RESUMO
Sterol regulatory element-binding protein (SREBP) transcription factors are central regulators of lipid homeostasis and are essential for lipid metabolic reprogramming that supports tumor growth in multiple cancers. SREBP pathway inhibitors have been identified, but bioavailable compounds are lacking. To address this need, we designed a novel approach for screening a collection of 4,474 FDA-approved drugs. SREBPs are conditionally essential and required under low lipid conditions. Leveraging this property, we screened for drugs that inhibited pancreatic cancer cell growth in lipid-poor, but not lipid-rich, medium. The primary screen identified 83 drugs that inhibited cell growth in a lipid-dependent manner. Secondary assays examining SREBP target gene expression, SREBP proteolytic cleavage, and effects on human breast cancer cells identified 13 FDA-approved drugs that inhibit SREBP pathway activation. Taken together, we demonstrated that our screening approach can identify SREBP inhibitors from a small library of compounds. This high-throughput screening platform enables screening of large compound collections to discover novel small molecule SREBP inhibitors.
Assuntos
Ensaios de Triagem em Larga Escala , Proteínas de Ligação a Elemento Regulador de Esterol , Humanos , Ensaios de Triagem em Larga Escala/métodos , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Linhagem Celular Tumoral , Transdução de Sinais/efeitos dos fármacos , Estados Unidos , Bibliotecas de Moléculas Pequenas/farmacologia , Bibliotecas de Moléculas Pequenas/química , Aprovação de Drogas , Proliferação de Células/efeitos dos fármacos , United States Food and Drug Administration , Antineoplásicos/farmacologiaRESUMO
Three sesquiterpenoids were isolated and purified from the 95% ethanol extract of Atractylodis Macrocephalae Rhizoma by column chromatography on silica gel, Sephadex LH-20, ODS, and high-performance liquid chromatography(HPLC). Their chemical structures were identified on the basis of spectroscopic analysis and physiochemical properties as(7Z)-8ß,13-diacetoxy-eudesma-4(15),7(11)-diene(1), 7-oxo-7,8-secoeudesma-4(15),11-dien-8-oic acid(2), and guai-10(14)-en-11-ol(3). Compounds 1 and 2 are new compounds and compound 3 was obtained from Compositae family for the first time. Compounds 1, 2, and 3 showed weak inhibitory activities against sterol regulatory element-binding proteins(SREBPs).
Assuntos
Atractylodes , Medicamentos de Ervas Chinesas , Sesquiterpenos de Eudesmano , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Atractylodes/química , Medicamentos de Ervas Chinesas/química , Rizoma/química , Sesquiterpenos de Eudesmano/análise , Sesquiterpenos de Eudesmano/farmacologiaRESUMO
AIMS: Potential anti-obesity effects of quinizarin, a plant anthraquinone, were investigated using 3 T3-L1 preadipocyte cells and high-fat diet (HD)-induced obese mice. MAIN METHOD: Cell viability was determined using the MTT assay. Triglyceride (TG) and lipid accumulation were determined using a TG assay kit and Oil Red O staining, respectively. Adipogenic, lipogenic, and lipolytic gene and protein expression was measured by RT-PCR or Western blot. Serum biochemical indices, including cholesterol and blood glucose, in HD-fed obese mice were determined using corresponding assay kits. Histological analysis was performed with haematoxylin and eosin (H&E) staining. RESULTS: Quinizarin (0-10 µM) significantly reduced intracellular TG and lipid droplets during the differentiation of preadipocytes. Quinizarin significantly suppressed the expression of adipocyte differentiation marker proteins, such as CCAAT/enhancer-binding protein ß (C/EBP-ß), C/EBP-α, PPAR-γ, and aP2, and lipogenic marker proteins, including SREBP1c, SREBP2, fatty acid synthase (FAS), and acetyl-CoA carboxylase 1 (ACC1), reduced ACC2 expression and increased carnitine palmitoyltransferase 1 (CPT1) expression. Oral administration of quinizarin (15-30 mg/kg/day) to HD-fed mice for 6 weeks reduced the body weight gain and size of liver adipocytes and epididymal fat tissues, with significant reductions in liver TG and serum total cholesterol, blood glucose, LDL, and HDL levels. SIGNIFICANCE: The results of this study indicated that quinizarin exerts anti-obesity effects by inhibiting both adipogenesis and lipogenesis and stimulating lipolysis in vitro and in vivo mainly by downregulating the SREBP signalling pathway; thus, it might be a potent candidate as a health-beneficial food or therapeutic agent to prevent or treat obesity.
