ABSTRACT
Accumulating evidence indicates that mitochondrial dysfunction and oxidative stress play a pivotal role in the initiation and progression of nonalcoholic fatty liver disease (NAFLD). In this study, we found that blueberry-derived exosomes-like nanoparticles (BELNs) could ameliorate oxidative stress in rotenone-induced HepG2 cells and high-fat diet (HFD)-fed C57BL/6 mice. Preincubation with BELNs decreased the level of reactive oxygen species (ROS), increased the mitochondrial membrane potential, and prevented cell apoptosis by inducing the expression of Bcl-2 and heme oxygenase-1 (HO-1) and decreasing the content of Bax in rotenone-treated HepG2 cells. We also found that preincubation with BELNs accelerated the translocation of Nrf2, an important transcription factor of antioxidative proteins, from the cytoplasm to the nucleus in rotenone-treated HepG2 cells. Moreover, administration of BELNs improved insulin resistance, ameliorated the dysfunction of hepatocytes, and regulated the expression of detoxifying/antioxidant genes by affecting the distribution of Nrf2 in the cytoplasm and nucleus of hepatocytes of HFD-fed mice. Furthermore, BELNs supplementation prevented the formation of vacuoles and attenuated the accumulation of lipid droplets by inhibiting the expression of fatty acid synthase (FAS) and acetyl-CoA carboxylase 1 (ACC1), the two key transcription factors for de novo lipogenesis in the liver of HFD-fed mice. These findings suggested that BELNs can be used for the treatment of NAFLD because of their antioxidative activity.
Subject(s)
Biological Products/pharmacology , Blueberry Plants , Exosomes/metabolism , Mitochondria/drug effects , Non-alcoholic Fatty Liver Disease/pathology , Oxidative Stress/drug effects , Acetyl-CoA Carboxylase/drug effects , Animals , Apoptosis/drug effects , Disease Models, Animal , Fatty Acid Synthases/drug effects , Heme Oxygenase-1/drug effects , Hep G2 Cells , Humans , Insulin Resistance/physiology , Membrane Potential, Mitochondrial/drug effects , Mice, Inbred C57BL , NF-E2-Related Factor 2/drug effects , Nanoparticles , Proto-Oncogene Proteins c-bcl-2/drug effects , Reactive Oxygen Species/metabolismABSTRACT
Coronaviruses rely on host membranes for entry, establishment of replication centers, and egress. Compounds targeting cellular membrane biology and lipid biosynthetic pathways have previously shown promise as antivirals and are actively being pursued as treatments for other conditions. Here, we test small molecule inhibitors that target the PI3 kinase VPS34 or fatty acid metabolism for anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) activity. Our studies determine that compounds targeting VPS34 are potent SARS-CoV-2 inhibitors. Mechanistic studies with compounds targeting multiple steps up- and downstream of fatty acid synthase (FASN) identify the importance of triacylglycerol production and protein palmitoylation as requirements for efficient viral RNA synthesis and infectious virus production. Further, FASN knockout results in significantly impaired SARS-CoV-2 replication that can be rescued with fatty acid supplementation. Together, these studies clarify roles for VPS34 and fatty acid metabolism in SARS-CoV-2 replication and identify promising avenues for the development of countermeasures against SARS-CoV-2.
Subject(s)
Antiviral Agents/pharmacology , COVID-19/virology , Class III Phosphatidylinositol 3-Kinases/antagonists & inhibitors , Lipid Metabolism/drug effects , SARS-CoV-2/drug effects , SARS-CoV-2/physiology , Virus Replication/drug effects , Aminopyridines/pharmacology , Animals , Caco-2 Cells , Cell Line , Chlorocebus aethiops , Class III Phosphatidylinositol 3-Kinases/metabolism , Fatty Acid Synthases/drug effects , Fatty Acid Synthases/genetics , Gene Knockout Techniques , Humans , Lipoylation/drug effects , Pyrimidines/pharmacology , RNA, Viral/metabolism , Triglycerides/metabolism , Vero CellsABSTRACT
The regulation of lipogenesis and lipolysis mechanisms related to consumption of lipid has not been studied in swimming crab. The aims of the present study were to evaluate the effects of dietary lipid levels on growth, enzymes activities and expression of genes of lipid metabolism in hepatopancreas of juvenile swimming crab. Three isonitrogenous diets were formulated to contain crude lipid levels at 5·8, 9·9 and 15·1 %. Crabs fed the diet containing 15·1 % lipid had significantly lower growth performance and feed utilisation than those fed the 5·8 and 9·9 % lipid diets. Crabs fed 5·8 % lipid had lower malondialdehyde concentrations in the haemolymph and hepatopancreas than those fed the other diets. Highest glutathione peroxidase in haemolymph and superoxide dismutase in hepatopancreas were observed in crabs fed 5·8 % lipid. The lowest fatty acid synthase and glucose 6-phosphate dehydrogenase activities in hepatopancreas were observed in crabs fed 15·1 % lipid, whereas crabs fed 5·8 % lipid had lower carnitine palmitoyltransferase-1 activity than those fed the other diets. Crabs fed 15·1 % lipid showed lower hepatopancreas expression of genes involved in long-chain-PUFA biosynthesis, lipoprotein clearance, fatty acid uptake, fatty acid oxidation, lipid anabolism and lipid catabolism than those fed the other diets, whereas expression of some genes of lipoprotein assembly and fatty acid oxidation was up-regulated compared with crabs fed 5·8 % lipid. Overall, high dietary lipid level can inhibit growth, reduce antioxidant enzyme activities and influence lipid metabolic pathways to regulate lipid deposition in crab.
