Oligomeric amyloid-β peptide affects the expression of genes involved in steroid and lipid metabolism in primary neurons

Amyloid-β peptide (Aβ) is the principal component of plaques in the brains of patients with Alzheimer's disease (AD), and the most toxic form of Aβ may be as soluble oligomers. We report here the results of a microarray study of gene expression profiles in primary mouse cortical neurons in response to oligomeric Aβ(1-42). A major and unexpected finding was the down-regulation of genes involved in the biosynthesis of cholesterol and other steroids and lipids (such as Fdft1, Fdps, Idi1, Ldr, Mvd, Mvk, Nsdhl, Sc4mol), the expression of which was verified by quantitative real-time RT-PCR (qPCR). The ATP-binding cassette gene Abca1, which has a major role in cholesterol transport in brain and other tissues and has been genetically linked to AD, was notably up-regulated. The possible involvement of cholesterol and other lipids in Aβ synthesis and action in Alzheimer's disease has been studied and debated extensively but remains unresolved. These new data suggest that Aβ may influence steroid and lipid metabolism in neurons via multiple gene-expression changes.     

The PPARα agonist fenofibrate suppresses B-cell lymphoma in mice by modulating lipid metabolism

Obesity is associated with an increased risk for malignant lymphoma development. We used Bcr/Abl transformed B cells to determine the impact of aggressive lymphoma formation on systemic lipid mobilization and turnover. In wild-type mice, tumor size significantly correlated with depletion of white adipose tissues (WAT), resulting in increased serum free fatty acid (FFA) concentrations which promote B-cell proliferation in vitro. Moreover, B-cell tumor development induced hepatic lipid accumulation due to enhanced hepatic fatty acid (FA) uptake and impaired FA oxidation. Serum triglyceride, FFA, phospholipid and cholesterol levels were significantly elevated. Consistently, serum VLDL/LDL-cholesterol and apolipoprotein B levels were drastically increased. These findings suggest that B-cell tumors trigger systemic lipid mobilization from WAT to the liver and increase VLDL/LDL release from the liver to promote tumor growth. Further support for this concept stems from experiments where we used the peroxisome proliferator-activated receptor α (PPARα) agonist and lipid-lowering drug fenofibrate that significantly suppressed tumor growth independent of angiogenesis and inflammation. In addition to WAT depletion, fenofibrate further stimulated FFA uptake by the liver and restored hepatic FA oxidation capacity, thereby accelerating the clearance of lipids released from WAT. Furthermore, fenofibrate blocked hepatic lipid release induced by the tumors. In contrast, lipid utilization in the tumor tissue itself was not increased by fenofibrate which correlates with extremely low expression levels of PPARα in B-cells. Our data show that fenofibrate associated effects on hepatic lipid metabolism and deprivation of serum lipids are capable to suppress B-cell lymphoma growth which may direct novel treatment strategies. This article is part of a Special Issue entitled Lipid Metabolism in Cancer.     

Upregulation of miR-181a impairs lipid metabolism by targeting PPARα expression in nonalcoholic fatty liver disease

Recent studies have reported elevated expression of miR-181a in patients with non-alcoholic fatty liver disease (NAFLD), suggesting that it may play an important role in liver lipid metabolism and insulin resistance. We aimed to investigate the effect of miR-181a in lipid metabolism and find new treatments for NAFLD. The expression level of miR-181a in NAFLD patient serum and a palmitic acid (PA)-induced NAFLD cell model was examined by Q-PCR. Oil red O staining and triglyceride assays were used to assess lipid accumulation in hepatocytes. Western blotting was used to detect the protein expression levels of peroxisome proliferator-activated receptor-α (PPARα) and the fatty acid β-oxidation-related genes. Direct interactions were validated by dual-luciferase reporter gene assays. MiR-181a expression was significantly upregulated in the serum of NAFLD patients and PA-induced hepatocytes. Inhibition of miR-181a expression resulted in the increased expression of PPARα and its downstream genes, and PA-induced lipid accumulation in hepatocytes was also inhibited. Upregulation of miR-181a resulted in the downregulation of its direct target PPARα and downstream gene expression of PPARα as well as aggravated lipid accumulation in hepatocytes. At the same time, the increased expression of PPARα can offset lipid accumulation in hepatocytes induced by miR-181a mimics. This study demonstrates that reducing the expression of miR-181a may improve lipid metabolism in NAFLD. The downregulation of miR-181a expression can be a therapeutic strategy for NAFLD by modulating its target PPARα.     

