Klarmann B, Wixler V, Lorenz R, Hempel K
Institut für Medizinische Strählenkunde und Zellforschung, Universität Würzburg, Germany.
Int J Radiat Biol. 1995 Apr;67(4):421-30. doi: 10.1080/09553009514550481.
The frequency of H-2Kk and HPRT-deficient T cells was measured in the H-2Kb, kDd,k genotype mouse 8-10 weeks after X-ray exposure at doses up to 6 Gy to compare the mutant frequency (MF) of an autosomal gene with that of an X-chromosomal gene. H-2K mutants were enriched by magnetic cell separation (MACS) using the H-2Kk-specific monoclonal antibody H100.5/28 and were isolated by limiting dilution cloning. Finally, the mutant phenotype was verified by flow cytometric analysis in a representative number of clones. The frequency of HPRT-deficient T cells rises from 2.5 x 10(-6) at 0 Gy to a maximum of 1.13 x 10(-4) at 4 Gy, and decreases to 2.9 x 10(-5) at 6 Gy. The H-2K- MF in the non-irradiated mouse was 8.4 x 10(-7). It increases with dose to a maximum of 8.1 x 10(-6) at 4 Gy and declines to 3.3 x 10(-6) at 6 Gy. The H-2K- MF measured depends on the monoclonal antibody used for the isolation of mutants. In a pilot study with another H-2Kk-specific monoclonal antibody (11.4.1), the spontaneous MF was four times higher than in experiments with the H100.5/28 monoclonal antibody. The expression of other class 1 antigens was investigated in H-2K- clones. The H-2Dd antigen had also disappeared in six of 41 clones from irradiated animals. This gene is situated at a distance of 1500 kb from the K-locus. The H-2Kb antigen was present in every investigated clone. In the discussion a model is presented that explains the shape of the dose-response curve of MF by selection against mutants in vivo systems under homeostasis. The results of the present investigation indicate that observed X-ray mutagenicity depends on many factors and that several genes have to be explored before reliable risk estimates are possible.
在对H-2Kb、kDd、k基因型小鼠进行高达6 Gy的X射线照射8至10周后，测定H-2Kk和次黄嘌呤磷酸核糖转移酶（HPRT）缺陷型T细胞的频率，以比较常染色体基因与X染色体基因的突变频率（MF）。使用H-2Kk特异性单克隆抗体H100.5/28通过磁性细胞分选（MACS）富集H-2K突变体，并通过有限稀释克隆进行分离。最后，通过流式细胞术分析在代表性数量的克隆中验证突变体表型。HPRT缺陷型T细胞的频率从0 Gy时的2.5×10⁻⁶上升至4 Gy时的最高值1.13×10⁻⁴，并在6 Gy时降至2.9×10⁻⁵。未照射小鼠中的H-2K⁻ MF为8.4×10⁻⁷。它随剂量增加，在4 Gy时达到最高值8.1×10⁻⁶，并在6 Gy时降至3.3×10⁻⁶。所测得的H-2K⁻ MF取决于用于分离突变体的单克隆抗体。在一项使用另一种H-2Kk特异性单克隆抗体（11.4.1）的初步研究中，自发MF比使用H100.5/28单克隆抗体的实验高四倍。在H-2K⁻克隆中研究了其他I类抗原 的表达。在来自受照射动物的41个克隆中的6个中，H-2Dd抗原也消失了。该基因位于距K基因座1500 kb处。每个研究的克隆中都存在H-2Kb抗原。在讨论中提出了一个模型，该模型通过在稳态下的体内系统中针对突变体进行选择来解释MF剂量反应曲线的形状。本研究结果表明，观察到的X射线致突变性取决于许多因素，并且在进行可靠的风险评估之前必须探索多个基因。
