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F A Neethling

Publications and source records attributed to F A Neethling.

36 records · Page 2Linked to original sources
PubMed1994

Evidence that intravenously administered alpha-galactosyl carbohydrates reduce baboon serum cytotoxicity to pig kidney cells (PK15) and transplanted pig hearts.

Methods of inhibiting the hyperacute antibody-mediated rejection that occurs when pig organs are transplanted into primates have been investigated using the baboon as a potential recipient. Baboons were treated with different regimens that included combinations of (1) splenectomy, (2) pharmacologic immunosuppression (CsA, cyclophosphamide, corticosteroids +/- methotrexate), and (3) intravenous infusion of oligosaccharides. The cytotoxicity of the serum was then assessed on cultures of pig kidney cells (PK15). Unmodified serum caused approximate 65-100% pig cell destruction. Splenectomy and/or pharmacologic immunosuppression, and infusions of dextran, dextrose or mannitol, did not result in any reduction of cytotoxicity. Infusions of melibiose and/or arabinogalactan, both of which have terminal non-reducing alpha-galactose, however, decreased relative PK15 cell damage significantly in a dose-dependent manner. At high concentrations (< or = 50 g/hr), complete inhibition of cytotoxicity was achieved in 4 of 15 baboons. The extracorporeal immunoadsorption of baboon serum utilizing immunoaffinity columns of melibiose also resulted in a significant reduction (of approximately 80%) in cytotoxic effect. In 1 baboon, melibiose and arabinogalactan infusion delayed vascular rejection of a pig cardiac xenograft from 10 min to about 12 hr, at which time the baboon died from the toxic effects of the carbohydrate infusion. These observations (1) add further support to the role that anti-alpha-galactosyl antibodies play in the hyperacute rejection of pig tissues transplanted into primates, and (2) demonstrate that serum cytotoxicity can be reduced by the intravenous infusion of alpha-galactosyl oligosaccharides or by extracorporeal immunoadsorption using these carbohydrates.

Animals↗
PubMed1994

Protection of pig kidney (PK15) cells from the cytotoxic effect of anti-pig antibodies by alpha-galactosyl oligosaccharides.

Anti-pig antibodies in human and baboon serum are believed to be directed against alpha-galactosyl (alpha Gal) epitopes expressed on various pig cells, including vascular endothelia. We have investigated the effect of human sera on the PK15 pig kidney cell line, which abundantly expresses alpha Gal epitopes. To quantitate cell viability, we have used a staining method that differentiates live cells from dead ones. Various carbohydrates (n = 28) were added individually to serum at concentrations of 0.125-50 mg/ml. Unmodified serum caused approximate 100% PK15 cell death within 60 min. Carbohydrates that were not alpha Gal based did not significantly protect PK15 cells. Of the alpha Gal-based carbohydrates, only B disaccharide protected PK15 cells from both human and baboon serum (76% and 93% protection, respectively, at 1 mg/ml). Three alpha Gal oligosaccharides provided approximately 80-90% protection against both human and baboon sera at a concentration of 10 mg/ml. Three other closely related structures protected only against baboon serum (> 80%) at high concentration (50 mg/ml), suggesting a difference in anti-pig antibody affinity between baboon and man. Specific anti-alpha Gal antibody-depleted serum caused < 10% pig cell death, whereas the antibodies eluted from the alpha Gal columns caused > 70% pig cell death. In conclusion, this study provides further evidence that (1) alpha Gal structures are the targets for human and baboon anti-pig antibodies, and (2) there may be a therapeutic role for the infusion of specific alpha Gal carbohydrates, or for antibody removal using alpha Gal immunoaffinity columns, in order to prevent hyperacute rejection of pig organs in man.

Animals↗
PubMed1993

Binding and specificity of major immunoglobulin classes of preformed human anti-pig heart antibodies.

Preformed human anti-pig antibodies isolated from perfused pig hearts were used to analyze the binding of various immunoglobulin classes to cultured pig kidney cells. All anti-pig immunoglobulins (i.e., IgG, IgA, and IgM) were localized on the cell surface by the use of an indirect immunofluorescence technique. Anti-pig immunoglobulins also competed for the pig cell surface epitopes with Griffonia simplicifolia lectin (GS-I-B4), which is specific for alpha-galactosyl residues. This study provides further evidence that preformed human antibodies recognizing alpha-galactosyl-containing epitopes (anti-gal antibodies) could be an important factor in hyperacute rejection of pig organs.

Animals↗
PubMed1993

Identification of alpha-galactosyl and other carbohydrate epitopes that are bound by human anti-pig antibodies: relevance to discordant xenografting in man.

