The Mechanism Of Rhodopsin A In Regulating Hippo/YAP Signaling Pathway In Hepatocellular Carcinoma HepG₂ Cells And Its Anticancer Effect In Combination With Oleanolic Acid

Objective:In this study,we aimed to investigate the effect and mechanism of HA on the biological behavior of HepG2 cells and to explore the effect of HA and OA on HepG2 cells.Methods:MTT and Transwell assay were performed for the biological behavior of HepG2 cells.RT-PCR and Western Blot were performed to detect the expression of YAP and TEAD at mRNA and protein level.Immunofluorescence assay was performed for the localization and fluorescence intensity of YAP in HepG2 cells.In order to study the relationship between YAP and HA,YAP overexpression or YAP knockdown cell lines were constructed using plasmids containing YAP cDNA or siYAP interfering RNA.MTT assay was used to detect the effect of HA on HepG2.Results:after treatment of HepG2 cells with HA,cells became smaller,rounded and adhesion decreased,and many cells were floating.With the increasing of HA concentration,the number of cells decreased gradually.MTT and Transwell results showed that HA could obviously inhibite the growth and migration ability of HepG2 cells.The inhibition effect was positively correlated with the drug concentration.RT-PCR assay showed YAP and TEAD had no significant change at mRNA level after HepG2 cells were treated with HA.Western Blot and Immunofluorescence results showed that the reduction of YAP expression level.In addition,Immunofluorescence result also showed that HA inhibited the nuclear accumulation of YAP.When YAP was overexpressed,the inhibitory effect of HA on HepG2 cells was obviously reduced.When YAP was knocked down,the inhibitory effect of HA on HepG2 cells was obviously increased.After treatment of HepG2 cells with HA combined with OA,the results showed that the combined effect of HA and OA was more obvious than that of HA alone or OA alone,and the content of YAP in cells decreased more distinctly.Conclusion:HA inhibits the proliferation of liver cancer HepG2 cells through a non-photoactivation pathway,and its mechanism of action is related to the Hippo/YAP signaling pathway.The combination of HA and OA has a more significantly inhibition effect.