紧密连接蛋白1与重症急性胰腺炎大鼠胰腺微血管损伤关系的实验研究

目的研究重症急性胰腺炎（severe acute pancreatitis,SAP）大鼠紧密连接蛋白1（ZO-1）随时间的变化及其与SAP微血管损伤和胰腺组织病理学评分的关系。方法将48只Wistar大鼠随机均分为假手术组（SO组）和SAP组。SO组大鼠开腹后仅翻动胰腺;SAP组大鼠开腹后,以逆行胰胆管微泵注射5%牛磺胆酸钠法制备SAP模型。2组大鼠分别于手术后6、12及24 h各处死8只大鼠,取腹主动脉血测定外周血中淀粉酶、胰蛋白酶、白介素-8（IL-8）、肿瘤坏死因子-α（TNF-α）及ZO-1蛋白水平;取胰腺组织行HE染色,观察其病理学变化并评分;同时行免疫组化染色检测ZO-1蛋白的表达情况。结果同时点与SO组比较,各时点SAP组的血清淀粉酶、胰蛋白酶、IL-8、TNF-α及ZO-1蛋白水平均较高（P〈0.05）。SAP-6 h组和12 h组的血清淀粉酶水平均低于24 h组（P〈0.05）;SAP组内大鼠的胰蛋白酶、IL-8和ZO-1蛋白水平均随时点延长逐渐升高,各时点组间两两比较差异均有统计学意义（P〈0.05）;SAP组内3个时点组间TNF-α水平的差异无统计学意义（P〉0.05）。多重线性回归模型结果显示,SAP组血清ZO-1蛋白水平与胰腺组织病理学评分（b=0.96,P〈0.05）、血清淀粉酶水平（b=0.87,P〈0.05）、胰蛋白酶水平（b=0.72,P〈0.05）及IL-8水平（b=0.69,P〈0.05）均呈正相关,而与TNF-α水平的关系无统计学意义（P〉0.05）。HE染色结果显示,各时点SAP组大鼠的胰腺组织损伤程度重于SO组,且随时间延长,SAP大鼠胰腺组织的病理学损伤加重;SAP-12 h组和24 h组的胰腺组织病理学评分均高于6 h组（P〈0.05）。SP免疫组化染色结果显示,随时间延长,SAP组大鼠胰腺腺泡细胞间及毛细血管壁的ZO-1蛋白颗粒数量减少,且在毛细血管中的表达不连续。结论在SAP的进程中,血清中ZO-1蛋白的水平上升,同时其在胰腺组织中的表达呈下调趋势,表明其与SAP时的胰腺微血管损伤有关。

Experimental Study of Relationship of Zonula Occludens-1 and Microvascular Injury in Rats with Severe Acute Pancreatitis

Objective To investigate the changes of expression of zonula occludens-1（ZO-1） in rats with severe acute pancreatitis（SAP）, and to study the relationship between the ZO-1 protein and microvascular injury in rats with SAP. Methods Forty-eight Wistar rats were randomly divided into sham-operation（SO） group and SAP group, each group enrolled 24 rats. Pancreas of rats in SO group were fl ipped only after laparotomies, but rats of SAP group were injected with 5% sodium taurocholate by retrograding cholangiopancreatography micro pump to produce the SAP model.At 6, 12, and 24 hours after operation, 8 rats were sacrificed to get abdominal aortic blood for testing the levels of peripheral blood amylase, trypsin, interleukin-8（IL-8）, tumor necrosis factor-α（TNF-α）, and ZO-1 protein. At the same time, pancreatic tissues were got to perform HE staining and immunohistochemical staining for observation of the pathological changes and the expression of ZO-1 protein respectively. Results Compared with SO group at the same time, the levels of peripheral blood amylase, trypsin, IL-8, TNF-α, and ZO-1 protein were all higher in SAP group（P〈0.05）. The level of amylase in SAP-24 hours group was higher than those of 6 hours group and 12 hours group（P〈0.05）, the levels of trypsin, IL-8, and ZO-1 protein in SAP group increased over time（P〈0.05）, but levels of TNF-α in 3 time points of SAP group did not differ with each other significantly（P 0.05）. Results of regression showed that in the SAP group, the level of ZO-1 protein in serum was significantly positive correlated with pathological score of pancreatic tissue（b=0.96, P〈0.05）, levels of serum amylase（b=0.87, P〈0.05）, trypsin（b=0.72, P〈0.05）, and serum IL-8（b=0.69, P〈0.05）, but was not significantly correlated with level of TNF-α（P〈0.05）. HE staining results showed that damage of pancreatic tissues became worse over time in SAP group, and the pathological score of SAP-6 hours group was lower than those of 12