Mechanism of thiamine uptake by human colonocytes: studies with cultured colonic epithelial cell line NCM460

Thiamine (vitamin B-1) is essential for normal cellular functions and growt h. Mammals cannot synthesize thiamine and thus must obtain the vitamin via intestinal absorption. The intestine is exposed to a dietary thiamine sourc e and a bacterial source in which the vitamin is synthesized by the normal microflora of the large intestine. Very little is known about thiamine upta ke in the large intestine. The aim of this study was, therefore, to address this issue. Our results with human-derived colonic epithelial NCM460 cells as a model system showed thiamine uptake to be 1) temperature- and energy dependent, 2) Na+ independent, 3) increased with increasing buffer pH from 5 to 8 and after cell acidification but inhibited by amiloride, 4) saturabl e as a function of concentration, 5) inhibited by thiamine structural analo gs but not by unrelated organic cations, and 6) inhibited by modulators of a Ca2+/calmodulin-mediated pathway. NCM460 cells and native human colonic m ucosa expressed the recently cloned human thiamine transporter THTR-1 (prod uct of the SLC19A2 gene) at both mRNA and protein levels. These results dem onstrate for the first time that human NCM460 colonocytes possess a specifi c carrier-mediated system for thiamine uptake that appears to be under the regulation of an intracellular Ca2+/calmodulin-mediated pathway. It is sugg ested that bacterially synthesized thiamine in the large intestine may cont ribute to thiamine nutrition of the host, especially toward cellular nutrit ion of the local colonocytes.