灵芝多糖对腹膜透析大鼠Smad3/CTGF通路及腹膜纤维化的影响

Effect of Ganoderma Lucidum Polysaccharide on Smad3/CTGF Pathway and Peritoneal Fibrosis in Peritoneal Dialysis Rats

目的:探讨灵芝多糖对腹膜透析大鼠信号转导蛋白3(Smad3)/结缔组织生长因子(CTGF)通路及腹膜纤维化的影响。方法:采用5/6肾切除手术制备慢性肾功能衰竭大鼠模型,将造模成功的72只SD大鼠随机分为模型组、氯沙坦钾组(25mg·kg^(-1))、灵芝多糖低、高剂量组(40,80 mg·kg^(-1)),每组18只;另取18只大鼠作为正常对照组。氯沙坦钾组和灵芝多糖低、高剂量组灌胃给予相应药物,正常对照组和模型组给予等体积生理盐水,1次/d,连续给药4周。给药结束后,检测腹膜功能及厚度,观察腹膜结构病理变化,实时荧光定量PCR(qRT-PCR)法及蛋白免疫印迹(WB)法检测腹膜组织信号转导蛋白3(Smad3)、结缔组织生长因子(CTGF) mRNA和蛋白水平。结果:模型组腹膜间皮细胞为柱形、圆形,间皮细胞下基质增厚,部分脱落,且有大量炎症细胞浸润,伴纤维素样物质沉积;氯沙坦钾组和灵芝多糖高剂量组腹膜结构有所改善,间皮细胞排列较为整齐,柱形、圆形已向扁平间皮细胞转换,炎症细胞浸润减少,纤维素样物质沉积减少。与模型组比较,氯沙坦钾组、灵芝多糖低、高剂量组腹膜厚度、葡萄糖转运量、腹膜组织Smad3、CTGF mRNA和蛋白表达水平显著降低(P<0.05),超滤量水平显著升高(P<0.05);且灵芝多糖高、低剂量组间差异有统计学意义(P<0.05)。与氯沙坦钾组比较,灵芝多糖低剂量组腹膜厚度、葡萄糖转运量、腹膜组织Smad3、CTGF mRNA和蛋白表达水平显著升高(P<0.05),超滤量水平显著降低(P<0.05);灵芝多糖高剂量组差异无统计学意义(P>0.05)。结论:灵芝多糖对腹膜透析大鼠腹膜纤维化具有明显的抑制作用,其机制可能与灵芝多糖可抑制腹膜组织Smad3、CTGF mRNA和蛋白表达进而抑制Smad3/CTGF通路的激活有关。

Objective: To investigate the effect of ganoderma lucidum polysaccharide on Smad3/connective tissue growth factor(CTGF) pathway and peritoneal fibrosis in peritoneal dialysis rats. Methods: Chronic renal failure rat models were established by 5/6 nephrectomy in 72 SD rats. The successfully modeled rats were randomly divided into model group,losartan potassium group(25 mg·kg^(-1)),and ganoderma lucidum polysaccharide low and high dose groups(40 and 80 mg·kg^(-1)) with 18 rats in each group. Another 18 rats were taken as normal control group. Losartan potassium group,ganoderma lucidum polysaccharide low and high dose groups were given corresponding drug by gavage,and the normal control group and model group were given equal volume of normal saline,once a day for 4 weeks. After the administration,the function and thickness of peritoneum were tested and the pathological changes of peritoneum were observed. Real-time quantitative PCR(qRT-PCR) and Western blot(WB) methods were used to determine the mRNA and protein levels of Smad3 and CTGF in peritoneum. Results: The peritoneal mesothelial cells in the model group were columnar and round,and the submesothelial cells were thickened and partially shed,and there were a large number of inflammatory cells infiltrated with fibrin-like material deposition. The peritoneal structure of losartan potassium group and ganoderma polysaccharide high-dose group was improved,the mesothelial cells were arranged more neatly,the columnar and round shapes had been transformed into flat mesothelial cells,the infiltration of inflammatory cells was reduced,and the deposition of cellulose-like substances was reduced. Compared with those in the model group,the peritoneal thickness,glucose transport volume,peritoneal tissue Smad3,CTGF mRNA and protein expression levels of losartan potassium group and ganoderma lucidum polysaccharide low and high dose groups were significantly decreased(P<0.05),the ultrafiltration volume was significantly increased(P<0.05),and there were statistically significant differences between ganoderma lucidum polysaccharide low and high dose groups(P<0.05). Compared with those in losartan potassium group,the peritoneal thickness,glucose transport volume,peritoneal tissue Smad3,CTGF mRNA and protein expression levels of ganoderma lucidum polysaccharide low dose group were significantly increased(P<0.05),the ultrafiltration volume was significantly decreased(P<0.05),and there were no significant differences between the two ganoderma lucidum polysaccharide groups(P>0.05). Conclusion: Ganoderma lucidum polysaccharides can significantly inhibit peritoneal fibrosis in peritoneal dialysis rats,and the mechanism may be related to the expression inhibition of peritoneal tissue Smad3,CTGF mRNA and protein,and then the activation inhibition of Smad3/CTGF pathway by ganoderma lucidum polysaccharides.