3D-Cultured Adipose-Derived Stem Cell Spheres Using Calcium-Alginate Scaffolds for Osteoarthritis Treatment in a Mono-Iodoacetate-Induced Rat Model.

Osteoarthritis (OA) is a degenerative disease that causes pain, cartilage deformation, and joint inflammation. Mesenchymal stem cells (MSCs) are potential therapeutic agents for OA treatment. However, the 2D culture of MSCs could potentially affect their characteristics and functionality. In this study, calcium-alginate (Ca-Ag) scaffolds were prepared for human adipose-derived stem cell (hADSC) proliferation with a homemade functionally closed process bioreactor system; the feasibility of cultured hADSC spheres in heterologous stem cell therapy for OA treatment was then evaluated. hADSC spheres were collected from Ca-Ag scaffolds by removing calcium ions via ethylenediaminetetraacetic acid (EDTA) chelation. In this study, 2D-cultured individual hADSCs or hADSC spheres were evaluated for treatment efficacy in a monosodium iodoacetate (MIA)-induced OA rat model. The results of gait analysis and histological sectioning showed that hADSC spheres were more effective at relieving arthritis degeneration. The results of serological and blood element analyses of hADSC-treated rats indicated that the hADSC spheres were a safe treatment in vivo. This study demonstrates that hADSC spheres are a promising treatment for OA and can be applied to other stem cell therapies or regenerative medical treatments.

Muscle and Bone Impairment in Infantile Nephropathic Cystinosis: New Concepts.

Cystinosis Metabolic Bone Disease (CMBD) has emerged during the last decade as a well-recognized, long-term complication in patients suffering from infantile nephropathic cystinosis (INC), resulting in significant morbidity and impaired quality of life in teenagers and adults with INC. Its underlying pathophysiology is complex and multifactorial, associating complementary, albeit distinct entities, in addition to ordinary mineral and bone disorders observed in other types of chronic kidney disease. Amongst these long-term consequences are renal Fanconi syndrome, hypophosphatemic rickets, malnutrition, hormonal abnormalities, muscular impairment, and intrinsic cellular bone defects in bone cells, due to CTNS mutations. Recent research data in the field have demonstrated abnormal mineral regulation, intrinsic bone defects, cysteamine toxicity, muscle wasting and, likely interleukin-1-driven inflammation in the setting of CMBD. Here we summarize these new pathophysiological deregulations and discuss the crucial interplay between bone and muscle in INC. In future, vitamin D and/or biotherapies targeting the IL1ß pathway may improve muscle wasting and subsequently CMBD, but this remains to be proven.

6-gingerol ameliorates metabolic disorders by inhibiting hypertrophy and hyperplasia of adipocytes in high-fat-diet induced obese mice.

OBJECTIVES: Accumulating studies revealed that 6-gingerol, a compound extracted mainly from ginger, treats obesity by preventing hyperlipidemia in vivo induced by high-fat-diet (HFD). The present study intends to further evaluate the efficacy of 6-gingerol in the treatment of obesity and investigate its potential mechanism. METHODS: Obese mice were established by HFD induction. Bioinformatic analysis was used to predict the possible pathways enrolled by the application of 6-gingerol. Body weight and the levels of blood glucose and lipids were examined and analyzed for the evaluation of the therapeutic effect of 6-gingerol. The size and amounts as well as the status of adipocytes were determined by histological staining. The expression levels of related proteins in adipose tissue were assessed by immunohistochemical staining, immunofluorescent staining, and Western blot analysis. In addition, the expression levels of related mRNA were assessed by RT-qPCR. RESULTS: HFD induced obesity was significantly curbed by 6-gingerol treatment. Here inhibition mechanism of 6-gingerol is demonstrated on excessive hypertrophy and hyperplasia of adipocytes in white adipose tissue (WAT), which may be related to the regulation of adipocytokines, such as PPARγ, C/EBPα, FABP4 and adiponectin, and the TLR3/IL-6/JAK1/STAT3 axis. Moreover, 6-gingerol treatment suppressed the expressions of IL-1ß and CD68 in the liver and AKT/INSR/IRS-1 in epididymal WAT. CONCLUSION: The results suggested that 6-gingerol could alleviate metabolic inflammation in the liver and insulin resistance to treat obesity. The mechanism is mainly involved in the inhibition of excessive hypertrophy and hyperplasia of adipocytes.

