Trigonelline induces browning in 3T3-L1 white adipocytes.

Trigonelline, a major alkaloid component of fenugreek, has been demonstrated to have several biological activities, including antidiabetic and anticancer effects. This study aimed to examine the possible application of trigonelline as an anti-obesity compound based on an investigation of its enhancement of lipid catabolism and induction of browning in white adipocytes. Trigonelline induces browning of 3T3-L1 white adipocytes by enhancing the expressions of brown-fat signature proteins and genes as well as beige-specific genes, including Cd137, Cited1, Tbx1, and Tmem26. Trigonelline also improves lipid metabolism in white adipocytes by decreasing adipogenesis and lipogenesis as well as promotes lipolysis and fatty acid oxidation. Moreover, trigonelline increases the expression of Cox4, Nrf1, and Tfam genes that are responsible for mitochondrial biogenesis. Mechanistic studies revealed that the browning effect of trigonelline in 3T3-L1 white adipocytes is mediated by activating ß3-AR and inhibiting PDE4, thereby stimulating the p38 MAPK/ATF-2 signaling pathway. Considering its high bioavailability in humans and the results of this study, trigonelline may have potential as an anti-obesity compound.

4ß-Hydroxywithanolide E and withanolide E from Physalis peruviana L. inhibit adipocyte differentiation of 3T3-L1 cells through modulation of mitotic clonal expansion.

Obesity is a risk factor for many diseases, including type 2 diabetes and cardiovascular disease, and is related to the rising morbidity and mortality. Discovery of agents targeting adipogenesis, especially from natural sources, is important for the treatment of obesity. Here, we aimed to identify anti-adipogenic substances in methanol extracts of Physalis peruviana and to investigate their effect, along with underlying mechanisms. Activity-guided fractionation of the extract revealed 4ß-hydroxywithanolide E (HWE) and withanolide E (WE) as the adipogenesis inhibitors. Both compounds suppressed mRNA expression of central adipogenic transcription factors, peroxisome proliferator-activated receptor γ, and CCAAT/enhancer-binding protein α in the early stage of adipocyte differentiation. The inhibitory action of these two withanolides on adipogenesis was largely limited to this stage. The proliferation of preadipocytes was markedly suppressed by treatment with HWE and WE for 24 and 48 h in the differentiation medium, and cell-cycle arrest in the G0/G1 phase was observed. Therefore, our results suggested that withanolides from P. peruviana to be novel anti-adipogenic compounds that modulate mitotic clonal expansion.

Anti-obesity Effect of Fermented Persimmon Extracts via Activation of AMP-Activated Protein Kinase.

There is significant cultivation of persimmon (Diospyros kaki) in East Asia, a plant whose fruit has abundant nutrients, including vitamins, polyphenols, and dietary fiber. Persimmon dietary supplements can benefit health by amelioration of diabetes, cardiovascular disease, and obesity. There are also persimmon-based beverages produced via fermentation, such as wines and vinegars, and increasing consumption of these products in East Asia. Although there is great interest in functional foods, the health effects of fermented persimmon extract (FPE) are completely unknown. We examined the effects of FPE on the metabolic parameters of mice fed a high-fat diet (HFD). Our results indicated that FPE supplementation led to an approx. 15% reduction of body weight, reduced abdominal and liver fat, and reduced serum levels of triglycerides, total cholesterol, and glucose. FPE also blocked the differentiation of murine 3T3-L1 pre-adipocyte cells into mature adipocytes. We suggest that gallic acid is a major bioactive component of FPE, and that AMP-activated protein kinase mediates the beneficial effects of FPE and gallic acid.

Antiobesity and Cholesterol-Lowering Effects of Dendropanax morbifera Water Extracts in Mouse 3T3-L1 Cells.

