Integrated omics analysis unraveled the microbiome-mediated effects of Yijin-Tang on hepatosteatosis and insulin resistance in obese mouse

BackgroundGut microbiota play important roles in insulin homeostasis and the pathogenesis of non-alcoholic fatty liver diseases (NAFLD). Yijin-Tang (YJT), a traditional Korean and Chinese medicine, is used in the treatment of gastrointestinal diseases and obesity-related disorders such as insulin resistance (IR) and NAFLD.PurposeOur aim was to identify the microbiome-mediated effects of YJT on IR and associated NAFLD by integrating metagenomics and hepatic lipid profile.MethodsC57BL/6J mice were fed a normal chow diet (NC) or high-fat/high-cholesterol (HFHC) diet with or without YJT treatment. Hepatic lipid profiles were analyzed using liquid chromatography/mass spectrometry, and the composition of gut microbiota was investigated using 16S rRNA sequencing. Then, hepatic lipid profiles, gut microbiome, and inflammatory marker data were integrated using multivariate analysis and bioinformatics tools.ResultsYJT improved NAFLD, and 39 hepatic lipid metabolites were altered by YJT in a dose-dependent manner. YJT also altered the gut microbiome composition in HFHC-fed mice. In particular, Faecalibaculum rodentium and Bacteroides acidifaciens were altered by YJT in a dose-dependent manner. Also, we found significant correlation among hepatic phosphatidylglycerol metabolites, F. rodentium, and γδ-T cells. Moreover, interleukin (IL)-17, which is secreted by the γδ-T cell when it recognizes lipid antigens, were elevated in HFHC mice and decreased by YJT treatment. In addition, YJT increased the relative abundance of B. acidifaciens in NC or HFHC-fed mice, which is a gut microbiota that mediates anti-obesity and anti-diabetic effects by modulating the gut environment. We also confirmed that YJT ameliorated the gut tight junctions and increased short chain fatty acid (SCFA) levels in the intestine, which resulted in improved IR.ConclusionThese data demonstrated that gut microbiome and hepatic lipid profiles are regulated by YJT, which improved the IR and NAFLD in mice with diet-induced obesity.     

Lipid-Overloaded Enlarged Adipocytes Provoke Insulin Resistance Independent of Inflammation

In obesity, adipocyte hypertrophy and proinflammatory responses are closely associated with the development of insulin resistance in adipose tissue. However, it is largely unknown whether adipocyte hypertrophy per se might be sufficient to provoke insulin resistance in obese adipose tissue. Here, we demonstrate that lipid-overloaded hypertrophic adipocytes are insulin resistant independent of adipocyte inflammation. Treatment with saturated or monounsaturated fatty acids resulted in adipocyte hypertrophy, but proinflammatory responses were observed only in adipocytes treated with saturated fatty acids. Regardless of adipocyte inflammation, hypertrophic adipocytes with large and unilocular lipid droplets exhibited impaired insulin-dependent glucose uptake, associated with defects in GLUT4 trafficking to the plasma membrane. Moreover, Toll-like receptor 4 mutant mice (C3H/HeJ) with high-fat-diet-induced obesity were not protected against insulin resistance, although they were resistant to adipose tissue inflammation. Together, our in vitro and in vivo data suggest that adipocyte hypertrophy alone may be crucial in causing insulin resistance in obesity.     

Food-derived regulatory factors against obesity and metabolic syndrome

Abstract

Obesity is a key factor in metabolic syndrome. The study of metabolic syndrome focuses on the anti-weight gain properties of physiological mechanisms and food components. Abnormal energy metabolism is a major risk factor of metabolic syndrome. Chronic inflammation is a feature of obesity; cytokines from hypertrophied adipocytes cause inflammation in both adipose tissue and blood vessels, resulting in symptoms of metabolic syndrome. Tumor necrosis factor-α causes insulin resistance in adipocytes and regression of brown adipocytes, resulting in abnormal energy metabolism. Functional foods can serve as a strategy for prevention and treatment of obesity linked with metabolic processes in white and brown adipose tissues. Diet-induced thermogenesis caused by certain food components stimulates burning of stored fat within adipose tissues. A mechanistic understanding of dietary thermogenesis via the sympathetic nerve system will prove valuable for the development of precise strategies for the practical prevention of metabolic syndrome.     