Phospholipid Derivatives of Cinnamic Acid Restore Insulin Sensitivity in Insulin Resistance in 3T3-L1 Adipocytes.

BACKGROUND: Insulin resistance (IR) is a condition in which the physiological amount of insulin is insufficient to evoke a proper response of the cell, that is, glucose utilization. Metformin is the first choice for therapy, thanks to its glycemic efficacy and general tolerability. In addition, various natural compounds from plant extracts, spices, and essential oils have been shown to provide health benefits regarding insulin sensitivity. In the present study, we analyzed the effect of phospholipid derivatives of selected natural aromatic acids on insulin action and their potential use to overcome insulin resistance. METHODS: The 3T3-L1 fibroblasts were differentiated into mature adipocytes; next, insulin resistance was induced by palmitic acid (16:0). Cells were further cultured with phenophospholipids at appropriate concentrations. To assess insulin sensitivity, we measured the insulin-stimulated glucose uptake, using a glucose uptake test. RESULTS: We showed that cinnamic acid (CA) and 3-methoxycinnamic acid (3-OMe-CA) restored the proper insulin response. However, 1,2-dicinnamoyl-sn-glycero-3-phosphocholine (1,2-diCA-PC) and 1-cinnamoyl-2-palmitoyl-sn-glycero-3-phosphocholine (1-CA-2-PA-PC) improved insulin sensitivity in insulin-resistant adipocytes even stronger, exhibiting more beneficial effects. CONCLUSIONS: The binding of aromatic acids to phosphatidylcholine increases their beneficial effect on insulin sensitivity in adipocytes and expands their potential practical application as nutraceutical health-promoting agents.

Chrysanthemum morifolium Flower Extract Ameliorates Obesity-Induced Inflammation and Increases the Muscle Mitochondria Content and AMPK/SIRT1 Activities in Obese Rats.

Decreased energy expenditure and chronically positive energy balance contribute to the prevalence of obesity and associated metabolic dysfunctions, such as dyslipidemia, hepatic fat accumulation, inflammation, and muscle mitochondrial defects. We investigated the effects of Chrysanthemum morifolium Ramat flower extract (CE) on obesity-induced inflammation and muscle mitochondria changes. Sprague-Dawley rats were randomly divided into four groups and fed either a normal diet, 45% high-fat diet (HF), HF containing 0.2% CE, or 0.4% CE for 13 weeks. CE alleviated HF-increased adipose tissue mass and size, dyslipidemia, hepatic fat deposition, and systematic inflammation, and increased energy expenditure. CE significantly decreased gene expression involved in adipogenesis, pro-inflammation, and the M1 macrophage phenotype, as well as glycerol-3-phosphate dehydrogenase (GPDH) and nuclear factor-kappa B (NF-kB) activities in epididymal adipose tissue. Moreover, CE supplementation improved hepatic fat accumulation and modulated gene expression related to fat synthesis and oxidation with an increase in adenosine monophosphate-activated protein kinase (AMPK) activity in the liver. Furthermore, CE increased muscle mitochondrial size, mitochondrial DNA (mtDNA) content, and gene expression related to mitochondrial biogenesis and function, including sirtuin 1 (SIRT1), peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), and PGC-1α-target genes, along with AMPK-SIRT1 activities in the skeletal muscle. These results suggest that CE attenuates obesity-associated inflammation by modulating the muscle AMPK-SIRT1 pathway.

