Tsg101

The proteins were isolated with RIPA lysis buffer containing protease and phosphatase inhibitors, PMSF, separated by 10–15% SDS-PAGE, and transferred to a PVDF membrane (Millipore, USA). Next, the PVDF membranes were blocked with 5% milk for 1 h and incubated overnight at 4°C with primary antibodies against CD9 (1 : 1,000; Cell Signaling Technology), TSG101 (1 : 1,000; Boster), CD81 (1 : 800; Proteintech), calnexin (1 : 1,000; Boster), α-SMA (1 : 1,000; Boster), vimentin (1 : 1,000; Cell Signaling Technology), collagen I (1 : 1,000; Boster), periostin (1 : 1,000; Proteintech), CD31 (1 : 1,000; Arigo), Angptl4 (1 : 800; Cell Signaling Technology), cyclin D1 (1 : 800; Cell Signaling Technology), β-catenin (1 : 800; Cell Signaling Technology), and GAPDH (1 : 2,000; Cell Signaling Technology). The next day, the secondary antibodies (1 : 3,000; Cell Signaling Technology) were incubated with the membranes for 1 h at 37°C. The bands were visualized by enhanced chemiluminescent image analysis.

Protein was extracted with RIPA Lysis Buffer (Beyotime Biotechnology, China) with protease and phosphatase Inhibitors. Western blot was performed according to the standard procedure. The antibodies used in this research were listed as follows: Vimentin (1:1000; CST), E-cadherin (1:1000; CST), SMAD3 (1:1000; CST), p-SMAD3 (1:1000; CST), TFG-β2 (1:500; Proteintech), β-actin (1:2000; Boster), ALIX (1:400; Boster), TSG101 (1:400; Boster).