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G Andres

Publications and source records attributed to G Andres.

At least 19 recordsLinked to original sources
PubMed2000

Role of platelet-activating factor in functional alterations induced by xenoreactive antibodies in porcine endothelial cells.

BACKGROUND: Platelet-activating factor (PAF) is a phospholipid mediator of inflammation which has been implicated in rejection. The interaction of anti-alpha-galactosyl natural antibodies (anti-alpha gal Abs) with endothelial cells is the initial step for the development of xenograft rejection. In our study, we stimulated porcine aortic endothelial cells (PAEC) with anti-alpha gal IgG to investigate the synthesis of PAF from PAEC and its biological consequences. METHODS AND RESULTS: PAF was extracted and chromatographically purified from cultured PAEC stimulated with baboon anti-alpha gal Abs. The Abs induced a dose-dependent synthesis of PAF peaking after 30 min of incubation, and decreasing thereafter. Concomitant cell shape change, motility, and cytoskeleton redistribution were observed. These events were prevented by addition of a panel of PAF-receptor antagonists. An SV40 T-large antigen-immortalized PAEC line was engineered to express PAF acetyl-hydrolase (PAF-AH) cDNA, the major PAF-inactivating enzyme. These transfected cells exposed to anti-alpha gal Abs showed reduced cell contraction and motility compared with empty vector-transfected cells. Moreover, in PAEC stimulated with anti-alpha gal Abs, the synthesis of PAF promoted the adhesion of a monocytic cell line as shown by the inhibitory effect of PAF-receptor antagonists and of PAF-AH expression. Finally, studies on cell monolayer demonstrated an enhanced permeability 48 hr after exposure to anti-alpha gal Abs, and this increase was prevented by PAF-inactivation and by PAF-receptor blockade. CONCLUSIONS: These results demonstrate that on stimulation with anti-alpha gal Abs, PAEC synthetize PAF which can contribute to several vascular events involved in xenograft rejection.

Animals↗
PubMed2000

alpha-galactosyl epitopes on glycoproteins of porcine renal extracellular matrix.

BACKGROUND: The pig is the donor animal of choice for human xenotransplantation. In the most relevant pig-to-baboon model, pig organs transplanted into baboons are hyperacutely rejected by natural xenoantibodies, which mainly bind to alpha-galactosyl (alphaGal) epitopes expressed at the surface of endothelial cells. Recent advances in controlling hyperacute rejection have led to improved survival of these xenografts, and it is now important to identify alphaGal binding sites in other cells and tissues that may be subject to immunologic attack. To this end, we have studied whether alphaGal antibodies bind to glycated proteins of the extracellular matrix in the kidney and other organs most likely to be used for human xenotransplantation. METHODS: High-titer anti-alphaGal antibodies, similar to human natural xenoantibodies, were prepared in baboons, and their reactivity with components of pig extracellular matrix was tested by serology and immunohistology. RESULTS: The antibodies recognized epitopes of immobilized murine, bovine or porcine thyroglobulin, laminin, heparan sulfate proteoglycans, and fibronectin. In sections of pig tissue, the antibodies bound to endothelial and certain epithelial cells, as shown in previous studies, and also to mesenchymal cells, basement membranes, and extracellular matrices, in which they colocalized with matrix glycoproteins, especially laminin and heparan sulfate proteoglycans. CONCLUSIONS: These results suggest that when pig xenografts can be made to survive for prolonged periods, the reactivity of alphaGal antibody with matrix molecules can induce basement membrane and matrix lesions similar to those induced in laboratory animals by antilaminin and antiheparan sulfate proteoglycans antibodies.

Animals↗
PubMed1999

Interaction of baboon anti-alpha-galactosyl antibody with pig tissues.

As barriers to xenotransplantation are surmounted, such as suppression of hyperacute rejection allowing improved graft survival, it becomes important to define longer-term host-xenograft interactions. To this end we have prepared in baboons high titer anti-alpha-Galactosyl (alphaGal) and anti-porcine aortic endothelial cell antibodies, similar to human natural xenoantibodies and reactive with epitopes of thyroglobulin, laminin, and heparan sulfate proteoglycans. When injected into pigs with a protocol similar to that used in the rat to show the nephritogenic potential of heterologous anti-laminin and anti-heparan sulfate proteoglycan antibodies, baboon immunoglobulins bound first to renal vascular endothelium, and later to interstitial cells, especially fibroblasts and macrophages, and to antigens in basement membranes and extracellular matrix, where they colocalized with laminin- and heparan sulfate proteoglycan-antibodies, and with bound Griffonia simplicifolia B4. A similar binding was observed in other organs. The pigs did not develop an acute complement-dependent inflammation, but rather chronic lesions of the basement membranes and the extracellular matrix. Incubation of renal fibroblasts with baboon anti-alpha-Galactosyl antibodies resulted in increased synthesis of transforming growth factor-beta and collagen, suggesting a possible basis for the fibrotic response. The results demonstrate that in this experimental model a consequence of alphaGal antibody interaction with porcine tissues, is immunoreactivity with alphaGal on matrix molecules and interstitial cells, priming mechanisms leading to fibrosis resembling that in chronic allograft rejection. The possibility that similar lesions may develop in long-surviving pig xenografts is discussed.

