Apoptotic cell death and radiosensitive phase of cell cycle: A flow cytometric study

Abstract

The relationship between apoptotic cell death and radiosensitive phases of the cell cycle was studied using flow cytofluorometry. Pregnant mice were exposed to a single dose (0.1 Gy) of whole-body X-irradiation on Embryonic day 13 (E 13). The forebrains of the embryos were obtained at different times, either immediately after irradiation (0 hr) or 6 hr after irradiation. After fixation and washing, cellular DNA was measured by flow cytometry following cell staining with Diamidino-2-Phenylindole (DAPI) and sulforhodamine 101. A normal pattern of DNA-histograms was observed in the mice immediately after irradiation (0 hr), and no cell death could be found at that time. However, 6 hr after irradiation, another peak consisted of apoptotic cells in which DNA content lower than that in G1-phase cells appeared in the histogram of cells. The percentages of cells with DNA content equivalent to G1(-), S(-), and G2(+)M(-)phase cells for the control and 0 hr groups were: 72:18:10, and 6 hr after irradiation were 60:15:15, respectively. The number of cells with fractional DNA content (apoptotic cells) was about 10 percent. Because there was a significant difference in G1-phase cells between the control and 6 hr groups, from 72 to 60 percent, it was more likely that cells in G1 were selectively killed by irradiation in the mouse forebrain of E 13.

流動細胞光度測定法を用いてアポトーシス細胞死と細胞周期の放射線感受性相の関係を研究した。妊娠マウスは胚13日(E13)に単回量(1.0Gy)の全身性X線照射に暴露された。胚の前脳を照射直後(0時間)と6時間後の異なる時期に採取した。固定および洗浄の後、細胞DNAはジアミジノ-2-フェニルインドール(DAPI)およびスルフォロダミン101による細胞染色後に流動細胞光度測定法で測定された。照射直後(0時間)はDNA-ヒストグラムの正常パターンが観察され、その時間には細胞死がなかった。しかし、照射6時間後にはアポトーシスから成る他のピークがあり、そのDNA含量は細胞ヒストグラム中に見られるG1相細胞より低かった。対照群および0時間群のG1(-)、S(-)、およびG(2+)M(-)相細胞に相当するDNA含量の細胞%は72:18:10、および照射6時間後の値は60:15:15であった。分別DNA含量(アポトーシス細胞)を有する細胞数は約10%であった。対照群と6時間群の間にはG1相細胞に著しい差、72%から60%、があるので、E13マウス前脳の放射線照射によってG1相にある細胞が選択的に死んだと思われる。