含蛋白转导域的SARA/SBD融合蛋白对腹膜透析大鼠腹膜结构和功能的作用

Effect of PTD-SARA/SBD Fusion Protein on Peritoneal Morphology and Function of Peritoneal Dialysis Rats

目的:探讨含蛋白转导域的SARA融合蛋白(PTD-SAR/SBD)对腹膜透析大鼠腹膜纤维化的抑制作用。方法:每日腹腔注射4.25%葡萄糖腹膜透析液(PDF)制备腹膜透析大鼠模型。28只Sprague-Dawley大鼠随机分为正常对照组(n=8);腹膜透析模型组(n=10);PTD-SARA/SBD蛋白干预组(n=10)。4周后行腹膜平衡试验,检测超滤量、葡萄糖吸收率;留取壁层腹膜组织行HE染色;免疫组织化学方法检测大鼠壁层腹膜间皮细胞转分化指标E-cadherin和Twist的表达;Western blotting测定E-cadherin、twist,以及CollagenⅠ、TGF-β1、P-Smad3、t-Smad3的表达。结果:①与正常对照组比较,模型组大鼠壁层腹膜增厚,糖转运率增加,超滤量降低(P<0.01);免疫组织化学与western blotting检测结果显示E-cadherin表达下调,Twist表达上调;CollagenⅠ、P-Smad3、TGF-β1表达增加。②与模型组比较,PTD-SARA/SBD蛋白干预组大鼠壁层腹膜糖转运率降低,超滤量增加(P<0.05);E-cadherin表达上调,Twist表达下调;CollagenⅠ、TGF-β1、P-Smad3表达减少,各组t-Smad3无明显变化。结论:PTD-SARA/SBD融合蛋白通过抑制TGF-β/Smads信号通路部分逆转腹膜间皮细胞转分化从而改善腹膜结构和功能,为防治腹膜透析所致腹膜纤维化奠定基础。

Objective： To explore the prevention role of PTD-SARA/SBD fusion protein on peritoneal fibrosis induced by peritoneal dialysis in rats. Methods： 28 male Sprague-Dawley rats were randomly divided into three groups： normal control group（n=8）; peritoneal dialysis model group（n=10）; PTD-SARA/SBD group （n=10）. The animals were infused by intra-peritoneal injection and were killed at the end of 4 weeks. A 4 hour peritoneal equilibration test was performed after four-weeks peritoneal dialysis in each group. The dialysate samples were subjected to determination of 4h ultrafiltration volume （UF） or glucose transporion. The parietal peritoneum tissues of rats were harvested and stained with H-E for observation of the peritoneal structure. The expression of E-cadherin, Twist, TGF-β1, t-Smad3, P-Smad3, and Collagen Ⅰ in parietal peritoneum was detected with immunohistochemistry or Western blotting assay. Results： Compared with the normal Control group, the UF were lower, and glucose transporion were higher in model group （P〈0.01）. The results of western blotting and Immunohistochemistry showed that the level of E-cadherin was down-regulatied, and Twist expression was up regulation in parietal peritoneum tissues, and TGF-β1, P-Smad, Collagen Ⅰ expression increased in the model group; Administration of PTD-SARA/SBD fusion protein improved the function and structure changes of the peritoneum, improved the level of E-cadherin and decreased the level of Twist. The expression of TGF-β1, P-Smad3, Collagen Ⅰin parietal peritoneum were decreased in PTD-SARA/SBD group. Conclusion： PTD-SARA/SBD fusion protein may reverse the epithehal mesenehymal transition by inhibiting TGF-β/Smads signaling and improve the peritoneal morphology and function, which lays a foundation of further study on prevention and treatment of peritoneal fibrosis induced by peritoneal dialysis.