Induction of apoptosis by sphingosine in human leukemic HL-60 cells: a possible endogenous modulator of apoptotic DNA fragmentation occurring during phorbol ester-induced differentiation.
Induction of apoptosis by sphingosine in human leukemic HL-60 cells: a possible endogenous modulator of apoptotic DNA fragmentation occurring during phorbol ester-induced differentiation.
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发表时间：
1995-02
期刊：
影响因子：
11.2
通讯作者：
Hideki Ohta;E. Sweeney;A. Masamune;Yutaka Yatomi;Sen-itiroh Hakomori;Yasuyuki Igarashi
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文献类型：
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作者：
Hideki Ohta;E. Sweeney;A. Masamune;Yutaka Yatomi;Sen-itiroh Hakomori;Yasuyuki Igarashi
本研究旨在探讨鞘氨醇在HL-60早幼粒白血病细胞凋亡调控中的作用。HL-60细胞暴露于鞘氨醇或其甲基化衍生物N，N-二甲基鞘氨醇6小时，引起核小体间DNA片段化和细胞凋亡特征性的定型形态学变化（即，细胞收缩、核浓缩和凋亡小体的形成），以及蛋白激酶C的药理学抑制剂如1-（5-异喹啉磺酰基）-2-甲基哌嗪和星形孢菌素。使用流式细胞术方法测量鞘氨醇和N，N-二甲基鞘氨醇引起的细胞凋亡。用鞘氨醇（10 μ M）和N，N-二甲基鞘氨醇（10 μ M）处理6小时的培养物中凋亡细胞的百分比分别为55.6 +/- 7.8%和84.2 +/-11.6%。通过用5 nM 4 β-佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA）处理诱导HL-60细胞向巨噬细胞分化。核小体间DNA片段，这是一个标志性的细胞凋亡，首次检测后10小时暴露于PMA和增加更长的治疗。在细胞分化过程中，细胞中鞘氨醇浓度随着凋亡细胞比例的增加而增加。PMA诱导HL-60细胞分化48 h后鞘氨醇含量增加约3.3倍。分化的HL-60细胞表现出显着增加的转化外源添加[3 H]神经酰胺[3 H]鞘氨醇，表明神经酰胺酶活性的升高。此外，暴露于鞘氨醇导致c-myc mRNA的下调。这些观察结果表明，鞘氨醇在PMA诱导的髓系白血病细胞分化过程中诱导凋亡DNA片段化的可能作用。鞘氨醇可能作为内源性调节剂介导凋亡信号。
The present studies were undertaken to characterize the potential role of sphingosine in the regulation of apoptosis in HL-60 promyelocytic leukemia cells. A 6-h exposure of HL-60 cells to sphingosine or its methylated derivative, N,N-dimethylsphingosine, caused internucleosomal DNA fragmentation and stereotypical morphological changes characteristic of apoptosis (i.e., cell shrinkage, nuclear condensation, and the formation of apoptotic bodies), as well as that to pharmacological inhibitors of protein kinase C such as 1-(5-isoquinolinesulfonyl)-2-methylpiperazine and staurosporine. Apoptosis by sphingosine and N,N-dimethylsphingosine was measured using a flow cytometric method. The percentages of apoptotic cells in cultures treated with sphingosine (10 microM) and N,N-dimethylsphingosine (10 microM) for 6 h were 55.6 +/- 7.8% and 84.2 +/- 11.6%, respectively. HL-60 cells were induced to differentiate toward macrophages by treatment with 5 nM 4 beta-phorbol 12-myristate 13-acetate (PMA). Internucleosomal DNA fragmentation, which was a hallmark of apoptosis, was first detected after 10-h exposure to PMA and increased with longer treatment. Sphingosine concentrations in the cells increased concomitantly with the increasing proportion of apoptotic cells during cell differentiation. Sphingosine level in HL-60 cells differentiated by treatment with PMA for 48 h was about 3.3-fold greater than that in untreated cells. Differentiated HL-60 cells exhibited a markedly increased conversion of exogenously added [3H]ceramide to [3H]sphingosine, indicating elevation of ceramidase activity. Moreover, exposure to sphingosine resulted in down-regulation of c-myc mRNA. These observations suggest the possible role of sphingosine in induction of apoptotic DNA fragmentation during PMA-induced differentiation in myeloid leukemia cells. Sphingosine may function as an endogenous modulator mediating the apoptotic signal.