Transgenic expression of human heme oxygenase-1 in pigs confers resistance against xenograft rejection during ex vivo perfusion of porcine kidneys
Correction(s) for this article
-
Corrigendum
- Volume 19Issue 3Xenotransplantation
- pages: 212-212
- First Published online: June 15, 2012
Björn Petersen
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Both authors contributed equally to this work.
Search for more papers by this authorWolf Ramackers
Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery
Both authors contributed equally to this work.
Search for more papers by this authorAndrea Lucas-Hahn
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorErika Lemme
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorPetra Hassel
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorAnna-Lisa Queißer
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorDoris Herrmann
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorBrigitte Barg-Kues
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorJoseph W. Carnwath
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorJohannes Klose
Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery
Search for more papers by this authorLars Friedrich
Clinic for Anaesthesiology and Intensive Care, Hanover Medical School, Hannover, Germany
Search for more papers by this authorWiebke Baars
Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery
Search for more papers by this authorReinhard Schwinzer
Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery
Search for more papers by this authorMichael Winkler
Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery
Search for more papers by this authorHeiner Niemann
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorBjörn Petersen
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Both authors contributed equally to this work.
Search for more papers by this authorWolf Ramackers
Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery
Both authors contributed equally to this work.
Search for more papers by this authorAndrea Lucas-Hahn
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorErika Lemme
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorPetra Hassel
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorAnna-Lisa Queißer
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorDoris Herrmann
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorBrigitte Barg-Kues
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorJoseph W. Carnwath
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this authorJohannes Klose
Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery
Search for more papers by this authorLars Friedrich
Clinic for Anaesthesiology and Intensive Care, Hanover Medical School, Hannover, Germany
Search for more papers by this authorWiebke Baars
Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery
Search for more papers by this authorReinhard Schwinzer
Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery
Search for more papers by this authorMichael Winkler
Transplant Laboratory, Clinic for Visceral- and Transplantation Surgery
Search for more papers by this authorHeiner Niemann
Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut (FLI), Neustadt, Germany
Search for more papers by this author[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.
Supporting Information
Figure S1. PED-SV15 cells were transfected with the pTSG1 vector containing human HO-1 (PED-SV15, hHO-1 cells) or no insert (PED-SV15, vector control). Wild-type PED-SV15 cells and transfectants were lysed, the proteins were separated by 12.5% SDS–PAGE, transferred to PVDF membrane, and probed with an antibody detecting the FLAG-tag (polyclonal rabbit antibody, Rockland Immunochemicals, Gilbertsville, USA). As a protein-loading control, the same blots were stripped and reblotted with an anti-α-actin antibody (polyclonal mouse IgG2; Sigma, Deisenhofen, Germany). The anti-FLAG antibody cross-reacted with a protein of about 28 to 30 kDa in all samples (◀). The hHO-1/FLAG fusion protein (←) corresponding to a relative molecular mass of 32 kDa was only detected in PED-SV15-hHO-1 cells.
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