Subject(s)
Brachyura/growth & development , Brachyura/metabolism , Dietary Fats/administration & dosage , Gene Expression/drug effects , Lipid Metabolism/drug effects , Animal Feed/analysis , Animals , Antioxidants/metabolism , Brachyura/enzymology , Brachyura/genetics , Diet/veterinary , Dietary Fats/pharmacology , Fatty Acid Synthases/drug effects , Fatty Acids/metabolism , Glucosephosphate Dehydrogenase/drug effects , Glutathione Peroxidase/drug effects , Hepatopancreas/drug effects , Hepatopancreas/enzymology , Hepatopancreas/metabolism , Malondialdehyde/metabolism , Oxidation-Reduction/drug effects , Superoxide Dismutase/drug effects , SwimmingABSTRACT
Chikungunya virus (CHIKV) is a rapidly emerging mosquito-borne RNA virus that causes epidemics of debilitating disease in tropical and sub-tropical regions with autochtonous transmission in regions with temperate climate. Currently, there is no licensed vaccine or specific antiviral drug available against CHIKV infection. In this study, we examine the role, in the CHIKV viral cycle, of fatty acid synthase (FASN) and stearoyl-CoA desaturase (SCD1), two key lipogenic enzymes required for fatty acid production and early desaturation. We show that both enzymes and their upstream regulator PI3K are required for optimal CHIKV infection. We demonstrate that pharmacologic manipulation of FASN or SCD1 enzymatic activity by non-toxic concentrations of cerulenin or CAY10566 decreases CHIKV genome replication. Interestingly, a similar inhibitory effect was also obtained with Orlistat, an FDA-approved anti-obesity drug that targets FASN activity. These drugs were also effective against Mayaro virus (MAYV), an under-studied arthritogenic Old world Alphavirus endemic in South American countries with potential risk of emergence, urbanization and dispersion to other regions. Altogether, our results identify FASN and SCD1 as conserved druggable cofactors of Alphavirus genome replication and support the broad-spectrum activity of drugs targeting the host fatty acids metabolism.
Subject(s)
Alphavirus/drug effects , Fatty Acid Synthases/metabolism , Stearoyl-CoA Desaturase/metabolism , Virus Replication/drug effects , Alphavirus/genetics , Alphavirus Infections/drug therapy , Animals , Antiviral Agents/pharmacology , Cell Line , Cerulenin/pharmacology , Chikungunya Fever/drug therapy , Chikungunya virus/drug effects , Chikungunya virus/genetics , Chlorocebus aethiops , Fatty Acid Synthases/drug effects , Genome, Viral , HEK293 Cells , Humans , Orlistat/pharmacology , Stearoyl-CoA Desaturase/drug effects , Vero CellsABSTRACT
Despite advances in diagnostic and therapeutic management, oral squamous cell carcinoma (OSCC) patient survival rates have remained relatively unchanged. Thus, identifying early triggers of malignant progression is critical to prevent OSCC development. Traditionally, OSCC initiation is elicited by the frequent and direct exposure to multiple tobacco-derived carcinogens, and not by the nicotine contained in tobacco products. However, other nicotine-containing products, especially the increasingly popular electronic cigarettes (e-cigs), have unknown effects on the progression of undiagnosed tobacco-induced oral premalignant lesions, specifically in regard to the effects of nicotine. Overexpression of fatty acid synthase (FASN), a key hepatic de novo lipogenic enzyme, is linked to poor OSCC patient survival. Nicotine upregulates hepatic FASN, but whether this response occurs in oral dysplastic keratinocytes is unknown. We hypothesized that in oral dysplastic keratinocytes, nicotine triggers a migratory phenotype through FASN-dependent epidermal growth factor receptor (EGFR) activation, a common pro-oncogenic event supporting oral carcinogenesis. We report that in oral dysplastic cells, nicotine markedly upregulates FASN leading to FASN-dependent EGFR activation and increased cell migration. These results raise potential concerns about e-cig safety, especially when used by former tobacco smokers with occult oral premalignant lesions where nicotine could trigger oncogenic signals commonly associated with malignant progression.
Subject(s)
Carcinoma, Squamous Cell/drug therapy , ErbB Receptors/drug effects , Keratinocytes/drug effects , Nicotine/pharmacology , Carcinogenesis/drug effects , Carcinogenesis/pathology , Carcinoma, Squamous Cell/pathology , Cell Line, Tumor , Cell Movement/drug effects , Electronic Nicotine Delivery Systems/methods , ErbB Receptors/metabolism , Fatty Acid Synthases/drug effects , Fatty Acid Synthases/metabolism , Humans , Keratinocytes/metabolism , Mouth Neoplasms/drug therapy , Mouth Neoplasms/pathology , Precancerous Conditions/drug therapy , Precancerous Conditions/pathology , Up-RegulationABSTRACT
Increased de novo synthesis of fatty acids is a rather unique and targetable mechanism of human prostate cancer. We have shown previously that oral administration of sulforaphane (SFN) significantly inhibits the incidence and/or burden of prostatic intraepithelial neoplasia and well-differentiated adenocarcinoma in TRansgenic Adenocarcinoma of Mouse Prostate (TRAMP) mice. The present study used cellular models of prostate cancer and archived plasma/adenocarcinoma tissues and sections from the TRAMP study to demonstrate inhibition of fatty acid synthesis by SFN treatment in vitro and in vivo. Treatment of androgen-responsive (LNCaP) and castration-resistant (22Rv1) human prostate cancer cells with SFN (5 and 10 µM) resulted in downregulation of protein and mRNA levels of acetyl-CoA carboxylase 1 (ACC1) and fatty acid synthase (FASN), but not ATP citrate lyase. Protein and mRNA levels of carnitine palmitoyltransferase 1A (CPT1A), which facilitates fatty acid uptake by mitochondria for ß-oxidation, were also decreased following SFN treatment in both cell lines. Immunohistochemistry revealed a significant decrease in expression of FASN and ACC1 proteins in prostate adenocarcinoma sections of SFN-treated TRAMP mice when compared with controls. SFN administration to TRAMP mice resulted in a significant decrease in plasma and/or prostate adenocarcinoma levels of total free fatty acids, total phospholipids, acetyl-CoA and ATP. Consistent with these results, number of neutral lipid droplets was lower in the prostate adenocarcinoma sections of SFN-treated TRAMP mice than in control tumors. Collectively, these observations indicate that prostate cancer chemoprevention by SFN in TRAMP mice is associated with inhibition of fatty acid metabolism.
