR4-/- islets, purified rat beta
cells and human islets to investigate the cell types responsible for TLR2/6- and TLR4-receptor mediated
effects on islet inflammation and beta cell function. First, we depleted resident islet macrophages from
mouse and human islets and sorted them out by flow cytometry (FACS) for gene expression analysis. We
showed that resident islet macrophages contributed to TLR2/6- and TLR4-induced islet cytokine/chemokine expression. However, TLR ligands also reduced insulin gene expression independently of resident
islet macrophages, and via indirect actions on beta cells. Next, effects of TLR2/6 and TLR4-actived bone
marrow derived macrophages (BMDMs) on beta cell function were determined. TLR ligand activated
BMDMs reduced islet insulin secretion and Ins1/2 and Pdx1 mRNA expression. To inhibit TLR ligand
effects a TLR2 neutralizing antibody and a TLR4 small molecule inhibitor, TAK-242 were used. TLR
neutralization or inhibition reversed ligand effects on islet inflammation and beta cell gene expression.
We conclude that while the effects of TLR2/6 and TLR4 signalling on insulin mRNA can be macrophage
independent, resident islet macrophages are major contributors to islet inflammation in response to
TLR2/6 and TLR4 ligands and promote chemokine-induced attraction of circulating monocytes that
further impair beta cell function. Future work will focus on the role of islet endothelial cells on insulin
mRNA regulation in response to TLR2/6 and TLR4 ligands.
80
Trainee: Wai Hang Cheng
Board #: 57
Session #: 7
Title: Toward developing relevant models of mild traumatic brain injury
Author(s): Wai Hang Cheng, Dhananjay Namjoshi, Kurt McInnes, Peter Cripton, Cheryl Wellington
ABSTRACT:
Traumatic brain injury (TBI) is the leading cause of death and disability in developed countries for people
under 45 years of age. Mild TBI (mTBI), or concussion, is common in contact sports such as football and
hockey. Although traditionally considered relatively benign, recent studies show that mTBI may have
significant long-term consequences. For example, TBI is one of the strongest environmental risk factors
for dementia. Therefore, developing relevant experimental models of mTBI is of increasing importance.
MTBI does not involve skull penetration and direct destruction of brain tissue. Rather, sudden
acceleration or deceleration of the brain within the bony skull is responsible for coup-contracoup
contusions and diffuse axonal injury (DAI) due to uneven shear-strain effects experienced by grey vs
white matter. Our laboratories are investigating two mTBI models; the weight drop (WD) model and
the supine impactor (SI) model. The WD model, which impacts the skull with a mass under gravitational
acceleration, limits head movement during impact. In contrast, the SI model allows free movement of
the head during impact, and may thus result in more severe DAI. In this study, we intend to compare the
behavioral and histological effects of mTBI induced by the WD and SI models.
Young adult B/6 mice will be used for WD or SI-induced mTBI at similar impact energy. In both
models, mice will receive two TBI at 24 hours apart. At day 1, 2 and 7 post-injury, the Rotarod test will
be performed to evaluate motor performance. At day 7 post-injury, Novel Object Recognition test will
be performed to assess object memory. Mice will then be sacrificed for histological analysis of neuronal
damage, microglia activation and disruption of axonal functions. As the WD model restricts head
movement during impact, we predict that the SI model may better resemble clinical cases, and induce
more severe neuronal and axonal damages, and resulting in greater memory deficits. This study may
shed light on how biomechanical parameters influence mTBI outcomes, and may assist in translating
findings from mTBI studies from bench to bedside.
81
Trainee: Nicole Krentz
Board #: 58
Session #: 7
Supervisor: Francis Lynn
Department:Surgery
Title: Investigating the cell cycle of multipotent progenitor cells during pancreatic development
Author(s): Nicole A J Krentz, Francis C Lynn
ABSTRACT:
Background:
Diabetes mellitus is a metabolic syndrome characterized by the inability to properly
maintain blood glucose levels due to a loss or dysfunction of insulin-producing pancreatic
beta cells. Future regenerative medicine strategies for diabetes treatment may involve
the use of differentiated human embryonic stem cells (hESC) as an unlimited source of
pancreatic beta cells. Current differentiation protocols have been unable to produce
functional beta cells in vitro; thus there is a need for a greater understanding of
pancreatic development. The Cpa1+ multipotent progenitor gives rise to all three cell
types of the pancreas, exocrine, endocrine and ductal cells, early in development before
becoming restricted to only the exocrine lineage by E13.5. To date, the mechanism
behind the restriction in differentiation potential of these multipotent progenitors is
unknown. Previous work in our lab suggests that lengthening the G1 phase of the cell
cycle can promote progenitor cell differentiation. We hypothesize that a potential
mechanism the drives the differentiation potential of these multipotent progenitors is the
length of the G1-phase and that by manipulating this length we may be able to improve
current hESC differentiation protocols.
Objective: To characterize the cell cycle of Cpa1+ multipotent progenitor cells and to determine if
G1 lengthening in Cpa1+ cells is responsible for their change in differentiation potential.
Method: We will use the fluorescent, ubiquitination-based cell cycle indicator (FUCCI) mice that
express the orange fluorescent protein, mKO2, in the G1-phase and the green fluorescent protein, mAG, in the S/G2/M-phases of the cell cycle to investigate the parameters
of the cell cycle during pancreatic development. In parallel, we will also use the CD1
mouse and cumulative EdU labeling to determine the total cell cycle length, as well as
the lengths of the G1-, S-, G2-, and M-phases of the cell cycle in the Cpa1+ multipotent
progenitor.
Result: FUCCI mice express orange and green fluorescent proteins in a cell cycle dependent
manner that is easily detectable and quantifiable by confocal and flow cytometry.
Cumulative EdU labeling, a method that has been successful for quantification of the cell
cycle during neurogenesis, is a suitable methodology to investigate the cell cycle during
pancreatic development.
Conclusion: Using two mouse models, FUCCI and CD1, we will be able to quantify the length of the
G1-, S-, G2-, and M-phases of the cell cycle as well as total cell cycle length of the Cpa1+
multipotent progenitor. We suggest that the differentiation potential of these
multipotent progenitors during development is dependent on the length of the cell cycle
parameters.
82
Trainee: Phoebe Lu
Board #: 59
Session #: 7
Title: Cooperative activities of the Asf1 histone chaperone and the SWR1-C complex in the
regulation of heterochromatin-proximal regions
Author(s): Phoebe Y T Lu, Michael S Kobor
ABSTRACT:
Post-translational modification of histones, nucleosome movement by ATP-dependent chromatin
remodeling complexes, and replacement of canonical histones by histone variants are the three main
mechanisms that contribute to the dynamic nature of chromatin structure in yeast. SWR1-C and NuA4
are two chromatin-modifying complexes that have a tight, functional relationship in the cell. They act
sequentially in regulating histone variant H2A.Z biology to create specific chromatin neighbourhoods.
SWR1-C is an ATPase dependent chromatin remodeler that exchanges H2A.Z/H2B for H2A/H2B in the
chromatin; Whereas NuA4 is a histone acetyltransferase that acetylate H4, H2A, H2A.Z as well as other
non-histone substrates. SWR1-C and NuA4 share four subunits that may play a central role in targeting
sites of H2A.Z deposition and acetylation. The only non-essential shared subunit is the YEATS domain
containing protein, Yaf9. In budding yeast, the closest structural relative to the Yaf9 YEATS domain is
Asf1, a highly conserved histone chaperone for H3/H4 heterodimer.
