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1.
Nat Commun ; 12(1): 1889, 2021 03 25.
Artigo em Inglês | MEDLINE | ID: mdl-33767172

RESUMO

Plasma low-density lipoprotein (LDL) is primarily cleared by LDL receptor (LDLR). LDLR can be proteolytically cleaved to release its soluble ectodomain (sLDLR) into extracellular milieu. However, the proteinase responsible for LDLR cleavage is unknown. Here we report that membrane type 1-matrix metalloproteinase (MT1-MMP) co-immunoprecipitates and co-localizes with LDLR and promotes LDLR cleavage. Plasma sLDLR and cholesterol levels are reduced while hepatic LDLR is increased in mice lacking hepatic MT1-MMP. Opposite effects are observed when MT1-MMP is overexpressed. MT1-MMP overexpression significantly increases atherosclerotic lesions, while MT1-MMP knockdown significantly reduces cholesteryl ester accumulation in the aortas of apolipoprotein E (apoE) knockout mice. Furthermore, sLDLR is associated with apoB and apoE-containing lipoproteins in mouse and human plasma. Plasma levels of sLDLR are significantly increased in subjects with high plasma LDL cholesterol levels. Thus, we demonstrate that MT1-MMP promotes ectodomain shedding of hepatic LDLR, thereby regulating plasma cholesterol levels and the development of atherosclerosis.


Assuntos
Apolipoproteína B-100/sangue , Apolipoproteínas E/sangue , Aterosclerose/patologia , Lipoproteínas LDL/sangue , Metaloproteinase 14 da Matriz/metabolismo , Receptores de LDL/metabolismo , Animais , Apolipoproteínas E/genética , Linhagem Celular Tumoral , Ésteres do Colesterol/metabolismo , Dependovirus/genética , Feminino , Células HEK293 , Células Hep G2 , Humanos , Masculino , Metaloproteinase 14 da Matriz/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout
2.
Int J Mol Sci ; 22(4)2021 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-33557318

RESUMO

Previous studies on ablation of several key genes of meibogenesis related to fatty acid elongation, omega oxidation, and esterification into wax esters have demonstrated that inactivation of any of them led to predicted changes in the meibum lipid profiles and caused severe abnormalities in the ocular surface and Meibomian gland (MG) physiology and morphology. In this study, we evaluated the effects of Soat1 ablation that were expected to cause depletion of the second largest class of Meibomian lipids (ML)-cholesteryl esters (CE)-in a mouse model. ML of the Soat1-null mice were examined using liquid chromatography high-resolution mass spectrometry and compared with those of Soat1+/- and wild-type mice. Complete suppression of CE biosynthesis and simultaneous accumulation of free cholesterol (Chl) were observed in Soat1-null mice, while Soat1+/- mutants had normal Chl and CE profiles. The total arrest of the CE biosynthesis in response to Soat1 ablation transformed Chl into the dominant lipid in meibum accounting for at least 30% of all ML. The Soat1-null mice had clear manifestations of dry eye and MG dysfunction. Enrichment of meibum with Chl and depletion of CE caused plugging of MG orifices, increased meibum rigidity and melting temperature, and led to a massive accumulation of lipid deposits around the eyes of Soat1-null mice. These findings illustrate the role of Soat1/SOAT1 in the lipid homeostasis and pathophysiology of MG.


Assuntos
Ésteres do Colesterol/metabolismo , Modelos Animais de Doenças , Disfunção da Glândula Tarsal/patologia , Glândulas Tarsais/patologia , Esterol O-Aciltransferase/fisiologia , Lágrimas/metabolismo , Animais , Homeostase , Masculino , Disfunção da Glândula Tarsal/etiologia , Disfunção da Glândula Tarsal/metabolismo , Glândulas Tarsais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout
3.
Artigo em Inglês | MEDLINE | ID: mdl-33010453

RESUMO

Lipid droplets (LDs) are ubiquitous fat storage organelles composed of a neutral lipid core, comprising triacylglycerols (TAG) and sterol esters (SEs), surrounded by a phospholipid monolayer membrane with several decorating proteins. Recently, LD biology has come to the foreground of research due to their importance for energy homeostasis and cellular stress response. As aberrant LD accumulation and lipid depletion are hallmarks of numerous diseases, addressing LD biogenesis and turnover provides a new framework for understanding disease-related mechanisms. Here we discuss the potential role of LDs in neurodegeneration, while making some predictions on how LD imbalance can contribute to pathophysiology in the brain.


