Hirano T, Abe K, Gotoh M, Oka K
Department of Clinical Pharmacology, Tokyo College of Pharmacy, Japan.
Br J Cancer. 1995 Dec;72(6):1380-8. doi: 10.1038/bjc.1995.518.
Certain anti-cancer agents are known to induce apoptosis in human tumour cells. However, these agents are intrinsically cytotoxic against cells of normal tissue origin, including myelocytes and immunocytes. Here we show that a naturally occurring flavone of citrus origin, tangeretin (5,6,7,8,4'-pentamethoxyflavone), induces apoptosis in human promyelocytic leukaemia HL-60 cells, whereas the flavone showed no cytotoxicity against human peripheral blood mononuclear cells (PBMCs). The growth of HL-60 cells in vitro assessed by [3H]thymidine incorporation or tetrazolium crystal formation was strongly suppressed in the presence of tangeretin; the IC50 values range between 0.062 and 0.173 microM. Apoptosis of HL-60 cells, assessed by cell morphology and DNA fragmentation, was demonstrated in the presence of > 2.7 microM tangeretin. Flow cytometric analysis of tangeretin-treated HL-60 cells also demonstrated apoptotic cells with low DNA content and showed a decrease of G1 cells and a concomitant increase of S and/or G2/M cells. Apoptosis was evident after 24 h of incubation with tangeretin, and the tangeretin effect as assessed by DNA fragmentation or growth inhibition was significantly attenuated in the presence of Zn2+, which is known to inhibit Ca(2+)-dependent endonuclease activity. Ca2+ and Mg2+, in contrast, promoted the effect of tangeretin. Cycloheximide significantly decreased the tangeretin effect on HL-60 cell growth, suggesting that protein synthesis is required for flavonoid-induced apoptosis. Tangeretin showed no cytotoxicity against either HL-60 cells or mitogen-activated PBMCs even at high concentration (27 microM) as determined by a dye exclusion test. Moreover, the flavonoid was less effective on growth of human T-lymphocytic leukaemia MOLT-4 cells or on blastogenesis of PBMCs. These results suggest that tangeretin inhibits growth of HL-60 cells in vitro, partially through induction of apoptosis, without causing serious side-effects on immune cells.
已知某些抗癌药物可诱导人肿瘤细胞凋亡。然而，这些药物对包括骨髓细胞和免疫细胞在内的正常组织来源的细胞具有内在的细胞毒性。在此我们表明，一种天然存在的源自柑橘的黄酮类化合物，橘红素（5,6,7,8,4'-五甲氧基黄酮），可诱导人早幼粒细胞白血病HL-60细胞凋亡，而该黄酮类化合物对人外周血单个核细胞（PBMC）无细胞毒性。在存在橘红素的情况下，通过[3H]胸苷掺入或四氮唑晶体形成评估的HL-60细胞体外生长受到强烈抑制；IC50值在0.062至0.173微摩尔之间。在存在>2.7微摩尔橘红素的情况下，通过细胞形态和DNA片段化评估证明HL-60细胞发生凋亡。对经橘红素处理的HL-60细胞进行流式细胞术分析也显示出DNA含量低的凋亡细胞，并显示G1期细胞减少以及S期和/或G2/M期细胞相应增加。与橘红素孵育24小时后凋亡明显，并且在已知可抑制Ca(2+)依赖性核酸内切酶活性的Zn2+存在下，通过DNA片段化或生长抑制评估的橘红素作用明显减弱。相比之下，Ca2+和Mg2+促进了橘红素的作用。放线菌酮显著降低了橘红素对HL-60细胞生长的作用，表明黄酮类化合物诱导的凋亡需要蛋白质合成。通过染料排除试验确定，即使在高浓度（27微摩尔）下，橘红素对HL-60细胞或丝裂原激活的PBMC均无细胞毒性。此外，该黄酮类化合物对人T淋巴细胞白血病MOLT-4细胞的生长或PBMC的增殖作用较小。这些结果表明，橘红素在体外抑制HL-60细胞生长，部分是通过诱导凋亡，而不会对免疫细胞造成严重副作用。
