Study on mitosis of dyserythropoietic erythroblasts using monoclonal antibodies

从先天性红细胞生成障碍性贫血患者的脐带血或静脉血的单核细胞培养人成红细胞。III型（CDA III）。按Wada等的方法进行缺氧培养。例如，由我们修改用荧光单克隆抗体法研究了两种参与细胞有丝分裂的多肽：α-微管蛋白和增殖细胞核抗原（PCNA）.α-微管蛋白：在CDA Ⅲ型成红细胞中，发现了大量的富含α-微管蛋白的细胞突起，它们呈长钉状或杆状，而在正常成红细胞中，这种细胞突起的数量是，这些细胞突起可能反映了我们在以前的研究中使用长时移电影显微术观察到的CDA Ⅲ型成红细胞的强烈的突起形成活动。其中一些过程可能是有丝分裂后细胞间桥的残余。在多核或节核CDA III型成红细胞中，α-微管蛋白未见明显异常.增殖细胞核抗原：流式细胞仪检测成红细胞中增殖细胞核抗原（PCNA）的含量，同时用荧光单克隆抗体和碘化丙啶检测DNA。正常红细胞PCNA表达量在S期逐渐增加，G_2期达高峰。CDA Ⅲ期成红细胞PCNA表达量在S期也有类似的增加。在少数DNA含量大于4C的CDA Ⅲ型成红细胞中，PCNA含量与DNA含量呈平行关系，其意义有待进一步研究。

Human eythroblasts were cultured from mononuclear cells from cord blood or venous blood of a patient with congenital dyserythropoietic anemia. type III (CDA III). They were cultured in hypoxic condition according to the method of Wada et. al., modified by us. Two polypeptides involved in cell mitosis : alpha-tubulin and proliferating cell nuclear antigen (PCNA), were studied by fluorescent mononuclear antibody method.1. alpha-tubulin : In CDA III erythroblasts, plentry of alpha-tubulin-rich cell prosesses, which were spike-shape or rodlike, were found, while in normal erythroblasts, number of such cell processes were These cell processes may be reflecting the vigorous process-forming activity of CDA III erythroblasts observed by us in the previous study using long-term time-lapse cinemicrography of them. Some of these processes might possibly be remnants of the post-mitotic intercellular bridges. In multinuclear or segment-nuclear CDA III erythroblasts, no apparent abnormalities of alpha-tubulin was observed.2. Proliferating cell nuclear antigen : The amount of PCNA in erythroblasts was measured simultaneously with DNA by FACS, using fluorescent mononuclear antibody and propidium iodide. The amount of PCNA of normal erythroblasts increased gradually during S-phase and reached maximum at G_2-phase. The amount of PCNA of CDA III erythroblasts showed similar increase during S-phase. In small number of CDA III erythroblasts with DNA more than 4C, the amount of PCNA tended to be large paralleling to their amount of DNA.Further studies will be needed to analyze the significance of this finding.