| Hepatocellular carcinoma(HCC)is the third most common cause of cancer-associated mortality worldwide.There are>250,000 new HCC cases and an estimated 600,000 HCC mortalities each year worldwide.Although the combination of surgery and chemotherapy has increased the survival time of patients with HCC in recent years,a significant number of patients still relapse due to the resistance of tumor cells to chemotherapeutic agents.Unfortunately,the mechanisms underlying HCC chemoresistance remain unclear.Therefore,the identification of the genes and proteins that regulate chemoresistance is essential for the exploration of novel targeted therapies for HCC,which will be beneficial for a large group of patients.Yes-associated protein(YAP),a key component of the Hippo signaling pathway,serves roles in development,growth,repair and homeostasis.Accumulating evidence has implicated that YAP serves an important and multifaceted role in cancer progression,including gastric cancer,pediatric cancerand breast cancer.YAP also been demonstrated to be overexpressed in various types of solid tumors,including gastric cancer,pediatric cancer and breast cancer,and its overexpression is associated with tumor initiation,invasion and metastasis.These characteristics make YAP an attractive therapeutic target in cancer treatment.In HCC,YAP has been revealed to be involved in proliferation,migration,invasion and drug resistance.However,the underlying mechanisms remain unclear.So the purpose of this study is to demonstrate the mechanism of YAP regulating proliferation and invasion of HCC,and aslo to investigate the correlation between YAP and chemo-resistance in HCC.Methods1.HCC tissues and non-tumours tissues were enrolled in our study.The expression of YAP protein was detected by qRT-PCR and Western-blot methods.2.Hepatocelluar carcinoma cell lines SMMC-7721,normal live cell line THL-E3 were cultured in vitro,the expression of YAP were analyzed by qRT-PCR and Western-blot methods.3.Knockdown YAP in SMMC-7721 cells through LiporfectamineTM 2000 methods.The proliferation of cells was detected through MTT,BrdU and FCM methods.The invasive capacity of cells was analyzed through Materigel-Transwell methods.Meanwhile,qRT-PCR and Western-blot methods were used to detect the cell cycle associated proteins and apoptosis-associated proteins.4.Knockdown YAP in SMMC-7721 cells through LiporfectamineTM 2000 methods,the phosphorylation levels of PI3K/AKT signaling was analyzed through Western-blot.5.Knockdown YAP in SMMC-7721 cells through LiporfectamineTM 2000 methods,then the cells were incubated with different concentrations of LY294002 or DDP or LY294002 in combination with DDP,the proliferation of cells were analyzed with MTT methods.6.The HCC xenograft mouse model were constructed using PDTX methods,the mice were injected with normal saline,shYAP virus,DDP or shYAP virus in combination with DDP,the tumor volumes were calculated to detect the downregulation of YAP on HCC in vivo.Results1.Expression of YAP was increased in HCC tissues compared with non-tumors tissues.2.Expression of YAP in SMMC-7721 cells were both increased compared with THL-E3 normal live cell line.3.YAP knockdown inhibits hepatocellular carcinoma cell proliferation,cell invasion and also inhibits the expression of p21 and c-myc proteins,but increased the expression of Caspase-3 and Bax proteins in HepG2 and SMMC-7721 cells.4.Knockdown of YAP modulated phosphoinositide 3-kinase/AKT signaling.5.Knockdown of YAP enhances sensitivity of hepatocellular carcinoma cells to CDDP.6.Knockdown of YAP increases the anti-tumor activity of CDDP.ConclusionsThe findings suggest that the inhibition of YAP,alone or in combination with traditional chemotherapy,may be considered as an effective anti-HCC strategy. |
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