Therapeutic mechanism of Changji'an Formula in diarrhea-predominant irritable bowel syndrome model rats based upon tryptophan metabolism and ERK signaling pathways
OBJECTIVE To explore whether or not Changji'an Formula(CJAF)treats diarrhea-predominant irritable bowel syndrome(IBS-D)through interfering with tryptophan metabolism and its downstream NMDAR1/ERK/BDNF signaling pathway.METHODS Twenty-four Sprague-Dawley(SD)male rats were randomized into 4 groups of normal,model,CJAF(16.74 g·kg-1)and pinaverium bromide(18 mg·kg-1)(n=6 each).Three-factor method was employed for establishing a rat model of IBS-D.After successful modeling,CJAF was dosed by gavage once daily for 2 weeks.Finally the rats were sacrificed for tissue harvesting.The model was evaluated by drying method to determine the water content of feces,abdominal wall with-drawal reflex for assessing the visceral sensitivity,determination of sugar-water preference and hematoxylin-eosin(HE)stain.The plasma content of tryptophan metabolite was determined by LC-MS.Quantitative polymerase chain reaction(q-PCR)was utilized for detecting the mRNA expressions of NMDAR1,CREB1 and BDNF.Western blot was utilized for detecting the pro-tein expressions of IDO1,NMDAR1,ERK1/2,pERK,CREB1 and BDNF.And immunofluorescence was employed for detect-ing the expression of IDO1.Enzyme-linked immunosorbent assay(ELISA)was used for determining the expression of IFN-γ.RESULTS As compared with model group,CJAF and Pinaverium Bromide groups had lower fecal water content,lower AWR score and higher sugar-water preference value(P＜0.05).Plasma levels of TRP and KA spiked,the levels of Kyn and QA declined and the expression of IFN-γ was down-regulated(P＜0.05).And mRNA/protein expressions of IDO1,NMDAR1,pERK,CREB1 and BDNF dropped in colon(P＜0.05).CONCLUSION CJAF may improve the symptoms of pain and diar-rhea in IBS-D model rats,Its mechanism is probably correlated with the regulation of tryptophan-kynurenine metabolism,an inhi-bition of IDO1 expression and the regulation of NMDAR1/ERK/BDNF signaling pathway.
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