α1,3-半乳糖基转移酶敲除猪心、肾糖脂的结构特征及其与人及狒狒抗体的反应性。
Structural characterization of alpha1,3-galactosyltransferase knockout pig heart and kidney glycolipids and their reactivity with human and baboon antibodies.
机构信息
Department of Surgery, Institute of Clinical Sciences, Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden.
出版信息
Xenotransplantation. 2010 Jan-Feb;17(1):48-60. doi: 10.1111/j.1399-3089.2009.00564.x.
BACKGROUND
alpha1,3-galactosyltranferase knockout (GalT-KO) pigs have been established to avoid hyperacute rejection in GalT-KO pig-to-human xenotransplantation. GalT-KO pig heart and kidney glycolipids were studied focusing on elimination of Gal-antigens and whether novel antigens would appear. Non-human primates are used as pre-clinical transplantation experimental models. Therefore, sera from baboons transplanted with GalT-KO hearts were compared with human serum regarding reactivity with pig glycolipids.
METHODS
Neutral and acidic glycolipids were isolated from GalT-KO and WT pig hearts and kidneys. Glycolipid immune reactivity was tested on TLC plates using human affinity-purified anti-Gal Ig, anti-blood group monoclonal antibodies, lectins, and human serum as well as baboon serum collected before and after GalT-KO pig heart transplantations. Selected glycolipid fractions, isolated by HPLC, were structurally characterized by mass spectrometry and proton NMR spectroscopy.
RESULTS
GalT-KO heart and kidney lacked alpha3Gal-terminated glycolipids completely. Levels of uncapped N-acetyllactosamine precursor compounds, blood group H type 2 core chain compounds, the P1 antigen and the x(2) antigen were increased. Human serum antibodies reacted with Gal-antigens and N-glycolylneuraminic acid (NeuGc) in WT organs of which only the NeuGc reactivity remained in the GalT-KO tissues. A clear difference in reactivity between baboon and human antibodies with pig glycolipids was found. This was most pronounced for acidic, not yet identified, compounds in GalT-KO organs which were less abundant or lacking in the corresponding WT tissues.
CONCLUSIONS
GalT-KO pig heart and kidney completely lacked Gal glycolipid antigens whilst glycolipids synthesized by competing pathways were increased. Baboon and human serum antibodies showed a different reactivity pattern to pig glycolipid antigens indicating that non-human primates have limitations as a human pre-clinical model for immune rejection studies.
背景
为避免α1,3-半乳糖基转移酶敲除（GalT-KO）猪异种移植中的超急性排斥反应，已建立了 GalT-KO 猪。研究了 GalT-KO 猪心脏和肾脏糖脂，重点是消除 Gal 抗原以及是否会出现新抗原。非人类灵长类动物被用作临床前移植实验模型。因此，比较了移植 GalT-KO 心脏的狒狒的血清与人血清对猪糖脂的反应性。
方法
从 GalT-KO 和 WT 猪心脏和肾脏中分离出中性和酸性糖脂。使用人亲和纯化的抗 Gal Ig、抗血型单克隆抗体、凝集素以及人血清和移植 GalT-KO 猪心脏前后采集的狒狒血清，在 TLC 平板上测试糖脂免疫反应性。通过质谱和质子 NMR 光谱学对通过 HPLC 分离的选定糖脂级分进行结构表征。
结果
GalT-KO 心脏和肾脏完全缺乏α3Gal 末端糖脂。未封闭的 N-乙酰乳糖胺前体化合物、血型 H 型 2 核心链化合物、P1 抗原和 x(2) 抗原的水平增加。人血清抗体与 WT 器官中的 Gal 抗原和 N-糖基神经氨酸（NeuGc）反应，而 GalT-KO 组织中仅保留了 NeuGc 反应性。发现狒狒和人抗体与猪糖脂之间的反应性存在明显差异。GalT-KO 器官中酸性、尚未鉴定的化合物最为明显，这些化合物在相应的 WT 组织中含量较少或不存在。
结论
GalT-KO 猪心脏和肾脏完全缺乏 Gal 糖脂抗原，而通过竞争途径合成的糖脂增加。狒狒和人血清抗体对猪糖脂抗原表现出不同的反应模式，表明非人类灵长类动物作为免疫排斥研究的人类临床前模型存在局限性。
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