Medium-chain fatty acids enhance expression and histone acetylation of genes related to lipid metabolism in insulin-resistant adipocytes

BackgroundThe expressions of genes related to lipid metabolism are decreased in adipocytes with insulin resistance. In this study, we examined the effects of fatty acids on the reduced expressions and histone acetylation of lipid metabolism-related genes in 3T3-L1 adipocytes treated with insulin resistance induced by tumor necrosis factor (TNF)-α.MethodsShort-, medium-, and long-chain fatty acid were co-administered with TNF-α in 3T3-L1 adipocytes. Then, mRNA expressions and histone acetylation of genes involved in lipid metabolism were determined using mRNA microarrays, qRT-PCR, and chromatin immunoprecipitation assays.ResultsWe found in microarray and subsequent qRT-PCR analyses that the expression levels of several lipid metabolism-related genes, including Gpd1, Cidec, and Cyp4b1, were reduced by TNF-α treatment and restored by co-treatment with a short-chain fatty acid (C4: butyric acid) and medium-chain fatty acids (C8: caprylic acid and C10: capric acid). The pathway analysis of the microarray showed that capric acid enhanced mRNA levels of genes in the PPAR signaling pathway and adipogenesis genes in the TNF-α-treated adipocytes. Histone acetylation around Cidec and Gpd1 genes were also reduced by TNF-α treatment and recovered by co-administration with short- and medium-chain fatty acids.General significanceMedium- and short-chain fatty acids induce the expressions of Cidec and Gpd1, which are lipid metabolism-related genes in insulin-resistant adipocytes, by promoting histone acetylation around these genes.     

Bilobalide attenuates hypoxia induced oxidative stress,inflammation, and mitochondrial dysfunctions in 3T3-L1 adipocytes via its antioxidant potential

Abstract

Excessive expansion of white adipose tissue leads to hypoxia which is considered as a key factor responsible for adipose tissue dysfunction in obesity. Hypoxia induces inflammation, insulin resistance, and other obesity related complications. So the hypoxia-signalling pathway is expected to provide a new target for the treatment of obesity-associated complications. Inhibition or downregulation of the HIF-1 pathway could be an effective target for the treatment of obesity related hypoxia. In the present study, we evaluated the effect of hypoxia on functions of 3T3-L1 adipocytes emphasising on oxidative stress, antioxidant status, inflammation and mitochondrial functions. We have also evaluated the protective role of bilobalide, a bioactive from Gingko biloba, on hypoxia induced alterations. The results revealed that hypoxia significantly altered all the vital parameters of adipocyte biology like HIF-1α expression (103.47% ↑), lactate and glycerol release (184.34% and 69.1% ↑, respectively), reactive oxygen species (ROS) production (432.53% ↑), lipid and protein oxidation (376.6% and 566.6% ↑, respectively), reduction in antioxidant enzymes (superoxide dismutase and catalase) status, secretion of inflammatory markers (TNF-α, IL-6, IL-1β and IFN-γ) and mitochondrial functions (mitochondrial mass, membrane potential, permeability transition pore integrity, superoxide generation). Bilobalide significantly protected adipocytes from adverse effects of hypoxia in a dose-dependent manner by attenuating oxidative stress, inflammation and protecting mitochondria. Acriflavine (HIF-1 inhibitor) was used as positive control. On the basis of this study, a detailed investigation is needed to delineate the mechanism of action of bilobalide to develop it as therapeutic target for obesity.     

Oxidative stress in adipose tissue as a primary link in pathogenesis of insulin resistance

Obesity is a leading risk factor of diabetes mellitus type 2 (DM2), impairments of lipid metabolism and cardiovascular diseases. Dysfunctions of the accumulating weight of the visceral fat are primarily linked to pathogenesis of systemic insulin resistance. The review considers modern viewpoints on biochemical mechanisms underlying formation of oxidative stress in adipocytes at obesity, as one of key elements responsible for impairments of their metabolism triggering formation of systemic insulin resistance.     

