Nonalcoholic fatty liver disease (NAFLD) is one of the most common liver diseases which lacks ideal treatment options. Kaempferol and kaempferide, two natural flavonol compounds isolated from Hippophae rhamnoides L., were reported to exhibit a strong regulatory effect on lipid metabolism, for which the mechanism is largely unknown. In the present study, we investigated the effects of kaempferol and kaempferide on oleic acid (OA)-treated HepG2 cells, a widely used in vitro model of NAFLD. The results indicated an increased accumulation of lipid droplets and triacylglycerol (TG) by OA, which was attenuated by kaempferol and kaempferide (5, 10 and 20 μM). Western blot analysis demonstrated that kaempferol and kaempferide reduced expression of lipogenesis-related proteins, including sterol regulatory element-binding protein 1 (SREBP1), fatty acid synthase (FAS) and stearoyl-CoA desaturase 1 (SCD-1). Expression of peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT enhancer binding proteins β (C/EBPβ), two adipogenic transcription factors, was also decreased by kaempferol and kaempferide treatment. In addition, western blot analysis also demonstrated that kaempferol and kaempferide reduced expression of heme oxygenase-1 (HO-1) and nuclear transcription factor-erythroid 2-related factor 2 (Nrf2). Molecular docking was performed to identify the direct molecular targets of kaempferol and kaempferide, and their binding to SCD-1, a critical regulator in lipid metabolism, was revealed. Taken together, our findings demonstrate that kaempferol and kaempferide could attenuate OA-induced lipid accumulation and oxidative stress in HepG2 cells, which might benefit the treatment of NAFLD.

The high incidence of diabetes mellitus has caused widespread concern around the world, and has quickly become one of the most prevalent and costly chronic diseases. Alpha-glucosidase has been considered as the main target enzyme in the prevention and treatment of type 2 diabetes, and the inhibition of alpha-glucosidase activity can control postprandial blood glucose levels of diabetics and keep blood glucose levels normal. However, most of the current antidiabetic drugs in the clinic have many side effects. Recently, the screening of alpha-glucosidase inhibitors, from natural products, with excellent activity and less side effects has been successfully achieved, which also proposes more requirements and challenges for the approaches of alpha-glucosidase inhibitor (AGI) screening because of the trace level of active compounds in natural products, large-scale sample screening, and complex matrix of natural products. In this review, we comprehensively summarized the available strategies for the assay of alpha-glucosidase activity and AGI screening for the first time, including classic strategies such as high throughput screening methods (e.g. capillary electrophoresis strategies, HPLC/LC-MS strategies), emerging sensing strategies (e.g. colorimetric sensing, fluorescence sensing, electrochemical sensing, and surface plasmon resonance sensing) and other strategies. Finally, future opportunities and challenges in future research are tentatively proposed.

Dexamethasone is a glucocorticoid analog, which is reported to induce insulin resistance and to exacerbate diabetic symptoms. In this study, we investigated the association between mitochondrial dysfunction and the pathophysiology of dexamethasone-induced insulin resistance. An insulin resistance model in 3T3-L1 adipocyte was established by 48-h treatment of 1 μM dexamethasone, followed with the detection of mitochondrial function. Results showed that dexamethasone impaired insulin-induced glucose uptake and caused mitochondrial dysfunction. Abnormality in mitochondrial function was supported by decreased intracellular ATP and mitochondrial membrane potential (MMP), increased intracellular and mitochondrial reactive oxygen species (ROS) and mtDNA damage. Mitochondrial dynamic changes and biogenesis were suggested by decreased Drp1, increased Mfn2, and decreased PGC-1, NRF1, and TFam, respectively. The mitochondrial DNA (mtDNA) copy number exhibited no change while the mitochondrial mass increased. In agreement, studies in isolated mitochondria from mouse liver also showed dexamethasone-induced reduction of mitochondrial respiratory function, as suggested by decreased mitochondrial respiration controlling rate (RCR), lower MMP, declined ATP synthesis, opening of the mitochondrial permeability transition pore (mPTP), damage of mtDNA, and the accumulation of ROS. In summary, our study suggests that mitochondrial dysfunction occurs along with dexamethasone-induced insulin resistance in 3T3 L1 adipocytes and might be a potential mechanism of dexamethasone-induced insulin resistance.