Assuntos
Adipócitos/metabolismo , Antraquinonas/farmacologia , Proteínas Estimuladoras de Ligação a CCAAT/antagonistas & inibidores , Diferenciação Celular/fisiologia , Lipogênese/fisiologia , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Células 3T3-L1 , Adipócitos/efeitos dos fármacos , Adipogenia/efeitos dos fármacos , Adipogenia/fisiologia , Animais , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Diferenciação Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/fisiologia , Lipogênese/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismoRESUMO
Vitamin D3 metabolites inhibit the expression of lipogenic genes by impairing sterol regulatory element-binding protein (SREBP), a master transcription factor of lipogenesis, independent of their canonical activity through a vitamin D receptor (VDR). Herein, we designed and synthesized a series of vitamin D derivatives to search for a drug-like small molecule that suppresses the SREBP-induced lipogenesis without affecting the VDR-controlled calcium homeostasis in vivo. Evaluation of the derivatives in cultured cells and mice led to the discovery of VDR-silent SREBP inhibitors and to the development of KK-052 (50), the first vitamin D-based SREBP inhibitor that has been demonstrated to mitigate hepatic lipid accumulation without calcemic action in mice. KK-052 maintained the ability of 25-hydroxyvitamin D3 to induce the degradation of SREBP but lacked in the VDR-mediated activity. KK-052 serves as a valuable compound for interrogating SREBP/SCAP in vivo and may represent an unprecedented translational opportunity of synthetic vitamin D analogues.
Assuntos
Desenho de Fármacos , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo , Vitamina D/análogos & derivados , Animais , Peso Corporal/efeitos dos fármacos , Células CHO , Cricetinae , Cricetulus , Reação de Cicloadição , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Fígado Gorduroso/tratamento farmacológico , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Lipogênese/efeitos dos fármacos , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Receptores de Calcitriol/antagonistas & inibidores , Receptores de Calcitriol/metabolismo , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Proteínas de Ligação a Elemento Regulador de Esterol/genética , Vitamina D/metabolismo , Vitamina D/farmacologia , Vitamina D/uso terapêuticoRESUMO
Regulatory T cells (Treg cells) are essential for immune tolerance1, but also drive immunosuppression in the tumour microenvironment2. Therapeutic targeting of Treg cells in cancer will therefore require the identification of context-specific mechanisms that affect their function. Here we show that inhibiting lipid synthesis and metabolic signalling that are dependent on sterol-regulatory-element-binding proteins (SREBPs) in Treg cells unleashes effective antitumour immune responses without autoimmune toxicity. We find that the activity of SREBPs is upregulated in intratumoral Treg cells. Moreover, deletion of SREBP-cleavage-activating protein (SCAP)-a factor required for SREBP activity-in these cells inhibits tumour growth and boosts immunotherapy that is triggered by targeting the immune-checkpoint protein PD-1. These effects of SCAP deletion are associated with uncontrolled production of interferon-γ and impaired function of intratumoral Treg cells. Mechanistically, signalling through SCAP and SREBPs coordinates cellular programs for lipid synthesis and inhibitory receptor signalling in these cells. First, de novo fatty-acid synthesis mediated by fatty-acid synthase (FASN) contributes to functional maturation of Treg cells, and loss of FASN from Treg cells inhibits tumour growth. Second, Treg cells in tumours show enhanced expression of the PD-1 gene, through a process that depends on SREBP activity and signals via mevalonate metabolism to protein geranylgeranylation. Blocking PD-1 or SREBP signalling results in dysregulated activation of phosphatidylinositol-3-kinase in intratumoral Treg cells. Our findings show that metabolic reprogramming enforces the functional specialization of Treg cells in tumours, pointing to new ways of targeting these cells for cancer therapy.
Assuntos
Metabolismo dos Lipídeos , Neoplasias/imunologia , Neoplasias/metabolismo , Transdução de Sinais , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/imunologia , Animais , Colesterol/metabolismo , Ácido Graxo Sintases/metabolismo , Ácidos Graxos/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Ácido Mevalônico/metabolismo , Camundongos , Fosfatidilinositol 3-Quinase/metabolismo , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/metabolismo , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo , Linfócitos T Reguladores/enzimologia , Regulação para CimaRESUMO
Cancer is a multifaceted global disease. Transformation of a normal to a malignant cell takes several steps, including somatic mutations, epigenetic alterations, metabolic reprogramming and loss of cell growth control. Recently, the mevalonate pathway has emerged as a crucial regulator of tumor biology and a potential therapeutic target. This pathway controls cholesterol production and posttranslational modifications of Rho-GTPases, both of which are linked to several key steps of tumor progression. Inhibitors of the mevalonate pathway induce pleiotropic antitumor-effects in several human malignancies, identifying the pathway as an attractive candidate for novel therapies. In this review, we will provide an overview about the role and regulation of the mevalonate pathway in certain aspects of cancer initiation and progression and its potential for therapeutic intervention in oncology.
