参附注射液对脓毒症大鼠肠黏膜屏障的影响及机制研究

Effects and Mechanism Study of Shenfu Injection on Intestinal Mucosal Barrier Function in Rats with Sepsis

目的观察参附注射液对脓毒症大鼠肠黏膜屏障功能的影响。方法将40只大鼠随机分为假手术组、脓毒症组、参附注射液低剂量组和高剂量组,每组各10只。行盲肠结扎穿孔术制备重度脓毒症模型,假手术组仅行开腹手术后即予关闭腹腔。术后10 min内经尾静脉推注药物:模型组给予生理盐水10 mL/kg;参附低剂量组给予5 mL/kg参附注射液+生理盐水5 m L/kg稀释液;参附高剂量组给予10 mL/kg参附注射液。8 h后处死大鼠,以ELISA检测各组大鼠血清肿瘤坏死因子-ɑ(TNF-ɑ)及白细胞介素-10(IL-10)浓度、回肠组织丙二醛(MDA)含量和超氧化物歧化酶(SOD)活性,HE染色观察回肠组织病理改变(Chiu′s评分),TUNEL法检测回肠黏膜上皮细胞凋亡率(AI),免疫组化检测促凋亡蛋白Bax与抗凋亡蛋白Bcl-2,Western blotting检测跨膜紧密连接蛋白occludin水平。结果与假手术组比较,模型组INF-α、MDA、Chiu′s评分、AI及Bax均显著增高,IL-10、SOD、Bcl-2、ooccludin均显著降低,差异均有统计学意义(P<0.05);与模型组比较,参附低剂量组、参附高剂量组TNF-α、MDA、Chiu′s评分、AI均显著降低,IL-10、SOD显著增高,差异均有统计学意义(P<0.05);与参附低剂量组比较,参附高剂量组TNF-α、MDA、Chiu′s评分、AI、Bax均显著降低,IL-10、SOD、Bcl-2、occludin均显著增高,差异均有统计学意义(P<0.05)。结论参附注射液可显著改善重度脓毒症模型大鼠的肠黏膜屏障功能,其作用与参附注射液的剂量成正相关。

Objective:To observe the effect of Shenfu Injection on the intestinal mucosal barrier function in rats with sepsis.Methods:40 rats were randomly divided into the sham operation group(Sham),sepsis group(CLP),Shenfu Injection low-dose group(LSF)and high-dose group(HSF),with 10 rats in each group.In the Sham group,the abdominal cavity was closed after only laparotomy,and 10 m L/kg of normal saline was injected via the tail vein within 10 minutes;the other 3 groups all underwent cecal ligation and perforation to prepare severe sepsis models,and cecal ligation was performed in the rats.Different drugs were injected via tail vein according to body weight within 10 minutes after perforation:CLP group was given normal saline 10 mL/kg;LSF group was given 5mL/kg Shenfu Injection+normal saline 5 mL/kg diluent;HSF group was given 10 m L/kg Shenfu Injection.After 8hours,the rats were sacrificed.The serum TNF-αand IL-10 concentrations,ileal tissue malondialdehyde(MDA)content and superoxide dismutase(SOD)activity were detected by ELISA.The change of ileal tissue pathology was observed by HE staining(Chiu′s score),TUNEL method was used to detect ileal mucosal epithelial cell apoptosis(AI),immunohistochemistry was used to detect pro-apoptotic protein Bax and anti-apoptotic protein Bcl-2,and Western blotting was used to detect the level of transmembrane tight junction protein occludin.Results:Compared with the Sham group,TNF-α,MDA,Chiu′s score,AI and Bax in the CLP group were significantly increased,IL-10,SOD,Bcl-2,and ooccludin were significantly decreased,and the differences were statistically significant(P<0.05);compared with the CLP group,TNF-α,MDA,Chiu′s scores and AI in the LSF group and HSF groupwere significantly reduced,IL-10 and SOD increased significantly,and the difference was statistically significant(P<0.05);compared with the LSF group,the TNF-α,MDA,Chiu′s score,AI,and Bax in the HSF group were sig-nificantly reduced,IL-10,SOD,Bcl-2,and ooccludin increased significantly,and the difference was statisticallysignificant(P<0.05).Conclusion:Shenfu Injection can significantly improve the intestinal mucosal barrier func-tion of severe sepsis model rats,and its effect is positively related to the dose of Shenfu Injection.