Human anti-pig antibodies were obtained by perfusing pig hearts (n = 4) and kidneys (n = 8) with human AB or O plasma followed by elution with 3 M NaSCN. The antibodies were screened against a panel of 132 synthetic carbohydrates conjugated to bovine serum albumin using an enzyme-linked immunoassay. An anti-immunoglobulin antibody was also used to detect immunoglobulin deposits on pig tissues. Four carbohydrate molecules with a terminal alpha-galactose residue bound all but one of the human anti-pig kidney antibodies and most of the anti-pig heart antibodies. These were: (i) alpha Gal(1-->3)beta Gal(1-->4)beta GlcNac (linear B type 2); (ii) alpha Gal(1-->3)beta Gal(1-->4)beta Glc (linear B type 6); (iii) alpha Gal(1-->3)beta Gal(B disaccharide); and (iv) alpha Gal(alpha-D-galactose). Immunoglobulin deposition was documented post-plasma perfusion in all pig hearts and particularly strongly in all pig kidneys. These results suggest that human anti-pig antibodies are mainly directed against alpha-galactosyl structures. Extracorporeal immunoadsorption of human plasma through columns of the specific synthetic carbohydrate(s) might lead to depletion of anti-pig antibodies and allow discordant xenografting in man. Alternatively, the infusion of the specific carbohydrate(s) for a period of several days might result in neutralization of the anti-pig antibodies and allow accommodation to take place.

ABO Blood-Group System↗
PubMed1993

Specific intravenous carbohydrate therapy. A new concept in inhibiting antibody-mediated rejection--experience with ABO-incompatible cardiac allografting in the baboon.

Heterotopic allografting of ABO-incompatible donor hearts in recipient baboons "hyperimmunized" against the incompatible A or B antigen (n = 3) was followed by hyperacute antibody-mediated vascular rejection within a mean of 19 min. The A and B epitopes against which these antibodies are directed are carbohydrates that can be synthesized. The continuous i.v. infusion of the specific synthetic A or B trisaccharide, beginning immediately pre-transplant and continued posttransplant for several days, prolonged allograft survival to a mean of 8 days (n = 2) and prevented antibody-mediated rejection, graft failure resulting from acute cellular rejection. The addition of triple pharmacologic immunosuppressive therapy (n = 4) resulted in prolongation of graft survival to a mean of > 28 days, with one heart still beating at 52 days; all grafts showed features of cellular rejection. "Accommodation" would appear to have developed in several baboons as graft function continued for periods of up to 39 days after discontinuation of carbohydrate therapy. Specific i.v. carbohydrate therapy should allow organ allografting to be performed across the ABO blood group barrier in humans. Furthermore, if the carbohydrate epitopes on the organs of discordant animals (e.g., the pig) against which human xenoreactive antibodies are directed can be confirmed, then this form of therapy might allow successful discordant organ xenotransplantation in man.

ABO Blood-Group System↗
PubMed1991

Immunological compatibility between the chacma baboon and man.

Predictions of an increasing shortage of donor organs for the future has led to a resurgence of interest in xenotransplantation. We have methodically assessed the immunological compatibility of humans against the chacma baboon with a view to narrowing the gap of concordance by careful immunological screening. The necessity of major blood group compatibility in xenotransplantation is now established. While no group O universal donor exists in the baboon, groups A (45%), B (15%), and AB (40%) are well represented. Baboon histocompatibility antigens could not be precisely defined using human antisera. This does not necessarily imply lack of homology between the species, as we have shown specific crossreactivity of numerous antihuman monoclonal antibodies with baboon leukocytes. Normal humans do not exhibit preformed agglutinins to erythrocytes of the chacma baboon (Papio ursinus orientalis)) but cytotoxic antibodies are occasionally found. Sera from allosensitized patients may contain crossreacting hemagglutinins, leukoagglutinins and complement-dependent cytotoxic antibodies. Binding of human immunoglobulin-G and -M to baboon targets was demonstrated by flow cytometry. Negative crossmatch combinations for antibodies of the IgG subclass were easily found, but IgM antibodies from allosensitized patients were polyspecific in their action. In vitro assessment of lymphocyte mediated cytotoxicity showed that preformed cellular immunity between the species was rare. The response of human lymphocytes to xenoantigen stimulation in mixed lymphocyte cultures showed a normal distribution, permitting the selection of low-responding combinations. Screening for viruses, especially HTLV-1 and Coxsackie-BL34, is important. These findings demonstrate a closer degree of concordance than has previously been suspected.

Animals↗