Cyanidin inhibits adipogenesis in 3T3-L1 preadipocytes by activating the PLC-IP₃ pathway

Cyanidin is the most abundant anthocyanin found in red-purple plants and possesses anti-obesity properties. However, its mechanism of action in adipocytes remains unknown. The objective of this study was to elucidate how cyanidin inhibits adipocyte formation in 3T3-L1 preadipocytes. Cells were cultured in adipogenic differentiation medium supplemented with cyanidin and examined for adipogenesis, cell viability, and adipocyte gene expression using Oil Red O staining, MTT assay, and RT-qPCR. Real-time Ca²⁺ imaging analysis was performed in living cells to elucidate cyanidin's mechanism of action. The results demonstrated that cyanidin (1-50 μM) supplementation to the adipogenic medium inhibited adipogenesis by downregulating adipogenic marker gene expression (PPARγ, C/EBPα, adiponectin, and aP2) without affecting cell viability after 4 days of treatment. Stimulation of cells with cyanidin (30-100 μM) increased intracellular Ca²⁺ in a concentration dependent manner with peak calcium increases at 50 μM. Pretreatment of cells with the phospholipase C (PLC) inhibitor U73122, inositol triphosphate (IP₃) receptor blocker 2-APB, and depletion of endoplasmic reticulum Ca²⁺ stores by thapsigargin abolished the Ca²⁺ increases by cyanidin. These findings suggested that cyanidin inhibits adipocyte formation by activating the PLC-IP₃ pathway and intracellular Ca²⁺ signaling. Our study is the first report describing the mechanism underlying the anti-obesity effect of cyanidin.