Vidair C A, Doxsey S J, Dewey W C
Department of Radiation Oncology, University of California, San Francisco 94143-0806.
J Cell Physiol. 1993 Mar;154(3):443-55. doi: 10.1002/jcp.1041540302.
To identify the cellular target(s) responsible for thermal killing in the G1 phase of the cell cycle, synchronous cultures of Chinese hamster ovary cells (CHO) were heat shocked and studied for one cell cycle by time-lapse videomicroscopy and immunocytochemistry. At the first mitosis post-heating, the fraction of cells giving rise to multinucleated progeny approximately equaled the nonclonogenic fraction. In addition, the cells yielding multinucleated progeny were delayed in prophase-metaphase relative to the cells yielding two uninucleated progeny (clonogenic cells). To study the basis for the delay in prophase-metaphase and subsequent formation of multinucleated cells, cells in mitosis were examined by immunofluorescence for spindle abnormalities. Multipolar mitotic spindles and chromosome misalignment were induced by heat. All multiple spindle poles induced by heat stained for pericentriolar material (PCM), the microtubule nucleating material of centrosomes. Heated cells in mitosis also contained additional foci of PCM which were not associated with the spindle. Cells made thermotolerant by a nonlethal heat shock were resistant to both thermal killing and the induction of multiple foci of PCM. Quantitative analysis revealed a good correlation between the fraction of cells with multipolar spindles, the fraction with more than two foci of PCM, and the nonclonogenic fraction. These data indicate that heat-induced alterations to the PCM of centrosomes resulted in multipolar mitotic spindles, delay in prophase-metaphase, and formation of multinucleated cells which were nonclonogenic. These results identify the centrosome as a G1 target for cell killing.
为了确定在细胞周期G1期负责热杀伤的细胞靶点，对中国仓鼠卵巢细胞（CHO）的同步培养物进行热休克处理，并通过延时视频显微镜和免疫细胞化学对一个细胞周期进行研究。在加热后的第一次有丝分裂时，产生多核后代的细胞比例大致等于非克隆形成细胞的比例。此外，相对于产生两个单核后代的细胞（克隆形成细胞），产生多核后代的细胞在前期到中期出现延迟。为了研究前期到中期延迟以及随后多核细胞形成的基础，通过免疫荧光检查有丝分裂期的细胞是否存在纺锤体异常。热诱导产生多极有丝分裂纺锤体和染色体排列紊乱。热诱导产生的所有多个纺锤体极均对中心粒周围物质（PCM）染色，PCM是中心体的微管成核物质。有丝分裂期的受热细胞还含有与纺锤体无关的额外PCM聚集点。通过非致死性热休克产生耐热性的细胞对热杀伤和多个PCM聚集点的诱导均具有抗性。定量分析显示，具有多极纺锤体的细胞比例、具有两个以上PCM聚集点的细胞比例与非克隆形成细胞比例之间存在良好的相关性。这些数据表明，热诱导中心体PCM的改变导致多极有丝分裂纺锤体、前期到中期延迟以及非克隆形成的多核细胞的形成。这些结果确定中心体是细胞杀伤的G1靶点。
