VSAP磷酸化介导紧密连接蛋白参与保护脓毒症肠上皮屏障功能

    Study on the VSAP phosphorylation mediating the tight junction protein to protect the intestinal epithelial barrier function in sepsis

    • 摘要:
      目的 验证紧密连接蛋白ZO-1、Claudin-5和Occludin保护脓毒症肠屏障功能与血管扩张刺激磷蛋白(vasodilator-stimulated phosphoprotein, VASP)相关。
      方法 盲肠结扎穿孔术构建脓毒症小鼠模型,实验分为:对照组、假手术组、模型组和VASP磷酸化(p-VASP)激动剂Bucladesine组。ELISA检测血清中二胺氧化酶(DAO)和D-乳酸(D-LA)含量。HE染色观察小肠黏膜组织形态。TUNEL染色检测小肠上皮细胞凋亡。Western blotting检测Bax、Bcl-2、p-VASP、ZO-1、Occludin和claudin-5蛋白表达水平。
      结果 与对照组和假手术组相比,模型组中小肠绒毛数量减少、绒毛断裂、TUNEL阳性细胞增加、DAO和D-LA含量升高(P < 0.05),Bax蛋白、p-VASP蛋白表达水平升高(P < 0.05),ZO-1、Occludin和claudin-5蛋白、Bcl-2蛋白表达水平降低(P < 0.05)。与模型组相比,Bucladesine组中小肠绒毛数量增加、绒毛断裂减少、TUNEL阳性细胞减少、DAO和D-LA含量降低(P < 0.05),p-VASP蛋白表达水平降低(P < 0.05),ZO-1、Occludin和claudin-5蛋白表达水平升高(P < 0.05)。
      结论 抑制脓毒症小鼠p-VASP后小肠上皮屏障功能恢复,这与上调紧密连接蛋白表达水平有关。

       

      Abstract:
      Objective To verify that the tight junction proteins ZO-1, Claudin-5 and Occludin in protecting intestinal barrier function in sepsis were associated with vasodilator-stimulated phosphoprotein(VASP).
      Methods A mouse model of sepsis was constructed by cecum ligation perforation, and the rats were divided into the control group, sham-operated group, model group and Bucladesine of VASP phosphorylation agonist group.The serum levels of diamine oxidase(DAO) and D-lactic acid(D-LA) were measured by ELISA.HE staining was performed to observe the histomorphology of the small intestinal mucosa.TUNEL staining was used to detect the apoptosis of small intestinal epithelial cells.Bax, Bcl-2, p-VASP, ZO-1, Occludin and claudin-5 protein expression levels were detected by Western blotting.
      Results Compared with the control and sham-operated groups, the number of small intestinal villi reduced, the villi broken, the TUNEL-positive cells increased, the levels of DAO and D-LA significantly increased(P < 0.05), the levels of p-VASP protein expression significantly increased(P < 0.05), and the levels of ZO-1, Occludin and claudin-5 protein expression significantly decreased in the model group(P < 0.05).Compared with the model group, the number of small intestinal villi increased, the villi breakage decreased, the TUNEL-positive cells decreased, the DAO and D-LA content significantly decreased(P < 0.05), the p-VASP and Bax protein expression levels significantly decreased(P < 0.05), and the expression levels of ZO-1, Occludin, claudin-5 and Bcl-2 protein increased in the Bucladesine group(P < 0.05).
      Conclusions The function of small intestinal epithelial barrier recovers after inhibiting the p-VASP in sepsis mice, which is related to up-regulation of tight junction protein expression.

       

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