Garlic attenuates nitric oxide production in rat cardiac myocytes through inhibition of inducible nitric oxide synthase and the arginine transporter CAT-2 (cationic amino acid transporter-2)

It is now accepted that allicin, the main biologically active compound in garlic, exhibits antioxidant activity. The present study was designed to test the hypothesis that the antioxidant activity of garlic can be partially attributed to the inhibition of nitric oxide (NO) production by cytokine-induced NO synthase (iNOS). Cardiac myocytes cultured from neonatal Wistar rats were stimulated by lipopolysaccharide (LPS) and incubated for 24h with various concentrations of allicin. This resulted in marked inhibition of nitrite production. Interestingly, a low concentration of allicin (10μM) was significantly more potent in abrogating the effect of LPS on nitrite production than a higher concentration (40μM). Allicin decreased steady-state iNOS mRNA levels, and this effect was maximal when a lower concentration was used (10μM compared with 40μM). In order to explore additional effects of allicin on NO generation that might counteract the effect on iNOS, we assessed the effects of higher allicin concentrations on arginine transport. Allicin inhibited the uptake of 1mM extracellular arginine in a concentration-dependent manner. The expression of the two arginine transporters that are expressed in cardiac myocytes [CAT-1 (cationic amino acid transporter-1) and CAT-2] was studied using reverse transcription-PCR. A concentration of 200μM allicin abolished the expression of CAT-2 mRNA, 100μM significantly attenuated it, whereas 50μM had no effect. Allicin had no effect on steady-state CAT-1 mRNA levels. Our results suggest that allicin inhibits iNOS activity through two different mechanisms: at lower concentrations it decreases iNOS mRNA levels, whereas at higher concentrations it inhibits arginine transport through down-regulation of CAT-2 mRNA.