Obesity is the most common metabolic disease in developed countries and has become a global epidemic in recent years. Obesity is associated with various metabolic abnormalities, including glucose intolerance, insulin resistance, type 2 diabetes, dyslipidemia, and hypertension. Leaves from the plant Dendropanax morbiferus are beneficial to health as they contain high levels of vitamin C and tannin. There have been seminal studies on the anticancer, antimicrobial, antidiabetes, and antihyperglycemic effects of treatments with D. morbiferus trees. Herein, we investigated the toxicity of D. morbiferus water (DLW) extracts in vitro, and demonstrated no toxicity at 5-500 µg/mL in 24-72-h experiments with 3T3-L1 cells. The DLW increased cell viability at 48 h and inhibited adipogenesis in 3T3-L1 cells by reducing intracellular triglyceride levels and glucose uptake. In addition, mRNA and protein expression levels of adipogenesis-related genes were lowered by DLW, suggesting antiobesity effects in mouse 3T3-L1 cells. Because few studies have demonstrated cholesterol-lowering effects of D. morbiferus, we investigated the activities of adipogenic transcriptional factors following treatments of 3T3-L1 cells with D. morbiferus and observed increased CEBPα, CEBPß, PPARγ, and SREBP1 activities in the cells, indicating that DLW extracts inhibit adipogenesis.

Heme oxygenase-1 mediates anti-adipogenesis effect of raspberry ketone in 3T3-L1 cells.

BACKGROUND: Obesity is caused by excessive accumulation of body fat and is closely related to complex metabolic diseases. Raspberry ketone (RK), a major aromatic compound in red raspberry, was recently reported to possess anti-obesity effects. However, its mechanisms are unclear. AIM: Adipogenesis plays a critical role in obesity and, therefore, this study aimed to investigate the effect and mechanisms of action of RK on adipogenesis in 3T3-L1 preadipocytes. MATERIALS AND METHODS: 3T3-L1 preadipocytes were differentiated in medium containing insulin, dexamethasone, and 1-methyl-3-isobutylxanthine. Adipocyte lipid contents were determined using oil-red O staining while adipogenic transcription factor and lipogenic protein expressions were determined using western blotting. RESULTS: RK (300-400µM) strongly inhibited lipid accumulation during 3T3-L1 preadipocyte differentiation into adipocytes. RK reduced the CCAAT/enhancer-binding protein-α (C/EBP-α), peroxisome proliferation-activated receptor-γ (PPAR-γ), fatty acid synthase (FAS), and fatty acid-binding protein 4 (FABP4) expressions and increased heme oxygenase-1 (HO-1), Wnt10b, and ß-catenin expressions in 3T3-L1 adipocytes. Additionally, RK inhibited lipid accumulation, and adipogenic transcription factor and lipogenic protein expressions were all decreased by inhibiting HO-1 or ß-catenin using tin protoporphyrin (SnPP) or ß-catenin short-interfering RNA (siRNA), respectively. Furthermore, Wnt10b and ß-catenin expressions were negatively regulation by SnPP. CONCLUSION: RK may exert anti-adipogenic effects through modulation of the HO-1/Wnt/beta-catenin signaling pathway.

Chemical Compounds in Natural Medicines That Affect Macropharges and Adipocyte Cells.

Macrophages play major roles in inflammation, immunity and host defense mechanisms. Once activated they produce and release cytokines, oxygen and nitrogen species, and eicosanoids. The best characterized stimuli to induce the transcription of genes encoding pro-inflammatory proteins in macrophages in vitro is bacterial lipopolysaccharide (LPS). LPS could be used alone or in combination with recombinant mouse interferon-γ (IFN-γ). Such stimulation results in cytokine release and the synthesis of enzymes such as cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS). The nitric oxide (NO) radical is known to play a central role in inflammatory and immune reactions for self-protection. However, the excessive production of NO may lead to tissue damage. In inflammatory diseases such as rheumatoid arthritis, excessive NO production by activated macrophages has been observed. Adipose tissue is composed of various cell types such as mature adipocytes, preadipocytes, fibroblasts, endothelial cells, vascular cells, and macrophages. Recent studies indicate that obesity is associated with low-grade chronic inflammation of adipose tissues, and that such inflammation is one of the potential mechanisms leading to the insulin resistance. It has been demonstrated that obese adipose tissue is characterized by the increased infiltration of macrophages. Therefore, we attempted to identify natural anti-inflammatory compounds that not only inhibit the secretion of NO from RAW 264.7 cells, but also inhibit triglyceride accumulation in 3T3-L1 adipocytes. This review describes the NO prpduction inhibitory activity or the TG accumulation inhibitory activity of the compounds obtained from 18 plants and a fungi that have been used as traditional medicines.

