Volume 18, Issue 6 pp. 355-368
Original Article

Transgenic expression of human heme oxygenase-1 in pigs confers resistance against xenograft rejection during ex vivo perfusion of porcine kidneys

Björn Petersen

Björn Petersen

Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany

Both authors contributed equally to this work.

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Wolf Ramackers

Wolf Ramackers

Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery

Both authors contributed equally to this work.

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Andrea Lucas-Hahn

Andrea Lucas-Hahn

Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany

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Erika Lemme

Erika Lemme

Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany

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Petra Hassel

Petra Hassel

Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany

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Anna-Lisa Queißer

Anna-Lisa Queißer

Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany

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Doris Herrmann

Doris Herrmann

Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany

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Brigitte Barg-Kues

Brigitte Barg-Kues

Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany

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Joseph W. Carnwath

Joseph W. Carnwath

Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany

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Johannes Klose

Johannes Klose

Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery

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Andreas Tiede

Andreas Tiede

Haematology, Haemostasis and Oncology

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Lars Friedrich

Lars Friedrich

Clinic for Anaesthesiology and Intensive Care, Hanover Medical School, Hannover, Germany

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Wiebke Baars

Wiebke Baars

Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery

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Reinhard Schwinzer

Reinhard Schwinzer

Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery

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Michael Winkler

Michael Winkler

Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery

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Heiner Niemann

Heiner Niemann

Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany

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First published: 15 December 2011
Citations: 102
Address reprint requests to Heiner Niemann, Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Hoeltystrasse 10, Mariensee, D-31535 Neustadt, Germany (E-mail: [email protected])

[Correction added after initial online publication on December 15, 2011: Addendum has been included.]

Abstract

Petersen B, Ramackers W, Lucas-Hahn A, Lemme E, Hassel P, Queißer A-L, Herrmann D, Barg-Kues B, Carnwath JW, Klose J, Tiede A, Friedrich L, Baars W, Schwinzer R, Winkler M, Niemann H. Transgenic expression of human heme oxygenase-1 in pigs confers resistance against xenograft rejection during ex vivo perfusion of porcine kidneys. Xenotransplantation 2011; 18: 355–368. © 2011 John Wiley & Sons A/S.

Abstract:

Background: The major immunological hurdle to successful porcine-to-human xenotransplantation is the acute vascular rejection (AVR), characterized by endothelial cell (EC) activation and perturbation of coagulation. Heme oxygenase-1 (HO-1) and its derivatives have anti-apoptotic, anti-inflammatory effects and protect against reactive oxygen species, rendering HO-1 a promising molecule to control AVR. Here, we report the production and characterization of pigs transgenic for human heme oxygenase-1 (hHO-1) and demonstrate significant protection in porcine kidneys against xenograft rejection in ex vivo perfusion with human blood and transgenic porcine aortic endothelial cells (PAEC) in a TNF-α-mediated apoptosis assay.

Methods: Transgenic and non-transgenic PAEC were tested in a TNF-α-mediated apoptosis assay. Expression of adhesion molecules (ICAM-1, VCAM-1, and E-selectin) was measured by real-time PCR. hHO-1 transgenic porcine kidneys were perfused with pooled and diluted human AB blood in an ex vivo perfusion circuit. MHC class-II up-regulation after induction with IFN-γ was compared between wild-type and hHO-1 transgenic PAEC.

Results: Cloned hHO-1 transgenic pigs expressed hHO-1 in heart, kidney, liver, and in cultured ECs and fibroblasts. hHO-1 transgenic PAEC were protected against TNF-α-mediated apoptosis. Real-time PCR revealed reduced expression of adhesion molecules like ICAM-1, VCAM-1, and E-selectin. These effects could be abrogated by the incubation of transgenic PAECs with the specific HO-1 inhibitor zinc protoporphorine IX (Zn(II)PPIX, 20 μm). IFN-γ induced up-regulation of MHC class-II molecules was significantly reduced in PAECs from hHO-1 transgenic pigs. hHO-1 transgenic porcine kidneys could successfully be perfused with diluted human AB-pooled blood for a maximum of 240 min (with and without C1 inh), while in wild-type kidneys, blood flow ceased after ∼60 min. Elevated levels of d-Dimer and TAT were detected, but no significant consumption of fibrinogen and antithrombin was determined. Microthrombi could not be detected histologically.

Conclusions: These results are encouraging and warrant further studies on the biological function of heme oxygenase-I expression in hHO-1 transgenic pigs in the context of xenotransplantation.

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