Subject(s)
Anticarcinogenic Agents/pharmacology , Fatty Acids/metabolism , Isothiocyanates/pharmacology , Prostatic Neoplasms/prevention & control , Adenocarcinoma/metabolism , Adenocarcinoma/prevention & control , Animals , Chemoprevention/methods , Fatty Acid Synthases/drug effects , Fatty Acid Synthases/metabolism , Humans , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Mice, Transgenic , Prostate/drug effects , Prostate/metabolism , Prostatic Intraepithelial Neoplasia/metabolism , Prostatic Intraepithelial Neoplasia/prevention & control , Prostatic Neoplasms/metabolism , SulfoxidesABSTRACT
Oral squamous cell carcinoma (OSCC) is an aggressive and treatment-resistant malignancy in both feline and human patients. Recent work has demonstrated aberrant expression of fatty acid synthase (FASN) and an increased capacity for lipogenesis in human OSCC and other cancers. In human OSCC, inhibition of FASN decreased cell viability and growth in vitro, and diminished tumour growth and metastasis in murine preclinical models. This study aimed to characterize FASN as a therapeutic target in feline OSCC. Immunohistochemistry revealed high FASN expression in primary feline OSCC tumours, and FASN expression was detected in OSCC cell lines (3 feline and 3 human) by immunoblotting and quantitative real-time-polymerase chain reaction (qRT-PCR). Orlistat, a FASN inhibitor, substantially reduced cell viability in both feline and human OSCC lines, although feline cell lines consistently displayed higher sensitivity to the drug. FASN mRNA expression among cell lines mirrored sensitivity to orlistat, with feline cell lines expressing higher levels of FASN. Consistent with this observation, diminished sensitivity to orlistat treatment and decreased FASN mRNA expression were observed in feline OSCC cells following incubation under hypoxic conditions. Treatment with orlistat did not potentiate sensitivity to carboplatin in the cell lines investigated; instead, combinations of the 2 drugs resulted in additive to antagonistic effects. Our results suggest that FASN inhibition is a viable therapeutic target for feline OSCC. Furthermore, cats may serve as a spontaneous large animal model for human oral cancer, although differences in the regulation of lipogenesis between these 2 species require further investigation.
Subject(s)
Carcinoma, Squamous Cell/veterinary , Cat Diseases/enzymology , Fatty Acid Synthases/metabolism , Mouth Neoplasms/veterinary , Animals , Carcinoma, Squamous Cell/drug therapy , Carcinoma, Squamous Cell/enzymology , Cat Diseases/drug therapy , Cats , Cell Line, Tumor , Fatty Acid Synthases/antagonists & inhibitors , Fatty Acid Synthases/drug effects , Humans , Immunoblotting/veterinary , Lactones/pharmacology , Mouth Neoplasms/drug therapy , Mouth Neoplasms/enzymology , Orlistat , Real-Time Polymerase Chain Reaction/veterinaryABSTRACT
BACKGROUND: Manganese superoxide dismutase (MnSOD) is an important antioxidant enzyme affected in heart/muscle-specific MnSOD-deficient mice (H/M-SOD2-/-), which develop progressive congestive heart failure and exhibit pathology typical of dilated cardiomyopathy. METHODS: In this study we investigated the beneficial effects of epigallocatechin gallate (EGCG) on the cardiac remodeling and telomere biology in H/M-SOD2-/- mice. H/M-SOD2-/- mice were divided into three groups: those receiving normal drinking water (KO), a low dose of EGCG (L: 10mg/L), and a high dose of EGCG (H: 100mg/L) beginning at eight weeks of age and lasting for eight weeks. RESULTS: The mice in the KO group exhibited significantly dilated cardiac remodeling with reduced contractility, which was prevented by the administration of EGCG. Although the mortality of KO mice was about 50% at 16 weeks of age, the mice that received EGCG had a high survival rate. The cardiac dilatation with reduced cardiac contraction in KO mice was prevented by EGCG treatment. The levels of myocardial oxidative stress and free fatty acids were lower in the group treated with EGCG compared with the KO group. The increased expression of nitric oxide synthase 2, nitrotyrosine, fatty acid synthase, Toll-like receptor 4, and Sirt1 in the KO mice were prevented by EGCG treatment. The shortening of the telomere length, decreased telomerase activity in KO mice were also prevented by EGCG. CONCLUSIONS: H/M-SOD2-/- mice receiving EGCG have a lower mortality rate and exhibit less inflammation and a better preserved cardiac function and telomere biology.