Fucoxanthin Suppresses Lipid Accumulation and ROS Production During Differentiation in 3T3-L1 Adipocytes.

Fucoxanthin, a pigment from the chloroplasts of marine brown algae, has a number of effects against obesity, diabetes, inflammation and cancer and provides cerebrovascular protection. In this study, we investigated the inhibitory effects of fucoxanthin on lipid accumulation and reactive oxygen species (ROS) production during adipogenesis. Treatment with fucoxanthin suppresses protein levels of the adipogenic transcription factors CCAAT/enhancer-binding protein alpha C/EBPα and peroxisome proliferator-activated receptor-γ and of their target protein, fatty acid binding protein 4. Lipogenesis-related enzymes, such as diglyceride acyltransferase 1 and lysophosphatidic acid acyltransferase-θ, were downregulated by fucoxanthin. The ROS-producing enzyme nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 4 (NOX4) and the NADPH-generating enzyme glucose-6-phosphate dehydrogenase also decreased following fucoxanthin treatment. The adipokine adiponectin and the ROS-scavenging enzymes superoxide dismutase 2, glutathione reductase and catalase were dose-dependently increased by fucoxanthin. Furthermore, lipolysis-related enzymes and superoxide dismutase 1 were slightly decreased, because of the suppression of lipid-generating factors and the cytosolic enzyme NOX4. To confirm these results, we investigated lipid accumulation and ROS production in zebrafish, where fucoxanthin suppressed lipid and triglyceride accumulation, as well as ROS production. Our data suggest that fucoxanthin inhibits lipid accumulation and ROS production by controlling adipogenic and lipogenic factors and ROS-regulating enzymes. Copyright © 2016 John Wiley & Sons, Ltd.

Arginase inhibition ameliorates adipose tissue inflammation in mice with diet-induced obesity.

This study examined whether oral administration of an arginase inhibitor regulates adipose tissue macrophage infiltration and inflammation in mice with high fat diet (HFD)-induced obesity. Male C57BL/6 mice (n = 30) were randomly assigned to control (CTL, n = 10), HFD only (n = 10), and HFD with arginase inhibitor N(ω)-hydroxy-nor-l-arginine (HFD with nor-NOHA, n = 10) groups. Plasma and mRNA levels of cytokines in epididymal adipose tissues (EAT), macrophage infiltration into EAT, and macrophage phenotype polarization were measured in the animals after 12 weeks. Additionally, the effects of nor-NOHA on adipose tissue macrophage infiltration and mRNA expression of cytokines were measured in co-cultured 3T3-L1 adipocytes and RAW 264.7 macrophages. Macrophage infiltration into the adipocytes was significantly suppressed by nor-NOHA treatment in adipocyte/macrophage co-culture system and mice with HFD-induced obesity. Pro-inflammatory cytokines, including monocyte chemoattractant protein-1 (MCP-1), tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6), were significantly downregulated, and the anti-inflammatory cytokine IL-10 was significantly upregulated in nor-NOHA-treated co-cultured cells. In the mice with HFD-induced obesity, plasma and mRNA levels of MCP-1 significantly reduced after supplementation with nor-NOHA. In addition, oral supplement of nor-NOHA modified M1/M2 phenotype ratio in the EAT. Oral supplementation of an arginase inhibitor, nor-NOHA, altered M1/M2 macrophage phenotype and macrophage infiltration into HFD-induced obese adipose tissue, thereby improved adipose tissue inflammatory response. These results may indicate that arginase inhibition ameliorates obesity-induced adipose tissue inflammation.

miR-20a regulates adipocyte differentiation by targeting lysine-specific demethylase 6b and transforming growth factor-ß signaling.

