ASL Overexpression Potentiate Apoptosis Evasion In Hepatocellular Carcinoma Via BCL2-Associated X Protein-Dependent

Background: Hepatocellular carcinoma(HCC)is one of the most common cancers in China.HCC is the fouth most commonly diagnosed cancer and the second-leading cause of cancer death.HCC is an insidious disease and most HCC patients are diagnosed with advanced stages,which renders surgical resection not applicable.The effect of currently chemotherapy regimens are not ideal which cause the poor efficacy and high mortality of HCC.Therefore,it is important to fully and deeply study the molecule mechanism of HCC development for early diagnosis,metastasis prediction,treatment of HCC.Argininosuccinate lyase(ASL)is an important part of urea cycle.ASL involved in the development of a variety of tumors,however,the relationship between ASL and the growth of HCC is not clear.Object:1.To investigate the effect of ASL on proliferation and apoptosis of HCC cells.2.to illustrate the mechanism of ASL regulation of HCC cells apoptosis.Methods:1.qRT-PCR and Western blot were applied to detect mRNA and protein levels of multiple liver cancer cell lines,immortalized liver cancer cell line(MIHA),and primary liver cancer cell line(PHH).Then,the mRNA and protein levels of ASL in the liver cancer tissues and adjacent tissues of 24 patients with liver cancer were also measured by qRT-PCR and Western blot,respectively.2.The effects of ASL silencing on the proliferation of HCC cells were detected by trypan blue staining,the effects of ASL silencing on the colony formation of HCC cells were detected by plate colony formation assay,and the effects of ASL silencing on the unanchored growth ability of HCC cells were detected by soft agar assay.Flow cytometry and western blot were applied to detect the effect of ASL silencing on apoptosis of liver cancer cells.3.The changes of mRNA levels of apoptosis-related genes in HCC cells with ASL silenced were analyzed by qRT-PCR.The components of the Bax mediated apoptotic pathway were detected by western blot.We used flow cytology and Western blot to detect the effect of silencing Bax on the apoptosis of liver cancer cells induced by ASL overexpression.Results:1.In this study,we found that the mRNA and protein levels of ASL in liver cancer cell lines were significantly higher than those in other cell lines,and the mRNA and protein levels of ASL in liver cancer tissues of 24 patients with liver cancer were also significantly higher than those in adjacent tissues.2.ASL silencing on hepatocellular carcinoma cells can inhibit the proliferation of hepatocellular carcinoma cells.Silencing ASL can inhibit the unanchored growth of HCC cells.Silencing ASL inhibited the colony formation ability of HCC cells.We also found that silencing ASL can effectively induce apoptosis of liver cancer cells.The mRNA and protein levels of Bax-related apoptosis pathway were induced by ASL silence in HCC cells.The growth inhibition of HCC cells induced by ASL silence was substantially restored by knock down of Bax protein.Bax silencing markedly deceased apoptosis that was induced by ASL silence.Conclusion: In summary,this study demonstrated that ASL acts as a carcinogene in the progression of HCC by regulating Bax signaling pathway.Therefore,we propose that ASL may be a potential therapeutic target for HCC treatment.