Biomedical subjects

J L Platt

Publications and source records attributed to J L Platt.

At least 127 records · Page 7Linked to original sources

PubMed1996

Reduction in the level of Gal(alpha1,3)Gal in transgenic mice and pigs by the expression of an alpha(1,2)fucosyltransferase.

Hyperacute rejection of a porcine organ by higher primates is initiated by the binding of xenoreactive natural antibodies of the recipient to blood vessels in the graft leading to complement activation. The majority of these antibodies recognize the carbohydrate structure Gal(alphal,3)Gal (gal epitope) present on cells of pigs. It is possible that the removal or lowering of the number of gal epitopes on the graft endothelium could prevent hyperacute rejection. The Gal(alpha1,3) Gal structure is formed by the enzyme Galbeta1,4GlcNAc3-alpha-D-galactosyltransferase [alpha(1,3)GT; EC 2.4.1.51], which transfers a galactose molecule to terminal N-acetyllactosamine (N-lac) present on various glycoproteins and glycolipids. The N-lac structure might be utilized as an acceptor by other glycosyltransferases such as Galbeta1,4GlcNAc 6-alpha-D-sialyltransferase [alpha(2,6)ST], Galbeta1,4GlcNAc 3-alpha-D-Sialyltransferase [alpha(2,3)ST], or Galbeta 2-alpha-L-fucosyltransferase [alpha(1,2)FT; EC 2.4.1.691, etc. In this report we describe the competition between alpha(1,2)FT and alpha(1,3)GT in cells in culture and the generation of transgenic mice and transgenic pigs that express alpha(1,2)Fr leading to synthesis of Fucalpha,2Galbeta- (H antigen) and a concomitant decrease in the level of Gal(alpha1,3)Gal. As predicted, this resulted in reduced binding of xenoreactive natural antibodies to endothelial cells of transgenic mice and protection from complement mediated lysis.

Animals↗

PubMed1996

Characterization of transgenic pigs expressing functionally active human CD59 on cardiac endothelium.

The critical shortage of human donor organs has generated interest in the potential for porcine to human xenotransplantation. The initial immunological barrier to xenotransplantation is hyperacute rejection, which is mediated by xenoreactive antibodies and complement, and results in rapid and irreversible tissue destruction. While endogenous complement regulatory proteins (CRPs) protect cells from injury caused by autologous complement, they are relatively species specific and most likely ineffectual in this setting. This has led to the hypothesis that expression of human CRPs in transgenic pigs may affect susceptibility to complement-mediated tissue injury in a porcine-to-human xenograft. Using specific lines of transgenic pigs that express low levels of human CD59, a CRP that acts at the terminal stage of the complement cascade, we present evidence that shows that the human CD59 protein inhibits membrane attack complex assembly and reduces tissue damage when the heart is transplanted to a baboon. Examination by immunohistochemistry of transgenic porcine hearts after transplantation revealed markedly reduced deposition of C5b and MAC, but a similar level of C3 deposition as compared with transplanted control hearts. This finding supports the concept that the species specific function of CRPs contributes to the humoral barrier to xenotransplantation and, given the low level of human CD59 protein expression in the porcine heart, argues that the human protein contributes a unique rather than an additive function in regulation of complement in a xenogeneic setting.

Animals↗

PubMed1996

Identification and characterization of a galactosyl peptide mimetic. Implications for use in removing xenoreactive anti-A Gal antibodies.

Gal alpha 1,3 Gal is thought to be the major antigenic epitope present on pig tissues to which XNAs bind. Removal of antibodies directed against that structure may be critical to the success of pig to human xeno-transplantation. As a first step toward the development of ligands capable of removing XNAs, we have used a phage-displayed peptide library to identify a six-amino-acid peptide that binds to the lectin GS-1-B4 (which binds the carbohydrate Gal alpha 1,3 Gal). This peptide blocks the binding of GS-1-B4 to pig aortic endothelial cells. The carbohydrate Gal alpha 1,3 Gal competes with the binding of GS-1-B4 to the peptide, suggesting that they may bind the same site. Using a RBC agglutination assay, we show that this peptide inhibits the agglutination of pig RBCs by heat-inactivated human serum at concentrations similar to that of Gal alpha 1,3 Gal.

Amino Acid Sequence↗

PubMed1996

Human natural killer cells induce morphologic changes in porcine endothelial cell monolayers.

