Li Jianzhong, Ren Jiafa, Liu Xin, Jiang Lei, He Weichun, Yuan Weiping, Yang Junwei, Dai Chunsun
Center for Kidney Disease, Second Affiliated Hospital, Nanjing Medical University, Nanjing, Jiangsu, China.
State Key Laboratory of Experimental Hematology, Institute of Hematology and Blood Disease Hospital, Chinese Academy of Medical Science and Peking Union Medical College, Tianjin, China.
Kidney Int. 2015 Sep;88(3):515-27. doi: 10.1038/ki.2015.119. Epub 2015 May 13.
The mammalian target of rapamycin (mTOR) was recently identified in two structurally distinct multiprotein complexes: mTORC1 and mTORC2. Previously, we found that Rictor/mTORC2 protects against cisplatin-induced acute kidney injury, but the role and mechanisms for Rictor/mTORC2 in TGFβ1-induced fibroblast activation and kidney fibrosis remains unknown. To study this, we initially treated NRK-49F cells with TGFβ1 and found that TGFβ1 could activate Rictor/mTORC2 signaling in cultured cells. Blocking Rictor/mTORC2 signaling with Rictor or Akt1 small interfering RNAs markedly inhibited TGFβ1-induced fibronection and α-smooth muscle actin expression. Ensuing western blotting or immunostaining results showed that Rictor/mTORC2 signaling was activated in kidney interstitial myofibroblasts from mice with unilateral ureteral obstruction. Next, a mouse model with fibroblast-specific deletion of Rictor was generated. These knockout mice were normal at birth and had no obvious kidney dysfunction or kidney morphological abnormality within 2 months of birth. Compared with control littermates, the kidneys of Rictor knockout mice developed less interstitial extracellular matrix deposition and inflammatory cell infiltration at 1 or 2 weeks after ureteral obstruction. Thus our study suggests that Rictor/mTORC2 signaling activation mediates TGFβ1-induced fibroblast activation and contributes to the development of kidney fibrosis. This may provide a therapeutic target for chronic kidney diseases.
雷帕霉素哺乳动物靶蛋白(mTOR)最近在两种结构不同的多蛋白复合物中被鉴定出来:mTORC1和mTORC2。此前,我们发现Rictor/mTORC2可预防顺铂诱导的急性肾损伤,但Rictor/mTORC2在转化生长因子β1(TGFβ1)诱导的成纤维细胞活化和肾纤维化中的作用及机制尚不清楚。为了研究这一问题,我们首先用TGFβ1处理NRK-49F细胞,发现TGFβ1可激活培养细胞中的Rictor/mTORC2信号。用Rictor或Akt1小干扰RNA阻断Rictor/mTORC2信号可显著抑制TGFβ1诱导的纤连蛋白和α-平滑肌肌动蛋白表达。随后的蛋白质印迹或免疫染色结果显示,单侧输尿管梗阻小鼠肾间质肌成纤维细胞中的Rictor/mTORC2信号被激活。接下来,构建了成纤维细胞特异性缺失Rictor的小鼠模型。这些基因敲除小鼠出生时正常,出生后2个月内无明显肾功能障碍或肾脏形态异常。与对照同窝小鼠相比,输尿管梗阻1或2周后,Rictor基因敲除小鼠的肾脏间质细胞外基质沉积和炎性细胞浸润较少。因此,我们的研究表明,Rictor/mTORC2信号激活介导TGFβ1诱导的成纤维细胞活化,并促进肾纤维化的发展。这可能为慢性肾脏病提供一个治疗靶点。