Assuntos
Antineoplásicos/farmacologia , Transformação Celular Neoplásica/metabolismo , Colesterol/biossíntese , Ácido Mevalônico/metabolismo , Neoplasias/metabolismo , Antineoplásicos/uso terapêutico , Proliferação de Células , Transformação Celular Neoplásica/efeitos dos fármacos , Transformação Celular Neoplásica/patologia , Ensaios Clínicos como Assunto , Progressão da Doença , Geraniltranstransferase/antagonistas & inibidores , Geraniltranstransferase/metabolismo , Humanos , Hidroximetilglutaril-CoA Redutases/metabolismo , Redes e Vias Metabólicas/efeitos dos fármacos , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo , Resultado do Tratamento , Proteínas rho de Ligação ao GTP/metabolismoRESUMO
HMG-CoA reductase (HMGCR) is the rate-limiting enzyme in the cholesterol biosynthetic pathway, and is the target of cholesterol-lowering drugs, statins. Previous studies have demonstrated that the enzyme activity is regulated by sterol-induced degradation in addition to transcriptional regulation through sterol-regulatory-element-binding proteins (SREBPs). While 25-hydroxycholesterol induces both HMGCR degradation and SREBP inhibition in a nonselective manner, lanosterol selectively induces HMGCR degradation. Here, to clarify the structural determinants of selectivity for the two activities, we established a luciferase-based assay monitoring HMGCR degradation and used it to profile the structure-activity/selectivity relationships of oxysterols and (oxy)lanosterols. We identified several sterols that selectively induce HMGCR degradation and one sterol, 25-hydroxycholest-4-en-3-one, that selectively inhibits the SREBP pathway. These results should be helpful in designing more potent and selective HMGCR degraders.
Assuntos
Hidroximetilglutaril-CoA Redutases/metabolismo , Lanosterol/metabolismo , Oxisteróis/metabolismo , Células HEK293 , Humanos , Lanosterol/farmacologia , Estrutura Molecular , Oxisteróis/farmacologia , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo , Relação Estrutura-AtividadeRESUMO
Background Trypanosoma cruzi is an intracellular parasite that causes debilitating chronic Chagas cardiomyopathy (CCM), for which there is no effective drug or vaccine. Previously, we demonstrated increased cardiac lipid accumulation and endoplasmic reticulum stress in mice with CCM. Increased endoplasmic reticulum stress may lead to uncontrolled SREBP (sterol regulatory element-binding protein) activation and lipotoxicity in the myocardium during the intermediate stage of infection and result in progression to chronic CCM. Therefore, we investigated whether inhibiting SREBP activation modulates CCM progression in T cruzi-infected mice. Methods and Results T cruzi-infected cultured cardiomyocytes (3:1 multiplicity of infection; 24 hours postinfection) were incubated with betulin (3 µmol/L per mL), an SREBP inhibitor, for 24 hours. Quantitative polymerase chain reaction and Western blotting analyses demonstrated a significant reduction in SREBP activation, lipid biosynthesis, and endoplasmic reticulum stress in betulin-treated infected cells compared with untreated cells. T cruzi infected (103 trypomastigotes of the Brazil strain) Swiss mice were fed a customized diet containing betulin during the intermediate stage (40 days postinfection) until the chronic stage (120 DPI). Cardiac ultrasound imaging and histological and biochemical analyses demonstrated anatomical and functional improvements in betulin-treated, infected mice compared with untreated controls: we observed a significant reduction in cholesterol/fatty acid synthesis that may result in the observed cardiac reduction in cardiac lipid accumulation, mitochondrial and endoplasmic reticulum stress, and ventricular enlargement. Conclusions Our study (in vitro and vivo) demonstrates that inhibition of cardiac SREBP activation reduces cardiac damage during T cruzi infection and modulates CCM in a murine Chagas model.