Subject(s)
Antioxidants/administration & dosage , Catechin/analogs & derivatives , Heart Failure/drug therapy , Animals , Catechin/administration & dosage , Disease Models, Animal , Fatty Acid Synthases/drug effects , Fatty Acids, Nonesterified/metabolism , Mice , Mice, Knockout , Myocardium/metabolism , Nitric Oxide Synthase Type II/drug effects , Oxidative Stress/drug effects , Sirtuin 1/drug effects , Superoxide Dismutase/genetics , Telomerase/drug effects , Telomere Shortening/drug effects , Toll-Like Receptor 4/drug effects , Tyrosine/analogs & derivatives , Tyrosine/drug effects , Ventricular Remodeling/drug effectsABSTRACT
OBJECTIVE: Fatty acid synthase (FAS) is a key lipogenic enzyme that is highly expressed in endometrial cancer. Orlistat is a weight loss medication that has been shown to be a potent inhibitor of FAS. The goal of this study was to evaluate the anti-tumorigenic potential of orlistat in endometrial cancer cell lines. METHODS: The endometrial cancer cell lines ECC-1 and KLE were used. Cell proliferation was assessed by MTT assay after treatment with orlistat. Cell cycle progression was evaluated by Cellometer and apoptosis was assessed using the Annexin V assay. Reactive oxygen species (ROS) was measured using the DCFH-DA assay. Western immunoblotting was performed to determine changes in FAS, cellular stress, cell cycle progression, and the AMPK/mTOR pathways. RESULTS: Orlistat inhibited cell proliferation by 61 % in ECC-1 cells and 57 % in KLE cells at a dose of 500 µM. Treatment with orlistat at this concentration resulted in G1 arrest (p < 0.05) but did not affect apoptosis. Orlistat increased ROS and induced the expression of BIP (1.28-fold in ECC-1 compared to control, p < 0.05; 1.92-fold in KLE, p < 0.05) and PERK (2.25-fold in ECC-1, 1.4-fold in KLE, p < 0.05). Western immunoblot analysis demonstrated that orlistat decreased expression of important proteins in fatty acid metabolism including FAS (67 % in ECC-1, 15 % in KLE), acetyl-CoA carboxylase (40 % in ECC-1, 35 % in KLE), and carnitine palmitoyltransferase 1A (CPT1A) (65 % in ECC-1, 25 % in KLE) in a dose-dependent manner. In addition, orlistat at a dose of 500 µM increased expression of phosphorylated-AMPK (1.9-fold in ECC-1, p < 0.01; 1.5-fold in KLE, p < 0.05) and decreased expression of phosphorylated-Akt (25 % in ECC-1, p < 0.05; 37 % in KLE, p < 0.05) and phosphorylated-S6 (68 % in ECC-1, 56 % in KLE). CONCLUSIONS: Orlistat inhibits cell growth in endometrial cancer cell lines through inhibition of fatty acid metabolism, induction of cell cycle G1 arrest, activation of AMPK and inhibition of the mTOR pathway. Given that patients with endometrial cancer have high rates of obesity, orlistat should be further investigated as a novel strategy for endometrial cancer treatment.
Subject(s)
Endometrial Neoplasms/drug therapy , Fatty Acid Synthases/drug effects , Lactones/therapeutic use , Cell Line, Tumor , Cell Proliferation , Endometrial Neoplasms/pathology , Fatty Acid Synthases/antagonists & inhibitors , Female , Humans , Lactones/administration & dosage , Lactones/pharmacology , Orlistat , Reactive Oxygen SpeciesABSTRACT
Fatty acid synthase (FAS) is overexpressed in many human cancers including breast cancer and is considered to be a promising target for therapy. Sea buckthorn has long been used to treat a variety of maladies. Here, we investigated the inhibitory effect of sea buckthorn procyanidins (SBPs) isolated from the seeds of sea buckthorn on FAS and FAS overexpressed human breast cancer MDA-MB-231 cells. The FAS activity and FAS inhibition were measured by a spectrophotometer at 340 nm of nicotinamide adenine dinucleotide phosphate (NADPH) absorption. We found that SBP potently inhibited the activity of FAS with a half-inhibitory concentration (IC50) value of 0.087 µg/ml. 3-4,5-Dimethylthiazol-2-yl-2,3-diphenyl tetrazolium bromide (MTT) assay was used to test the cell viability. SBP reduced MDA-MB-231 cell viability with an IC50 value of 37.5 µg/ml. Hoechst 33258/propidium iodide dual staining and flow cytometric analysis showed that SBP induced MDA-MB-231 cell apoptosis. SBP inhibited intracellular FAS activity with a dose-dependent manner. In addition, sodium palmitate could rescue the cell apoptosis induced by SBP. These results showed that SBP was a promising FAS inhibitor which could induce the apoptosis of MDA-MB-231 cells via inhibiting FAS. These findings suggested that SBP might be useful for preventing or treating breast cancer.
Subject(s)
Breast Neoplasms/enzymology , Fatty Acid Synthases/antagonists & inhibitors , Hippophae , Phytotherapy/methods , Proanthocyanidins/pharmacology , Apoptosis/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , Fatty Acid Synthases/drug effects , Flow Cytometry , Hippophae/chemistry , Humans , SeedsABSTRACT
BACKGROUND: Grape skin, a rich source of phytochemicals, has been reported to possess remarkable anti-obesity activity. Fatty acid synthase (FAS) is a key enzyme catalyzing the synthesis of fatty acid de novo, and has been considered as an anti-obesity target. To elucidate the anti-obesity mechanism of grape skin, we investigated the effects of grape skin extract (GSE) and resveratrol, one of the phytochemicals in GSE, on FAS and FAS over-expressed 3 T3-L1 preadipocyte. METHODS: Purified FAS was obtained from chicken liver. Dried grape skin was extracted by 50% ethanol and partitioned by ethyl acetate. Inhibitory effects of GSE and resveratrol on FAS including fast-binding inhibition, time-dependent inhibition, and enzyme kinetics were determined. Inhibitory effects of GSE and resveratrol on 3T3-L1 preadipocyte were also measured. RESULTS: GSE inhibited the overall reaction and ß-ketoacyl reductase (KR) reaction of FAS with IC50 values of 4.61 µg/ml and 20.3 µg/ml. For inhibition by resveratrol, the relevant IC50 values were 11.1 µg/ml and 21.9 µg/ml, respectively. And both GSE and resveratrol showed time-dependent inhibition for FAS, with the kobs values of 0.028 min-1, and 0.040 min-1 respectively. They inhibited the overall reaction of FAS competitively with acetyl-CoA, noncompetitively with malonyl-CoA and in a mixed manner with NADPH. Moreover, the inhibition on KR domain by resveratrol was time-dependent with kobs value of 0.106 min-1. In 3 T3-L1 preadipocytes, resveratrol reduced lipid accumulation remarkably. CONCLUSIONS: GSE and resveratrol are potent FAS inhibitors and they bound reversibly to the KR domain of FAS to inhibit the reduction of the saturated acyl groups in fatty acid synthesis. Based on the valid data and deliberate analysis, we proposed that GSE and resveratrol have great medical potential and officinal value in treating obesity and related diseases.
