Sorafenib通过抑制TGF-β/Smad途径延缓肾纤维化的研究
Sorafenib ameliorates renal fibrosis through inhibition of TGF-β/Smad pathway

摘要：目的 探讨sorafenib减轻肾纤维化的作用及机制。方法 用低、中、高剂量sorafenib分别给单侧输尿管梗阻(UUO)模型大鼠灌胃或干预经TGF-β1刺激的NRK-52E细胞。HE染色观察各组肾组织纤维化情况，免疫荧光染色检测肾组织及NRK-52E细胞α-SMA、E-cadherin的表达情况。用流式细胞术测定各组NRK-52E细胞周期。Western blot检测各组NRK-52E细胞中Smad3和p-Smad3的表达变化。结果 HE染色结果显示，与UUO模型组相比，sorafenib治疗组肾间质纤维化明显减轻，小管萎缩、炎细胞浸润较轻(P<0.05)；与对照组比较，sorafenib治疗组E-cadherin在NRK-52E细胞和肾组织中表达均增加，而α-SMA表达均降低(P<0.05)；流式细胞术分析发现细胞周期停滞于G0/G1期的细胞数明显增加，而进入G2、S期的细胞数明显减少(P<0.05)；与对照组比较，sorafenib干预组p-Smad3蛋白在NRK-52E细胞中表达降低，且与sorafenib剂量呈正相关(P<0.05)。结论 sorafenib具有抗肾脏纤维化作用，主要通过TGF-β/Smad途径发挥作用，可能为治疗肾纤维化提供一种早期干预的新手段。
ABSTRACT: Objective To investigate the effect of sorafenib in ameliorating renal fibrosis and its possible mechanisms. Methods Rats were subjected to unilateral ureteral obstruction (UUO) and intragastrically administered sorafenib. NRK-52E cells were treated with transforming growth factor-β1 (TGF-β1) and sorafenib. HE staining was used to visualize renal fibrosis. α-SMA and E-cadherin expressions in kidney tissue and NRK-52E cells were performed using immunofluorescence. The cell cycle of NRK-52E cells was determined by flow cytometry analysis. Smad3 and p-Smad3 protein expressions in NRK-52E cells were detected by Western blot analysis. Results HE staining showed that kidney interstitial fibrosis, tubular atrophy, and inflammatory cell infiltration in the sorafenib-treated UUO groups were significantly decreased compared with the vehicle-treated UUO group (P<0.05). Compared with those in UUO and TGF-β-stimulated NRK-52E groups, the expression of a-SMA decreased but E-cadherin expression increased in the UUO kidneys and NRK-52E cells of the sorafenib-treated groups (P<0.05). After 24h stimulation with TGF-β1 5ng/mL, the number of cell cycles arrested in G0/G1 phase was significantly increased and the number of cells that entered G2,S phase decreased (P<0.05). Compared with that in TGF-β-stimulated NRK-52E groups, p-Smad3 decreased in the sorafenib-treated groups (P<0.05). Conclusion Our results suggest that sorafenib may be useful for the treatment of renal fibrosis through suppressing TGF-β/Smad3 signaling