BACKGROUND: Several types of microRNAs (miRNAs) have recently been defined as important regulators in adipocyte differentiation, the role of other miRNAs in the processes and the mechanisms involved remain to be explored. METHODS: miR-20a expression was quantified in primary cultured marrow stromal cells and adipogenic cell lines after adipogenic treatment. Effects of miR-20a on adipocyte differentiation were studied following supplementing or depleting miR-20a in murine 3T3-L1 preadipocytes, ST2 stromal cells and C3H10T1/2 mesenchymal cells. Bioinformatics prediction of miRNA targets was performed, and potential targets of miR-20a were verified by using dual luciferase activity assays. Gain-of-function and loss-of-function studies were performed to examine the effects of the target genes on adipocyte differentiation. RESULTS: miR-20a was induced in primary cultured marrow stromal cells and established adipogenic lines after adipogenic treatment. Supplementing miR-20a activity suppressed the growth of 3T3-L1 preadipocytes and induced 3T3-L1, ST2 and C3H10T1/2 cells to differentiate into mature adipocytes, along with the induction of adipocyte-specific transcription factors peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer binding protein-α (C/EBPα), C/EBPß and the marker gene adipocyte protein 2 (aP2). Conversely, inhibition of the endogenous miR-20a repressed 3T3-L1, ST2 and C3H10T1/2 cells to fully differentiate. Transforming growth factor-ß receptor II (Tgfbr2) and lysine-specific demethylase 6b (Kdm6b) were shown to be direct targets of miR-20a. Supplementing miR-20a activity in ST2 reduced levels of KDM6B and TGFBR2 proteins, while suppression of endogenous miR-20a increased KDM6B and TGFBR2. While TGF-ß signaling is a well-documented inhibitor of adipogenesis, the effects of Kdm6b on adipocyte formation need to be clarified. We demonstrated that overexpression of Kdm6b inhibited, while knockdown of Kdm6b promoted the differentiation of the ST2 cells into mature adipocytes. CONCLUSION: The present work provides evidence that mouse miR-20a promotes adipocyte progenitor cells to differentiate and this function may depend upon its inhibitory effects on Kdm6b and TGF-ß signaling.

Aculeatin, a coumarin derived from Toddalia asiatica (L.) Lam., enhances differentiation and lipolysis of 3T3-L1 adipocytes.

Toddalia asiatica (L.) Lam. (T. asiatica) has been utilized traditionally for medicinal purposes such as the treatment of diabetes. Currently, the extract is considered to be a good source of anti-diabetic agents, but the active compounds have yet to be identified. In this study, we investigated the effects of fractionated T. asiatica extracts on the differentiation of 3T3-L1 preadipocytes and identified aculeatin as a potential active agent. When 3T3-L1 preadipocytes were treated with aculeatin isolated from T. asiatica in the presence of insulin, aculeatin increased cellular triglyceride levels and glycerol-3-phosphate dehydrogenase activity. This indicated that aculeatin could enhance the differentiation of preadipocytes into adipocytes. Further analyses using a DNA microarray and real-time quantitative reverse-transcription PCR showed an increase in the expression of peroxisome proliferator-activated receptor-γ target genes (Pparg, Ap2, Cd36, Glut4 and Adipoq) by aculeatin, suggesting that aculeatin enhances the differentiation of 3T3-L1 cells by modulating the expression of genes critical for adipogenesis. Interestingly, after treatment of differentiated adipocytes with aculeatin, glucose uptake and lipolysis were enhanced. Overall, our results suggested that aculeatin is an active compound in T. asiatica for enhancing both differentiation and lipolysis of adipocytes, which are useful for the treatment of lipid abnormalities as well as diabetes.