Role of thyroid receptor β in lipid metabolism

Thyroid hormones (THs) exert their actions by binding to thyroid hormone receptors (TRs) and thereby affect tissue differentiation, development, and metabolism in most tissues. TH-deficiency creates a less favorable lipid profile (e.g. increased plasma cholesterol levels), whereas TH-excess is associated with both positive (e.g. reduced plasma cholesterol levels) and negative (e.g. increased heart rate) effects. TRs are encoded by two genes, THRA and THRB, which, by alternative splicing, generate several isoforms (e.g. TRα1, TRα2, TRβ1, and TRβ2). TRα, the major TR in the heart, is crucial for heart rate and for cardiac contractility and relaxation, whereas TRβ1, the major TR in the liver, is important for lipid metabolism. Selective modulation of TRβ1 is thus considered as a potential therapeutic target to treat dyslipidemia without cardiac side effects. Several selective TH analogs have been tested in preclinical studies with promising results, but only a few of these compounds have so far been tested in clinical studies. This review focuses on the role of THs, TRs, and selective and non-selective TH analogs in lipid metabolism. This article is part of a Special Issue entitled: Translating nuclear receptors from health to disease.     

The function of porcine PPARγ and dietary fish oil effect on the expression of lipid and glucose metabolism related genes

Peroxisome-proliferator-activated receptor γ (PPARγ) plays a critical role in regulation of adipocyte differentiation and insulin sensitivity. To become functional, PPARγ must be activated by binding an appropriate ligand. Polyunsaturated fatty acids (PUFA) are potential ligands for PPARγ. The current experiment was designed to determine the potential for PUFA, particularly eicosapentaenoic acid and docosahexaenoic acid, to activate the function of porcine PPARγ in vivo. Transgenic mice, expressing porcine PPARγ in skeletal muscle were generated and fed with a high-saturated fat (beef tallow) or high-unsaturated fat (fish oil) diet for 4 months. When transgenic mice were fed a fish oil supplemented diet, the expression of adipogenic and glucose uptake genes was increased, leading to reduced plasma glucose concentration. The PPARγ transgene increased the expression of Glut4 in the muscle. This result suggests that there was increased glucose utilization and, therefore, a reduced blood glucose concentration in the transgenic mice. Also, the plasma adiponectin was elevated by fish oil treatment, suggesting a role of adiponectin in mediating the PUFA effect. These results suggest that PUFA may serve as a natural regulator of glucose uptake in vivo and these effects are mainly through PPARγ function.     

Does overfeeding enhance genotype effects on energy metabolism and lipid deposition in breast muscle of ducks?

