DNA永生化链和肿瘤干细胞关系的初步研究

Preliminary study on relationship between DNA immortal strands and tumor stem cells

目的:探讨DNA永生化链的分配方式与肿瘤干细胞之间的关系.方法:采用启动和促进两阶段化学诱癌方法在小鼠背部诱导皮肤肿瘤;用溴脱氧尿嘧啶核苷(BrdU)掺入法检测DNA永生化链在发育期、成年期小鼠皮肤成体干细胞及皮肤肿瘤干细胞中的滞留时间;图像分析DNA含量方法检测肿瘤细胞倍体变化.结果:诱癌4mo后,40只小鼠中有17只背部分别长出1～3个皮肤肿瘤.BrdU阳性细胞变化情况检测显示,24h后发育期、成年期标记的小鼠皮肤上皮细胞和肿瘤组织的细胞均为阳性,阳性部位为细胞核;60d后发育期标记的小鼠皮肤上皮中仍有少数细胞为阳性,它们主要存在于毛囊的隆突部位(bulge),少数位于皮肤基底层.标记后30d检测成年期标记的小鼠皮肤上皮细胞,结果均为阴性.标记后30d检测肿瘤组织发现仍有少数细胞BrdU阳性,这些肿瘤细胞主要分布在肿瘤实质组织和间质组织的交界部位;标记后60d检测,肿瘤组织中不再有BrdU阳性细胞.图像分析DNA含量方法发现上述17例小鼠肿瘤都是非整倍体核型.结论:小鼠皮肤成体干细胞存在DNA永生化链,其在有丝分裂过程中发生非随机分配,肿瘤干细胞中可能存在DNA永生化链,其分配方式趋向随机分配.

s AIM: To study the relationship between DNA immortal strands and tumor stem cells. METHODS: Initiation and promotion two-stage chemical carcinogenesis method was used to induce cutaneous tumors in the mouse back skin. DNA label retaining method and immunolabelling assay were used to chase the changes of BrdU positive cells in mouse skin and tumor tissues. DNA ploidy image analysis was performed to detect the karyotype of tumor cells. RESULTS: Four months after tumor induction, there were 1-3 tumors in the back of 17 mice. During the chase period, immunolabelling results showed that an extremely high percentage of keratinocytes and cutaneous tumor cells incorporated the BrdU at 24 h after final BrdU labeling. Some keratinocytes of mice labeled with BrdU during juvenile period were BrdU positive for at least 60 d after the final BrdU injection. Most of these cells were located within the bulge of hair follicle and some scattered in the basal layer of skin. In contrast, the keratinocytes labeled with BrdU during adult period were BrdU negative completely at 30 d after the final BrdU injection. BrdU positive cells were observed in tumor tissues for at least 30 d but not more than 60 d after the final BrdU injection. DNA ploidy image analysis showed that all the tumor cells were aneuploidy. CONCLUSION: We have confirmed both the existence of immortal DNA strands and the cosegregation of chromosomes that bear them in mouse epidermal stem cells. Our findings suggest that there may be immortal DNA strands which intend to distribute randomly in tumor stem cells.