Subject(s)
Enzyme Inhibitors/pharmacology , Fatty Acid Synthases/antagonists & inhibitors , Fatty Acid Synthases/drug effects , Plant Extracts/pharmacology , Vitis/chemistry , 3T3-L1 Cells , Adipocytes/drug effects , Animals , Cell Survival/drug effects , Enzyme Inhibitors/chemistry , Fatty Acid Synthases/metabolism , Lipids/analysis , Lipogenesis/drug effects , Mice , Oxidoreductases/antagonists & inhibitors , Oxidoreductases/drug effects , Oxidoreductases/metabolism , Plant Extracts/chemistry , Resveratrol , Stilbenes/chemistry , Stilbenes/pharmacologyABSTRACT
The effect of frequent protein malnutrition on liver function has not been intensively examined. Thus, the effects of alternating 5 days of a protein and amino acid-free diet followed by 5 days of a complete diet repeated three times (3 PFD-CD) on female mouse liver were examined. The expression of carbonic anhydrase III (CAIII), fatty acid synthase (FAS), glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and glutathione S-transferase P1 (GSTP1) in liver were assessed by proteomics, reverse transcriptase-polymerase chain reaction and Northern blotting. The activities of liver GSTs, glutathione reductase (GR) and catalase (CAT), as well as serum glutamic-oxaloacetic transaminase (SGOT) and glutamic-pyruvic transaminase (SGPT) were also tested. Additionally, oxidative damage was examined by measuring of protein carbonylation and lipid peroxidation. Liver histology was examined by light and electron microscopy. Compared with control mice, 3 PFD-CD increased the content of FAS protein (+90%) and FAS mRNA (+30%), while the levels of CAIII and CAIII mRNAs were decreased (-48% and -64%, respectively). In addition, 3 PFD-CD did not significantly change the content of GSTP1 but produced an increase in its mRNA level (+20%), while it decreased the activities of both CAT (-66%) and GSTs (-26%). After 3 PFD-CD, liver protein carbonylation and lipid peroxidation were increased by +55% and +95%, respectively. In serum, 3 PFD-CD increased the activities of both SGOT (+30%) and SGPT (+61%). In addition, 3 PFD-CD showed a histological pattern characteristic of hepatic damage. All together, these data suggest that frequent dietary amino acid deprivation causes hepatic metabolic and ultrastructural changes in a fashion similar to precancerous or cancerous conditions.
Subject(s)
Dietary Proteins/administration & dosage , Liver/metabolism , Liver/pathology , Malnutrition/metabolism , Oxidative Stress/drug effects , Alanine Transaminase/blood , Alanine Transaminase/drug effects , Alanine Transaminase/metabolism , Animals , Aspartate Aminotransferases/blood , Aspartate Aminotransferases/drug effects , Aspartate Aminotransferases/metabolism , Carbonic Anhydrase III/drug effects , Carbonic Anhydrase III/metabolism , Catalase/drug effects , Catalase/metabolism , Fatty Acid Synthases/drug effects , Fatty Acid Synthases/metabolism , Female , Glutathione Peroxidase/drug effects , Glutathione Peroxidase/metabolism , Glutathione Reductase/drug effects , Glutathione Reductase/metabolism , Glutathione Transferase/drug effects , Glutathione Transferase/metabolism , Glyceraldehyde 3-Phosphate Dehydrogenase (NADP+)/drug effects , Glyceraldehyde 3-Phosphate Dehydrogenase (NADP+)/metabolism , Lipid Peroxidation , Liver/drug effects , Mice , Mice, Inbred BALB C , Protein Carbonylation/drug effectsABSTRACT
ETHNOPHARMACOLOGICAL RELEVANCE: Coix seed has been traditionally used to treat cancers in folk medicine. AIM OF THE STUDY: Study the anticancer action mechanism of Coix seed extract. MATERIALS AND METHODS: After the treatment with Coix seed extract (10 microl/ml), the residual activity of fatty acid synthase (FAS) as overall reaction, beta-ketoacyl reduction, enoyl reduction, and acetyl acetyl coenzyme A (AcAcCoA) reduction was separately detected at 340 nm in the UV-190 spectrophotometer. After rats were administrated Coix seed extract (2.5, 5.0, and 10.0 ml/kg) intragastrically for 10 days consecutively, activities of FAS, malate dehydrogenase (MDH), lipid protein lipase (LPL), hepatic lipase (HL), triglyceride (TG), and glucose-6-phosphate dehydrogenase (G-6-PD) in the plasma, liver and fatty tissues were determined. RESULTS: Experiments in vitro showed that the inhibition of Coix seed extract on FAS activity was significant and dose dependent, and two active sites inhibited were beta-ketoacyl reductases (KR) and enoyl reductase (ER). Experiments in vivo showed that Coix seed extract inhibited FAS activity in the liver, and elevated LPL and HL activity in the plasma, and effected G-6-PD activity. CONCLUSIONS: The study supports that FAS is a novel target for anticancer activity, and provides a theoretical foundation for the wide application of Coix seed extract in traditional medicine.