Previously, we found that the growth of cells lacking both ASF1 and YAF9 is significantly compromised
compared to either gene deletion alone, indicating that these two proteins interact genetically and act
in parallel pathways in the cell. In this study, we found that the synergistic interaction between YAF9 and
ASF1 was a consequence of Yaf9’s contribution to both H2A.Z deposition and NuA4 acetyltransferase
activity. Further analysis revealed that this interaction was mediated by Asf1’s ability to regulate H3K56
acetylation by Rtt109. To better understand the mechanism underlying the genetic interaction between
ASF1 and YAF9, we thought to elucidate their activities at the well-characterized HMR locus and at the
telomeric region on the right arm of chromosome III. Interestingly, we found that both ASF1 and YAF9
are required to prevent silencing around the HMR locus whereas only YAF9 was required for silencing at
the telomere. We also determined that H2A.Z enrichment at heterochromatin boundaries depended on
ASF1 or YAF9. Presumably, the repressive effects seen at subtelomeric genes were due to the failure of
H2A.Z nucleosomes to restrict the spread of silencing factors. Even though loss of ASF1 leads to a
down-regulation of gene expression around the HMR, this observation was not consistent with the
spread of SIR complex proteins. Taken together, our results suggest that Asf1 and SWR1-C cooperate
together at the HMR boundary to regulate H2A.Z biology, but have distinct functions at the telomere.
83
Trainee: Ashish Marwaha
Board #: 60
Session #: 7
Title: Tregs from T1D subjects with a susceptible IL-2R gene SNP respond aberrantly to
IL-2 stimulation
Author(s): Ashish Marwaha, Sze Wing Wong, Spencer Staiger, Aaron Hirschfield, John Priatel,
Constadina Panagiotopoulos, Stuart Turvey, Jane Buckner, Rusung Tan
ABSTRACT:
Regulatory T cells (Tregs) require the cytokine interleukin-2 (IL-2) for survival and maintenance of their
suppressive function. IL-2 binds to the high-affinity IL-2 receptor (IL-2R), leading to phosphorylation of
signal transducer and activator of transcription 5 (STAT5) and downstream transcription of effector
proteins. Activation of the STAT5 signaling pathway is required to maintain high expression of the
transcription factor FOXP3, which sustains the suppressive function of Tregs. The IL-2 receptor gene
has single nucleotide polymorphisms (SNPs), which are strongly associated with Type 1 diabetes (T1D)
incidence.
We hypothesized that T1D subjects with SNPs in the IL-2R gene possess Tregs that respond aberrantly
to IL-2 and therefore do not optimally induce signaling in the STAT5 pathway. PBMC from T1D subjects
with a homozygous susceptible (GG), heterozygous (AG) and homozygous wild-type (AA) genotype for
a SNP located in the rs3118470 locus of the IL-2R gene were stimulated with IL-2, prior to fixation and
staining for CD4, FOXP3, CD25 and pSTAT5. Our results indicate that Tregs from individuals with the
homozygous susceptible GG genotype have reduced levels of pSTAT5 compared to those with the AA
genotype (p=0.0133). This provides evidence that Tregs from T1D subjects with SNPs in the rs3118470
locus of the IL-2R gene have diminished STAT5 phosphorylation in response to IL-2. This is a potential
mechanism of defective Treg function in a genetically identifiable subset of children with T1D.
84
Trainee: Cecil Ming Yeung Chau
Board #: 61
Session #: 7
Supervisor: Ruth Grunau
Title: Neonatal pain-related stress interacts with 5HTTLPR and COMT Rs4680 genotype to
predict internalizing behavior in girls born very preterm
Author(s): Cecil MY Chau, Ivan L Cepeda, Tim F Oberlander, Angela M Devlin, Anne R Synnes,
Ruth E Grunau
ABSTRACT:
Background:
Risk for developing depressive and anxiety symptoms (internalizing behavior) is related
to serotonin transporter (5HTTLPR) and catechol-O-methyltransferase (COMT)
genotypes. Internalizing behaviors are prevalent in children born very preterm, and are
associated with greater exposure to neonatal procedural pain-related stress. However,
the role of these gene variants in interaction with neonatal pain-related stress and
concurrent maternal stress is unknown.
Objective: To examine whether the 5HTTLPR and COMT rs4680 variants interact with neonatal
pain-related stress and concurrent maternal stress to predict internalizing behavior at
age 7 years in children born very preterm.
Design/
N=100 children born ≤32 weeks gestation, (50 girls, 50 boys) were seen at mean age
Method:
7.7 years (SD=.31). Excluded: IVH 3/4, PVL, motor/sensory/intellectual impairment/
autism. From prospective neonatal chart review from birth to term equivalent,
pain-related stress was calculated as the number of skin-breaking procedures (adjusted
for confounders). 5HTTLPR and COMT rs4680 were genotyped by real-time PCR of
saliva genomic DNA [COMT (Val/Val=.29; Val/Met=.5; Met/Met=.21); 5HTTLPR (ll=.23;
ls=.57; ss=.20)]. Mothers completed Child Behavioral Check List (CBCL), Parenting Stress
Index (PSI), Beck Depression Index (BDI-2). The association between genotypes, number
of skin-breaks, concurrent maternal parenting stress and depressive symptoms, in
relation to child internalizing was examined with generalized linear modeling,
adjusting for neonatal clinical confounders (gestational age, SNAP-II at day 1, number
of days on mechanical ventilation, number of surgeries, morphine exposure adjusted for
daily weight).
Result: Greater neonatal pain-related stress predicted higher internalizing in girls with COMT
rs4680 Val/Val and Val/Met genotypes (p=.02). In contrast, greater concurrent maternal
parenting stress predicted higher internalizing in girls with at least one 5HTTLPR short
allele (p=.006). Boys n.s.
85
Trainee: Tuan-Anh Nguyen
Board #: 62
Session #: 7
Supervisor: Dan Rurak
Department: Obstetrics & Gynaecology
Title: Postnatal outcomes in lambs exposed antenatally to fluoxetine
Author(s): Tuan-Anh T Nguyen, Timothy Chow, Wayne Riggs, Dan Rurak
ABSTRACT:
Introduction: Studies have demonstrated Selective Serotonin Reuptake Inhibitors (SSRIs) cause
changes in fetal behavioral states and neuroendocrine function, alteration of fetal middle
cerebral artery flow characteristics and blunted fetal heart rate variability (HRV). Yet,
the long-term effects of prenatal SSRIs exposure on neurobehavioral development are
uncertain. In this study, we investigated the postnatal outcomes in lambs exposed to
fluoxetine (FX) during late gestation.
Material and Daily 50mg FX were given to pregnant ewes (N=5) via implanted catheters for 14 days
Method: or until delivery.
FX exposure averaged the last 12 ± 1.3 days of gestation. Sleep-activity cycle and
neonatal behaviors were determined by actiwatch and continuous digital video
recording (DVR). Birth weight, daily weight, heart rate and tissue sPo2 were obtained.
HRV and FX were measured in lambs on postnatal day 2, 5, 10, 14. FX was also
measured in ewes and lambs at birth. Activities and HRV were analyzed by two-way
ANOVA repeated measures with Bonferroni’s correction. Differences in other variables
were compared by t-test (P< 0.05).
Result: Lamb:ewe [FX] against the time from the last FX dose in the ewes (R? = 0.4384)
declined logarithmically. No significant differences in birth weight, daily weight gain,
heart rate and sPo2 were found between control and FX-exposed lambs. However, FX
lambs (N=7) were more active than the control lambs (N=7) indicated by significantly
faster first time activities (standing-walking-suckling) and by an increase in subsequent
activities obtained from actiwatches and DRV observations from postnatal day 4 to
postnatal day 14. Interestingly, the increased activity in FX-exposed lambs was not due
to increased suckling bouts but to increased non-suckling bouts. HRV on postnatal day
2 were significantly lower in FX lambs vs control.
