397 Ex Vivo Adenoviral IL-10 Gene Therapy in a Pig Lung Transplantation Survival Model

Abstracts Management, Medical University of Vienna, Vienna, Austria; 6 Department of Cardiology, Medical University of Vienna, Vienna, Austria. Purpose: Early reperfusion of the culprit coronary artery within a narrow time window has significantly improved early mortality after acute myocardial infarction (AMI). However, a lack of adequate reperfusion at the microvascular level is often a limiting prognostic factor. The myocardial area corresponding to this impaired microvascular perfusion is termed area at risk (AAR) and is closely linked to platelet activation and aggregation. Our group has recently found that paracrine factors secreted from apoptotic peripheral blood mononuclear cells (APOSEC) attenuate AMI. The aim of this study was to determine the influence of APOSEC on AAR in a porcine AMI model. Methods and Materials: Cell culture supernatants derived from irradiated apoptotic peripheral blood mononuclear cells (APOSEC) were injected intravenously after induction of myocardial infarction. AAR was determined by magnetic resonance imaging and triphenyltetrazolium chloride (TTC) staining. Platelet function was monitored in vitro and in vivo by means of ELISA, flow cytometry, light transmission aggregometry, and western blot. Results: Treatment of AMI with APOSEC resulted in a significant reduction of AAR (26.2⫾2.6 vs. 15.6⫾2.9 % LV; p⫽0.02). This reduction was linked to improved microvascular perfusion (Myocardial blush grade: 1.3⫾0.3 vs. 2.5⫾0.3; p⫽0.033). Platelet activation markers (P-selectin, CD40L, PF-4, TSP-1) were reduced in plasma samples obtained during the procedure, suggesting an anti-aggregatory capacity of APOSEC. This finding was confirmed by in vitro tests showing that activation and aggregation of purified platelets obtained from healthy donors were significantly impaired by co-incubation with APOSEC. Conclusions: Our data give first evidence that APOSEC reduces the extent of AAR during AMI. As a possible mode of action VASP mediated inhibition of platelets was suggested. 396 Water-Soluble Carbon Monoxide-Releasing Molecule Attenuates Allograft Airway Rejection T. Ohtsuka, T. Goto, M. Anraku, M. Kohno, H. Nomori. Section of Thoracic Surgery, Keio University, Tokyo, Japan. Purpose: Acute rejection after lung transplantation is the main risk factor for the development of bronchiolitis obliterans syndrome (BOS). Carbon monoxide (CO) can provide beneficial antiapoptotic and anti-inflammatory effects in the context of ischemia–reperfusion injury (IRI), and may serve to limit tissue inflammation and injury in the setting of airway transplant rejection. Here we tested the ability of carbon monoxide releasing molecules (CORMs) to prevent airway rejection in established mouse orthotopic and heterotopic trachea transplant models. Methods and Materials: Tracheal grafts from wild type BALB/c or C57BL/6 were transplanted to C57BL/6 recipients. Experimental groups were treated with multiple doses of tricarbonyl dichloro ruthenium(II) dimer (CORM-2), at 10 mg/kg 1 hour before transplantation, and 1, 3, and 5 days after transplantation (n ⫽ 6 per group). Histopathologic evaluation of luminal obliteration was blindly reviewed at days 7 in orthotopic model, at days 14 in heterotopic model.