Apigenin isolated from Daphne genkwa Siebold et Zucc. inhibits 3T3-L1 preadipocyte differentiation through a modulation of mitotic clonal expansion.

AIMS: Obesity develops when energy intake chronically exceeds total energy expenditure. We sought to assess whether the flavonoid-rich fraction of crude extracts from Daphne genkwa Siebold et Zuccarini (GFF) might inhibit adipogenesis of 3T3-L1 cells. MAIN METHODS: Cell viability of 3T3-L1 preadipocytes was assessed by MTT assays, and lipid accumulation was measured by Oil Red O. Adipogenesis related factors were checked by Western blot analysis. Flow cytometry was used to analyze the mitotic cell cycle during the mitotic clonal expansion phase. KEY FINDINGS: Among five flavonoids isolated from GFF, only apigenin potently inhibited the differentiation of 3T3-L1 cells. Apigenin reduced CCAAT/enhancer binding protein (C/EBP) α and peroxisome proliferator-activated receptor γ levels. Apigenin-treated 3T3-L1 cells failed to undergo clonal expansion during the early phase of adipocyte differentiation. Apigenin arrested cell cycle progression at the G0/G1 phase. This effect was associated with a marked decrease in cyclin D1 and cyclin-dependent kinase 4 expression, with the concomitant and sustained expression of p27(Kip1). In addition, apigenin inhibited the DNA-binding activity of C/EBPß in differentiating 3T3-L1 cells by down-regulating the 35kDa isoform of C/EBPß (liver-enriched activating protein) and up-regulating the expression of two different sets of C/EBP inhibitors: C/EBP homologous protein and the phospho-liver-enriched inhibitory protein isoform of C/EBPß. SIGNIFICANCE: These findings suggest that apigenin can prevent 3T3-L1 preadipocyte differentiation by the inhibition of the mitotic clonal expansion and the adipogenesis related factors and upregulation of the expression of multiple C/EBPß inhibitors.

Adipocyte fatty acid-binding protein is released from adipocytes by a non-conventional mechanism.

Adipocyte fatty acid-binding protein (AFABP) is an adipokine, which induces insulin resistance. However, AFABP does not possess any secretion-directed signals and the mechanisms for AFABP release have not been thoroughly assessed so far. In the current study, mechanisms for AFABP secretion were elucidated in 3T3-L1 adipocytes in vitro in the presence or absence of hormonal stimulation, calcium ionophore and secretion inhibitors by cell fractionation experiments, immunoblotting and ELISAs. We demonstrate that AFABP secretion is upregulated during adipocyte differentiation. AFABP secretion is not influenced by treatment with protein secretion inhibitors that block vesicular traffic at the endoplasmic reticulum and the Golgi apparatus. AFABP is secreted partially by adipocyte-derived microvesicles (ADMs), an established mechanism for unconventional secretion from adipocytes. Both total and ADM-secreted AFABP are downregulated by insulin and upregulated by the calcium ionophore ionomycin. Furthermore, murine RAW 264.7 macrophages secrete AFABP and AFABP release from these cells is upregulated by lipopolysaccharide treatment. Taken together, these results suggest that AFABP is actively released by unconventional mechanisms and by ADMs from 3T3-L1 adipocytes. Furthermore, AFABP secretion from fat cells is regulated by insulin and intracellular calcium.

Protein tyrosine phosphatase 1B (PTP1B) inhibitors from Morinda citrifolia (Noni) and their insulin mimetic activity.