Conclusion: Prenatal FX exposure appears to have behavioral and cardiovascular effects in the
offspring beyond birth. Behavioral effects still persisted long after FX level were
undetectable. This supports a mechanism of SSRIs-induced alteration in fetal brain
development in human infants exposed to the drugs in utero.
86
Trainee: Anna Poon
Board #: 63
Session #: 7
Supervisor: Daniel Goldowitz
Title: Genetics underlying neural progenitor cell proliferation in the adult mouse brain: Insights
from Quantitative Trait Locus (QTL) analyses
Author(s): Anna Poon, Daniel Goldowitz
ABSTRACT:
Adult neurogenesis is a potential reservoir for neuronal plasticity in the adult brain. The subgranular zone
(SGZ) in the dentate gyrus and the subventricular zone (SVZ)-rostral migratory stream (RMS) pathway to
the olfactory bulb are the sites of neuronal production. We sought to take a phenotype-driven approach
to define the genes that underlie adult neurogenesis.
In this study, we systematically quantified neural progenitor cell (NPC) proliferation in the RMS of a
diverse panel of mouse strains, including nine common inbred strains and two recombinant inbred (RI)
lines (i.e. 61 BXDs and 27 AXB/BXAs) using a 1 hour injection of bromodeoxyuridine (BrdU) prior to
sacrifice and tissue processing to demonstrate BrdU-positive cells. At 2 months of age in the inbred
strains, significant inter-strain differences are observed with a heritability estimated to be 0.58. The RMS
of RI lines had even wider differences in the numbers of proliferating cells, suggesting that individual
RI lines have inherited discrete genetic loci that underlie cell proliferation in the RMS. Quantitative trait
locus (QTL) mapping of the phenotypic differences revealed two significant QTL on Chr 11 and Chr 6
regulating NPC proliferation, in the AXB and BXD RI lines, respectively. Informatic analysis indicated a
subset of genes in the mapped QTL regions are associated with cell proliferation.
To determine whether the differences among the inbred strain persist at older ages, we extended our
proliferative survey of the RMS in the nine inbred mouse strains from 2 to 18 months of age. Significant
inter-strain differences are detected at 2 and 12 months, but not at 18 of age, as all strains had a low
level of proliferation at these ages.
In summary, our study demonstrates that the genetic diversity among the inbred strains is a rich resource
for unraveling genetic loci and the complex genetic architecture regulating NPC proliferation at different
ages.
87
POSTER SESSION EIGHT
postdoctoral Fellows
MODERATOR:
PARTICIPANTS:
Wendy Robinson
Jiadi Wen
Matthias Görges
Jacqueline Lai
Guobin Sun
Alice Wang
Hong Yang
Maryam Zarepour
POSTDOCTORAL FELLOWS | POSTER SESSION EIGHT
Trainee: Jiadi Wen
Board #: 64
Session #: 8
Supervisor: Evica Rajcan-Separovic
Title: Screening for and functional assessment of miscarriage CNVs
Author(s): Wen J , Hanna C W , Martell S, Leung P C K, Robinson W P, Stephenson M D,
Rajcan-Separovic E
ABSTRACT:
Detection of unique copy number variations (CNVs) in miscarriages suggests that their integral genes
have a role in maintaining early pregnancy. Previously, we identified 11 unique miscarriage CNVs in
~30% of studied miscarriages, which were predominantly familial in origin. Using qPCR, we have now
screened for copy number changes of 13 genes from these miscarriage CNVs in 250 couples with
recurrent miscarriage (RM). RM couples showed more frequent copy number changes in PNPLA4
(Xp22.31) (0.6% vs 0.3%) and STX6 (1q25.3) (0.2% vs 0%) compared to controls. For three genes (TIMP2,
OFD1 and TRAPP2C) the RNA and protein expression was altered in miscarriages that had CNVs, in
keeping with their genomic copy number changes. In depth analysis of a maternal duplication disrupting
the TIMP2 gene was performed in four miscarriages from one female with RM, due to the involvement of
TIMP2 in placental remodeling and embryo development, and the previous suggestion that it is
maternally expressed in placenta. Methylation of upstream promoter sites in the miscarriages and control
early pregnancy tissues, as well allelic expression levels, are consistent with preferential maternal
expression of the TIMP2 gene in early pregnancy; however, the regulation of its expression and
imprinting appears complex. Further functional studies of miscarriage CNVs may elucidate their role in
causing miscarriage.
91
Trainee: Matthias Görges
Board #: 65
Session #: 8
Supervisor: Mark Ansermino
Department: Anesthesia, Pharmacology & Therapeutics
Title: Stroke volume variability doesn’t predict cardiac output decrease when prone positioning
children for scoliosis surgery
Author(s): Matthias Görges, Zoe E Brown, Erin Cooke, Heng Gan, Guy A Dumont, J Mark Ansermino
ABSTRACT:
Background:
For a posterior surgical approach to scoliosis repair, patients are placed in the prone position.
This can be associated with immediate significant decreases in blood pressure [1]. Changes
in cardiac index when turning adult patients to the prone position for spinal surgery are well
documented [2, 3], whilst pilot data are available in children [4]. Targeted preload optimization, with the goal of achieving a stroke volume variation of less than 14%, was reported to
be effective in preventing significant cardiac index decreases induced by positioning adults
prone [5]. We investigated the ability of using stroke volume variability to predict cardiac index
changes in children during prone positioning in an observational study.
Method: Following REB approval and written informed parental consent, with subject assent where
appropriate, 29 ASA I-II patients, aged 12-18 years undergoing primary scoliosis repair, were
recruited into this observational study. A standardized total intravenous anesthetic technique
with propofol and sufentanil was used. In addition to routine monitoring, including an arterial
line, an age appropriate transesophageal doppler probe (CardioQ DP240 or CardioQ KDP72,
Deltex Medical Limited, West Sussex, UK) was positioned post induction of anesthesia [6, 7].
Cardiac index (CI) and stroke volume (SV) were recorded for a minute at three time points:
post induction of anesthesia, following a 5ml/kg crystalloid fluid bolus, and immediately after
the patient was turned prone. Data were plotted and analyzed using MATLAB (The Mathworks
Inc, Natick, MA).
Result: Stroke volume variability (SVV) after a 5ml/kg crystalloid fluid bolus and immediately prior to
positioning the patient prone, had a median of 12.7 % (range 5.3 to 36.9%), with 17/29 (59%)
having a SVV<14%. Prone positioning caused a median decrease in CI of 19.3% (range -10.3
to 40.9 %) and produced a median SVV of 18.0% (range 4.1 to 42.8 %). Both changes with
prone positioning are statistically significant (p=7.78x10-6, ?2=19.99 for SVV and p=1.2x10-3,
?2=10.51 for CI respectively). The CI drop in patients with SVV <14% did not significantly differ from patients with higher SVV (p=0.29, ?2=1.08). Figure 1 shows the temporal relationships
of CI and SVV for all 29 patients. There was no correlation between SVV and the change in
cardiac index (r2=0.047, see scatterplot in Figure 2).
Conclusion: Prone positioning produced significant drop in CI and increase in SVV. SVV before prone positioning does not predict the magnitude of CI change when positioning children prone, and minor fluid loading (5cc/kg) does not prevent prominent drops in CI, even when it reduces SVV
to less than 14% before prone positioning. Future work is needed to identify predictors of CI
change with prone positioning as well as strategies to mitigate the decrease in CI to improve
safety of positioning children prone.
