View Abstract

A4197 - Endothelial Specific S1PR1 Deletion Exacerbates Bleomycin-Induced Vascular Permeability and Pulmonary Fibrosis
Author Block: R. S. Knipe1, C. K. Probst1, J. J. Spinney1, A. Franklin1, B. S. Shea2, B. D. Medoff1, A. M. Tager1, Dr. Tager passed away August 11, 2017; 1Pulmonary and Critical Care Medicine, Massachusetts General Hospital, Boston, MA, United States, 2Pulmonary, Critical Care and Sleep Medicine, Rhode Island Hospital, Providence, RI, United States.
Rationale:
Idiopathic pulmonary fibrosis is a progressive scarring lung disease which leads to lung tissue destruction and respiratory failure. The current paradigm of fibrogenesis involves aberrant or overexuberant wound-healing responses to repetitive lung injury. Increased vascular permeability is a cardinal response to tissue injury. Sphingosine 1-phosphate (S1P) is a major endogenous regulator of vascular permeability. S1P signals through one of its receptors, sphingosine-1-phosphate receptor 1 (S1PR1), on endothelial cells, inducing cytoskeletal rearrangements which stabilize paracellular junctions and maintain barrier function. We previously demonstrated that pharmacologic antagonism of S1PR1 promotes vascular permeability and pulmonary fibrosis (Shea BS et al, AJRCMB 2010;43:662-73). Here we genetically delete endothelial S1PR1 in vivo to investigate its role in pulmonary fibrosis.
Methods:
Mice with the sphingosine-1-phosphate receptor 1 (S1PR1) gene floxed (S1PR1 f/f) were bred to mice expressing inducible endothelial specific Cre recombinase (CDH5-CreERT2), then challenged with intratracheal (IT) bleomycin (1.2 U/kg) after Tamoxifen induced deletion. Vascular permeability was measured in the lungs 7 days after IT Bleomycin. Lung Evans blue index was calculated as [lung]:[plasma] concentrations of Evans blue dye. Pulmonary fibrosis was quantified by hydroxyproline assay in lungs at day 14 after low dose IT bleomycin (0.5 U/kg). Fibrosis and extravascular coagulation were observed with Trichrome staining on histology.
Results:
At day 7 post-bleomycin challenge, vascular leak was markedly increased in lungs of CDH5-CreERT2/ S1PR1 f/f mice compared to controls. Lung Evans blue index (0.180 +/- 0.014) was also increased in these mice compared to controls (0.037 +/- 0.005, p = 0.0004). Furthermore, at day 7, extensive fibrin deposition was present on histology in the CDH5-CreERT2/ S1PR1 f/f mice, and at day 14, fibrosis was significantly increased in the lungs of these mice compared to controls. Hydroxyproline values for CDH5-CreERT2/ S1PR1 f/f mice at day 14 was 352.2 mcg/lungs +/- 12.7 versus 265.0 mcg/lungs +/- 16.9 for control mice, p = 0.002.
Conclusions:
Endothelial specific deletion of S1PR1 markedly exacerbated vascular leak in response to IT bleomycin challenge and increased the fibrotic response to low dose IT bleomycin. Histologic analysis at day 7 post bleomycin demonstrated widespread fibrin deposition, preceding fibrotic remodeling. These results suggest that exaggeration of vascular permeability increases pulmonary fibrosis in the bleomycin mouse model. We hypothesize that loss of vascular integrity is a critical pro-fibrotic mechanism, through the leakage of plasma proteins into airspaces. Protecting endothelial barrier function through S1PR1 agonism could be a novel and effective anti-fibrotic therapeutic strategy.