Effect of Walnut Meal Peptides on Hyperlipidemia and Hepatic Lipid Metabolism in Rats Fed a High-Fat Diet.

As a natural active substance that can effectively improve blood lipid balance in the body, hypolipidemic active peptides have attracted the attention of scholars. In this study, the effect of walnut meal peptides (WMP) on lipid metabolism was investigated in rats fed a high-fat diet (HFD). The experimental results show that feeding walnut meal peptides counteracted the high-fat diet-induced increase in body, liver and epididymal fat weight, and reduce the serum concentrations of total cholesterol, triglycerides, and LDL-cholesterol and hepatic cholesterol and triglyceride content. Walnut meal peptides also resulted in increased HDL-cholesterol while reducing the atherosclerosis index (AI). Additionally, the stained pathological sections of the liver showed that the walnut meal peptides reduced hepatic steatosis and damage caused by HFD. Furthermore, walnut meal peptide supplementation was associated with normalization of elevated apolipoprotein (Apo)-B and reduced Apo-A1 induced by the high-fat diet and with favorable changes in the expression of genes related to lipid metabolism (LCAT, CYP7A1, HMGR, FAS). The results indicate that walnut meal peptides can effectively prevent the harmful effects of a high-fat diet on body weight, lipid metabolism and liver fat content in rats, and provide, and provide a reference for the further development of walnut meal functional foods.

Viburnum stellato-tomentosum Extract Suppresses Obesity and Hyperglycemia through Regulation of Lipid Metabolism in High-Fat Diet-Fed Mice.

The potential biological activities of Viburnum stellato-tomentosum (VS), a plant mainly found in Costa Rica, have yet to be reported. Supplementation of VS extract for 17 weeks significantly decreased body weight gain, fat weight, fasting glucose, insulin, homeostasis model assessment of insulin resistance (HOMA-IR), and triglyceride levels in high-fat diet (HFD)-fed C57BL/6J mice. The molecular mechanisms underlying the anti-obesity and glucose-lowering effects of VS extract were investigated. VS extract suppressed adipocyte hypertrophy by regulating lipogenesis-related CCAAT/enhancer-binding protein α (C/EBPα) and insulin sensitivity-related peroxisome proliferator-activated receptor γ (Pparg) expression in adipose tissue (AT) and hepatic steatosis by inhibiting C/EBPα and lipid transport-related fatty acid binding protein 4 (FABP4) expression. VS extract enhanced muscular fatty acid ß-oxidation-related AMP-activated protein kinase (AMPK) and PPARα expression with increasing Pparg levels. Furthermore, VS extract contained a much higher content of amentoflavone (AMF) (29.4 mg/g extract) compared to that in other Viburnum species. AMF administration decreased Cebpa and Fabp4 levels in the AT and liver, as well as improved insulin signaling-related insulin receptor substrate 1 (Irs1) and glucose transporter 1 (Glut1) levels in the muscle of HFD-fed mice. This study elucidated the in vivo molecular mechanisms of AMF for the first time. Therefore, VS extract effectively diminished obesity and hyperglycemia by suppressing C/EBPα-mediated lipogenesis in the AT and liver, enhancing PPARα-mediated fatty acid ß-oxidation in muscle, and PPARγ-mediated insulin sensitivity in AT and muscle.

Transplantation of encapsulated human Leydig-like cells: A novel option for the treatment of testosterone deficiency.

Previous studies have demonstrated that the transplantation of alginate-poly-ʟ-lysine-alginate (APA)-encapsulated rat Leydig cells (LCs) provides a promising approach for treating testosterone deficiency (TD). Nevertheless, LCs have a limited capacity to proliferate, limiting the efficacy of LC transplantation therapy. Here, we established an efficient differentiation system to obtain functional Leydig-like cells (LLCs) from human stem Leydig cells (hSLCs). Then we injected APA-encapsulated LLCs into the abdominal cavities of castrated mice without an immunosuppressor. The APA-encapsulated cells survived and partially restored testosterone production for 90 days in vivo. More importantly, the transplantation of encapsulated LLCs ameliorated the symptoms of TD, such as fat accumulation, muscle atrophy and adipocyte accumulation in bone marrow. Overall, these results suggest that the transplantation of encapsulated LLCs is a promising new method for testosterone supplementation with potential clinical applications in TD.