In this study, we have investigated the early in vitro effects of natural killer (NK) cells on porcine aortic endothelial cell (PAEC) monolayers. Incubation of effector cells containing about 70% CD56+ cells on PAEC monolayer led to time-dependent changes in PAEC monolayer morphology. As little as 20 min of incubation resulted in changes in PAEC shape and in the appearance of gaps between the cells. These effects have been observed for up to 6 hr, but not before 20 min or after 6 hr. When NK-depleted effector cells were used, no morphological changes were observed in comparison with the same effectors before depletion; if CD56+ cells were added back, the effects were comparable with those on nondepleted effector cells. There was no detectable NK cell-mediated cytolytic activity during the 1-6 hr of incubation of peripheral blood lymphocytes with PAEC monolayers. These data indicate that NK cells may participate in endothelial cell changes leading to xenograft rejection.

Animals↗

PubMed1996

Shedding of heparan sulfate proteoglycan by stimulated endothelial cells: evidence for proteolysis of cell-surface molecules.

Activation of endothelial cells by cytokines and endotoxin causes procoagulant and pro-inflammatory changes over a period of hours. We postulated that the same functional state might be achieved more rapidly by changes in the metabolism of heparan sulfate, which supports many of the normal functions of endothelial cells. We previously found that binding of anti-endothelial cell antibodies and activation of complement on endothelial cells causes the rapid shedding of endothelial cell heparan sulfate. Here we report the biochemical mechanism responsible for the release of the heparan sulfate. Stimulation of endothelial cells by anti-endothelial cell antibodies and complement resulted in the release of 35S-heparan sulfate proteoglycan and partially degraded 35S-heparan sulfate chains. Degradation of the 35S-heparan sulfate chains was not necessary for release since heparin and suramin prevented cleavage of the heparan sulfate but did not inhibit release from stimulated endothelial cells. The 35S-heparan sulfate proteoglycan released from endothelial cells originated from the cell surface and had a core protein similar in size (70.5 kD) to syndecan-1. Release was due to proteolytic cleavage of the protein core by serine and/or cysteine proteinases since the release of heparan sulfate was inhibited 87% by antipain and 53% by leupeptin. Release of heparan sulfate coincided with a decrease of approximately 7 kD in the mass of the protein core and with a loss of hydrophobicity of the proteoglycan, consistent with the loss of the hydrophobic transmembrane domain. The cleavage and release of cell-surface 35S-heparan sulfate proteoglycan might be a novel mechanism by which endothelial cells may rapidly acquire the functional properties of activated endothelial cells.

Animals↗

PubMed1996

Transplantation of discordant xenografts: a challenge revisited.

Six years ago, Jeffrey Platt and colleagues reviewed the biological hurdles to transplanting organs between species. The ensuing years have allowed the concepts advanced at that time to be tested leading to significant progress in understanding the immunology of xenotransplantation and in developing strategies for potential clinical application. Here, William Parker and colleagues review that progress.

Animals↗

PubMed1996

Isohemagglutinins and xenoreactive antibodies: members of a distinct family of natural antibodies.

Just as anti-blood group A and anti-blood group B antibodies pose a strong humoral barrier to the transplantation of allogeneic organs or blood, xenoreactive natural antibodies directed against Gal alpha 1-3Gal pose a barrier to the transplantation of xenogeneic organs or blood. We tested the idea that, although "natural" iso-hemagglutinins and xenoreactive natural antibodies recognize distinct structures, they have a similar origin and function. Anti-A antibodies, anti-B antibodies, and xenoreactive natural antibodies were present in serum at similar concentrations and varied with age, gender, and the concentration of total IgM in serum in a similar manner. Anti-A antibodies, anti-B antibodies, and xenoreactive natural antibodies, unlike some elicited antibodies, had a high degree of thermal lability and bound more avidly at lower temperatures. The natural antibodies manifest remarkable homogeneity and high functional avidity for determinants on a cell surface but only a weak affinity for monovalent ligands. These findings suggest that anti-A antibodies, anti-B antibodies, and xenoreactive natural antibodies specific for Gal alpha 1-3Gal have a common origin and function and, given similar antigen density on target cells, provide similar humoral barriers to transplantation or transfusion and that these antibodies may be members of a common "family" of antibodies.

ABO Blood-Group System↗

PubMed1996

The influence of concordant xenografts on the humoral and cell-mediated immune responses to subsequent allografts in primates.

