Epigenetic Mechanisms of Environmentally-mediated Cardiovascular Disease Risk in the WHI
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Epigenetic Mechanisms of Environmentally-mediated
Cardiovascular Disease Risk in the WHI
Andrea Baccarelli, MD, PhD, MPHLab of Environmental Epigenetics
Harvard School of Public Health
A. Baccarelli, Harvard School of Public Health
Epigenetics Programming of gene expression
that: does not depend on the DNA code (relatively) stable, i.e., replicated
through: cell mitosis meiosis, i.e. transgenerational (limited
evidence in humans) Characteristics of Epigenetic
Programming Modifiable (can be reprogrammed) Active or poised to be activated:
Potentially associated with current health states or predict future events
A Symphonic Example
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DNA Phenotype
Epigenetics
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A Symphonic Example
A. Baccarelli, Harvard School of Public Health From the Royal Society of Chemistry website
(www.rsc.org)
DNA Methylation
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Changes during mitosis Fidelity of “transcription” of DNA
methylation varies between 97-99.9% De-novo methylation: 3-5% mitosis Much more dynamic compared to DNA
sequence!
DNA methylation is known to be modified: through aging oxidative stress, inflammation,
micronutrients
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Triggers of cardiovascular events at the population level
Asymptomatic Angina MI
CardiovascularRisk Factors
Triggers
Prev
entio
nO
ppor
tuni
ties
Primordial Primary Secondary
Subclinical Disease
Racial &
Socioeconomic Disparities
Individual interventions
Societal interventions
Sudden or CHD Death
10 20 30 50 60 7040 80 90Age
Atherosclerosis time-course
• Diabetes mellitus• Hypertension• Smoking• Dyslipidemia• Inflammation• Physical inactivity
• Traffic exposure• Physical exertion• Alcohol• Coffee consumption• Air pollution• Emotions (positive/negative)• Heavy meal• Cocaine abuse• Sexual activity, etc.
Preclinical Clinical
High population attributable fraction (PAF)
High relative or absolute individual risk
Exposure to traffic and air pollution are estimated to trigger more heart attacks than:-Heavy alcohol consumpion (2.5 fold)-Cocaine abuse (10 fold)
Long term exposure to air pollution leads to:-Atherosclerosis-Cardiovascular morbidity and death
Figure from Editorial byBaccarelli & BenjaminThe Lancet 2011
Coding & non-coding DNA in the human genome
Protein coding genes 1-2%Unique non-coding DNA ≈45%
Pseudogenes <1% Repetitive sequences ≈50%LINEs (e.g., LINE-1): 21%SINEs (e.g., Alu): 11%Both LINEs & SINEs are retrotranspons
Heavily methylate
d
Chromosomal stabilityLimits retrotranspositionLimits inflammation
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A. Baccarelli, Harvard School of Public Health
Effects of Air Pollution on Repetitive Element DNA Methylation
Tarantini, et al. Environment Health Perspect, 2009
Effects of PM10 found both at the beginning and at the end of the work week
2525
.526
26.5
2727
.5A
LU D
NA
met
hyla
tion
(%m
c)100 250 500 1500
average PM10 (µg/m³)
7678
8082
84LI
NE
-1 D
NA
met
hyla
tion
(%m
c)
100 250 500 1000 1500average PM10 (µg/m³)
βunadj=-0.30; P=0.07βadj=-0.34; P=0.04
βunadj=-0.18; P=0.04βadj=-0.19; P=0.04
LINE-1 Alu
β for an increment equal to the difference between the 90th and 10th
percentile of PM10Mixed models: unadjusted or adjusted for age, BMI, smoking, cigarettes/day
A. Baccarelli, Harvard School of Public Health
10
20
30
40
50
60
Cum
ulat
ive
inci
denc
e of
isch
emic
hea
rt di
seas
e or
stro
ke (%
)
24 36 48 60 72 84 96Months from baseline examination
Low (68.1-77.4 %5mC)High (77.5-86.2 %5mC)
LINE-1 methylation
10
20
30
40
50
60
Cum
ulat
ive
inci
denc
e of
isch
emic
hea
rt di
seas
e (%
)
24 36 48 60 72 84 96Months from baseline examination
Low (68.1-77.4 %5mC)High (77.5-86.2 %5mC)
LINE-1 methylation
5
10
15
20
25
30
Cum
ulat
ive
inci
denc
e of
stro
ke (%
)
24 36 48 60 72 84 96Months from baseline examination