HM-chromanone inhibits adipogenesis by regulating adipogenic transcription factors and AMPK in 3T3-L1 adipocytes.

Portulaca oleracea L. is used as a folk medicine in many countries because of its wide range of pharmacological effects. HM-chromanone, isolated from P. oleracea using bioassay-guided fractionation and HPLC, belongs to the homoisoflavonoid group and has been shown to exert several biological effects. In this study, we evaluated whether HM-chromanone inhibits adipogenesis by regulating adipogenic transcription factors in 3T3-L1 adipocytes. The results showed that HM-chromanone suppresses adipocyte differentiation and adipogenesis in a dose-dependent manner in 3T3-L1 adipocytes. The HM-chromanone-treated adipocytes exhibited lower triglyceride accumulation and leptin secretion, and higher glycerol and adiponectin secretion than the control adipocytes. Microscopic observation using oil red O staining revealed a dose-dependent reduction in the number of lipid droplets in the HM-chromanone-treated adipocytes compared to the control group. HM-chromanone significantly down-regulated the protein expression of major adipogenic transcription factors sterol regulatory element binding protein-1c (SREBP-1c), peroxisome proliferator-activated receptor γ (PPARγ), and CCAAT/enhancer binding protein α (C/EBPα) and markedly inhibited several key adipogenic enzymes including fatty acid synthase (FAS) and acetyl-CoA carboxylase (ACC). In addition, adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) were both more activated in the HM-chromanone-treated adipocytes than in the control adipocytes. HM-chromanone also promoted the phosphorylation of 5' Adenosine monophosphate-activated protein kinase (AMPK), which inhibits adipogenesis through the regulation of adipogenic transcription factors. These results suggest that HM-chromanone may be an effective anti-adipogenesis agent that functions via the suppression of adipogenic transcription factors and the activation of AMPK.

Comparison of main chemical composition of Plantago asiatica L. and P. depressa Willd. seed extracts and their anti-obesity effects in high-fat diet-induced obese mice.

BACKGROUND: Nowadays, the pharmacological effects of Plantaginis semen was getting more and more attention because of the great effect of treating diuresis, hypertension, hyperlipidemia, and hyperglycemia. According to the Chinese Pharmacopoeia, Plantaginis semen is the seed of Plantago asiatica L. or P. depressa Willd. This was verified by examining chemical composition differences in a preliminary experiment, predicting their differences in pharmacology. PURPOSE: In this study, we aimed to compared the the differences in main components and anti-obesity effects of Plantago asiatica L. seed extract (PASE) and P. depressa Willd. seed extract (PDSE). STUDY DESIGN AND METHODS: The ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) analysis was used to characterize and compare the differences chemical constituents of PASE and PDSE. The difference therapeutic effects between PASE and PDSE on obesity and associated metabolic disorders was investigated by high-fat (HF) diet induced mice model. RESULTS: The fingerprint of Plantaginis semen were established by screening and identified 15 main components, including iridoids, phenethanol glycosides, flavonoids, guanidines, and fatty acids. Pentahydroxy flavanone was observed only in PDSE but not in PASE. The quantitative analysis results indicated that the main bioactive components in PASE were geniposidic acid and acteoside; their concentrations were three times higher in PASE than in PDSE. In anti-obesity effects, the result show the levels of fasting blood glucose were improved in both PASE and PDSE when compared with the HF group, while the PASE is show a significant effect then the PDSE group and improved the glucose tolerance but not in PDSE. The results also displayed that the Plantaginis semen did not modify food intake or body weight but decreased abdominal white/brown adipocyte size, serum total cholesterol (TC), triglyceride (TG), low density lipoprotein cholesterol (LDL-c), hepatic TG and TC, fecal TG and TC concentrations when compared with the HF group. Among these indicators, serum TG, liver TG, fecal TC and TG levels were significantly improved in PASE compared with PDSE. The results indicated that PASE treatment more effectively improved lipid and glucose metabolism in HF diet-induced obese mice than did PDSE. CONCLUSION: As Plantaginis semen sources, P. asiatica L. seeds demonstrated more bioactive components and favorable metabolic disorder treatment outcomes than did P. depressa Willd. seeds.

