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Release of pig leukocytes during pig kidney perfusion and characterization of pig lymphocyte carbohydrate xenoantigens
Author(s) -
Magnusson Stefan,
Månsson JanEric,
Strokan Valeri,
Jussila Rainer,
Kobayashi Takaaki,
Rydberg Lennart,
Romano Egidio,
Breimer Michael E.
Publication year - 2003
Publication title -
xenotransplantation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.052
H-Index - 61
eISSN - 1399-3089
pISSN - 0908-665X
DOI - 10.1034/j.1399-3089.2003.02052.x
Subject(s) - xenotransplantation , antigen , microbiology and biotechnology , biology , miniature pig , glycolipid , miniature swine , immune system , lymphocyte , perfusion , transplantation , chemistry , andrology , biochemistry , immunology , medicine , genetics
The Galα1–3Gal (αGal) antigen is considered the main xenoantigen in the pig to human species combination but other porcine antigens have to be considered such as the swine lymphocyte antigen (SLA), the blood group A/O and the Hanganutziu–Deicher (H–D) antigens. The H–D antigens are N ‐glycolyl‐neuraminic acid (NeuGc) terminated gangliosides that are widely distributed in mammalian species but absent in humans. Upon exposure to a vascularized pig organ, the human recipient can be immunized by direct interaction with the pig tissue or/and by transfer of tissue/cells from the organ into the recipient. In the present work, we describe the release of cells from porcine kidneys upon perfusion and the expression of glycolipid based αGal, blood group A/O and H–D antigens in pig lymphocytes. Pig kidneys were flushed with 20 ml of NaCl or Lidocain containing 5000 U heparin, and thereafter perfused with 3000‐ml perfusion solution and the cells released were counted and examined microscopically. Neutral glycolipid and ganglioside fractions were extracted from purified pig lymphocytes. The extracted components were characterized by thin layer chromatography, degradation and mass spectrometry. The expression of αGal and H–D epitopes on cells released from pig kidneys and purified pig lymphocytes were studied by immune electron microscopy. A total amount of about 300 × 10 6 leukocytes, mainly lymphocytes were released in the perfusate from the kidneys, of which about 100 × 10 6 cells were eluated in the 600 to 2400 ml perfusate fraction. Immunelectron microscopical analysis with Griffonia simplicifolia isolectin B 4 showed staining of pig leukocytes and other cells, morphologically similar to endothelial cells, released in the perfusate. The purified porcine lymphocytes contained 930 μg neutral glycolipid (4.2 μg/mg cell protein) of which 95% was glycolipids with one to four sugar residues. Immunostaining of the neutral glycolipid fractions revealed αGal terminated compounds migrating in the five and 10 to 12 sugar regions and blood group A compounds in the six and eight sugar regions. Two major gangliosides NeuGc‐GM 3 and NeuGc‐GD 3 were found in the pig lymphocytes. In a patient extracorporeally xenoperfused with a pig kidney, an increased staining of both αGal terminated structures as well as the H–D reactive gangliosides were found in the post‐perfusion serum samples. In summary, leukocytes, mainly lymphocytes are released from pig kidneys during perfusion which may contribute to immunization of human xenograft recipients.