PURPOSE: Ischemia-reperfusion injury after lung transplantation (LTx) is associated with early graft dysfunction, increased morbidity, and mortality. MicroRNA (miR) are identified as critical regulators of gene expression which could provide potential targets for novel gene therapy. Herein, we aim to examine the feasibility of using the ex vivo lung perfusion (EVLP) platform to study miR expression change in human lungs in response to cold ischemia and ex-vivo reperfusion (CI/EVR). METHODS: Prior to initiating the clinical EVLP program at our institution, our group has established a biobank of human lung tissue according to an approved (cellular) EVLP research protocol. Twenty-four donor lungs that did not meet the ideal criteria for LTx or did not match to an available recipient were used for this study. Samples were obtained after cold ischemia (median 6, interquartile range 5-13 hrs), and after 2 hrs of EVLP. Control samples were obtained from different 6 donors that were stored directly without cold ischemia or reperfusion. MicroRNA expression profiling of lung tissue was performed using next generation sequencing. MicroRNAs with the most significant fold change were validated and detected in the tissues using In-situ hybridization assay. RESULTS: Assessment of inflammatory response of the EVLP model using cytokines array showed significant up-regulation of IL-1β, IL-6, IL-8, IL-10, and TNF-α in the perfusate after 2 hours EVLP. MicroRNA sequencing identified significant expression change of a total of 21 miR after cold ischemia and 47 miR after ex-vivo reperfusion when compared to control group. Validation using quantitative PCR showed significant up-regulation of miR-17 & miR548b after CI/EVR. Downstream analysis identified abundant inflammatory and immunologic targets for miR-17 and miR-548b that are known mediators of lung injury. Expression of miR-17 in the lung tissue had a significant negative correlation with IL-8 levels in the perfusate (R = -0.8, p=0.001). In-situ hybridization assays detected positive expression of miR-17 & miR548b in alveolar epithelial cells after CI/EVR. R. CONCLUSION: In this study, we were able to utilize EVLP model to study miR signature in human lungs after CI/EVR. The finding that alveolar epithelial cells up-regulate miR-17 & miR548b in response to CI/EVT is novel and warrant further investigation.
|Journal||The Journal of heart and lung transplantation : the official publication of the International Society for Heart Transplantation|
|Publication status||Published - 2020 Apr 1|
ASJC Scopus subject areas
- Pulmonary and Respiratory Medicine
- Cardiology and Cardiovascular Medicine