L-arginine attenuates Interleukin-1β (IL-1β) induced Nuclear Factor Kappa-Beta (NF-κB) activation in Caco-2 cells.
BACKGROUND
Specific nutrients like L-arginine (L-Arg) ameliorate intestinal inflammation, however the exact mechanisms of this effect are unclear. We hypothesized the anti-inflammatory effects of L-Arg require active transport and metabolism by inducible nitric oxide synthase (iNOS) to generate nitric oxide (NO). To test this hypothesis we examined the effects of L-Arg, L-Arg transport activity, NO production and iNOS inhibitor on IL-1β-mediated NF-κB-activation in Caco-2 cells.


METHODS
Caco-2 cells were cultured, transfected with a NF-κB promoter luciferase vector, incubated ± L-Arg, ± IL-1β and luciferase activity was measured. Using siRNA we inhibited the L-Arg cationic amino acid transporter system y+ (CAT1) expression and examined its effects on L-Arg transport activity and IL-1β-mediated NF-κB-activation. Finally, the effects of sodium nitroprusside (SNP, a NO donor) and Nω-nitro-L-arginine (NNA, an iNOS inhibitor) on IL-1β-mediated NF-κB-activation were examined.


RESULTS
IL-1β increased NF-κB luciferase activity (8-fold) and NF-κB expression (mRNA and protein), both of these were significantly decreased by L-Arg. System y+ CAT1 siRNA decreased CAT1 expression, L-Arg transport activity and attenuated the inhibitory effects of L-Arg on NF- κB activity. SNP attenuated the IL-1β-induced increase in NF-κB luciferase activity and expression, whereas NNA diminished the inhibitory effects of L-Arg on IL-1β-inducible NF- κB luciferase activity.


CONCLUSIONS
The inhibitory effects of L-Arg on IL-1β-mediated NF-κB-activation in Caco-2 cells involve L-Arg transport activity by CAT1, regulation of IL-1β-mediated increases in NF-κB expression, changes in iNOS expression and NO production. Our data suggest the inhibitory effects of L-Arg on NF-κB activation are mediated in part by iNOS since SNP preserves and NNA attenuates the effects of L-Arg on IL-1β-mediated NF-κB-activation and expression.
Department of Surgery, SUNY Upstate Medical University, Syracuse, New York, United States of America.
Qinghe
Q
Meng
Department of Surgery, SUNY Upstate Medical University, Syracuse, New York, United States of America.
Mitchell
M
Cooney
Department of Surgery, SUNY Upstate Medical University, Syracuse, New York, United States of America.
Natesh
N
Yepuri
Department of Surgery, SUNY Upstate Medical University, Syracuse, New York, United States of America.
Robert N
RN
Cooney
D001120:Arginine / Q000494:pharmacology*; D018938:Caco-2 Cells; D027201:Cationic Amino Acid Transporter 1 / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D053583:Interleukin-1beta / Q000494:pharmacology*; D015850:Interleukin-6 / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000037:antagonists & inhibitors; D034741:RNA, Small Interfering
D027201:Cationic Amino Acid Transporter 1; D004791:Enzyme Inhibitors; D053583:Interleukin-1beta; D015850:Interleukin-6; D016328:NF-kappa B; D034741:RNA, Small Interfering; D009569:Nitric Oxide; D001120:Arginine; D052247:Nitric Oxide Synthase Type II