Aberrant methylation and expression of TNXB promotes chondrocyte apoptosis and extracullar matrix degradation in hemophilic arthropathy via AKT signaling

Abstract

Recurrent joint bleeding in patients with hemophilia frequently results in hemophilic arthropathy (HA). Drastic degradation of articular cartilage is a major characteristic of HA, but its pathological mechanisms has not yet been clarified. Here, we conducted a genome-wide DNA methylation study with the goal of identifying critical genes for HA cartilage degeneration.DNA was isolated from human osteoarthritis (N = 5) and HA (N = 5) articular cartilages and analyzed using the Infinium Human Methylation 850 BeadChip array. Adeno-associated virus-mediated shRNA and siRNA were used to knock down Tenascin XB (TNXB) in chondrocytes and F8 -/- male mice, respectively. Then histopathological analysis, qPCR, Western blotting and immunofluorescence assays were conducted to detected chondrocyte homeostasis and HA progression.We found that Dnmt1 and Dnmt3a protein levels were increased in cartilages from HA patients compared with OA patients. Genome-wide DNA methylation analysis identified 1228 differentially methylated regions (DMRs) associated with HA. Functional enrichment analyses then revealed that DMR genes (DMGs) were related to extracellular matrix organization. Among these DMGs, TNXB protein expression was down-regulated in human and mouse HA cartilages. Further, the loss of Tnxb in F8 -/- mouse cartilage provides a disease-promoting role in HA by augmenting cartilage matrix degeneration and subchondral bone loss. Tnxb knockdown also promoted chondrocyte apoptosis and inhibited phosphorylation of AKT. Importantly, AKT agonist showed a chondroprotective effect following Tnxb knockdown.Our study demonstrated that TNXB is a central mediator of cartilage matrix degradation following joint bleeding, which functions by regulating the activation of AKT. These mechanistic insights allow targeted development of potentially new strategies for cartilage protection in HA.