Oleanolic acid (OA), asiatic acid (AA), and maslinic acid (MA) are ubiquitous isomeric triterpene phytochemicals with many pharmacological effects. To improve their application value, we used lipopolysaccharide (LPS) to induce RAW264.7 cells and studied the differences in the anti-inflammatory effects of the triterpenes according to their structural differences. MTT, Griess, and immunofluorescence assays, ELISA, flow cytometry, and Western blotting, were performed. The release of LPS-induced pro-inflammatory mediators, such as nitric oxide (NO), inducible nitric oxide synthase (iNOS), and interleukin (IL-6), was significantly inhibited by OA, AA, and MA at the same concentration, and AA and MA promoted the production of anti-inflammatory factor IL-10. OA, AA, and MA inhibited LPS-induced NF-κB nuclear translocation in RAW264.7 cells. OA and AA inhibited the phosphorylation of ERK1/2, P38, and JNK1/2 in LPS-stimulated RAW264.7 cells. Moreover, OA increased LPS-induced Nrf2 expression and decreased Keap1 expression in RAW264.7 cells. OA, AA, and MA inhibited LPS-stimulated intracellular reactive oxygen species (ROS) production and alleviated mitochondrial membrane potential depletion. Overall, our data suggested that OA, AA, and MA exhibited significant anti-inflammatory effects in vitro. In particular, OA and AA take effects through the MAPKs, NF-κB, and Nrf2 signaling pathways.

Aporphine alkaloids from the leaves of Nelumbo nucifera Gaertn are substances of great interest because of their important pharmacological activities, particularly anti-diabetic, anti-obesity, anti-hyperlipidemic, anti-oxidant, and anti-HIV’s activities. In order to produce large amounts of pure alkaloid for research purposes, a novel method using high-speed counter-current chromatography (HSCCC) was developed. Without any initial cleanup steps, four main aporphine alkaloids, including 2-hydroxy-1-methoxyaporphine, pronuciferine, nuciferine and roemerine were successfully purified from the crude extract by HSCCC in one step. The separation was performed with a simple two-phase solvent system composed of n-hexane-ethyl acetate-methanol-acetonitrile-water (5:3:3:2.5:5, v/v/v/v/v). In each operation, 100 mg crude extracts was separated and yielded 6.3 mg of 2-hydroxy-1-methoxyaporphine (95.1% purity), 1.1 mg of pronuciferine (96.8% purity), 8.5 mg of nuciferine (98.9% purity), and 2.7 mg of roemerine (97.4%) respectively. The chemical structure of four aporphine alkaloids are identified by means of electrospray ionization MS (ESI-MS) and nuclear magnetic resonance (NMR) analysis. Moreover, the effects of four separated aporphine alkaloids on insulin-stimulated glucose consumption were examined in 3T3-L1 adipocytes. The results showed that 2-hydroxy-1-methoxyaporphine and pronuciferine increased the glucose consumption significantly as rosiglitazone did.

α-glucosidase is a major enzyme that is involved in starch digestion and type 2 diabetes mellitus. In this study, the inhibition of hypericin by α-glucosidase and its mechanism were firstly investigated using enzyme kinetics analysis, real-time interaction analysis between hypericin and α-glucosidase by surface plasmon resonance (SPR), and molecular docking simulation. The results showed that hypericin was a high potential reversible and competitive α-glucosidase inhibitor, with a maximum half inhibitory concentration (IC₅₀) of 4.66 ± 0.27 mg/L. The binding affinities of hypericin with α-glucosidase were assessed using an SPR detection system, which indicated that these were strong and fast, with balances dissociation constant (KD) values of 6.56 × 10⁻⁵ M and exhibited a slow dissociation reaction. Analysis by molecular docking further revealed that hydrophobic forces are generated by interactions between hypericin and amino acid residues Arg-315 and Tyr-316. In addition, hydrogen bonding occurred between hypericin and α-glucosidase amino acid residues Lys-156, Ser-157, Gly-160, Ser-240, His-280, Asp-242, and Asp-307. The structure and micro-environment of α-glucosidase enzymes were altered, which led to a decrease in α-glucosidase activity. This research identified that hypericin, an anthracene ketone compound, could be a novel α-glucosidase inhibitor and further applied to the development of potential anti-diabetic drugs.