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1.
Transplantation ; 79(7): 777-82, 2005 Apr 15.
Article in English | MEDLINE | ID: mdl-15818319

ABSTRACT

BACKGROUND: Nonhuman primates are potential permissive animals for studying the risk of in vivo infection with porcine endogenous retrovirus (PERV). Anti-alphaGal natural antibodies are considered one of the barriers for preventing PERV infection, and it has been postulated that reduction of these antibodies could increase the risk of this infection. The aim of this study was to investigate the role of GAS 914, which depletes anti-alphaGal antibodies, in the potential in vivo transfer of PERV after pig-to-baboon organ xenotransplantation. METHODS: Twenty-seven baboons underwent xenotransplantation with hDAF or hMCP/hDAF transgenic pig organs, including heterotopic heart (n = 14) and kidney (n = 13) transplants. All of them received GAS 914 along with different immunosuppression protocols. PERV sequences were investigated by reverse-transcriptase polymerase chain reaction and by polymerase chain reaction assays in samples obtained at autopsy. The presence of PERV-specific antibodies and/or pig xenomicrochimerism was also evaluated. RESULTS: PERV RNA was not detected in any baboon plasma sample. In addition, all plasma samples were negative for PERV antibodies. However, PERV DNA sequences were detected in peripheral blood mononuclear cells from 6 of 14 (43%) animals investigated. Porcine mitochondrial DNA was also found in all of these positive samples and in six of the eight (75%) samples with negative PERV DNA, indicating that the detection of PERV sequences was attributable to xenochimerism. PERV-positive cells as a result of xenochimerism were also found in eight of nine (89%) spleen and lymph node tissue samples tested. CONCLUSIONS: Sustained depletion of anti-alphaGal antibodies does not augment the risk of PERV infection in pig-to-baboon organ transplantation.


Subject(s)
Endogenous Retroviruses/physiology , Galactose/immunology , Papio/immunology , Retroviridae Infections/immunology , Retroviridae Infections/virology , Swine/virology , Transplantation, Heterologous , Trisaccharides/immunology , Animals , Antibodies/immunology , Cell Line , Chimera , Endogenous Retroviruses/genetics , Endogenous Retroviruses/isolation & purification , Heart Transplantation/immunology , Humans , Kidney Transplantation/immunology , Polymerase Chain Reaction , RNA, Messenger/analysis , RNA, Messenger/genetics , Retroviridae Infections/therapy , Species Specificity
2.
Cytometry A ; 62(1): 54-60, 2004 Nov.
Article in English | MEDLINE | ID: mdl-15455411

ABSTRACT

BACKGROUND: A flow cytometry complement-mediated cytotoxicity assay (FCCA) using fluorescein diacetate (FDA) and propidium iodide (PI) to measure antibody-dependent toxicity is useful to determine the success of xenotransplant organs. We evaluated the validity of different mathematical models as a measure of cytotoxicity in FCCA. METHODS: Sera from untreated baboons (n = 7) and from immunosuppressed animals (n = 5) undergoing different xenotransplantation protocols with pig organs were tested by endogenous FCCA and a similar assay also using exogenous complement, and the results were compared with those of a complement-dependent hemolytic assay to detect anti-pig antibodies (APHA). The influence of PI/FDA staining and the use of several mathematical models were analyzed. RESULTS: For both groups of animals, we observed high correlations between the endogenous and exogenous FCCA pathways and between calculations based on PI and FDA staining. Of the four mathematical models tested--the Von Krogh equation, two exponential models, and area under the curve--the Von Krogh equation was the most appropriate in terms of goodness of fit and concordance with APHA. CONCLUSIONS: FDA/PI FCCA is useful to measure endogenous and exogenous complement-mediated cytotoxicities, and it has advantages related to identification of potential new xenoantibodies. Although all four mathematical models produced acceptable solutions, the Von Krogh equation was the best option.


Subject(s)
Antibodies, Heterophile/immunology , Antibody-Dependent Cell Cytotoxicity , Complement System Proteins/immunology , Cytotoxicity Tests, Immunologic/methods , Flow Cytometry/methods , Transplantation, Heterologous/immunology , Animals , Cell Line , Fluoresceins/chemistry , Humans , Papio , Propidium/chemistry , Staining and Labeling , Swine , Transplantation, Heterologous/pathology
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