Monensin inhibits the expression of sucrase-isomaltase in Caco-2 cells at the mRNA level

AbstractUsing L-[35S]methionine labeling, SDS-PAGE and Northern blot analysis of sucrase-isomaltase mRNA, two different concentrations of monensin were used to delineate in Caco-2 cells the effect of the drug on the conversion of the high mannose to the complex form of sucrase-isomaltase from its dual effect on the biosynthesis of the enzyme and on the rate of glucose consumption. At 0.1 μM the drug has no effect on the rate of glucose consumption and, although it inhibits the conversion of the high mannose to the complex form of the enzyme, it has no effect on the level of sucrase-isomaltase mRNA and on the amount of neosynthesized enzyme. At 1 μM, in addition to its inhibiting effect on the maturation of the enzyme, monensin provokes concomitantly an increase in the rate of glucose consumption and a decrease in the level of sucrase-isomaltase mRNA and in the amount of neosynthesized enzyme. All these effects are reversible within 48 h after removal of the drug

Enterocyte-like differentiation and polarization of the human colon carcinoma cell line Caco-2 in culture

International audienceThe buman colon carcinoma cell line Caco-2 grown in vitro under standard culture conditions in the absence of inducers of differentiation spontaneously exbibits signs of structural and functional differentiation and polarization. The differentiation is total at late confluency. Transmission and scanning electron microscopy show that the entire upper side of the monolayer is covered with typical brusb border microvilli which extend perpendicularly to the surface. The asymmetry of the cells and the presence of tight junctions suggest that the monolayer is polarized. Functionally the differentiation is characterized by high levels of activity of the brush border associated enzymes. The levels of alkaline phospbatase and sucrase·isomaltase are nearly 50% of those found in similar preparations of human small intestine, and 10% in the case of aminopeptidase. The presence of the enzymes is further confirmed by the immunofluorescence staining of the cells with anti-sucrase-isomaltase antibodies. The functional enterocytic differentiation of the brush border microvilli is associated with the formation of domes which are typical of transporting epithelial monolaycrs. ln the early proliferation phase of the culture 70% of the cells are structurally differentiated, but only a few of them are immunoreactive with anti-sucrase· isomaltase antibodies, and the enzyme activities are low