Ab104156

After extracting protein using RIPA buffer (Beyotime, Shanghai, China), the protein was quantified using BCA Protein Assay Kit (Tiangen, Beijing, China). Then, the protein samples were separated by 10% SDS-PAGE and transferred to PVDF membranes (Beyotime). Following blocking with 5% skimmed milk for 2 h at room temperature, the membranes interacted with primary antibodies against proliferating cell nuclear antigen (PCNA; 1:1000, ab18197, Abcam), Bcl-2 associated X protein (Bax; 1:1000, ab104156, Abcam), HDAC9 (1:20000, ab109446, Abcam), or GAPDH (1:2500, ab9485, Abcam) overnight at 4°C. After washing, the membranes were probed with HRP-coupled secondary antibody (1:25000, ab205718, Abcam) at room temperature for 2 h. Finally, the signal intensity was measured using ECL reagent (Absin, Shanghai, China).

The extraction of total protein was done utilizing RIPA buffer (Beyotime) and quantified utilizing the BCA protein assay kit (Tiangen, Beijing, China). An equal amount of protein was resolved with sodium dodecyl sulfonate-polyacrylamide gel (Solarbio, Beijing, China) and then transferred onto polyvinylidene difluoride membranes (Sigma-Aldrich). After blocking in 5% defatted milk for 1 h at indoor temperature, the membranes were incubated with primary antibodies against CD9 (ab223052; Abcam, Cambridge, MA, USA), CD63 (ab68418; Abcam), CyclinD1 (ab226977; Abcam), Bax (ab104156; Abcam), matrix metalloprotein 13 (MMP13; ab39012; Abcam), aggrecan (ab36861; Abcam), TRAF6 (ab137452; Abcam) or GAPDH (ab37168; Abcam) overnight at 4 °C and indicated secondary antibody (ab6789; Abcam) for 1.5 h at indoor temperature. The ECL kit (Beyotime) was employed for chemiluminescence imaging.

Cells were lysed using RNA immunoprecipitation (RIPA) (Solarbio), and protein samples were collected via centrifugation. Proteins of 20 μg were loaded on sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to nitrocellulose membranes (Solarbio). The membranes were blocked in 5% fat-free milk and incubated with primary antibodies anti-TRPC6 (ab62461, Abcam, Cambridge, MA, USA), anti-BAX (ab104156, Abcam), anti-BCL2 (ab194583, Abcam), or anti-β-actin (ab8227, Abcam) and the secondary antibody (ab205718, Abcam). Next, the bands were exposed to enhanced chemiluminescence (ECL) reagent (Solarbio).