氨氯吡咪诱导卵巢癌细胞凋亡

Apoptosis of ovarian carcinoma cell line induced by amiloride

目的观察利尿药氨氯吡咪（ａｍｉｌｏｒｉｄｅ）诱导卵巢癌细胞凋亡及其可能的机理。方法用光镜及荧光显微镜从形态上了解凋亡的发生，用琼脂糖凝胶电泳进一步检测发生凋亡的特征性ＤＮＡ降解。流式细胞术定量分析用不同剂量氨氯吡咪处理的卵巢癌细胞诱导的凋亡发生率，同时检测被处理细胞细胞内ｐＨ。结果氨氯吡咪处理的卵巢癌细胞在形态学上显示出凋亡细胞的特征，琼脂糖凝胶电泳显示特征性ＤＮＡ阶梯图。流式细胞术检测表明，０．０１～５μｍｏｌ／Ｌ氨氯吡咪能诱导１８．７％～６１．１％卵巢癌细胞发生凋亡，凋亡发生随用药剂量及药物作用时间增加而增多。氨氯吡咪处理的卵巢癌细胞细胞内ｐＨ值降低，并出现一群酸化细胞，经分选而分离出的酸化细胞逐渐表现出染色质的浓缩、致密及ＤＮＡ降解等凋亡特征。这群酸化细胞与凋亡细胞有很好的相关性（ｒ＝０．９６）。结论氨氯吡咪能诱导卵巢癌细胞凋亡，可能与其降低细胞内ｐＨ有关。

Objective To investigate induction of apoptosis and its possible mechanism in amiloride treated ovarian carcinoma cell line. Methods Morphological changes of apoptotic cells were investigated by light and fluorescence microsopy. DNA fragmentation was analysed by agarose gel electrophoresis. In addition, the number of hypodiploid cells (apoptotic cells) was quantitatively assessed by flow cytometry and intracellular pH was also analysed. Results Amiloride treated ovarian carcinoma cells showed morphological characateristics of apoptosis. A ladder like pattern of DNA fragmentation was demonstrated on agarose gel electrophoresis. Amiloride at 0.01～5 μmol/L could induce apoptosis in 18.7%～61.6% of ovarian carcinoma cells. The apoptosis inducing effect of amiloride was dose and time dependent. In addition, amiloride induced intracellular acidification in a subpopulation of the treated cells. Furthermore, these isolated acidified cells revealed chromatin condensation as well as DNA degradation with characteristics of apoptosis. There was good correlation between apoptotic cells and acidic cells. Conclusion Amiloride triggers apoptosis of ovarian carcinoma cells; intracellular acidification may be involved in the mechanism of apoptosis.