title: ATRA induces podocyte differentiation and alters nephrin and podocin expression in vitro and in vivo.
abstract: BACKGROUND
  Podocytes are terminally differentiated and highly specialized epithelial cells. The factors governing podocyte differentiation are poorly understood. We tested the hypothesis that all-trans retinoic acid (ATRA), a vitamin A derivative, induces podocyte differentiation in vitro and in vivo.
  
  
  METHODS
  We tested the effects of ATRA on podocytes. Primary rat, primary mouse, and immortalized mouse podocytes were exposed to ATRA (1, 5, 10, 20, 40, 50, 80, 160, and 200 micromol/L) or control (ethanol) for 72 hours. Cell morphology was examined by electron microscopy, the expression of podocyte specific proteins was measured by immunoflourescence and Western blot analysis, cell number and apoptosis were measured by 3-[4,5] dimethylthiazol-2,5-diphenyltetrazolium bromide (MTT) assay and Hoechst staining, respectively. To determine if ATRA alters podocyte differentiation in vivo, experimental injury was induced in C57BL6 mice using the antiglomerular antibody. Animals were given either daily intraperitoneal ATRA (16 mg/kg) or vehicle (corn oil). For end points, we measured proteinuria, podocyte-specific protein immunostaining, and proliferation [proliferating cell nuclear antigen (PCNA)] at days 5 and 14 (N= 5/group/time point).
  
  
  RESULTS
  ATRA induced podocyte process formation in vitro, and significantly increased the expression of nephrin and podocin. This coincided with a reduction in proliferation. ATRA also significantly prevented the decrease in staining for synaptopodin, nephrin, and podocin in experimental animals (P < 0.05 vs. control). This was accompanied by reduced proteinuria and decreased podocyte proliferation (P < 0.05 vs. control).
  
  
  CONCLUSIONS
  ATRA induces podocyte differentiation in vitro and in vivo and alters the expression of certain podocyte-specific proteins. Further studies are ongoing to delineate the mechanism of this effect.
authors.affiliation: Department of Medicine, Division of Nephrology, University of Washington, Seattle, Washington, USA.
authors.forename: Michael R
authors.initials: MR
authors.lastname: Vaughan
authors.forename: Jeffrey W
authors.initials: JW
authors.lastname: Pippin
authors.forename: Siân V
authors.initials: SV
authors.lastname: Griffin
authors.forename: Ron
authors.initials: R
authors.lastname: Krofft
authors.forename: Margaret
authors.initials: M
authors.lastname: Fleet
authors.forename: Leah
authors.initials: L
authors.lastname: Haseley
authors.forename: Stuart J
authors.initials: SJ
authors.lastname: Shankland
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002461:Cell Line, Transformed; D005921:Glomerulonephritis / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D066298:In Vitro Techniques; D047908:Intracellular Signaling Peptides and Proteins; D007678:Kidney Glomerulus / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D011507:Proteinuria / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D051381:Rats; D017207:Rats, Sprague-Dawley; D014212:Tretinoin / Q000494:pharmacology*
chemical_list: D000970:Antineoplastic Agents; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; C407223:NPHS2 protein; C113267:nephrin; D014212:Tretinoin