Assuntos
Doença de Alzheimer/metabolismo , Autofagia/genética , Demência Frontotemporal/metabolismo , Gotículas Lipídicas/metabolismo , Lipólise/genética , Atrofia Muscular Espinal/metabolismo , Sistema Nervoso/metabolismo , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Ésteres do Colesterol/metabolismo , Demência Frontotemporal/genética , Demência Frontotemporal/patologia , Regulação da Expressão Gênica , Humanos , Lisossomos/metabolismo , Atrofia Muscular Espinal/genética , Atrofia Muscular Espinal/patologia , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Sistema Nervoso/patologia , Neuroglia/metabolismo , Neuroglia/patologia , Neurônios/metabolismo , Neurônios/patologia , Fosfolipídeos/metabolismo , Triglicerídeos/metabolismo
4.
Artigo em Inglês | MEDLINE | ID: mdl-33031913

RESUMO

We previously demonstrated that AKR vs. DBA/2 mouse bone marrow derived macrophages have higher levels of free cholesterol and lower levels of esterified cholesterol after cholesterol loading, and that AKR, but not DBA/2, macrophages induced C/EBP homologous protein (CHOP) expression after cholesterol loading. We earlier determined that the free and esterified cholesterol level effect is due to a truncation in the sterol O-acyltransferase 1 (Soat1) gene, encoding acetyl-coenzyme A acetyltransferase 1 (ACAT1). Here we examined the mechanism for the differential induction of CHOP by cholesterol loading. CHOP was induced in both strains after incubation with tunicamycin, indicating both strains have competent endoplasmic reticulum stress pathways. CHOP was induced when DBA/2 macrophages were cholesterol loaded in the presence of an ACAT inhibitor, indicating that the difference in free cholesterol levels were responsible for this strain effect. This finding was confirmed in macrophages derived from DBA/2 embryonic stem cells. Cholesterol loading of Soat1 gene edited cells, mimicking the AKR allele, led to increased free cholesterol levels and restored CHOP induction. The upstream pathway of free cholesterol induced endoplasmic reticulum stress was investigated; and, RNA-dependent protein kinase-like endoplasmic reticulum kinase (PERK) and inositol-requiring enzyme 1 α protein kinase (IRE1α) pathways were required for maximal CHOP expression.


Assuntos
Colesterol/farmacologia , Estresse do Retículo Endoplasmático/genética , Macrófagos/metabolismo , Esterol O-Aciltransferase/genética , Fator de Transcrição CHOP/genética , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Colesterol/metabolismo , Ésteres do Colesterol/metabolismo , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/efeitos dos fármacos , Células-Tronco Embrionárias/metabolismo , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Endorribonucleases/genética , Endorribonucleases/metabolismo , Feminino , Fêmur/citologia , Fêmur/metabolismo , Regulação da Expressão Gênica , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Masculino , Camundongos Endogâmicos AKR , Camundongos Endogâmicos DBA , Camundongos Knockout para ApoE , Cultura Primária de Células , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Especificidade da Espécie , Esterol O-Aciltransferase/metabolismo , Fator de Transcrição CHOP/metabolismo , Tunicamicina/farmacologia , eIF-2 Quinase/genética , eIF-2 Quinase/metabolismo
5.
Artigo em Inglês | MEDLINE | ID: mdl-33069870

