2.6. Western blotting

HUVECs were lysed in ice‐cold RIPA lysis buffer (Beyotime) supplemented with 1% protease and phosphatase inhibitor cocktail (Thermo Scientific). Equal amounts of proteins were separated using SDS‐PAGE and transferred to polyvinylidene fluoride membrane. Among the different experiments, the membranes were blocked and incubated with primary antibodies for MG53 (1:1000), phosphorylated FAK^Y397 (1:1000), phosphorylated Src^Y416 (1:1000), phosphorylated Akt^T308 (1:1000), phosphorylated ERK1/2 (1:1000), total FAK (1:1000), total Src (1:1000), total Akt (1:1000) and total ERK1/2 (1:1000) overnight at 4℃. Then, the membranes were exposed to horseradish peroxidase‐conjugated species‐specific respective goat IgG (Cell Signaling Technology) for 60 minutes at room temperature. The protein bands were visualized with chemiluminescence, and ImageJ software was used to measure the density of the bands.