Fisetin Alleviates Hepatic and Adipocyte Fibrosis and Insulin Resistance in Diet-Induced Obese Mice.

The present study aimed to investigate the protective role of the flavonoid fisetin (FI) on inflammation-mediated metabolic diseases, especially tissue fibrosis and insulin resistance (IR) in high-fat diet (HFD)-induced obese mice. C57BL/6J mice were fed with normal-fat diet, HFD (40 kcal% fat), or HFD +0.02% (w/w) FI for 16 weeks. Dietary FI supplementation improved hepatic steatosis by restricting lipogenesis, while promoting lipolysis in the liver. FI also prevented adiposity via an increase in the expression of genes involved in FA oxidation and a decrease in the expression of genes involved in lipogenesis in white adipose tissue. In addition, FI increased brown adipose tissue (BAT) and skeletal muscle weights, thermogenic gene mRNA expression in BAT, and tricarboxylic acid cycle-related gene expression in skeletal muscle, which may be linked to the prevention of nonalcoholic fatty liver disease as well as adiposity. Moreover, FI supplementation decreased excessive reactive oxygen species production by increasing paraoxonase activity, adipokine dysregulation, proinflammatory cytokine production, and extracellular matrix amassment in the liver. FI supplementation ameliorated IR, in part, by normalizing pancreatic islet dysfunction, and it declined hepatic gluconeogenesis and proinflammatory responses. Taken together, the present findings indicate that FI can protect against HFD-induced inflammation-mediated disorders, including fibrosis and IR.

(-)-Epicatechin protects thoracic aortic perivascular adipose tissue from whitening in high-fat fed mice.

High adipose tissue (AT) accumulation in the body increases the risk for many metabolic and chronic diseases. This work investigated the capacity of the flavonoid (-)-epicatechin to prevent undesirable modifications of AT in mice fed a high-fat diet. Studies were focused on thoracic aorta perivascular AT (taPVAT), which is involved in the control of blood vessel tone, among other functions. Male C57BL/6J mice were fed for 15 weeks a high-fat diet with or without added (-)-epicatechin (20 mg per kg body weight per d). In high-fat diet fed mice, (-)-epicatechin supplementation: (i) prevented the expansion of taPVAT, (ii) attenuated the whitening of taPVAT (according to the adipocyte morphology, diameter, and uncoupling-protein 1 (UCP-1) levels) and (iii) blunted the increase in plasma glucose and cholesterol. The observed taPVAT modifications were not associated with alterations in the aorta wall thickness, aorta tumor necrosis factor-alpha (TNF-α) and NADPH-oxidase 2 (NOX2) expression, and endothelial nitric oxide synthase (eNOS) phosphorylation levels. In summary, our results indicate (-)-epicatechin as a relevant bioactive protecting from the slow and silent development of metabolic and chronic diseases as they are associated with excessive fat intake.

Antiobesity Effects of Gentiana lutea Extract on 3T3-L1 Preadipocytes and a High-Fat Diet-Induced Mouse Model.

