温阳振衰颗粒对肾间质纤维化模型大鼠细胞焦亡及肾间质纤维化的影响

Effects of Wenyang Zhenshuai Granules on Pyroptosis and Renal Interstitial Fibrosis in Renal Interstitial Fibrosis Model Rats

目的观察温阳振衰颗粒对肾间质纤维化大鼠细胞焦亡及肾间质纤维化的影响,从NLRP3/Caspase-1通路探讨其干预机制。方法50只SD大鼠随机分为空白组、假手术组、模型组、抑制剂组和温阳振衰颗粒组,每组10只。以结扎并剪断左侧输尿管建立肾间质纤维化大鼠模型,分别予相应药物干预,连续14 d。试剂盒检测血肌酐(SCr)、血尿素氮(BUN)含量,HE和Masson染色观察肾组织病理变化,免疫组化检测肾组织转化生子因子(TGF)-β1、α-平滑肌肌动蛋白(α-SMA)、Ⅰ型胶原蛋白(Col-Ⅰ)表达,Westernblot和RT-PCR检测肾组织NOD样受体蛋白3(NLRP3)、半胱氨酸蛋白酶-1(Caspase-1)、白细胞介素(IL)-18、IL-1β蛋白和mRNA表达,免疫荧光双染法检测细胞焦亡。结果与假手术组比较,模型组大鼠SCr、BUN含量显著增加(P<0.01),肾小管扩张,炎性细胞浸润,间质纤维化明显,肾组织TGF-β1、α-SMA、Col-Ⅰ蛋白表达显著升高(P<0.01),NLRP3、Caspase-1、IL-18、IL-1β蛋白和mRNA表达显著升高(P<0.01),细胞焦亡率显著增加(P<0.01);与模型组比较,抑制剂组和温阳振衰颗粒组大鼠SCr、BUN含量显著减少(P<0.01),肾小管、肾小球损伤程度减轻,肾组织TGF-β1、α-SMA、Col-Ⅰ蛋白表达显著降低(P<0.01),NLRP3、Caspase-1、IL-18、IL-1β蛋白和mRNA表达显著降低(P<0.01),细胞焦亡率显著减少(P<0.01);抑制剂组与温阳振衰颗粒组比较差异无统计学意义。结论温阳振衰颗粒可能通过NLRP3/Caspase-1通路介导细胞焦亡,进而抑制肾纤维化,改善肾功能。

Objective To observe the effects of Wenyang Zhenshuai Granules on pyroptosis and renal interstitial fibrosis in renal interstitial fibrosis model rats;To discuss its intervention mechanism from the NLRP3/Caspase-1 pathway.Methods Totally 50 SD rats were randomly divided into blank group,sham-operation group,model group,inhibitor group and Wenyang Zhenshuai Granules group,with 10 rats in each group.The renal fibrosis model was established by ligating and cutting the left ureter.The rats were treated with corresponding drug respectively,for consecutive 14 days.Kit was used to detect serum creatinine(SCr)and blood urea nitrogen(BUN)contents;HE and Masson staining were used to observe the pathological change of renal tissue;the expressions of TGF-β1,α-SMA,and ColⅠwere detected by immunohistochemistry;the NOD-like receptor protein 3(NLRP3),cysteine protease-1(Caspase-1),interleukin(IL)-18,IL-1βprotein and mRNA expression in renal tissue were detected by Western blot and RT-PCR;double immunofluorescence staining was used to detect the pyrocytosis.Results Compared with the sham-operated group,the contents of SCr and BUN of the rats in model group significantly increased(P<0.01),the renal tubules dilated,infiltrated of inflammatory cells,and obvious interstitial fibrosis,the expression of TGF-β1,α-SMA,and ColⅠprotein in renal tissue significantly increased(P<0.01),the protein and mRNA expressions of NLRP3,Caspase-1,IL-18,IL-1βsignificantly increased(P<0.01),and the pyroptosis rate significantly increased(P<0.01).Compared with the model group,the contents of SCr and BUN of the rats in inhibitor group and Wenyang Zhenshui Granules group significantly reduced(P<0.01),the injury degree of renal tubules and glomeruli were alleviated,the expressions of TGF-β1,α-SMA and Col-Ⅰprotein in renal tissue significantly decreased(P<0.01),the protein and mRNA expressions of NLRP3,Caspase-1,IL-18,IL-1βsignificantly decreased(P<0.01),and the pyroptosis rate significantly reduced(P<0.01).There was no significant difference between the inhibitor group and Wenyang Zhenshuai Granules group.Conclusion Wenyang Zhenshuai Granules may mediate pyroptosis through NLRP3/Caspase-1 pathway,thereby inhibiting renal fibrosis and improving renal function.