XOMA 052, an anti‐IL‐1β monoclonal antibody,prevents IL‐1β‐mediated insulin resistance in 3T3‐L1 adipocytes

Objective:

Interleukin‐1β (IL‐1β) has recently been implicated as a major cytokine that is involved in the pancreatic islet inflammation of type 2 diabetes mellitus. This inflammation impairs insulin secretion by inducing beta‐cell apoptosis. Recent evidence has suggested that in obesity‐induced inflammation, IL‐1β plays a key role in causing insulin resistance in peripheral tissues.

Design and Methods:

To further investigate the pathophysiological role of IL‐1β in causing insulin resistance, the inhibitory effects of IL‐1β on several insulin‐dependent metabolic processes in vitro has been neutralized by XOMA 052. The role IL‐1β plays in insulin resistance in adipose tissue was assessed using differentiated 3T3‐L1 adipocytes and several parameters involved in insulin signaling and lipid metabolism were examined.

Results and Conclusion:

IL‐1β inhibited insulin‐induced activation of Akt phosphorylation, glucose transport, and fatty acid uptake. IL‐1β also blocked insulin‐mediated downregulation of suppressor of cytokine signaling‐3 expression. Co‐preincubation of IL‐1β with XOMA 052 neutralized nearly all of these inhibitory effects in 3T3‐L1 adipocytes. These studies provide evidence, therefore, that IL‐1β is a key proinflammatory cytokine that is involved in inducing insulin resistance. These studies also suggest that the monoclonal antibody XOMA 052 may be a possible therapeutic to effectively neutralize cytokine‐mediated insulin resistance in adipose tissue.     

PPARδ activation attenuates hepatic steatosis in Ldlr?/? mice by enhanced fat oxidation,reduced lipogenesis,and improved insulin sensitivity

PPARδ regulates systemic lipid homeostasis and inflammation, but its role in hepatic lipid metabolism remains unclear. Here, we examine whether intervening with a selective PPARδ agonist corrects hepatic steatosis induced by a high-fat, cholesterol-containing (HFHC) diet. Ldlr^−/− mice were fed a chow or HFHC diet (42% fat, 0.2% cholesterol) for 4 weeks. For an additional 8 weeks, the HFHC group was fed HFHC or HFHC plus GW1516 (3 mg/kg/day). GW1516-intervention significantly attenuated liver TG accumulation by induction of FA β-oxidation and attenuation of FA synthesis. In primary mouse hepatocytes, GW1516 treatment stimulated AMP-activated protein kinase (AMPK) and acetyl-CoA carboxylase (ACC) phosphorylation in WT hepatocytes, but not AMPKβ1^−/− hepatocytes. However, FA oxidation was only partially reduced in AMPKβ1^−/− hepatocytes, suggesting an AMPK-independent contribution to the GW1516 effect. Similarly, PPARδ-mediated attenuation of FA synthesis was partially due to AMPK activation, as GW1516 reduced lipogenesis in WT hepatocytes but not AMPKβ1^−/− hepatocytes. HFHC-fed animals were hyperinsulinemic and exhibited selective hepatic insulin resistance, which contributed to elevated fasting FA synthesis and hyperglycemia. GW1516 intervention normalized fasting hyperinsulinemia and selective hepatic insulin resistance and attenuated fasting FA synthesis and hyperglycemia. The HFHC diet polarized the liver toward a proinflammatory M1 state, which was reversed by GW1516 intervention. Thus, PPARδ agonist treatment inhibits the progression of preestablished hepatic steatosis.     

Suppression of lipin-1 expression increases monocyte chemoattractant protein-1 expression in 3T3-L1 adipocytes

Lipin-1 plays a crucial role in the regulation of lipid metabolism and cell differentiation in adipocytes. Expression of adipose lipin-1 is reduced in obesity, and metabolic syndrome. However, the significance of this reduction remains unclear. This study investigated if and how reduced lipin-1 expression affected metabolism. We assessed mRNA expression levels of various genes related to adipocyte metabolism in lipin-1-depleted 3T3-L1 adipocytes by introducing its specific small interfering RNA. In lipin-1-depleted adipocytes, mRNA and protein expression levels of monocyte chemoattractant protein-1 (MCP-1) were significantly increased, although the other genes tested were not altered. The conditioned media from the cells promoted monocyte chemotaxis. The increase in MCP-1 expression was prevented by treatment with quinazoline or salicylate, inhibitors of nuclear factor-κB activation. Because MCP-1 is related to adipose inflammation and systemic insulin resistance, these results suggest that a reduction in adipose lipin-1 in obesity may exacerbate adipose inflammation and metabolism.     

