Epidermal growth factor inhibits glycylsarcosine transport and hPepT1 expression in a human intestinal cell line

The human intestinal cell line Caco-2 was used as a model system to study t he effects of epidermal growth factor (EGF) on peptide transport. EGF decre ased apical-to-basolateral fluxes of [C-14]glycylsarcosine ([C-14]Gly-Sar) up to 50.2 +/- 3.6% (n = 6) of control values. Kinetic analysis of the flux es showed that maximal flux (V-max) of transepithelial transport decreased from 3.00 +/- 0.17 nmol.cm(-2).min(-1) in control cells to 0.50 +/- 0.07 nm ol.cm(-2).min(-1) in cells treated with 5 ng/ ml EGF (n = 6, P < 0.01). The apparent Michaelis-Menten constant (K-m) was 2.71 +/- 0.31 mM (n = 6) in c ontrol cells and 1.89 +/- 0.28 mM (n 5 6, not significantly different from control) in EGF-treated cells. Similarly, apical uptake of [C-14]Gly-Sar de creased in cells treated with EGF, with an ED50 value of 0.36 +/- 0.06 ng/ ml (n = 6) EGF and a maximal inhibition of 80 +/- 0.02% (n = 6). V-max decr eased from 2.61 +/- 0.4 to 1.06 +/- 0.1 nmol.cm(-2).min(-1) (n = 3, P < 0.0 5), whereas K-m remained constant. Basolateral Gly-Sar uptake showed no cha nges in V-max or K-m after EGF treatment (n 5 3). RT-PCR showed a decrease in hPepT1 mRNA (using glucose-6-phosphate dehydrogenase mRNA as control) in cells treated with EGF. Western blotting indicated a decrease in hPepT1 pr otein in cell lysates. We conclude that EGF treatment decreases Gly-Sar tra nsport in Caco-2 cells by decreasing the number of peptide transporter mole cules in the apical membrane.