Low (68.1-77.4 %5mC)High (77.5-86.2 %5mC)
LINE-1 methylation
aFigure 1
b
Adj. Hazard Ratio3.6 (95%CI 1.8-7.0)
p<0.001
HR=2.8 (95%CI 1.3-5.9), p=0.009 for IHDHR=4.3 (95%CI 0.7-25.8), p=0.11 for strokeBaccarelli et al., Epidemiology
2010
DNA Methylation and Incidence of non-fatal IHD or Stroke in the NAS
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2
4
6
8
10
12M
orta
lity
(%)
[Dea
ths
from
isch
emic
hea
rt di
seas
e or
stro
ke]
0 12 24 36 48 60 72 84 96Months from baseline examination
Low (68.1-77.4 %5mC)High (77.5-86.2 %5mC)
LINE-1 methylation
Adj. Hazard Ratio2.9 (95%CI 1.4-6.3)
P=0.006
HR=3.5 (95%CI 1.4-8.8), p=0.007 for IHDHR=2.2 (95%CI 0.5-10.0), p=0.30 for strokeBaccarelli et al., Epidemiology
2010
DNA Methylation and Deathfrom IHD or Stroke in the NAS
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Potential Roles of Epigenetics
Baccarelli, Rienstra & Benjamin Circulation: Cardiovascular Genetics 2010
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Conclusions and Research Needs DNA methylation:
Sensitive to the environment Predicts risk of cardiovascular disease
Limitations: Limited power Unreplicated Limited to repeat elements (or other candidate sequence) environmentally homogeneous, single-city populations of
white men Research needs:
Investigating a diverse population Studies of women Characterizing DNA methylation changes over time and their
association with CVD-related risk factors
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R01 ES020836MPIs, Whitsel, Hou, Baccarelli
Epigenetic mechanisms of PM-mediated
cardiovascular risk
Pending (Score=19; Percentile 11%)
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Region Type Regions CpG sites covered on 450K BeadChip array
Average # of CpG sites per region
CpG Island 26,153 139,265 5.08N Shore 25,770 73,508 2.74S Shore 25,614 71,119 2.66N Shelf 23,896 49,093 1.97S Shelf 23,968 48,524 1.94
Remote/Unassigned - 104,926 -Total 485,553
CpG shelves, shores & islands classification (UCSC CpGi annotation)
N Shelf N Shore CpG Island
5’ UTR 3’ UTRTSS1500 TSS200
S Shore S Shelf
Illumina 450K BeadChip CoverageThe 450K BeadChip covers a total of 77,537 CpG Islands and CpG Shores (N+S)
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Study Population Two-stage, longitudinal study of associations
between PM air pollution, DNA methylation, and CVD risk factors: exam site- and race-stratified, randomly selected
6% minority oversample approximately 4,300 Women’s Health Initiative clinical
trial (WHI CT) women fasting blood draws and resting, standard, twelve-
lead electrocardiograms (ECGs) repeated at three-year intervals from 1993 to 2004.
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Study Design Stage 1 (Discovery)
interrogation, discovery and ranking of >450,000 DNA methylation sites potentially sensitive to PM in 1999-2001 blood samples from 800 participants
Stage II (Validation) Longitudinal validation of the 10 most PM-sensitive DNA methylation
sites identified by Stage 1 in up to three blood samples collected serially from the remaining 3,500 participants (1993-2004)
Focus on the temporal relationship between PM and DNA methylation at those sites, and that between site-specific DNA methylation and CVD risk factors.
Other Features: Phenomics framework to incorporate phenotypes Epigenetic data analyses adjusted for both ancestral admixture and
multiple comparisons External validation with cohorts with different participants
characteristics (ARIC, NAS)
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DNA methylation in the WHI
The WHI team for the Environmental Epigenomics proposal
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Eric Whitsel, University of North Carolina (PI) Lifang Hou, Northwestern University (PI) Andrea Baccarelli, Harvard University (PI) Yun Li, University of North Carolina Duanping Liao, Penn State Simon Lin, Northwestern University Lesley Tinker, Fred Hutchinson Linda Van Horn, Northwestern University