REQUIREMENT FOR NA+-DEPENDENT ASCORBIC-ACID TRANSPORT IN OSTEOBLAST FUNCTION

被引:46
作者
FRANCESCHI, RT
WILSON, JX
DIXON, SJ
机构
[1] UNIV MICHIGAN, SCH MED, DEPT BIOL CHEM, ANN ARBOR, MI 48109 USA
[2] UNIV WESTERN ONTARIO, FAC DENT, DIV ORAL BIOL, LONDON, ON N6A 5C1, CANADA
来源
AMERICAN JOURNAL OF PHYSIOLOGY-CELL PHYSIOLOGY | 1995年 / 268卷 / 06期
关键词
VITAMIN-C; SODIUM-ASCORBATE COTRANSPORT; BONE CELL DIFFERENTIATION;
D O I
10.1152/ajpcell.1995.268.6.C1430
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
Ascorbic acid is necessary for expression of the osteoblast phenotype. We examined whether Na+-dependent transport is required for MC3T3-E1 preosteoblast cells to respond to vitamin C and investigated the role of membrane transport in the intracellular accumulation and function of ascorbate. MC3T3-E1 cells were found to possess a saturable, stereoselective, Na+-dependent ascorbic acid transport activity that is sensitive to the transport inhibitors sulfinpyrazone, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, and phloretin. Transport activity showed no competition with glucose or 2-deoxyglucose and was not inhibited by cytochalasin B, indicating that it is distinct from known hexose transporters. On addition of 100 mu M ascorbic acid to the extracellular medium, intracellular concentrations of 10 mM were reached within 5-10 h and remained constant for up to 24 h. A good correlation was observed between intracellular ascorbic acid concentration and rate of hydroxyproline synthesis. Although ascorbic acid was transported preferentially compared with D-isoascorbic acid, both isomers had equivalent activity in stimulating hydroxyproline formation once they entered cells. Marked stereoselectivity for extracellular L-ascorbic acid relative to D-isoascorbic acid was also seen when alkaline phosphatase and total hydroxyproline were measured after 6 days in culture. Moreover, ascorbic acid transport inhibitors that prevented intracellular accumulation of vitamin blocked the synthesis of hydroxyproline. Thus Na+-dependent ascorbic acid transport is required for MC3T3-E1 cells to achieve the millimolar intracellular vitamin C concentrations necessary for maximal prolyl hydroxylase activity and expression of the osteoblast phenotype.
引用
收藏
页码:C1430 / C1439
页数:10
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