Tubular epithelial cells play a central role in the pathogenesis of chronic nephropathies. Previous toxicogenomic studies have demonstrated that cyclosporine- (CsA-) induced epithelial phenotypic changes (EPCs) are reminiscent of an incomplete epithelial to mesenchymal transition (EMT) in a TGF-β-independent manner. Furthermore, we identified endoplasmic reticulum (ER) stress as a potential mechanism that may participate in the modulation of tubular cell plasticity during CsA exposure. Because c-jun-N-terminal kinase (JNK), which is activated during ER stress, is implicated in kidney fibrogenesis, we undertook the current study to identify the role of JNK signaling in EPCs induced by CsA. In primary cultures of human renal epithelial cells, CsA activates JNK signaling, and the treatment with a JNK inhibitor reduces the occurrence of cell shape changes, E-cadherin downregulation, cell migration, and Snail-1 expression. Our results suggest that CsA activates JNK signaling, which, in turn, may participate in the morphological alterations through the regulation of Snail-1 expression. 1. Introduction The pathogenesis of cyclosporine (CsA) nephrotoxicity is not fully understood, and many mechanisms have been proposed [1, 2], such as the epithelial to mesenchymal transition (EMT), a phenomenon that has been implicated in the initiation and development of chronic nephropathies [3–6]. Under the pressure of numerous facilitating factors, tubular cells can lose their epithelial phenotype, express mesenchymal markers, acquire migratory and invasive potential, and secrete extracellular matrix. EMT promotes the generation of myofibroblasts, and growing evidence has implicated this process in diseased kidneys, leading to renal fibrosis [4, 7–9]. A better understanding of the role of tubular cells and EMT in the development of kidney fibrosis may lead to the development of specific biomarkers of early graft damage and to the characterization of novel therapeutic targets. Transcriptomic analysis of tubular cell response to CsA has shown that CsA induces EMT in vitro in a transforming growth factor β- (TGF-β-) dependent manner [10]. In a previous study, we have shown that CsA promotes TGF-β-independent epithelial phenotypic changes (EPCs). However, we have not observed de novoα-smooth muscle actin (α-SMA) expression, suggesting that CsA induces an incomplete EMT. In addition, we have identified the endoplasmic reticulum (ER) stress as a potential mechanism that may participate in the modulation of tubular cell plasticity in CsA-induced EPCs [11]. However, the precise
References
[1]
E. A. Burdmann, T. F. Andoh, L. Yu, and W. M. Bennett, “Cyclosporine nephrotoxicity,” Seminars in Nephrology, vol. 23, no. 5, pp. 465–476, 2003.
[2]
J. M. Campistol and S. H. Sacks, “Mechanisms of nephrotoxicity,” Transplantation, vol. 69, no. 12, pp. SS5–SS10, 2000.
[3]
A. A. Eddy, “Molecular basis of renal fibrosis,” Pediatric Nephrology, vol. 15, no. 3-4, pp. 290–301, 2000.
[4]
Y. Liu, “Epithelial to mesenchymal transition in renal fibrogenesis: pathologic significance, molecular mechanism, and therapeutic intervention,” Journal of the American Society of Nephrology, vol. 15, no. 1, pp. 1–12, 2004.
[5]
F. Strutz and M. Zeisberg, “Renal fibroblasts and myofibroblasts in chronic kidney disease,” Journal of the American Society of Nephrology, vol. 17, no. 11, pp. 2992–2998, 2006.
[6]
M. Zeisberg, F. Strutz, and G. A. Müller, “Renal fibrosis: an update,” Current Opinion in Nephrology and Hypertension, vol. 10, no. 3, pp. 315–320, 2001.
[7]
A. Djamali, S. Reese, J. Yracheta, T. Oberley, D. Hullett, and B. Becker, “Epithelial-to-mesenchymal transition and oxidative stress in chronic allograft nephropathy,” American Journal of Transplantation, vol. 5, no. 3, pp. 500–509, 2005.
[8]
A. Hertig, J. Verine, B. Mougenot et al., “Risk factors for early epithelial to mesenchymal transition in renal grafts,” American Journal of Transplantation, vol. 6, no. 12, pp. 2937–2946, 2006.
[9]
M. Iwano, D. Plieth, T. M. Danoff, C. Xue, H. Okada, and E. G. Neilson, “Evidence that fibroblasts derive from epithelium during tissue fibrosis,” Journal of Clinical Investigation, vol. 110, no. 3, pp. 341–350, 2002.
[10]
T. McMorrow, M. M. Gaffney, C. Slattery, E. Campbell, and M. P. Ryan, “Cyclosporine A induced epithelial-mesenchymal transition in human renal proximal tubular epithelial cells,” Nephrology Dialysis Transplantation, vol. 20, no. 10, pp. 2215–2225, 2005.
[11]
N. Pallet, N. Bouvier, A. Bendjallabah et al., “Cyclosporine-induced endoplasmic reticulum stress triggers tubular phenotypic changes and death,” American Journal of Transplantation, vol. 8, no. 11, pp. 2283–2296, 2008.
