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A5785 - Fibroblasts from Patients with Pulmonary Fibrosis Suppress Inflammatory Responses in Pulmonary Endothelial Cells
Author Block: A. E. Wyman1, M. E. Tulapurkar2, P. Karki2, T. T. T. Nguyen2, N. W. Todd2, I. G. Luzina2, S. P. Atamas2, A. A. Birukova2, K. G. Birukov2; 1Baltimore VA Medical Center, Baltimore, MD, United States, 2University of Maryland School of Medicine, Baltimore, MD, United States.
RATIONALE: Acute exacerbations of pulmonary fibrosis (AE-PF) are episodes of severe decline in respiratory function characterized by radiographic and pathologic features of acute lung injury superimposed on pulmonary fibrosis (PF). AE-PF are significant predictors of disease progression in patients with PF with very high mortality. The pathophysiology of AE-PF is poorly understood. We began to study mechanisms of AE-PF by investigating interactions between endothelial cells (EC) and fibroblasts in different cell culture models and comparing EC responses to acute inflammatory stimuli in the presence of fibroblasts from patients with idiopathic pulmonary fibrosis (IPF) and healthy controls. We have recently shown that PF is associated with decreased levels of the longevity-promoting molecules sirtuins (SIRTs), with a particularly notable decline in SIRT7 in fibroblasts from patients with PF compared to controls. To gain insight into aging-related mechanisms in AE-PF, we investigated effects of SIRT manipulation on inflammatory responses in endothelial cells and EC-fibroblast interactions. METHODS: Normal human pulmonary artery endothelial cells (HPAEC) were cultured with conditioned media from normal human lung (NHLF) and IPF lung (IPFLF) fibroblasts and co-cultured with normal and diseased fibroblasts, either directly by seeding HPAEC on fibroblasts or by separation with a membrane using trans-well inserts. To simulate an aging phenotype in HPAEC-fibroblast interactions, NHLF were transfected with control or SIRT7 siRNA and co-cultured with HPAEC. HPAEC or HPAEC-fibroblast co-cultures were stimulated with lipopolysaccharide (LPS) or TNF-α, and ICAM-1, IL-8, and IL-6 levels measured by western blotting (WB) or ELISA. RESULTS: Basal and LPS-induced levels of ICAM-1, IL-8, and IL-6 were reduced in HPAEC cultured with fibroblast-conditioned media or fibroblasts from patients with IPF compared to controls. Silencing SIRT7 in HPAEC alone or in NHLF co-cultured with HPAEC had similar, although more pronounced effects, with lower soluble ICAM-1, IL-8, and IL-6 levels. CONCLUSION: Compared to normal lung fibroblasts, fibroblasts from patients with pulmonary fibrosis suppress endothelial cell acute inflammatory responses in EC-fibroblast co-culture models. Silencing SIRT7 in endothelial cells or in NHLF co-cultured with endothelial cells has similar effects, suggesting that an aging phenotype may contribute to suppressed acute inflammatory activation of vascular endothelium in exacerbations of PF. These findings may represent a change in PF exacerbations from an acute to low-grade subacute or chronic inflammatory process that leads to a persistent fibro-proliferative phase and lung remodeling. These potential mechanisms are currently being investigated by our group.