Assuntos
Cardiomiopatia Chagásica/tratamento farmacológico , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Metabolismo dos Lipídeos/efeitos dos fármacos , Miócitos Cardíacos/efeitos dos fármacos , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Triterpenos/farmacologia , Trypanosoma cruzi/patogenicidade , Animais , Linhagem Celular , Cardiomiopatia Chagásica/metabolismo , Cardiomiopatia Chagásica/parasitologia , Cardiomiopatia Chagásica/patologia , Doença Crônica , Modelos Animais de Doenças , Interações Hospedeiro-Parasita , Masculino , Camundongos Endogâmicos C3H , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/parasitologia , Miócitos Cardíacos/patologia , Ratos , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismoRESUMO
Vitamin D3 metabolites are capable of controlling gene expression in mammalian cells through two independent pathways: vitamin D receptor (VDR) and sterol regulatory element-binding protein (SREBP) pathways. In the present study, we dissect the complex biological activity of vitamin D by designing synthetic vitamin D3 analogs specific for VDR or SREBP pathway, i.e., a VDR activator that lacks SREBP inhibitory activity, or an SREBP inhibitor devoid of VDR activity. These synthetic vitamin D probes permitted identification of one of the vitamin D-responsive genes, Soat1, as an SREBP-suppressed gene. The chemical probes developed in the present study may prove useful in dissecting the intricate interplay of vitamin D actions, thereby providing insights into how vitamin D target genes are regulated.
Assuntos
Sondas Moleculares/química , Vitamina D/farmacologia , Descoberta de Drogas , Humanos , Receptores de Calcitriol/metabolismo , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo , Vitamina D/metabolismoRESUMO
Covalent conjugates of multiple nutrients often exhibit greater biological activities than each individual nutrient and more predictable safety profiles than completely unnatural chemical entities. Here, we report the construction and application of a focused chemical library of 308 covalent conjugates of a variety of small-molecule nutrients. Screening of the library with a reporter gene of sterol regulatory element-binding protein (SREBP), a master regulator of mammalian lipogenesis, led to the discovery of a conjugate of docosahexaenoic acid (DHA), glucosamine, and amino acids as an inhibitor of SREBP (molecule 1, DHG). Mechanistic analyses indicate that molecule 1 impairs the SREBP activity by inhibiting glucose transporters and thereby activating AMP-activated protein kinase (AMPK). Oral administration of molecule 1 suppressed the intestinal absorption of glucose in mice. These results suggest that such synthetic libraries of nutrient conjugates serve as a source of novel chemical tools and pharmaceutical seeds that modulate energy metabolism.
Assuntos
Nutrientes/farmacologia , Bibliotecas de Moléculas Pequenas/farmacologia , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Aminoácidos/síntese química , Aminoácidos/farmacologia , Animais , Células CACO-2 , Ácidos Graxos Insaturados/síntese química , Ácidos Graxos Insaturados/farmacologia , Genes Reporter , Glucosamina/síntese química , Glucosamina/farmacologia , Glucose/metabolismo , Humanos , Absorção Intestinal/efeitos dos fármacos , Masculino , Camundongos Endogâmicos ICR , Nutrientes/síntese química , Bibliotecas de Moléculas Pequenas/síntese química , Proteínas de Ligação a Elemento Regulador de Esterol/genética , Vitaminas/síntese química , Vitaminas/farmacologiaRESUMO
Sterol regulatory element-binding proteins (SREBPs) are transcription factors that regulate the expression of genes involved in fatty acid and cholesterol biosynthetic pathways. The present study showed that the flavonoid chrysin impairs the fatty acid synthase promoter. Chrysin reduces the expression of SREBP target genes, such as fatty acid synthase, in human hepatoma Huh-7 cells and impairs de novo synthesis of fatty acids and cholesterol. Moreover, it reduces the endogenous mature, transcriptionally active forms of SREBPs, which are generated by the proteolytic processing of precursor forms. In addition, chrysin reduces the enforced expressing mature forms of SREBPs and their transcriptional activity. The ubiquitin-proteasome system is not involved in the chrysin-mediated reduction of SREBPs mature forms. These results suggest that chrysin suppresses SREBP activity, at least partially, via the degradation of SREBPs mature forms. Abbreviations: ACC1: acetyl-CoA carboxylase 1; DMEM: Dulbecco's modified Eagle's medium; FAS: fatty acid synthase; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; 25-HC: 25-hydroxycholesterol; HMGCS: HMG-CoA synthase; LDH: lactate dehydrogenase; LPDS: lipoprotein-deficient serum; PI3K: phosphatidylinositol 3-kinase; SCD1: stearoyl-CoA desaturase; SREBPs: sterol regulatory element-binding proteins.
