Abstract:

Objective To discuss the effect of Yes-associated protein （YAP） silencing on the proliferation， migration， and invasion capabilities of the human cervical cancer （CC） SiHa cells. Methods The human CC SiHa cells were cultured in vitro， and the lentiviral YAP shRNA was transfected into the SiHa cells to establish stably transfected YAP-shRNA experimental group （sh-YAP group） and empty plasmid control group （control group）. Western blotting method was used to detect the silencing effect of YAP； immunofluorescence method was used to detect the microfilament number and morphology of actin filaments （F-actin） in the cells in both groups； CCK-8 method was used to detect the survival rates of the cells in two groups； Transwell chamber assay and wound healing assay were used to detect the numbers of migration and invasion cells and scratch healing rates of the cells in two groups； Western blotting method was used to detect the expression levels of epithelial-mesenchymal transition （EMT） markers （E-cadherin and Snail）， DNA damage repair-related proteins （γ-H2AX）， and apoptosis-related proteins ［c-MYC and B-cell lymphoma-2 （Bcl-2）］ in the cells in two groups. Results The results of lentiviral YAP shRNA transfection into SiHa cells showed that the expression level of YAP protein in the SiHa cells was significantly decreased （P<0.05）. The immunofluorescence results showed that after YAP silencing， the F-actin in SiHa cells was sparse and regularly arranged， with a reduced number of cells and a shriveled appearance. The CCK-8 results showed that compared with control group， the survival rate of the SiHa cells in sh-YAP group was significantly decreased cultured for 24 and 48 h （P<0.01）. The results of Transwell chamber assay and the wound healing assay showed that compared with control group， the numbers of migration and invasion SiHa cells in sh-YAP group were significantly decreased （P<0.01）， and the cell scratch healing rates were signifiantly decreased（P<0.05）. The Western blotting results showed that compared with control group， the expression level of E-cadherin protein in the cells in sh-YAP group was increased （P<0.05）， and the expression levels of c-MYC， Bcl-2， and γ-H2AX proteins were decreased （P<0.05 or P<0.01）. Conclusion YAP gene silencing leads to the depolymerization of F-actin in the human CC SiHa cells and regulates the apoptosis and DNA damage repair， potentially reversing the EMT process， thereby inhibiting the proliferation and migration of the tumor cells.

Key words: Cervical neoplasm, Yes-associated protein, Cell proliferation, Cell migration