Mmp 13

ASIC1a (1:1,000 dilution, Bioss), Collagen-1 (1:1,000 dilution, Proteintech), α-SMA (1:1,000 dilution, Bioss), β-Actin (1:1,000 dilution, Proteintech), CaM (1:1,000 dilution, Affinity), CaMK II (1:1,000 dilution, Affinity), NFAT (1:1,000 dilution, Affinity), MMP-13 (1:1,000 dilution, Bimake.cn), NF-κB (1:1,000 dilution, Bimake.cn), KN93, and colchicine were purchased from MedChemExpress (MCE, United States). Spider-venompeptide (PcTx-1) was purchased from Abcam (Cambridge, United States). High-sugar DMEM culture medium and phosphate buffered saline (PBS) were purchased from HyClone (United States). Horseradish enzyme labeled anti-rabbit IgG and horseradish enzyme labeled anti-mouse IgG were purchased from Bioss (Beijing, China). PVDF membrane was purchased from Millipore (United States). Fluo-3AM calcium fluorescent probe was purchased from beyotime (Shanghai, China).Lipofectamine 2000, TRIzol Reagent, and Opti-MEM were purchased from Invitrogen (Invitrogen, United States). Amiloride was purchased from Sigma-Aldrich (St. Louis, MO, United States). Rapid Gel Preparation Kits were purchased from EpiZyme (EpiZyme, China). Cell cycle detection kit and CCK-8 cell proliferation toxicity assay kit were purchased from BestBio (Beijing, China).

Supernatants were collected for determination of total proteins after cells had been lysed on ice using the RIPA lysis buffer containing 10% phenylmethanesulfonyl fluoride. Protein concentrations were determined using the BCA Protein Assay Kit (Cat. no. 23227, Pierce, USA). Then, they were separated using 10% SDS-PAGE and transferred to a PVDF membrane. The membrane was incubated with primary antibodies and probed with respective secondary antibodies. The following antibodies were used: Cyclin D1(#2978, CST, USA), PCNA (SC-25280, USA), P-Erk1/2(#9101, CST, USA), Erk1/2(#4695, CST, USA), Stat3(#4904, CST, USA), P-Stat3(#9145, CST, USA), P-Stat3(#9134, CST, USA), MMP13(SC-30073, USA), MMP9(A5725, Bimake, China) and GAPDH (#2118s, CST, USA).