| Background:Primary hepatocellular carcinoma(HCC)is one of the most common malignant tumors today.The degree of malignancy is high,the treatment effect is not ideal,and the mortality rate ranks third in malignant tumors.China is a high-risk area for liver cancer.About half of the world’s new liver cancer occurs every year in China,and about 90% of liver cancer in China is hepatitis B-related liver cancer.About 300,000 people die from liver cancer every year in China,giving people health.And the impact of the national economy should not be underestimated.The treatment of liver cancer is comprehensive treatment,including surgical resection,interventional therapy,ablation therapy,radiotherapy,chemotherapy,antiviral therapy and targeted therapy and immunotherapy in recent years,but the overall 5-year survival rate of liver cancer patients is about 12%.Even after surgical resection,the 5-year survival rate is usually lower than 38%-61%.In recent years,there has been no significant improvement.The treatment of liver cancer has reached the bottleneck stage.It is necessary to re-examine the current liver cancer treatment plan from the pathogenesis of liver cancer.In-depth study of the molecular mechanism of liver cancer development,the search for new markers and therapeutic targets for the diagnosis and treatment of liver cancer has very important theoretical and practical significance.The Hedgehog(Hh)signaling pathway has a complex role in the development of HCC and is associated with a variety of cancer characteristics.It has recently been found that blocking the Hh signaling pathway delays Hepatocyte Carcinogenesis induced by the hepatitis B virus-associated protein Hbx.Recent studies have reported that the expression of Hbx protein in human hepatitis B-associated hepatoma cells is associated with activation of Hh pathway;and the effect of Hbx on the growth of hepatoma cells by in vivo and in vitro experiments depends,at least in part,on the activation of Hh pathway.Tip: Hbx is involved in the regulation of Hh signalingpathway activity,but it is still unclear how Hbx regulates Hh signaling pathway.Objectives:1.Does hepatitis B virus-associated protein Hbx play a role in the abnormal activation of Hh signaling pathway in hepatoma cells?2.Does the hepatitis B virus-related protein Hbx promote the abnormal activation of Hh signaling pathway and promote the malignant phenotype of liver cancer cells?Methods:1.After transfection of Hbx plasmid in Huh7 hepatoma cell line,RNA was extracted and Hh signaling pathway-related target genes were detected.2.RNA was extracted from the liver of C57 mice infected with HBV,and Hh signaling pathway related indicators were detected.3.The HA-Hbx and Gli2-Flag plasmids were co-transfected into liver cancer cells for 24 hr,and the cells were lysed.The interaction between exogenous Hbx and Gli2 was detected by co-immunoprecipitation with the tag antibody.4.Gli-BS-Luc and Hbx expression plasmids were co-transfected into cells,and the regulation of Hb signaling pathway by Hbx was detected by Luciferase reporter gene assay.5.Hepatocellular carcinoma cell lines stably expressing Hbx were constructed,and the proliferation of hepatoma cells was observed by cell growth curve,plate colony formation assay and Edu cell proliferation assay.The invasion and migration ability of liver cancer cells was examined by cell invasion and migration experiments.Results:1.In the Huh7 hepatoma cell line,overexpression of Hbx leads to activation of the Hh signaling pathway and high expression of target genes downstream of the signaling pathway.2.The RNA level of Gli2 was significantly increased in mice infected with HBV compared with WT mice.3.The results of protein immunoprecipitation showed that Hbx protein interacts with Gli2 protein,a key component of Hh signaling pathway,in intact cells.4.The Hbx gene is a Gli2 upstream target gene and can regulate the Gli2 gene to enhance its expression.5.The proliferation,invasion and migration ability of hepatoma cell lines overexpressing Hbx were significantly enhanced compared with the control group.Conclusion:There is a correlation between HBs-related protein Hbx and Hh signaling pathway.Overexpression of Hbx in hepatocellular carcinoma cells can lead to abnormal activation of Hh signaling pathway and high expression of downstream target genes.Hbx is an upstream target gene of Gli2,a key gene of Hh signaling pathway,which regulates the expression of Gli2.There is an interaction between Hbx and Gli2 in intact cell physiology.Stable expression of Hbx promotes proliferation,migration and invasion of liver cancer cells. |
