期刊
JOURNAL OF CELLULAR BIOCHEMISTRY
卷 84, 期 2, 页码 389-400出版社
WILEY
DOI: 10.1002/jcb.10027
关键词
alkaline phosphatase; RBE4 cells; insulin; receptor mediated endocytosis; PI-PLC; blood brain barrier
The physiological function of alkaline phosphatase (ALP) remains controversial. It was recently suggested that this membrane-bound enzyme has a role in the modulation of transmembranar transport systems into hepatocytes and Caco-2 cells. ALP activity expressed on the apical surface of blood-brain barrier cells, and its relationship with I-125-insulin internalization were investigated under physiological conditions using p-nitrophenylphosphate (p-NPP) as substrate. For this, an immortalized cell line of rat capillary cerebral endothelial cells (RBE4 cells) was used. ALP activity and I-125-insulin internalization were evaluated in these cells. The results showed that RBE4 cells expressed ALP, characterized by an ecto-oriented active site which was functional at physiological pH. Orthovanaclate (100 muM), an inhibitor of phosphatase activities, decreased both RBE4-ALP activity and I-125-insulin internalization. in the presence Of L-arginine (1 mM) or adenosine (100 muM) RBE4-ALP activity and I-125-insulin, internalization were significantly reduced. However, D-arginine (1 mM) had no significant effect. Additionally, RBE4-ALP activity and I-125-insulin internalization significantly increased in the presence of the bioflavonoid kaempferol (100 muM), of the phorbol ester PMA (80 nM), IBMX (1 mM) progesterone (200 and 100 muM), beta -estradiol (100 muM), iron (100 muM), or in the presence of all-trans retinoic acid (RA) (10 muM). The ALP inhibitor levamisole (500 muM) was able to reduce I-125-insulin internalization to 69.1 +/- 7.1% of control. Our data showed a positive correlation between ecto-ALP activity and I-125-insulin incorporation (r=0.82; P <0.0001) in cultured rat brain endothelial cells, suggesting that insulin entry into the blood-brain barrier may be modulated through ALP. (C) 2001 Wiley-Liss, Inc.
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