Fibrocytes, y-Herpesvirus and Periostin Augment Bleomycin-induced Lung Fibrosis.

Abstract

This dissertation has 4 distinct projects. First, we tested whether Pseudomonas aeruginosa, or Influenza A, (H1N1) could augment bleomycin-induced fibrosis, yet found neither of these agents had the exacerbating effects of the gamma-herpesvirus (γHV-68). Second, we explored the role of the matricellular protein, periostin, in circulating fibrocytes. Wild-type (WT) fibroblasts were treated with conditioned medium from fibrocytes isolated from bleomycin-treated WT or periostin-/- mice. We saw less α-smooth muscle actin expression in cells treated with medium from periostin-/- fibrocytes. In vitro analysis demonstrated co-regulation of mesenchymal activation by TGFβ and periostin and indicates beta 1 integrin as a potential receptor for periostin on fibrocytes. Furthermore, periostin influenced production of connective tissue growth factor from fibrocytes. Treatment of WT lung mesenchymal cells with periostin led to increased X-linked inhibitor of apoptosis (XIAP) protein expression. TGF-β1 and bleomycin treatments increased XIAP and cellular IAP (cIAP) 1 and 2 in murine lungs and mesenchymal cells. Functional blockade of XIAP and the cIAPs with AT-406, an orally bioavailable second mitochondria-derived activator of caspases (Smac) mimetic, abrogated bleomycin-induced lung fibrosis when given both prophylactically and therapeutically. Surprisingly, we compared fibrosis in XIAP deficient mice (XIAP-/y) and littermate controls but found no difference. XIAP-/y mesenchymal cells had increased resistance to Fas-mediated apoptosis, but AT-406 treatment restored sensitivity to Fas-mediated apoptosis in XIAP-/y mesenchymal cells in vitro and increased apoptosis of mesenchymal cells in vivo indicating that the increased apoptosis resistance in XIAP-/y mesenchymal cells was the result of increased cIAP expression. Biomarkers in IPF would be clinically valuable. We used an unbiased proteomic approach to identify a six-analyte index, scaled 0 to 11, based on markers of immune function, proteolysis and angiogenesis [high levels of ficolin-2 (FCN2), cathepsin-S (Cath-S), legumain (LGMN) and soluble vascular endothelial growth factor receptor 2 (VEGFsR2), but low levels of inducible T cell costimulator (ICOS) or trypsin 3 (TRY3)] that predicted better progression-free survival in IPF. This index could be useful for clinical decision making in idiopathic pulmonary fibrosis (IPF).