Investigation On The Roles Of SiRNA-Id-1 In The Proliferation Of HepG2 Cell And The Regulation Of ERK1/2 Pathway

Objective: My project aims To study the effect of siRNA-Id-1 on the cell grow on HepG2 and investigate the relationship between Id-1 induces cell proliferation and the effects of extracellular signal regulated protein kinase.Method: established siRNA-Id-1 in vitro.after stable transfection into HepG2 cells by Lipofectamine 2000,HepG2 cell was setted to four groups with different treatments: siRNA-Id-1group,control SiRNA group, lipo2000 group, and blank group.the expression of Id-1 and ERK1/2 was analyzed by RT-PCR and western blotting respectively, cell growth was measured with MTT assay,cell cycle of the transfected cells was examined by flow cytometry.Result:Part I:After HepG2 cells were transfected with siRNA-Id-1 24 hours later,The results of RT-PCR analysis discovered that the expression of Id-1mRNA was suppressed significantly. Also Western bloting analysis suggested the result that the protein content of Id-1 was significantly decreased when Compared to the control siRNA group,lipo2000 group and blank group.Part II1. The results of MTT Experiment show this: The cell proliferation was inhibited when transfected with siRNA-Id-1, the inhibition ratio was higher than the other groups with markedly statistics difference (P<0.05).2. Flow cytometry analysis showed that HepG2 cells in G0/G1 phase increased after transfected with siRNA-Id- 1,but cells in S and G2/M phases decreased. which was significantly different from the control siRNA group,lipo 2000 group and blank group (P<0.05).Conclusion:Part I: During HepG2 cell was treated with siRNA-Id-1, the expression of protein Id-1 was silenced,and ERK1/2 genetic expression decrease at the same time.It hints that Id-1and ERK1/2 may get a certain relationship in the proliferation of HepG2 cell.Part II: RNA interference could inhibit the proliferation of HepG2 cell. and the inhibiton of proliferation has close relation with the decrease of Id-1 and ERK1/2 protein.it shows that Id-1 may promote the cell cycle,increase the proliferation rate via Ras-Raf-ERK1/2 signaling pathway. Id-1 may participate in the development of HCC by activating the ERK1/2 signaling pathways,...