Transport of thiamine in human intestine: mechanism and regulation in intestinal epithelial cell model Caco-2

The present study examined the intestinal uptake of thiamine (vitamin B-1) using the human-derived intestinal epithelial cells Caco-2 as an in vitro m odel system. Thiamine uptake was found to be 1) temperature and energy depe ndent and occurred with minimal metabolic alteration; 2) pH sensitive; 3) N a+ independent; 4) saturable as a function of concentration with apr appare nt Michaelis-Menten constant of 3.18 +/- 0.56 mu M and maximal velocity of 13.37 +/- 0.94 pmol.mg protein(-1).3 min(-1); 5) inhibited by the thiamine structural analogs amprolium and oxythiamine, but not by unrelated organic cations tetraethylammonium, N-methylnicotinamide, and choline; and 6) inhib ited in a competitive manner by amiloride with an inhibition constant of 0. 2 mM. The role of specific protein kinase-mediated pathways in the regulati on of thiamine uptake by Caco-2 cells was also examined using specific modu lators of these pathways. The results showed possible involvement of a Ca2/calmodulin (CaM)-mediated pathway in the regulation of thiamine uptake. No role for protein kinase C- and protein tyrosine kinase-mediated pathways i n the regulation of thiamine uptake was evident. These results demonstrate the involvement of a carrier-mediated system for thiamine uptake by Caco-2 intestinal epithelial cells. This system is Na+ independent and is differen t from the transport systems of organic cations. Furthermore, a CaM-mediate d pathway appears to play a role in regulating thiamine uptake in these cel ls.