Assuntos
Flavonoides/farmacologia , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo , Animais , Linhagem Celular Tumoral , Colesterol/biossíntese , Ácido Graxo Sintases/genética , Ácidos Graxos/biossíntese , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Regiões Promotoras Genéticas , Proteólise , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Proteínas de Ligação a Elemento Regulador de Esterol/genéticaRESUMO
Nonalcoholic steatohepatitis (NASH) is a rapidly growing cause of chronic liver damage, cirrhosis, and hepatocellular carcinoma. How fatty liver pathogenesis is subject to epigenetic regulation is unknown. We hypothesized that chromatin remodeling is important for the pathogenesis of fatty liver disease. AT-rich interactive domain-containing protein 1A (ARID1A), a DNA-binding component of the SWItch/sucrose nonfermentable adenosine triphosphate-dependent chromatin-remodeling complex, contributes to nucleosome repositioning and access by transcriptional regulators. Liver-specific deletion of Arid1a (Arid1a liver knockout [LKO]) caused the development of age-dependent fatty liver disease in mice. Transcriptome analysis revealed up-regulation of lipogenesis and down-regulation of fatty acid oxidation genes. As evidence of direct regulation, ARID1A demonstrated direct binding to the promoters of many of these differentially regulated genes. Additionally, Arid1a LKO mice were more susceptible to high-fat diet-induced liver steatosis and fibrosis. We deleted Pten in combination with Arid1a to synergistically drive fatty liver progression. Inhibition of lipogenesis using CAT-2003, a potent sterol regulatory element-binding protein inhibitor, mediated improvements in markers of fatty liver disease progression in this Arid1a/Pten double knockout model. Conclusion: ARID1A plays a role in the epigenetic regulation of hepatic lipid homeostasis, and its suppression contributes to fatty liver pathogenesis. Combined Arid1a and Pten deletion shows accelerated fatty liver disease progression and is a useful mouse model for studying therapeutic strategies for NASH.
Assuntos
Proteínas de Ligação a DNA/deficiência , Lipogênese , Fígado/metabolismo , Hepatopatia Gordurosa não Alcoólica/etiologia , Fatores de Transcrição/deficiência , Animais , Proteínas de Ligação a DNA/antagonistas & inibidores , Proteínas de Ligação a DNA/genética , Ácidos Graxos/metabolismo , Camundongos , Camundongos Knockout , Terapia de Alvo Molecular , Hepatopatia Gordurosa não Alcoólica/metabolismo , Oxirredução , PTEN Fosfo-Hidrolase/deficiência , PTEN Fosfo-Hidrolase/genética , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genéticaRESUMO
Fatostatin, a chemical inhibitor of the sterol regulatory elementbinding protein (SREBP) pathway, has been reported to possess high antitumor activity against prostate and pancreatic cancer. The main aim of the present study was to investigate the effects and mechanism of fatostatin in endometrial carcinoma (EC). In the present study, we determined that fatostatin inhibited EC cell viability and colony formation capacity, decreased the invasive and migratory capacities of EC cells, induced EC cell cycle arrest at the G2/M phase and stimulated caspasemediated apoptosis of EC cells. In addition, fatostatin significantly decreased the protein expression levels of nuclear SREBPs and their downstream genes and increased the protein expression levels of cleaved caspase9, caspase3 and PARP in EC cells. In addition, the mRNA expression levels of SREBPcontrolled downstream genes were also significantly downregulated. The quantification assays of fatty acids and total cholesterol revealed that the levels of free fatty acids and total cholesterol in EC cells were decreased. The present study indicated that fatostatin exhibited antitumor effects by blocking SREBPregulated metabolic pathways and inducing caspasemediated apoptosis in EC and may be a potent therapeutic strategy for the treatment of EC.
