Arginine-induced stimulation of protein synthesis and survival in IPEC-J2 cellsis mediated by mTOR but not nitric oxide.

摘要

Arginine is an indispensable amino acid in neonates and is required for growth.Neonatal intestinal epithelial cells (IEC) are capable of arginine transport,catabolism, and synthesis and express nitric oxide (NO) synthase to produce NOfrom arginine. Our aim was to determine whether arginine directly stimulates IEC growth and protein synthesis and whether this effect is mediated via mammaliantarget of rapamycin (mTOR) and is NO-dependent. We studied neonatal porcine IEC(IPEC-J2) cultured in serum- and arginine-free medium with increasing arginineconcentrations for 4 or 48 h. Our results show that arginine enhances IPEC-J2cell survival and protein synthesis, with a maximal response at a physiologicalconcentration (0.1-0.5 mM). Addition of arginine increased the activation ofmTOR, p70 ribosomal protein S6 (p70 S6) kinase, and eukaryotic initiation factor 4E-binding protein 1 (4E-BP1) in a time- and dose-dependent manner. Thearginine-induced protein synthesis response was not inhibited by the NOinhibitors nitro-l-arginine methyl ester (l-NAME) and aminoguanidine, despiteinducible NO synthase expression in IPEC-J2 cells. Moreover, protein synthesiswas not increased or decreased in some cases by addition of an NO donor(S-nitroso-N-acetylpenicillamine), arginine precursors (proline and citrulline)in the absence of arginine, or insulin; S-nitroso-N-acetylpenicillaminesuppressed phosphorylation of mTOR, p70 S6 kinase, and 4E-BP1. We found amarkedly higher arginase activity in IPEC-J2 cells than in primary pig IEC.Furthermore, mTOR inhibition by rapamycin partially (42%) reduced thearginine-induced protein synthesis response and phosphorylation of mTOR and4E-BP1. We conclude that arginine-dependent cell survival and protein synthesissignaling in IPEC-J2 cells are mediated by mTOR, but not by NO.