[12]
F. Urano, X. Wang, A. Bertolotti et al., “Coupling of stress in the ER to activation of JNK protein kinases by transmembrane protein kinase IRE1,” Science, vol. 287, no. 5453, pp. 664–666, 2000.
[13]
M. Diry, C. Tomkiewicz, C. Koehle et al., “Activation of the dioxin/aryl hydrocarbon receptor (AhR) modulates cell plasticity through a JNK-dependent mechanism,” Oncogene, vol. 25, no. 40, pp. 5570–5574, 2006.
[14]
J. C. Pastor-Pareja, F. Grawe, E. Martín-Blanco, and A. García-Bellido, “Invasive cell behavior during Drosophila imaginal disc eversion is mediated by the JNK signaling cascade,” Developmental Cell, vol. 7, no. 3, pp. 387–399, 2004.
[15]
Y. Xia and M. Karin, “The control of cell motility and epithelial morphogenesis by Jun kinases,” Trends in Cell Biology, vol. 14, no. 2, pp. 94–101, 2004.
[16]
F. Y. Ma, R. S. Flanc, G. H. Tesch et al., “A pathogenic role for c-Jun amino-terminal kinase signaling in renal fibrosis and tubular cell apoptosis,” Journal of the American Society of Nephrology, vol. 18, no. 2, pp. 472–484, 2007.
[17]
K. Giehl, Y. Imamichi, and A. Menke, “Smad4-independent TGF-β signaling in tumor cell migration,” Cells Tissues Organs, vol. 185, no. 1–3, pp. 123–130, 2007.
[18]
D. Anglicheau, N. Pallet, M. Rabant et al., “Role of P-glycoprotein in cyclosporine cytotoxicity in the cyclosporine-sirolimus interaction,” Kidney International, vol. 70, no. 6, pp. 1019–1025, 2006.
[19]
N. Pallet, E. Thervet, D. L. Corre et al., “Rapamycin inhibits human renal epithelial cell proliferation: effect on cyclin D3 mRNA expression and stability,” Kidney International, vol. 67, no. 6, pp. 2422–2433, 2005.
[20]
K. J. Livak and T. D. Schmittgen, “Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCT method,” Methods, vol. 25, no. 4, pp. 402–408, 2001.
[21]
C. Xu, B. Bailly-Maitre, and J. C. Reed, “Endoplasmic reticulum stress: cell life and death decisions,” Journal of Clinical Investigation, vol. 115, no. 10, pp. 2656–2664, 2005.
[22]
T. Borsellol, P. G. H. Clarkel, L. Hirt et al., “A peptide inhibitor of c-Jun N-terminal kinase protects against excitotoxicity and cerebral ischemia,” Nature Medicine, vol. 9, no. 9, pp. 1180–1186, 2003.
[23]
C. P. Lim, N. Jain, and X. Cao, “Stress-induced immediate-early gene, egr-1, involves activation of p38/JNK1,” Oncogene, vol. 16, no. 22, pp. 2915–2926, 1998.
[24]
N. Pallet, M. Rabant, Y. C. Xu-Dubois et al., “Response of human renal tubular cells to cyclosporine and sirolimus: a toxicogenomic study,” Toxicology and Applied Pharmacology, vol. 229, no. 2, pp. 184–196, 2008.
[25]
C. Slattery, E. Campbell, T. McMorrow, and M. P. Ryan, “Cyclosporine A—induced renal fibrosis: a role for epithelial-mesenchymal transition,” American Journal of Pathology, vol. 167, no. 2, pp. 395–407, 2005.
[26]
M. H. De Borst, J. Prakash, W. B. W. H. Melenhorst et al., “Glomerular and tubular induction of the transcription factor c-Jun in human renal disease,” Journal of Pathology, vol. 213, no. 2, pp. 219–228, 2007.
[27]
F. Y. Ma, M. Sachchithananthan, R. S. Flanc, et al., “Mitogen activated protein kinases in renal fibrosis,” Frontiers in Bioscience, vol. 1, pp. 171–187, 2009.
[28]
H. McNeill, “Planar cell polarity and the kidney,” Journal of the American Society of Nephrology, vol. 20, no. 10, pp. 2104–2111, 2009.
[29]
H. Peinado, D. Olmeda, and A. Cano, “Snail, ZEB and bHLH factors in tumour progression: an alliance against the epithelial phenotype?” Nature Reviews Cancer, vol. 7, no. 6, pp. 415–428, 2007.
[30]
J. Yoshino, T. Monkawa, M. Tsuji, M. Inukai, H. Itoh, and M. Hayashi, “Snail1 is involved in the renal epithelial-mesenchymal transition,” Biochemical and Biophysical Research Communications, vol. 362, no. 1, pp. 63–68, 2007.
[31]
Y. Li, Y. Liu, Y. Xu, J. J. Voorhees, and G. J. Fisher, “UV irradiation induces Snail expression by AP-1 dependent mechanism in human skin keratinocytes,” Journal of Dermatological Science, vol. 60, no. 2, pp. 105–113, 2010.
