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目的:探讨黄芪甲苷(AS-IV)抑制转化生长因子β1(TGF-β1)诱导肾小管上皮细胞凋亡的机制。方法:体外采用TGF-β1(10 ng/m1)刺激人近端小管上皮细胞(HK-2),采用不同浓度的AS-IV(50μg/rl、100μg/ml、200μg/ml)进行干预治疗24h收集细胞。体内建立小鼠单侧输尿管梗阻(UUO)模型,C57BL/6小鼠随机分为假手术组(Sham组)、模型组(UUO组)和治疗组(AS-IV组)。采用单侧(右)输尿管结扎建立UUO模型,Sham组仅游离输尿管但不结扎,AS-IV组连续7天给予AS-Ⅳ[20mg/(kg·d)]灌胃,UUO组和Sham给予等剂量溶剂灌胃,术后第14天采集肾脏组织。TUNEL法检测凋亡,CCK法观察细胞活性,Western印迹检测TGF-β1和被切割的半胱氨酸蛋白酶3(cleaved-caspase 3)表达,并观察c-Jun氨基末端激酶(JNK)丝裂原活化蛋白激酶(MAPK)磷酸化情况。结果:AS-Ⅳ改善UUO诱导的肾组织TGF-β1表达及细胞凋亡(P<0.01),抑制TCF-β1诱导的HK-2细胞活性下降及细胞凋亡,200μg/mlAS-Ⅳ作用最明显(P<0.01)。体内体外实验均可见,AS-IV减少caspase 3活化,抑制JNK MAPK磷酸化。结论:AS-IV可抑制TGF-β1的表达及其诱导的肾小管上皮细胞凋亡,其机制可能与JNK MAPK通路有关。
Objective: To investigate the mechanism of Astragaloside IV (AS-IV) inhibiting the apoptosis of renal tubular epithelial cells induced by transforming growth factor-β1 (TGF-β1). Methods: Human proximal tubular epithelial cells (HK-2) were stimulated with TGF-β1 (10 ng / ml) and AS-IV (50 μg / ml, 100 μg / ml and 200 μg / ml) Collect cells. The unilateral ureteral obstruction (UUO) model was established in vivo. C57BL / 6 mice were randomly divided into sham operation group (Sham group), model group (UUO group) and treatment group (AS-IV group). UUO model was established by unilateral (right) ureteral ligation, while the Sham group was only free of the ureter but not ligated. The AS-IV group was administered with AS-IV [20mg / (kg · d) The dose of solvent gavage, kidney tissue collected after 14 days. The apoptosis was detected by TUNEL method. The cell viability was observed by CCK method. The expression of TGF-β1 and cleaved-caspase 3 was detected by Western blotting. The c-Jun N-terminal kinase (JNK) mitogen Activated protein kinase (MAPK) phosphorylation. Results: AS-Ⅳ could improve UUO-induced TGF-β1 expression and apoptosis (P <0.01), inhibit TCF-β1-induced HK-2 cell activity and apoptosis, and 200μg / ml AS- (P <0.01). In vitro and in vivo experiments were seen, AS-IV reduced caspase 3 activation, inhibition of JNK MAPK phosphorylation. CONCLUSION: AS-IV can inhibit the expression of TGF-β1 and induce the apoptosis of renal tubular epithelial cells, which may be related to the JNK MAPK pathway.