Loss of the alpha-isoform of calcineurin is sufficient to induce nephrotoxicity and altered expression of transforming growth factor-beta.
BACKGROUND
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.


METHODS
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.


RESULTS
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.


CONCLUSIONS
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.
Gooch
Jennifer L
JL
Department of Medicine, Division of Nephrology, Emory University School of Medicine, Atlanta, GA 30322, USA. jgooch@emory.edu
Roberts
Brian R
BR
Cobbs
Scott L
SL
Tumlin
James A
JA
D000818:Animals; D000906:Antibodies / Q000276:immunology / Q000494:pharmacology; D019703:Calcineurin / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D005786:Gene Expression Regulation; D007527:Isoenzymes / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D007674:Kidney Diseases / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D051379:Mice; D018345:Mice, Knockout; D016212:Transforming Growth Factor beta / Q000276:immunology / Q000378:metabolism*
D000906:Antibodies; D007527:Isoenzymes; D016212:Transforming Growth Factor beta; D019703:Calcineurin