Obesity is one of the most common metabolic diseases resulting in metabolic syndrome. In this study, we investigated the antiobesity effect of Gentiana lutea L. (GL) extract on 3T3-L1 preadipocytes and a high-fat-diet (HFD)-induced mouse model. For the induction of preadipocytes into adipocytes, 3T3-L1 cells were induced by treatment with 0.5 mM 3-isobutyl-1-methylxanthine, 1 mM dexamethasone, and 1 µg/mL insulin. Adipogenesis was assessed based on the messenger ribonucleic acid expression of adipogenic-inducing genes (adiponectin (Adipoq), CCAAT/enhancer-binding protein alpha (Cebpa), and glucose transporter type 4 (Slc2a4)) and lipid accumulation in the differentiated adipocytes was visualized by Oil Red O staining. In vivo, obese mice were induced with HFD and coadministered with 100 or 200 mg/kg/day of GL extract for 12 weeks. GL extract treatment inhibited adipocyte differentiation by downregulating the expression of adipogenic-related genes in 3T3-L1 cells. In the obese mouse model, GL extract prevented HFD-induced weight gain, fatty hepatocyte deposition, and adipocyte size by decreasing the secretion of leptin and insulin. In conclusion, GL extract shows antiobesity effects in vitro and in vivo, suggesting that this extract can be beneficial in the prevention of obesity.

Juzentaihoto extract suppresses adipocyte hypertrophy and improves hyperglycemia in KKAy mice.

Juzentaihoto is a herbal medicine with reported anti-inflammatory effects, and it is predicted to improve inflammation and insulin sensitivity within obesity. In the present study, juzentaihoto hot water extract (JTT) was administered to obese type 2 diabetic model mice (KKAy) for 56 days. In addition, the effects of JTT on the adipose tissue, glucose metabolism, and blood lipids were evaluated for examining its impact on insulin sensitivity and obesity. As a result of JTT administration, KKAy mice exhibited suppressed adipocyte hypertrophy, decreased the mRNA levels of tumor necrosis factor α, and increased the mRNA levels of adiponectin in epididymal fat tissue. In addition, fasting blood glucose levels, blood triglyceride, and total cholesterol decreased. In summary, these data indicated that JTT administration suppressed the production of inflammatory cytokines and increased adiponectin levels in the adipose tissue. Therefore, with improved insulin sensitivity, blood glucose, and lipid decreased.

Obesity and NRF2-mediated cytoprotection: Where is the missing link?

The expanding dimensions of the global health crisis of overweight population has defined the term "globesity". Among the most common pathological conditions connected with excessive adiposity are hyperglycemia, insulin resistance, dyslipidemia and hypertension which result in chronic non-communicable diseases (NCD) such as metabolic syndrome (MetS), type 2 diabetes (T2D), and nonalchoholic steatohepatitis (NASH). The contribution of inflammatory-immune reactions in obesity and its related co-morbidities is unequivocal. Increased levels of free fatty acids (FFA), reactive oxygen species (ROS) and reactive nitrogen species (RNS) overloads the homeostatic system resulting in pro-inflammatory adipokines secretion, immune-activation and chronic inflammation in obesity. The cellular mechanisms of defense against oxidative stress are orchestrated by the transcription factor nuclear factor erythroid 2 p45-related factor 2 (NRF2). Excessive oxidative stress in the cell activates NRF2 which upregulates genes encoding major cytoprotective enzymes such as NAD(P)H:quinone oxidoreductase 1 (NQO1), heme oxygenase 1 (HO1), and glutathione S-transferases (GST). The present review aims to clarify the interconnections between chronic inflammation, oxidative overload and NRF2-mediated cytoprotection as potential therapeutic approach in obesity.

