柴胡皂苷A调节MLCK/MLC2信号通路对SAP大鼠肠损伤的影响

Effect of saikosaponin A regulating MLCK/MLC2 signaling pathway on intestinal injury in SAP rats

目的探讨柴胡皂苷A调节肌球蛋白轻链激酶(MLCK)/肌球蛋白轻链2(MLC2)信号通路对重症急性胰腺炎(SAP)大鼠肠损伤的影响。方法随机选择10只大鼠作为假手术组,其余大鼠注射牛磺胆酸钠溶液构建SAP大鼠模型。将造模成功的SAP大鼠模型随机平分为SAP组、柴胡皂苷A组(腹腔注射10.0 mg/kg的柴胡皂苷A)、iE-DAP组(腹腔注射3.5 mg/kg MLCK/MLC2通路激活剂iE-DAP)、柴胡皂苷A+iE-DAP组(腹腔注射10.0 mg/kg柴胡皂苷A+3.5 mg/kg iE-DAP),每组均10只大鼠,每天1次,连续注射1周,假手术组和SAP组注射等量生理盐水。酶联免疫吸附试验(ELISA)检测各组大鼠血清淀粉酶(AMY)、脂肪酶(LIP)、二胺氧化酶(DAO)、白细胞介素(IL)-1β、IL-6和肿瘤坏死因子-α(TNF-α)水平;HE染色检测各组大鼠回肠组织病理形态变化。ELISA检测各组大鼠回肠组织中氧化应激指标水平。蛋白质免疫印迹检测肠道屏障相关蛋白及MLCK/MLC2通路相关蛋白表达。结果与SAP组相比,柴胡皂苷A组AMY、LIP、DAO、IL-1β、IL-6和TNF-α水平显著降低,而iE-DAP组AMY、LIP、DAO、IL-1β、IL-6和TNF-α水平显著升高,差异有统计学意义(P<0.05)。与SAP组相比,柴胡皂苷A组大鼠回肠组织结构得到改善,回肠组织病理评分显著降低(P<0.05)。与SAP组相比,柴胡皂苷A组谷胱甘肽、超氧化物歧化酶水平显著升高,丙二醛水平显著降低,差异有统计学意义(P<0.05)。与SAP组相比,柴胡皂苷A组MLCK、p-MLC2/MLC2蛋白水平显著降低,差异有统计学意义(P<0.05)。结论柴胡皂苷A可能通过下调MLCK/MLC2信号通路对SAP大鼠肠损伤起到改善作用。

Objective To investigate the effect of saikosaponin A regulating myosin light chain kinase(MLCK)/myosin light chain 2(MLC2)signaling pathway on intestinal injury in rats with severe acute pancreatitis(SAP).Methods A total of 10 rats were randomly selected as sham operation group,and the other rats were injected with sodium taurine cholate solution to construct SAP rat model.SAP rat models were randomly divided into SAP group,saikosaponin A group(10.0 mg/kg intraperitoneal injection of saikosaponin A)and iE-DAP group(3.5 mg/kg intraperitoneal injection of MLCK/MLC2 pathway activator iE-DAP),saikosaponin A+iE-DAP group(intraperitoneally injected with 10.0 mg/kg saikosaponin A+3.5 mg/kg iE-DAP),10 rats in each group were injected once a day for 1 week,sham operation group and SAP group were injected with the same amount of normal saline.The serum levels of amylase(AMY),lipase(LIP),diamine oxidase(DAO),interleukin(IL)-1β,IL-6 and tumor necrosis factor-α(TNF-α)were detected by enzyme-linked immunosorbent assay(ELISA).HE staining was used to det ect the pathological morphology of ileum tissue in each group.The levels of oxidative stress indexes in ileum tissue were detected by ELISA.Intestinal barrier related proteins and MLCK/MLC2 pathway related proteins were detected by western blot.Results Compared with SAP group,the levels of AMY,LIP,DAO,IL-1β,IL-6 and TNF-αin saikosonin A group were significantly decreased,while the levels of AMY,LIP,DAO,IL-1β,IL-6 and TNF-αin iE-DAP group were significantly increased,with statistical significance(P<0.05).Compared with SAP group,the structure of ileum tissue was improved and the pathological score of ileum tissue was significantly decreased in SA group(P<0.05).Compared with SAP group,the levels of glutathione and superoxide dismutase in SA group were significantly increased,and the levels of malondialdehyde were significantly decreased in SA group,with statistical significance(P<0.05).Compared with SAP group,the protein levels of MLCK,p-MLC2/MLC2 in SA group were significantly decreased,and the difference was statistically significant(P<0.05).Conclusion Saikosaponin A may improve intestinal injury in SAP rats by down-regulating the MLCK/MLC2 signaling pathway.