探讨血必净注射液治疗重症急性胰腺炎肺损伤的机制

Study on the mechanism of Xuebijing injection in treating severe acute pancreatitis with lung injury

目的:探讨血必净(XBJ)注射液对重症急性胰腺炎(SAP)大鼠肺损伤模型的治疗机制。方法:将80只雄性Sprauge-Dawley(SD)大鼠随机分为4组:对照组、SAP模型组、XBJ低剂量治疗组、XBJ高剂量治疗组,每组20只。除对照组,各组大鼠均经胰胆管匀速逆行注射4.5%牛磺胆酸钠溶液(0.1 mL/100 g,0.05 mL/min)以诱发SAP,造模成功30 min后,将XBJ低、高剂量治疗组大鼠经尾静脉注射XBJ(剂量分别为5、20 mL/kg),对照组及模型组给予等体积生理盐水。24 h后,从每组中随机抽取10只大鼠,经尾静脉注射Evans blue检测肺组织毛细血管通透性。处死余下的各组大鼠,取腹水测量其体积;取部分胰腺及肺组织计算组织干湿重比值;苏木精-伊红(HE)染色观察胰腺、肺脏的病理改变,并进行病理评分;酶联免疫吸附法(ELISA)检测腹水淀粉酶含量;Western印迹法检测肺组织中ROCK1、MYPT1、pMLC的相对表达量。结果:SAP模型组较对照组腹水量及腹水淀粉酶含量明显升高(t=-21.73、-40.72,均P<0.01);SAP模型组、XBJ低剂量组、XBJ高剂量组腹水量及腹水淀粉酶含量逐渐降低(F=39.66、141.78,均P<0.01)。SAP模型组较对照组,胰腺干湿重比值、肺脏干湿重比值明显降低(t=19.83、15.47,均P<0.01),肺组织中Evans blue渗出量明显增高(t=-27.9,P<0.01);SAP模型组、XBJ低剂量组、XBJ高剂量组胰腺干湿重比值、肺脏干湿重比值逐渐升高(F=75.19、15.47,均P<0.01),肺组织中Evans blue渗出量逐渐降低(F=99.52,P<0.01)。对照组大鼠胰腺组织结构完整,SAP模型组大鼠胰腺病理得分明显升高(t=-42.79,P<0.01);XBJ低剂量组、XBJ高剂量组大鼠胰腺病理得分逐渐降低(F=175.43,P<0.01)。对照组大鼠肺组织结构完整,SAP模型组大鼠病理得分明显升高(t=-37.57,P<0.01);XBJ低剂量组、XBJ高剂量组大鼠肺组织病变减轻,病理得分逐渐降低(F=126.00,P<0.01)。SAP模型组较对照组肺组织中ROCK1、pMLC蛋白表达量明显升高(t=-16.97、-13.53,均P<0.01),MYPT1表达量显著降低(t=23.30,P<0.01);SAP模型组、XBJ低剂量组、XBJ高剂量组肺组织中ROCK1、pMLC蛋白表达量逐渐降低(F=84.89、50.84,均P<0.01),MYPT1表达量显著升高(F=48.68,P<0.01)。结论:XBJ通过降低多种细胞因子及DAMPs形成,抑制ROCK1-MYPT1-pMLC信号通路活化,对SAP肺损伤起到治疗的作用。

Objective:To investigate the therapeutic mechanism of Xuebijing(XBJ)injection on lung injury model of severe acute pancreatitis(SAP)in rats.Methods:A total of 80 male Spruge Dawley(SD)rats were randomly divided into 4 groups:control group,SAP model group,XBJ low-dose treatment group and XBJ high-dose treatment group,with 20 rats in each group.Except for the control group,the rats in all groups were injected with 4.5%sodium taurocholate solution(0.1 mL/100 g,0.05 mL/min)at uniform speed retro-grade through the pancreatic bile duct to induce SAP.30 min after successful molding,the rats in the low-dose and high-dose XBJ treat-ment groups were injected with XBJ injection through the tail vein(the dose was 5 and 20 mL/kg,respectively).Control group and mod-el group were given equal volume normal saline.After 24 h,10 rats were randomly selected from each group,and Evans blue was injected into the tail vein to detect the capillary permeability of lung tissue.The remaining rats were killed and their volume was measured with ascites.The dry-wet weight ratio was calculated by taking part of pancreas and lung tissues.Hematoxylin-eosin(HE)staining was used to observe the pathological changes of pancreas and lung,and pathological score was performed.The amylase content of ascites was detected by enzyme-linked immunosorbent assay(ELISA).The relative expression levels of ROCK1,MYPT1 and pMLC in lung tissues were detected by Western blotting.Results:Compared with the control group,the water volume and amylase content of ascites in SAP model group were significantly increased(t=-21.73,-40.72,both P<0.01).Compared with SAP model group,XBJ low-dose group and XBJ high-dose group,the water volume and amylase content of ascites gradually decreased(F=39.66,141.78,both P<0.01).Compared with the control group,the dry-wet weight ratio of pancreas and dry-wet weight ratio of lung were significantly de-creased in SAP model group(t=19.83,15.47,both P<0.01),and the Evans blue exudation amount in lung tissue was significantly in-creased(t=-27.9,P<0.01).Compared with SAP model group,XBJ low-dose group and XBJ high-dose group,the dry-wet weight ratio of pancreas and dry-wet weight ratio of lung were gradually increased(F=75.19,15.47,both P<0.01),and the Evans blue exudation amount in lung tissue was gradually decreased(F=99.52,P<0.01).The pancreatic tissue structure of the control group was intact,and the pancreatic pathological score of the SAP model group was significantly increased(t=-42.79,P<0.01).The pathological scores of the pancreas in the low-dose group and the high-dose group of XBJ decreased gradually(F=175.43,P<0.01).The lung structure of the con-trol group was intact,and the pathological score of the SAP model group was significantly increased(t=-37.57,P<0.01).Compared with the low-dose group of XBJ and the high-dose group of XBJ,the lung tissue lesions were reduced,and the pathological scores were de-creased gradually(F=126.00,P<0.01).Compared with the control group,the expression levels of ROCK1 and pMLC protein in SAP model group were significantly increased(t=-16.97,-13.53,both P<0.01),and the expression level of MYPT1 was significantly decreased(t=23.30,P<0.01).Compared with SAP model group,XBJ low-dose group and XBJ high-dose group,the expression lev-els of ROCK1 and pMLC protein in lung tissue gradually decreased(F=84.89,50.84,both P<0.01),while the expression levels of MYPT1 were significantly increased(F=48.68,P<0.01).Conclusion:XBJ plays a therapeutic role in SAP lung injury by reducing the formation of various cytokines and DAMPs,inhibiting the activation of ROCK1-MYPT1-pMLC signaling pathway.