title: Estrogen receptor alpha expression in podocytes mediates protection against apoptosis in-vitro and in-vivo.
abstract: OBJECTIVE
  Epidemiological studies have demonstrated that women have a significantly better prognosis in chronic renal diseases compared to men. This suggests critical influences of gender hormones on glomerular structure and function. We examined potential direct protective effects of estradiol on podocytes.
  
  
  METHODS
  Expression of estrogen receptor alpha (ERα) was examined in podocytes in vitro and in vivo. Receptor localization was shown using Western blot of separated nuclear and cytoplasmatic protein fractions. Podocytes were treated with Puromycin aminonucleoside (PAN, apoptosis induction), estradiol, or both in combination. Apoptotic cells were detected with Hoechst nuclear staining and Annexin-FITC flow cytometry. To visualize mitochondrial membrane potential depolarization as an indicator for apoptosis, cells were stained with tetramethyl rhodamine methylester (TMRM). Estradiol-induced phosphorylation of ERK1/2 and p38 MAPK was examined by Western blot. Glomeruli of ERα knock-out mice and wild-type controls were analysed by histomorphometry and immunohistochemistry.
  
  
  RESULTS
  ERα was consistently expressed in human and murine podocytes. Estradiol stimulated ERα protein expression, reduced PAN-induced apoptosis in vitro by 26.5±24.6% or 56.6±5.9% (flow cytometry or Hoechst-staining, respectively; both p<0.05), and restored PAN-induced mitochondrial membrane potential depolarization. Estradiol enhanced ERK1/2 phosphorylation. In ERα knockout mice, podocyte number was reduced compared to controls (female/male: 80/86 vs. 132/135 podocytes per glomerulus, p<0.05). Podocyte volume was enhanced in ERα knockout mice (female/male: 429/371 µm(3) vs. 264/223 µm(3) in controls, p<0.05). Tgfβ1 and collagen type IV expression were increased in knockout mice, indicating glomerular damage.
  