Assuntos
Proliferação de Células/efeitos dos fármacos , Neoplasias do Endométrio/tratamento farmacológico , Piridinas/administração & dosagem , Proteínas de Ligação a Elemento Regulador de Esterol/genética , Tiazóis/administração & dosagem , Apoptose/efeitos dos fármacos , Caspases/genética , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Neoplasias do Endométrio/genética , Neoplasias do Endométrio/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Redes e Vias Metabólicas/efeitos dos fármacos , Células-Tronco/efeitos dos fármacos , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidoresRESUMO
There is increased expression of liver x receptor (LXR) target genes and reduced low density lipoprotein receptor (LDLR) during spontaneous luteolysis in primates. The LXRs are nuclear receptors that increase cholesterol efflux by inducing transcription of their target genes. Transcription of LDLR is regulated by sterol regulatory element binding proteins (SREBPs). Human chorionic gonadotropin (hCG) prevents luteolysis and stimulates progesterone synthesis via protein kinase A (PKA). Thus, our primary objectives are: 1) Determine the effects of LXR activation and SREBP inhibition on progesterone secretion and cholesterol metabolism, and 2) Determine whether hCG signaling via PKA regulates transcription of LXR and SREBP target genes in human luteinized granulosa cells. Basal and hCG-stimulated progesterone secretion was significantly decreased by the combined actions of the LXR agonist T0901317 and the SREBP inhibitor fatostatin, which was associated with reduced intracellular cholesterol storage. Expression of LXR target genes in the presence of T0901317 was significantly reduced by hCG, while hCG promoted transcriptional changes that favor LDL uptake. These effects of hCG were reversed by a specific PKA inhibitor. A third objective was to resolve a dilemma concerning LXR regulation of steroidogenic acute regulatory protein (STAR) expression in primate and non-primate steroidogenic cells. T0901317 induced STAR expression and progesterone synthesis in ovine, but not human cells, revealing a key difference between species in LXR regulation of luteal function. Collectively, these data support the hypothesis that LXR-induced cholesterol efflux and reduced LDL uptake via SREBP inhibition mediates luteolysis in primates, which is prevented by hCG.
Assuntos
Gonadotropina Coriônica/farmacologia , Receptores X do Fígado/metabolismo , Células Lúteas/metabolismo , Progesterona/metabolismo , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo , Animais , Colesterol/metabolismo , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Hidrocarbonetos Fluorados/farmacologia , Receptores X do Fígado/antagonistas & inibidores , Receptores X do Fígado/genética , Células Lúteas/efeitos dos fármacos , Modelos Biológicos , Fosfoproteínas/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , Receptores de LDL/metabolismo , Ovinos , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Proteínas de Ligação a Elemento Regulador de Esterol/genética , Sulfonamidas/farmacologia , Tiazóis/farmacologia , Transcrição Gênica/efeitos dos fármacosRESUMO
Nowadays, more and more attention has been paid to osteoporosis caused by diabetes mellitus. Elevated levels of pro-inflammatory cytokines in diabetic patients activate the activity of osteoclasts through the RANKL/OPG pathway. The nuclear transcription factor SREBP2, a master regulator of cholesterol metabolism, has been found involved in osteoclastogenesis. In our previous study, we have identified anhydroicaritin as a potent inhibitor of transcription factor SREBPs, which improves dyslipidemia and insulin resistance. In this study, we demonstrated that anhydroicaritin could also decrease the level of SREBP2 and its target genes in osteoclasts induced by RANKL without significant cytotoxicity. Moreover, anhydroicaritin suppressed RANKL-induced osteoclasts differentiation. In STZ-induced diabetic mice model, we found that the osteoclasts were largely increased accompanied with deterioration of bone structure. Anhydroicaritin decreased the level of blood glucose and alleviated insulin resistance. More importantly, anhydroicaritin inhibited osteoclast differentiation and rescued diabetes-induced bone loss in vivo. In conclusion, anhydroicaritin, a potent SREBP2 inhibitor, inhibits the osteoclasts formation and improves diabetes-induced bone loss.
