Gotoh Y, Takashima H, Noguchi K, Nishimura H, Tokushima M, Shirai T, Kimoto M
Department of Immunology, Saga Medical School, Japan.
J Immunol. 1993 Jun 1;150(11):4777-87.
We have tried to demonstrate the existence of a mixed haplotype MHC class II molecule in (NZB x NZW)F1 (B/WF1) mice. When a large panel of keyhole limpet hemocyanin-specific T cell clones derived from B/WF1 mice was analyzed, several clones were shown to be restricted by a F1-specific A beta Z/A alpha d class II molecule. Autoreactive A beta Z/A alpha d-specific T cell clones were also obtained. The ability of the association and expression of A beta Z with A alpha d was confirmed by hybridoma and transfection experiments. Hybridoma cell lines created by fusion of NZW (H-2z) spleen cells with M12.C3 (a A beta d- variant cell line derived from M12.4.1 (H-2d) B lymphoma) cells expressed A beta Z determinants. Transfection of A beta Z genomic DNA to M12.C3 cells resulted in the expression of A beta Z determinants. These hybridoma cell lines and transfectants were able to stimulate A beta Z/A alpha d-specific T cell clones, suggesting the expression of A beta Z/A alpha d molecules on the cell surface. However, attempts to demonstrate the existence of mixed haplotype MHC class II molecules in B/WF1 mice by two-dimensional (nonequilibrium pH gradient gel electrophoresis/SDS-PAGE) gel electrophoresis analysis with the use of anti-class II mAb failed to demonstrate the existence of mixed haplotype A beta Z/A alpha d or A beta d/A alpha z class II molecules in B/WF1 mice. Analysis of mixture of TA beta Z cell and B/WF1 spleen cell lysates immunoprecipitated by anti-A beta Z mAb suggested that the amount of haplotype mixed A beta Z/A alpha d molecules in B/WF1 spleen cells is less than 1/10 that of haplotype matched A beta/A alpha pairs. Our results suggest that, although undetectable by biochemical analysis, small amounts of mixed haplotype A beta Z/A alpha d molecules exist in B/WF1 spleen cells. Also, T cell clones which recognize them exists in B/WF1 mice. Because autoimmune symptoms of B/WF1 mice are shown to be related to heterozygosity at the H-2 region, autoreactive T cell clones which recognize the mixed haplotype A beta Z/A alpha d class II molecule might be involved for the induction of autoimmunity in B/WF1 mice.
我们试图证明在(新西兰黑鼠×新西兰白鼠)F1(B/WF1)小鼠中存在混合单倍型MHC II类分子。当分析一大组源自B/WF1小鼠的钥孔戚血蓝蛋白特异性T细胞克隆时,发现有几个克隆受F1特异性AβZ/Aαd II类分子的限制。还获得了自身反应性AβZ/Aαd特异性T细胞克隆。通过杂交瘤和转染实验证实了AβZ与Aαd的缔合及表达能力。用NZW(H-2z)脾细胞与M12.C3(源自M12.4.1(H-2d)B淋巴瘤的Aβd变异细胞系)细胞融合产生的杂交瘤细胞系表达AβZ决定簇。将AβZ基因组DNA转染到M12.C3细胞中导致AβZ决定簇的表达。这些杂交瘤细胞系和转染子能够刺激AβZ/Aαd特异性T细胞克隆,表明AβZ/Aαd分子在细胞表面表达。然而,尝试通过使用抗II类单克隆抗体的二维(非平衡pH梯度凝胶电泳/SDS-PAGE)凝胶电泳分析来证明B/WF1小鼠中存在混合单倍型MHC II类分子,但未能证明B/WF1小鼠中存在混合单倍型AβZ/Aαd或Aβd/Aαz II类分子。用抗AβZ单克隆抗体免疫沉淀的TAβZ细胞和B/WF1脾细胞裂解物混合物的分析表明,B/WF1脾细胞中混合单倍型AβZ/Aαd分子的量不到单倍型匹配的Aβ/Aα对的1/10。我们的结果表明,尽管通过生化分析无法检测到,但B/WF1脾细胞中存在少量混合单倍型AβZ/Aαd分子。此外,B/WF1小鼠中存在识别它们的T细胞克隆。由于B/WF1小鼠的自身免疫症状显示与H-2区域的杂合性有关,识别混合单倍型AβZ/Aαd II类分子的自身反应性T细胞克隆可能参与了B/WF1小鼠自身免疫的诱导。