References: [1] Can J Anaesth 2011;58:451-5; [2] Spine 2006;31:1388-93; [3] Acta Anaesthesiol Scand
1991;35:741-4; [4] Proc CAS Annual Meeting 2012; #1341754; [5] Anaesthesia 2012; doi:
10.1111/j.1365-2044.2012.07116.x; [6] Intensive Care Med 2001;27:201-5; [7] J Clin Monit
Comput 2008;22:299-307
92
Trainee: Jacqueline Lai
Board #: 66
Session #: 8
Supervisor: Jan Dutz
Department: Dermatology & Skin Sciences
Title: Microparticulate-encapsulated antigen with split topical CpG oligodeoxynucleotide
adjuvant as a single-injection immunization strategy
Author(s): Jacqueline Lai, Tullio Esposito, Urs Hafeli, Jan Dutz
ABSTRACT:
Antigen-presenting cells (APCs) are responsible for presenting foreign antigens to, and activating T cells
to fight off infection. Manipulation of APCs to modulate immune responses to immunization has been
a focus in vaccinology. Keratinocytes and skin-resident plasmacytoid dendritic cells can be activated via
binding of their Toll-like receptor 9 to CpG oligodeoxynucleotides (ODN) – synthetic DNA that mimic
unmethylated CpG bacterial DNA. Here, we encapsulated the ovalbumin (OVA) antigen in polylactide
co-glycolide microparticles (PLG-OVA) and show that mice immunized subcutaneously with PLG-OVA
generate antigen-specific cytotoxic T cells when CpG ODN is used as an adjuvant. Using flow
cytometric analysis, 2.3% antigen-specific cells are detected within the CD8+ cytotoxic T cell population
in the peripheral blood when CpG ODN is administered topically (P < 0.05 compared to PLG-OVA or
CpG ODN alone). In addition, 0.8% of CD8+ splenocytes produce interferon-gamma when re-stimulated
with the OVA (SIINFEKL) peptide in vitro (P < 0.001). The microparticles show a triphasic release of OVA
which simulates two doses of antigen injected separately. Using the release properties of PLG-OVA and
a second topical application of adjuvant, we enhance the generation of antigen-specific cytotoxic T cells
in the blood from 0.4 to 0.9% (P < 0.05) and OVA-specific total IgG and IgG2c production, effectively
generating a single-injection vaccine with split topical adjuvant administration that induces robust
cytotoxic T cell and antibody responses. Generating single-injection vaccines may enhance compliance
with vaccination in the general population and may also decrease the cost of administration.
93
Trainee: Guobin Sun
Board #: 67
Session #: 8
Title: The PTPα-BCAR3-Cas signaling complex promotes malignant properties of
rhabdomyosarcoma RD cells
Author(s): Sun G, Cheng SYS, Le HT, Pallen CJ
ABSTRACT:
The ubiquitously expressed scaffolding protein Cas has been implicated in promoting cell migration
and survival, and mediates tumor cell invasive motility by integrating signals from growth factor receptors and integrins. Recently we demonstrated that a novel PTPα-BCAR3-Cas complex promotes Cas
recruitment to focal adhesions (FAs) to enhance Cas tyrosine phosphorylation and downstream migration
signaling. The role of Cas in tumor cell invasion and migration is best established in breast cancer, where
Cas and BCAR3 are often overexpressed in association with enhanced metastasis and poor prognosis.
In this study, we translate our findings regarding PTP?-mediated regulation of Cas to investigate the
significance of this module in conferring cancer cell properties such as migration, survival, and
invasion. The rhabdomyosarcoma RD cell line, which we determined to have high expression of BCAR3
and Cas, was selected as a model system to investigate the role of the PTPα-BCAR3-Cas complex using
an approach of siRNA-mediated silencing of PTPα and/or BCAR3. We confirm that in RD cells the
formation of this complex is dependent on PTPα Tyr789 phosphorylation and the presence of BCAR3.
Depletion of PTPα or BCAR3 does not induce apoptosis in RD cells but significantly reduces migration
and invasion, suggesting that BCAR3-mediated PTPα-Cas association is required for RD cell migration
and invasion. Furthermore, expression of BCAR3 mutants which cannot interact with PTPα
phosphoTyr789 or Cas also inhibited RD cell invasion through dominant-negative effects. This study
reveals the upstream mechanisms that regulate Cas-mediated tumor progression and provides insight for
the development of molecular targeting strategies to inhibit Cas-dependent tumor metastasis.
94
Trainee: Alice Wang
Board #: 68
Session #: 8
Title: The C-terminal docking domain of the histone variant H2A.Z is required but not
sufficient for its functions
Author(s): Alice Wang, Michael S Kobor
ABSTRACT:
Eukaryotic chromosomes contain distinct neighbourhoods with unique properties and cellular functions.
These are specified by differences in the underlying chromatin structures. Nucleosomes are the f
undamental building blocks of chromatin, each consisting of 146bp of DNA wrapped around an
octameric histone core containing 2 copies each of H2A, H2B, H3 and H4. Several principal mechanisms
can alter the chromatin structure, including the replacement of canonical histones with histone variants.
Whereas canonical histones are deposited randomly into chromatin during DNA replication, histone
variants are deposited in a replication-independent manner by a class of specialized deposition
complexes in specific locations, thus creating distinct chromosome neighborhoods. One such variant,
H2A.Z, is conserved from yeast to humans and replaces canonical H2A in around 10% of nucleosomes,
predominantly in promoter and heterchromatic regions. H2A.Z has specific roles in gene regulation,
heterochromatin boundary formation, and genomic integrity. The precise features of H2A.Z required to
specify an identity different from canonical H2A remain to be fully explored. Here we explore the
difference between the C-terminal docking domain of the two histones and whether this region confers
H2A.Z-specific properties. We establish that the C-terminus is required but not sufficient for H2A.Z
functions in budding yeast Saccharomyces cerevisiae.
95
Trainee: Hong Yang
Board #: 69
Session #: 8
Supervisor: Stuart Turvey
Title: Amino acid structure determining the immune responses of peptide-gold nanoparticle
hybrids
Author(s): Hong Yang, Yingzhe Zhou, Shan-Yu Fung, Licun Wu, Tom Waddell, Tiago Machuca,
Kevin Tsai, Rusung Tan, Mark de Perrot, Stuart Turvey, Mingyao Liu
ABSTRACT:
Manipulation of the immune responses using nano-devices provides a new strategy to fight deadly
diseases and to improve current medical interventions. It has been shown that nanoparticles can boost
the antigen specific immune responses for vaccine development and can be used to improve the
existing immune suppressive therapies for transplantation and autoimmune diseases. Despite these
encouraging achievements, little is known systemically regarding the influence of the nano-devices on
the immune system. Thus, efforts should be made to better understand how the various nano-devices
interact with the immune system. In this study, we constructed a peptide-gold nanoparticle hybrid
system with tuneable surface chemistry (via different peptide ligands), and use it as a model nano–device
to address this question. We first studied the interaction between the hybrids and the murine bone
marrow derived dendritic cells (BMDCs). It was found that the hybrids did not affect the cell morphology
and viability; however, they significantly increased the expression of cytokines and co-stimulatory
molecules, indicating the activation of BMDCs. The activation was determined by peptide ligands with
single amino acid alteration. Secondly, these activated DCs can further enhance the T cell proliferation
in an allogenic mixed lymphocyte reaction assay. This work allows us to understand how immune cells
respond to nanoparticles with different surface moieties. More importantly, it may provide a principle for
utilizing nanoscale materials in immune modulation.