Assuntos
Benzopiranos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Diabetes Mellitus Experimental/complicações , Osteoclastos/efeitos dos fármacos , Osteoporose/patologia , Ligante RANK/farmacologia , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Animais , Benzopiranos/uso terapêutico , Linhagem Celular , Camundongos , Osteoclastos/citologia , Osteoclastos/patologia , Osteoporose/complicações , Osteoporose/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacosRESUMO
Flaviviruses are positive-strand RNA viruses distributed all over the world that infect millions of people every year and for which no specific antiviral agents have been approved. These viruses include the mosquito-borne West Nile virus (WNV), which is responsible for outbreaks of meningitis and encephalitis. Considering that nordihydroguaiaretic acid (NDGA) has been previously shown to inhibit the multiplication of the related dengue virus and hepatitis C virus, we have evaluated the effect of NDGA, and its methylated derivative tetra-O-methyl nordihydroguaiaretic acid (M4N), on the infection of WNV. Both compounds inhibited the infection of WNV, likely by impairing viral replication. Since flavivirus multiplication is highly dependent on host cell lipid metabolism, the antiviral effect of NDGA has been previously related to its ability to disturb the lipid metabolism, probably by interfering with the sterol regulatory element-binding proteins (SREBP) pathway. Remarkably, we observed that other structurally unrelated inhibitors of the SREBP pathway, such as PF-429242 and fatostatin, also reduced WNV multiplication, supporting that the SREBP pathway may constitute a druggable target suitable for antiviral intervention against flavivirus infection. Moreover, treatment with NDGA, M4N, PF-429242, and fatostatin also inhibited the multiplication of the mosquito-borne flavivirus Zika virus (ZIKV), which has been recently associated with birth defects (microcephaly) and neurological disorders. Our results point to SREBP inhibitors, such as NDGA and M4N, as potential candidates for further antiviral development against medically relevant flaviviruses.
Assuntos
Antivirais/farmacologia , Masoprocol/análogos & derivados , Masoprocol/farmacologia , Febre do Nilo Ocidental/tratamento farmacológico , Vírus do Nilo Ocidental/crescimento & desenvolvimento , Infecção por Zika virus/tratamento farmacológico , Zika virus/crescimento & desenvolvimento , Animais , Linhagem Celular , Chlorocebus aethiops , Células HeLa , Humanos , Metabolismo dos Lipídeos/efeitos dos fármacos , Piridinas/farmacologia , Pirrolidinas/farmacologia , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Tiazóis/farmacologia , Células Vero , Replicação Viral/efeitos dos fármacos , Vírus do Nilo Ocidental/efeitos dos fármacos , Zika virus/efeitos dos fármacosRESUMO
The therapeutic use of statins has been associated to a reduced risk of Parkinson's disease (PD) and may hold neuroprotective potential by counteracting the degeneration of dopaminergic neurons. Transcriptional activation of the sterol regulatory element-binding protein (SREBP) is one of the major downstream signaling pathways triggered by the cholesterol-lowering effect of statins. In a previous study in neuroblastoma cells, we have shown that statins consistently induce the upregulation of presynaptic dopaminergic proteins and changes of their function and these effects were accompanied by downstream activation of SREBP. In this study, we aimed to determine the direct role of SREBP pathway in the modulation of dopaminergic phenotype. We demonstrate that treatment of SH-SY5Y cells with U18666A, an SREBP activator, increases the translocation of SREBPs into the nucleus, increases the expression of SREBP-1, SREBP-2, and of the presynaptic dopaminergic markers such as vesicular monoamine transporter 2, synaptic vesicle glycoprotein 2 A and 2 C, synaptogyrin-3, and tyrosine hydroxylase. The addition of SREBP inhibitor, PF-429242, blocks the increase of U18666A-induced expression of SREBPs and presynaptic markers. Our results, in line with previously reported effects of statins, demonstrate that direct stimulation of SREBP translocation is associated to differentiation toward a dopaminergic-like phenotype and suggest that SREBP-mediated transcriptional activity may lead to the restoration of the presynaptic dopamine markers and may contribute to neuroprotection of dopaminergic neurons. These findings further support the potential protective role of statin in PD and shed light upon SREBP as a potential new target for developing disease-modifying treatment in PD.
Assuntos
Androstenos/farmacologia , Dopaminérgicos/farmacologia , Neurônios Dopaminérgicos/efeitos dos fármacos , Terminações Pré-Sinápticas/efeitos dos fármacos , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Anticolesterolemiantes/farmacologia , Linhagem Celular Tumoral , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Neurônios Dopaminérgicos/citologia , Neurônios Dopaminérgicos/fisiologia , Expressão Gênica/efeitos dos fármacos , Humanos , Glicoproteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Terminações Pré-Sinápticas/fisiologia , Pirrolidinas/farmacologia , RNA Mensageiro/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Sinaptogirinas/metabolismo , Proteínas Vesiculares de Transporte de Monoamina/metabolismoRESUMO
Cholesterol is important for normal brain function. The brain synthesizes its own cholesterol, presumably in astrocytes. We have previously shown that diabetes results in decreased brain cholesterol synthesis by a reduction in sterol regulatory element-binding protein 2 (SREBP2)-regulated transcription. Here we show that coculture of control astrocytes with neurons enhances neurite outgrowth, and this is reduced with SREBP2 knockdown astrocytes. In vivo, mice with knockout of SREBP2 in astrocytes have impaired brain development and behavioral and motor defects. These mice also have altered energy balance, altered body composition, and a shift in metabolism toward carbohydrate oxidation driven by increased glucose oxidation by the brain. Thus, SREBP2-mediated cholesterol synthesis in astrocytes plays an important role in brain and neuronal development and function, and altered brain cholesterol synthesis may contribute to the interaction between metabolic diseases, such as diabetes and altered brain function.
