A Global Increase in 5‐Hydroxymethylcytosine Levels Marks Osteoarthritic Chondrocytes

Abstract

Objective. To investigate the role of the newly discovered epigenetic mark 5-hydroxymethylcytosine (5hmC) and its regulators in altered gene expression in osteoarthritis (OA).Methods. Cartilage was obtained from OA patients undergoing total knee arthroplasty and from control patients undergoing anterior cruciate ligament reconstruction. Global levels of 5hmC and 5-methylcytosine (5mC) were investigated using immunoblotting, enzyme-linked immunosorbent assays, and cellular staining. Gene expression changes were monitored by quantitative polymerase chain reaction (PCR) analysis. Levels of locusspecific 5hmC and 5mC at CpG sites in the matrix metalloproteinase 1 (MMP-1), MMP-3, ADAMTS-5, and hypoxanthine guanine phosphoribosyltransferase 1 (HPRT-1) promoters were quantified using a glucosylation and enzyme digestion-based method followed by quantitative PCR analysis. Global and locus-specific 5hmC levels and gene expression changes were monitored in normal chondrocytes stimulated with inflammatory cytokines to identify the effect of joint inflammation.Results. A global 5-6-fold increase in 5hmC concomitant with a loss of TET1 was observed in human OA chondrocytes compared to normal chondrocytes. Enrichment of 5hmC was observed in promoters of enzymes critical to OA pathology, MMP-1 and MMP-3. Short-term treatment of normal chondrocytes with inflammatory cytokines induced a rapid decrease in TET1 expression but no global or locus-specific 5hmC enrichment.Conclusion. This study provides the first evidence of an epigenetic imbalance of the 5hmC homeostasis in OA leading to TET1 down-regulation and 5hmC accumulation. Our experiments identify 5hmC and its regulators as potential diagnostic and therapeutic targets in OA.

DNA methyltransferase 3b regulates articular cartilage homeostasis by altering metabolism

“…Whether canonical and/or noncanonical NF-κB signaling mechanisms play a role in regulating Dnmt3b expression has yet to be determined. In addition, IL-1β has also been shown to downregulate TET1 in chondrocytes (38,39). TET1 belongs to the TET family of proteins that catalyze DNA demethylation, and there is growing interest in how these TET-mediated epigenetic processes control cartilage homeostasis.…”

A B-cell targeting virus disrupts potentially protective genomic methylation patterns in lymphoid tissue by increasing global 5-hydroxmethylcytosine levels

“…Bar = 100 μM. development [9] and the pathogenesis of cancers like leukaemia [15,16] and inflammatory diseases such as osteoarthritis [27]. In light of these findings, and given the predilection of IBDV for the BF, we investigated potential changes in genomic methylation during key stages of B-cell development and after IBDV infection.…”

The first international workshop on the epigenetics of osteoarthritis

“…Alterations in DNA methylation patterns have been observed in OA chondrocytes, particularly a loss of DNA methylation at the promoters and the concurrent "unsilencing" of various OA-associated genes including MMP3, 9, and 13, ADAMTS4, IL-1β, and iNOS (14,33,34). Recently, Professor Bhutani's team reported a remarkable dysregulation of 5hmC homeostasis in patients with OA, with increases in global 5hmC levels observed in OA chondrocytes when compared to normal chondrocytes (35). 5hmC gain at specific sites in the promoters of key OA genes was associated with increased gene expression.…”