Clausen O P, Elgjo K, Kirkhus B, Pedersen S, Bolund L
J Invest Dermatol. 1983 Dec;81(6):545-9. doi: 10.1111/1523-1747.ep12523189.
Epidermal basal cells from hairless mice were isolated after pulse labeling with tritiated DNA precursors and subjected to DNA flow cytometry combined with cell sorting. Cells were sorted from a window in the middle of the S phase, collected on glass slides, and subjected to autoradiography. Unlabeled cells in the middle of the S phase were found in normal mouse epidermis after optimal pulse labeling with tritiated thymidine [( 3H]dThd), in accordance with previous results. The proportion of unlabeled S phase cells was considerably increased among basal cells from mice treated with growth-inhibitory epidermal extracts. Reanalysis and re-sorting of cells previously sorted from mid S showed that unlabeled cells could not be accounted for by G1 contamination. Furthermore, labeling with precursors incorporated into DNA by "de novo" metabolic pathway [( 3H]Urd) did not reduce the proportion of unlabeled S phase cells, either when given alone or when given in combination with the precursor for DNA incorporated by the "salvage" pathway [( 3H]dThd). This strongly indicates that the unlabeled S phase cells do not synthesize DNA continuously, or are synthesizing DNA at a rate below the level of detection. A reduced proportion of unlabeled S phase cells was found in regenerating epidermis. This may be explained by a dilution effect caused by the 3-fold increase in the total number of cells within S phase at this condition. The observation that essentially all cells in mid S phase were labeled during 4 days of continuous labeling with [3H]dThd, indicates that cells in S phase that remain unlabeled after optimal pulse labeling are cycling, albeit slowly. Two-parameter sorting based on DNA and light scatter indicated that slowly cycling cells are larger than the average. These cells may represent a subpopulation of basal cells going through their last division cycle before differentiation.
用氚标记的DNA前体进行脉冲标记后，从无毛小鼠中分离出表皮基底细胞，并将其进行DNA流式细胞术结合细胞分选。从S期中期的一个窗口对细胞进行分选，收集到载玻片上，然后进行放射自显影。在用氚标记的胸腺嘧啶核苷[（3H）dThd]进行最佳脉冲标记后，在正常小鼠表皮中发现了S期中期未标记的细胞，这与之前的结果一致。在用生长抑制性表皮提取物处理的小鼠的基底细胞中，未标记的S期细胞的比例显著增加。对先前从S期中期分选的细胞进行重新分析和重新分选表明，未标记的细胞不能用G1期污染来解释。此外，无论是单独给予还是与通过“补救”途径掺入DNA的前体[（3H）dThd]联合给予，通过“从头”代谢途径掺入DNA的前体[（3H）尿苷]标记都不会降低未标记的S期细胞的比例。这有力地表明，未标记的S期细胞不是持续合成DNA，或者是以低于检测水平的速率合成DNA。在再生表皮中发现未标记的S期细胞的比例降低。这可能是由于在这种情况下S期细胞总数增加了3倍而导致的稀释效应。在用[3H]dThd连续标记4天期间，基本上所有S期中期的细胞都被标记，这一观察结果表明，在最佳脉冲标记后仍未标记的S期细胞正在循环，尽管速度很慢。基于DNA和光散射的双参数分选表明，缓慢循环的细胞比平均细胞大。这些细胞可能代表了在分化之前经历最后一个分裂周期的基底细胞亚群。