Assuntos
Astrócitos/metabolismo , Composição Corporal/fisiologia , Encéfalo/metabolismo , Colesterol/metabolismo , Metabolismo Energético/fisiologia , Proteína de Ligação a Elemento Regulador de Esterol 2/deficiência , Animais , Composição Corporal/genética , Linhagem Celular Tumoral , Metabolismo Energético/genética , Feminino , Técnicas de Silenciamento de Genes , Glioma/patologia , Glucose/metabolismo , Hiperinsulinismo/metabolismo , Masculino , Aprendizagem em Labirinto , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Comportamento de Nidação , Neuritos/ultraestrutura , Oxirredução , Ratos , Teste de Desempenho do Rota-Rod , Proteína de Ligação a Elemento Regulador de Esterol 2/genética , Proteína de Ligação a Elemento Regulador de Esterol 2/fisiologia , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Proteínas de Ligação a Elemento Regulador de Esterol/genéticaRESUMO
Sterol regulatory element-binding proteins (SREBPs) are transcription factors that control lipid homeostasis. SREBP activation is regulated by a negative feedback loop in which sterols bind to SREBP cleavage-activating protein (SCAP), an escort protein essential for SREBP activation, or to insulin-induced genes (Insigs) (endoplasmic reticulum [ER] anchor proteins), sequestering the SREBP-SCAP-Insig complex in the ER. We screened a chemical library of endogenous molecules and identified 25-hydroxyvitamin D (25OHD) as an inhibitor of SREBP activation. Unlike sterols and other SREBP inhibitors, 25OHD impairs SREBP activation by inducing proteolytic processing and ubiquitin-mediated degradation of SCAP, thereby decreasing SREBP levels independently of the vitamin D receptor. Vitamin D supplementation has been proposed to reduce the risk of metabolic diseases, but the mechanisms are unknown. The present results suggest a previously unrecognized molecular mechanism of vitamin D-mediated lipid control that might be useful in the treatment of metabolic diseases.
Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Metabolismo dos Lipídeos/efeitos dos fármacos , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Vitamina D/análogos & derivados , Animais , Células CHO , Células Cultivadas , Cricetulus , Relação Dose-Resposta a Droga , Regulação para Baixo/efeitos dos fármacos , Células HEK293 , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Conformação Molecular , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas de Ligação a Elemento Regulador de Esterol/genética , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo , Relação Estrutura-Atividade , Vitamina D/química , Vitamina D/metabolismo , Vitamina D/farmacologiaRESUMO
The formation of the myelin membrane of the oligodendrocyte in the CNS is a fundamental process requiring the coordinated synthesis of many different components. The myelin membrane is particularly rich in lipids, however, the regulation of this lipid synthesis is not understood. In other cell types, including Schwann cells, the myelin-forming cells of the PNS, lipid synthesis is tightly regulated by the sterol regulatory element-binding protein (SREBP) family of transcription factors, but this has not been previously shown in oligodendrocytes. We investigated SREBPs' role during oligodendrocyte differentiation in vitro. Both SREBP-1 and SREBP-2 were expressed in oligodendrocyte precursor cells and differentiating oligodendrocytes. Using the selective site-1 protease (S1P) inhibitor PF-429242, which inhibits the cleavage of SREBP precursor forms into mature forms, we found that preventing SREBP processing inhibited process growth and reduced the expression level of myelin basic protein, a major component of myelin. Further, process extension deficits could be rescued by the addition of exogenous cholesterol. Blocking SREBP processing reduced mRNA transcription and protein levels of SREBP target genes involved in both the fatty acid and the cholesterol synthetic pathways. Furthermore, de novo levels and total levels of cholesterol synthesis were greatly diminished when SREBP processing was inhibited. Together these results indicate that SREBPs are important regulators of oligodendrocyte maturation and that perturbation of their activity may affect myelin formation and integrity. Cover Image for this issue: doi: 10.1111/jnc.13781.