As part of our ongoing search for new antidiabetic agents from medicinal plants, we found that a methanol extract of Morinda citrifolia showed potential stimulatory effects on glucose uptake in 3T3-L1 adipocyte cells. Bioassay-guided fractionation of this active extract yielded two new lignans (1 and 2) and three new neolignans (9, 10, and 14), as well as 10 known compounds (3-8, 11-13, and 15). The absolute configurations of compounds 9, 10, and 14 were determined by ECD spectra analysis. Compounds 3, 6, 7, and 15 showed inhibitory effects on PTP1B enzyme with IC50 values of 21.86 ± 0.48, 15.01 ± 0.20, 16.82 ± 0.42, and 4.12 ± 0.09 µM, respectively. Furthermore, compounds 3, 6, 7, and 15 showed strong stimulatory effects on 2-NBDG uptake in 3T3-L1 adipocyte cells. This study indicated the potential of compounds 3, 6, 7, and 15 as lead molecules for antidiabetic agents.

Insulin sensitizing effect of 3 Indian medicinal plants: an in vitro study.

OBJECTIVE: Measurement of glucose uptake into peripheral tissue is an important mechanism to assess Insulin sensitivity. The present in vitro study was conducted to evaluate the Insulin sensitizing activity of Phyllanthus emblica (Pe), Tinospora cordifolia (Tc) and Curcuma longa (Cl) by assessing glucose uptake activity in a 3T3L1 adipocyte model. MATERIALS AND METHODS: The 3T3 L1 fibroblast cells were differentiated to adipocytes, using a cocktail of insulin, isobutyl-1-methylxanthine and dexamethazone. These adipocytes were initially treated with different concentrations of the selected plants following which 2-deoxy glucose uptake was estimated using a radioactive assay. The effects of plants on glucose uptake both in the presence and absence of insulin was evaluated and compared with pioglitazone, a known insulin sensitizer. RESULTS: Pe and Tc per se significantly stimulated glucose uptake in 3T3-L1 adipocytes in a dose dependent manner with maximal effect at higher concentrations (200 µg/ml). The effect of both Pe and Tc at 200 µg/ml was comparable to insulin and greater than pioglitazone. Cl per se stimulated glucose uptake with maximal effect at 50 µg/ml. However, this effect was lesser as compared to insulin with higher concentrations inhibiting glucose uptake. When combined with insulin, an antagonist effect was observed between Pe, Tc and insulin indicating a possible plant-drug interaction while Cl in combination with insulin showed an increase in the glucose uptake as compared to Cl alone. CONCLUSION: The results suggest that one of the mechanisms for the anti-diabetic effect of Pe, Cl and Tc may be through an insulin sensitizing effect (stimulation of glucose uptake into adipocytes). Further studies using other target sites viz. skeletal muscle and hepatocytes models and in an insulin resistant state would help substantiate this conclusion.

Unraveling the adipocyte inflammomodulatory pathways activated by North American ginseng.

BACKGROUND: North American (NA) ginseng (Panax quinquefolius) is a popular natural health product (NHP) that has been demonstrated to regulate immune function, inflammatory processes and response to stress and fatigue. Recent evidence suggests that various extracts of NA ginseng may have different bioactivities because of distinct profiles of ginsenosides and polysaccharides. To date, the bioactive role of ginseng on adipocytes remains relatively unexplored. OBJECTIVE: The goal of this work was to study the extract-specific bioactivity of NA ginseng on differentiated preadipocyte gene expression and adipocytokine secretion. METHODS: In vitro differentiated 3T3-L1 preadipocytes were treated with 25 and 50 µg ml of either crude ethanol (EtOH) or aqueous (AQ) NA ginseng extracts, or polysaccharide and ginsenoside extracts isolated from the AQ extract. Global gene expression was studied with microarrays and the resulting data were analyzed with functional pathway analysis. Adipocytokine secretion was also measured in media. RESULTS: Pathway analysis indicated that the AQ extract, and in particular the polysaccharide extract, triggered a global inflammomodulatory response in differentiated preadipocytes. Specifically, the expression of Il-6 (interleukin 6), Ccl5 (chemokine (C-C motif) ligand 5), Nfκb (nuclear factor-kappaB) and Tnfα (tumor necrosis factor alpha) was increased. These effects were also reflected at the protein level through the increased secretion of IL-6 and CCL5. No effect was seen with the EtOH extract or ginsenoside extract. Using a specific toll-like receptor 4 (TLR4) inhibitor reduced the upregulation of inflammatory gene expression, indicating the relevance of this pathway for the signaling capacity of NA ginseng polysaccharides. CONCLUSION: This work emphasizes the distinct bioactivities of different ginseng extracts on differentiated preadipocyte signaling pathways, and highlights the importance of TLR4 for mediating the inflammomodulatory role of ginseng polysaccharides.

