基于RNA⁃Seq探讨清解化攻方干预重症急性胰腺炎大鼠肠屏障损伤的潜在作用机制

RNA⁃Seq⁃based exploration of the potential mechanism of action of Qingjie Huagong decoction in intervening intestinal barrier injury in rats with severe acute pancreatitis

目的:采用高通量RNA⁃Seq探讨清解化攻方干预重症急性胰腺炎大鼠肠屏障损伤的潜在作用机制。方法:采用逆行胰胆管注射牛黄胆酸钠方法建立SAP模型,HE观察空白组、模型组、中药组大鼠胰腺和回肠组织病理改变情况,ELISA法检测大鼠血清血淀粉酶、脂肪酶、DAO、内毒素水平;RNA⁃seq测序检测各组大鼠回肠组织差异性表达基因,筛选清解化攻方干预重症急性胰腺炎大鼠肠屏障损伤的核心基因、生物学功能及信号通路。结果:与空白组比较,模型组大鼠胰腺和回肠组织均有明显的病理改变,经清解化攻方干预得到显著改善;ELISA结果提示清解化攻方能够显著改善SAP大鼠血清淀粉酶、脂肪酶、DAO、内毒素含量(P<0.05)。RNA⁃Seq结果显示SAP组与BG组差异基因有2623个,经QJHGD处理后表达明显下调的基因有175个,主要有Tnip3、Lpo等,而上调的基因有320个,主要有aicda、Tnn等,参与调节氧化应激等生物过程,并干预TJ、细胞凋亡、Toll样受体、NOD样受体、NF⁃κB等多条信号通路而发挥作用。结论:清解化攻方可能通过调节Tnip3、aicda等关键基因及调控TJ、细胞凋亡、Toll样受体等多条信号通路起到抑制重症急性胰腺炎肠屏障损伤作用。

Objective:To explore the potential mechanism of action of Qingjie Huagong decoction(QJHGD)in intervening the intestinal barrier damage in rats with severe acute pancreatitis by using high⁃throughput RNA⁃Seq.Methods:A SAP model was established by retrograde pancreaticobiliary injection of sodium oxybate,HE was used to observe the histopathological chang⁃es in the pancreas and ileum of rats in the blank group,model group and traditional Chinese medicine group,and the levels of se⁃rum amylase,lipase,DAO and endotoxin in rats were detected by ELISA method;the differentially expressed genes were detect⁃ed in ileal tissues of rats in each group by RNA⁃seq sequencing,so as to screen the core genes,biological functions and signaling pathways of the QJHGD for intervening in the intestinal barrier damage in rats with severe acute pancreatitis.Results:Compared with the blank group,the pancreas and ileum tissues of rats in the model group showed obvious pathological changes,which were significantly improved by the intervention of QJHGD;the ELISA results suggested that QJHGD could significantly improve the serum amylase,lipase,DAO and endotoxin levels in rats with SAP(P<0.05).The RNA⁃Seq results showed that there were 2623 genes differentiated in the SAP group from the BG group,and 175 genes were significantly down⁃regulated after QJHGD treatment,mainly Tnip3 and Lpo,while 320 genes were up⁃regulated,mainly aicda and Tnn,and so on.There were 175 genes whose expression was significantly down⁃regulated after QJHGD treatment,mainly Tnip3,Lpo,etc.,while 320 genes were up⁃regulated,mainly aicda,Tnn,etc.,which were involved in the regulation of biological processes such as oxidative stress and intervened in multiple signaling pathways such as TJ,apoptosis,Toll⁃like receptor,NOD⁃like receptor,NF⁃κB and so on,and played a role in the regulation of oxidative stress.Conclusion:QJHGD may inhibit the intestinal barrier injury of severe acute pan⁃creatitis by regulating key genes such as Tnip3 and aicda,as well as multiple signaling pathways such as TJ,apoptosis,and Toll⁃like receptor.