RESUMO

Although most mammalian fatty acids (FAs) are straight-chain, there also exist branched-chain FAs such as iso- and anteiso-FAs, especially in the meibomian glands. Meibum lipids, which are secreted from the meibomian glands and are important for dry eye prevention, contain abundant branched-chain lipids, such as cholesteryl esters and wax esters with chain-lengths of ≥C21 (very-long-chain; VLC). However, the exact tissue distribution of branched-chain lipids or the enzymes involved in the production of branched-chain VLCFAs has remained poorly understood. Here, we revealed that FA elongases ELOVL1, ELOVL3, and ELOVL7, of the seven mammalian ELOVL isozymes, elongated saturated branched-chain acyl-CoAs. ELOVL3 was highly active toward iso-C17:0 and anteiso-C17:0 acyl-CoAs and elongated them up to iso-C23:0 and anteiso-C25:0 acyl-CoAs, respectively. ELOVL1 elongated both iso- and anteiso-C23:0 acyl-CoAs to C25:0 acyl-CoAs. By establishing a liquid chromatography-tandem mass spectrometry method capable of separating branched- and straight-chain lipids, we showed that essentially all of the cholesteryl esters and 88% of the wax esters in the mouse meibomian glands are branched. In Elovl1 mutant mice, the levels of ≥C24:0 branched-chain cholesteryl esters and ≥C25:0 branched-chain wax esters were decreased, indicating that ELOVL1 indeed elongates branched-chain VLC acyl-CoAs in vivo. In addition, substantial amounts of ceramides containing branched-chain FAs were present in the skin, meibomian glands, and liver. Our findings provide new insights into the molecular mechanisms that create FA and lipid diversity.


Assuntos
Acil Coenzima A/metabolismo , Aminoácidos de Cadeia Ramificada/metabolismo , Elongases de Ácidos Graxos/metabolismo , Glândulas Tarsais/metabolismo , Aciltransferases/genética , Aciltransferases/metabolismo , Aldeído Oxirredutases/genética , Aldeído Oxirredutases/metabolismo , Aminoácidos de Cadeia Ramificada/classificação , Animais , Ceramidas/classificação , Ceramidas/metabolismo , Ésteres do Colesterol/classificação , Ésteres do Colesterol/metabolismo , Elongases de Ácidos Graxos/genética , Regulação da Expressão Gênica , Células HEK293 , Humanos , Lipidômica/métodos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Pele/metabolismo , Esterol O-Aciltransferase/genética , Esterol O-Aciltransferase/metabolismo , Especificidade por Substrato
6.
Am J Physiol Endocrinol Metab ; 319(1): E102-E104, 2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32369415

RESUMO

Glucocorticoids belong to the superfamily of steroid hormones that are synthesized from the common precursor cholesterol. Adrenal gland-derived glucocorticoids, e.g., cortisol in humans and corticosterone in rodents, contribute to various processes essential for normal daily life. Glucocorticoid deficiency, also referred to as primary adrenal insufficiency, therefore, often becomes evident early in life and can be present with hypoglycemia, a failure to thrive, recurrent development of infections, and neurological problems, such as seizures and coma. The majority of congenital primary adrenal insufficiency cases are caused by deleterious mutations in genes involved in the intracellular mobilization of cholesterol and the subsequent conversion of cholesterol into glucocorticoids. A significant number of glucocorticoid deficiency cases, however, cannot be explained by known genetic variations. This perspective highlights existing literature regarding the importance of lipoprotein-derived cholesterol acquisition through scavenger receptor class B, type I (SR-BI/SCARB1) for the maintenance of an optimal adrenal glucocorticoid function in mice and humans. On the basis of the reviewed findings, it is suggested that the SCARB1 gene should be included in the standard glucocorticoid deficiency genetic screening panel to 1) facilitate knowledge development on the relative contribution of SR-BI-mediated cholesterol acquisition to steroid hormone synthesis in humans and 2) open up the possibility to reclassify glucocorticoid deficiency patients without a currently known genetic cause for concomitant treatment optimization.


