Ecl chemiluminescence kit

Total proteins extracted by RIPA buffer (Beyotime Biotechnology, China) were separated by SDS-polyacrylamide gel electrophoresis (SDS-PAGE), and then transferred to polyvinylidene fluoride (PVDF) membranes (Millipore ISEQ00010). The membranes were first incubated with primary antibodies followed by secondary antibodies. The primary antibodies against NLRP3 (Cat# ab214185) and Caspase-1(Cat# ab1872), were from Abcam (Cambridge, UK). Antibodies against GAPDH (Cat#Mab5465-100) and HRP-linked anti-rabbit Immunoglobulin G (IgG) (Cat#GAR007) were from MultiSciences Biotech Co. (Hangzhou, China). Antibody against FOXO3a (Cat#D19A7) was from Cell Signaling Technology (Danvers, MA, USA). The primary antibodies against CD63 (catalog # ab134045), CD9 (catalog # ab92726), and TSG101 (catalog # ab125011) were from Abcam (Cambridge, MA, USA). Another primary antibody against CD63 (catalog # sc-5275) was from Santa Cruz Biotechnology (Dallas, TX, USA). The protein signals were detected using an ECL chemiluminescence kit (Biological Industries) and the luminescence was visualized using a BioRad luminescent imaging system.

Protein samples were extracted from UMSCs or exosomes using RIPA buffer (Beyotime Biotechnology) containing a cocktail of protease inhibitors (Beyotime Biotechnology). Equal amounts of protein were separated by SDS‐polyacrylamide gel electrophoresis and then transferred to PVDF membranes. The membranes were blocked with 5% bovine serum albumin (BSA) for 1 hour at room temperature and then incubated with primary antibodies followed by a horseradish peroxidase‐conjugated secondary antibody. The primary antibodies used were B2M (1:5000; Abcam Inc) and Bim (1:1000; Cell Signaling Technology Inc). The proteins were visualized using an ECL chemiluminescence kit (Biological Industries), and the luminescence was detected using a BioRad luminescent imaging system.

Total proteins extracted by RIPA buffer (Beyotime Biotechnology, China) were separated by SDS-polyacrylamide gel electrophoresis (SDS-PAGE), and then transferred to polyvinylidene fluoride (PVDF) membranes (Millipore ISEQ00010). The membranes were first incubated with primary antibodies followed by secondary antibodies. The primary antibodies against p16 (Cat#32050) and p21 (Cat#30427) were from Signalway Antibody (Baltimore, MD, USA). Antibody against GAPDH (Cat#60004-1-lg) was from Proteintech (Rosemount, IL, USA). Antibodies against HuR (Cat#sc-5261) and CD63 (Cat#sc-5275) were obtain from Santa Cruz Biotechnology (Dallas, TX, USA). The primary antibodies against CD9 (Cat#ab92726), were from Abcam (Cambridge, MA, USA). Antibodies against β-TrCP (Cat#4394), Histone H3 (Cat#9715) and ubiquitin (Cat#3933) were from Cell Signaling Technology (Danvers, MA, USA). The protein signals were detected using an ECL chemiluminescence kit (Biological Industries) and the luminescence was visualized using a BioRad luminescent imaging system.