The Effect Of Kui Jie'an On INOS Expression In Ulcerative Colitis Model And The Research On RNAi Technique

Objective: 1) To construct acute ulcerative colitis models of rats and interfere with Kui Jie'an, detected the difference of iNOS mRNA expression to study the mechanism of Kui Jie'an; 2) To establish a platform of RNA interference technique, which play the basic role in research on the occurrence of ulcerative colitis and the mechanism of Kui Jie'an. Methods: Using 2, 4-dinitrochlorobenzene(DNCB) and acetic acid to induce ulcerative colitis model,rats were divided randomly into 5 groups: normal group, model group, normal salt(NS) negative control group, 5-ASA positive control group, Kui Jie'an intervention group. Using semi-quantitative RT-PCR to detect the difference of iNOS mRNA expression in rats' colons. Construct an eukaryotic expression recombonant containing the rat iNOS gene and a recombinant plasmid carrying siRNA to rat iNOS gene. After transfecting the two plasmids into COS-7 cells together, detected the inhibitive effect by Western blot. Results: 1) Pathological section of colon showed that there were ulcerations in model group; 2) The expression of iNOS mRNA showed the statistical significance between model group and the other groups (P<0.05). But the expression between Kui Jie'an intervention group and normal salt group didn't show the statistical significance (P>0.05). 3) Identifcation by enzyme cutting and sequencing showed that the two plasmids were constructed successfully. Cotransfection showed that the expression of iNOS-GFP fusion protein was decreased in COS-7 cells. Conclusion: 1) The construction of model was successful; 2) Concluded that the effect of Kui Jie'an on ulcerative colitis didn't relate to iNOS expression directly; 3) Regulated the expression of rat iNOS gene in COS-7 cells using RNA interference (RNAi) technique successfully.