Vrana J A, Rao A S, Wang Z, Jarvis W D, Grant S
Department of Medicine, Medical College of Virginia, Richmond VA, 23298, USA.
Int J Oncol. 1998 Apr;12(4):927-34. doi: 10.3892/ijo.12.4.927.
The goal of the present study was to determine whether partial restoration of the differentiation-inducing capacity of the PKC activator bryostatin 1 by the calcium ionophore A23187 is accompanied by enhancement of apoptosis in ara-C-pretreated human leukemia cells. When HL-60 cells were exposed to ara-C (10 or 100 microM;6 h) followed by bryostatin 1 alone (10 nM; 24 h), no increase in apoptosis was noted. In contrast, subsequent exposure of ara-C-pretreated cells to A23187 (250 nM; 24 h) increased apoptosis by approximately 100%. When ara-C-pretreated cells were incubated with A23187 and bryostatin 1, no further potentiation of cell death (compared to cells exposed to A23187 alone) was observed. Nevertheless, the combination of bryostatin 1 and A23187 substantially increased inhibition of clonogenicity in cells preincubated with ara-C (e.g., by > or = 2 logs). This effect was associated with morphological and functional evidence (i.e., plastic adherence) of enhanced leukemic cell maturation. The differentiating capacity of the combination of bryostatin 1 and A23187 was significantly weaker than that of the phorbol diester, PMA (10 nM), and unaccompanied (at 24 h) by induction of the cyclin-dependent kinase inhibitors (CDKIs) p21WAF1/CIP1 and p27KIP1. However, the extent of apoptosis was comparable in cells exposed to ara-C followed by PMA or bryostatin 1 + A23187, suggesting that differentiation per se is not solely responsible for enhancement of cell death in ara-C-pretreated cells. Coadministration of bryostatin 1 and the organotellurium compound AS101, which mimics the actions of A23187 in some systems, after ara-C also led to enhanced antiproliferative effects which were unaccompanied by an increase in apoptosis. Finally, exposure of cells to ara-C followed by other differentiation-inducing agents, including dimethylsulfoxide and sodium butyrate also resulted in increases in cell death in this cell line. These findings indicate that the inability of bryostatin 1 to potentiate apoptosis in ara-C-pretreated HL-60 cells may involve factors other than an inadequate differentiation stimulus. They also suggest that loss of leukemic self-renewal capacity following exposure to cytotoxic and differentiation-inducing agents may involve mechanisms other than, or in addition to, potentiation of apoptosis, particularly cellular maturation.
本研究的目的是确定钙离子载体A23187对蛋白激酶C（PKC）激活剂苔藓抑素1的诱导分化能力的部分恢复，是否会伴随阿糖胞苷预处理的人白血病细胞凋亡的增强。当HL-60细胞暴露于阿糖胞苷（10或100 microM；6小时），随后单独给予苔藓抑素1（10 nM；24小时）时，未观察到凋亡增加。相反，将阿糖胞苷预处理的细胞随后暴露于A23187（250 nM；24小时）可使凋亡增加约100%。当阿糖胞苷预处理的细胞与A23187和苔藓抑素1一起孵育时，未观察到细胞死亡的进一步增强（与单独暴露于A23187的细胞相比）。然而，苔藓抑素1和A23187的组合显著增强了对用阿糖胞苷预孵育的细胞克隆形成能力的抑制（例如，抑制程度≥2个对数）。这种效应与白血病细胞成熟增强的形态学和功能证据（即贴壁）相关。苔藓抑素1和A23187组合的诱导分化能力明显弱于佛波酯PMA（10 nM），并且在24小时时不伴有细胞周期蛋白依赖性激酶抑制剂（CDKIs）p21WAF1/CIP1和p27KIP1的诱导。然而，暴露于阿糖胞苷后再给予PMA或苔藓抑素1 + A23187的细胞中凋亡程度相当，这表明分化本身并非阿糖胞苷预处理细胞中细胞死亡增强的唯一原因。在阿糖胞苷之后同时给予苔藓抑素1和有机碲化合物AS101（在某些系统中其作用类似于A23187）也导致抗增殖作用增强，但未伴随凋亡增加。最后，将细胞暴露于阿糖胞苷后再给予其他诱导分化剂，包括二甲基亚砜和丁酸钠，也导致该细胞系中细胞死亡增加。这些发现表明，苔藓抑素1无法增强阿糖胞苷预处理的HL-60细胞的凋亡可能涉及除分化刺激不足之外的其他因素。它们还表明，暴露于细胞毒性和诱导分化剂后白血病自我更新能力的丧失可能涉及除凋亡增强之外的其他机制，或除凋亡增强之外还涉及细胞成熟等机制。
