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A2197 - The Ovarian Cancer G-Protein Coupled Receptor 1 Protects Against the Development of Pulmonary Fibrosis
Author Block: D. Nagel, W. Ku, R. Clough, W. Narrow, P. J. Sime, R. M. Kottmann; Pulmonary and Critical Care, University of Rochester Medical Center, Rochester, NY, United States.
Rationale: We previously demonstrated that Transforming Growth Factor Beta (TGF-β)-induced extracellular acidification is a critical mechanism that drives myofibroblast differentiation via activation of latent TGF-β. This induces expression of lactate dehydrogenase thereby enhancing production of lactate which reduces extracellular pH and activates additional latent TGF-β. Mechanisms by which fibroblasts sense changes in extracellular pH are not well understood. One possibility involves a family of proton sensing G-protein coupled receptors which includes the Ovarian Cancer G-Protein Coupled Receptor 1 (OGR1). Our preliminary data show that OGR1 expression is significantly reduced in lung tissue from patients with pulmonary fibrosis, and expression is negatively regulated by TGF-β. We therefore hypothesize that OGR1 is a critical negative regulator of myofibroblast differentiation and pulmonary fibrosis.
Methods: C57Bl6 mice and OGR1-/- mice obtained from Novartis Pharmaceuticals Corporation (Basel, Switzerland) were administered 2 U/kg bleomycin or saline via oropharyngeal aspiration and sacrificed 28 days later. Bronchoalveolar Lavage Fluid (BALF) was analyzed for cell count and differential, and lung tissue was analyzed for hydroxyproline content, histologic evidence of fibrosis and fibrotic gene expression. Lung fibroblasts were isolated from OGR-/- and WT mice and cultured with/without 1 ng/mL of TGF-β. Myofibroblast differentiation was examined by measurement of alpha Smooth Muscle Actin (α-SMA) expression via Western Blot and collagen gene expression by qRT-PCR.
Results: OGR1-/- mice exposed to saline alone exhibited a three-fold increase in lung hydroxyproline compared to saline treated WT mice. Saline treated OGR1-/- mice also exhibited significant increases in Col1A1 gene expression compared to control mice. Bleomycin induced a significant increase in lung hydroxyproline in both OGR1-/- and WT mice but there was no difference between these groups suggesting a maximal response in OGR1-/- mice was achieved with saline alone. There were no differences in BALF cell counts between OGR1-/- mice and WT mice. Lastly, OGR -/- lung fibroblasts displayed greater α-SMA and Col1A1 expression compared to WT fibroblasts when exposed to TGF-β.
Conclusions: Our data supports the hypothesis that OGR1 expression negatively regulates myofibroblast differentiation and pulmonary fibrosis. Saline exposure alone induces a significant increase in lung collagen content in OGR1-/- mice compared to WT controls. OGR1-/- MLFs exhibit enhanced expression of αSMA and Col1A1 compared to WT control MLFs. These data suggest that the presence and activation of OGR1 by protons or other potential endogenous ligands significantly inhibits the pro-fibrotic effects of bleomycin and TGFβ. Therefore OGR1 may represent a novel anti-fibrotic therapeutic target.