Assuntos
Doença de Addison/genética , Colesterol/metabolismo , Glucocorticoides/biossíntese , Receptores Depuradores Classe B/genética , Doença de Addison/congênito , Doença de Addison/diagnóstico , Doença de Addison/metabolismo , Animais , Ésteres do Colesterol/metabolismo , Testes Genéticos , Humanos , Lipoproteínas HDL/metabolismo , Camundongos , Camundongos Knockout , Receptores Depuradores Classe B/deficiência , Receptores Depuradores Classe B/metabolismo
7.
BMC Mol Cell Biol ; 21(1): 32, 2020 Apr 22.
Artigo em Inglês | MEDLINE | ID: mdl-32321446

RESUMO

BACKGROUND: Neutral cholesterol ester hydrolase 1 (NCEH1) catalyzes the hydrolysis of cholesterol ester (CE) in macrophages. Genetic ablation of NCEH1 promotes CE-laden macrophages and the development of atherosclerosis in mice. Dysregulation of NCEH1 levels is involved in the pathogenesis of multiple disorders including metabolic diseases and atherosclerosis; however, relatively little is known regarding the mechanisms regulating NCEH1. Retinoic acid receptor-related orphan receptor α (RORα)-deficient mice exhibit several phenotypes indicative of aberrant lipid metabolism, including dyslipidemia and increased susceptibility to atherosclerosis. RESULTS: In this study, inhibition of lipid droplet formation by RORα positively regulated NCEH1 expression in macrophages. In mammals, the NCEH1 promoter region was found to harbor putative RORα response elements (ROREs). Electrophoretic mobility shift, chromatin immunoprecipitation, and luciferase reporter assays showed that RORα binds and responds to ROREs in human NCEH1. Moreover, NCEH1 was upregulated through RORα via a phorbol myristate acetate-dependent mechanism during macrophage differentiation from THP1 cells. siRNA-mediated knockdown of RORα significantly downregulated NCEH1 expression and accumulated lipid droplets in human hepatoma cells. In contrast, NCEH1 expression and removal of lipid droplets were induced by RORα agonist treatments and RORα overexpression in macrophages. CONCLUSION: These data strongly suggested that NCEH1 is a direct RORα target, defining potential new roles for RORα in the inhibition of lipid droplet formation through NCEH1.


Assuntos
Gotículas Lipídicas/metabolismo , Macrófagos/metabolismo , Receptores do Ácido Retinoico/metabolismo , Esterol Esterase/metabolismo , Animais , Aterosclerose/enzimologia , Aterosclerose/etiologia , Aterosclerose/genética , Aterosclerose/metabolismo , Linhagem Celular Tumoral , Ésteres do Colesterol/metabolismo , LDL-Colesterol/farmacologia , Imunoprecipitação da Cromatina , Técnicas de Silenciamento de Genes , Humanos , Macrófagos/enzimologia , Camundongos , Camundongos Knockout , Regiões Promotoras Genéticas , Ligação Proteica , RNA Interferente Pequeno , Receptores do Ácido Retinoico/agonistas , Receptores do Ácido Retinoico/genética , Esterol Esterase/genética , Acetato de Tetradecanoilforbol/farmacologia , Regulação para Cima
8.
Cancer Sci ; 111(7): 2275-2283, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32314495

