Western blotting analysis
After treatment with PA and/or samples, the cells were harvested and
lysed with radioimmunoprecipitation assay lysis buffer. Cells containing
10 μg of protein were separated on sodium dodecyl sulfate-polyacrylamide
gel electrophoresis gels. The gels loaded with proteins were transferred
to polyvinylidene difluoride membranes, followed by blocking in skim
milk. The membranes were incubated with primary antibody and kept at 4
°C overnight. Subsequently, the membrane was washed and mounted with the
indicated secondary antibody based on the primary antibody for 1 h
incubation. The membrane was imaged with a chemiluminescence solution in
the dark and visualized by the Tanon 5200 automatic chemiluminescence
imaging system (Shanghai Tianneng Technology Co., LTD, China). The
intensities of proteins were calculated by ImageJ software.