Lack of Galactose-α-1,3-Galactose Expression on Porcine Endothelial Cells Prevents Complement-Induced Lysis but Not Direct Xenogeneic NK Cytotoxicity

Baumann, Bettina C.; Forte, Pietro; Hawley, Robert J.; Rieben, Robert; Schneider, Mårten K J; Seebach, Jorg Dieter

doi:10.4049/jimmunol.172.10.6460

Cited by 82 publications

(74 citation statements)

References 62 publications

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“…6) and displayed an EC-typical phenotype including the expression of adhesion receptors important for human leukocyte adhesion (24). The observations that the immortalized cells in this study showed the same results as the primary PAECs, lends support to the validity of previous results obtained using these cell lines as a reliable model for in vitro studies on the role of Gal in pig-to-human xenotransplantation (16,24,45), where the immortalized Gal Ϫ/Ϫ were used. The fact that immortalized and primary Gal ϩ/ϩ PAEC induced comparable cytokine production patterns in human cells, except for only three cytokines (IFN-␥, TNF-␣, and VEGF) exclusively detected when using immortalized cells, support the assumption that the primary and immortalized cells differ only slightly phenotypically, and that these cytokines might not be physiologically relevant in a clinically setting.…”

Section: Discussionsupporting

confidence: 76%

“…The immortalized PAEC line PEDSV.15 (Gal ϩ/ϩ ) was generated as previously described in detail (27). The Gal Ϫ/Ϫ cell line PED2*3.51 was generated from PEDSV.15 by homologous recombination, panning, and limiting dilution cloning (28). All cells were cultured in DMEM (Invitrogen Life Technologies) supplemented with 10% FCS (BioWhittaker), 1 mM sodium pyruvate, 2 mM L-glutamine, 0.1 mM nonessential amino acids, 20 mM HEPES, 50 g/ml gentamicin (all obtained from Invitrogen Life Technologies), and 0.25 g/ml amphotericin B (BioWhittaker).…”

Section: Cell Culturementioning

confidence: 99%

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Cytokine Secretion Depends on Galα(1,3)Gal Expression in a Pig-to-Human Whole Blood Model

Sæthre

Schneider

Lambris

et al. 2008

The Journal of Immunology

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Transplants from α1,3-galactosyltransferase (Gal) gene-knockout pigs to nonhuman primates are largely protected from hyperacute but not acute humoral xenograft rejection. The present study investigates the role of Gal in cytokine responses using a novel pig-to-human whole blood in vitro model, developed for species-specific analysis of porcine and human cytokines. Porcine (n = 7) and human (n = 27) cytokines were measured using ELISA or multiplex technology, respectively. Porcine aortic endothelial cells from control (Gal+/+) and Gal-deficient (Gal−/−) pigs were incubated with human lepirudin anticoagulated whole blood from healthy donors. E-selectin expression was measured by flow cytometry. The C3 inhibitor compstatin and a C5aR antagonist were used to study the role of complement. Cytokine species specificity was documented, enabling detection of 2 of 7 porcine cytokines and 13 of 27 human cytokines in one single sample. Gal+/+ porcine aortic endothelial cells incubated with human whole blood showed a marked complement C5b-9 dependent up-regulation of E-selectin and secretion of porcine IL-6 and IL-8. In contrast, Gal−/− cells responded with E-selectin and cytokine expression which was so weak that the role of complement could not be determined. Human IL-6, IL-8, IFN-γ, MIP-1α, MIP-1β, eotaxin, and RANTES were detected in the Gal+/+ system, but virtually no responses were seen in the Gal−/− system (p = 0.03). The increase in human cytokine release was largely complement dependent and, in contrast to the porcine response, mediated through C5a. Species-specific analysis of cytokine release revealed a marked, complement-dependent response when Gal+/+ pig cells were incubated with human whole blood, compared with Gal−/− cells which induced virtually no cytokine release.

