Original Pre-Clinical Science
Sustained function of genetically modified porcine lungs in an ex vivo model of pulmonary xenotransplantation

https://doi.org/10.1016/j.healun.2013.07.001Get rights and content

Background

Xenotransplantation could provide a solution to the donor shortage that is currently the major barrier to solid-organ transplantation. The ability to breed pigs with multiple genetic modifications provides a unique opportunity to explore the immunologic challenges of pulmonary xenotransplantation.

Methods

Explanted lungs from wild-type and 3 groups of genetically modified pigs were studied: (i) α1,3-galactosyltransferase gene knockout (GTKO); (ii) GTKO pigs expressing the human complementary regulatory proteins CD55 and CD59 (GTKO/CD55-59); and (iii) GTKO pigs expressing both CD55-59 and CD39 (GTKO/CD55-59/CD39). The physiologic, immunologic and histologic properties of porcine lungs were evaluated on an ex vivo rig after perfusion with human blood.

Results

Lungs from genetically modified pigs demonstrated stable pulmonary vascular resistance and better oxygenation of the perfusate, and survived longer than wild-type lungs. Physiologic function was inversely correlated with the degree of platelet sequestration into the xenograft. Despite superior physiologic profiles, lungs from genetically modified pigs still showed evidence of intravascular thrombosis and coagulopathy after perfusion with human blood.

CONCLUSIONS

The ability to breed pigs with multiple genetic modifications, and to evaluate lung physiology and histology in real-time on an ex vivo rig, represent significant advances toward better understanding the challenges inherent to pulmonary xenotransplantation.

Section snippets

Generation of transgenic pigs

GTKO pigs were generated by homologous recombination and cloning by nuclear transfer, as previously described.5 These pigs lack expression of the Gal epitope on all organs and tissues but are otherwise phenotypically normal. Transgenic pigs coexpressing the human complement regulators CD55 and CD59 were generated by conventional oocyte microinjection.6 Transgenic pigs expressing human CD39 were produced by transfection of pig fetal fibroblasts followed by cloning using stable transfectants as

Establishing the ex vivo rig

For the 10 experiments, the mean time from instillation of pneumoplegia to lung explant and ice preservation was 20 ± 2 minutes. Cannulation to the ex vivo rig and initial perfusion with Steen solution took a further 88 ± 2 minutes, after which it took 74 ± 4 minutes to establish stable physiologic ventilator and perfusion function. Wild-type porcine lungs demonstrated stable physiologic function for 6 hours while being continuously perfused with Steen solution (data not shown).

Lung survival and performance

Wild-type lungs

Discussion

In this series of preliminary experiments on ex vivo performance of porcine lungs after addition of human blood, we have demonstrated that genetic modification of the donor results in superior graft function compared with wild-type lungs. Specifically, we were able to demonstrate superior oxygenation of the perfusate and reduced pulmonary vascular resistance when human blood was perfused through GTKO, GTKO/CD55-59 and GTKO/CD55-59/CD39 lungs compared with wild-type porcine lungs. Although there

Disclosure statement

The authors have no conflicts of interest to disclose.

This study was supported by a Norman E. Shumway International Society for Heart and Lung Transplantation (ISHLT) Career Development Award (to G.W.) and by the Margaret Pratt Foundation.

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