The Effect And Mechanism Of WSP On DSS-induced Ulcerative Colitis Through TLR4/NF-κB Signal Pathway

Objective:The rat model of ulcerative colitis(UC)was reproduced by dextran sodium sulfate(DSS)solution,and the effect of Water-soluble propolis on UC rats was investigated based on TLR4/NF-κB signal pathway,and its possible mechanism was studied.Method:Grouping: Male SD rats were random Ly divided into 6 groups(n=36),normal control group(NC group),ulcerative colitis group(DSS group),propolis lowdose group(L-WSP group,50 mg/kg),propolis medium-dose group(M-WSP group,100 mg/kg),propolis high-dose group(H-WSP group,200 mg/kg),and sulfasalazine group(Sulfa group,100 mg/kg).Model preparation: first weigh 5g of DSS powder and dilute it to 100 m L with double distilled water.Except for NC group,rats in other groups freely drink 5% DSS solution for 7 days to replicate the ulcerative colitis model.After that,rats in NC group and DSS group were orally gavaged with the same amount of distilled water,and rats in other groups were gavaged with the above concentration,at the same time point,for 7 days,once a day.During the modeling process,observe the status of rats in each group and record the daily disease activity index(DAI).After modeling,take the complete colon tissue and measure its length;The colon tissue of rats in each group was frozen and the morphological changes of colon tissue were observed by HE staining;The contents of TNF-ɑ and IL-6 in serum and colon,as well as the contents of NOX1 and NOX2 in colon were detected by ELISA.The activities of SOD,CAT and MDA in colon of rats in each group were detected by the kit;The expression of ZO-1 and Occludin protein in colon tissue was detected by immunohistochemistry;The content of TLR4,TAK1 and NF-κB m RNA and the expression level of TLR4,TAK1 and NF-κB p65 protein in colon were detected by Q-PCR and Western Blot respectively.Result:1.Compared with NC group,the rats in DSS group showed signs of depression,emaciation,decreased appetite,drowsiness and severe bloody stool,and the DAI score was significantly higher(P<0.05);Compared with the DSS group,the mental state of the rats in the drug treatment group in each group gradually improved,began to be active,the weight and appetite gradually increased,the blood and stool condition improved,and the DAI score decreased significantly(P<0.05).2.HE results showed that The colon length of rats in DSS group was significantly shorter than that in NC group(P<0.05);The colon length of rats in L-WSP group,MWSP group,H-WSP group and Sulfa group was significantly longer than that in DSS group(P<0.05);Compared with L-WSP group,the colon length of rats in M-WSP group,H-WSP group and Sulfa group increased significantly(P<0.05)。3.HE results showed that compared with NC group,the colonic mucosa epithelium of DSS group was severely damaged,the intestinal mucosa edema and exfoliation,serious erosion,and the number of intestinal gland cells and goblet cells morphology and structure destruction decreased.Compared with DSS group,the structure and morphology of colon tissue in L-WSP and M-WSP groups recovered,and the number and morphology of colon mucosal epithelial cells,gland cells and goblet cells in HWSP and Sulfa groups were close to normal rats.4.The indexes of IL-6 and TNF-ɑ in colon and serum of rats in DSS group were significantly higher than those in NC group(P<0.05);Compared with DSS group,the indexes of IL-6 and TNF-ɑ in colon and serum of drug treatment group were significantly decreased(P<0.05);Compared with L-WSP group,the indexes of IL-6 and TNF-ɑ in colon and serum of rats in M-WSP group,H-WSP group and Sulfa group also decreased significantly(P<0.05).Compared with NC group,the activities of SOD and CAT in colon tissue of rats in DSS group were significantly lower than that in NC group,and the content of MDA was increased(P<0.05);Compared with DSS group,the activities of SOD and CAT in colon tissue of rats in M-WSP group,H-WSP group and Sulfa group increased significantly,while the contents of MDA in colon tissue of rats in L-WSP group,M-WSP group,H-WSP group and Sulfa group decreased(P<0.05);Compared with L-WSP group,the activity of SOD in colon tissue of rats in H-WSP group and Sulfa group increased,and the content of MDA decreased significantly(P<0.05).The contents of NOX1 and NOX2 in colon tissue of rats in DSS group were significantly higher than those in NC group(P<0.05);The contents of NOX1 and NOX2 in colon tissue of L-WSP group,M-WSP group,H-WSP group and Sulfa group were significantly lower than those of DSS group(P>0.05);Compared with L-WSP group,the content of NOX1 in colon tissue of H-WSP group and Sulfa group decreased significantly(P<0.05).5.The expression of ZO-1 and Occludin in colon tissue of rats in DSS group was significantly lower than that in NC group(P<0.05);The expression of ZO-1 and Occludin in colon tissue of L-WSP group,M-WSP group,H-WSP group and Sulfa group was significantly higher than that of DSS group(P<0.05);Compared with L-WSP,the expression of ZO-1 and Occludin in colon tissue of M-WSP group,H-WSP group and Sulfa group increased significantly(P<0.05).6.The expression of TLR4,TAK1 and NF-κB m RNA in colon tissue of rats in DSS group was significantly higher than that in NC group(P<0.05);Compared with DSS group,the expression of TAK1 and NF-κB m RNA in colon tissue of L-WSP group was significantly decreased,and the expression of TLR4,TAK1 and NF-κB m RNA in colon tissue of M-WSP group,H-WSP group and Sulfa group were significantly decreased(P<0.05);Compared with L-WSP group,the expression of TLR4 and NF-κB m RNA in colon tissue of H-WSP group and Sulfa group decreased significantly(P<0.05).7.Compared with NC group,the expression of TLR4,TAK1 and NF-κB p65 in colon tissue of rats in DSS group was significantly higher(P<0.05);Compared with DSS group,the expression of TLR4,TAK1 and NF-κB p65 in colon tissue of L-WSP group decreased significantly,while that of M-WSP group,H-WSP group and Sulfa group decreased significantly(P<0.05).Conclusion :1.The improvement effect of WSP on enteritis injury in UC rats is mainly through reducing the production of inflammatory factors and increasing the activity of antioxidant enzymes,thus improving the integrity of the colon mucosal barrier in UC rats and alleviating its pathological damage;2.WSP reduces the level of intestinal inflammation and oxidative stress,restores the integrity of intestinal barrier,and significantly reduces the colon injury induced by DSS in UC rats,which may be related to the down-regulation of TLR4/TAK1/NF-κB signaling pathway related proteins.