title: GSK3, snail, and adhesion molecule regulation by cyclosporine A in renal tubular cells.
abstract: Tubular cell injury and fibrosis are key features of calcineurin inhibitor nephrotoxicity, but the molecular processes involved are not fully understood. In cultured murine MCT and human kidney 2 proximal tubular cells, gene expression and protein levels were studied by real-time polymerase chain reaction, Western blot, and confocal microscopy. Protein function was evaluated by pharmacological inhibitors and confirmed by small interfering RNA (siRNA) gene targeting. In renal tubular cells, cytotoxic concentrations of cyclosporine A (CsA) inhibited both gene and protein expression of adherent and tight junction proteins (E-cadherin, ZO-1, claudin-1, and β-catenin) and increased vimentin expression, without involvement of transforming growth factor β1 or caspase activity. CsA upregulated transcriptional repressors (Snail, Slug, and Twist) of the adherent and tight junction proteins were studied. Snail siRNA targeting prevented the downregulation of E-cadherin by CsA. CsA promoted glycogen synthase kinase 3 (GSK3) phosphorylation and increased Snail half-life. The GSK3 inhibitor lithium upregulated Snail and decreased E-cadherin expression in a Snail-dependent manner. Moreover, targeting GSK3 activity by siRNA also upregulated Snail. Furthermore, GSK3 siRNA had a negative impact on CsA-induced upregulation of Snail. Tacrolimus also inhibited GSK3 and mimicked CsA responses in tubular cells. We conclude that calcineurin inhibitors may directly decrease the expression of epithelial adhesion molecules by repressing GSK3 and stabilizing Snail. This offers potential pharmacological targets for prevention of nephrotoxicity.
authors.affiliation: Division of Nephrology and Hypertension, Laboratory of Nephrology and Vascular Pathology, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz, Universidad Autónoma de Madrid, 28040 Madrid, Spain.
authors.forename: Sergio
authors.initials: S
authors.lastname: Berzal
authors.forename: Matilde
authors.initials: M
authors.lastname: Alique
authors.forename: Marta
authors.initials: M
authors.lastname: Ruiz-Ortega
authors.forename: Jesús
authors.initials: J
authors.lastname: Egido
authors.forename: Alberto
authors.initials: A
authors.lastname: Ortiz
authors.forename: Adrián M
authors.initials: AM
authors.lastname: Ramos
mesh_terms: D008841:Actin Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D015820:Cadherins / Q000378:metabolism; D019703:Calcineurin / Q000378:metabolism; D065095:Calcineurin Inhibitors; D015815:Cell Adhesion Molecules / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D062445:Claudin-1; D016572:Cyclosporine / Q000633:toxicity*; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007166:Immunosuppressive Agents / Q000633:toxicity*; D007687:Kidney Tubules, Proximal / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D018613:Microscopy, Confocal; D010750:Phosphoproteins / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D034622:RNA Interference; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D000071250:Snail Family Transcription Factors; D013997:Time Factors; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D053773:Transforming Growth Factor beta1 / Q000378:metabolism; D014746:Vimentin / Q000378:metabolism; D062826:Zonula Occludens-1 Protein; D051176:beta Catenin / Q000378:metabolism
chemical_list: C568108:CLDN1 protein, human; D015820:Cadherins; D065095:Calcineurin Inhibitors; D015815:Cell Adhesion Molecules; D062445:Claudin-1; C568113:Cldn1 protein, mouse; D007166:Immunosuppressive Agents; D008565:Membrane Proteins; D010750:Phosphoproteins; D047428:Protein Kinase Inhibitors; C000604407:SNAI1 protein, human; C000604449:Snai2 protein, mouse; D000071250:Snail Family Transcription Factors; C569042:TJP1 protein, human; C569043:Tjp1 protein, mouse; D014157:Transcription Factors; D053773:Transforming Growth Factor beta1; D014746:Vimentin; D062826:Zonula Occludens-1 Protein; D051176:beta Catenin; D016572:Cyclosporine; D038362:Glycogen Synthase Kinase 3; D019703:Calcineurin