RESUMO

Nasopharyngeal carcinoma (NPC) is a common malignant tumor and a major cause of mortality and morbidity in southern China. However, the mechanism is still elusive. Here, we focused on studying the role of squalene epoxidase (SQLE), a key enzyme of cholesterol biosynthesis, in the progression of NPC. Clinical study revealed that SQLE expression was significantly upregulated in NPC tissues compared to normal tissues from mRNA level and patients with high expression of SQLE showed a poor prognosis. In vitro experiments showed that SQLE overexpression led to a significant proliferation of cells whereas SQLE knockdown showed an opposite result. In vivo studies also showed that SQLE promoted tumor growth in nude mice. Further study revealed that SQLE promoted NPC proliferation by cholesteryl ester accumulation instead of cholesterol. Mechanism studies indicated that cholesteryl ester promoted NPC cell proliferation by activating the PI3K/AKT pathway and inhibition of this pathway in SQLE-overexpressed or cholesteryl ester-treated cells resulted in a significant reduction of NPC cell proliferation. These results indicate that the oncogenic effect of SQLE in NPC mainly resulted from cholesteryl ester accumulation and PI3K/AKT is a promising target for NPC with SQLE overexpression.


Assuntos
Ésteres do Colesterol/metabolismo , Carcinoma Nasofaríngeo/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Esqualeno Mono-Oxigenase/metabolismo , Animais , Antineoplásicos/farmacologia , Apoptose/genética , Biomarcadores Tumorais , Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos Nus , Carcinoma Nasofaríngeo/genética , Carcinoma Nasofaríngeo/mortalidade , Carcinoma Nasofaríngeo/patologia , Prognóstico , Esqualeno Mono-Oxigenase/antagonistas & inibidores , Esqualeno Mono-Oxigenase/genética , Ensaios Antitumorais Modelo de Xenoenxerto
9.
Mikrochim Acta ; 187(4): 225, 2020 03 13.
Artigo em Inglês | MEDLINE | ID: mdl-32170399

RESUMO

Synthesis and functionalization of magnetite nanoparticles (Fe3O4) was achieved with the view to covalently bind both cholesterol oxidase and cholesterol esterase biorecognition agents for the development of free and total cholesterol biosensors. Prior to enzyme attachment, Fe3O4 was functionalized with 3-aminopropyltriethoxysilane (APTES) and polyamidoamine (PAMAM) dendrimer. Characterization of the material was performed by FT-IR and UV spectroscopy, SEM/EDX surface analysis and electrochemical investigations. The response to cholesterol and its palmitate ester was examined using cyclic voltammetry. Optimum analytical performance for the free cholesterol biosensor was obtained using APTES-functionalized magnetite with a sensitivity of 101.9 µA mM-1 cm-2, linear range 0.1-1 mM and LOD of 80 µM when operated at 37 °C. In the case of the total cholesterol biosensor, the best analytical performance was obtained using PAMAM dendrimer-modified magnetite with sensitivity of 73.88 µA mM-1 cm-2 and linear range 0.1-1.5 mM, with LOD of 90 µM. A stability study indicated that the free cholesterol biosensors retained average activity of 98% after 25 days while the total cholesterol biosensors retained 85% activity upon storage over the same period. Graphical abstract Schematic representation of cholesterol esterase and oxidase loaded magnetic nanoparticles (Fe3O4@APTES or Fe3O4@APTES-PAMAM) generating hydrogen peroxide from cholesterol palmitate.


Assuntos
Técnicas Biossensoriais , Ésteres do Colesterol/análise , Colesterol/análise , Técnicas Eletroquímicas , Nanopartículas de Magnetita/química , Colesterol/metabolismo , Ésteres do Colesterol/metabolismo , Colesterol Oxidase/química , Colesterol Oxidase/metabolismo , Humanos , Estrutura Molecular , Esterol Esterase/química , Esterol Esterase/metabolismo
10.
Artigo em Inglês | MEDLINE | ID: mdl-32061751