Metabolic disease prevention and suppression of fat accumulation by Salacia reticulata.

In Ayurvedic medicine, Salacia reticulata is known to be useful against various metabolic diseases, including diabetes and obesity. In this study, we attempted to clarify the antiobesity mechanism and the safety of S. reticulata in vivo and in vitro. We gave ordinary MF feed, alone or mixed with S. reticulata (0.3 or 1.0%), to Tsumura Suzuki obesity diabetes (TSOD) mice (spontaneous obese type II diabetes model mice) and Tsumura Suzuki non-obese (TSNO) mice (the corresponding reference animals), ad libitum for 2 months. As compared with the TSNO control mice, the TSOD control mice became obese due to fat accumulation and developed various signs of metabolic diseases. The TSOD mouse group receiving S. reticulata showed the following effects: suppression of body weight increase and fat accumulation, alleviation of abnormal lipid metabolism and abnormal glucose tolerance, and suppression of intrahepatic fat accumulation. Also, S. reticulata prevented the mesenteric adipocyte hypertrophy recognized in TSOD mice. In the TSNO controls, the feed containing 1.0% S. reticulata exerted a suppressing effect on body weight increase and fat accumulation, but the feed containing 0.3% S. reticulata did not show any effect at all. In an in vitro experiment using mouse-derived adipocyte precursor 3T3-L1 cells, S. reticulata significantly suppressed fat accumulation in the differentiation induction phase and maturation phase. This suggested that the metabolic disease-preventing effects of S. reticulata, including the antiobesity effect, may involve suppression of differentiation and accumulation in the adipocytes.

Homoisoflavonoids from the fibrous roots of Polygonatum odoratum with glucose uptake-stimulatory activity in 3T3-L1 adipocytes.

The EtOAc-soluble fraction of a 90% MeOH extract of the fibrous roots of Polygonatum odoratum was found to potentiate insulin-stimulated glucose uptake in differentiated 3T3-L1 adipocytes. Bioassay-guided fractionation yielded nine homoisoflavonoids (1-9), four of which were new (1-4), together with an isoflavone glycoside (10) and a flavanone glycoside (11). The structures of new compounds were elucidated on the basis of extensive 1D and 2D NMR spectroscopy, and the absolute configurations were deduced by CD spectra. All 11 compounds showed effects of sensitizing adipocytes for insulin in a cell-based glucose uptake assay using 3T3-L1 adipocytes. The results indicate that homoisoflavonoids may be potential insulin sensitizers.

Chlorophyll a, an active anti-proliferative compound of Ludwigia octovalvis, activates the CD95 (APO-1/CD95) system and AMPK pathway in 3T3-L1 cells.

