Dynamic epigenetic mechanisms regulate age-dependent SOX9 expression in mouse articular cartilage

While the developmental role of the SOX9 transcription factor in chondrocyte differentiation and cartilage formation is well documented, age-dependent SOX9 expression in articular chondrocytes (ACs) and its regulatory mechanisms remain unclear. This study aimed to explore epigenetic regulatory mechanisms for age-related changes in SOX9 expression in ACs of mice, spanning from the developmental stage to 18 months of age. Sox9 mRNA and protein were highly expressed in ACs during joint development but significantly decreased after 2 months of age. Histopathological features of osteoarthritis were not observed in examined hip and shoulder joints by 18 months of age. Epigenetic studies revealed that DNA methylation levels were increased at specific CpG islands of the Sox9 gene at 6 and 12 months; treatment of cultured ACs from 6-month-old mice with 5-azacytidine (an inhibitor of DNA methylation) elevated the level of Sox9 expression in ACs by lowering DNA methylation levels in the Sox9 promoter region. Histone 3 lysine 4 dimethylation (H3K4me2, a histone modification for transcriptional activation) in the Sox9 promoter region was decreased with age, which was associated with the age-dependent decrease in SOX9 expression in ACs. Knockdown of lysine-specific demethylase-1 up-regulated SOX9 expression in ACs of adult mice through increased recruitment of H3K4me2 in the Sox9 promoter region. Our results suggest that SOX9 expression in mouse ACs is significantly decreased after the completion of joint development. These age-dependent changes in SOX9 expression are dynamically regulated by DNA methylation and histone methylation.