Complement dependent early immunological responses during ex vivo xenoperfusion of hCD46/HLA-E double transgenic pig forelimbs with human blood.

Bongoni, Anjan K.; Kiermeir, David; Jenni, Hansjörg; Bähr, Andrea; Ayares, David; Klymiuk, Nikolai; Wolf, Eckhard; Vögelin, Esther; Constantinescu, Mihai A.; Seebach, Jörg D.; Rieben, Robert (2014). Complement dependent early immunological responses during ex vivo xenoperfusion of hCD46/HLA-E double transgenic pig forelimbs with human blood. Xenotransplantation, 21(3), pp. 230-243. Wiley 10.1111/xen.12090

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BACKGROUND Besides α1,3-galactosyltransferase gene (GGTA1) knockout, several transgene combinations to prevent pig-to-human xenograft rejection are currently being investigated. In this study, the potential of combined overexpression of human CD46 and HLA-E to prevent complement- and NK-cell-mediated xenograft rejection was tested in an ex vivo pig-to-human xenoperfusion model. METHODS α1,3-Galactosyltransferase knockout heterozygous, hCD46/HLA-E double transgenic (transgenic) as well as wild-type pig forelimbs were ex vivo perfused with whole, heparinized human and autologous pig blood, respectively. Blood samples were analyzed for the production of porcine and/or human inflammatory cytokines as well as complement activation products. Biopsy samples were examined for deposition of human and porcine C3b/c, C4b/c, and C6 as well as CD62E (E-selectin) and CD106 (VCAM-1) expression. Apoptosis was measured in the porcine muscle tissue using TUNEL assays. Finally, the formation of thrombin-antithrombin (TAT) complexes was measured in EDTA plasma samples. RESULTS No hyperacute rejection was seen in this model. Extremity perfusions lasted for up to 12 h without increase in vascular resistance and were terminated due to continuous small blood losses. Plasma levels of porcine cytokines IL1β, IL-6, IL-8, IL-10, TNF-α, and MCP-1 as well as human complement activation markers C3a (P = 0.0002), C5a (P = 0.004), and soluble C5b-9 (P = 0.03) were lower in blood perfused through transgenic as compared to wild-type limbs. Human C3b/c, C4b/c, and C6 as well as CD62E and CD106 were deposited in tissue of wild-type limbs, but significantly lower levels (P < 0.0001) of C3b/c, C4b/c, and C6 deposition as well as CD62E and CD106 expression were detected in transgenic limbs perfused with human blood. Transgenic porcine tissue was protected from xenoperfusion-induced apoptosis (P < 0.0001). Finally, TAT levels were significantly lower (P < 0.0001) in transgenic limb as compared to wild-type limb xenoperfusions. CONCLUSION Transgenic hCD46/HLA-E expression clearly reduced humoral xenoresponses since all, the terminal pathway of complement activation, endothelial cell activation, muscle cell apoptosis, inflammatory cytokine production, as well as coagulation activation, were all downregulated. Overall, this model represents a useful tool to study early immunological responses during pig-to-human vascularized xenotransplantation in the absence of hyperacute rejection.

Item Type:

Journal Article (Original Article)

Division/Institute:

04 Faculty of Medicine > Department of Orthopaedic, Plastic and Hand Surgery (DOPH) > Clinic of Plastic and Hand Surgery > Plastic, Reconstructive and Aesthetic Surgery
04 Faculty of Medicine > Department of Orthopaedic, Plastic and Hand Surgery (DOPH) > Clinic of Plastic and Hand Surgery > Hand Surgery
04 Faculty of Medicine > Department of Orthopaedic, Plastic and Hand Surgery (DOPH) > Clinic of Plastic and Hand Surgery
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 > Forschungsgruppe Handchirurgie
09 Interdisciplinary Units > Microscopy Imaging Center MIC

UniBE Contributor:

Bongoni, Anjan; Kiermeir, David; Vögelin, Esther; Constantinescu, Mihai Adrian and Rieben, Robert

Subjects:

600 Technology > 610 Medicine & health

ISSN:

0908-665X

Publisher:

Wiley

Language:

English

Submitter:

Jörg Arnoldi

Date Deposited:

05 May 2014 13:24

Last Modified:

20 Feb 2019 09:40

Publisher DOI:

10.1111/xen.12090

PubMed ID:

24635052

Uncontrolled Keywords:

HLA-E, apoptosis, complement, endothelial cells, hCD46, pig-to-human xenotransplantation, transgenic pig

BORIS DOI:

10.7892/boris.49948

URI:

https://boris.unibe.ch/id/eprint/49948

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