Gooch Jennifer L, Roberts Brian R, Cobbs Scott L, Tumlin James A
Department of Medicine, Division of Nephrology, Emory University School of Medicine, Atlanta, GA 30322, USA.
Transplantation. 2007 Feb 27;83(4):439-47. doi: 10.1097/01.tp.0000251423.78124.51.
Use of calcineurin inhibitors is frequently limited by fibrosis, closely linked with increased transforming growth factor (TGF)-beta. However, mechanisms of extracellular matrix expansion and TGFbeta regulation following calcineurin inhibition are unknown. Mice lacking specific calcineurin catalytic subunit isoforms may offer important insight into this pathway.
We compared mice lacking the alpha or beta isoform to a model of cyclosporin nephrotoxicity. Histological features common with cyclosporin nephrotoxicity including matrix expansion, arteriole hyalinization, and inflammation were assessed. Next, regulation specifically of fibronectin and TGFbeta was examined in vivo and in vitro. Finally, the role of TGFbeta in upregulation of fibronectin with loss of calcineurin activity was examined.
Loss of the alpha isoform results in histologic features and matrix expansion similar to cyclosporin, whereas loss of the beta does not. Fibronectin and TGFbeta are increased and renal function is impaired in alpha-null and aged alpha+/-. In primary alpha-/- renal fibroblasts, nuclear translocation of the calcineurin substrate NFATc is normal but regulation is lost in beta-null fibroblasts, confirming that the isoforms have distinct functions. Consistent with in vivo findings, alpha-null cells have increased fibronectin and TGFbeta. However, neutralizing TGFbeta antibody did not reduce fibronectin accumulation.
Our data show that calcineurin-alpha is key to regulation of fibrosis and TGFbeta and loss of this isoform reproduces features of cyclosporine nephrotoxicity in vivo and in vitro. In addition, we show that upregulation of TGFbeta and fibronectin likely result from a shared mechanism, but changes in fibronectin expression are independent of TGFbeta in renal fibroblasts.
钙调神经磷酸酶抑制剂的使用常因纤维化而受到限制，纤维化与转化生长因子（TGF）-β的增加密切相关。然而，钙调神经磷酸酶抑制后细胞外基质扩张和TGF-β调节的机制尚不清楚。缺乏特定钙调神经磷酸酶催化亚基异构体的小鼠可能为该途径提供重要见解。
我们将缺乏α或β异构体的小鼠与环孢素肾毒性模型进行了比较。评估了与环孢素肾毒性共有的组织学特征，包括基质扩张、小动脉玻璃样变和炎症。接下来，在体内和体外研究了纤连蛋白和TGF-β的特异性调节。最后，研究了TGF-β在钙调神经磷酸酶活性丧失导致纤连蛋白上调中的作用。
α异构体的缺失导致组织学特征和基质扩张与环孢素相似，而β异构体的缺失则不会。在α基因敲除和老年α+/-小鼠中，纤连蛋白和TGF-β增加，肾功能受损。在原代α-/-肾成纤维细胞中，钙调神经磷酸酶底物NFATc的核转位正常，但在β基因敲除的成纤维细胞中调节丧失，证实这些异构体具有不同的功能。与体内研究结果一致，α基因敲除细胞中纤连蛋白和TGF-β增加。然而，中和TGF-β抗体并未减少纤连蛋白的积累。
我们的数据表明，钙调神经磷酸酶-α是纤维化和TGF-β调节的关键，该异构体的缺失在体内和体外重现了环孢素肾毒性的特征。此外，我们表明TGF-β和纤连蛋白的上调可能源于共同机制，但肾成纤维细胞中纤连蛋白表达的变化独立于TGF-β。