Subject(s)
Antineoplastic Agents, Phytogenic/pharmacology , Coix/chemistry , Fatty Acid Synthases/drug effects , Plant Extracts/pharmacology , 3-Oxoacyl-(Acyl-Carrier-Protein) Reductase , Alcohol Oxidoreductases/antagonists & inhibitors , Animals , Antineoplastic Agents, Phytogenic/administration & dosage , Antineoplastic Agents, Phytogenic/isolation & purification , Dose-Response Relationship, Drug , Drug Delivery Systems , Ducks , Fatty Acid Synthases/metabolism , Glucosephosphate Dehydrogenase/drug effects , Glucosephosphate Dehydrogenase/metabolism , Lipase/drug effects , Lipase/metabolism , Lipoprotein Lipase/drug effects , Lipoprotein Lipase/metabolism , Liver/drug effects , Liver/enzymology , Medicine, Traditional , Plant Extracts/administration & dosage , Rats , Rats, Wistar , Seeds , Triglycerides/metabolismABSTRACT
Gene therapy with adenoviral vectors is a promising new approach for the treatment of refractory advanced breast cancer. Strategies to restrict adenoviral-mediated therapeutic gene expression are important to avoid harming normal cells. Fatty acid synthase (FAS) is overexpressed in several human cancers. FAS is highly expressed in infiltrating breast cancer tissue, and always associated with malignant phenotypes and poor prognosis. In this study, expression of the FAS was evaluated in three breast cancer cell lines. A 680 bp-FAS promoter was cloned and its transcriptional activity was analyzed in breast cancer cell lines. We made a recombinant adenovirus construct carrying herpes simplex virus thymidine kinase (HSV-TK) driven by human FAS promoter (Ad-FAS-TK) and analyzed its target cytotoxicity in vitro and in vivo against human breast cancer cells combined with prodrug ganciclovir (GCV). The results show that the expression of FAS varies in the three breast cancer cell lines examined (respectively, SK-Br3>MCF-7>MDA-MB-231), but FAS promoter can initiate relative high transcriptional activities in all three kinds of cancer cells while little in normal fibroblast cells. Furthermore, FAS promoter can drive the therapeutic gene in a wider range of human breast cancers than cerbB2 promoter and exhibit a stronger activity than midkine (MK) promoter. Combination of Ad-FAS-TK and GCV treatment exhibited strong-targeted cytotoxic effect on breast cancer cells but showed little activity in normal fibroblast cells. The tumorigenic capability of breast cancer cells treated with Ad-FAS-TK/GCV was completely inhibited in vitro and in vivo assays. In conclusion, adenoviral-mediated suicide gene therapy controlled by tumor associated-FAS promoter can induce specific cytotoxic effect on human breast cancer cells in vitro and in vivo. So it is a promising target for the development of gene therapy against breast cancers.
Subject(s)
Adenoviruses, Human , Breast Neoplasms/genetics , Fatty Acid Synthases/therapeutic use , Genetic Therapy , Recombinant Fusion Proteins/therapeutic use , Tumor Suppressor Proteins/therapeutic use , Antineoplastic Agents/therapeutic use , Breast Neoplasms/drug therapy , Breast Neoplasms/therapy , DNA-Binding Proteins/administration & dosage , DNA-Binding Proteins/therapeutic use , Fatty Acid Synthases/drug effects , Female , Ganciclovir/therapeutic use , Gene Expression , Genes, Transgenic, Suicide , Genetic Vectors , Humans , In Vitro Techniques , Nuclear Proteins/administration & dosage , Nuclear Proteins/therapeutic use , Phenotype , Prognosis , Recombinant Fusion Proteins/administration & dosage , Treatment Failure , Treatment Outcome , Tumor Suppressor Proteins/administration & dosageABSTRACT
Although antipsychotics are established drugs in schizophrenia treatment, they are admittedly known to induce side effects favoring the onset of obesity and worsening its complications. Despite potential involvement of histamine receptor antagonism, or of other neurotransmitter systems, the mechanism by which antipsychotic drugs increase body weight is not elucidated. The aim of the present study was to investigate whether chronic antipsychotic treatments can directly alter the regulation of two main functions of white adipose tissue: lipolysis and glucose utilization. The influence of a classical antipsychotic (haloperidol) was compared to that of two atypical antipsychotics, one known to favor weight gain (olanzapine), the other not (ziprasidone). Cell size, lipolytic capacity and glucose transport activity were determined in white adipocytes of rats subjected to 5-week oral treatment with these antipsychotics. Gene expression of adipocyte proteins involved in glucose transport or fat storage and mobilization, such as glucose transporters (GLUT1 and GLUT4), leptin, matrix metallo-proteinase-9 (MMP9), hormone-sensitive lipase (HSL) and fatty acid synthase (FAS) was also evaluated. Adipocytes from chronic olanzapine-treated rats exhibited decreased lipolytic activity, lowered HSL expression and increased FAS expression. These changes were concomitant to enlarged fat deposition and adipocyte size. Alterations were observed in adipocytes from olanzapine-treated rats whereas the other antipsychotics did not induce any notable disorder. Our results therefore show evidence of an effect of chronic antipsychotic treatment on rat adipocyte metabolism. Thus, impairment of fat cell lipolysis should be considered as a side effect of certain antipsychotics, leading, along with the already documented hyperphagia, to the excessive weight gain observed in patients under prolonged treatment..