96
Trainee: Maryam Zarepour
Board #: 70
Session #: 8
Supervisor: Bruce Vallance
Title: Muc2 limits pathogen burdens and epithelial barrier dysfunction during Salmonella
Typhimurium colitis
Author(s): Maryam Zarepour, Kirandeep Bhullar, Marinieve Montero, Caixia Ma, Tina Haung,
Anna Velcich, Lijun Xia, Bruce A Vallance
ABSTRACT:
Salmonella Typhimurium is a model organism used to test the virulence factors involved in Salmonella
pathogenesis. To study Salmonella’s ability to cause intestinal inflammation, most groups use a colitis
model relying on streptomycin to displace intestinal commensal microbes, resulting in heavy pathogen
colonization of cecal tissues and severe inflammation. Although intestinal mucus is the first line of
defense in the mouse GI tract, its role in providing host defense against Salmonella is still unclear. The
mucus barrier is made up of the highly glycosylated mucin Muc2, which is secreted by goblet cells.
Glycosylation involves the actions of several enzymes, for example, Core 3- O derived glycans are
synthesized by Core 3 ?1,3-N-acetylglucosaminyltransferase (C3GnT). Mice lacking these glycans still
produce the Muc2 protein, but display a thinner mucus barrier, and show increased susceptibility to
chemical induced colitis.
By comparing Salmonella induced colitis and mucus dynamics in Muc2-/- mice, C3GnT-/- mice and
C57BL/6 mice, we observed that mucus secretion increased during infection in C3GnT-/- and C57BL/6,
with Salmonella found within the mucus layer. In contrast, Muc2-/- mice showed dramatic susceptibility to Salmonella infection, heavier cecal pathogen burdens and increased barrier disruption compared
with C57BL/6 mice. As a result, Muc2 -/- mice displayed high rates of morbidity and mortality. However,
C3GnT -/- mice, they carry WT pathogen burdens but developed exaggerated barrier disruption like
Muc2 -/- mice. These data suggest that the intestinal mucus layer plays a critical role in controlling
Salmonella intestinal burdens, whereas core-3 glycosylation plays an important role in controlling
intestinal epithelial barrier function.
97
POSTER SESSION NINE
postdoctoral Fellows
MODERATOR:
PARTICIPANTS:
Megan Levings
Bat-Chen Friedman
Anita Coté
Judith de Niet
Shan-Yu Fung
Klaske van Heusden
Whitney Weikum
POSTDOCTORAL FELLOWS | POSTER SESSION NINE
Trainee: Bat-Chen Friedman
Board #: 71
Session #: 9
Supervisor: Ran Goldman
Title: Viral etiology for acute wheezing episodes in children with high risk for subsequent
asthma
Author(s): Bat-Chen Friedman, Eva Thomas, Peter Tilley, Ran D Goldman
ABSTRACT:
Background:
Viral-induced wheezing episodes are common in pre-school children and results in
considerable distress and health care utilization. The aim of our study was to document
the viral etiology of acute wheezing episodes in young children and to evaluate the
virally-induced illness in children with or without predisposition for asthma.
Method: Children 6 to 36 months of age who presented to Emergency Department (ED) at
BC Children’s Hospital with episode of a viral illness and wheezing were prospectively
enrolled following parental consent. We excluded children with underlying chronic
diseases. Nasopharyngeal aspirates were tested using a multiplex reverse transcriptionPCR assay for the detection of 11 viral pathogens: influenza A/B, respiratory syncytial
virus (RSV), parainfluenza, enteroviruses, human rhinovirus (HRV), human metapneumovirus (HMPV), human coronaviruses, adenoviruses and bocavirus. Asthma predisposition
was calculated using a modified Asthma Predictive Index (API).
Result: We enrolled 116 children (mean age 15.5 months) from January 2011 to March 2012.
103(89%) were included in data analysis. 68(66%) were male, 16(16%) were premature
and 36(35%) had at least 3 wheezing episodes in the prior year. Duration of symptoms
was a mean of 3.5 days. All patients received beta-agonist inhalations, 71(72%) received
oral corticosteroids, 35(34%) had chest x-ray done and 16 (16%) were hospitalized. HRV
was detected in 39(38%) samples, RSV in 44(43%), Bocavirus in 13(13%), Parainfluenza
in 4(4%), HMPV in 4(4%), and Coronaviruses in 3(3%). Co-infection with two or more
pathogens was detected in 12(11%) of the samples; half of them involved HRV. In 6(6%)
no viral pathogen was detected. No significant differences were found in clinical
presentation between pathogens. Corticosteroids were more commonly used in children
with HRV infection (86% vs. 63% in non HRV). Fifty seven (56%) of children had a positive
API. There was a trend towards higher rate of HRV infection in children with a positive
API (41% vs 34%, p=NS).
Conclusion: HRV infection is a major cause of wheezing in young healthy children and account for
almost 40% of the patients seen in the ED for acute episode. Further research should
determine if HRV has a role in predicting asthma.
101
Trainee: Anita Coté
Board #: 72
Session #: 9
Title: Comt rs4680 variant and cardiometabolic side-effects in children treated with
second-generation antipsychotics
Author(s): Anita T Coté, Constadina Panagiotopoulos, Angela M Devlin
ABSTRACT:
Second-generation antipsychotics (SGA) are used to treat children for a wide-range of mental health
conditions but are associated with side effects such as rapid weight gain; increased waist circumference;
and elevated fasting plasma glucose, and blood pressure. However, not all children are susceptible to
these side-effects. The catechol-O-methyltransferase gene (COMT) is involved in metabolism and energy
homeostasis, and has been associated with weight gain in adults with mental health conditions. The aim
of this study is to investigate the interaction of the COMT V158M (rs4680) variant and SGA treatment
on cardiometabolic side-effects in a cross-sectional population of SGA-treated (n=147) and SGA–naïve (n=177) children. SGA-treated children had higher (P<0.05) BMI z-scores, fasting plasma glucose,
HOMA-IR, triglycerides, and total and LDL-cholesterol concentrations than SGA-naïve children. The
COMT rs4680 variant genotype frequencies were not different between SGA-treated (MM 24.2%, VM
50.5%, VV 25.3%) and SGA-naïve (MM 24.6%, VM 45.6%, VV 29.8%) children. An interaction between
SGA-treatment and COMT V158M genotype was observed such that MM genotype had higher SBP in
SGA-treated children, and VV genotype was associated with higher SBP in SGA-naïve children. COMT
VV genotype was associated with higher HDL cholesterol independent of SGA-treatment. No associations between COMT V158M and psychiatric diagnoses were found. These findings suggest that the
COMT V158M (rs4680) variant may predict children at risk for developing high blood pressure with SGA
treatment.
102
Trainee: Judith de Niet
Board #: 73
Session #: 9
Title: Adolescents’ intake of sugar-sweetened beverages and less healthful food in schools
Author(s): Judith E de Niet, Louise C Mâsse, Patti-Jean Naylor, Elizabeth Saewyc
ABSTRACT:
Purpose:
The school environment represents an important setting to address childhood
obesity. This study evaluated whether intake of sugar-sweetened beverages (SSB) and
less healthful foods was highest among adolescents who attended schools with less
healthy nutrition environments.
Method: Data from students (n=11,385) in grades 7-12 who completed the 2008 British
Columbia Adolescent Health Survey was linked to school principal data (n=174).
Adolescents’ intake assessed previous day consumption of SSB, fruits/vegetables, and
low-nutrient energy dense snacks (cookies, cakes, candies) and foods (pizza, hotdog,
French fries). A Food Consumption Index (FCI) was computed to denote overall
consumption of less healthful foods. Principals completed a survey measuring the school
nutrition environment including assessment of nutrition guidelines (schools and districts),
nutrition capacity/resources, participation in nutrition programs, and support for
adopting stringent nutrition guidelines. In addition, principals reported on the
availability of SSB and less healthful foods at school. Multi-level mixed-effects linear and
logic regression analyses with relevant covariates were used for the analyses.
Result: Adolescents in schools with increased availability of SSB and less nutrition guidelines
(e.g., fewer requirements for nutrition education and fewer guidelines for using foods as
rewards) were significantly more likely to consume SSB (p=.003). In contrast, the school
environment and availability of less healthful foods at school were not associated with
adolescent intake of less healthful foods.
