Mitogen-Activated Protein Kinases (MAPK) mediates Epidermal Growth Factor (EGF) up regulation of arginine transport in cultured intestinal epithelia

Abstract

Introduction. Intestinal luminal growth factor EGF plays a key role in regulating intestinal epithelial growth and differentiation as well as amino acid absorption. However, little is known about the intracellular regulatory mechanisms involved. The purpose of this in vitro study was to delineate the intracellular signaling pathways involved in the EGF stimulation of arginine transport in cultured intestinal epithelial cells. Methods. IEC-6 cells were grown to sub-confluence and incubated in EGF (0–200 ng/ml), ± inhibitors of MAPK ERK1 (PD 98059, 0–50 μM) or MAPK p38 (SB 203580, 0–10 μM). 3H-arginine (5 μm to 10 mM) transport activity, arginine transporter MCAT1 mRNA, and MAPK levels were measured. Data are means ± SD and analyzed by ANOVA with P < 0.05. Results. EGF stimulated arginine transport activity in a dose- and time-dependent fashion. Prolonged EGF exposure (100 ng/ml, 48 h) resulted in a 48% increase of arginine transport and a 2.5-fold increase in transporter MCAT1 mRNA levels. The transport activity increase was the result of an increase of maximal transport velocity of both Systems y + (V max 40 ± 7 control versus 74 ± 13 EGF, nmol/mg/min, P < 0.05) and L (V max 9.8 ± 1.2 control versus 17 ± 2.3 EGF, P < 0.05). The transport affinity of both systems was not affected ( P = NS). EGF increased the phopspho-MER1, phospho-p44/42, and phospho-p38 levels, the active forms of MAPK, without affecting the total MAPK levels. PD 98059 or SB 203580 individually attenuated the EGF-induced arginine transport activity and transporter mRNA levels. Conclusion. EGF stimulates arginine transport activity and transporter gene expression in IEC-6 cells via an intracellular MAPK signaling cascade. TABLE—ABSTRACT P14 Treatment Control PD 98059 SB 203580 Control 0.213 ± 0.11 0.238 ± 0.03 0.225 ± 0.05 EGF 0.309 ± 0.03 ∗ 0.256 ± 0.02 ∗∗ 0.245 ± 0.03 ∗∗ Note. Unit: nmole/mg/min; ∗ p < 0.05 versus control; ∗∗ p < 0.05 versus EGF alone.