Functional Roles of Tumor Necrosis Factor-Alpha and Interleukin 1-Beta in Hypoxia and Reoxygenation Heather E. Merry, MD, Patrick Phelan, MD, Matthew.

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Functional Roles of Tumor Necrosis Factor-Alpha and Interleukin 1-Beta in Hypoxia and Reoxygenation Heather E. Merry, MD, Patrick Phelan, MD, Matthew Doaks, BS, Minqing Zhao, MD, PhD, Michael S. Mulligan, MD The Annals of Thoracic Surgery Volume 99, Issue 4, Pages 1200-1205 (April 2015) DOI: 10.1016/j.athoracsur.2014.11.042 Copyright © 2015 The Society of Thoracic Surgeons Terms and Conditions

Fig 1 Baseline chemokine content in experimental media. There was no significant difference in monocyte chemotactic protein 1 (MCP-1) or CINC content between the activated alveolar macrophage (AM) media, the tumor necrosis factor-alpha (TNF-α)–depleted and interleukin 1-beta (IL-1β)–depleted media. The Annals of Thoracic Surgery 2015 99, 1200-1205DOI: (10.1016/j.athoracsur.2014.11.042) Copyright © 2015 The Society of Thoracic Surgeons Terms and Conditions

Fig 2 Electromobility shift assay for nuclear factor-kappa B (NFκB) nuclear translocation in pulmonary artery endothelial cells. There was a marked increase in NFκB translocation relative to the control (Ctrl) media with activated alveolar macrophage (AM) media at 15 minutes of reoxygenation (p < 0.001). However, with tumor necrosis factor-alpha (TNF-α) depletion there was suppression of NFκB below the level of control media (p < 0.005). (HR = hypoxia–reperfusion; IL-1β = interleukin 1-beta; OD = optical density.) The Annals of Thoracic Surgery 2015 99, 1200-1205DOI: (10.1016/j.athoracsur.2014.11.042) Copyright © 2015 The Society of Thoracic Surgeons Terms and Conditions

Fig 3 Electromobility shift assay for nuclear factor-kappa B (NFκB) nuclear translocation in type 2 pneumocytes. There was a similar pattern of NFκB activity as seen in the pulmonary artery endothelial cells, with suppression of translocation with tumor necrosis factor-alpha (TNF-α) depletion. (AM = alveolar macrophage; Ctrl = control media; HR = hypoxia–reperfusion; IL-1β = interleukin 1-beta.) The Annals of Thoracic Surgery 2015 99, 1200-1205DOI: (10.1016/j.athoracsur.2014.11.042) Copyright © 2015 The Society of Thoracic Surgeons Terms and Conditions

Fig 4 Electromobility shift assay and densitometry for early growth response protein 1 (EGR-1) and activator protein 1 (AP-1). In pulmonary artery endothelial cells there was a suppression of EGR-1 translocation with interleukin 1-beta (IL-1β) depletion, and in type 2 pneumocytes there was suppression of AP-1 translocation relative to control (Ctrl) and activated alveolar macrophage (AM) media. Tumor necrosis factor-alpha (TNF-α) depletion normalized AP-1 translocation in, type 2 pneumocytes; however, in pulmonary artery endothelial cells there was a 25% increase relative to the control media. (A) AP-1 translocation at 15 minutes of reoxygenation in type 2 pneumocytes. (B) EGR-1 translocation at 15 minutes reoxygenation in pulmonary artery endothelial cells. (HR = hypoxia–reperfusion; OD = optical density.) The Annals of Thoracic Surgery 2015 99, 1200-1205DOI: (10.1016/j.athoracsur.2014.11.042) Copyright © 2015 The Society of Thoracic Surgeons Terms and Conditions