RESUMO

Carboxyl ester lipase (Cel), is a lipolytic enzyme secreted by the pancreas, which hydrolyzes various species of lipids in the gut. Cel is also secreted by mammary gland during lactation and exists in breast milk. It facilitates dietary fat digestion and absorption, thus contributing to normal infant development. This study aimed to examine whether the Cel in zebrafish embryos has a similar role of maternal lipid utilization as in human infants, and how Cel contributes to the utilization of yolk lipids in zebrafish. The cel1 and cel2 genes were expressed ubiquitously in the blastodisc and yolk syncytial layer before 24 hpf, and in the exocrine pancreas after 72 hpf. The cel1 and cel2 morphants exhibited developmental retardation and yolk sac retention. The total cholesterol, cholesterol ester, free cholesterol, and triglyceride were reduced in the morphants' body while accumulated in the yolk (except triglyceride). The FFA content of whole embryos was much lower in morphants than in standard controls. Moreover, the delayed development in cel (cel1/cel2) double morphants was partially rescued by FFA and cholesterol supplementation. Delayed and weakened cholesterol ester transport to the brain and eyes was observed in cel morphants. Correspondingly, shrunken midbrain tectum, microphthalmia, pigmentation-delayed eyes as well as down-regulated Shh target genes were observed in the CNS of double morphants. Interestingly, cholesterol injections reversed these CNS alterations. Our findings suggested that cel genes participate in the lipid releasing from yolk sac to developing body, thereby contributing to the normal growth rate and CNS development in zebrafish.


Assuntos
Carboxilesterase/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Transtornos do Crescimento/genética , Saco Vitelino/enzimologia , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/embriologia , Animais , Animais Geneticamente Modificados , Carboxilesterase/genética , Sistema Nervoso Central/embriologia , Colesterol/metabolismo , Ésteres do Colesterol/metabolismo , Modelos Animais de Doenças , Embrião não Mamífero , Desenvolvimento Embrionário , Técnicas de Silenciamento de Genes , Transtornos do Crescimento/embriologia , Transtornos do Crescimento/enzimologia , Proteínas Hedgehog/metabolismo , Humanos , Metabolismo dos Lipídeos , Morfolinos/administração & dosagem , Morfolinos/genética , Pâncreas Exócrino/embriologia , Pâncreas Exócrino/enzimologia , Triglicerídeos/metabolismo , Saco Vitelino/embriologia , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética
11.
Neuron ; 105(5): 837-854.e9, 2020 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-31902528

RESUMO

Loss-of-function (LOF) variants of TREM2, an immune receptor expressed in microglia, increase Alzheimer's disease risk. TREM2 senses lipids and mediates myelin phagocytosis, but its role in microglial lipid metabolism is unknown. Combining chronic demyelination paradigms and cell sorting with RNA sequencing and lipidomics, we find that wild-type microglia acquire a disease-associated transcriptional state, while TREM2-deficient microglia remain largely homeostatic, leading to neuronal damage. TREM2-deficient microglia phagocytose myelin debris but fail to clear myelin cholesterol, resulting in cholesteryl ester (CE) accumulation. CE increase is also observed in APOE-deficient glial cells, reflecting impaired brain cholesterol transport. This finding replicates in myelin-treated TREM2-deficient murine macrophages and human iPSC-derived microglia, where it is rescued by an ACAT1 inhibitor and LXR agonist. Our studies identify TREM2 as a key transcriptional regulator of cholesterol transport and metabolism under conditions of chronic myelin phagocytic activity, as TREM2 LOF causes pathogenic lipid accumulation in microglia.


Assuntos
Encéfalo/metabolismo , Colesterol/metabolismo , Macrófagos/metabolismo , Glicoproteínas de Membrana/genética , Microglia/metabolismo , Bainha de Mielina/metabolismo , Fagocitose/genética , Receptores Imunológicos/genética , Acetil-CoA C-Acetiltransferase/antagonistas & inibidores , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Animais , Ésteres do Colesterol/metabolismo , Modelos Animais de Doenças , Citometria de Fluxo , Humanos , Células-Tronco Pluripotentes Induzidas , Metabolismo dos Lipídeos/genética , Lipidômica , Receptores X do Fígado/agonistas , Camundongos , Camundongos Knockout , Camundongos Knockout para ApoE , RNA-Seq
12.
PLoS One ; 15(1): e0228024, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31978092

