Mechanism Of Qingchang Huashi Recipe By Activating AhR To Regulate IL-22 To Repair Intestinal Mucosal Injury In Colitis Model Mice

Objective:To explore the therapeutic effect of Qingchang Huashi recipe on colitis model mice,and explore from the perspective of aromatic hydrocarbon receptor(AhR)and intestinal mucosal barrier,to explore the effect of Qingchang Huashi recipe on IL-22 secretion and mucin and tight junction protein expression,and to explain alleviate the mucosal damage mechanism of colitis model mice,so as to provide objective basis for clinical application and drug research and development.Methods:1.Twenty four male SPF C57BL/6 mice were randomly divided into normal group,model group,Qingchang Huashi recipe low-dose group(12g/kg)and Qingchang Huashi recipe high-dose group(24g/kg).Since the research group has discussed the pharmacodynamic effect of Qingchang Huashi recipe group and 5-aminosalicylic acid group on colitis model mice in the early stage,the 5-aminosalicylic acid group will not be set up for the purpose of dose evaluation.Except for the normal group,mice in each group drank 2.5%DSS distilled water for 1 week to establish UC model.The low-dose Qingchang Huashi recipe group and the high-dose Qingchang Huashi prescription group were intragastrically administered with 12 g/kg and 24 g/kg of traditional Chinese medicine,while the other groups were given 0.9%normal saline.The mice were given gavage for 10 days,and the mice were killed on the 11th day.The disease index(DAI)and body mass index were measured.The colon length of mice was measured after administration.Observation of colonic pathology by HE staining.Western Blot was used to detect protein expressions of ZO-1,Claudin-4 and AhR.Q-PCR was used to detect the mRNA of IL-6,IL-1β,TNF-α and AhR in colon.2.Forty male SPF C57BL/6 mice were randomly divided into normal group,model group,Qingchang Huashi recipe group(12g/kg),Qingchang Huashi recipe+TMF(5 mg/kg)group and TMF group.Except for the normal group,mice in each group drank 2.5%DSS distilled water for 1 week to establish UC model.Qingchang Huashi recipe group,Qingchang Huashi recipe+TMF group were given 12 g/kg traditional Chinese medicine by gavage,0.9%normal saline was given to other groups.In addition to Qingchang Huashi recipe+TMF group and TMF group,TMF was injected intraperitoneally in TMF group according to 5 mg/kg,and 0.9%normal saline was injected intraperitoneally in the other groups.The mice were killed on the 11th day.The disease index(DAI)and body mass index were measured.The colon length of mice was measured at the end of administration;the pathological changes of colon were observed by HE staining;the expressions of ZO-1,claudin-4,AhR,and CYP1A1 were detected by Western blot.The mRNA levels of IL-1β、TNF-α、IL-22 and AhR were detected by Q-PCR.The expression of IL-22 in colon tissue was detected by immunofluorescence.Results:1.Compared with normal group,DAI score of model group rised(P<0.01);colon length was shortened(P<0.01);pathological damage of colonic mucosa was serious;protein expression of ZO-1,AhR and claudin-4 reduced(P<0.01;P<0.05;P<0.01);mRNA expression of AhR reduced(P<0.05);mRNA expression of IL-6,IL-1β,TNF-α rised(P<0.01;P<0.05;P<0.01).Compared with model group,DAI score of Qingchang Huashi recipe group(12g/kg)reduced(P<0.01),length of colon became longer(P<0.01),pathological condition of colonic mucosa was improved,protein expression of ZO-1 and claudin-4 was rised(P<0.01;P<0.01);mRNA expression of IL-6,IL-1β and TNF-α was reduced(P<0.01;P<0.01);protein expression of AhR was rised(P<0.01);mRNA expression of AhR was rised(P<0.01).Compared with model group,DAI score of high-dose Qingchang Huashi recipe group(24g/kg)reduced(P<0.01),length of colon became longer(P<0.01),pathological condition of colonic mucosa was improved,protein expression of ZO-1 and claudin-4 was rised(P<0.01;P<0.01);mRNA expression of IL-6,IL-1β and TNF-α reduced(P<0.01;P<0.05;P<0.01).;protein and mRNA expression of AhR rised(P<0.01).2.Compared with normal group,DAI score of model group rised(P<0.01);colon length shortened(P<0.01);pathological damage of colonic mucosa was serious;protein expression of ZO-1,AhR,CYP1A1,claudin-4 reduced(P<0.01);mRNA expression of IL-22、AhR reduced(P<0.01;P<0.05);mRNA expression of IL-1β、TNF-α rised(P<0.05;P<0.01);IL-22 expression in colon was continuously destroyed by immunofluorescence.Compared with model group,DAI score of Qingchang Huashi recipe group(12g/kg)reduced(P<0.01);colon became longer(P<0.01),colon mucosa pathology improved;protein expression of ZO-1,AhR,CYP1A1,claudin-4 rised(P<0.01);mRNA expression of IL-1β,TNF-α reduced(P<0.01;P<0.01);mRNA expression of IL-22、AhR rised(P<0.01;P<0.05);immunofluorescence of IL-22 in colon tissue significantly rised.It show recovery is continuous and distribution is uniform.Conclusion:Qingchang Huashi recipe can promote the recovery of colitis model mice and repair intestinal mucosal damage,and effect of Qingchang Huashi recipe low dose group(12g/kg)is better than Qingchang Huashi recipe high dose group(24g/kg).Qingchang Huashi recipe can activate AhR,up regulate the downstream gene CYP1A1,promote secretion of IL-22,inhibit level of inflammation,increase expression of mucin and tight junction protein,repair the intestinal mucosal barrier,so as to alleviate mucosal damage of colitis model mice and achieve therapeutic effect of UC.