Multiple genetically modified GTKO/hCD46/HLA-E/hβ2−mg porcine hearts are protected from complement activation and natural killer cell infiltration during ex vivo perfusion with human blood

Abicht, Jan-Michael; Sfriso, Riccardo; Reichart, Bruno; Längin, Matthias; Gahle, Katja; Puga Yung, Gisella L.; Seebach, Jörg D.; Rieben, Robert; Ayares, David; Wolf, Eckhard; Klymiuk, Nikolai; Baehr, Andrea; Kind, Alexander; Mayr, Tanja; Bauer, Andreas (2018). Multiple genetically modified GTKO/hCD46/HLA-E/hβ2−mg porcine hearts are protected from complement activation and natural killer cell infiltration during ex vivo perfusion with human blood. Xenotransplantation, 25(5), e12390. Wiley 10.1111/xen.12390

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Background In pig‐to‐human xenotransplantation, early cellular rejection reactions are mediated by natural killer cells (NK cells). Human NK cells are inhibited by HLA‐E via CD94/NKG2A receptors. To protect porcine grafts against human NK cell responses, transgenic GTKO pigs expressing hCD46 and HLA‐E have been generated. The aim of this study was to test the effect of this genetic modification on xenogeneic, and in particular human NK cell response, using an ex vivo perfusion model of pig hearts with human blood. Methods Cardiopleged and explanted genetically modified (gm) pig hearts (GTKO/hCD46/HLA‐E/hβ2‐microglobulin) and wild‐type (wt) controls (n = 6 each) were reperfused and tested in an 8 hours ex vivo perfusion system using freshly drawn human blood. Cardiac function was evaluated during a 165‐minute period in working heart mode. Myocardial damage, antibody deposition, complement activation, and coagulation parameters were evaluated histologically at the end of perfusion. The number of NK cells in the perfusate was determined by flow cytometry at baseline and at 8 hours; tissue infiltration by NK cells was quantified by immunofluorescence microscopy using NKp46 staining of frozen sections. Results Deposition of IgG (1.2 ± 1 × 107 vs 8.8 ± 2.9 × 106; P < .01), IgM (4.4 ± 3.7 × 106 vs 1.7 ± 1.2 × 106; P < .01), and the complement activation product C4b/c (3.5 ± 1.3 × 106 vs 2.3 × 106 ± 9.4 × 105; P > .01) was lower in gm than wt hearts. NK cell percentages of leukocytes in the perfusate decreased from 0.94 ± 0.77% to 0.21 ± 0.25% (P = .04) during xenoperfusion of wt hearts. In contrast, the ratio of NK cells did not decrease significantly in the gm hearts. In this group, NK cell myocardial infiltration after 480 minutes of perfusion was lower than in wt organs (2.5 ± 3.7 × 104/mm3 vs 1.3 ± 1.4 × 105/mm3; P = .0001). The function of gm hearts was better preserved compared to wt organs, as demonstrated by higher cardiac index during the first 2 hours of ex vivo perfusion. Conclusion GTKO, hCD46, and HLA‐E expression in porcine hearts reduced complement deposition, complement dependent injury, and myocardial NK cell infiltration during perfusion with human blood. This tested combination of genetic modifications may minimize damage from acute human‐anti‐pig rejection reactions and improve myocardial function after xenotransplantation.

Item Type:

Journal Article (Original Article)

Division/Institute:

04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR)
04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > Forschungsbereich Mu50 > Forschungsgruppe Herz und Gefässe
04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > Forschungsbereich Mu50
09 Interdisciplinary Units > Microscopy Imaging Center MIC

Graduate School:

Graduate School for Cellular and Biomedical Sciences (GCB)

UniBE Contributor:

Sfriso, Riccardo and Rieben, Robert

Subjects:

600 Technology > 610 Medicine & health
500 Science > 570 Life sciences; biology

ISSN:

0908-665X

Publisher:

Wiley

Language:

English

Submitter:

Riccardo Sfriso

Date Deposited:

31 Oct 2018 09:49

Last Modified:

23 Oct 2019 10:37

Publisher DOI:

10.1111/xen.12390

PubMed ID:

29536572

BORIS DOI:

10.7892/boris.120729

URI:

https://boris.unibe.ch/id/eprint/120729

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