基于脂质组学技术研究急性缺血性脑卒中临床诊断标志物

目的:急性缺血性脑卒中(Acute ischemic stroke, AIS)是由于血流减少导致的脑功能突然丧失。由于AIS发病机制是异质性和多因素的,我们建立全面的脂质组学方法来阐明AIS进程相关的脂质变化和复杂的脂质代谢网络。方法:选取26例AIS患者血液标本和27例健康志愿者血清作为研究对象,进行总脂抽提,通过基于LC-MS策略的非靶向脂质组学方法进行规模性、整体性的脂质组学分析。结果:对AIS患者和健康志愿者血浆进行大规模脂质定性定量分析,通过Progenesis~? QI软件分析Xevo~? G2-XS QTOF质谱系统MSE采集的子离子数据,精确定量到1054个脂质特征差异,准确定性得到368个脂质分子,多变量统计分析中差异脂质组成能将AIS患者和健康志愿者区分开来,通路富集分析图显示差异脂质主要参与甘油磷脂代谢的紊乱。结论:AIS患者血浆脂质组成与健康志愿者存在显著差异,差异表达的脂质可能与AIS发生有关。这些发现有助于开发新的诊断标志物和AIS治疗靶点。     

FFA‐Induced Adipocyte Inflammation and Insulin Resistance: Involvement of ER Stress and IKKβ Pathways

Free‐fatty acids (FFAs) are well‐characterized factor for causing production of inflammatory factors and insulin resistance in adipocytes. Using cultured adipocytes, we demonstrate that FFAs can activate endoplasmic reticulum (ER) stress pathway by examination of ER stress sensor activation and marker gene expression. Chemical chaperone tauroursodeoxycholic acid (TUDCA) can reduce FFA‐induced adipocyte inflammation and improve insulin signaling whereas overexpression of spliced X‐box protein 1 (XBP‐1s) only attenuates FFA‐induced inflammation. PKR‐like eukaryotic initiation factor 2α kinase (PERK) is one of the three major ER stress sensor proteins and deficiency of PERK alleviates FFA‐induced inflammation and insulin resistance. The key downstream target of FFA‐induced ER stress is IκB kinase β (IKKβ), a master kinase for regulating expression of inflammatory genes. Deficiency of PERK attenuates FFA‐induced activation of IKKβ and deficiency of IKKβ alleviates FFA‐induced inflammation and insulin resistance. Consistently, overexpression of IKKβ in 3T3‐L1 CAR adipocytes causes inflammation and insulin resistance. In addition, IKKβ overexpression has profound effect on adipocyte lipid metabolism, including inhibition of lipogenesis and promotion of lipolysis. Furthermore, increased endogenous IKKβ expression and activation is also observed in isolated primary adipocytes from mice injected with lipids or fed on high‐fat diet (HFD) acutely. These results indicate that ER stress pathway is a key mediator for FFA‐induced inflammation and insulin resistance in adipocytes with PERK and IKKβ as the critical signaling components.     

Human milk derived peptide AOPDM1 attenuates obesity by restricting adipogenic differentiation through MAPK signalling