Animals↗
PubMed1999

Alpha-galactosyl antibody redistributes alpha-galactosyl at the surface of pig blood and endothelial cells.

The interaction of antibodies with cell surface antigens may induce redistribution of immune complexes, followed by antigen depletion, with increased resistance to injurious effect of antibody and complement (antigenic modulation). Human natural antibodies to Gal alpha 1,3Gal beta 1,4GlcNAc-R (alpha Gal) epitopes expressed at the surface of pig cells are a major obstacle to xenotransplantation. Recent studies have shown that these antibodies do not modulate alpha Gal, but the morphological consequences of the antigen-antibody interaction are unknown. Pig blood and endothelial cells, were exposed to baboon alpha-Gal antibodies, and studied by immunofluorescence and phase contrast microscopy, flow cytometry, and inhibition enzyme-linked immunosorbent assay. In cells studied at 4 degrees C or fixed, alpha Gal was diffusely expressed at the surface. After cross-linking at 37 degrees C, antigenic modulation did not occur, but granular redistribution of alpha Gal immune complexes was seen in all cell types. In other systems a similar redistribution is known to induce perturbation of the plasma membrane/cytoskeletal structure with changes in adhesive properties, gene regulation, and T cell activation, which could be important if pig xenografts will be made to survive for prolonged periods.

Animals↗
PubMed1999

Membranous glomerulonephritis induced in the pig by antibody to angiotensin-converting enzyme: considerations on its relevance to the pathogenesis of human idiopathic membranous glomerulonephritis.

In the course of studies on the humoral consequences of swine to primate xenotransplantation, the investigators induced formation of glomerular subepithelial immune deposits and tubular lesions in pigs injected with heterologous antibody to angiotensin-converting enzyme. This study describes the morphology of the lesions, discusses their mechanism, explains their relevance for understanding the pathogenesis of human idiopathic membranous glomerulonephritis, and proposes future directions for investigations.

Animals↗
PubMed1998

Experimental Goodpasture's syndrome in Wistar-Kyoto rats immunized with alpha3 chain of type IV collagen.

BACKGROUND: Glomerulonephritis and lung hemorrhage of autoimmune Goodpasture syndrome develop due to immune reactions against epitope(s) of the non-collagenous (NC1) domain of alpha3-chain of type IV collagen [alpha3(IV) NC1]. Whether thymic mechanisms have a role in the loss of tolerance to the Goodpasture epitope has not been established. We studied the renal and pulmonary effects of immunization with different forms (monomer, dimer, or hexamer) of alpha3(IV) NC1 collagen in Wistar-Kyoto (WKY) rats, and assessed whether the intrathymic inoculation of the antigen may protect against anti-GBM disease. METHODS: WKY rats were immunized with bovine alpha3(IV) monomer, dimer, or hexamer, or with alpha3(IV) NC1 synthetic peptide. Renal function, kidney and lung immunohistology, and circulating and tissue bound antibodies to type IV collagen chains were analyzed. Effects of intrathymic inoculation of antigen on subsequent disease induction were analyzed in WKY rats given alpha3(IV) NC1 dimer or GBM preparation intrathymically 48 hours before immunization. RESULTS: Proteinuria, linear IgG deposition in GBM, and crescentic glomerulonephritis developed in WKY rats immunized with alpha3(IV) NC1 dimer or hexamer. Lesions were dose-dependent upon injections of 10 to 100 microgram dimer. The alpha3(IV) NC1 monomer induced less severe proteinuria and no crescents. Pulmonary hemorrhage was detectable in 35% of rats immunized with 25 to 100 microgram alpha3(IV) NC1 dimer; alpha3(IV) synthetic peptide (36 carboxyl terminal) did not induce disease. Rats injected intrathymically with up to 100 microgram alpha3(IV) NC1 dimer or with GBM 48 hours before immunization were not protected against subsequent development of proteinuria and glomerulonephritis. CONCLUSIONS: These findings document that glomerulonephritis and lung hemorrhage can be elicited in WKY rats by immunization with alpha3(IV) NC1. Failure of the intrathymic inoculation of antigen to prevent disease suggests that immunological tolerance cannot be achieved by this intervention, in contrast to other autoimmune conditions, and may imply independent roles for cellular and humoral nephritogenic pathways in anti-GBM nephritis.

