益气、化瘀、化痰中药制剂及其合剂对肾间质纤维化大鼠TGF-β1/Smad通路及TRB3的影响

Effect of Yiqi，Huayu，Huatan preparation and their mixed preparation on TGFβ1/Smad pathway and TRB3 in rats with renal interstitial fibrosis

摘要:

目的：观察益气、化瘀、化痰中药制剂及其合剂———益气化瘀化痰制剂对单侧输尿管结扎（unilateral ureteral obstruc－tion，UUO）模型大鼠肾间质纤维化的影响及其可能机制。方法：42只雌性SD大鼠，随机分为假手术组、模型组、替米沙坦组、益气组、化瘀组、化痰组及益气化瘀化痰组，每组6只。除假手术组外，其余各组采用UUO法建立肾间质纤维化模型。UUO术后3周，模型组及假手术组以等量生理盐水灌胃，其余各组以相应制剂灌胃治疗，每日1次，连续用药12周后处死大鼠。检测大鼠血清尿素氮（blood urea nitrogen，BUN）、肌酐（serum creatinine，Scr）水平。取大鼠术侧肾组织，HE染色观察组织形态学改变，Masson染色观察胶原纤维沉积情况，Western blot检测转化生长因子-β1（transforming growth factor-β1，TGF-β1）、α-平滑肌肌动蛋白（α-smooth muscle actin，α-SMA）、E-钙黏蛋白（E-cadherin）蛋白表达水平；qRT-PCR 检测TGF-β1、Smad3、Smad7、TRB3、α-SMA、E-cadherin mRNA 表达水平。结果：与假手术组比较，模型组肾间质纤维化明显，BUN、Scr水平明显升高（P<0.01），而各药物干预组肾间质纤维化程度及BUN、Scr水平均较模型组明显改善，其中益气化瘀化痰组、替米沙坦组及益气组改善较明显（P<0.05）。与模型组相比，益气化瘀化痰组、替米沙坦组及益气组TGF-β1、Smad3、TRB3、α-SMA mRNA表达明显降低（P<0.05），E-cadherin mRNA表达明显升高（P<0.05）。各组间Smad7 mRNA表达无显著差异。TGF-β1、α-SMA、E-cadherin的蛋白表达与其mRNA表达趋势一致。结论：益气、化瘀、化痰及益气化瘀化痰中药制剂和替米沙坦均能改善UUO大鼠肾间质纤维化，且以益气化瘀化痰制剂、替米沙坦及益气制剂的改善作用较为明显，其机制可能是通过调控TRB3、TGF-β1/Smad信号通路，进而抑制肾小管上皮细胞向间充质细胞转化。

Abstract:

Objective：To observe the effect of Yiqi Huayu Huatan preparation and their mixed preparation on renal interstitial fibrosis in rats with unilateral ureteral obstruction（UUO） and investigate the possible mechanism. Methods：Totally 42 female sprague-dawley（SD） rats were randomly divided into sham operation group，model group，telmisartan group，Yiqi group，Huayu group，Huatan group and Yiqi Huayu Huatan group，with 6 rats in each group. Except for the sham operation group，the model was established by ligating left ureter of the rats. After 3 weeks，sham operation group and model group were treated with equal amount of normal saline，the other groups were treated with the corresponding decoction，once a day. After 12 weeks，the rats were sacrificed. The serum samples were collected for determination of concentrations of BUN and Scr. The morphological changes of renal tissue were observed by HE staining. The collagen fiber was observed by Masson staining. The protein expression levels of TGF-β1，α-SMA and E-cad－herin were detected by Western blot. The mRNA expression levels of TGF-β1，Smad3，Smad7，TRB3，α-SMA and E-cadherin were detected by qRT-PCR. Results：Compared with those in sham operation group，the renal interstitial fibrosis，the con－centrations of BUN and Scr were significantly increased in the model group（P<0.01），while the collagen fiber deposition and the levels of BUN and Scr were significantly reduced by Yiqi Huayu Huatan preparation，telmisartan and Yiqi preparation（P<0.05）. Compared with those in the model group，the expressions of TGF-β1，Smad3，TRB3，and α-SMA in Yiqi Huayu Huatan，telmisartan and Yiqi group were significantly decreased（P<0.05），while the expression of E-cadherin was significantly increased（P<0.05）. There was no significant difference in the expression of Smad7 among the groups. Conclusion：Yiqi，Huayu，Huatan，Yiqi Huayu Huatan preparation and telmisartan could alleviate renal interstitial fibrosis，but Yiqi Huayu Huatan preparation，telmisartan and Yiqi prepa－ration showed better curative effect. This may be related to the inhibition of epithelial-mesenehymal transition（EMT） mediated by TRB3 and TGF-β1/Smad signal pathway.