BackgroundEmerging evidence revealed peptides within breast milk may be an abundant source of potential candidates for metabolism regulation. Our previous work identified numerous peptides existed in breast milk, but its function has not been validated. Thus, our study aims to screen for novel peptides that have the potential to antagonize obesity and diabetes.MethodsA function screen was designed to identify the candidate peptide and then the peptide effect was validated by assessing lipid storage. Afterwards, the in vivo study was performed in two obese models: high-fat diet (HFD)-induced obese mice and obese ob/ob mice. For mechanism study, a RNA-seq analysis was conducted to explore the pathway that account for the biological function of peptide.ResultsBy performing a small scale screening, a peptide (AVPVQALLLNQ) termed AOPDM1 (anti-obesity peptide derived from breast milk 1) was identified to reduce lipid storage in adipocytes. Further study showed AOPDM1 suppressed adipocyte differentiation by sustaining ERK activity at later stage of differentiation which down-regulated PPARγ expression. In vivo, AOPDM1 effectively reduced fat mass and improved glucose metabolism in high-fat diet (HFD)-induced obese mice and obese ob/ob mice.ConclusionsWe identified a novel peptide AOPDM1 derived from breast milk could restrict adipocyte differentiation and ameliorate obesity through regulating MAPK pathway.General significanceOur findings may provide a potential candidate for the discovery of therapeutic drugs for obesity and type 2 diabetes.     

The role of the lipidome in obesity-mediated colon cancer risk

Obesity is a state of chronic inflammation influenced by lipids such as fatty acids and their secondary oxygenated metabolites deemed oxylipids. Many such lipid mediators serve as potent signaling molecules of inflammation, which can further alter lipid metabolism and lead to carcinogenesis. For example, sphingosine-1-phosphate activates cyclooxygenase-2 in endothelial cells resulting in the conversion of arachidonic acid (AA) to prostaglandin E₂ (PGE₂). PGE₂ promotes colon cancer cell growth. In contrast, the less studied path of AA oxygenation via cytochrome p450 enzymes produces epoxyeicosatetraenoic acids (EETs), whose anti-inflammatory properties cause shrinking of enlarged adipocytes, a characteristic of obesity, through the liberation of fatty acids. It is now thought that EET depletion occurs in obesity and may contribute to colon cell carcinogenesis. Meanwhile, gangliosides, a type of sphingolipid, are cell surface signaling molecules that contribute to the apoptosis of colon tumor cells. Many of these discoveries have been made recently and the mechanisms are still not fully understood, leading to an exciting new chapter of lipidomic research. In this review, mechanisms behind obesity-associated colon cancer are discussed with a focus on the role of small lipid signaling molecules in the process. Specifically, changes in lipid metabolite levels during obesity and the development of colon cancer, as well as novel biomarkers and targets for therapy, are discussed.     

PPARadigms and PPARadoxes: expanding roles for PPARgamma in the control of lipid metabolism

The nuclear receptor PPARgamma is a central regulator of adipose tissue development and an important modulator of gene expression in a number of specialized cell types including adipocytes, epithelial cells, and macrophages. PPARgamma signaling pathways impact both cellular and systemic lipid metabolism and have links to obesity, diabetes, and cardiovascular disease. The ability to activate this receptor with small molecule ligands has made PPARgamma an attractive target for intervention in human metabolic disease. As our understanding of PPARgamma biology has expanded, so has the therapeutic potential of PPARgamma ligands. Recent studies have provided insight into the paradoxical relationship between PPARgamma and metabolic disease and established new paradigms for the control of lipid metabolism. This review focuses on recent advances in PPARgamma biology in the areas of adipocyte differentiation, insulin resistance, and atherosclerosis.     

基于肠道微生态探讨黄连素在代谢性疾病中的抗炎作用

肠道菌群与能量代谢密切相关,其组成和代谢紊乱可通过多种途径导致胰岛素抵抗,肥胖和2型糖尿病。黄连素因具有减重、降糖、调脂等作用被广泛用于肥胖、2型糖尿病及非酒精性脂肪性肝病等代谢性疾病的辅助治疗;研究表明,黄连素可调节肠道菌群的组成和代谢,改善肠道微生态环境,从而改善胰岛素抵抗和代谢。本文综述了黄连素通过肠道菌群-炎症轴在干预代谢性疾病的研究进展,以期为代谢性疾病的治疗寻找新的策略,并为今后该领域的深入研究提供指导意义。     

Insulin resistance in the New Zealand obese mouse (NZO): lipolysis and lipogenesis in isolated adipocytes