Results: Allografts treated with CORM-2 revealed a striking reduction of

S141 thickening in epithelial and subepithelial airway layers (p ⬍ 0.05) in orthotopic trachea transplantation model compared with allografts treated with vehicle. In heterotopic trachea transplantation model, allografts treated with CORM-2 showed a reduction of luminal obliteration and fibrosis (p ⬍ 0.05) compared with allografts treated with vehicle (Figure 1). Conclusions: These data implicate CORM-2-derived CO as a critical negative modulator in experimental BOS, and may represent a target for the therapeutic intervention of acute and chronic lung allograft rejection. 397 Ex Vivo Adenoviral IL-10 Gene Therapy in a Pig Lung Transplantation Survival Model T.N. Machuca, R. Bonato, M. Cypel, J. Yeung, Z. Guan, S. Juvet, D.M. Hwang, M. Chen, T. Saito, T.K. Waddell, M. Liu, S. Keshavjee. Latner Thoracic Surgery Research Laboratories, University Health Network - University of Toronto, Toronto, ON, Canada. Purpose: Clinical translation of adenoviral human IL-10 (AdhIL-10) gene therapy could potentially help to prevent or reduce ischemia-reperfusion injury and allograft rejection. We have recently further refined AdhIL-10 gene delivery using the ex vivo lung perfusion (EVLP) system. The aims of this pre-clinical study were to assess: 1) safety, 2) efficacy of transgene expression and 3) effects of ex vivo AdhIL-10 on allograft function in a large animal survival model. Methods and Materials: Pig donor lungs were preserved according to standard practice with Perfadex for 6h at 4oC and then randomly assigned to 2 groups (n⫽3/group): Control ⫽ 12 hours EVLP, and Gene therapy ⫽ 12 hours EVLP ⫹ AdhIL-10 intra-bronchially. The left lung was then transplanted. The recipient animals were immunosuppressed with cyclosporine, azathioprine and prednisone and sacrificed 7 days post-transplant. Results: Animals in both groups followed the same good clinical postoperative course through the 7 days. Daily blood analysis demostrated normal hematology (CBC), kidney function (urea, creatinine), liver enzymes (AST, ALT), LDH and AP in both groups. ELISA for hIL-10 on daily plasma samples ranged from 507 to 1660 pg/ml and on day 7 in lung homogenates from 1.1 to 55.4 pg/mg protein confirming adequate transgene expression in the AdhIL-10 group. Inflammatory cytokine analysis demonstrated a trend towards lower levels of IL-1beta (p⫽0.05) and IFN-gamma (p⫽0.1) at day 7 in the AdhIL-10 group. Histologic assessment of transplanted lungs at day 7 demonstrated lower grades of acute rejection in the gene therapy group. Conclusions: AdhIL-10 gene therapy administered ex vivo to the donor lung leads to effective gene expression, excellent post transplant lung function and no signs of vector-related toxicity in a large animal survival model. Ex vivo AdhIL-10 demostrates the potential to attenuate early inflammatory and immune mediated events and decrease the severity of acute rejection in this pre-clinical lung transplantation survival study. 398 Successful 24-Hour Ex-Vivo Maintenance of Swine Lungs Using the Organ Care System (OCS™) M. Avsar,1 G. Warnecke,1 F. Ius,1 A. Haverich,1 I. Abdel Fattah,2 A. Hassanein,3 M. Iraniha,4 H. O’Connor.4 1Thoracic and Cardiovascular Surgery, Hannover Medical School, Hannover, Germany; 2Thoracic and Cardiovascular Surgery, Cairo University Medical School, Cairo, Egypt; 3Thoracic and Cardiovascular Surgery, Ain Shams University School of Medicine, Cairo, Egypt; 4Department of Clinical Development, TransMedics, Inc., Andover, MA. Purpose: Over the past decade there has been a global focus on ex-vivo lung perfusion (EVLP) as a technique that could address the major limitations of cold storage in lung transplantation to increase the utilization of existing donors and improve short & long-term outcomes. The OCS™ Lung system is an integrated portable system designed to apply all the reconditioning and evaluation capabilities of EVLP immediately after organ retrieval. In addition, it eliminates ischemic injury to the lungs. Methods and Materials: 7 bilateral adult swine lungs were perfused and ventilated on the OCS™ lung device for 24 hours using OCS perfusion solution supplemented with swine RBCs. The organs were assessed for oxygenation capacity, dynamic compliance and bronchoscopy at 1, 6, 8 &