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Survey of glycoantigens in cells from α1‐3galactosyltransferase knockout pig using a lectin microarray
Author(s) -
Miyagawa Shuji,
Takeishi Shunsaku,
Yamamoto Aki,
Ikeda Kosuke,
Matsunari Hitomi,
Yamada Masao,
Okabe Masaru,
Miyoshi Eiji,
Fukuzawa Masahiro,
Nagashima Hiroshi
Publication year - 2010
Publication title -
xenotransplantation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.052
H-Index - 61
eISSN - 1399-3089
pISSN - 0908-665X
DOI - 10.1111/j.1399-3089.2009.00565.x
Subject(s) - griffonia simplicifolia , xenotransplantation , microbiology and biotechnology , lectin , biology , andrology , transplantation , medicine
Miyagawa S, Takeishi S, Yamamoto A, Ikeda K, Matsunari H, Yamada M, Okabe M, Miyoshi E, Fukuzawa M, Nagashima H. Survey of glycoantigens in cells from α1‐3galactosyltransferase knockout pig using a lectin microarray.
Xenotransplantation 2010; 17: 61–70. © 2010 John Wiley & Sons A/S. Abstract: Background: Glycoantigens represent major obstacles to successful xenotransplantation. Even after the α1‐3galactosyltransferase (GalT) gene knockout (GalT‐KO) pigs were produced, non‐Gal antigens continue to be present. This study reports on lectin blot analyses for endothelial cells (EC) and fibroblasts from GalT‐KO pigs. Methods: Differences in glycoantigens that are produced on cell surfaces in humans and pigs were surveyed. Differences between ECs and fibroblasts from wild‐type and GalT‐KO pigs were also examined. EC and fibroblasts from GalT‐KO pigs (heterozygous and homozygous) with N ‐acetylglucosaminyltransferase‐III (GnT‐III), a wild‐type EC from the sibling, human EC lines, HUVEC (human EC from umbilical veins), & HAOEC (human EC from aortas), and human fibroblast line were used. EC and fibroblasts were cultured in gelatin‐coated dishes for several days. After sonication and centrifugation, the supernatant protein from each cell was labeled with Cy3, applied to a lectin array and scanned with an SC Profiler, and analyzed using an Array Pro Analyzer. Results: The pig EC showed higher signals in Euonymus Europaeus (EEL) & Griffonia simplicifolia I‐B 4 (GSI‐B4), binds α‐Gal, and in Wisteria Floribunda (WFA), Helix pomatia (HPA), Glycine max (SBA), & Griffonia simplicifolia I‐A 4 (GSI‐A4), binds GalNAc including the Thomsen‐Friedenreich precursor (Tn)‐antigen, while the human EC showed strong signals in Ulex europaeus I (UEA‐I), Maackia amurensis (MAL), Erythrina cristagalli (ECA), & Trichosanthes japonica I (TJA‐I) instead. The EC from the GalT‐KO pig signals for EEL & GSI‐B4 disappeared and those for Bauhinia purpurea alba (BPL), HPA, SBA, & GSI‐A4 were greatly diminished as well, while it up‐regulated signals for Sambucus Nigra (SNA), Sambucus sieboldiana (SSA), & TJA‐I, bind α2‐6 sialic acid, compared to the wild‐type pig EC. Concerning fibroblasts, the signals for HPA, SBA, & GSI‐A4 were the most intense in the wild‐type, and the intensities for homozygous‐KO were less, approaching those of humans. In addition, the order of the intensities, as detected by Arachis hypogaea (PNA) & Maclura pomifera (MPA), binding Galβ1‐2GalNAc, indicates that the Thomsen‐Friedenreich (T)‐antigen is likely present on pig fibroblasts. Conclusion: It is possible that the T‐antigen and Tn‐antigen related to GalNAc are non‐Gal antigens, but, fortunately, not only α‐Gal but also GalNAc were found to be decreased in the KO‐pig.