Conclusion: Preliminary evidence suggests that the school environment is associated with adolescent
intake of SSB. The extent to which school policies may reduce intake of SSB remains to be
investigated.
103
Trainee: Shan-Yu Fung
Board #: 74
Session #: 9
Supervisor: Stuart Turvey
Title: A potent anti-inflammatory agent targeting Toll-Like receptor signaling identified from
marine sponge extracts
Author(s): Shan-Yu Fung, Vladimir Sofiyev, Julia Schneiderman, Aaron F Hirschfeld, Rachel E Victor, Kate Woods, Nicole J de Voogd, Raymond J Andersen, Stuart E Turvey
ABSTRACT:
Background:
While Toll-like receptors (TLRs) play a critical role in innate immunity, activation of TLR
signaling pathways is also associated with many harmful inflammatory diseases.
Identification of novel anti-inflammatory molecules targeting TLR signaling pathways is
anticipated to empower the development of new treatment approaches for acute and
chronic inflammation.
Method: We performed high throughput screening to isolate TLR inhibitors from crude
marine sponge extracts. The top hits were validated by looking at their cytotoxicity and
dose-dependent activity. The leading compound was identified after several cycles of
screening and validation. We then conducted structure-activity analysis of the leading
compound to potentially improve its bio-activity. In addition, we studied different TLR
signaling pathways with TLR reporter cells and the inhibitory mechanism of girolline by
Western Blotting. Its anti-inflammatory activity on human peripheral blood mononuclear
cells and macrophages was evaluated by looking at the cytokine production via ELISA.
Result: Through the unbiased screening and validation, we identified girolline as a potent
anti-inflammatory agent through inhibition of TLR signaling. We demonstrated that
girolline inhibits signaling through both MyD88-dependent and –independent TLRs
(i.e. TLR2, 3, 4, 5 and 7), in variety of relevant cell types, including human peripheral
blood mononuclear cells and macrophages. The molecular target of girolline was
determined to be downstream in the TLR signaling cascade, distal to TRAF6, but not
directly affecting the transcription factor NF-kappaB activation. In addition, the
structure-activity analysis indicated that the whole chemical structure of girolline is
necessary for its bio-activity.
Conclusion: Together these data identify the sponge natural product girolline as a potent
anti-inflammatory agent targeting TLR signaling.
104
Trainee: Klaske van Heusden
Board #: 75
Session #: 9
Supervisor: Guy Dumont
Department: Electrical & Computer Engineering
Title: Clinical evaluation of closed-loop administration of propofol guided by the NeuroSENSE monitor in children
Author(s): K van Heusden, G A Dumont, K Soltesz, C Petersen, N West, A Umedaly, J M Ansermino
ABSTRACT:
Introduction: In closed-loop administration of propofol, drug infusion is automatically adjusted based
on feedback of a measure of the clinical effect. This is expected to improve stability of
the depth of anesthesia, reduce drug overdosing and reduce the effect of interpatient
variability. The objective of this study was 1) to evaluate the clinical feasibility of closedloop delivery of propofol guided by the NeuroSENSE monitor during induction and
maintenance of anesthesia in children and 2) to assess the performance of a simple
robustly tuned proportional-integral-derivative (PID) controller during moderately
stimulating procedures.
Method: Following REB approval, and informed consent/assent, 45 children aged 7-17 (see Table
1), ASA I-II, requiring anesthesia for elective upper or lower gastrointestinal endoscopic
investigations were enrolled for evaluation of the closed-loop system. The system uses
feedback from the NeuroSENSE monitor that provides the WAVcns index as a measure
of the depth of hypnosis [2]. Propofol is delivered by an Alaris TIVA infusion pump. In
addition to the robust PID controller, the control system contains necessary safety layers
and alarms. After initial testing of a robust PID controller for children [3], the PID controller was retuned to improve speed of induction of anesthesia and the rate of response to
stimulation. Both induction and maintenance of anesthesia were closed-loop controlled.
A bolus of remifentanil (0.5 mcg/kg) was administered before the start of induction of
anesthesia, followed by a continuous infusion (0.03 mcg/kg/min).
Result: Two cases were excluded. Typical performance measures including induction time
(Tind) are calculated according to the definitions from [4], see Table 1. Blood plasma
concentrations (Cp) calculated using the Paedfusor model ([5], defined up to 16y) and
the Schnider model (17y, [6]) at Tind varied from 2.3 to 7.7 mcg/ml. Figure 1 shows the
observed interpatient PK/PD variability.
Conclusion: Robust PID control of propofol infusion guided by the NeuroSENSE monitor can automate induction of anesthesia with limited overshoot in children age 7-17y. It can provide
adequate and stable depth of hypnosis during maintenance of anesthesia in moderately
stimulating procedures, despite the large interpatient variability in PK/PD behavior in
children. Further investigations to improve the performance of the automated system
include the addition of control of analgesic agents and individualization of the control
strategy.
105
Trainee: Whitney Weikum
Board #: 76
Session #: 9
Supervisor: Tim Oberlander
Title: Impact of maternal depression and prenatal antidepressant exposure on infant
habituation
Author(s): Whitney M Weikum, Janet F Werker, Ruth E Grunau, Ursula Brain, Linda C Mayes,
Takao Hensch, Tim F Oberlander
ABSTRACT:
Background:
Depression during pregnancy is common and serotonin reuptake inhibitors (SRIs) are
increasingly prescribed to manage mood disturbances. Both exposure to maternal
depression and SRIs alter serotonin levels. Serotonin is central to attention regulation
and information processing. Rates of habituation are used to assess information
processing capacity in infancy.
Objective: Examine impact of depression and prenatal SRI exposure on infant habituation rates
during the first year of life. Design/Methods: At 3, 6 and 10 months, infants of mothers
with no depression symptoms (Non-exposed), depression symptoms with no
medications (Depressed-only) and depression symptoms and prescribed SRIs (SRI-exposed) completed infant-controlled habituation tasks. Maternal depression was defined
as Hamilton Rating Scale for Depression (HAMD) score >8. In our first cohort, at both 6
and 10 months, infants participated in an auditory habituation procedure (syllable /da/
repeated) and a visual habituation procedure (faces silently talking). To confirm a
developmental trajectory, a second younger cohort (3 or 6 months; n=64), tested on a
different habituation task (photos of faces) was examined.
Results: At 10 months (n=76) there was no difference (time to reach habituation criterion)
between the three groups. At 6 months (n=80), the time to reach the habituation
criterion was significantly shorter among Depressed-only infants on both tasks (p<.05),
even when controlling for third trimester maternal mood, and mood at the time of the
study. For the 3-6 month task, again, there was no difference between the Non-exposed
and SRI-exposed, but the Depressed-only showed faster habituation rates compared to
the SRI-exposed (p<.05).
Conclusions: At 3 and 6 months, postnatal maternal depression appears to decrease habituation
rates. While this might reflect improved information processing, it may also be nonoptimal attentional engagement. Despite similar postnatal levels of depression
symptoms, no difference was observed between the SRI-exposed and Non-exposed
infants at any age. This suggests that exposure to SRIs may ‘program’ a future infant
response that compensates for a postnatal environment with a depressed mother. Since
differences in the Depressed-only infants’ habituation rates disappear by 10 months,
this finding suggests that infants are particularly susceptible to their mother’s depressed
mood in early infancy.
