The Effects Of RNAi On The Growth Of Hepatoma Carcinoma Cell HepG2 In Nude Mice And Erk1／2 Signaling Transduction Pathway

Objective:The purpose of this thesis is to detect the the effects of si-ID-1 on the growth of hepatoma carcinoma cell HepG2 in nude mice and to analyse the expression of ID-1 and p-ERK1/2 in transplanted liver cancer of nude mice.The other aim is to investigate the function of ID-1 and p-ERK1/2 in transplanted liver cancer of nude mice.Methods:5×106/0.2ml HepG2 cells were transplanted subcutaneously in the right axillary fossa of BALB/C nude mice,and fifteen transplanted liver cancer models were produced.A few days later nude mice with a tumor diameter of about 8mm in mean diameter were separately divided into three different group,including treatment group and a negative control group and a blank control group.Each group included 5 nude mice.Treatment group were injected with methylation of the SiRNA-ID-1 4nmol/100ul after 1 day,4 days,7 days,11 days,15 days,while the negative control group was injected with CONTROL-SiRNA4nmol/100ul, blank control group was injected with normal saline 100ul.Tumors'dimensions of three groups were measured per weeks and tumor growth curve were maked.After 28 days,nude mice were killed and routine pathological slides of tumor tissue were observed under light microscope.The expression of ID-1 and ERK1/2 was analyzed by real time RT-PCR.The expression of protein expression of ID-1 and p-ERK1/2 were examined with immunohistochemical methods.Result:Tumor formation rate in nude mice is 100%.Subcutaneous nodules could be touch after 5 days.With injection of methylated Si-ID-1, the treatment group of nude mice tumor volume significantly smaller than the negative control group and the blank control group after 1 week. The growth rate of the treatment group was also slowed down while the tumor volume of negative control group and blank control group continued to grow, the volume increased every week. Pathology showed the tumor cells in the experimental group were performance of degeneration and necrosis,most of the nucleus completely dissolved and the cell structure disappeared,tumor tissue of necrosis were homogeneous and red dye.T he cells around tumor in treatment group were less residual.However the tumor cells in the negative control group and blank control group, pathology showed tumor cells arranged densely,the growth were actively, nucleus were deep-stained and there are more mitotic figures.Compared with the two control groups,the mRNA level of ID-1 and p-ERK1/2 dramatically reduced (P<0.01)in treatment group.Immunohistochemical examination showed that the positive response rate compared with negative control group and blank control group is significantly reduced.There were significant statistical difference between the treatment group and the negative control group and the blank control group separately.Conclusion ID-1 and p-ERK1/2 in hepatoma carcinoma cell HepG2 in nude mice were highly expressed.The application of siRNA targeting methylation inhibited ID-1 gene could make the transplanted liver cancer of nude mice smaller in size and growth rate.The expression of ID-1 and p-ERK1/2 in tumor issues was decreased.We speculate that due to ID-1 over-expression led to ERK1/2 continue activate.Cells at the transcriptional level changed, and which due to cell proliferation and apoptosis uncontrolled.They are the prime molecular mechanism of cancerous liver cells.The si-ID-1 inhibited HepG2 cell proliferation and induced apoptosis mechanism may be through down the expression of ID-1 to block ERK1/2 pathway activation.ID-1 gene may be a new therapy target for liver cancer.