Konishi H, Steinbach G, Terry N H, Lee J J, Dubin J A, Glober G A, Fujita K, Spaulding D, Cass L, Hittelman W N
Department of Gastrointestinal Medical Oncology and Digestive Diseases, University of Texas M.D. Anderson Cancer Center, Houston 77030, USA.
Cancer Res. 1996 Feb 1;56(3):434-7.
Measurements of cell cycle phase fractions, particularly S-phase, are useful for studies of cell biology and carcinogenesis. Up-regulation of histone gene expression is tightly coupled to the G1-S-phase transition of the cell cycle, and mRNA levels rise 30-100-fold during S-phase. Labeling of histone H3 mRNA using in situ hybridization (ISH) was assessed as a measure of S-phase cells and compared with that found using in vivo 5-bromodeoxyuridine (BrdUrd) labeling in formalin-fixed rat colonic crypts under baseline, modified 72-h starvation, and 24-h refeeding conditions. The labeling index scored in single-labeled sections by histone H3 ISH tightly correlated with that found by in vivo BrdUrd labeling (r = 0.99, p < 0.0001) and clearly discriminated between the control, starved, and refed states (P < 0.001). In 180 crypt sections double labeled using histone H3 ISH and BrdUrd, 92% of 1572 labeled cells exhibited both nuclear BrdUrd and cytoplasmic histone H3 label. It is concluded that histone H3 ISH is an accurate measure of the S-phase fraction and provides an alternative to in vivo BrdUrd labeling in rat colon. This finding warrants validation in human studies.
细胞周期阶段分数的测量，尤其是S期分数，对于细胞生物学和癌症发生的研究很有用。组蛋白基因表达的上调与细胞周期的G1-S期转变紧密相关，并且在S期mRNA水平会升高30-100倍。使用原位杂交（ISH）对组蛋白H3 mRNA进行标记，以此作为S期细胞的一种测量方法，并与在基线、改良的72小时饥饿和24小时再喂养条件下，在福尔马林固定的大鼠结肠隐窝中使用体内5-溴脱氧尿苷（BrdUrd）标记所得到的结果进行比较。通过组蛋白H3 ISH在单标记切片中评分的标记指数与通过体内BrdUrd标记所得到的结果紧密相关（r = 0.99，p < 0.0001），并且能够清楚地区分对照、饥饿和再喂养状态（P < 0.001）。在使用组蛋白H3 ISH和BrdUrd进行双重标记的180个隐窝切片中，1572个标记细胞中有92%同时呈现出核BrdUrd和细胞质组蛋白H3标记。得出的结论是，组蛋白H3 ISH是S期分数的一种准确测量方法，并且为大鼠结肠中的体内BrdUrd标记提供了一种替代方法。这一发现有待在人体研究中得到验证。
