Leal M A, Aller P, Mas A, Calle C
Departamento de Bioquímica y Biología Molecular, Facultad de Medicina, Universidad Complutense, Madrid, Spain.
Exp Cell Res. 1995 Apr;217(2):189-94. doi: 10.1006/excr.1995.1078.
We have studied the effect of 10(-8) M 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] on insulin binding, insulin receptor gene expression, and isotype mRNA pattern in U-937 human promonocytic cells. Binding assays indicated that 1,25-(OH)2D3 increased total receptor number without altering receptor affinity, while the dissociation rate of the hormone bound to its receptor remained unchanged. RNA blot assays indicated that 1,25-(OH)2D3 increased the levels of the two major insulin receptor mRNAs (11 and 8.5 kb) present in these cells. Experiments with the RNA synthesis inhibitor actinomycin D suggested that 1,25-(OH)2D3 did not alter the stability of total insulin receptor mRNA. Experiments with the protein synthesis inhibitor cycloheximide indicated that the increase in the amounts of this mRNA occurs as a direct response to the action of the hormone. Finally, polymerase chain reaction assays revealed that the insulin receptor mRNA isotype lacking the exon 11 (A isotype) was the only one present in both untreated and 1,25-(OH)2D3-treated U-937 cells. This indicates that 1,25-(OH)2D3 does not alter the splicing of the primary insulin receptor transcript in this cell line.
我们研究了10(-8)M 1,25-二羟基维生素D3[1,25-(OH)2D3]对U-937人原单核细胞中胰岛素结合、胰岛素受体基因表达及同种型mRNA模式的影响。结合试验表明，1,25-(OH)2D3增加了总受体数量，而未改变受体亲和力，同时激素与其受体结合的解离速率保持不变。RNA印迹试验表明，1,25-(OH)2D3增加了这些细胞中存在的两种主要胰岛素受体mRNA（11kb和8.5kb）的水平。用RNA合成抑制剂放线菌素D进行的实验表明，1,25-(OH)2D3未改变总胰岛素受体mRNA的稳定性。用蛋白质合成抑制剂环己酰亚胺进行的实验表明，这种mRNA量的增加是对激素作用的直接反应。最后，聚合酶链反应试验显示，缺乏第11外显子的胰岛素受体mRNA同种型（A同种型）是未处理和经1,25-(OH)2D3处理的U-937细胞中唯一存在的同种型。这表明1,25-(OH)2D3不会改变该细胞系中胰岛素受体初级转录本的剪接。