The marked stimulatory effect of insulin on the metabolism of [U-¹⁴C]glucose to CO₂, glyceride-glycerol, and fatty acid observed with adipocytes from normal New Zealand yellow (NZY) mice and young (nonobese) New Zealand obese (NZO) mice was greatly diminished in cells obtained from adult obese NZO mice. Adipocytes from obese NZO mice had lower basal rates of CO₂ formation and fatty acid synthesis than cells from NZY or young NZO mice. Glyceride-glycerol was labeled to a similar extent under basal conditions in adipocytes from all three groups of mice, implying that the basal rate of glucose transport and the enzymes of the glycolytic pathway are intact in obese NZO adipocytes. Both basal and epinephrine-stimulated lipolysis were impaired in adipocytes from obese NZO mice when compared with cells from NZY and young NZO mice. Epinephrine-stimulated lipolysis was markedly less sensitive to the inhibitory effect of insulin in adipocytes from obese NZO mice than in NZY and young NZO controls. These studies suggest that adipocytes from young, nonobese NZO mice do not exhibit resistance to epinephrine and insulin, and that hormone resistance and decreased rates of metabolism accompany the onset and evolution of obesity.     

Enhanced Lipid Oxidation and Maintenance of Muscle Insulin Sensitivity Despite Glucose Intolerance in a Diet-Induced Obesity Mouse Model

Background

Diet-induced obesity is a rising health concern which can lead to the development of glucose intolerance and muscle insulin resistance and, ultimately, type II diabetes mellitus. This research investigates the associations between glucose intolerance or muscle insulin resistance and tissue specific changes during the progression of diet-induced obesity.

Methodology

C57BL/6J mice were fed a normal or high-fat diet (HFD; 60% kcal fat) for 3 or 8 weeks. Disease progression was monitored by measurements of body/tissue mass changes, glucose and insulin tolerance tests, and ex vivo glucose uptake in intact muscles. Lipid metabolism was analyzed using metabolic chambers and ex vivo palmitate assays in intact muscles. Skeletal muscle, liver and adipose tissues were analyzed for changes in inflammatory gene expression. Plasma was analyzed for insulin levels and inflammatory proteins. Histological techniques were used on muscle and liver cryosections to assess metabolic and morphological changes.

Principal Findings/Conclusions

A rapid shift in whole body metabolism towards lipids was observed with HFD. Following 3 weeks of HFD, elevated total lipid oxidation and an oxidative fiber type shift had occurred in the skeletal muscle, which we propose was responsible for delaying intramyocellular lipid accumulation and maintaining muscle’s insulin sensitivity. Glucose intolerance was present after three weeks of HFD and was associated with an enlarged adipose tissue depot, adipose tissue inflammation and excess hepatic lipids, but not hepatic inflammation. Furthermore, HFD did not significantly increase systemic or muscle inflammation after 3 or 8 weeks of HFD suggesting that early diet-induced obesity does not cause inflammation throughout the whole body. Overall these findings indicate skeletal muscle did not contribute to the development of HFD-induced impairments in whole-body glucose tolerance following 3 weeks of HFD.     

Expression and Regulation of Soluble Epoxide Hydrolase in Adipose Tissue

Obesity is an increasingly important public health issue reaching epidemic proportions. Visceral obesity has been defined as an important element of the metabolic syndrome and expansion of the visceral fat mass has been shown to contribute to the development of insulin resistance and cardiovascular disease. To identify novel contributors to cardiovascular and metabolic abnormalities in obesity, we analyzed the adipose proteome and identified soluble epoxide hydrolase (sEH) in the epididymal fat pad from C57BL/6J mice that received either a regular diet or a “western diet.” sEH was synthesized in adipocytes and expression levels increased upon differentiation of 3T3‐L1 preadipocytes. Although normalized sEH mRNA and protein levels did not differ in the fat pads from mice receiving a regular or a “western diet,” total adipose sEH activity was higher in the obese mice, even after normalization for body weight. Furthermore, peroxisome proliferator–activated receptor γ (PPARγ) agonists increased the expression of sEH in mature 3T3‐L1 adipocytes in vitro and in adipose tissue in vivo. Considering the established role for sEH in inflammation, cardiovascular diseases, and lipid metabolism, and the suggested involvement of sEH in the development of type 2 diabetes, our study has identified adipose sEH as a potential novel therapeutic target that might affect the development of metabolic and cardiovascular abnormalities in obesity.