Signaling mechanisms involved in the effects of mucosal leptin on glucose absorption and Na+-K+ ATPase in Caco-2 cells -

Abstract

The aim of this project was to determine the effect of leptin on intestinal glucose absorption using Caco-2 cells as a model. Cells grown on transwell filters were incubated 23 days after confluence in presence of leptin and [14C] 3-O-methyl-D-glucose and the amount of radioactive glucose in the lower chamber was measured at different time points after confluence. The Na+ -K+ ATPase activity was also assayed by measuring the amount of inorganic phosphate liberated in presence and absence of inhibitors of the ATPase. The expression levels of leptin receptors and glucose transporters were monitored in isolated apical and basolateral cell membranes. As Caco-2 cells differentiated after confluence, the amount of glucose absorbed increased along with an increase in the level of Na+-dependent glucose transporter (SGLT1) and facilitative glucose transporter (GLUT2). In fully differentiated Caco-2 cells which resemble enterocytes, leptin reduced glucose absorption by decreasing both the numbers of glucose transporters and the activity of the Na+ -K+ ATPase. To elucidate the signaling pathway of leptin, a panel of specific inhibitors was used to inactivate different putative signaling mediators individually and in different combinations. The results show that leptin inhibits glucose absorption by the sequential activation of protein kinase C (PKC), p38mitogen activated protein kinase (p38MAPK), phosphatidylinositol-3-OH-kinase (PI3K) and extracellular signal-regulated kinase (ERK). Leptin reduces the activity of the Na+ -K+ ATPase by activating p38MAPK via an inhibition of its inhibitor, PKC. ERK and PI3K were not found to be directly involved in this pathway. It is concluded that leptin regulates body weight not only by reducing appetite but also by reducing nutrient absorption in particular glucose.