  
  CONCLUSIONS
  Podocytes express ERα, whose activation leads to a significant protection against experimentally induced apoptosis. Possible underlying mechanisms include stabilization of mitochondrial membrane potential and activation of MAPK signalling. Characteristic morphological changes indicating glomerulopathy in ERα knock-out mice support the in vivo relevance of the ERα for podocyte viability and function. Thus, our findings provide a novel model for the protective influence of female gender on chronic glomerular diseases.
authors.affiliation: Department of General Pediatrics and Neonatology, University Children's Hospital Duesseldorf, Duesseldorf, Germany. Sebastian.Kummer@med.uni-duesseldorf.de
authors.forename: Sebastian
authors.initials: S
authors.lastname: Kummer
authors.forename: Stefanie
authors.initials: S
authors.lastname: Jeruschke
authors.forename: Lara Vanessa
authors.initials: LV
authors.lastname: Wegerich
authors.forename: Andrea
authors.initials: A
authors.lastname: Peters
authors.forename: Petra
authors.initials: P
authors.lastname: Lehmann
authors.forename: Annette
authors.initials: A
authors.lastname: Seibt
authors.forename: Friederike
authors.initials: F
authors.lastname: Mueller
authors.forename: Nadezda
authors.initials: N
authors.lastname: Koleganova
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Halbenz
authors.forename: Claus Peter
authors.initials: CP
authors.lastname: Schmitt
authors.forename: Markus
authors.initials: M
authors.lastname: Bettendorf
authors.forename: Ertan
authors.initials: E
authors.lastname: Mayatepek
authors.forename: Marie-Luise
authors.initials: ML
authors.lastname: Gross-Weissmann
authors.forename: Jun
authors.initials: J
authors.lastname: Oh
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000235:genetics; D015153:Blotting, Western; D002478:Cells, Cultured; D004958:Estradiol / Q000494:pharmacology; D047628:Estrogen Receptor alpha / Q000235:genetics / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D007678:Kidney Glomerulus / Q000378:metabolism; D053078:Membrane Potential, Mitochondrial; D051379:Mice; D018345:Mice, Knockout; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics / Q000378:metabolism; D050199:Podocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D011692:Puromycin Aminonucleoside / Q000494:pharmacology; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology; D048051:p38 Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism
chemical_list: D047628:Estrogen Receptor alpha; D004958:Estradiol; D011692:Puromycin Aminonucleoside; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D048051:p38 Mitogen-Activated Protein Kinases
references.citation: Neurosci Lett. 2007 Jan 22;412(1):68-72
references.pmid: 17196335
references.citation: Cytometry A. 2008 Feb;73(2):129-38
references.pmid: 18163486
references.citation: J Clin Pathol. 2006 Apr;59(4):424-8
references.pmid: 16497871
references.citation: Nephrol Dial Transplant. 2009 Dec;24(12):3739-44
references.pmid: 19574340
references.citation: J Cell Biochem. 2008 Dec 15;105(6):1342-51
references.pmid: 18846505
references.citation: Nephrol Dial Transplant. 1996 Jun;11(6):1003-11
references.pmid: 8671960
references.citation: Ther Ggw. 1955 Oct;94(10):364-6
references.pmid: 13291284
references.citation: Kidney Int. 2006 Jun;69(12):2131-47
references.pmid: 16688120
references.citation: Steroids. 2008 Oct;73(9-10):859-63
references.pmid: 18272190
references.citation: Kidney Int. 1998 Sep;54(3):687-97
references.pmid: 9734594
references.citation: J Neurochem. 2009 Jul;110(1):1-11
references.pmid: 19457121
references.citation: Exp Cell Res. 1997 Oct 10;236(1):248-58
references.pmid: 9344605
references.citation: J Clin Invest. 2001 Sep;108(6):807-16
references.pmid: 11560950
references.citation: Mol Endocrinol. 2005 Apr;19(4):833-42
references.pmid: 15695368
references.citation: J Clin Endocrinol Metab. 2010 Jul;95(7 Suppl 1):s1-s66
references.pmid: 20566620
references.citation: Endocr Rev. 2001 Apr;22(2):153-83
references.pmid: 11294822
references.citation: Apoptosis. 2003 Mar;8(2):115-28
references.pmid: 12766472
references.citation: Nephrol Dial Transplant. 2008 Jul;23(7):2247-53
references.pmid: 18182409
references.citation: Nephrol Dial Transplant. 2006 Sep;21(9):2577-82
references.pmid: 16720595
references.citation: J Am Soc Nephrol. 2002 Dec;13(12):3005-15
references.pmid: 12444221
references.citation: Am J Physiol Renal Physiol. 2001 May;280(5):F777-85
references.pmid: 11292619
references.citation: Lab Invest. 1975 Nov;33(5):554-61
references.pmid: 52758
references.citation: Kidney Int. 2011 Feb;79(4):404-13
references.pmid: 20962747
references.citation: Kidney Int. 2008 Aug;74(3):370-6
references.pmid: 18496507
references.citation: Adv Ren Replace Ther. 2003 Jan;10(1):3-14
references.pmid: 12616458
references.citation: Life Sci. 2011 Jan 17;88(3-4):178-86
references.pmid: 21074543
references.citation: J Biol Chem. 1995 Dec 1;270(48):28924-31
references.pmid: 7499422
references.citation: Proc Natl Acad Sci U S A. 2004 Feb 17;101(7):2076-81
references.pmid: 14764897
references.citation: Am J Hypertens. 1992 Jul;5(7):437-43
references.pmid: 1637515
references.citation: Am J Physiol Renal Physiol. 2006 Feb;290(2):F319-23
references.pmid: 16159896
references.citation: Pediatr Nephrol. 2012 Aug;27(8):1213-9
references.pmid: 21766172
references.citation: Kidney Int. 2007 Aug;72(4):464-72
references.pmid: 17495854
references.citation: Am J Kidney Dis. 1995 Apr;25(4):515-33
references.pmid: 7702046
references.citation: Science. 1995 Nov 24;270(5240):1326-31
references.pmid: 7481820
references.citation: Circ Res. 2002 May 31;90(10):1087-92
references.pmid: 12039798
references.citation: Mol Endocrinol. 2003 Jun;17(6):1039-53
references.pmid: 12624116
references.citation: Kidney Int. 1998 Jun;53(6):1696-705
references.pmid: 9607202
references.citation: Steroids. 2004 Aug;69(8-9):523-9
references.pmid: 15288764
references.citation: Kidney Int. 1991 Jul;40(1):29-34
references.pmid: 1921152
references.citation: J Am Soc Nephrol. 2004 Feb;15(2):348-58
references.pmid: 14747381
references.citation: Experientia. 1971 Aug;27(8):956-7
references.pmid: 4947885
references.citation: Kidney Int. 2001 Oct;60(4):1309-23
references.pmid: 11576345
references.citation: Am J Physiol Renal Physiol. 2004 Sep;287(3):F586-91
references.pmid: 15149972
references.citation: Virchows Arch. 2004 Jun;444(6):561-6
references.pmid: 15107991
references.citation: J Am Soc Nephrol. 2002 Nov;13(11):2737-47
references.pmid: 12397044
references.citation: Kidney Int. 1996 Oct;50(4):1278-87
references.pmid: 8887289
references.citation: Life Sci. 1987 Sep 28;41(13):1629-34
references.pmid: 3650658
references.citation: J Am Soc Nephrol. 2000 Feb;11(2):319-329
references.pmid: 10665939
references.citation: Development. 2000 Oct;127(19):4277-91
references.pmid: 10976058