β-Carotene Suppresses Lipopolysaccharide-induced Nitric Oxide Production in Microglia via Retinoic Acid Receptor-dependent Mechanisms

Carotenoids are one of the common food ingredients with potential anti-inflammatory effects. Screening of carotenoids for their anti-inflammatory effects has predominantly been conducted using macrophages, whereas comparative analyses of their effects on excessive activation of microglia remain limited. This study aimed to evaluate the anti-inflammatory effects of eight different carotenoids in lipopolysaccharide (LPS)-stimulated microglia, focusing on nitric oxide (NO) production suppression and underlying molecular mechanisms. Mouse microglia cell line MG6 was pretreated with carotenoids, followed by stimulation with LPS. Among the tested carotenoids, β-carotene, echinenone, and siphonaxanthin exhibited strong NO-inhibitory effects. Western blot analysis indicated that the effect of siphonaxanthin was mediated by NF-κB inhibition. Lipidomic analysis revealed distinct effects of β-carotene and siphonaxanthin on LPS-stimulated microglia, indicating possible differences in their inhibitory mechanisms. Gene expression analysis showed that β-carotene increased the expression of elongation of very long chain fatty acids (ELOVL) 3, ELOVL4, and fatty acid synthase, while restoring ELOVL6 mRNA expression. These results were consistent with the lipidomic analysis. Pharmacological inhibition studies demonstrated that β-carotene exerted its inhibitory effects via retinoic acid receptor (RAR)-dependent mechanisms, although the involvement of RAR-independent pathways cannot be excluded. Echinenone exhibited RAR-dependent suppression of NO production comparable to that of β-carotene. These findings suggest that carotenoids modulate neuroinflammation through multiple mechanisms, including NF-κB inhibition (e.g. siphonaxanthin), RAR activation (e.g. β-carotene and echinenone), and possibly alternative signaling pathways (e.g. astaxanthin), although the detailed mechanisms remain to be elucidated. Since RAR activation has been shown to improve cognitive function in mouse models of Alzheimer's disease, β-carotene and echinenone may have therapeutic potential in reducing neuroinflammation and preserving cognitive function. Further in vivo studies are necessary to validate these effects.