Animals↗
PubMed1996

In vivo modulation of CD26 (dipeptidyl peptidase IV) in the mouse: effects of polyreactive and monoreactive antibodies.

We previously reported that intravenous injections in rabbits or guinea pigs of divalent antibodies to purified protein or carbohydrate antigens located mainly on endothelial cells induce acute pulmonary edema, which is often lethal. Surviving animals develop resistance to the injurious effect of subsequent injection of antibodies (adaptation), associated with shedding of antigen-antibody complexes from endothelial cells. In the present study, we investigated and compared in mice the effects of 3-day multiple injections of two different rabbit antibody (IgG) preparations against antigens expressed mainly at the surface of epithelial cells. The first preparation contained antibodies to a single transmembrane protein, CD26 (dipeptidyl peptidase IV [DPP IV]) (monoreactive anti-DPP IV IgG); the second contained antibodies against multiple antigens of the renal tubular brush border (BB), including DPP IV (polyreactive anti-BB IgG). Both IgG preparations caused loss of DPP IV from the organs studied, as shown by reduction in enzyme activity in tissue homogenates and by immunofluorescence microscopy, which showed loss of DPP IV from cell surface. However, the monoreactive anti-DPP IV IgG induced considerably greater reduction than polyreactive anti-BB IgG. Loss of DPP IV from the cell surface probably occurred by shedding of immune complexes into vascular and extravascular fluids, including bile and urine. The results may have relevance to hyperacute rejection of xenografts, as from pigs to primates. Since human natural antibodies that bind to porcine cells are polyreactive, a new prophylactic strategy for hyperacute rejection might be based on down-regulation of the major xenogeneic antigen, alpha-galactosyl, by injecting donor animals with monoreactive alpha-galactosyl antibodies before transplantation.

Amino Acid Sequence↗
PubMed1996

Identification of a novel inducible cell-surface ligand of CD5 on activated lymphocytes.

CD5 is a 67-kD glycoprotein that is expressed on most T lymphocytes and on a subset of mature B cells. Although its physiologic function is unknown, several lines of evidence suggest that CD5 may play a role in the regulation of T cell activation and in T cell-antigen presenting cell interactions. Using a CD5-immunoglobulin fusion protein (CD5Rg, for receptorglobulin) we have uncovered a new CD5 ligand (CD5L) expressed on the surface of activated splenocytes. Stimulation of murine splenocytes with anti-CD3 and anti-CD28 antibodies induce transient expression of CD5L on B lymphocytes that lasts for approximately 72 h. Binding of CD5Rg to activated splenocytes is trypsin resistant and independent of divalent cations. However, it is pronase sensitive and dependent on N-linked glycosylation of CD5, since treatment of CD5Rg with PNGaseF on N-glycanase completely abrogates its ability to bind activated splenocytes. It addition to splenocytes, CD5L is expressed on activated murine T cell clones. Immunoprecipitation, antibody, and recombinant protein blocking studies indicate that CD5L is distinct from CD72, which has been proposed to be a CD5 ligand. To determine whether CD5-CD5L interaction might play a role in vivo, we tested the effect of CD5Rg in a murine model of antibody-mediated membranous glomerulonephritis. Injection of CD5Rg was found to abrogate development of the disease. Taken together, our results help identify a novel ligand of CD5 and propose a role for CD5 in the regulation of immune responses.

Animals↗
PubMed1996

Distinct regulatory roles of lymphocyte costimulatory pathways on T helper type-2 mediated autoimmune disease.

