| PurposeTo explore the effects of TLR4 regulating the proliferation of tumor-related protein CDK4,CDK6 on proliferation in HBV-related Hepatic carcinoma. Methods1.The expression of TLR4,CDK4 and CDK6 protein in 36 cases of HBV-related Hepatic carcinoma tissues and corresponding adjacent tissue were detected by Western blot, and the correlation between them in carcinoma tissues were analyzed.2.Three specific interference TLR4 siRNA sequence and a negative control sequence were builded and transfected into Hepatoma cells Hep-3B by liposomes.The expression of TLR4 at protein level was measured by Western blot assay before and after transfection, then selecting the best interference sequence.3.The experiment were divided into the si-RNA(-) group,TLR4-siRNA group and the control group(normal Hep-3B). The expression of TLR4,CDK4 and CDK6 at protein level were measured by Western blot assay before and after transfection.The proliferation in Hepatoma cells was also observed by MTT assay and colony experimental after transfection successfully. Results1.The overall level of expression of TLR4,CDK4 and CDK6 proteins in HBV-related Hepatic carcinoma were significantly higher than those in the corresponding adjacent tissues(P<0.05).And there was a positive correction between TLR4 and CDK4 with correlation coefficient(r) was 0.66(P<0.05), the positive correction between TLR4 and CDK6 was also found(r=0.57,P<0.05).2.Compared with the other two interfere sequences or normal cell, the best effect in reducing the protein levels of TLR4 was obtained with using the TLR4-siRNA-03 interference sequence(P<0.05).3.The proteins levels of CDK4 and CDK6 were significantly decreases in the TLR4-siRNA group(P<0.05), which compared to the control group after reducing the protein level of TLR4 by transfecting the TLR4-siRNA to the liver cancer cells.What is more, the proliferation of the Hepatoma cells Hep-3B and the colony formation rate(18.77±4.61)% were both decreased(P<0.05) in the group of TLR4-siRNA. Neither proliferation nor colony formation rate was changed between the groups of the control and the siRNA(-)(P>0.05). Conclusion1. The overall expression of TLR4, CDK4 and CDK6 proteins in HBV-related Hepatic carcinoma was higher than those in the corresponding adjacent tissues, and TLR4 has positively correlated with CDK4 and CDK6 in carcinoma tissues.2. TLR4 may regulates proliferation of HBV-related Hepatic carcinoma through controlling the expression of the proliferation of tumor- related protein CDK4 and CDK6. |
