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Mary E. Huerter, MD, MA, Ashish K

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1 Attenuation of Pulmonary Ischemia-Reperfusion Injury by Adenosine A2B Receptor Antagonism 
Mary E. Huerter, MD, MA, Ashish K. Sharma, MBBS, PhD, Yunge Zhao, MD, PhD, Eric J. Charles, MD, Irving L. Kron, MD, Victor E. Laubach, PhD  The Annals of Thoracic Surgery  Volume 102, Issue 2, Pages (August 2016) DOI: /j.athoracsur Copyright © 2016 The Society of Thoracic Surgeons Terms and Conditions

2 Fig 1 Lung function after ischemia-reperfusion (IR) is improved by ATL802. Mice exhibited significant lung dysfunction after IR, which was significantly attenuated by ATL802 treatment. Compared with untreated mice after IR, ATL802 treatment significantly increased pulmonary compliance and significantly decreased airway resistance and pulmonary artery pressure (∗p < 0.01 versus the sham procedure; #p < 0.03 versus IR; n = 4–9 per group). The Annals of Thoracic Surgery  , DOI: ( /j.athoracsur ) Copyright © 2016 The Society of Thoracic Surgeons Terms and Conditions

3 Fig 2 Cytokine production after ischemia-reperfusion (IR) is significantly attenuated by ATL802. IR resulted in significantly elevated levels of proinflammatory cytokines (KC, interleukin [IL]-6, IL-1β, monocyte chemoattractant protein 1 [MCP-1], macrophage inflammatory protein (MIP)-1α, and regulated on activation, normal T cell expressed and secreted [RANTES]) in bronchoalveolar lavage fluid, which were all significantly attenuated by ATL802 treatment (∗p < 0.05 versus the sham procedure; #p < 0.05 versus IR; n = 4 to 5 per group). The Annals of Thoracic Surgery  , DOI: ( /j.athoracsur ) Copyright © 2016 The Society of Thoracic Surgeons Terms and Conditions

4 Fig 3 ATL802 attenuates infiltration and activation of neutrophils after ischemia-reperfusion (IR). Lung sections were immunostained for neutrophils (red), and representative images are shown on top (×20 magnification). Quantification of neutrophils per high-powered field (HPF) demonstrated significant neutrophil infiltration in lungs after IR, which was significantly attenuated by ATL802 treatment (∗p = versus the sham procedure; #p = versus IR; n = 4 per group). In addition, myeloperoxidase levels in bronchoalveolar lavage (BAL) fluid were significantly elevated after IR, which was significantly attenuated by ATL802 treatment (∗∗p = versus the sham procedure, ##p = versus IR; n = 4–6 per group). The Annals of Thoracic Surgery  , DOI: ( /j.athoracsur ) Copyright © 2016 The Society of Thoracic Surgeons Terms and Conditions

5 Fig 4 Pulmonary vascular permeability after ischemia-reperfusion (IR) is attenuated by ATL802. Vascular permeability was assessed using Evans blue dye extravasation technique. IR resulted in significant vascular permeability, which was significantly reduced by ATL802 treatment (∗p < versus the sham procedure; #p <  versus IR; n = 5 per group). The Annals of Thoracic Surgery  , DOI: ( /j.athoracsur ) Copyright © 2016 The Society of Thoracic Surgeons Terms and Conditions

6 Fig 5 ATL802 improves lung function after ex vivo lung perfusion (EVLP) of warm ischemic lungs. Lungs underwent 15 minutes of warm ischemia, 1 hour of cold preservation, and 1 hour of EVLP using either standard Krebs-Henseleit (KH) buffer or Steen solution. EVLP with Steen solution significantly improved lung function (elevated pulmonary compliance and reduced pulmonary artery pressure) versus KH buffer and was further and significantly improved by supplementing the Steen solution with 100 nM ATL802 (∗p < versus EVLP KH; #p < versus EVLP Steen; n = 9 per group). The Annals of Thoracic Surgery  , DOI: ( /j.athoracsur ) Copyright © 2016 The Society of Thoracic Surgeons Terms and Conditions

7 Fig 6 Secretion of interleukin (IL)-8 by alveolar type 2 epithelial cells after hypoxia-reoxygenation (HR) is attenuated by ATL802. A549 cells exhibited significant production of IL-8 by HR versus normoxia and was significantly attenuated by ATL802 (100 and 200 nM) (∗p < versus normoxia; #p < versus HR; n = 8 replicates per group). The Annals of Thoracic Surgery  , DOI: ( /j.athoracsur ) Copyright © 2016 The Society of Thoracic Surgeons Terms and Conditions

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