Study on mitosis of dyserythropoietic erythroblasts using monoclonal antibodies

从1例先天性红细胞生成性贫血患者的脐血或静脉血中分离培养人红细胞。类型III(CDA III)。按Wada ET方法在低氧条件下培养。Al.，由我们修改。采用荧光单核抗体方法对参与细胞有丝分裂的两种多肽：α-微管蛋白和增殖细胞核抗原进行了研究。α-微管蛋白：在CDA-III红细胞中，发现富含α-微管蛋白的细胞突起呈棘状或棒状，而在正常红细胞中，这些细胞突起的数量较多，这些细胞突起可能反映了我们在先前研究中用长期延时摄影观察到的CDA-III红细胞强烈的过程形成活性。其中一些过程可能是有丝分裂后细胞间桥的残留物。在多核或节段核CDA III红细胞中，未观察到明显的α-微管蛋白异常。增殖细胞核抗原：用荧光单核抗体和碘化丙啶，用流式细胞仪同时检测红细胞增殖细胞核抗原和DNA的含量。正常红细胞增殖细胞核抗原的数量在S期逐渐增加，在G2期达到最大。CDAⅢ期红细胞增殖细胞核抗原的数量在S期也有类似的增加。在少数DNA>4C的CDA III红细胞中，增殖细胞核抗原的数量与其DNA的数量趋于平行，这一发现的意义还有待进一步的研究。

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.