title: TGF-β1 induces proteinase-activated receptor 2 (PAR2) expression in endometriotic stromal cells and stimulates PAR2 activation-induced secretion of IL-6.
abstract: BACKGROUND
  Proteinase-activated receptor 2 (PAR2) is a G-protein-coupled receptor that is activated by several serine proteases. PAR2 activation in endometriotic stromal cells (ESCs) has been implicated in the development of endometriosis but the regulatory mechanism of PAR2 expression in ESC is unknown. Our objective was to study the mechanism by which PAR2 expression may be regulated in endometriotic lesions.
  
  
  METHODS
  Primary cultures of ESCs were treated with transforming growth factor-β (TGF-β) 1, tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), and the expression of PAR2 was examined by real-time quantitative PCR. ESCs pretreated with or without TGF-β1 were treated with PAR2 agonist peptide (PAR2AP) and the secretion of the pro-endometriotic cytokine, IL-6, was measured using a specific enzyme-linked immunosorbent assay. Effects of TGF-β type 1 inhibitor, SB431542, and PAR2 small interfering RNA (siRNA) on the TGF-β1 stimulation of PAR2 gene expression and PAR2AP-induced IL-6 secretion were also evaluated. To study intracellular signaling, effects of inhibitors of mitogen-activated protein kinases (MAPKs) and phosphoinositide 3-kinase (PI3K) and of Smad4 siRNA on the TGF-β1-induced PAR2 gene expression were studied.
  
  
  RESULTS
  Only TGF-β1, but neither TNF-α nor IL-1β, increased gene expression of PAR2. Activation of PAR2 with PAR2AP increased the secretion of IL-6 from ESCs. As expected, TGF-β1 pretreatment dose-dependently enhanced the PAR2AP-induced increase in IL-6 secretion from ESCs. Treatment of ESCs with the TGF-β type 1 inhibitor, SB431542, inhibited both TGF-β1-stimulation of PAR2 gene expression and PAR2AP-induced IL-6 secretion. Transfection of ESCs with PAR2 siRNA produced a similar inhibition of IL-6 secretion. The TGF-β1-induced increase in PAR2 gene expression was repressed by inhibition of p38 MAPK, p42/44 MAPK or PI3K, but not by knockdown of Smad4 expression.
  
  
  CONCLUSIONS
  In view of significant roles of PAR2 and IL-6 in endometriosis, the TGF-β1-induced increase in PAR2 expression may be an elaborate mechanism that augments the progression of the disease.
authors.affiliation: Department of Obstetrics and Gynecology, University of Tokyo, Tokyo, Japan.
authors.forename: Ako
authors.initials: A
authors.lastname: Saito
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Osuga
authors.forename: Osamu
authors.initials: O
authors.lastname: Yoshino
authors.forename: Masashi
authors.initials: M
authors.lastname: Takamura
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Hirata
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Hirota
authors.forename: Kaori
authors.initials: K
authors.lastname: Koga
authors.forename: Miyuki
authors.initials: M
authors.lastname: Harada
authors.forename: Yuri
authors.initials: Y
authors.lastname: Takemura
authors.forename: Tetsu
authors.initials: T
authors.lastname: Yano
authors.forename: Yuji
authors.initials: Y
authors.lastname: Taketani
mesh_terms: D004715:Endometriosis / Q000378:metabolism* / Q000473:pathology; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D053583:Interleukin-1beta / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000187:drug effects; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D034622:RNA Interference; D044462:Receptors, Proteinase-Activated / Q000378:metabolism*; D051901:Smad4 Protein / Q000037:antagonists & inhibitors; D017154:Stromal Cells / Q000378:metabolism*; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D053583:Interleukin-1beta; D015850:Interleukin-6; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D044462:Receptors, Proteinase-Activated; C497796:SMAD4 protein, human; D051901:Smad4 Protein; D053773:Transforming Growth Factor beta1; D014409:Tumor Necrosis Factor-alpha