Researches On The Effect Of RNAi Targeting COX-2 On The Proliferation And Invasion Of Human Colon Carcinoma Cell

Objective: To observe the COX-2 expression after transiently transfecting human colon carcinoma cell HT-29 using siRNA, to approach the effect of RNAi targeting COX-2 on the proliferation and apoptosis of colon carcinoma cell.Methods: Three groups were divided,siRNA group, negative control group and blank group.For 72 hours after transfecting HT-29 cells with specific anti-COX-2 siRNA or N.C.FAM siRNA , MTT method was used to detect the proliferation of the cells every day in three days afer transfection and the cell growth curve was depicted, cell cycle and apoptosis were carried out on cells via flow cytometry.The level of COX-2 mRNA in the cells was measured by real time PCR and the PGE2 concentration in the cells culture supernatant was determined by ELISA.Results: Compared with two control groups, cell proliferation of siRNA group was obviously suppressed, the cells of siRNA group grew slowly, in G0-G1 phase were obviously increased(P<0.01) , in S and G2-M phases were obviously decreased (P<0.01), visible cell apoptosis was detected(P<0.01).The expression of COX-2 mRNA and the PGE2 concentration in the cell culture supernatant of siRNA group were reduced by 76.90%and 57.50%(P<0.01),but negative control group and blank group had no obvious difference (P >0.05).Conclusions: (1) In our country ,this is the first time to study the effect of slencing COX-2 by RNAi to the proliferation of human colon carcinoma.(2) Silencing COX-2 could effectively inhibit the proliferation of human colon carcinoma,the cells stayed in G0-G1 phase and were induced to cell apoptosis .(3) Silencing COX-2 may become a new way of therapy for human colon carcinoma.Part 2 Researches on the effect of RNAi targeting COX-2 on the invasion of human colon carcinoma cellObjective: To investigate the effect of RNAi targeting COX-2 on the invasion of human colon carcinoma cell.Methods: Three groups were divided,siRNA group, negative control group and blank group. For 48 hours after transfecting HT-29 cells with specific anti-COX-2 siRNA or N.C.FAM siRNA,the level of MMP-2 mRNA in the cells was measured by real time PCR, immunnohistochemistry was taken to detect the expression of MMP-2 protein,the invasiveness of HT-29 cell in vitro was measured quantitatively by the invasion test of Boyden chamber .Results: The expression of MMP-2 mRNA and protein of siRNA group were obviously suppressed compared with blank group and negative control group (P<0.01).The invasiveness of HT-29 cell was also reduced by 54.16%(P<0.01),but negative control group and blank group have no obvious difference (P >0.05).Conclusion: (1) In our country, this is the first time to study the effect of slencing COX-2 by RNAi to the invasiveness of human colon carcinoma. (2) Silencing COX-2 could effectively inhibit the expression of MMP-2mRNA,the protein level of MMP-2 was decreased and the invasiveness of colon carcinoma was degraded.(3) Down regulation of MMP-2 may be one of the important mechanisms to inhibit the invasiveness of colon carcinoma.