The herbal extract ALS-L1023 from Melissa officinalis alleviates visceral obesity and insulin resistance in obese female C57BL/6J mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Melissa officinalis L. (Labiatae; lemon balm) has traditionally been used as a medicinal herb to treat stress, anxiety, and insomnia. Current reports suggest that not only chronic stress stimulates angiogenesis, but angiogenesis also regulates adipogenesis and obesity. Because the herbal extract ALS-L1023 from Melissa officinalis inhibits angiogenesis, we hypothesized that ALS-L1023 could suppress visceral obesity and insulin resistance in obese female C57BL/6J mice, a mouse model of obese premenopausal women. MATERIALS AND METHODS: The mice were grouped and fed for 16 weeks as follows: 1) low-fat diet (LFD), 2) high-fat diet (HFD), or 3) HFD supplemented with 0.4 or 0.8% ALS-L1023. Variables and determinants of visceral obesity, insulin resistance, and pancreatic dysfunction were then assessed via blood analysis, histology, immunohistochemistry, and real-time polymerase chain reaction. RESULTS: ALS-L1023 decreased weight gain, visceral adipocyte size, and serum lipid levels in HFD-fed obese mice. ALS-L1023 also normalized hyperglycemia and hyperinsulinemia and concomitantly reduced blood glucose levels during oral glucose tolerance tests. The pancreatic islet size and insulin-positive ß-cell area were significantly reduced in ALS-L1023-treated mice compared with untreated obese controls, reaching a level similar to that of LFD-fed lean mice. ALS-L1023 suppressed pancreatic lipid accumulation, infiltration of inflammatory cells, and collagen levels. ALS-L1023 treatment altered the pancreatic expression of genes involved in steatosis, inflammation, and fibrosis. CONCLUSIONS: Our findings indicate that the herbal extract ALS-L1023 from Melissa officinalis not only inhibits visceral obesity, but also attenuates the increased fasting blood glucose, impaired glucose tolerance, and pancreatic dysfunction seen in female obese mice. These results suggest that ALS-L1023 may be effective in the prevention of visceral obesity and insulin resistance in obese premenopausal women.

Paeonol, an Ingredient of Kamishoyosan, Reduces Intracellular Lipid Accumulation by Inhibiting Glucocorticoid Receptor Activity in 3T3-L1 Cells.

Excessive triglyceride accumulation in lipid-metabolizing tissues is associated with an increased risk of a variety of metabolic diseases. Kamishoyosan (KSS) is a Kampo composed of 10 constituent herbs, and contains moutan cortex (MC) and paeonol (PN) as the major ingredient of MC. Here, we demonstrate the molecular mechanism underlying the effect of KSS on the differentiation of mouse preadipocytes (3T3-L1 cells). KSS inhibited the accumulation of triglycerides in a dose-dependent manner in 3T3-L1 cells that were induced to differentiate into adipocytes. We also found that MC and PN were responsible for the anti-adipogenetic effect of KSS and significantly suppressed the expression of CCAAT/enhancer-binding proteins-δ (C/EBP-δ) mRNA 3 days after the induction of differentiation. Thus, PN may contribute to the anti-adipogenetic property of MC in 3T3-L1 cells. In addition, PN inhibited dexamethasone (Dex)-induced glucocorticoid receptor (GR) promoter activity. Taken together, these results suggest that PN suppresses C/EBP-δ expression by inhibiting Dex-induced GR promoter activity at the early stage of differentiation and, consequently, delays differentiation into mature adipocytes. Our results suggest that the habitual intake of Kampo-containing PN contributes to the prevention of the onset of metabolic diseases by decreasing the excessive accumulation of triglycerides in lipid-metabolizing tissues.

Adipocyte-derived extracellular vesicles modulate appetite and weight through mTOR signalling in the hypothalamus.

