Abstract
Epigenetic modifications are heritable changes in gene expression without changes in DNA sequence. DNA methylation has been implicated in the control of several cellular processes including differentiation, gene regulation, development, genomic imprinting and X-chromosome inactivation. Methylated cytosine residues at CpG dinucleotides are commonly associated with gene repression; conversely, strategic loss of methylation during development could lead to activation of lineage-specific genes. Evidence is emerging that bone development and growth are programmed; although, interestingly, bone is constantly remodelled throughout life. Using human embryonic stem cells, human fetal bone cells (HFBCs), adult chondrocytes and STRO-1+ marrow stromal cells from human bone marrow, we have examined a spectrum of developmental stages of femur development and the role of DNA methylation therein. Using pyrosequencing methodology we analysed the status of methylation of genes implicated in bone biology; furthermore, we correlated these methylation levels with gene expression levels using qRT-PCR and protein distribution during fetal development evaluated using immunohistochemistry. We found that during fetal femur development DNA methylation inversely correlates with expression of genes including iNOS (NOS2) and COL9A1, but not catabolic genes including MMP13 and IL1B. Furthermore, significant demethylation was evident in the osteocalcin promoter between the fetal and adult developmental stages. Increased TET1 expression and decreased expression of DNA (cytosine-5-)-methyltransferase 1 (DNMT1) in adult chondrocytes compared to HFBCs could contribute to the loss of methylation observed during fetal development. HFBC multipotency confirms these cells to be an ideal developmental system for investigation of DNA methylation regulation. In conclusion, these findings demonstrate the role of epigenetic regulation, specifically DNA methylation, in bone development, informing and opening new possibilities in development of strategies for bone repair/tissue engineering.
Highlights
Epigenetic regulation of gene expression is an important mechanism implicated in cell stemness, differentiation and function
In order to explore a potential link between DNA methylation changes in gene expression observed during fetal development, we have selected genes that we have previously reported to be associated with osteoarthritis (OA) [12,13,14]
Using human embryonic stem cells, human fetal bone cells (HFBCs), adult chondrocytes and a STRO-1+ skeletal stem cell containing population of adult bone marrow, we have examined a spectrum of developmental stages of femur development
Summary
Epigenetic regulation of gene expression is an important mechanism implicated in cell stemness, differentiation and function. Epigenetic modifications are heritable changes in gene expression that are not encoded by the DNA sequence. Methylated cytosine residues at CpG dinucleotides are commonly associated with gene repression [1]. The major role of DNA methylation is to protect host DNA against degradation by restriction enzymes. Eukaryote DNA methylation has been implicated in the control of several cellular processes including differentiation, gene regulation, development, genomic imprinting, X-chromosome inactivation and suppression of repetitive element transcription and transposition [2,3,4]. Current research has focused on the role of epigenetic processes in fetal programming and the potential consequences, including enhanced susceptibility, to chronic diseases in adulthood [5,6]
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