Protocol for the EMPHASIS study; epigenetic mechanisms linking maternal pre-conceptional nutrition and children’s health in India and Sub-Saharan Africa

Abstract

Background Animal studies have shown that nutritional exposures during pregnancy can modify epigenetic marks regulating fetal development and susceptibility to later disease, providing a plausible mechanism to explain the developmental origins of health and disease. Human observational studies have shown that maternal peri-conceptional diet predicts DNA methylation in offspring. However, a causal pathway from maternal diet, through changes in DNA methylation, to later health outcomes has yet to be established. The EMPHASIS study (Epigenetic Mechanisms linking Pre-conceptional nutrition and Health Assessed in India and Sub-Saharan Africa, ISRCTN14266771) will investigate epigenetically mediated links between peri-conceptional nutrition and health-related outcomes in children whose mothers participated in two randomized controlled trials of micronutrient supplementation before and during pregnancy. Methods The original trials were the Mumbai Maternal Nutrition Project (MMNP, ISRCTN62811278) in which Indian women were offered a daily snack made from micronutrient-rich foods or low-micronutrient foods (controls), and the Peri-conceptional Multiple Micronutrient Supplementation Trial (PMMST, ISRCTN13687662) in rural Gambia, in which women were offered a daily multiple micronutrient (UNIMMAP) tablet or placebo. In the EMPHASIS study, DNA methylation will be analysed in the children of these women (~1,100 children aged 5-7 y in MMNP and 298 children aged 7-9 y in PMMST). Cohort-specific and cross-cohort effects will be explored. Differences in DNA methylation between allocation groups will be identified using the Illumina Infinium MethylationEPIC array, and by pyrosequencing top hits and selected candidate loci. Associations will be analysed between DNA methylation and health-related phenotypic outcomes, including size at birth, and children’s post-natal growth, body composition, skeletal development, cardio-metabolic risk markers (blood pressure, serum lipids, plasma glucose and insulin) and cognitive function. Pathways analysis will be used to test for enrichment of nutrition-sensitive loci in biological pathways. Causal mechanisms for nutrition-methylation-phenotype associations will be explored using Mendelian Randomization. Associations between methylation unrelated to supplementation and phenotypes will also be analysed. Conclusion The study will increase understanding of the epigenetic mechanisms underpinning the long-term impact of maternal nutrition on offspring health. It will potentially lead to better nutritional interventions for mothers preparing for pregnancy, and to identification of early life biomarkers of later disease risk.

Periconceptional environment predicts leukocyte telomere length in a cross-sectional study of 7–9 year old rural Gambian children

“…EMPHASIS cohort. The Gambian arm of the EMPHASIS study followed up children of mothers who were enrolled in the Peri-conceptional Multiple Micronutrient Supplementation Trial (PMMST) 54,55 (See Supplementary Methods for details). The present study utilised DNA collected from venous blood samples from the EMPHASIS children at age 7-9 yrs.…”

Racial/ethnic differences in the burden of type 2 diabetes over the life course: a focus on the USA and India

“…The cord blood of Indian babies also has higher concentrations of leptin and insulin, but lower concentrations of adiponectin, confirming a high-risk profile for future type 2 diabetes [34]. The contribution of genetics and epigenetics to this phenotype is currently being actively investigated [35,36].…”

Environmentally sensitive hotspots in the methylome of the early human embryo

“…We next directly explored the influence of genotype and environment at SoC-CpGs in the EMPHASIS (replication) cohort, for which we had data on 293 individuals measured at 286,552 polymorphic variants on the Illumina Global Screening Array (GSA) 12 . Following a similar strategy to that used in a recent study in the GUSTO cohort 31 , we performed a screen for genetic effects on DNAm.…”

“…Detailed descriptions of the Gambian cohorts analysed in the season of conception study are published elsewhere 11,12 . Briefly, for the younger cohort, blood samples from 233 children aged 2 years (median[IQR]: 731[729,733] days old) were collected from participants in the Early Nutrition and Immune Development ("ENID") study 11 .…”

“…In this study we assess the influence of seasonality on DNAm in two Gambian child cohorts 11,12 , enabling robust identification of loci showing consistent effects at the ages of 24 months and 8-9 years. Through prospective study designs, we capture conceptions throughout the year and use statistical models that make no prior assumptions about specific seasonal windows driving DNAm changes in offspring.…”