Differential regulation of L-arginine transport and inducible NOS in cultured vascular smooth muscle cells

Experiments were performed to characterize the uptake of L-arginine in rat aortic smooth muscle cells (SMC) and to examine whether inducers of nitric oxide synthase (NOS) could regulate the transport of L-arginine into these cells. L-Arginine transport by SMC was saturable, Na+ independent, strongly inhibited in the presence of other cationic amino acids, and could be stimulated by preloading the cells with cationic amino acids. Kinetic studies revealed the presence of both a high [Michaelis constant (Km) approximately 125 microM] and low (Km approximately 1.4 mM) affinity L-arginine transporter. Treatment of vascular SMC with interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF-alpha) resulted in parallel increases in L-arginine transport and nitric oxide (NO) synthesis, as measured by nitrite production. Increasing the concentration of IL-1 beta and TNF-alpha caused a progressive elevation in nitrite production but did not further stimulate L-arginine uptake. Treatment of SMC with the combination of TNF-alpha and interferon-gamma (IFN-gamma) synergistically enhanced nitrite release without having any additional effect on L-arginine transport. Both induction of L-arginine transport and NOS activity by these cytokines were blocked by cycloheximide. Finally, treatment of SMC with interferon-gamma and N6,2'-O-dibutyryl adenosine 3',5'-cyclic monophosphate selectively stimulated the formation of nitrite by SMC but had no effect on L-arginine transport. These results demonstrate that L-arginine transport by cultured vascular SMC is mediated by the system y+ carrier and that inducers of NOS differentially regulate the activity of this transporter.(ABSTRACT TRUNCATED AT 250 WORDS)