Cyanidin-3-O-glucoside suppresses adipogenesis in the early phase in 3T3-L1 cells through activation of thermogenic pathways

Background Adipocytes hyperplasia is a condition that mainly contributes to obesity development by engaging preadipocytes proliferation and differentiation into mature adipocytes in adipose tissue, via a process named adipogenesis. This process, through the involvement of different transcriptional factors and signaling cascades induces lipid droplets generation. During the early phase of this process, preadipocytes go to mitotic clonal expansion followed by the activation of peroxisome proliferator-activated receptor γ (PPARγ) leading to fatty acids synthesis and adipokine production. On the contrary, AMP-activated protein kinase (AMPK) acts as a key nutrient sensor on energy metabolism inducing catabolic metabolism (e.g., fatty acid β-oxidation, glucose uptake, and mitochondria biosynthesis). This mechanism, known as thermogenic process, is considered a valid target for the treatment of obesity. Therefore, the present work examined the in vitro beneficial effects of cyanidin-3-O-glucoside (C3G) on the inhibition of the adipogenic process in 3T3-L1 cells, evaluating molecular pathways involved in adipogenesis and thermogenesis, during the early and the late phase of adipocyte differentiation. Methods C3G at two different concentrations (5-10 μM) has been added to 3T3-L1 cells during the early phase (day 0 to day 4) or the late phase (day 4 to day 10) of the differentiation process. C3G effects on lipid accumulation were evaluated through Oil Red O staining, whereas the main markers involved in adipogenesis and thermogenesis were evaluated via Western blot and qPCR techniques. Results Data showed that addition of C3G during the early phase, rather than in the late phase, of the 3T3-L1 differentiation process resulted in a marked suppression of adipogenesis, as demonstrated by reduced lipidic accumulation and inhibition of the main markers involved in the adipogenic process (PPARγ, C/EBPβ, and FASN). Furthermore, C3G activated the thermogenic process in cells exposed during the early phase of differentiation via the upregulation of pAMPK, and of downstream genes (UCP1 and PGC1α). Conclusions These results clearly indicate that C3G inhibits adipocyte differentiation mainly during the early phase of adipogenesis via activation of the thermogenic process. Thus, since decreasing early adipogenic transcription markers and increasing transcription inhibitors related to adipogenesis contribute to the suppression of terminal adipogenic differentiation, C3G could be used to prevent/treat adipose tissue development and obesity. References Ali et al., 2013. Eur. J. Cell Biol. 2013, 92, 229–236. Engin 2017. Adv Exp Med Biol. 2017; 960:1-17. Montanari et al., 2017. Obes Rev. 2017 May;18(5):495-513. Salehi et al., 2020. Front Pharmacol. 2020 Aug 26;11:1300.