We assessed the role of CD40-CD40L, cytotoxic T lymphocyte (CTL)A4/CD28-B7s, and CD2-CD48/CD58 lymphocyte costimulatory pathways in the development of mercury chloride (HgCl2)-induced autoimmune disease in mice, which is believed to be mediated by T helper (Th) subset Th2. Inhibition of CD40-CD40-L and CTLA4/CD28-B7s interactions by anti-CD40-L antibody and soluble CTLA4-immunoglobulin (Ig) fusion protein, respectively, abrogated the autoimmune disease without affecting interleukin 4 (IL-4) production, showing the importance of physical contact between T and B lymphocytes in the Th2-mediated process. In contrast, two anti-CD2 antibodies that have been shown to induce immunosuppression of Th1-mediated events exacerbated the autoantibody response and augmented IgG1, IgE, and IL-4 production, transforming a mild mesangial glomerulopathy into a severe systemic immune complex disease. These observations demonstrate that manipulation of lymphocyte accessory counterreceptor interactions may affect the course of Th2-associated autoimmune disease and suggest that signals resulting from CD2 engagement play an essential role in the regulation of the Th1-Th2 effector equilibrium.

Animals↗
PubMed1996

Cellular mechanisms of adaptation of grafts to antibody.

New, more effective, strategies of immunosuppression, including those recently designed to induce durable T cell tolerance (by grafting allogeneic or xenogeneic haematopoietic cells into T lymphocyte-depleted recipients), leave humoral rejection as the main barrier to transplantation of vascularized organs between different species. Recent experimental work indicates that hyperacute rejection can be prevented by manipulations of antibodies and complement. In this paper, we review the mechanisms governing the interaction of antibodies with cell surface antigens in vitro and in vivo, and their cellular consequences. Evidence is presented that, in appropriate conditions, antibodies can protect by effecting modification of graft antigenicity (adaptation or accommodation).

Animals↗
PubMed1995

Inhibition of the CD40-CD40ligand pathway prevents murine membranous glomerulonephritis.

Several forms of glomerulonephritis are induced by antibodies against self or foreign antigens. Normal B lymphocyte antibody production requires T cell costimulatory signals provided in part by T cell surface expression of gp39/CD40ligand (CD40L) that engages the B cell receptor CD40 and induces B cell differentiation and immunoglobulin class switching. We assessed the effect of disrupting the CD40L-CD40 costimulatory pathway, using a CD40-Ig fusion protein, on the development of membranous glomerulonephritis (MGN) in the mouse. MGN is induced by mouse antibodies that recognize and bind to exogenously administered rabbit anti-mouse renal tubular brush border (RbAMBB) IgG immobilized in the glomerular capillary wall. MGN did not occur in nude mice, showing the need of the T cell function. C57Bl/10 mice immunized with RbAMBB and treated with CD40-Ig fusion protein displayed a delayed autologous response and absence of MGN lesions, while control fusion proteins failed to prevent the development of the disease. These observations provide evidence that disruption of the CD40-CD40L costimulatory pathway can prevent the development of MGN by suppressing T cell-dependent antibody production.

Animals↗
PubMed1995

Interaction of antibody with Forssman antigen in guinea pigs. A mechanism of adaptation to antibody- and complement-mediated injury.

Forssman antigen is a glycosphingolipid with antigenic specificity determined by extra-membrane haptenic sugars similar to blood group antigens and antigens that are the main barrier to xenogeneic organ transplantation. Herein, we describe the localization of Forssman antigen in guinea pig lungs and kidneys and the consequences of its interaction with antibodies in vitro and in vivo (Forssman reaction). Exposure of cultured guinea pig aortic endothelial cells to Forssman antibodies induced rapid redistribution of antigen-antibody complexes at the cell surface, followed by shedding that occurred by blebbing of plasma membrane as vesicles or fragments, and was associated with disappearance of antigen from the cell surface (antigenic modulation). Guinea pigs surviving frequent intravenous infections of increasing amounts of antibodies, for a total of 20 to 40 lethal doses, developed a partial or complete adaptation to generalized Forssman reaction, and adaptation was associated with partial or complete modulation of Forssman antigen at the surface of the pulmonary and, in minor degree, renal endothelial and epithelial cells. These findings support the hypothesis that modulation of endothelial carbohydrate antigens contributes to adaptation of highly vascularized organs exposed to tolerable levels of allo- or xenoantibodies.

Animals↗
PubMed1994

Effect of CTLA-4 chimeric protein on rat autoimmune anti-glomerular basement membrane glomerulonephritis.

