The role of epigenetic processes in bone development

Abstract

Osteoporosis is a progressive systemic skeletal disease caused by an imbalance of the bone remodelling process leading to a net loss of bone. Peak bone mass, the amount of bone tissue accrued at the end of skeletal maturation, is a strong predictor of osteoporotic fracture risk in later life. Genotype has been suggested to account for around 10% of the variance in peak bone mass, with increasing evidence suggesting the remaining variance may be explained by environmental exposures in prenatal life, such as maternal vitamin D status. The influence the prenatal environment on the trajectory of skeletal development in postnatal life has been suggested to occur through epigenetic mechanisms. Consistent with this, candidate gene studies have identified epigenetic marks in birth tissues predictive of later bone outcomes.
This project aimed to: (i) identify DNA methylation signatures at birth associated with childhood BMC and BMD, and (ii) determine whether maternal vitamin D supplementation altered DNA methylation in genes linked to improved BMC or BMD in postnatal development. To investigate these aims epigenome-wide association studies (EWAS) were performed using the Illumina Infinium MethylationEPIC Beadchip (EPIC) array to interrogate birth tissue DNA from the Maternal Vitamin D Osteoporosis Randomised Control Study (MAVIDOS RCT) and the Southampton Women’s Survey (SWS) studies along with umbilical cord mesenchymal stromal cells (UC-MSCs).
EWAS analysis identified methylation signatures at birth associated with childhood bone outcomes in both the MAVIDOS RCT and SWS studies. Within each study the top dmCpGs at each age group showed overlap between BMC and BMD outcomes, but with less overlap observed between age groups or between studies, indicating a tissue and age specificity to these changes. In addition, EPIC array interrogation of umbilical cord tissue DNA from the MAVIDOS RCT found maternal vitamin D supplementation induced significant differential methylation in three CpGs, including the potentially pro-osteogenic Solute Carrier Family 38 Member 10 and Tenascin C genes. The MAVIDOS RCT bone and intervention-related analyses together indicated differential methylation of multiple CpG sites in Protein Tyrosine Phosphatase Receptor Type N2 (PTPRN2), a gene involved in glucose homeostasis which also encodes miR-153, a negative regulator of insulin secretion and an anti-osteogenic factor in human bone marrow mesenchymal stromal cells. In the SWS cohort, differential methylation analysis of SWS umbilical cord blood DNA showed dmCpGs significantly associated with an insufficient maternal vitamin D status in late pregnancy. The most significant dmCpG was within Hexokinase 1, a rate-limiting gene in glycolysis. EWAS analysis of short-term vitamin D treated UC-MSCs also saw enrichment for dmCpGs in cellular respiration and glycolytic pathways.
In this study we identified associations between methylation of birth tissues and postnatal skeletal growth into childhood. Whilst the functional implications of these changes are not yet known, these dmCpGs, after further replication and validation, could potentially act as age and tissue-specific prospective markers of postnatal bone growth, helping to identify those with a lower peak bone mass and so a greater risk for osteoporosis later in life. These findings also suggest maternal vitamin D supplementation induced DNA methylation changes within foetal tissues in prenatal life may have the potential to influence the development of bone in postnatal life through epigenetic mechanisms. The effects of vitamin D on the epigenome may also be indirect through differential regulation of PTPRN2 expression or through influence on glucose homeostasis, both of which may also have implications for skeletal development.

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ORCID for Karen Lillycrop: orcid.org/0000-0001-7350-5489

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