低温条件对肿瘤细胞长期存活的影响

Long term hypothermic tumor cell culture and its effect on cell morphology

目的 :摸索哺乳动物细胞低温长期生存条件及培养程序 ,为太空搭载肿瘤细胞及建立太空生物医学研究平台提供研究基础 ;了解低温情况下肿瘤细胞长期生存的细胞形态、细胞周期变化 ;初步探讨低温长期培养在肿瘤疫苗制备中的意义。方法 :3种肿瘤细胞接种于自行设计的低温细胞长期生存系统中 ,摸索出低温下细胞培养程序。细胞株存活 1 5d后复温单克隆化 ,观察细胞生存情况及形态变化。其中对小鼠黑色素瘤B1 6细胞进行了低温 2 5d生存实验。结果 :肿瘤细胞可在低温细胞长期生存系统中成功存活 1 5d ,细胞形态无明显变化 ,细胞复温后可生长增殖 ,单克隆化。B1 6细胞可在低温不更换培养基的情况下生存 2 5d ,细胞形态发生明显变化。低温培养可使细胞周期阻滞 ,G1期细胞增多。结论 :成功建立了哺乳动物细胞低温条件下的长期生存方法 ,初步了解低温长期培养条件下肿瘤细胞形态及细胞周期的变化。

Objective:To investigate the condition and process of a long term hypothermic mammalian cell culture,and to lay the base for carrying mammalian cells to outer space and establishing outer space biomedical research platform. To understand the changes of the growth,multiplication,morphology and cell cycle of tumor cells,and to investigate the further applicability of hypothermic tumor cell culture in tumor vaccine production. Methods:Three tumor cell lines were maintained in the special designed hypothermic cell culture facility for 15 days and then monocloned after inoculated at 37℃ for 8 hr. The physiological functional changes of the tumor cells were observed such as cell morphology,cell multiplication and cell cycle. Especially,B16 cells were managed to survive and then be monocloned after 25 days' growth in hypothermia.Results:Tumor cells were successfully maintained in the hypothermia cell culture facility for 15 days without obvious changes of cell morphology. The cells could normally grow,multiple and be monocloned after return to 37℃. However,significant morphological changes of B16 cell were detected after cells grown in hypothermia for 25 days. Comparing with the control group,cell cycle was blocked,so that the number of cells in G1 phase increased markedly.Conclusion:We successfully established the hypothermic cell culture facility,ensuring technically mammalian cells to be carried by space craft and further research on space biomedicine. We also gained primarily understanding of the morphological and cell cycle changes of tumor cells cultured in hypothermia for a long time,offering new research clues of tumor vaccine production and hypothermia biology.