The interaction of the T cell receptor with the antigen/major histocompatibility class II complex is insufficient to induce optimal T cell activation. Co-stimulatory signals, including those provided by CD28/CTLA-4 on T cells and B7 molecules (B7-1, -2, and -3) on antigen-presenting cells, are also required. CD28-B7 interactions can be blocked by a soluble human CTLA-4 chimeric protein (CTLA4Ig). We tested the effect of administration of CTLA4Ig on experimental anti-glomerular basement membrane (GBM) autoimmune glomerulonephritis in Wistar-Kyoto rats induced by immunization with bovine GBM. The disease is characterized by development of antibody to the alpha 3 chain of type IV collagen (Goodpasture's antigen), deposition of rat IgG in GBM, infiltration of the kidney by T cells and macrophages, severe crescent formation and renal failure leading to death in 5-6 weeks. Animals injected with human CTLA4Ig from day 0 to day 14 or to day 35 had reduced disease severity. Beneficial effects were observed even when injections were begun after the onset of glomerulonephritis on day 14. However, the rats developed antibody to the human CTLA4Ig, associated with reduction in levels of circulating CTLA4Ig. The results provide evidence for CD28/CTLA-4 signaling in rat autoimmune glomerulonephritis, and suggest that more effective inhibition of B7-dependent T cell activation, such as might be achieved with homologous CTLA4Ig, could be useful in the treatment of autoimmune diseases.

Abatacept↗
PubMed1994

Organ distribution in rats of two members of the low-density lipoprotein receptor gene family, gp330 and LRP/alpha 2MR, and the receptor-associated protein (RAP).

We investigated immunohistochemically the distribution in rats of the homologous proteins gp330 and the LDL receptor-related protein (LRP/alpha 2MR), and a receptor-associated protein (RAP), and the sites to which soluble exogenous RAP binds. We found gp330 in a restricted group of epithelial cells, including renal proximal tubule cells, podocytes, Type II pneumocytes, cells of the parathyroid, thyroid, epididymis, lining of the uterus, ependyma, retina, ciliary body, yolk sac, and placenta. In these cells gp330 was detected mainly at the cell surface, except for parathyroid and retinal epithelial cells, where diffuse cell staining was found. LRP/alpha 2MR was widely distributed in interstitial cells, notably in fibroblasts and macrophages, and was also present in a selected group of epithelial or specialized cells, including hepatocytes, adrenal cortical cells, follicular cells of the ovary, cells of the choroid plexus, ciliary body, mesangial cells, and some neurons. In certain cells, notably hepatocytes and adrenal cortical cells, LRP/alpha 2MR was detected mainly on the surface, but in others, including macrophages, fibroblasts, and epithelial cells of the choroid plexus and ciliary body, staining throughout the cell was seen. The only cells that clearly expressed both LRP/alpha 2MR and gp330 were retinal and ciliary epithelial cells. RAP was found in intracellular vesicles in all cells that expressed gp330 or LRP/alpha 2MR. RAP was not definitely detected on the cell surface. Binding sites for RAP were found on the surface of those cells with surface gp330 or LRP, and also throughout the cytoplasm in cells with diffuse cellular LRP/alpha 2MR or gp330. Because of their different locations, we conclude that gp330 and LRP/alpha 2MR serve distinct functions in vivo, despite similarities in ligand-binding properties observed in vitro. Since RAP is found largely within cells, its major physiological function may be concerned with intracellular assembly or trafficking of the receptors to which it binds.

Amino Acid Sequence↗
PubMed1993

Antibody-mediated redistribution and shedding of endothelial antigens in the rabbit.

We report the results of studies performed in vitro and in vivo that were designed to explore individual, sequential, and concurrent Ag-antibody interactions at the surface of rabbit endothelial cells. Divalent heterologous antibodies to rabbit lung angiotensin-converting enzyme and to rabbit lung thrombomodulin were employed. In cultured monolayers, both antibodies redistributed the specific Ag and co-redistributed the immunologically unrelated Ag inducing partial or complete disappearance of the Ag from the cell surface (antigenic modulation) in 15 to 60 min. When injected into living rabbits, each antibody induced a rapid (1 to 3 min) redistribution and subsequent modulation of the specific and of the unrelated Ag at the surface of alveolar endothelial cells. Immune complexes, and the unrelated Ag, were shed in the circulation, attaining peak levels 3 to 4 min after the injection; were rapidly bound by platelets, E, and polymorphonuclear leukocytes; and were subsequently found in phagocytic cells in the spleen and in the liver. Thrombomodulin co-shed by angiotensin-converting enzyme antibody and, to a lesser degree, angiotensin-converting enzyme co-shed by thrombomodulin antibody, crossed the glomerular capillary walls and were reabsorbed by the epithelial cells of the proximal tubules within 2 to 3 min. The results show that immunologically unrelated Ag can be passively entrapped during formation of immune complexes at the cell surface, and provide new information on the kinetics of clearance of immune complexes containing endogenous, structural Ag.

Animals↗