Subject(s)
Adipocytes/drug effects , Antipsychotic Agents/pharmacology , Lipid Metabolism/drug effects , Weight Gain/drug effects , Adipocytes/cytology , Adipocytes/metabolism , Animals , Benzodiazepines/pharmacology , Cell Size/drug effects , Drug Administration Schedule , Fatty Acid Synthases/drug effects , Fatty Acid Synthases/genetics , Fatty Acid Synthases/metabolism , Gene Expression Regulation/drug effects , Glucose Transport Proteins, Facilitative/drug effects , Glucose Transport Proteins, Facilitative/genetics , Glucose Transport Proteins, Facilitative/metabolism , Haloperidol/pharmacology , Male , Obesity/chemically induced , Obesity/metabolism , Olanzapine , Piperazines/pharmacology , RNA/analysis , Rats , Rats, Sprague-Dawley , Statistics, Nonparametric , Sterol Esterase/drug effects , Sterol Esterase/genetics , Sterol Esterase/metabolism , Thiazoles/pharmacologyABSTRACT
High levels of fatty acid synthase (FAS) expression have been observed in several cancers, including breast, prostate, colon and lung carcinoma, compared with their respective normal tissue. We present data that show high levels of FAS protein in human and rat glioma cell lines and human glioma tissue samples, as compared to normal rat astrocytes and normal human brain. Incubating glioma cells with the FAS inhibitor cerulenin decreased endogenous fatty acid synthesis by approximately 50%. Cell cycle analysis demonstrated a time- and dose-dependent increase in S-phase cell arrest following cerulenin treatment for 24 h. Further, treatment with cerulenin resulted in time- and dose-dependent decreases in glioma cell viability, as well as reduced clonogenic survival. Increased apoptotic cell death and PARP cleavage were observed in U251 and SNB-19 cells treated with cerulenin, which was independent of the death receptor pathway. Overexpressing Bcl-2 inhibited cerulenin-mediated cell death. In contrast, primary rat astrocytes appeared unaffected. Finally, RNAi-mediated knockdown of FAS leading to reduced FAS enzymatic activity was associated with decreased glioma cell viability. These findings suggest that FAS might be a novel target for antiglioma therapy.
Subject(s)
Brain Neoplasms/enzymology , Brain/enzymology , Fatty Acid Synthases/metabolism , Glioma/enzymology , Animals , Apoptosis/drug effects , Apoptosis/physiology , Astrocytes/drug effects , Base Sequence , Blotting, Western , Brain Neoplasms/drug therapy , Cell Line, Tumor , Cerulenin/pharmacology , Collagen Type XI/drug effects , Collagen Type XI/metabolism , Dose-Response Relationship, Drug , Enzyme Inhibitors/pharmacology , Fatty Acid Synthases/drug effects , Fatty Acid Synthases/genetics , Flow Cytometry , Glioma/drug therapy , Humans , Molecular Sequence Data , Proto-Oncogene Proteins c-bcl-2/metabolism , RNA, Small Interfering , Radiation Tolerance/drug effects , Rats , TransfectionABSTRACT
AMP-activated protein kinase (AMPK) is a key sensor and regulator of intracellular and whole-body energy metabolism. We have identified a thienopyridone family of AMPK activators. A-769662 directly stimulated partially purified rat liver AMPK (EC50 = 0.8 microM) and inhibited fatty acid synthesis in primary rat hepatocytes (IC50 = 3.2 microM). Short-term treatment of normal Sprague Dawley rats with A-769662 decreased liver malonyl CoA levels and the respiratory exchange ratio, VCO2/VO2, indicating an increased rate of whole-body fatty acid oxidation. Treatment of ob/ob mice with 30 mg/kg b.i.d. A-769662 decreased hepatic expression of PEPCK, G6Pase, and FAS, lowered plasma glucose by 40%, reduced body weight gain and significantly decreased both plasma and liver triglyceride levels. These results demonstrate that small molecule-mediated activation of AMPK in vivo is feasible and represents a promising approach for the treatment of type 2 diabetes and the metabolic syndrome.
Subject(s)
Diabetes Mellitus, Type 2/drug therapy , Enzyme Activators/chemistry , Enzyme Activators/therapeutic use , Metabolic Syndrome/drug therapy , Multienzyme Complexes/metabolism , Protein Serine-Threonine Kinases/metabolism , Pyrones/chemistry , Pyrones/therapeutic use , Thiophenes/chemistry , Thiophenes/therapeutic use , AMP-Activated Protein Kinases , Animals , Biphenyl Compounds , Cell Line , Diabetes Mellitus, Type 2/metabolism , Disease Models, Animal , Dose-Response Relationship, Drug , Enzyme Activation/drug effects , Enzyme Activation/physiology , Enzyme Activators/pharmacology , Fatty Acid Synthases/drug effects , Fatty Acid Synthases/genetics , Fatty Acid Synthases/metabolism , Glucose-6-Phosphatase/drug effects , Glucose-6-Phosphatase/genetics , Glucose-6-Phosphatase/metabolism , Hepatocytes/drug effects , Hepatocytes/metabolism , Humans , In Vitro Techniques , Metabolic Syndrome/metabolism , Metformin/chemistry , Metformin/pharmacology , Metformin/therapeutic use , Mice , Mice, Obese , Molecular Weight , Multienzyme Complexes/drug effects , Phosphoenolpyruvate Carboxykinase (GTP)/drug effects , Phosphoenolpyruvate Carboxykinase (GTP)/genetics , Phosphoenolpyruvate Carboxykinase (GTP)/metabolism , Protein Serine-Threonine Kinases/drug effects , Pyrones/pharmacology , RNA, Messenger/drug effects , RNA, Messenger/genetics , RNA, Messenger/metabolism , Rats , Rats, Sprague-Dawley , Thiophenes/pharmacologyABSTRACT
OBJECTIVE: This study evaluated the physiologic properties of high protein buckwheat flour (PBF) by examining its effects on serum cholesterol and body fat in rats and on cholesterol gallstone formation in mice. METHODS: Animals were fed experimental diets that contained casein, buckwheat protein extract (BWP), or PBF as a protein source (net protein content 200 g/kg). RESULTS: In experiment 1, consumption of PBF and BWP for 10 d caused 33% and 31% decreases, respectively, in serum cholesterol of rats fed cholesterol-enriched diets when compared with consumption of casein (P < 0.05). Dietary PBF caused a significant decrease in liver cholesterol, whereas dietary BWP caused only a slight decrease (P > 0.05). Fecal excretion of neutral and acidic steroids in the PBF group was significantly higher than those in the BWP and casein groups. In experiment 2, consumption of PBF for 10 d significantly suppressed adipose tissue weight and hepatic activity of fatty acid synthase in rats fed cholesterol-free diets compared with consumption of casein (P < 0.05), whereas that of BWP for this period caused only a slight decrease in adipose tissue weight (P > 0.05). In experiment 3, dietary PBF and BWP significantly decreased the incidence of cholesterol gallstones and lithogenic index in mice fed cholesterol-enriched diets for 27 d, which was associated with increased fecal excretion of acidic steroids. CONCLUSION: This study demonstrated that PBF has strong activities against hypercholesterolemia, obesity, and gallstone formation, suggesting a potential usefulness of PBF as functional ingredient.
