The Effect Of LI-cadherin On The Growth Of Hep3B Hepatocarcinoma Cells And The Related Mechanisms

ObjectiveTo investigate the effect of LI-cadherin (CDH17) on the growth of human Hepatocarcinoma Cells,use small RNA technology down it’s expression in human Hepatocellular carcinoma cell line Hep3B and to explore the mechanisms.Methods1. SiRNA was transfected into Hep3B by Lipofectamine transfection method,using G418selection stable transfection of Hep3B-LI-cadherin-of SiRNA cell lines, and using fluorescence microscopy to observe the transfection efficiency.2. Detect the expression of LI-cadherin protein and cyclin Dl by Western Bloting method in each group.3. Detect the importance of Ll-cadherin protein in hepatocarcinoma growth by MTT method and draw cell growth curve.4. The distribution of cell cycle in each group were detected by PI/Rnase staining-Flow Cytometry(FCM). Results1. Green fluorescence could obviously be seen from the SiRNA transfected cells,which confirm that the transfection is successful.The proportion was figured of fluorescenct cells of all the cells in the field of vision, which was85%, in line with the basic conditions of this experiment.2. Western blot analysis was used to detect the expression of LI-cadherin protein when Common culture after transfection for72hours, which showed:There were no obviously difference between empty plasmid carrying group and control group (P>0.05); The expression of LI-cadherin of SiRNA transfection group were lower than control group and empty plasmid carrying group (P<0.05), indicating that LI-cadherin is effectively suppressed; The expression of CyclinD1were significantly higher than empty plasmid carrying group and control group.3. Six points in three groups of cells were detected by MTT showed that:The difference of growth rate was no statistically significance among SiRNA transfection groups, control groups and empty plasmid carrying group in12h、24h、36h(P>0.05), but in48h,72h and96h of SiRNA transfected groups was significantly faster growth than empty plasmid transfected group and the blank control group, the difference was statistically significant (P<0.05), while the control group and empty plasmid transfected group no statistically significant (P>0.05).4. SiRNA transfection group of cells were compared with blank control group and empty plasmid transfected group:the percentage of S phase and G2/M phase increased significantly, G0/G1phase decreased significantly, cell proliferation index (PI)more higher, the differences were statistically significant (P<0.05); Blank control group was compared with empty plasmid transfected group in the percentage of cells in each cell cycle and PI value have no statistically significant (P>0.05). Conclusion1. LI-cadherin-SiRNA was transfected into hepatocellular carcinoma cells Hep3B, and Li-cadherin-SiRNA-Hep3B cell lines was constructed successfully.2. Reducing expression of LI-cadherin protein in hepatic cancer cells Hep3B which was successful transfected by LI-cadherin-SiRNA, increasing expression of CyclinD1Protein.3. LI-cadherin can inhibit the growth of the hepatoma cell line Hep3B and its possible by down-regulating the cell cycle protein expression of Cyclin D1levels and affect the cell cycle of the distribution of the percentage content.