The Study Of The Effect On K Uichangning D Ecoction About TLR2、NF-κB、IKK-β And Cytokines Of IL- 13, IL- 1β And TNF-α In Ulcerative Colitis Rats

Objective:Build the model rats of Ulcerative Colitis by the means of TNBS. To observe the effects about TLR2,NF-κB,IKK-βand the Cytokine of IL-1β,TNF-α, IL-13 of Kui Chang Ning on the colonic mucosa in model rats with Ulcerative Colitis, explore its mechanism of action, and provide a good experimental base for further clinical research. Methods:Stage one(Grouping):Twenty male and twenty female adult Wistar rats with specific pathogen free(SPF) whose weight is 170 gram to 200 gram, were divided into 4 groups(blank group, model group, Chinese medicine group, western medicine) randomly, and each group has ten.Stage two(Establishing the model and delivering the drug): The rats except blank control group were anesthetized by Nembutal, then clystered with TNBS(80mg/kg) and 50% ethanol method, to induce the acute colitis. Fifteen days later, these rats were clystered with TNBS(30mg/kg) to establish the ulcerative colitis of relapse model. And at the fifteen day, the Chinese medicine group was gavaged with Kui Chang Ning(13.5g/kg), and the western medicine group with sulf- asalazine(0.3g/kg), while the blank group and model group were given saline(0.9%) by the scale of 10ml/kg. Each group was fed in continuous 7 days.Stage three(Testing the index): At the end of medication, all groups stop feeding about 24 hours. All the rats anesthetized with 0.3% Nembutal, then anatomize the rat to get the whole colon tissue, cleaning with ice normal saline, observation the hyperemia, edema, ulceration on colonic mucosa by naked eye. Fixing the colon tissue about 1cm length at 4 % paraformaldehyde,make it into dehydration,put them in paraffin and embedded sections. At last,routining HE staining the sections to observe the change of colon histomorphology in light microscope. Testing the expression levels of NF-κB and TLR2 by immunohistochemistry, IKK-β by Real-time PCR, TNF-α/ IL-1β/ IL-13 by ELISA with the left colon tissue。Result:1. The blank group rats have a better state in lustre of hair, response, body weight, stool than other groups. After treatment, compared with the rats in model group, Chinese medicine group’s rats have evident effect in hair, response, ingestion, body weight, stool, and so on.2. Compared with the rats in blank group, the level of TLR2 is increased significantly in model group(P< 0.01). Compared with the rats in model group, the level of TLR2 is decreased significantly in Chinese medicine group(P< 0.01). Compared with the rats in western medicine group, the level of TLR2 has no significant difference with Chinese medicine group(P> 0.05).3. Compared with the rats in blank group, the level of NF-κB is increased significantly in model group(P< 0.01). Compared with the rats in model group, the level of NF-κB is decreased significantly in Chinese medicine group(P< 0.01). Compared with the rats in western medicine group, the level of NF-κB has no significant difference with Chinese medicine group(P> 0.05).4. Compared with the rats in blank group, the level of IKK-β is increased significantly in model group(P< 0.01). Compared with the rats in model group, the level of IKK-β is decreased significantly in Chinese medicine group(P< 0.01). Compared with the rats in western medicine group, the change level of IKK-β in Chinese medicine group is not obviously, it has no statistical significance(P>0.05).5. Compared with the rats in blank group, the level of TNF-α is increased significantly in model group(P< 0.01). Compared with the rats in model group, the level of TNF-α is decreased significantly in Chinese medicine group(P< 0.01). Compared with the rats in western medicine group, the level of TNF-α is decreased significantly in Chinese medicine group(P< 0.01).6. Compared with the rats in blank group, the level of IL-13 is decreased significantly in model group(P< 0.01). Compared with the rats in model group, the level of IL-13 is increased significantly in Chinese medicine group(P< 0.01). Compared with the rats in western medicine group, the level of IL-13 is increased significantly in Chinese medicine group(P< 0.01).7. Compared with the rats in blank group, the level of IL-1β is increased significantly in model group(P< 0.01). Compared with the rats in model group, the level of IL-1β is decreased significantly in Chinese medicine group(P< 0.01). Compared with the rats in western medicine group, the level of IL-1β is decreased significantly in Chinese medicine group(P< 0.01). Conclusion:1. Though clysters with TNBS can successfully established Ulcerative Colitis rat model. Kui Chang Ning Formula treatment Ulcerative Colitis curative effect is accurate. Kui Cang Ning Formula can inhibit inflammatory cell infiltration; promote the repair of colonic mucosa to treatment the Ulcerative Colitis.2. Kui Chang Ning Formula though decreases the expression level of TLR2 in colonic mucosa of rats; adjust the immunoreactions and production of inflammatory mediator, to reduce the injury of colonic mucosa in rats. It may be an important target sites in cure the rats with Ulcerative Colitis.3. Kui Chang Ning Formula can inhibit the over-activation of NF-κB. The expression level of NF-κB is a sensitive index for evaluating the degree of damage of colonic mucosa in Ulcerative Colitis. Kui Cang Ning Formula can decrease the expression level of NF-κB; reduce the inflammatory response in colonic mucosa. It may be a mechanism in cure the rats with Ulcerative Colitis.4. Kui Chang Ning Formula can decrease the expression level of IKK-β; it may be a mechanism in decrease the activity of NF-κB to cure the rats with Ulcerative Colitis.5. Kui Chang Ning Formula though decrease the content of TNF-α to reduce the injury of colonic mucosa. 6. Kui Chang Ning Formula though increases the content of IL-13 to promote the repair of colonic mucosa.7. Kui Chang Ning Formula though decreases the content of IL-1β to reduce the injury of colonic mucosa, and inhibit the inflammatory response.