Modulation of uptake of organic cationic drugs in cultured human colon adenocarcinoma Caco-2 cells by an ecto-alkaline phosphatase activity

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Abstract
Alkaline phosphatase (ALP) refers to a group of nonspecific phosphomonoesterases located primarily in cell plasma membrane. It has been described in different cell lines that ecto‐ALP is directly or indirectly involved in the modulation of organic cation transport. We aimed to investigate, in Caco‐2 cells, a putative modulation of 1‐methyl‐4‐phenylpyridinium (MPP+) apical uptake by an ecto‐ALP activity. Ecto‐ALP activity and 3H‐MPP+ uptake were evaluated in intact Caco‐2 cells (human colon adenocarcinoma cell line), in the absence and presence of a series of drugs. The activity of membrane‐bound ecto‐ALP expressed on the apical surface of Caco‐2 cells was studied at physiological pH using p‐nitrophenylphosphate as substrate. The results showed that Caco‐2 cells express ALP activity, characterized by an ecto‐oriented active site functional at physiological pH. Genistein (250 µM), 3‐isobutyl‐1‐methylxanthine (1 mM), verapamil (100 µM), and ascorbic acid (1 mM) significantly increased ecto‐ALP activity and decreased 3H‐MPP+ apical transport in this cell line. Orthovanadate (100 µM) showed no effect on 3H‐MPP+ transport and on ecto‐ALP activity. On the other hand, okadaic acid (310 nM) and all trans‐retinoic acid (1 µM) significantly increased 3H‐MPP+ uptake and inhibited ecto‐ALP activity. There is a negative correlation between the effect of drugs upon ecto‐ALP activity and 3H‐MPP+ apical transport (r = −0.9; P = 0.0014). We suggest that apical uptake of organic cations in Caco‐2 cells is affected by phosphorylation/dephosphorylation mechanisms, and that ecto‐ALP activity may be involved in this process. J. Cell. Biochem. 87: 408–416, 2002.
Funding Information
  • FCT
  • POCTI
  • FEDER (32550/99)

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