Human DAF on pig cells protects against human and non-human primate sera cytotoxicity mediated by exogenous or endogenous complement, as determined by flow cytometry
Human DAF on pig cells protects against human and non-human primate sera cytotoxicity mediated by exogenous or endogenous complement, as determined by flow cytometry
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DOI：
10.1016/j.trim.2006.03.008
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发表时间：
2006-08-01
影响因子：
1.5
通讯作者：
Domenech, Nieves
中科院分区：
文献类型：
--
作者：
Diaz-Roman, Tomas M.;Manez, Rafael;Domenech, Nieves
提出通过转基因在猪细胞中表达人类补体调节蛋白（CRP），以防止补体激活以及猪灵长类移植后随之而来的猪组织和器官的排斥。将人类腐烂加速因子（hDAF）转基因猪的器官移植到非人灵长类动物中不会发生超急性排斥反应，但 hDAF 不能预防不道德异种移植排斥（AHXR）。猪细胞中人类补体调节蛋白的表达缺乏预防 AHXR 的功效的一个可能解释可能是人类和非人类补体调节系统之间的种间差异。我们检测了猪细胞上表达的转基因 hDAF 抑制灵长类动物血清体外补体活性的功效。使用流式细胞术补体介导的细胞毒性测定（FCCA）针对来自 hDAF 和非转基因猪的外周血淋巴细胞（PBL），使用内源和外源补体对来自七只未经处理的狒狒以及正常人和狒狒血清库的个体细胞毒性进行了测定。我们还通过 ELISA 分析了狒狒血清的抗 Gala 1-3 Gal (α Gal) 抗体滴度，并通过免疫荧光分析了 PBL 表面 hDAF 的表达。转基因 hDAF 表达能够保护猪细胞免受狒狒和人血清造成的损伤。 hDAF 的细胞表达降低了内源性和外源性补体介导的细胞毒性，尽管前者略高。体液细胞毒性与特定抗体无关，但与 hDAF 表达呈负相关。 hDAF 的存在可以比 NBS 更有效地保护猪细胞免受 NHS 裂解。这些结果在体外证实了 hDAF 在猪细胞中对异源补体介导的损伤的保护作用，但它们也表明，如果细胞表达低水平的 hDAF，则 hDAF 的保护程度会降低。 (C) 2006 Elsevier B.V. 保留所有权利。
Expression of human complement regulatory proteins (CRP) in pig cells through transgenesis was proposed to prevent complement activation and the ensuing rejection of pig tissues and organs following pig-to-primate transplantation. Transplantation in non-human primates of organs from transgenic pigs for human decay accelerating factor (hDAF) did not undergo hyperacute rejection, but hDAF could not prevent Immoral xenograft rejection (AHXR). A possible explanation for the lack of efficacy of the expression of human complement regulatory proteins in pig cells to prevent AHXR may be interspecies differences between human and non-human complement regulatory system. We assayed the efficacy of transgenic hDAF expressed on porcine cells to inhibit the in vitro complement activity of primate sera. The individual cytotoxicity of sera from seven untreated baboons and of pools of normal human and baboon sera was assayed with endogenous and exogenous complement using a flow-cytometry complement-mediated cytotoxicity assay (FCCA) against peripheral blood lymphocytes (PBL) from hDAF and non-transgenic pigs. We also analyzed the anti-Gala 1-3 Gal (alpha Gal) antibody titre of the baboon sera by ELISA and the expression of hDAF on the PBL surface by immunofluorescence. Transgenic hDAF expression was capable of protecting pig cells against injury produced by both baboon and human serum. Cellular expression of hDAF reduced cytotoxicity mediated by endogenous and exogenous complement, although the former was slightly higher. Humoral cytotoxicity was not related to a particular antibody but was inversely related to hDAF expression. The presence of hDAF protected pig cells against lysis by NHS more effectively than against NBS. These results confirm in vitro the protective role of hDAF in pig cells to heterologous complement mediated damage, but they also suggest that the extent of hDAF protection decreases, however, if cells express low levels of hDAF. (C) 2006 Elsevier B.V. All rights reserved.