AIM: Type 2 diabetes and obesity are diseases related to surplus energy in the body. Abnormal interaction between the hypothalamus and adipose tissues is a key trigger of energy metabolism dysfunction. Extracellular vesicles (EVs) regulate intercellular communication by transporting intracellular cargo to recipient cells thereby altering the function of recipient cells. This study aimed to evaluate whether adipocyte-derived EVs can act on hypothalamic neurons to modulate energy intake and to identify the EV-associated non-coding RNAs. METHODS: Confocal imaging was used to trace the uptake of labelled adipocyte-derived exosomes by hypothalamic anorexigenic POMC neurons. The effects of adipocyte-derived EVs on the mammalian target of rapamycin (mTOR) signalling pathway in POMC neurons were evaluated based on mRNA and protein expression in vitro using quantitative real-time PCR and western blotting. In addition, adipocyte-derived EVs were injected into recipient mice, and changes in mice body weight and daily food intake were monitored. The biological effects of the EV-associated MALAT1 on POMC neurons were explored. RESULTS: Adipocyte-derived EVs were successfully transferred into POMC neurons in vitro. Results showed that adipocytes of obese mice secreted MALAT1-containing EVs, which increased appetite and weight when administered to lean mice. Conversely, adipocyte-derived EVs from lean mice decreased food intake and weight when administered to obese mice. CONCLUSION: Adipocyte-derived EVs play important roles in mediating the interaction between adipocytes and hypothalamic neurons. Adipocyte-derived EVs can regulate POMC expression through the hypothalamic mTOR signalling in vivo and in vitro, thereby affecting body energy intake.

Cardamonin suppresses lipogenesis by activating protein kinase A-mediated browning of 3T3-L1 cells.

BACKGROUND: Obesity develops when dietary energy intake exceeds energy expenditure, and can be associated with metabolic syndrome. Recent studies have shown that dietary phytochemicals can promote energy expenditure by inducing the browning of white adipose tissue (WAT). PURPOSE: This study investigated whether cardamonin induces the browning of 3T3-L1 adipocytes through the activation of protein kinase A (PKA). METHODS: Anti-obesity potential of cardamonin was evaluated in 3T3-L1 adipocytes. Adipocyte-specific genes were observed using western blot, qPCR analysis and immunocytochemistry. RESULTS: Cardamonin treatment inhibited lipid droplet accumulation and reduced the expression of the adipogenic proteins C/EBPα and FABP4, and the lipogenic proteins LPAATθ, lipin 1, DGAT1, SREBP1, and FAS. Cardamonin also induced the expression of the browning marker genes PRDM16, PGC1α, and UCP1 at the mRNA and protein levels, and induced mRNA expression of CD137, a key marker of beige adipocytes. It also increased the expression of the ß-oxidation genes CPT1 and PPARα at the mRNA and protein levels. In addition, cardamonin increased PKA phosphorylation and the mRNA and protein expression of the downstream lipolytic enzymes adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL). CONCLUSION: Our findings demonstrate novel effects of cardamonin to stimulate adipocyte browning, suppress lipogenesis, and promote lipolysis, implying it may have potential as an anti-obesity agent.

Pyridocarbazole alkaloids from Ochrosia borbonica: lipid-lowering agents inhibit the cell proliferation and adipogenesis of 3T3-L1 adipocyte via intercalating into supercoiled DNA.

Bioassay-guided fractionation of an ethanolic extract of Ochrosia borbonica led to the isolation of two known pyridocarbazole alkaloids, ellipticine (1) and 9-methoxyellipticine (2), and six known monoterpenoid indole alkaloids (3-8). Lipid-lowering assay in 3T3-L1 cell model revealed that 1 and 2 could significantly inhibit the lipid droplet formation (EC50 = 0.41 and 0.92 µmol·L-1, respectively) and lower triglyceride levels by 50%-60% at the concentration of 1 µmol·L-1, being more potent than the positive drug luteolin (EC50 = 2.63 µmol·L-1). A mechanistic study indicated that 1 and 2 could intercalate into supercoiled DNA, which consequently inhibited the mitotic clonal expansion of 3T3-L1 cells at the early differentiation phase, leading to the retardance of following adipogenesis and lipogenesis. These findings suggest that 1 and 2 may serve as promising leads for further development of anti-obesity drugs.