H3k36me3

Chondrocytes were seeded at a density of 2 × 10⁵ cells and subjected to different treatments. Total protein of cells were extracted by SDS lysis buffer (Beyotime, China). Protein samples were subjected to 4–20% polyacrylamide gels (Genshare biological, China) electrophoresis and transferred to polyvinylidene fluoride (PVDF) membranes (Merck Millipore, USA). After being blocked with 5% skim milk, membranes were incubated with specific primary antibodies against COLII (1:1000, proteintech, UK), SOX9 (1:1000, Abcam, UK), COX-2 (1:1000, CST, UK), MMP13 (1:1000, proteintech, UK), H3K4me3 (1:1000, CST, UK), H3K9me3 (1:1000, Abcam, UK), H3K27me3 (1:1000, CST, UK), H3K36me3 (1:1000, ABclonal, USA), H3K79me3 (1:1000, ABclonal, USA), KMT2B (1:1000, ABclonal, USA), β-catenin (1:1000, CST, UK), Histone H3 (1:2000, CST, UK), ZBTB20 (1:1000, proteintech, UK) and GAPDH (1:10,000, proteintech, USA) at 4 °C overnight. The membranes were then probed with secondary antibodies and visualized on an Amersham 600 Chemiluminescence System by using chemiluminescence (ECL) substrate kit (Merck Millipore, USA). All the Western blot bands were quantified by Image-Pro Plus 6.0 software and normalized to an internal control (GAPDH or Histone H3).