冬凌草甲素对雨蛙素诱导的重症急性胰腺炎大鼠免疫紊乱和组织损伤的影响

Effect of oridonin on severe acute pancreatitis and tissue damage induced by ranin in rats

关键词:  冬凌草甲素  重症急性胰腺炎  组织损伤

Key words:  oridonin  severe acute pancreatitis  tissue damage

摘要:

[目的] 探讨冬凌草甲素（Ori）在重症急性胰腺炎（SAP）大鼠免疫紊乱和组织损伤中的调节作用。[方法] 将50只SD雄性大鼠随机分为对照组、模型组（SAP组）、SAP+Ori 5 mg/kg组、SAP+Ori 10 mg/kg组、SAP+Ori 20 mg/kg组，每组10只。采用化学诱导法建立SAP大鼠模型，皮下注射雨蛙素（25 μg/kg）诱导SAP；计算胰腺质量指数，酶联免疫吸附（ELISA）法测定血清脂肪酶、淀粉酶水平，苏木精-伊红（HE）染色观察胰腺组织病理形态学变化；免疫组化观察半胱氨酸天冬氨酸蛋白酶-3（Caspase-3）蛋白表达；蛋白免疫印记（Western Blot）法检测B淋巴细胞瘤-2（Bcl-2）相关X蛋白（Bax）/Bcl-2蛋白表达；ELISA法测定血清肿瘤坏死因子-α（TNF-α）、诱导型一氧化氮合酶（iNOS）、白细胞介素-10（IL-10）表达水平；实时定量荧光聚合酶链反应（RT-PCR）法检测胰腺组织白细胞介素-1β（IL-1β）、白细胞介素-4（IL-4）mRNA表达水平。[结果] 与对照组比较，SAP组大鼠胰腺组织功能明显下降（P＜0.05），组织形态紊乱，Caspase-3蛋白高表达，凋亡指标（Bax/Bcl-2）显著增高（P＜0.05），血清中TNF-α、iNOS、IL-10蛋白高表达（P＜0.05），胰腺组织中IL-1β、IL-4 mRNA高表达（P＜0.05）。与SAP组比较，5 mg/kg Ori给药治疗后，胰腺组织功能无明显差异；10、20 mg/kg Ori显著改善胰腺组织功能（P＜0.05），降低细胞凋亡率，下调Caspase-3、Bax蛋白表达，上调Bcl-2蛋白表达（P＜0.05），抑制促炎因子TNF-α、iNOS蛋白表达和IL-1β mRNA表达（＜i＞P＜/i＞＜0.05），促进抗炎因子IL-10蛋白表达和IL-4 mRNA表达（P＜0.05）。[结论] Ori抑制胰腺组织病理损伤、凋亡蛋白表达及促炎因子的分泌，缓解SAP大鼠免疫紊乱和组织损伤。

Abstract:

[Objective] To explore the regulatory role of oridonin in severe acute pancreatitis of immune disorders and tissue damage in rats. [Methods] Fifty SD male rats were randomly divided into healthy control group,model group(SAP),SAP+Ori5 mg/kg group,SAP+Ori10 mg/kg group,SAP+Ori20 mg/kg group,10 rats in each group. A rat model of severe acute pancreatitis was established by chemical induction,and acute pancreatitis was induced by subcutaneous injection of caerulein(25 μg/kg). Calculate pancreatic weight index,measure serum lipase and amylase levels by enzyme-linked immunosorbent assay(ELISA),observe the pathological changes of pancreatic tissue by hematoxylin-eosin(HE) staining;observe the expression of caspase-3 protein by immunohistochemistry;Western Blot detection of Bax/Bcl-2 protein expression;ELISA detection of serum tumor necrosis factor-α(TNF-α),inducible nitric oxide synthase(iNOS),interleukin 10(IL-10) expression levels;RT-PCR was used to detect the expression levels of interleukin-1β(IL-1β) and interleukin-4(IL-4) mRNA in pancreatic tissue. [Results] Compared with the control group,the pancreatic tissue function of the model group was significantly decreased(P<0.05),the tissue morphology was disordered,Caspase-3 protein was highly expressed,and the apoptosis index(Bax/Bcl-2) was significantly increased(P<0.05). TNF-α,iNOS,IL-10 protein were highly expressed in serum(P<0.05),and IL-1β and IL-4 mRNA were highly expressed in pancreatic tissue(P<0.05). Compared with the model group,there was no significant difference in pancreatic tissue function after 5 mg/kg oridonin was administered;10 and 20 mg/kg oridonin significantly improved pancreatic tissue function(P<0.05) and decreased apoptosis rate,down-regulation of caspase-3,bax protein expression,up-regulation of Bcl-2 protein expression(P<0.05),inhibition of pro-inflammatory factors TNF-α,iNOS protein expression and IL-1β mRNA expression(P<0.05),and promote anti- Inflammatory factor IL-10 protein expression and IL-4 mRNA expression(P<0.05). [Conclusion] Oridonin inhibits the pathological damage of pancreatic tissue,the expression of apoptotic proteins and the secretion of pro-inflammatory factors,and alleviates the immune disorder and tissue damage in rats with severe acute pancreatitis.