Abstract
BackgroundEpigenetic changes (i.e., chromatin modifications) occur during chondrogenesis and in osteoarthritis (OA). We investigated the effect of H3K27me3 demethylase inhibition on chondrogenesis and assessed its utility in cartilage tissue engineering and in understanding cartilage destruction in OA.MethodsWe used a high-content screen to assess the effect of epigenetic modifying compounds on collagen output during chondrogenesis of monolayer human mesenchymal stem cells (MSCs). The impact of GSK-J4 on gene expression, glycosaminoglycan output and collagen formation during differentiation of MSCs into cartilage discs was investigated. Expression of lysine (K)-specific demethylase 6A (UTX) and Jumonji domain-containing 3 (JMJD3), the HEK27Me3 demethylases targeted by GSK-J4, was measured in damaged and undamaged cartilage from patients with OA. The impact of GSK-J4 on ex vivo cartilage destruction and expression of OA-related genes in human articular chondrocytes (HACs) was assessed. H3K27Me3 demethylase regulation of transforming growth factor (TGF)-β-induced gene expression was measured in MSCs and HACs.ResultsTreatment of chondrogenic MSCs with the H3K27me3 demethylase inhibitor GSK-J4, which targets JMJD3 and UTX, inhibited collagen output; expression of chondrogenic genes, including SOX9 and COL2A1; and disrupted glycosaminoglycan and collagen synthesis. JMJD3 but not UTX expression was increased during chondrogenesis and in damaged OA cartilage, suggesting a predominant role of JMJD3 in chondrogenesis and OA. GSK-J4 prevented ex vivo cartilage destruction and expression of the OA-related genes MMP13 and PTGS2. TGF-β is a key regulator of chondrogenesis and articular cartilage homeostasis, and TGF-β-induced gene expression was inhibited by GSK-J4 treatment of both chondrogenic MSCs and HACs.ConclusionsOverall, we show that H3K27me3 demethylases modulate chondrogenesis and that enhancing this activity may improve production of tissue-engineered cartilage. In contrast, targeted inhibition of H3K27me3 demethylases could provide a novel approach in OA therapeutics.Electronic supplementary materialThe online version of this article (doi:10.1186/s13075-016-1053-7) contains supplementary material, which is available to authorized users.
Highlights
Epigenetic changes occur during chondrogenesis and in osteoarthritis (OA)
Using a focused library of 31 epigenetic inhibitors in chondrogenesis of human mesenchymal stem cell (MSC) (Fig. 1), we identified and confirmed the molecule GSK-J4 as an inhibitor of collagen output (0.63-fold; p = 0.0079) (Fig. 2a)
We show that inhibition of H3K27me3 demethylase activity by GSK-J4 disrupts in vitro chondrogenic differentiation of human MSCs but inhibits adult human articular cartilage degradation
Summary
Epigenetic changes (i.e., chromatin modifications) occur during chondrogenesis and in osteoarthritis (OA). Gene expression and cellular phenotype changes associated with OA and chondrogenesis are increasingly being attributed to altered activity of epigenetic modifying enzymes and consequent epigenetic regulation of target genes [1, 2]. Epigenetic modifications occur in gene promoters and include regulation of chromatin structure through cytosine methylation of DNA or through modifications such as acetylation and methylation of lysine residues in histone tails. Correct regulation of transforming growth factor (TGF)-β signalling is essential for cartilage development [4]. A disrupted TGF-β signalling response in both murine and human OA has been reported [7]. Epigenetic regulation of the TGF-β pathway in fibrosis has been reported, and TGF-β itself regulates expression of epigenetic modifying enzymes [8]
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