DNA methylation regulates sclerostin (SOST) expression in osteoarthritic chondrocytes by bone morphogenetic protein 2 (BMP-2) induced changes in Smads binding affinity to the CpG region of SOST promoter

Abstract

IntroductionSclerostin (SOST), a soluble antagonist of Wnt signaling, is expressed in chondrocytes and contributes to chondrocytes’ hypertrophic differentiation; however its role in osteoarthritis (OA) pathogenesis is not well known. Based on our previous findings on the interaction between Wnt/β-catenin pathway and BMP-2 in OA, we aimed to investigate the role of DNA methylation and BMP-2 on SOST’s expression in OA chondrocytes.MethodsSOST mRNA and protein expression levels were investigated using real-time polymerase chain reaction (PCR) and Western blot, respectively. The methylation status of SOST promoter was analysed using methylation-specific PCR (MSP), quantitative methylation-specific PCR (qMSP) and bisulfite sequencing analysis. The effect of BMP-2 and 5’-Aza-2-deoxycytidine (5-AzadC) on SOST’s expression levels were investigated and Smad1/5/8 binding to SOST promoter was assessed by Chromatin Immunoprecipitation (ChΙP).ResultsWe observed that SOST’s expression was upregulated in OA chondrocytes compared to normal. Moreover, we found that the CpG region of SOST promoter was hypomethylated in OA chondrocytes and 5-AzadC treatment in normal chondrocytes resulted in decreased SOST methylation, whereas its expression was upregulated. BMP-2 treatment in 5-AzadC-treated normal chondrocytes resulted in SOST upregulation, which was mediated through Smad 1/5/8 binding on the CpG region of SOST promoter.ConclusionsWe report novel findings that DNA methylation regulates SOST’s expression in OA, by changing Smad 1/5/8 binding affinity to SOST promoter, providing evidence that changes in DNA methylation pattern could underlie changes in genes’ expression observed in OA.

Inflammation and epigenetic regulation in osteoarthritis

“…Another study showed that COL9A1 promoter was hypermethylated in OA chondrocytes and that such hypermethylation attenuated SOX9 binding to the COL9A1 promoter (94). Changes in DNA methylation of the sclerostin ( SOST ) promoter were also identified in OA chondrocytes, thus explaining its upregulation in these cells (95). …”

Sclerostin expression and functions beyond the osteocyte

“…While articular cartilage is negative for sclerostin protein in healthy joint tissue (18), numerous studies have reported sclerostin protein expression in articular chondrocytes isolated from osteoarthritic joints (13, 20–22). Sclerostin expression has also been reported to be altered in rheumatic joint diseases (14, 23, 24);…”

“…OA joints exhibit increased canonical Wnt signaling, leading to increased aggrecanase and matrix metalloproteinase expression and cartilage destruction. While normal articular chondrocytes are negative for sclerostin protein (18), articular chondrocytes isolated from OA-affected joints express sclerostin (13, 20–22). This has been reported in surgically-induced OA models in sheep (20) and mice (13, 20), the STR/Ort OA mouse model (30), as well as human OA (13, 20–22).…”

Distinct Gene Expression Patterns Defining Human Osteoblasts' Response to BMP2 Treatment: Is the Therapeutic Success All a Matter of Timing?

“…These donors show a strong induction of RANKL expression (23.9-fold) upon rhBMP2 expression. This might be due to the high binding affinity of the activated Smad1/5/8 complex to the SOST promoter described by Papathanasiou et al [34] that is supposed to be strongly dependent on the DNA methylation status of the cells. This might be one explanation why mesenchymal stem cells lose their osteogenic differentiation potential with age-dependent increase in DNA methylation [35].…”