We evaluated the effects of genotype (Muscovy, Pekin and their crossbreed hinny and mule ducks) and feeding levels (overfeeding between 12 and 14 weeks of age vs ad libitum feeding) on energy metabolism and lipid deposition in breast muscle of ducks. Samples of breast muscle (Pectoralis major) were collected at 14 weeks of age from 8 birds per group. Overfeeding induced an accumulation of lipids in breast muscle (1.5- to 1.7-fold, depending on genotype) mainly induced by triglyceride deposition. It also induced a considerable increase in the amounts (expressed as g/100 g of tissue) of saturated and mono-unsaturated fatty acids (SFA, MUFA), while the amounts of poly-unsaturated fatty acids (PUFA) remained unchanged in hinny and Muscovy ducks or slightly increased in Pekin and mule ducks. In breast muscle, overfeeding decreased the activity of the main enzymes involved in lipogenesis from glucose (glucose-6-phosphate dehydrogenase, G6PDH, malic enzyme, ME, acetyl CoA carboxylase, ACX). Lipoprotein lipase (LPL) activity in Pectoralis major muscle was also significantly decreased (-21%). The ability of muscle tissues to catabolize long-chain fatty acids, as assessed by beta-hydroxyacyl CoA dehydrogenase (HAD) activity, was increased in Pectoralis major muscle, as was cytochrome-c oxidase (COX) activity. Hybrid and Pekin ducks exhibited higher levels of ACX and LPL activity in Pectoralis major muscle than Muscovy ducks, suggesting a greater ability to synthesise lipids in situ, and to take up circulating lipids. Total lipid content in breast muscle of hybrid and Pekin ducks was higher than in that of Muscovy ducks. In hybrid and Pekin ducks, lipid composition of breast muscle was characterized by higher amounts of triglycerides, SFA and MUFA than in Muscovy ducks. Finally, oxidative metabolism was greater in Pectoralis major muscles of hybrid and Pekin ducks than in Muscovy ducks, suggesting an adaptative strategy of muscle energy metabolism according to lipid level.     

Cholesterol enhances amyloid β deposition in mouse retina by modulating the activities of Aβ-regulating enzymes in retinal pigment epithelial cells

Cancer patients frequently develop autoantibodies. To test the hypothesis that the appearance of autoantibodies precedes the clinical diagnosis of cancer, we applied an immunoproteomic approach to compare autoantibody profiles before and after appearance of malignances. Proteins from A549 cells, a lung adenocarcinoma cell line, were separated by two dimensional electrophoresis and then immunoblotted with serum samples from 8 individuals who were eventually diagnosed with lung cancer. Compared with autoantibody profiles from 3 years prior to the appearance of malignances, 21 immunoreactive spots newly appeared or presented with stronger staining intensity when clinical diagnoses were made. Among them, 10 matched spots on 2-DE gels were identified by mass spectrometry analysis as 5 proteins. With immunoprecipitation analysis, the antigenicity of protein cathepsin D was confirmed, and notably, in lung cancer sera, the occurrences of autoantibodies against the specific forms of cathepsin D differed significantly from the control groups (p<0.05). Our findings suggest that harnessing immunity may have utility for early cancer marker discovery, and that comparing autoantibodies to specific forms of cathepsin D may be a promising early marker of lung cancer.     

Relationship between signal transduction and PPARα-regulated genes of lipid metabolism in rat hepatic-derived Fao cells

The goal of this study was to characterize phosphorylated proteins and to evaluate the changes in their phosphorylation level under the influence of a peroxisome proliferator (PP) with hypolipidemic activity of the fibrate family. The incubation of rat hepatic derived Fao cells with ciprofibrate leads to an overphosphorylation of proteins, especially one of 85 kDa, indicating that kinase (or phosphatase) activities are modified. Moreover, immunoprecipitation of ³²P-labeled cell lysates shows that the nuclear receptor, PP-activated receptor, α isoform, can exist in a phosphorylated form, and its phosphorylation is increased by ciprofibrate. This study shows that PP acts at different steps of cell signaling. These steps can modulate gene expression of enzymes involved in fatty acid metabolism and lipid homeostasis, as well as in detoxication processes.     

