Antiadipogenic Effect of Citrus Flavonoids: Evidence from RNA Sequencing Analysis and Activation of AMPK in 3T3-L1 Adipocytes.

Citrus fruits are rich in dietary flavonoids and have many health benefits, but their antiadipogenic mechanism of action and their impact on lipid metabolism remain unclear. In this study, we investigated the effect of citrus flavonoids, namely, hesperidin (HES), narirutin (NAR), nobiletin (NOB), sinensetin (SIN), and tangeretin (TAN), on preventing fat cell development by gene expression in 3T3-L1 adipocytes. Among the citrus flavonoids tested, HES and NAR significantly reduced fat storage and triglyceride levels and increased glucose uptake in 3T3-L1 adipocytes. Additionally, HES and NAR treatment increased the phosphorylation of AMP-activated protein kinase (AMPK) and acetyl-CoA carboxylase (ACC) while reducing the protein expression of 3-hydroxy-3-methylglutaryl CoA reductase (HMGCR). Furthermore, in silico docking revealed that flavonoids activate AMPK. RNA sequencing analysis demonstrated that citrus flavonoids normalized the expression of 40 genes, which were either upregulated by more than 2-fold or downregulated by less than 0.6-fold including Acadv1, Acly, Akr1d1, Awat1, Cyp27a1, Decr1, Dhrs4, Elovl3, Fasn, G6pc, Gba, Hmgcs1, Mogat2, Lrp5, Sptlc3, and Snca to levels comparable to the control group. Altogether, HES and NAR among five citrus flavonoids showed antiadipogenic effects by regulating the expression of specific lipid metabolism genes partially restored to control levels in 3T3-L1 cells.

Comprehensive Assessment of Antioxidant and Anti-Inflammatory Properties of Papaya Extracts.

Antioxidant and anti-inflammatory properties of papaya (Carica papaya) fruits were evaluated to provide comprehensive information associated with the bioactive compounds. 'Tainung No. 2' papaya fruits, cultivated in a greenhouse, Korea, were harvested at unripe and ripe stages and then divided into seed and peel-pulp. Total phenolic and flavonoid contents were determined using spectrophotometry, and individual phenolic compounds were relatively quantified by HPLC-DAD and fifteen standards. Antioxidant activities were measured using four assays: DPPH (2,2-diphenyl-1-picrylhydrazyl) and ABTS (2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid)) scavenging activities, inhibition of lipid peroxidation, and FRAP (ferric reducing antioxidant power). Anti-inflammatory activities were measured by the regulation of NF-κB signaling pathways with the measurements of ROS and NO productions as the degree of oxidative stress. Total phenol contents increased in seed and peel-pulp extracts during ripening; flavonoid contents increased only in seed extracts. Total phenolic contents were associated with ABTS radical scavenging activity and FRAP. Of fifteen phenolic compounds, chlorogenic acid, cynarin, eupatorine, neochlorogenic acid, and vicenin II were identified among papaya extracts. ROS and NO productions were inhibited in papaya extracts. Especially, NO productions were inhibited higher in ripe seed extracts than in other extracts, which would be associated with the suppression of NF-κB activation and iNOS expression. These results suggest that papaya fruit extracts, including seeds, peels, and pulps, could be potential raw materials for functional foods.

In Vitro and In Vivo Effects of Nobiletin on DRG Neurite Elongation and Axon Growth after Sciatic Nerve Injury.

Sciatic nerve injury (SNI) leads to sensory and motor dysfunctions. Nobiletin is a major component of polymethoxylated flavonoid extracted from citrus fruits. The role of nobiletin on sciatic nerve regeneration is still unclear. Thus, the purpose of this study was to investigate whether nobiletin increases DRG neurite elongation and regeneration-related protein expression after SNI. Cytotoxicity of nobiletin was measured in a concentration-dependent manner using the MTT assay. For an in vitro primary cell culture, the sciatic nerve on the middle thigh was crushed by holding twice with forceps. Dorsal root ganglion (DRG) and Schwann cells were cultured 3 days after SNI and harvested 36 h later and 3 days later, respectively. In order to evaluate specific regeneration-related markers and axon growth in the injured sciatic nerve, we applied immunofluorescence staining and Western blot techniques. Nobiletin increased cell viability in human neuroblastoma cells and inhibited cytotoxicity induced by exposure to H2O2. Mean neurite length of DRG neurons was significantly increased in the nobiletin group at a dose of 50 and 100 µM compared to those at other concentrations. GAP-43, a specific marker for axonal regeneration, was enhanced in injury preconditioned Schwann cells with nobiletin treatment and nobiletin significantly upregulated it in injured sciatic nerve at only 3 days post crush (dpc). In addition, nobiletin dramatically facilitated axonal regrowth via activation of the BDNF-ERK1/2 and AKT pathways. These results should provide evidence to distinguish more accurately the biochemical mechanisms regarding nobiletin-activated sciatic nerve regeneration.