The humoral and cell-mediated immune responses to subsequent allografts were determined in primate recipients after concordant xenotransplantation as a bridge to allotransplantation. Heterotopic heart transplants (n = 4) were performed from cynomolgus monkeys into ABH type-matched olive baboons followed 2 weeks later by allotransplantation from ABH type-matched baboon donors. Allografts were explanted at 8 weeks. All recipients underwent splenectomy at the time of xenotransplantation and received immunosuppression with cyclosporine, azathioprine, and methylprednisolone. Concordant xenotransplantation in these primates did not induce humoral or cell-mediated immune responses that jeopardized subsequent allografts. The degree of xenospecific immune reactivity, as determined by specific cytotoxicity of recipient T-cell lines derived from the xenograft and extent of histologic xenograft rejection, did not predict the severity of subsequent allograft rejection. In two of the four recipients, xenotransplantation induced an alloreactive humoral response against antigens expressed by the B cells of more than 50% of members from a panel of 12 unrelated baboons. In all recipients, priming with xenogeneic splenocytes in vitro induced an accelerated proliferative T-cell response to allogeneic lymphocytes from 16% of this panel. This study affirms the role of concordant xenografts as appropriate biologic bridges to human allotransplantation. However, our results suggest that xenoreactive baboon memory CD4 T cells may recognize major histocompatibility complex class II--like structures shared between the xenogeneic and allogeneic targets. The potential allorecognition induced by a xenograft may affect the process of subsequent allograft donor selection.

Animals↗

PubMed1996

Prospects for xenotransplantation of the liver.

Because of the tremendous success of clinical transplantation of the liver and other organs in the last three decades, the demand for organs for transplantation has risen sharply and there is now a substantial shortage. The utilization of organs from other species, xenotransplantation, is increasingly viewed as a potential solution to this problem. The major limitation to xenotransplantation is the formidable immunological barriers that prevent the successful transplantation of non-human organs into human recipients. Here we review current knowledge about the immunology of xenotransplantation and the limited clinical and experimental experience in xenotransplantation of the liver.

Adaptation, Physiological↗

PubMed1996

Xenotransplantation: recent progress and current perspectives.

A severe shortage of human transplant donors has sparked interest in the use of animals as a source of organs and tissues for transplantation. Clinical application of xenotransplantation is limited in large part by the severe immunological reaction of the recipient against the graft. This immunological reaction is mediated initially by components of natural immunity such as xenoreactive antibodies, complement and natural killer cells and later by elicited humoral and cellular immune responses which act in concert to disrupt the function of the endothelial lining of blood vessels. The past few years have brought considerable progress in elucidating the molecular and cellular basis of xenograft rejection and in developing strategies to overcome xenograft rejection.

Acute Disease↗

PubMed1996

Specificity of xenoreactive anti-Gal alpha 1-3Gal IgM for alpha-galactosyl ligands.

The transplantation of organs from lower animals such as pigs into humans is prevented by a severe rejection reaction initiated by complement fixing xenoreactive natural antibodies. Most anti-pig xenoreactive natural antibodies in humans are thought to recognize Gal alpha 1-3Gal beta 1-4GlcNAc and are also thought to recognize, albeit less avidly, Gal alpha 1-6Glc. Gal alpha 1-6Glc has been used as a ligand for purification of 'anti-Gal alpha 1-3Gal antibodies' and as a therapeutic or reagent to prevent the binding of these antibodies to porcine organs or cells. We tested the specificity of anti-Gal alpha 1-3Gal IgM for Gal alpha 1-6Glc and related saccharides. Based on inhibition of binding of xenoreactive anti-Gal alpha 1-3Gal IgM to porcine cells by soluble saccharides, anti-Gal alpha 1-3Gal IgM in a human serum was found to consist of a mixture of antibodies which have a similar affinity for Gal alpha 1-3Gal but varying affinities for Gal alpha 1-6Glc and other structures. Twenty to 40% of the anti-Gal alpha 1-3Gal IgM from the population tested did not recognize Gal alpha 1-6Glc. The binding of anti-Gal alpha 1-3Gal IgM to Gal alpha 1-6Glc varied widely from individual to individual, some samples lacking almost entirely anti-Gal alpha 1-3Gal IgM which bound to Gal alpha 1-6Glc.

Animals↗

PubMed1996