BMP6 increases TGF-β1 production by up-regulating furin expression in human granulosa-lutein cells

Bone morphogenetic protein 6 (BMP6) and transforming growth factor-β1 (TGF-β1) are key intraovarian regulators that play essential roles in regulating mammalian follicular function and promoting oocyte maturation. Furin, a member of the subtilisin-like proprotein convertase family, promotes the activation of diverse functional proteins by cleaving protein precursors in the secretory pathway. The aim of this study was to investigate the effect and underlying molecular mechanisms by which BMP6 regulates the expression of furin to increase TGF-β1 production. Primary and immortalized (SVOG) human granulosa-lutein (hGL) cells were used as study models. Our results show that BMP6 significantly up-regulated the expression of furin and increased the production of TGF-β1 in hGL cells. Using dual inhibition approaches (kinase receptor inhibitors and small interfering RNA-targeted knockdown), we demonstrate that both activin receptor-like (ALK)2 and ALK3 are involved in the BMP6-induced up-regulation of furin. Additionally, knockdown of furin abolished BMP6-induced increases in TGF-β1 production. Moreover, knockdown of endogenous SMAD4 reversed the BMP6-induced increase in furin expression. These results indicate that the ALK2/3-mediated canonical SMAD signaling pathway is required for the stimulatory effect of BMP6 on furin expression, which in turn increases the production of TGF-β1 in hGL cells. Our findings provide insights into the molecular interactions and mechanisms of two intrafollicular growth factors in hGL cells.     

Characterization and expression of chitinase and 1,3-β-glucanase genes in cotton

We have isolated cDNA clones representing mRNAs encoding chitinase and 1,3--glucanase in cotton (Gossypium hirsutum L.) leaves. The chitinase clones were sequenced and found to encode a 28,806 Da protein with 71% amino acid sequence similarity to the SK2 chitinase from potato (Solanum tuberosum). The 1,3--glucanase clones encoded a 37,645 Da protein with 57.6% identity to a 1,3--glucanase from soybean (Glycine max). Northern blot analyses showed that chitinase mRNA is induced in plants treated with ethaphon or salicylic acid, whereas the levels of 1,3--glucanase mRNA are relatively unaffected. Southern blots of cotton genomic DNA and genomic clones indicated chitinase is encoded by a small gene family of which two members, Chi 2;1 and Chi 2;2, were characterized. These genes share 97% sequence identity in their transcribed regions. The genes were found to have three exons which are 309, 154 and 550 bp long, and two introns 99 and 154 bp in length. The 5-flanking regions of Chi 2;1 and Chi 2;2 exhibit a large degree of similarity and may contain sequences important for gene response to chemical agents and fungal attack.     

The expression of three desaturase genes of Spirulina platensis in Escherichia coli DH5α – Heterologous expression of Spirulina-desaturase genes

The genes from a cyanobacterium--Spirulina platensis strain C1--that encode the acyl-lipid desaturases (desC, desA and desD) involved in gamma-linolenic (GLA) synthesis have been successfully expressed for the first time in Escherichia coli by employing a pTrcHisA expression system. In this report, the authors describe the expression of the three Spirulina N-terminal 6xHis-desaturases as well as the functional analysis of these recombinant proteins. The gene products of desC, desA and desD have approximate molecular masses of 37, 45, and 47 kDa, respectively. Enzymatic activity measurement of these products was carried out in vivo to demonstrate that (i) the expressed proteins are in functional form, and (ii) the cofactors of the host system can complement the system of Spirulina platensis. The study demonstrated that the gene products of desC and desA catalyzed the reactions in vivo where the enzyme substrates were provided in appropriate concentration. This indicates that the delta9 and delta12 desaturases were expressed in the heterologous host in their active form, and that these two reactions can be carried out in an E. coli host cell using its cofactors system. In contrast, delta6 desaturase activity can be detected only in vitro where electron carriers are provided. This suggests that while this enzyme is expressed in the heterologous host in its active form, its function in vivo is suppressed, as the electron carriers of the host system cannot complement the system of Spirulina platensis.     