RESUMO

Abnormal accumulation of acyl-CoA cholesterol acyltransferase-1 (ACAT-1) mediated cholesterol ester has been shown to contribute to cancer progression in various cancers including leukemia, glioma, breast, pancreatic and prostate cancers. However, the significance of ACAT-1 and cholesterol esters (CE) is relatively understudied in ovarian cancer. In this in vitro study, we assessed the expression and contribution of ACAT-1 in ovarian cancer progression. We observed a significant increase in the expression of ACAT-1 and CE levels in a panel of ovarian cancer cell lines (OC-314, SKOV-3 and IGROV-1) compared to primary ovarian epithelial cells (normal controls). To confirm the tumor promoting capacity of ACAT-1, we inhibited ACAT-1 expression and activity by treating our cell lines with an ACAT inhibitor, avasimibe, or by stable transfection with ACAT-1 specific short hairpin RNA (shRNA). We observed significant suppression of cell proliferation, migration and invasion in ACAT-1 knockdown ovarian cancer cell lines compared to their respective controls (cell lines transfected with scrambled shRNA). ACAT-1 inhibition enhanced apoptosis with a concurrent increase in caspases 3/7 activity and decreased mitochondrial membrane potential. Increased generation of reactive oxygen species (ROS) coupled with increased expression of p53 may be the mechanism(s) underlying pro-apoptotic action of ACAT-1 inhibition. Additionally, ACAT-1 inhibited ovarian cancer cell lines displayed enhanced chemosensitivity to cisplatin treatment. These results suggest ACAT-1 may be a potential new target for the treatment of ovarian cancer.


Assuntos
Progressão da Doença , Neoplasias Ovarianas/enzimologia , Neoplasias Ovarianas/patologia , Esterol O-Aciltransferase/metabolismo , Acetil-CoA C-Acetiltransferase , Apoptose/efeitos dos fármacos , Caspase 3/metabolismo , Caspase 7/metabolismo , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Ésteres do Colesterol/metabolismo , Cisplatino/farmacologia , Feminino , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Invasividade Neoplásica , RNA Interferente Pequeno/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Esterol O-Aciltransferase/antagonistas & inibidores , Ensaio Tumoral de Célula-Tronco
13.
Artigo em Inglês | MEDLINE | ID: mdl-31863970

RESUMO

Scavenger receptor class B type I (SR-BI) mediates the selective uptake of cholesteryl esters (CE) from high-density lipoproteins (HDL). An impaired SR-BI function leads to hyperalphalipoproteinemia with elevated levels of cholesterol transported in the HDL fraction. Accumulation of cholesterol in apolipoprotein B (apoB)-containing lipoproteins has been shown to alter skin lipid composition and barrier function in mice. To investigate whether these hypercholesterolemic effects on the skin also occur in hyperalphalipoproteinemia, we compared skins of wild-type and SR-BI knockout (SR-BI-/-) mice. SR-BI deficiency did not affect the epidermal cholesterol content and induced only minor changes in the ceramide subclasses. The epidermal free fatty acid (FFA) pool was, however, enriched in short and unsaturated chains. Plasma CE levels strongly correlated with epidermal FFA C18:1 content. The increase in epidermal FFA coincided with downregulation of cholesterol and FFA synthesis genes, suggesting a compensatory response to increased flux of plasma cholesterol and FFAs into the skin. Importantly, the SR-BI-/- epidermal lipid barrier showed increased permeability to ethyl-paraminobenzoic acid, indicating an impairment of the barrier function. In conclusion, increased HDL-cholesterol levels in SR-BI-/- mice can alter the epidermal lipid composition and lipid barrier function similarly as observed in hypercholesterolemia due to elevated levels of apoB-containing lipoproteins.


