Mechanism of Shengxue Tongbian Granules in treating slow transit constipation based on network pharmacology and experimental validation
Objective To predict the potential mechanism of action of Shengxue Tongbian Granules(SXTBG)in treating slow transit constipation(STC)by network pharmacology and to verify it by animal experiments.Methods TCMSP and BATMAN-TCM databases were used to screen out the active ingredients and targets of SXTBG,and GeneCards and OMIM databases were used to screen out STC disease-related targets,then an"active ingredient-disease target"network was established.GO and KEGG enrichment analyses were performed on intersection genes through R language.A STC rat model was established by gavage with loperamide hydrochloride suspension[3 mg/(kg·d)].Rats were randomized into normal group(equal volume of distilled water),model group(equal volume of distilled water),mosapride group[1.35 mg/(kg·d)],low-dose SXTBG group[1.44 g/(kg·d)],and high-dose SXTBG group[2.88 g/(kg·d)],with eight rats in each group.All the groups were treated by gavage once a day for 14 consecutive days.Intestinal propulsion rate of the rats after treatment was recorded.In the rat colon tissue,HE staining was used to observe its pathological changes,immunohistochemical staining to check the expression level of tyrosine kinase receptor(c-Kit),TUNEL staining to examine the apoptosis of interstitial cells of Cajal(ICC),and Western blot to test expressions of glucose regulated protein 78(GRP78),C/EBP homologous protein(CHOP),Bcl-2 associated X protein(Bax),B-cell lymphoma-2(Bcl-2),and cleaved cysteine aspartic acid specific protease-3(cleaved Caspase-3).Results A total of 74 active ingredients of SXTBG were screened out,with 492 corresponding targets.CASP3(Caspase-3),Bcl-2,and Bax apoptosis genes were one of the core targets of STC,and their related pathways of action were endoplasmic reticulum stress signaling pathways.Animal experiment results showed that compared with normal group,the intestinal propulsion rate of model group decreased(P<0.01);the colonic mucosa atrophied and became thinner,with loss of mucosal epithelial layer,severe damage to the glandular tissue in the lamina propria,and visible inflammatory changes;the expression of c-Kit in the colon tissue decreased(P<0.01);the green fluorescence of apoptotic cells increased,the red fluorescence of c-Kit decreased,and the ICC apoptosis index increased(P<0.01);the protein expression levels of GRP78,CHOP,Bax,and cleaved Caspase-3 in the colon tissue increased(P<0.01),while that of Bcl-2 decreased(P<0.01).Compared with model group,the intestinal propulsion rates of mosapride group and low-and high-dose SXTBG groups increased(P<0.01);the colonic mucosal structure was relatively intact,and the inflammation was alleviated,and the glandular arrangement was relatively neat;the expression of c-Kit in the colon tissue increased(P<0.01);the green fluorescence of apoptotic cells decreased,the red fluorescence of c-Kit increased,and the ICC apoptosis index decreased(P<0.01);the protein expression levels of GRP78,CHOP,Bax,and cleaved Caspase-3 in the colon tissue decreased(P<0.01),while that of Bcl-2 increased(P<0.01).Compared with mosapride group,high-dose SXTBG group showed an increase in the intestinal propulsion rate(P<0.05)and c-Kit expression(P<0.05),and a decrease in the ICC apoptosis index(P<0.05);the protein expression levels of GRP78,CHOP,Bax,and cleaved Caspase-3 in the colon tissue of low-and high-dose SXTBG groups decreased(P<0.01),while that of Bcl-2 increased in high-dose SXTBG group(P<0.01).Conclusion SXTBG can inhibit ICC apoptosis and exert therapeutic effects on STC by inhibiting the endoplasmic reticulum stress GRP78/CHOP signaling pathway and regulating the expressions of Bax,Bcl-2,and cleaved Caspase-3 apoptotic proteins.
万方数据
维普