106
POSTER SESSION TEN
subspecialty resident or Fellow
MODERATOR:
PARTICIPANTS:
Julie Bettinger
Mercedes Chan
Heng Gan
Anas Manouzi
Lana Shaiba
Tracy Tucker
Yuk Joseph Ting
SUBSPECIALTY RESIDENT OR FELLOW | POSTER SESSION TEN
Trainee: Mercedes Chan
Board #: 77
Session #: 10
Supervisor: Helen Foster
Title: Assessment of musculoskeletal abnormalities in children with mucopolysaccharidoses
using a simple musculoskeletal examination (paediatric gait, arms, legs and spine)
Author(s): Mercedes Chan, Ethan Sen, Elizabeth Hardy, Tim Rapley, Pauline Hensman, Ed Wraith,
Helen Foster
ABSTRACT:
Background:
Mucopolysaccharidoses (MPS) are rare inherited disorders resulting from glycosaminoglycans accumulation in cells. Children with MPS often have musculoskeletal (MSK)
abnormalities, from joint contractures to deforming abnormalities of the extremities and
spine. pGALS (paediatric Gait, Arms, Legs, and Spine), is a simple MSK assessment previously validated in school-age children to detect abnormal joints. We aimed to describe
the use of pGALS to identify patterns of MSK abnormalities in children with MPS.
Method: Videos of children with a spectrum of MPS performing pGALS were made at an MPS
specialist centre as part of their routine care, independent of the current study. Informed
consent for use of videos in research was obtained. A piloted proforma to record
abnormalities for each pGALS manoeuvre observed in the videos (scored as normal/
abnormal/not assessable) was used by 2 paediatric rheumatology (PRh) trainees and
1 specialist PRh physiotherapist, all blinded to MPS subtype. Videos were scored
independently by the 3 observers then re-scored for intra- and inter-observer
consistency. Data were pooled and analysed.
Result: 15 videos of children (9 boys, 6 girls, median age 11 years [4-19]) with MPS (10 MPS
attenuated type I; 4 MPS type II; 1 mannosidosis) were assessed. The most common
abnormalities detected using pGALS exam were restriction of shoulder, elbow, wrist,
TMJ excursion (likely impacted by many children having enlarged digits) [>75% cases]
and spinal deformity/restriction (2/3 cases). Mean intra-observer Kappa 0.74 (range
0.65-0.88) and inter-observer Kappa 0.62 (range 0.51-0.77). Hip manoeuvres within
pGALS were not clearly demonstrated in the videos.
Conclusion: In this observational study, pGALS identifies MSK abnormalities in children with MPS.
Restricted joint movement (and especially upper limb) was a consistent finding. We
acknowledge that further work is needed to include pGALS assessment of the hip and
also to test pGALS in an additional population of children with MPS; notably in further
children with attenuated MPS I as this subtype often has MSK abnormalities as the only
feature. The use of pGALS and awareness of patterns of joint involvement may be a
useful adjunct to facilitate earlier recognition of these rare conditions and facilitate
access to specialist care.
109
Trainee: Heng Gan
Board #: 78
Session #: 10
Supervisor: Mark Ansermino
Department: Anesthesia, Pharmacology & Therapeutics
Title: Calibration and evaluation of a novel low-cost pulse oximeter sensor
Author(s): Heng Gan, Christian Petersen, Martin J MacInnis, Guy Dumont, J Mark Ansermino
ABSTRACT:
Introduction: Pulse oximetry is vital for patient safety during anesthesia, yet remains worryingly
unaffordable in low-resource countries. Mobile phones are widely available even in rural
areas of the world. A novel pulse oximeter sensor, the AudioOx, interfaces directly with
the audio channel of the mobile phone, using the audio output to power the sensor, the
microphone input to record plethysmographic signal, the phone processor to interpret
plethysmographic data, and the phone display to report oxygen saturation (SpO2)
measurements. We calibrated and evaluated the AudioOx for accuracy, using two
commercially available pulse oximeters as references.
Method: With ethics and Health Canada approval, and informed consent, oximetry data were
collected in an observational study of healthy adult volunteers in an unrelated study
that required them to remain overnight in a hypoxia chamber (ambient oxygen 12-21%).
Subjects were fitted with three different pulse oximeters: a Nonin Xpod, a Masimo OEM
module and the AudioOx. Subjects and investigators were blinded to SpO2
measurements during data collection. The AudioOx was calibrated to SpO2 readings
from the Nonin Xpod. Subsequent AudioOx measurements were then compared to
concurrent SpO2 readings from the Nonin Xpod and the Masimo OEM module. Root
mean square accuracy (ARMS) and mean bias were calculated per the International
Organization for Standardization.
Result: Data were recorded from nine subjects, with SpO2 ranging from 70% to 100%. Data
from the first two subjects were used for calibration, and data from the subsequent
seven subjects for evaluation. With reference to the Nonin Xpod, the AudioOx had a
2.47% ARMS with a 0.70% mean bias (standard deviation 2.37%). With reference to the
Masimo OEM module, the AudioOx had a 4.34% ARMS with a -1.75% mean bias
(standard deviation 3.71%).
Discussion: The audio channel of a mobile phone can be used as an interface for a pulse oximeter
sensor, effectively reducing the cost of pulse oximetry to that of the sensor itself. The
simplicity and low cost of the AudioOx and its close correlation to reference SpO2
measurements promises an exciting new solution for disseminating pulse oximetry and
improving perioperative patient safety in low-resource countries.
110
Trainee: Anas Manouzi
Board #: 79
Session #: 10
Supervisor: Kevin Harris
Title: Coronary artery abnormalities identified with optical coherence tomography in children
with Kawasaki Disease
Author(s): Anas Manouzi, Martin Hosking, Anthony Fung, Astrid De Souza, Jim Potts, Kevin Harris
ABSTRACT:
Background:
Optical Coherence Tomography (OCT) is an intravascular imaging technique allowing
high- resolution imaging of the lumen and the superficial structure of the vessel wall
(resolution ~ 10 microns). The current evaluation of the cardiac sequelae of Kawasaki
Disease (KD) in children predominantly relies on the identification of coronary artery
dilation or stenosis based on echocardiography or angiography. The purpose of this
study is to demonstrate that OCT can be used in order to further characterize the
coronary vessel wall remodelling and intravascular lesions in children with KD.
Method: Patients with a diagnosis of KD with coronary artery dilation were assessed during their
planned follow-up catheterization. A series of modifications were made to the traditional
OCT protocol used in adults in order to adapt this technique to the pediatric population.
OCT recordings (Ilumien System, LightLabs, Massachussets) were performed in the right
coronary artery. Serial measurements of the cross sectional area (CSA) of the lumen and
intima were made at 1mm increments along the length of the vessel. Entire imaging data
was analyzed for the presence of mural thrombus and calcification.
Result: Cardiac catheterization was performed 11 months to 8.5 years after the diagnosis of
KD. Data was collect at 61 points along the right coronary artery in three patients with a
history of coronary artery aneurysms. The lumen area was normal (z-score >-2 and <+2)
at 93% of points. However, significant vessel wall abnormalities were noted in all children. Changes included marked intimal thickening, presence of calcification, presence
of white thrombus. Intimal thickening was most severe in areas of mild luminal narrowing
(r2 = 0.65). The CSA intima/lumen ratio was >1 at 48% of points analyzed consistent with
marked intimal thickening.
Conclusion: In this initial experience with OCT in children, we have shown that there are striking
coronary artery abnormalities in children with KD despite normal or near normal lumen
dimensions. Using conventional imaging techniques these changes would not be
identified. The prognostic significance of these changes is not presently known. This is
the first description of high- resolution, invivo imaging, of coronary changes in children
with KD.
