Ways D K, Dodd R C, Bennett T E, Gray T K, Earp H S
Department of Medicine, School of Medicine, East Carolina University, Greenville, North Carolina 27834-4354.
Endocrinology. 1987 Nov;121(5):1654-61. doi: 10.1210/endo-121-5-1654.
In the U937 human monoblastoid cell line, 1,25-dihydroxyvitamin D3[1,25(OH)2D3] through a specific interaction with the 1,25(OH)2D3 receptor promotes differentiation toward a more mature phenotype. In addition to this direct effect, 1,25(OH)2D3 also potentiates differentiation in response to lymphokines and (Bu)2cAMP. We examined the effect of 1,25(OH)2D3 on phorbol ester-stimulated differentiation. Either preincubation with or simultaneous exposure to 1,25(OH)2D3 enhanced phorbol ester-stimulated differentiation. Over a 72-h period, the increase in phorbol ester responsiveness was dependent on the duration of 1,25(OH)2D3 exposure. Enhancement of phorbol ester-induced differentiation was observed with 1,25(OH)2D3 concentrations ranging from 0.1-10 nM. The 1,25(OH)2D3 vitamin D metabolite was more potent than the 24,25-dihydroxyvitamin D3 and 25-hydroxyvitamin D3 metabolites in potentiating phorbol ester-induced differentiation. Phorbol esters can exert at least a portion of their effects on cellular function by activating protein kinase C. Thus, one mechanism by which 1,25(OH)2D3 could amplify signal transduction leading to potentiation of phorbol ester-stimulated differentiation would be by enhancing phorbol ester-stimulated phosphorylation. To examine this possibility, we measured protein kinase C-dependent substrate phosphorylation in extracts derived from cells pretreated with 1,25(OH)2D3. In extracts derived from cells treated with 1,25(OH)2D3, the protein kinase C-dependent phosphorylation of endogenous U937 substrates stimulated by calcium, phosphatidyl serine, and diolein was increased compared to that observed in vehicle-treated cells. The conditions required for 1,25(OH)2D3 to increase protein kinase C-dependent phosphorylation of endogenous substrates (concentration, duration of exposure, and metabolite specificity) were similar to those required to enhance phorbol ester-stimulated differentiation. Possibly mediating this enhanced phosphorylation was an increase in protein kinase C activity observed in extracts derived from 1,25(OH)2D3-treated cells.
在U937人单核细胞系中，1,25 - 二羟基维生素D3[1,25(OH)2D3]通过与1,25(OH)2D3受体的特异性相互作用，促进细胞向更成熟的表型分化。除了这种直接作用外，1,25(OH)2D3还能增强细胞对淋巴因子和双丁酰环磷腺苷（(Bu)2cAMP）的分化反应。我们研究了1,25(OH)2D3对佛波酯刺激分化的影响。预先用1,25(OH)2D3孵育或同时暴露于1,25(OH)2D3均能增强佛波酯刺激的分化。在72小时的时间段内，佛波酯反应性的增加取决于1,25(OH)2D3暴露的持续时间。在1,25(OH)2D3浓度范围为0.1 - 10 nM时，观察到佛波酯诱导的分化增强。1,25(OH)2D3这种维生素D代谢物在增强佛波酯诱导的分化方面比24,25 - 二羟基维生素D3和25 - 羟基维生素D3代谢物更有效。佛波酯可通过激活蛋白激酶C对细胞功能发挥至少部分作用。因此，1,25(OH)2D3增强信号转导从而增强佛波酯刺激的分化的一种机制可能是通过增强佛波酯刺激的磷酸化作用。为了检验这种可能性，我们测量了用1,25(OH)2D3预处理的细胞提取物中蛋白激酶C依赖性底物的磷酸化情况。在用1,25(OH)2D3处理的细胞提取物中，与用溶剂处理的细胞相比，由钙、磷脂酰丝氨酸和二油精刺激的内源性U937底物的蛋白激酶C依赖性磷酸化增加。1,25(OH)2D3增加内源性底物蛋白激酶C依赖性磷酸化所需的条件（浓度、暴露持续时间和代谢物特异性）与增强佛波酯刺激的分化所需的条件相似。在用1,25(OH)2D3处理的细胞提取物中观察到的蛋白激酶C活性增加可能介导了这种增强的磷酸化作用。
