| Objective:The effect of KYS on UC and the regulation of TLR4/MyD88/NF-?B p65 signaling pathway were studied by SPF mouse ulcerative colitis(UC)model,and the target and mechanism of KYS treatment of UC were explored.To lay the experimental foundation for its clinical application.Methods:A mouse model of ulcerative colitis(UC)was induced by a method of administering drinking water containing 3%dextran sulfate(DSS).Sixty SPF-level BALB/c male mice were randomly divided into 6 groups according to their body weight:normal(Ctrl)group,model(DSS)group,sulfasalazine(SASP)group(positive drug group),KYS-low dose(L).Group,KYS-medium dose(M)group,KYS-high dose(H)group,10 rats in each group.The Ctrl group was given normal drinking water for 15 days,and the remaining groups were given drinking water(3%DSS)for 7 days,and the 8th to 15th days were changed to normal drinking water.KYS-L,M,and H mice were given different concentrations of KYS solution(11.25 g/kg,22.5 g/kg,45 g/kg)daily from day 1 to day 14;SASP group mice were given daily.0.5 g/kg SASP suspension was intragastrically administered;the normal group and the model group were given normal saline per day;the volume of the mice in each group was consistent.The body weight and feed of the mice were weighed daily,and the activity,fur,and stool of the mice were observed and recorded.On the 14th night,the mice were fasted for 12 hours.On the 15th day,the mice were harvested.The mice were sacrificed by cervical dislocation and placed on ice.The colon specimens were taken from the anus and measured and recorded.Length,a sample of pathological and index tests made in mouse colon.The disease state of the mice was evaluated by disease activity index(DAI);the pathological condition of the colon was observed by HE staining,and the degree of colon tissue damage was evaluated by colon length change,colonic mucosal injury index(CMDI)and histological damage index(TDI).Colorimetric detection of myeloperoxidase(MPO)activity in colon tissue,indirectly reflecting colonic inflammation;serum endotoxin and inflammatory cytokine levels were detected by ELISA,inflammatory cytokines were detected by QPCR and Western Blot.TLR4/MyD88/NF-?B p65 signaling pathway-related gene and protein expression,comparing the therapeutic effects of SASP and KYS on mouse UC,and whether KYS treats UC by regulating TLR4/MyD88/NF-KB p65 signaling pathway.Results:Compared with the normal group of mice,the mice in the model group showed loose stools and bloody stools on the 5th day after modeling,the skin gloss disappeared,the activity decreased,the food intake and body weight decreased significantly(P<0.05),and the DAI increased significantly(P).<0.05);The mice in the model group were significantly shortened(P<0.05),the colonic mucosa was severely damaged,the CMDI was significantly increased(P<0.05),the colonic epithelial mucosal structure was deficient,and there was a large amount of inflammatory cell infiltration,and TDI increased significantly.(P<0.05);MPO activity in colon tissue increased significantly(P<0.05);ELISA results showed that serum LPS,TNF-?,IL-1?increased significantly in the model group(P<0.05);QPCR and Western Blot showed colon tissue TLR4 MyD88,NF-?B p65,I-?B gene and protein levels were up-regulated(P<0.05).Western Blot results showed phosphorylated NF-?B p65(p-NF-KB p65)and phosphorylated I-?B(pI-?B)levels were also significantly upregulated(P<0.05).Compared with the model group,the general conditions of SASP and KYS mice in different concentrations improved,the food intake and body weight increased significantly(P<0.05),DAI decreased significantly(P<0.05),and the colon length increased significantly(P<0.05),colonic mucosal injury recovered,CMDI increased significantly(P<0.05),colonic epithelial mucosal structure and inflammatory cell infiltration improved significantly,TDI decreased significantly(P<0.05),and mouse colon tissue MPO activity was down-regulated(P<0.05)P<0.05);ELISA results showed that the serum LPS,TNF-?,IL-1? in the model group decreased significantly(P<0.05);QPCR and Western Blot results showed that the colon tissue TLR4,MyD88,NF-?B p65,I-?B gene,The protein level decreased significantly(P<0.05).The results of Western Blot showed that the levels of phosphorylated NF-?B p65(p-NF-?B p65)and phosphorylated I-?B(pI-?B)were significantly down-regulated(P<0.05).There was no significant difference between the KYS-L,H and SASP groups,while the KYS-M group had the best effect.Conclusions:This study successfully constructed a DSS-induced UC mouse model;SASP and KYS interventions can significantly improve the UC symptoms caused by DSS,and KYS-M is the best and superior to the clinically used drug SASP;DSS induces activation of TLR/MyD88/NF-?B signaling pathway,SASP and KYS interventions can regulate this signaling pathway and reduce the production of inflammatory factors,and KYS-M is particularly significant;by regulating TLR4/MyD88/NF-?B p65 signaling pathway may be KYS treatment of UC mechanism. |
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