111
Trainee: Lana Shaiba
Board #: 80
Session #: 10
Supervisor: Hassan El-Fawal
Title: Proteomic and neuroimmune biomarkers of Hypoxic-Ischemic Encephalopathy (HIE) in
an experimental model
Author(s): Lana Shaibah, Michael Horgan, Shaker Mousa, Hassan El-Fawal
ABSTRACT:
Background:
HIE has devastating neurological outcomes. With a narrow window of intervention and
no accurate diagnostic modality, the discovery of biomarkers should address this
challenge. Neuroimmune interactions suggest that nervous system (NS)-specific antigens
provide a record of insult in the form of autoantibodies (NAb).
Objective:
Delineate the humoral hypoxic response and corresponding detection of NAb
biomarkers in HIE.
Design
Sprague-Dawley rat pups underwent left carotid ligation and 3hr of hypoxia (8% O2) on
Method: PN7. Littermates with sham surgery and normoxia were controls. Neurofilaments (NF)
and myelin basic protein (MBP) antigens were chosen since gray and white matter are
involved in HIE. Serum NAb against NF, MBP and vascular endothelium growth factor
(VEGF), by ELISA, and erythropoietin (EPO), VEGF, IL-2, IL-6 and TNFα, by multiplex,
were measured on 1, 4, 7, 14, 21, 28 and 36d (n=9-16/time point) post-surgery.
Comparisons between groups was by ANOVA and associations between biomarkers by
regression analysis. Results: Pro-inflammatory IL-2, IL-6 and TNF-α were elevated at 24
hours post-surgery in HIE rat pups and for the duration of the study. Levels of cytokines
correlated with each other (r=0.7-0.9; p<0.001). Furthermore, EPO, adjusted for body
weight, and VEGF were (p<0.001) elevated at 24hr, but began to decline at 14d. HIE
pups had detectable levels of IgM and IgG against NF and MBP at 24hr post-hypoxic
exposure which increased in a time-dependent manner. Although there was a decline
in IgG levels between 14 and 28d, levels rebounded by 36d. Anti-NF and anti-MBP IgG
isotype were correlated (r=0.6-0.9; p<0.001).
Conclusion: Presence of pro-inflammatory cytokines, consistent with pathophysiology, and early
increases in EPO and VEGF, consistent with compensatory attempts, were evident. The
later decline in EPO levels provides further evidence for its therapeutic role in HIE, while
the decline in VEGF corresponded to the appearance of anti-VEGF IgG. The early
predominance of IgG against NF and MBP suggests isotype switching from a natural
repertoire of IgM. Natural NAb and subsequent complement activation are implicated
in the progression of hypoxic-ischemic injury. This may explain the transient decline in
NAb, reflecting release of NS antigen due to secondary insult. This study of early NAb
detection indicates that they may prove useful as biomarkers of HIE and suggests a role
in the progression of the disorders.
112
Trainee: Tracy Tucker
Board #: 81
Session #: 10
Supervisor: Tanya Nelson
Title: One step closer to cost effective Fragile X syndrome population carrier screening;
validation of a novel approach to assessing repeat size
Author(s): Tracy Tucker, Tanya N Nelson, Sylvie Langlois, Christy Cline, Kent J Moore, Mac Schermer
ABSTRACT:
Fragile X syndrome (FXS) is the most common form of inherited intellectual disability and is caused by
an expansion of a CGG repeat in the 5’ end of the FMR1 gene. FXS represents approximately one third
of X-linked intellectual disability and has a carrier frequency in females estimated to be between 1 in
250 and 1 in 350. To date, the greatest limitation to population carrier screening has been the lack of a
simple, accurate and cost-effective test.
Objective:
To validate a novel approach to assessing FMR1 CGG repeat size(s) in a cohort of male
and female samples enriched for premutation and full mutation alleles.
Method:
184 archived samples, previously analyzed using the current gold standard of PCR
visualized by capillary electrophoresis and/or Southern blot, were selected.These included individuals with normal (n=20), intermediate (n=20), premutation (n=82), mosaic pre/
full mutation (n=2) and full mutation alleles (60). Samples were analyzed blinded to the
genotype. Genomic DNA was PCR amplified, purified and analyzed by microfluidic
technology and repeat size calculated. Calculated repeat sizes were compared to
genotypes obtained by the current gold standard.
Result:
All premutation and full mutations were amplified; preferential amplification of smaller
alleles resulted in the identification of additional pre/full mutation mosaics. No normal samples were scored as full mutations. However, accuracy of genotyping was not
as high as with the current gold standard. As a result, some small full mutations were
scored as large premutations, some female mosaic pre/full mutations were scored as
large
premutations, and some large intermediate alleles were scored as small premutations.
Precision (run-to-run variation) was also not as high as for capillary electrophoresis.
Conclusion: Results from this pilot study indicate that this PCR-only based approach has a very high
sensitivity and specificity. Since it is amenable to high throughput with fast turn around
time, these data suggest that this assay could be used for preconception population
screening, with a small percent of samples at the intermediate/premutation boundary
requiring further size refinement by capillary electrophoresis. Further, if used for
diagnostic testing, the very small percentage of samples identified as large premutations
would require methylation analysis. Additional studies to assess feasibility and cost
effectiveness of integration into a clinical workflow are ongoing.
113
Trainee: Yuk Joseph Ting
Board #: 82
Session #: 10
Supervisor: Susan Albersheim
Title: Is the metabolism of triglycerides altered in neonates with hypoxic-ischemic
encephalopathy undergoing therapeutic hypothermia?
Author(s): Joseph Y Ting, Deepak Manhas, Sheila Innis, Susan G Albersheim
ABSTRACT:
Background:
Therapeutic hypothermia (TH) has been widely applied to improve both the survival and
neuro-developmental outcomes among infants with moderate-to-severe hypoxic
ischemic encephalopathy (HIE). Generally enteral feeds are withheld and total parenteral
nutritional (TPN) provided during hypothermia and early rewarming. Hypertriglyceridemia
can be associated with neurologic complications ranging from altered mental status and
irritability to seizures. Recently published studies have demonstrated altered
pharmacokinetics of medications metabolized primarily by the hepatic microsomal enzymes or excreted unchanged by the kidneys. In mice, acute hypoxia has been found to
decrease plasma triglyceride (TG) clearance. Whether TG metabolism is affected in infants
with HIE undergoing TH remains unclear.
Objective:
To compare serum TG concentrations of neonates, on TPN, with HIE undergoing TH to
those without TH (HIE and non-HIE infants).
Design
We conducted a retrospective review of infants with HIE undergoing TH during Jan
Method: 2010-Nov 2011 admitted into our neonatal intensive care unit, who had serum TG
concentrations measured at 48-96 hours of life. In practice Intralipid is prescribed at 1g/
kg/day on day 1-2 and then advanced by 1g/kg/day to a maximum of 3g/kg/day. Measurement of TG is not included in the routine blood work in our HIE management protocol, and there has been variation in practice. We identified seven infants with HIE and TH
(group I). Five infants with HIE, but without TH were in group II. Seven infants without HIE
or cooling (gastroschisis) serve as controls (group III).
Results: 3/7 infants undergoing TH had TG≥1.7mmol/l, whereas none of the others had TG
outside this desirable range.
Conclusions: We found that infants with HIE undergoing TH and receiving TPN, exhibited early
elevated TG levels compared to other infants. We postulate that under hypothermic conditions after hypoxic-ischemic insult, there might be decreased lipoprotein and
hepatic lipase activities leading to decreased clearance of the infused lipid, exacerbated
by increased mobilization of adipose tissue TG. Further large-scale prospective trials
would be helpful in elucidating lipid metabolism in these infants. This finding
underscores the importance of considering the ramifications of each new intervention,
particularly when dramatically altering systems.
114
Science Making Miracles
www.cfri-training.ca
950 West 28th Avenue, Vancouver, BC Canada V5Z 4H4
Phone: 604.875.3194 | Fax: 604.875.2496 www.cfri.ca

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