Subject(s)
Adipose Tissue/metabolism , Cholesterol/blood , Dietary Proteins/administration & dosage , Digestion , Fagopyrum , Gallstones/prevention & control , Hypercholesterolemia/prevention & control , Animals , Bile/chemistry , Body Composition/drug effects , Caseins/administration & dosage , Cholesterol, Dietary/administration & dosage , Cholesterol, Dietary/metabolism , Dietary Proteins/pharmacology , Fatty Acid Synthases/drug effects , Fatty Acid Synthases/metabolism , Feces/chemistry , Flour , Gallstones/epidemiology , Liver/metabolism , Male , Mice , Rats , Rats, Sprague-Dawley , Steroids/analysisABSTRACT
In our previous study, we demonstrated that retrograded starch, a kind of resistant starch, of beans reduced serum lipid levels in rats. In this study, we examined whether retrograded starch in potato pulps could reduce serum lipid concentrations. Rats were given diets containing 15 g of retrograded starch in potato pulps from the Benimaru potato (BM) or Hokkaikogane potato (HK) in a 100 g diet for 4 wk. At the 4th week, the total cholesterol level in the serum in the BM group and serum triglyceride (TG) level in the HK group were significantly lower than those in the control group. In the BM group, the contents of fecal bile acids were significantly higher than those in the control group. On the other hand, in the HK group, the hepatic mRNA level of fatty acid synthase (FAS) was significantly lower than that in the control group. The FAS mRNA level correlated with the mRNA level of sterol regulatory element-binding protein-1c (SREBP-1c), a regulator of expression of FAS, positively. These results suggested that BM pulp promoted the excretion of bile acids, which resulted in a low concentration of serum cholesterol. On the other hand, HK pulp inhibited the synthesis of fatty acids at the mRNA levels of FAS and SREBP-1c, which might lead to a reduction of the serum TG level.
Subject(s)
Cholesterol/blood , Solanum tuberosum , Starch/pharmacology , Triglycerides/blood , Animals , Bile Acids and Salts/metabolism , Body Weight/drug effects , Cholestanol/metabolism , Diet , Fatty Acid Synthases/drug effects , Fatty Acid Synthases/metabolism , Feces , Hydrogen-Ion Concentration , Liver/drug effects , Liver/metabolism , Male , Organ Size/drug effects , RNA, Messenger/drug effects , RNA, Messenger/metabolism , Rats , Rats, Wistar , Starch/administration & dosage , Sterol Regulatory Element Binding Protein 1/drug effects , Sterol Regulatory Element Binding Protein 1/metabolism , Time FactorsABSTRACT
Adipocyte determination and differentiation-dependent factor 1 (ADD1) drives the expression of several lipogenic genes in mammals. Polyunsaturated fatty acids decrease ADD1 mRNA abundance in differentiating porcine adipocytes. The current study was designed to explore the mechanisms by which PUFA inhibit the expression of ADD1 in porcine adipocytes. Porcine preadipocytes were differentiated for 24 h with 0 or 100 microM of docosahexaenoic acid (DHA) and mixtures of different concentrations of antioxidants to investigate the effect of DHA and antioxidants on the ADD1 mRNA abundance. We found the relative mRNA abundance was decreased by the addition of 100 microM DHA to the medium for porcine differentiating adipocytes, and adding an antioxidant mixture to the medium prevented part of the decrease in ADD1 mRNA abundance. These data suggest that DHA decreased the steady-state transcription factor ADD1 mRNA through a mechanism related to fatty acid peroxidation. Indeed, adding 7.5 microM vitamin E (a natural antioxidant) also restored the concentrations of ADD1 and fatty acid synthase mRNA, which were decreased by DHA treatment; however, the DHA or the antioxidant treatment did not change the expression of antioxidation genes (superoxide dismutase 1 and glutathione peroxidase 1) in porcine stromal vascular cells. When supplemented with the eicosanoid synthesis pathway inhibitors, the inhibition of the expression of ADD1 by arachidonic acid was partially recovered. These results suggest that the mechanism by which PUFA decrease ADD1 mRNA is due to the metabolic product of eicosanoids and peroxidation of these PUFA.