Ex Vivo Lung Perfusion Improves the Inflammatory Signaling Profile of the Porcine Donor Lung Following Transplantation

Abstract

Background. Primary graft dysfunction and allograft rejection represent major caveats to successful lung transplantation. Reducing inflammation in donor lungs before transplantation may improve outcomes. Evidence exists that ex vivo lung perfusion (EVLP) can alter the donor lung environment, although the mechanisms remain unclear. This study aimed to characterize the inflammatory signaling profile of the lung following standard and EVLP transplant and delineate the immediate impact on the recipient circulation. Methods. Female recipient pigs (n = 12) were randomized to undergo left lung transplantation from male donors either using the gold standard protocol (static cold storage) or following 3 hours of EVLP. The relative phosphorylation of 44 phosphokinases and the relative expression of 35 apoptosis-related molecules were profiled within the donor lung 24 hours posttransplantation. Results. A global profile of mitochondrial salvage and cell survival was observed in the EVLP lung tissue compared with lungs undergoing standard transplantation. This included increased phosphorylation of downstream prosignaling kinases, including ERK1/2 and FAK. In addition, there was upregulated expression of the antiapoptotic proteins Bcl-2, HSP-70, LIVIN, and PON2 with downregulation of apoptosis inducing mitochondrial associated molecules, including clusterin, cytochrome C, and HTRA2/OMI. In the early postoperative period, there were significantly lower levels of circulating mitochondrial DNA in recipients receiving EVLP lungs compared with a standard transplant (P = 0.016). Genomic DNA did not differ between groups, with donor DNA undetectable at all time points. Conclusions. EVLP alters the inflammatory signaling profile of the donor lung before transplantation, with a global cell survival and antiapoptotic signature.

Challenging the Ex Vivo Lung Perfusion Procedure With Continuous Dialysis in a Pig Model

“…25 Notably, we observed that the cell survival and proliferation profiles were not altered by our different perfusion procedures. Interestingly, upregulation of the antiapoptotic BCL2 and downregulation of the proapoptotic Bax proteins were found 24 h posttransplantation in pig lungs treated with EVLP as compared to in classically transplanted lungs, 4 indicating that the survival pathways induced upon EVLP have a prolonged effect even after transplantation.…”

The potential of ex vivo lung perfusion on improving organ quality and ameliorating ischemia reperfusion injury

“…Collectively, these molecular studies suggesting beneficial effects of EVLP on IRI‐derived inflammation, provide mechanistic insights for various experimental studies that have demonstrated positive effects of EVLP on lung function after transplantation. Hence, porcine lungs transplanted using an EVLP showed a significantly lower acute lung injury score with less interstitial and intra‐alveolar edema, hemorrhage and hyaline formation when compared to lungs procured in the standard of care cold storage group 33 . Moreover, alveolar epithelial cell tight junction integrity, evaluated by zona occludens‐1 protein staining, had been preserved in the cell membranes of porcine lung tissue after EVLP 69 (Figure 1F).…”

“…33 In turn, pro-survival signaling proteins had been upregulated in the EVLP transplanted pig lung (compared with the standard of care) including, ERK1/2, FAK, PDGFRβ, STAT5/6, and members of the Src family of protein tyrosine kinases Fgr, Hck, and Lck indicating cellular proliferation. 33 Consistently, after the third hour of acellular normothermic EVLP of clinically rejected human lungs, the strongest predicted activation in a gene expression array had been cell viability and cell survival in addition to a cellular tight junction recovery seen by upregulation of the tight junction canonical pathway 67 which was confirmed in another study using immunohistochemistry. 69 Further underscoring the concept of tissue repair, EVLP of explanted rat lungs has been shown to modulate the pulmonary turnover of high molecular weight hyaluron (Figure 1E), a tissue protective glycosaminoglycan contributing to the maintenance of lung tissue homeostasis.…”

“…Hence, in porcine donor lungs EVLP preservation prior to transplantation has been associated with the upregulation of a broad range of anti‐apoptotic proteins, such as Bcl‐2, HSP‐70, LIVIN, and PON2, with downregulation of apoptosis inducing mitochondrial‐associated molecules, including clusterin, cytochrome C, and HTRA2/OMI following transplantation 33 . In turn, pro‐survival signaling proteins had been upregulated in the EVLP transplanted pig lung (compared with the standard of care) including, ERK1/2, FAK, PDGFRβ, STAT5/6, and members of the Src family of protein tyrosine kinases Fgr, Hck, and Lck indicating cellular proliferation 33 . Consistently, after the third hour of acellular normothermic EVLP of clinically rejected human lungs, the strongest predicted activation in a gene expression array had been cell viability and cell survival in addition to a cellular tight junction recovery seen by upregulation of the tight junction canonical pathway 67 which was confirmed in another study using immunohistochemistry 69 .…”

Minimizing Ischemia Reperfusion Injury in Xenotransplantation

“…EVLP was introduced to the clinic in 2011 for high-risk lungs (290). Preclinical studies in a porcine model have shown that EVLP may reduce IRI by decreasing the insult on the endothelium or by reducing the inflammatory cytokine load (291)(292)(293). Controlling reperfusion by means of reversing oxygen debt in the absence of mechanical stress/strain and in the absence of platelets and neutrophils are also beneficial attributes of EVLP (291,292).…”