Adenovirus-mediated adiponectin gene transfer in a rat model of orthotopic left lung transplantation
Sayed Himmat1, Shingo Takahara3, Lin Fu Zhu6, Xiuhua Wang1, Xiao Qi1, Sanaz Hatami1, Katie-Marie Buswell6, Rayan Edgar6, Amy Barr3, Jody Levasseur3, Keir Forgie2,5, Jason P Acker4, Jason Dyck3, Darren Freed2,5, Jayan Nagendran2,5.
1Surgery Department, University of Alberta, Edmonton, AB, Canada; 2Cardiac Surgery Department , University of Alberta, Edmonton, AB, Canada; 3Department of Pediatrics, University of Alberta, Edmonton, AB, Canada; 4Department of Laboratory Medicine and Pathology, University of Alberta, Edmonton, AB, Canada; 5Mazankowski Alberta Heart Institute, Edmonton, AB, Canada; 6Ray Rajotte Surgical Medical Research Institute, University of Alberta, Edmonton, AB, Canada
Introduction: Primary graft dysfunction (PGD) can occur early following lung transplantation (LT) and correlates with acute and chronic lung allograft rejection. Abrupt inflammatory and oxidative stress responses occur within minutes of LT. This ischemia-reperfusion injury (IRI) is believed to be responsible for initiating PGD. Interestingly, adiponectin (APN) a bioactive peptide, has significant anti-inflammatory and cytoprotective properties to pulmonary endothelium and airway epithelium. Thus, gene delivery of APN to donated lungs may decrease the incidence of PGD from IRI. We aim to assess the feasibility and early post-transplant effect of adenovirus-mediated adiponectin gene therapy to donor lungs.
Methods: Male Lewis rats (350-450 g) were randomly allocated into 3 groups (N=5 / group), where they underwent in vivo transbronchial instillation of 1) Control: 400 µL of PBS+10% glycerol, 2) Ad-mCherry: adenovirus-mCherry 400 µL of 1.5*1010 PFU, and 3) Ad-APN: adenovirus-adiponectin 400 µL of 1.5*1010 PFU. After 24 h, transduced lungs were procured, flushed with ice-cold low-potassium dextran, and preserved in cold ischemia for ~3 h. Then, left donor lungs were orthotopically transplanted into recipient male Sprague Dawley rats (350-450 g). Following 2 h of reperfusion, blood gas samples were analyzed from the aorta and left pulmonary vein. Finally, recipient rats were euthanized, plasma and lung tissue were collected for wet/dry ratio and further analysis.
Results: Target gene expression in Ad-APN and Ad-mCherry lungs was confirmed using tissue immunofluorescent staining. During the 2 h reperfusion, all recipient rats had stable oxygen saturation (SPO2% >90). The mean arterial partial pressure of oxygen (pO2) for groups 1-3 was 191.0, 193.4, and 231.5 mmHg; the estimated % contribution of donor lung to recipient pO2 was 73.2, 46.0, and 46.5, respectively. On average, wet/dry ratio of left donor lung was 8.7 for Control, 9.5 for Ad-mCherry, and 6.7 for the Ad-APN group. Plasma multiplex cytokine assay showed elevated levels of pro-inflammatory cytokines: IL-6, IL-1β, and IL-18 in Ad-mCherry compared to Control and Ad-APN. Monocyte chemoattractant protein (MCP-1) secretion was significantly higher in the Ad-mCherry group than APN (p = 0.02).
Conclusion: Adenovirus treated lungs with Ad-mCherry, and Ad-APN exhibited comparable lung function post-transplant. Of interest, APN expression modulated the vector-induced inflammation in the Ad-APN group. Furthermore, APN induced the downregulation of MCP-1 secretion by alveolar macrophages. This will impair the chemoattraction of recipient monocytes and neutrophils, therefore potentially attenuating lung allograft inflammation and IRI post transplantation.
right-click to download
