目的 探討茵梔黃顆粒治療小鼠膽汁淤積癥的作用及其機(jī)制。方法 將8周齡C57BL/6小鼠給予含1%膽酸的飼料2周，構(gòu)建小鼠膽汁淤積癥模型，將小鼠分為對照組、模型組、熊去氧膽酸（0.1 g/kg）組以及茵梔黃顆粒（5、10、20 g/kg）組，各組分別ig相應(yīng)藥物，對照組和模型組大鼠ig同體積生理鹽水，連續(xù)給藥4周。分別給予收集小鼠血清，全自動生化儀測定肝功能相關(guān)指標(biāo)，超高效液相色譜串聯(lián)質(zhì)譜（UPLC-MS/MS）測定膽汁酸成分；收集小鼠肝組織，蘇木精–伊紅（HE）染色觀察肝臟組織病理變化；RT-qPCR檢測肝組織炎癥因子和膽汁酸代謝關(guān)鍵基因的mRNA表達(dá)水平；Western blotting檢測肝組織膽汁酸代謝關(guān)鍵蛋白的表達(dá)情況；收集小鼠盲腸內(nèi)容物，Illumina Miseq測序平臺對V₃～V₄可變區(qū)進(jìn)行擴(kuò)增和測序，對小鼠腸道菌群結(jié)構(gòu)進(jìn)行分析。結(jié)果 與模型組相比，茵梔黃顆粒各組小鼠血清丙氨酸氨基轉(zhuǎn)移酶（ALT）、天冬氨酸氨基轉(zhuǎn)移酶（AST）、堿性磷酸酶（ALP）水平均顯著降低（P＜0.05、0.01、0.001）；且能夠顯著改善小鼠肝組織損傷。與模型組相比，茵梔黃顆粒各劑量組小鼠肝臟白細(xì)胞介素-10（IL-10）、腫瘤壞死因子-α（TNF-α）、白細(xì)胞介素-6（IL-6）、單核細(xì)胞趨化蛋白-1（MCP-1）mRNA表達(dá)水平均顯著降低，白細(xì)胞介素-1β（IL-1β）、膽固醇7-羥化酶（CYP7A1）、膽鹽輸出泵（Bsep）、牛磺膽酸鈉共轉(zhuǎn)運(yùn)蛋白（Ntcp）、膽汁酸轉(zhuǎn)運(yùn)蛋白多藥耐藥相關(guān)蛋白2（Mrp2）、UDP葡糖醛酸基轉(zhuǎn)移酶1家族多肽A1（Ugt1a1）mRNA表達(dá)水平均顯著升高（P＜0.05、0.01、0.001），并影響膽汁酸代謝關(guān)鍵蛋白的mRNA和蛋白表達(dá)。16S rDNA測序顯示，茵梔黃顆粒組小鼠腸道菌群中乳桿菌屬相對豐度顯著升高，羅斯氏菌屬和Allobaculum屬相對豐度顯著降低。膽汁酸代謝組學(xué)發(fā)現(xiàn)，茵梔黃顆粒能夠降低多種次級膽汁酸的含量。結(jié)論 茵梔黃顆粒改善膽汁淤積的作用機(jī)制可能與調(diào)控腸道菌群組成影響膽汁酸代謝有關(guān)。

Objective To investigate the effect and mechanism of Yinzhihuang Granules in treatment of cholestasis mice. Methods C57BL/6 mice at the age of 8 weeks were given a diet containing 1% cholic acid for 2 weeks to establish a mouse cholecystasis model, and the mice were divided into control group, model group, ursodeoxycholic acid (0.1 g/kg) group, and Yinzhihuang Granules (5, 10, 20 g/kg) groups, each group was given the corresponding drug intragaically, and the rats in the control group and model group were given the same volume of normal saline intragaically. The drug was administered continuously for 4 weeks. Serum was collected, liver function indexes were determined by automatic biochemical analyzer, and bile acid components were determined by UPLC-MS/MS. Liver tissues of mice were collected and stained with hematoxylin-eosin (HE) to observe the pathological changes of liver tissue. The mRNA expression levels of inflammatory factors and key genes of bile acid metabolism were detected by RT-qPCR. The expression of key proteins of bile acid metabolism in liver tissues was detected by Western blotting. The contents of mouse cecum were collected, and the V₃ — V₄ variable regions were amplified and sequenced by Illumina Miseq sequencing platform, and the structure of mouse intestinal flora was analyzed. Results Compared with model group, the serum levels of ALT, AST, and ALP in Yinzhihuang Granules group were significantly decreased (P < 0.05, 0.01, 0.001), and it can significantly improve the liver tissue damage of mice. Compared with model group, the mRNA expression levels of liver IL-10, TNF-α, IL-6, and MCP-1 in each dose group of Yinzhihuang Granules were significantly decreased. The mRNA expression levels of IL-1β, CYP7A1, Bsep, Ntcp, Mrp2, and Ugt1a1 were all positive significantly increased (P < 0.05, 0.01, 0.001), and affected the mRNA and protein expression of key proteins in bile acid metabolism. 16S rDNA sequencing showed that the abundance of Lactobacillus in gut microbiota of mice in Yinzhihuang Granules group was significantly increased, and the abundance of Roseburia and Allobaculum was significantly reduced. Bile acid metabolomics found that Yinzhihuang Granules could reduce the content of various secondary bile acid. Conclusion Mechanism of Yinzhihuang Granules on improving cholestasis liver injury may be related to regulating the composition of gut microbiota and affecting bile acid metabolism.

國家自然科學(xué)基金青年基金資助項目（81900468）；河南省醫(yī)學(xué)科技攻關(guān)計劃聯(lián)合共建項目（LHGJ20220419）；鄭州大學(xué)第一附屬醫(yī)院橫向課題（K2019-0148）