Reduced expression of β2 integrin genes in rat peripheral leukocytes by inhibiting postprandial hyperglycemia

β(2) integrins (CD11s/CD18) promote the attachment of leukocytes to vascular endothelial cells. We performed in this study sucrose loading to rats with moderate postprandial hyperglycemia with/without once-daily dosing of the α-glucosidase inhibitor, miglitol, for 4 days under 4-h fasting conditions. The streptozotocin (STZ)-treated rats showed moderate postprandial hyperglycemia on days 1 and 4. The gene expression was higher for CD11a with fasting and 3-h postprandial on day 1, and with fasting on day 4, for CD11b with fasting on day 1 and 3-h postprandial on day 4, and for CD18 with fasting on days 1 and 4 in peripheral leukocytes from the STZ-treated rats than in peripheral leukocytes from the saline-treated rats. Miglitol reduced postprandial hyperglycemia and the gene expression of CD11a with fasting and of CD11b 3-h postprandial on day 4. These results indicate that inhibiting postprandial hyperglycemia reduced the mRNA expression of β(2) integrins in peripheral leukocytes of moderately postprandial hyperglycemic rats.     

Resveratrol protects rats from Aβ-induced neurotoxicity by the reduction of iNOS expression and lipid peroxidation

Alzheimer disease (AD) is an age-dependent neurodegenerative disease characterized by the formation of β-amyloid (Aβ)-containing senile plaque. The disease could be induced by the administration of Aβ peptide, which was also known to upregulate inducible nitric oxide synthase (iNOS) and stimulate neuronal apoptosis. The present study is aimed to elucidate the cellular effect of resveratrol, a natural phytoestrogen with neuroprotective activities, on Aβ-induced hippocampal neuron loss and memory impairment. On adult Sprague-Dawley rats, we found the injection of Aβ could result in a significant impairment in spatial memory, a marked increase in the cellular level of iNOS and lipid peroxidation, and an apparent decrease in the expression of heme oxygenase-1 (HO-1). By combining the treatment with Aβ, resveratrol was able to confer a significant improvement in spatial memory, and protect animals from Aβ-induced neurotoxicity. These neurological protection effects of resveratrol were associated with a reduction in the cellular levels of iNOS and lipid peroxidation and an increase in the production of HO-1. Moreover, the similar neurological and cellular response were also observed when Aβ treatment was combined with the administration of a NOS inhibitor, N(G)-nitro-L-arginine methyl ester hydrochloride (L-NAME). These findings strongly implicate that iNOS is involved in the Aβ-induced lipid peroxidation and HO-1 downregulation, and resveratrol protects animals from Aβ-induced neurotoxicity by suppressing iNOS production.     

Effects of antisense-mediated inhibition of 11β-hydroxysteroid dehydrogenase type 1 on hepatic lipid metabolism

11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) converts inactive 11-keto derivatives to active glucocorticoids within tissues and may play a role in the metabolic syndrome (MS). We used an antisense oligonucleotide (ASO) to knock down 11β-HSD1 in livers of C57BL/6J mice consuming a Western-type diet (WTD). 11β-HSD1 ASO-treated mice consumed less food, so we compared them to ad libitum-fed mice and to food-matched mice receiving control ASO. Knockdown of 11β-HSD1 directly protected mice from WTD-induced steatosis and dyslipidemia by reducing synthesis and secretion of triglyceride (TG) and increasing hepatic fatty acid oxidation. These changes in hepatic and plasma lipids were not associated with reductions in genes involved in de novo lipogenesis. However, protein levels of both sterol regulatory element-binding protein (SREBP) 1 and fatty acid synthase were significantly reduced in mice treated with 11β-HSD1 ASO. There was no change in hepatic secretion of apolipoprotein (apo)B, indicating assembly and secretion of smaller apoB-containing lipoproteins by the liver in the 11β-HSD1-treated mice. Our results indicate that inhibition of 11β-HSD1 by ASO treatment of WTD-fed mice resulted in improved plasma and hepatic lipid levels, reduced lipogenesis by posttranslational regulation, and secretion of similar numbers of apoB-containing lipoproteins containing less TG per particle.