In vitro study of immunocompatibility of humanized genetically modified pig erythrocytes with human serum
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摘要:
目的 探讨野生型（WT）、四基因修饰（TKO/hCD55）和六基因修饰（TKO/hCD55/hCD46/hTBM）猪红细胞与人血清的免疫相容性和免疫差异。 方法 收集20名不同血型志愿者的血液，将WT、TKO/hCD55和TKO/hCD55/hCD46/hTBM猪红细胞、ABO-相容（ABO-C）及ABO-不相容（ABO-I）人红细胞分别暴露于不同血型人血清中，检测血凝集、抗原抗体结合（IgG、IgM）水平和补体依赖细胞毒性，评估2种基因修饰猪红细胞与人血清的免疫相容性。 结果 ABO-C组未出现明显凝集；WT组和ABO-I组凝集水平高于TKO/hCD55组和TKO/hCD55/hCD46/hTBM组（均为P<0.001）。WT组猪红细胞裂解水平高于ABO-C组、TKO/hCD55组和TKO/hCD55/hCD46/hTBM组；ABO-I组猪红细胞裂解水平高于TKO/hCD55组、TKO/hCD55/hCD46/hTBM组（均为P<0.01）。TKO/hCD55组猪红细胞IgM和IgG结合水平均低于WT组和ABO-I组；TKO/hCD55/hCD46/hTBM组猪红细胞IgG和IgM结合水平低于WT组，IgG低于ABO-I组（均为P<0.05）。 结论 基因修饰猪红细胞的免疫相容性优于野生型猪，并接近于ABO-C，人源化猪红细胞在血资源奇缺时可考虑作为血液来源。 Abstract:Objective To investigate the differences and the immunocompatibility of wild-type (WT), four-gene modified (TKO/hCD55) and six-gene modified (TKO/hCD55/hCD46/hTBM) pig erythrocytes with human serum. Methods The blood samples were collected from 20 volunteers with different blood groups. WT, TKO/hCD55, TKO/hCD55/hCD46/hTBM pig erythrocytes, ABO-compatible (ABO-C) and ABO-incompatible (ABO-I) human erythrocytes were exposed to human serum of different blood groups, respectively. The blood agglutination and antigen-antibody binding levels (IgG, IgM) and complement-dependent cytotoxicity were detected. The immunocompatibility of two types of genetically modified pig erythrocytes with human serum was evaluated. Results No significant blood agglutination was observed in the ABO-C group. The blood agglutination levels in the WT and ABO-I groups were higher than those in the TKO/hCD55 and TKO/hCD55/hCD46/hTBM groups (all P<0.001). The level of erythrocyte lysis in the WT group was higher than those in the ABO-C, TKO/hCD55 and TKO/hCD55/hCD46/hTBM groups. The level of erythrocyte lysis in the ABO-I group was higher than those in the TKO/hCD55 and TKO/hCD55/hCD46/hTBM groups (both P<0.01). The pig erythrocyte binding level with IgM and IgG in the TKO/hCD55 group was lower than those in the WT and ABO-I groups. The pig erythrocyte binding level with IgG and IgM in the TKO/hCD55/hCD46/hTBM group was lower than that in the WT group and pig erythrocyte binding level with IgG was lower than that in the ABO-I group (all P<0.05). Conclusions The immunocompatibility of genetically modified pig erythrocytes is better than that of wild-type pigs and close to that of ABO-C pigs. Humanized pig erythrocytes may be considered as a blood source when blood sources are extremely scarce. -
图 1 血凝集试验
注：A图为血凝集试验图像；B图为应用改良Marsh评分计算凝集水平，与ABO-I组比较，aP<0.001，与WT组比较，bP<0.001。
Figure 1. Hemagglutination assay
图 2 补体依赖细胞毒性试验
注：与ABO-I组比较，aP<0.01；与WT组比较，bP<0.01。
Figure 2. Complement-dependent cytotoxicity assay
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