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Section: Discussionsupporting

confidence: 76%

Section: Cell Culturementioning

confidence: 99%

Cytokine Secretion Depends on Galα(1,3)Gal Expression in a Pig-to-Human Whole Blood Model

Sæthre

Schneider

Lambris

et al. 2008

The Journal of Immunology

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“…It is unclear whether the preformed SLs are sufficient or the de novo formation of SLs is necessary for their lytic function. PECs have been widely used to activate xenogeneic NK cytotoxicity (20). When NK cells were allowed to drop onto a monolayer of PECs for 10 min, we observed a large augmentation in the secretory response elicited by Ca 2ϩ , as shown in Fig.…”

Section: Resultsmentioning

confidence: 77%

Rapid biogenesis and sensitization of secretory lysosomes in NK cells mediated by target-cell recognition

Liu

Yang

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

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Natural killer (NK) cells play important roles in defense againsttumor, viral infection, and cell-mediated xenograft rejection through secretion of secretory lysosomes. In this study, we used high time-resolution membrane capacitance measurement and fluorescence-imaging techniques to study the biogenesis and exocytosis of secretory lysosomes in a human NK cell line. We demonstrated a high-affinity Ca 2؉ -dependent exocytosis of secretory lysosomes, which is sensitized further after target-cell stimulation. Our data also suggest an unusual rapid and dramatic de novo formation of secretory lysosomes after target-cell recognition. The rapid biogenesis of secretory lysosomes was blocked by specific protein kinase C inhibitor but not by brefeldin A. We propose that target-cell recognition triggers rapid biogenesis and sensitization of secretory lysosomes in NK cells through activation of PKC.exocytosis ͉ lytic granules ͉ calcium ͉ immune defense

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“…The GalT- KO modification of pigs overcomes acute humoral rejection, but provides no advantage in preventing graft destruction by NK cells (either by direct or by indirect mechanisms). This presents an impending barrier to organ and cellular xenotransplantation, because the recognition of pig cells by NK cells is not dependent on the expression of Gal [35]. Being non-vascularized grafts, direct mechanisms are involved in the destruction of xenogeneic hepatocytes.…”

Section: Natural Killer (Nk) Cellsmentioning

confidence: 99%

Current status of hepatocyte xenotransplantation

Meier

Navarro-Alvarez

Morel

et al. 2015

International Journal of Surgery

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The treatment of acute liver failure, a condition with high mortality, comprises optimal clinical care, and in severe cases liver transplantation. However, there are limitations in availability of organ donors. Hepatocyte transplantation is a promising alternative that could fill the medical need, in particular as the bridge to liver transplantation. Encapsulated porcine hepatocytes represent an unlimited source that could function as a bioreactor requiring minimal immunosuppression. Besides patients with acute liver failure, patients with alcoholic hepatitis who are unresponsive to a short course of corticosteroids are a target for hepatocyte transplantation. In this review we present an overview of the innate immune barriers in hepatocyte xenotransplantation, including the role of complement and natural antibodies; the role of phagocytic cells and ligands like CD47 in the regulation of phagocytic cells; and the role of Natural Killer cells. We present also some illustrations of physiological species incompatibilities in hepatocyte xenotransplantation, such as incompatibilities in the coagulation system. An overview of the methodology for cell microencapsulation is presented, followed by proof-of-concept studies in rodent and nonhuman primate models of fulminant liver failure: these studies document the efficacy of microencapsulated porcine hepatocytes which warrants progress towards clinical application. Lastly, we present an outline of a provisional clinical trial, that upon completion of preclinical work could start within the upcoming 2-3 years

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Lack of Galactose-α-1,3-Galactose Expression on Porcine Endothelial Cells Prevents Complement-Induced Lysis but Not Direct Xenogeneic NK Cytotoxicity

Cited by 82 publications

References 62 publications

Cytokine Secretion Depends on Galα(1,3)Gal Expression in a Pig-to-Human Whole Blood Model

Cytokine Secretion Depends on Galα(1,3)Gal Expression in a Pig-to-Human Whole Blood Model

Rapid biogenesis and sensitization of secretory lysosomes in NK cells mediated by target-cell recognition

Current status of hepatocyte xenotransplantation

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