乳腺癌细胞休眠期至分裂间期染色质行为动力学特征分析

Analysis of dynamic characteristics of chromatin behaviors in breast cancer cell from dormancy phase to interphase

目的细胞周期失调会导致基因组不稳定、引起细胞恶性转化和癌变。本研究通过分析乳腺癌细胞休眠期(G0期)至分裂间期细胞染色质动态行为特征,为进一步研究癌细胞周期调控提供一定的实验参考。方法以小鼠乳腺癌4T1细胞为研究对象,利用DNA标记物和组蛋白相关荧光标记观察细胞染色质形态及其在细胞周期中的动态变化特征,通过激光共聚焦显微镜进行记录和分析。结果实验证实DAPI信号和GFP-H2B信号有比较好的共定位,两者均可以有效标记染色质。在细胞由G0期至分裂间期过程中,起始时可以观察到有大体积荧光标记斑块,在其中有多个着丝粒卫星DNA原位杂交斑点,而在1 h时荧光斑块逐步解离为小颗粒斑块甚至消失,到1 h 40 min时荧光标记斑块会再次出现,其体积显著小于观察起始,并规律性排布于核仁外周和核膜内侧。结论G0期乳腺癌细胞中染色质间存在聚集现象,这种聚集可能与着丝粒卫星DNA的作用相关。细胞在增值信号刺激下进入细胞周期时,染色质聚集体会逐步解聚并消失,在染色质完成复制并高度压缩形成染色体后,进一步分裂到两个子细胞。

Objective Dysregulation of the cell cycle can lead to genome instability,causing maligtin from G0 phase to the interphase of the breast cancercell linesinour study,it provides a certain experimental basis for further research on cancer cell cycle regulation.MethodsIn mouse breast cancer 4 T1 cellline,we applied DNA markers and histone-related fluorescent markers to reflect the chromatin morphology in the cell and its dynamic changes in the cell cycle,recording and analyzing experimental graphicsby laser confocal microscope.ResultsOur study showed that the DAPI signal and GFP-H2 B signal have better co-localization,and both can effectively label chromatin.During the period from the G0 phase to the interphase,a large volume of fluorescently labeled plaques was observed at the beginning,in which there were multiplecentromere satellite DNA in situ hybridization spots.The fluorescent plaques gradually dissociate into small plaques even disappeared but reappearedwith a smaller volume 1 h 40 mins later,whichwas regularly arranged on the periphery of the nucleolus and the inside of the nuclear membrane.ConclusionThere is aggregation between chromatin in G0 phase breast cancer cells,and it may be related to the function of centromeric satellite DNA.When entering the interphase under the stimulation of cell proliferation signal,the aggregate will gradually dissociate and disappear,after the chromatin is copied and highly compressed to form a chromosome,it further divides into two cells.