Ludwigia octovalvis is an aquatic plant widely distributed in Taiwan. It is traditionally used as a diuretic and is consumed as health drink. In this study, we evaluated the anti-proliferative activity of extracts and active constituent (chlorophyll a; CHL-a) of L. octovalvis in 3T3-L1 adipocytes; its mode of action on apoptosis was also investigated. Results showed that, among the different extracts and fractions, the ethylacetate layer (EAL) possessed the most potent anti-proliferative activity. Activity guided fractionation of the EAL obtained the bioactive constituent CHL-a (IC50: 24.10+/-0.83 nM). At concentrations 5-30 nM, CHL-a exhibited a dose-dependent accumulation of the Sub-G1 peak and caused cell cycle arrest at the G0/G1 phase. At 30 nM, it significantly reduced the cell viability, induced the appearance of DNA fragments, and enhanced the activation of caspase-3. Western blot data revealed that CHL-a decreased the level of Bcl-2, and increased the expression of CD95 (APO-1/CD95) and Bax. Furthermore, CHL-a up-regulated the AMPK and p-AMPK levels, and down-regulated the expression of PPAR-gamma. These results conclude that CHL-a possesses potent anti-proliferative activity, and its apoptotic effects on 3T3-L1 adipocytes are mediated through the activation of CD95 (APO-1/CD95) system and the AMPK signaling pathway.

Red mold rice prevents the development of obesity, dyslipidemia and hyperinsulinemia induced by high-fat diet.

OBJECTIVE: To investigate the influences of red mold rice (RMR) on obesity and related metabolic abnormalities. METHODS AND RESULTS: The 3T3-L1 cell line was used to examine the effects of RMR extracts on preadipocytes and on mature adipocytes. Both water and ethanol extracts of RMR had inhibitory effects on 3T3-L1 preadipocyte proliferation and differentiation. Water extracts of RMR enhanced the lipolysis activity in mature adipocytes, which negatively correlated with the triglyceride content within cells. RMR treatment did not affect heparin-releasable lipoprotein lipase activity in mature adipocytes. Furthermore, animal studies were carried out to explore the antiobesity effects of RMR. The control group of male Wistar rats were fed regular laboratory feed, whereas the other groups were fed the high-fat (HF) diet supplemented with lovastatin, rice or RMR (0.4 and 2%, w w(-1)). The relative caloric intakes of the control and HF groups were 3.34 and 4.85 kcal g(-1), respectively. After 6 weeks, rats treated with RMR at the 0.4 and 2% doses had lower weight gain and less fat pads mass accompanied with smaller fat cells than did the HF-diet rats. These effects probably resulted from an increase in the lipolysis activity of adipose tissue and a reduction in food/energy consumption. On the other hand, the RMR supplement significantly reduced serum total cholesterol, serum low-density lipoprotein (LDL) cholesterol, the ratio of LDL to high-density lipoprotein (HDL) cholesterol and serum insulin in the HF group. Moreover, the 2% RMR treatment significantly increased serum HDL cholesterol. CONCLUSION: This study reveals for the first time that RMR can prevent body fat accumulation and improve dyslipidemia. The antiobesity effects of RMR mainly derive from the lipolytic activity and mild antiappetite potency of RMR. In addition, extracts of RMR suppressed the proliferation and differentiation in 3T3-L1 preadipocytes, which might have contributed to the inhibition of new adipocyte formation or hyperplasia in adipose tissue.

[Effects of berberine on adiponectin mRNA expression in 3T3-L1 adipocyte].

OBJECTIVE: To explore the effects of berberine and insulin on adiponectin mRNA expression in 3T3-L1 adipocyte. METHOD: The 3T3-L1 adipocyte was treated with berberine and insulin for 48 hours, the level of adiponectin mRNA in 3T3-L1 adipocyte expression was determined with semiquantity RT-PCR using beta-actin as internal reference. RESULT: The level of adiponetin mRNA in 3T3-L1 adipocyte was increased after treated with berberine only (P < 0.05), the effect of berberine was inhibited by high concentration insulin (P < 0.05). CONCLUSION: In vitro, berberine increases the expression of adiponectin in 3T3-L1 adipocyte, insulin inhibits the effect of berberine.