加味白头翁汤调节AMPK/SIRT1/PGC-1α通路对溃疡性结肠炎小鼠肠黏膜屏障的作用及机制OA

Objective To investigate the effect and mechanism of Modified Baitouweng Decoction on the intestinal mucosal barrier in mice with dextran sulfate sodium(DSS)-induced ulcerative colitis(UC)based on the adenosine 5'-monophosphate-activated protein kinase(AMPK)/silent information regulator 1(SIRT1)/peroxisome proliferator-activated receptor gamma coactivator 1-alpha(PGC-1α)pathway.Methods BALB/c mice were randomly divided into blank group,model group,mesalazine group(519.97 mg·kg-1 by gavage),high-dose Modified Baitouweng Decoction group(16 g·kg-1 by gavage),low-dose Modified Baitouweng Decoction group(8 g·kg-1 by gavage),and high-dose decoction+AMPK inhibitor group(16 g·kg-1 Modified Baitouweng Decoction by gavage+5 mg·kg-1 Compound C by intraperitoneal injection),with 10 mice in each group.The UC mouse model was induced by free access to 3％DSS solution for 7 days.Concurrent with model induction,each treatment group received the corresponding intervention at the above doses and routes once daily for 7 consecutive days.The general condition and body weight changes of the mice were observed and recorded,and the disease activity index(DAI)was scored.Colon length was measured,and the colonic mucosal damage index(CMDI)was scored.Hematoxylin-eosin(HE)staining and alcian blue-periodic acid-Schiff(AB-PAS)staining were used to observe colonic histopathological changes.Serum levels of diamine oxidase(DAO),interleukin(IL)-10,IL-4,IL-1β,and tumor necrosis factor-α(TNF-α)were measured by ELISA.The expression level of mucin 2(MUC2)in colonic mucosal cells was detected by immunohistochemistry.The mRNA expression levels of AMPK,SIRT1,and PGC-1α in colon tissue were detected by qPCR.The protein expression levels of the AMPK/SIRT1/PGC-1α pathway as well as ZO-1,Claudin-1,and Occludin were detected by Western Blot.M1 and M2 macrophage polarization in colon tissue and the proportions of Treg and Th17 cells in peripheral blood were detected by flow cytometry.Results(1)Compared with the blank group,the model group showed significantly decreased body weight(P<0.01),significantly increased DAI and CMDI scores(P<0.01),and significantly shortened colon length(P<0.01).Colon tissue exhibited crypt structural abnormalities,inflammatory cell infiltration,a significantly increased quantitative score for mucosal tissue damage(P<0.01),and a significantly reduced goblet cell area(P<0.01).Serum levels of DAO,IL-1β,and TNF-α were significantly increased(P<0.01),while IL-10 and IL-4 levels were significantly decreased(P<0.01).MUC2 expression in colonic mucosal cells was significantly decreased(P<0.01).The mRNA expression of SIRT1 and PGC-1α in colon tissue was significantly downregulated(P<0.05,P<0.01),and the protein expression of p-AMPK/AMPK,SIRT1,PGC-1α,ZO-1,Claudin-1,and Occludin was significantly downregulated(P<0.01).The proportion of M1 phenotype macrophages in colon tissue was significantly increased(P<0.01),while the proportion of M2 phenotype macrophages was significantly decreased(P<0.05).The proportion of Treg cells in peripheral blood was significantly decreased(P<0.05),while the proportion of Th17 cells was significantly increased(P<0.05).(2)Compared with the model group,the high-dose Modified Baitouweng Decoction group and the mesalazine group showed significantly increased body weight(P<0.01),significantly decreased DAI and CMDI scores(P<0.01),and significantly increased colon length(P<0.05).Colon tissue showed no obvious structural damage or inflammatory cell infiltration,with relatively intact crypts,a significantly decreased quantitative score for mucosal tissue damage(P<0.01),and a significantly increased goblet cell area(P<0.01).Serum levels of DAO,IL-1β,and TNF-α were significantly decreased(P<0.01),while IL-10 and IL-4 levels were significantly increased(P<0.05,P<0.01).MUC2 expression in colonic mucosal cells was significantly increased(P<0.01).The mRNA expression of SIRT1 and PGC-1α in colon tissue was significantly upregulated(P<0.05,P<0.01),and the protein expression of p-AMPK/AMPK,SIRT1,PGC-1α,Occludin,ZO-1,and Claudin-1 was significantly upregulated(P<0.05,P<0.01).The proportion of M1 phenotype macrophages in colon tissue was significantly decreased(P<0.01),while the proportion of M2 phenotype macrophages was significantly increased(P<0.05).The proportion of Treg cells in peripheral blood was significantly increased(P<0.05,P<0.01),while the proportion of Th17 cells was significantly decreased(P<0.01).(3)Compared with the high-dose Modified Baitouweng Decoction group,the high-dose decoction+inhibitor group showed significantly decreased body weight(P<0.01),significantly increased DAI and CMDI scores(P<0.05,P<0.01),and significantly shortened colon length(P<0.05).Colon tissue exhibited markedly increased damage and necrosis,with evident crypt abnormalities and inflammatory cell infiltration,a significantly increased quantitative score for mucosal tissue damage(P<0.01),and a significantly reduced goblet cell area(P<0.05).Serum levels of DAO and TNF-α were significantly increased(P<0.01),while IL-10 and IL-4 levels were significantly decreased(P<0.05).MUC2 expression in colonic mucosal cells was significantly decreased(P<0.01).The mRNA expression of SIRT1 and PGC-1α in colon tissue was significantly downregulated(P<0.01),and the protein expression of p-AMPK/AMPK,SIRT1,PGC-1α,and Occludin was significantly downregulated(P<0.01).The proportion of M2 phenotype macrophages in colon tissue was significantly decreased(P<0.01).The proportion of Treg cells in peripheral blood was significantly decreased(P<0.05),while the proportion of Th17 cells was significantly increased(P<0.01).Conclusion Modified Baitouweng Decoction ameliorates general symptoms,alleviates colonic histopathological damage,inhibits inflammatory responses,reduces intestinal permeability,and promotes intestinal mucosal barrier repair in DSS-induced UC mice.These effects may be associated with activation of the AMPK/SIRT1/PGC-1α pathway,promotion of M2 macrophage polarization,and restoration of Treg/Th17 cell homeostasis.