title: ZEB1 is estrogen responsive in vitro in human foreskin cells and is over expressed in penile skin in patients with severe hypospadias.
abstract: OBJECTIVE
  We determined the effect of estrogen on ZEB1 in vitro and tested the hypothesis that ZEB1 is over expressed in the penile skin of subjects with hypospadias.
  
  
  METHODS
  Hs68 cells, a fibroblast cell line derived from human foreskin, were exposed to 0, 1, 10 and 100 nM estrogen, and the expression level of ZEB1 was assessed using reverse transcription real-time polymerase chain reaction, Western blot and immunocytochemical analysis. Next, preputial skin was prospectively collected from case and control subjects at hypospadias repair (37 cases) and circumcision (11). Hypospadias was classified as severe (13 cases) or mild (24) based on the position of the urethral meatus. ZEB1 expression was quantified using reverse transcription real-time polymerase chain reaction, Western blot and immunohistochemical analysis.
  
  
  RESULTS
  Estrogen increased ZEB1 expression at the mRNA and protein levels in Hs68 cells in a concentration dependent fashion (p <0.01). Subjects with severe hypospadias had significantly higher ZEB1 mRNA levels and protein expression compared to controls or subjects with mild hypospadias (both p <0.01). Subjects with severe hypospadias had increased expression of ZEB1 in the basal layers of the preputial epidermis.
  
  
  CONCLUSIONS
  Estrogen increases ZEB1 expression in a human foreskin fibroblast cell line in vitro. Furthermore, ZEB1 is significantly over expressed in the penile skin of subjects with severe hypospadias. We propose that ZEB1 overexpression may contribute to development of hypospadias and may mediate the effect of estrogen on developing external male genitalia.
authors.affiliation: Frank Hinman, Jr. Urological Research Laboratory, Department of Urology, University of California, San Francisco, California 94143, USA.
authors.forename: Liang
authors.initials: L
authors.lastname: Qiao
authors.forename: Gregory E
authors.initials: GE
authors.lastname: Tasian
authors.forename: Haiyang
authors.initials: H
authors.lastname: Zhang
authors.forename: Gerald R
authors.initials: GR
authors.lastname: Cunha
authors.forename: Laurence
authors.initials: L
authors.lastname: Baskin
mesh_terms: D015153:Blotting, Western; D016022:Case-Control Studies; D002460:Cell Line; D002478:Cells, Cultured; D002648:Child; D004967:Estrogens / Q000502:physiology*; D005347:Fibroblasts / Q000378:metabolism; D052816:Foreskin / Q000378:metabolism*; D018398:Homeodomain Proteins / Q000378:metabolism*; D006801:Humans; D007021:Hypospadias / Q000378:metabolism*; D007150:Immunohistochemistry; D008297:Male; D010413:Penis / Q000378:metabolism*; D010641:Phenotype; D011446:Prospective Studies; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012720:Severity of Illness Index; D014157:Transcription Factors / Q000378:metabolism*; D015854:Up-Regulation; D000071799:Zinc Finger E-box-Binding Homeobox 1
chemical_list: D004967:Estrogens; D018398:Homeodomain Proteins; D014157:Transcription Factors; C486307:ZEB1 protein, human; D000071799:Zinc Finger E-box-Binding Homeobox 1