抑制线粒体钙单向转运体对大鼠急性胰腺炎氧化应激的作用研究

Effects of inhibiting the mitochondrial calcium uniporter on the oxidative stress in rats with acute pancreatitis

摘要:

目的 探讨抑制线粒体钙单向转运体（MCU）减弱线粒体钙超载对雨蛙肽诱导的大鼠急性胰腺炎（AP）及胰腺氧化应激的影响。方法 将32只雄性SD大鼠分为对照组、急性胰腺炎组（AP组）、急性胰腺炎+钌红干预组（AP+RR组）、钌红组（RR组），每组8只。模型复制24 h后处置大鼠，检测各组大鼠血清淀粉酶（Amy）、脂肪酶（Lipase）、白细胞介素6（IL-6）水平，HE染色观察胰腺组织病理改变；倒置荧光显微镜下观察腺泡细胞活性氧（ROS）、线粒体内Ca²⁺、胞内游离Ca²⁺荧光强度；检测胰腺组织还原型谷胱甘肽（GSH）、丙二醛（MDA）含量；Western blotting检测胰腺组织MCU、沉默信息调节因子3（SIRT3）、MnSOD蛋白的表达；透射电镜观察腺泡细胞超微结构。结果 各组大鼠胰腺HE评分和血清Amy、Lipase、IL-6水平比较，经方差分析，差异有统计学意义（P <0.05），AP组较对照组高（P <0.05）。AP组MCU相对表达量，线粒体内Ca²⁺、胞内游离Ca²⁺浓度高于对照组、RR组（P <0.05），AP+RR干预组低于AP组（P <0.05）。AP组MnSOD、SIRT3、GSH低于对照组（P <0.05），ROS、MDA高于对照组（P <0.05），与AP组比较，AP+RR干预组ROS、MDA低于AP组（P <0.05），MnSOD、SIRT3、GSH高于AP组（P <0.05）。结论 抑制MCU可减弱腺泡钙超载，同时可能通过调节SIRT3/MnSOD途径改善雨蛙肽诱导的AP氧化应激，但对胰腺组织病理学和生化指标无明显改变。

Abstract:

Objective To investigate the effects of attenuating the mitochondrial calcium overload by inhibiting the mitochondrial calcium uniporter (MCU) on the oxidative stress in rats with acute pancreatitis (AP) induced by caerulein (CAE).Methods A total of 32 male SD rats were divided into control group, AP group, AP plus ruthenium red group (AP + RR group) and ruthenium red group (RR group), with 8 cases in each group. The rats were sacrificed 24 hours after establishing the models, and the serum levels of amylase (Amy), lipase and interleukin-6 (IL-6) were detected. The histopathological changes of the pancreas were observed by HE staining. The fluorescence intensity of reactive oxygen species (ROS), mitochondrial calcium and cytoplasmic calcium in the acinar cells was observed under the inverted fluorescence microscope. The contents of glutathione (GSH) and malondialdehyde (MDA) in the pancreatic tissues were measured. The relative protein expressions of MCU, SIRT3 and MnSOD in the pancreatic tissues were detected by Western blotting. The ultrastructure of acinar cells was observed under the transmission electron microscope.Results The HE scores and serum levels of Amy, lipase and IL-6 were different among the groups (P < 0.05), and they were higher in the AP group compared with those in the control group (P < 0.05). The relative expression of MCU, and the levels of mitochondrial and cytoplasmic calcium were higher in the AP group relative to those in the control group and RR group (P < 0.05), while they were lower in the AP + RR group compared with the AP group (P < 0.05). The expressions of MnSOD and SIRT3 and the content of GSH were lower (P < 0.05), but the fluorescence intensity of ROS and the content of MDA were higher in the AP group compared with the control group (P < 0.05). In addition, the fluorescence intensity of ROS and the content of MDA were lower (P < 0.05), but the expressions of MnSOD and SIRT3 and the content of GSH were higher in the AP + RR group compared with the AP group (P < 0.05).Conclusions The inhibition of MCU could reduce the calcium overload in acinar cells and attenuate the oxidative stress induced by caerulein in AP via regulating the SIRT3 / MnSOD pathway. However, it may not improve the histopathological and biochemical changes of the pancreatic tissues.