Brock W A, Swartzendruber D E, Grdina D J
Cancer Res. 1982 Dec;42(12):4999-5003.
Murine fibrosarcoma cells can be separated into subpopulations by centrifugation through 10 to 35% Renografin density gradients. Previous work has shown that the heavier cell populations are rich in chronically hypoxic cells. In this study, each subpopulation was characterized for thymidine incorporation, thymidine transport, thymidine triphosphate pool sizes, and thymidine triphosphate specific activities. The heavier cell populations have less accessibility to exogenous thymidine, and they have lower endogenous pools of thymidine triphosphate and synthesize lower levels of DNA than do the lighter cell populations. However, if the cells are removed from the tumors and labeled with [3H]thymidine in vitro, all subpopulations synthesize DNA at similar rates. Two-parameter flow cytometry using acridine orange staining following partial acid denaturation of chromatin identified a small quiescent population in the most dense cell fraction, but the small number of these cells cannot account for the results of the biochemical studies. It appears that the hypoxic cells in the fibrosarcoma tumors are noncycling or slowly cycling, are in all phases of the cell cycle, and recover their ability to synthesize DNA when cultured under in vitro conditions.
通过在10%至35%的泛影葡胺密度梯度中离心，可将小鼠纤维肉瘤细胞分离成亚群。先前的研究表明，较重的细胞群体富含慢性缺氧细胞。在本研究中，对每个亚群的胸苷掺入、胸苷转运、三磷酸胸苷池大小和三磷酸胸苷比活性进行了表征。较重的细胞群体对外源胸苷的摄取较少，其三磷酸胸苷的内源性池较小，且与较轻的细胞群体相比，其DNA合成水平较低。然而，如果将细胞从肿瘤中取出并在体外用[3H]胸苷标记，所有亚群合成DNA的速率相似。在染色质部分酸变性后使用吖啶橙染色的双参数流式细胞术在密度最高的细胞组分中鉴定出一小群静止细胞，但这些细胞数量太少，无法解释生化研究的结果。看来，纤维肉瘤肿瘤中的缺氧细胞处于非循环或缓慢循环状态，处于细胞周期的所有阶段，并且在体外培养条件下恢复其合成DNA的能力。
