TNFα regulates sugar transporters in the human intestinal epithelial cell line Caco-2

Abstract

Purpose: During intestinal inflammation TNFα levels are increased and as a consequence malabsorption of nutrients may occur. We have previously demonstrated that TNFα inhibits galactose, fructose and leucine intestinal absorption in animal models. In continuation with our work, the purpose of the present study was to investigate in the human intestinal epithelial cell line Caco-2, the effect of TNFα on sugar transport and to identify the intracellular mechanisms involved.

Methods: Caco-2 cells were grown on culture plates and pre-incubated during different periods with various TNFα concentrations before measuring the apical uptake of galactose, α-methyl-glucoside (MG) or fructose for 15 min. To elucidate the signaling pathway implicated, cells were pre-incubated for 30min with the PKA inhibitor H-89 or the PKC inhibitor chelerythrine, before measuring the sugar uptake. The expression in the apical membrane of the transporters implicated in the sugars uptake process (SGLT1 and GLUT5) was determined by Western blot.

Results: TNFα inhibited 0.1mM MG uptake after pre-incubation of the cells for 6-48h with the cytokine and in the absence of cytokine pre-incubation. In contrast, 5mM fructose uptake was stimulated by TNFα only after long pre-incubation times (24 and 48 h). These effects were mediated by the binding of the cytokine to its specific receptor TNFR1, present in the apical membrane of the Caco-2 cells. Analysis of the expression of the MG and fructose transporters at the brush border membrane of the cells, after 24h pre-incubation with the cytokine, revealed decrease on the amount of SGLT1 and increase on the amount of GLUT5 proteins. Short-term inhibition of MG transport by TNFα was not modified by H-89 but was blocked by chelerythrine.

Conclusions: SGLT1 and GLUT5 expression in the plasma membrane is regulated by TNFα in the human epithelial cell line Caco-2 cells, leading to alteration on sugars transport, suggesting that TNFα could be considered as a physiological local regulator of nutrients absorption in response to an intestinal inflammatory status.

Keywords: Caco-2 cells; GLUT5; PKC; SGLT1; TNF-α.