Assuntos
Proteínas de Transferência de Ésteres de Colesterol/deficiência , Epiderme/metabolismo , Erros Inatos do Metabolismo Lipídico/metabolismo , Ácido 4-Aminobenzoico/farmacocinética , Animais , Apolipoproteínas B/metabolismo , Antígenos CD36/genética , Proteínas de Transferência de Ésteres de Colesterol/genética , Proteínas de Transferência de Ésteres de Colesterol/metabolismo , Ésteres do Colesterol/sangue , Ésteres do Colesterol/metabolismo , Epiderme/patologia , Ácidos Graxos Insaturados/metabolismo , Feminino , Lecitinas/genética , Lecitinas/metabolismo , Erros Inatos do Metabolismo Lipídico/genética , Erros Inatos do Metabolismo Lipídico/patologia , Camundongos , Camundongos Endogâmicos C57BL
14.
Nat Rev Mol Cell Biol ; 21(4): 225-245, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31848472

RESUMO

Cholesterol homeostasis is vital for proper cellular and systemic functions. Disturbed cholesterol balance underlies not only cardiovascular disease but also an increasing number of other diseases such as neurodegenerative diseases and cancers. The cellular cholesterol level reflects the dynamic balance between biosynthesis, uptake, export and esterification - a process in which cholesterol is converted to neutral cholesteryl esters either for storage in lipid droplets or for secretion as constituents of lipoproteins. In this Review, we discuss the latest advances regarding how each of the four parts of cholesterol metabolism is executed and regulated. The key factors governing these pathways and the major mechanisms by which they respond to varying sterol levels are described. Finally, we discuss how these pathways function in a concerted manner to maintain cholesterol homeostasis.


Assuntos
Colesterol/biossíntese , Colesterol/metabolismo , Colesterol/fisiologia , Animais , Ésteres do Colesterol/metabolismo , Homeostase/fisiologia , Humanos , Metabolismo dos Lipídeos/fisiologia , Lipoproteínas/metabolismo
15.
Artigo em Inglês | MEDLINE | ID: mdl-31678516

RESUMO

SR-B1 belongs to the class B scavenger receptor, or CD36 super family. SR-B1 and CD36 share an affinity for a wide array of ligands. Although they exhibit similar ligand binding specificity, SR-B1 and CD36 have some very specific lipid transport functions. Whereas SR-B1 primarily facilitates the selective delivery of cholesteryl esters (CEs) and cholesterol from HDL particles to the liver and non-placental steroidogenic tissues, as well as participating in cholesterol efflux from cells, CD36 primarily mediates the uptake of long-chain fatty acids in high fatty acid-requiring organs such as the heart, skeletal muscle and adipose tissue. However, CD36 also mediates cholesterol efflux and facilitates selective lipoprotein-CE delivery, although less efficiently than SR-B1. Interestingly, the ability or efficiency of SR-B1 to mediate fatty acid uptake has not been reported. In this paper, using overexpression and siRNA-mediated knockdown of SR-B1, we show that SR-B1 possesses the ability to facilitate fatty acid uptake. Moreover, this function is not blocked by BLT-1, a specific chemical inhibitor of HDL-CE uptake activity of SR-B1, nor by sulfo-N-succinimidyl oleate, which inhibits fatty acid uptake by CD36. Attenuated fatty acid uptake was also observed in primary adipocytes isolated from SR-B1 knockout mice. In conclusion, facilitation of fatty acid uptake is an additional function that is mediated by SR-B1.


Assuntos
Adipócitos/metabolismo , Ácidos Graxos/metabolismo , Receptores Depuradores Classe B/metabolismo , Adipócitos/efeitos dos fármacos , Animais , Antígenos CD36/antagonistas & inibidores , Antígenos CD36/genética , Antígenos CD36/metabolismo , Células Cultivadas , Ésteres do Colesterol/metabolismo , HDL-Colesterol/metabolismo , Ciclopentanos/farmacologia , Técnicas de Silenciamento de Genes , Metabolismo dos Lipídeos/efeitos dos fármacos , Camundongos , Camundongos Knockout , Ácidos Oleicos/farmacologia , Cultura Primária de Células , RNA Interferente Pequeno/metabolismo , Receptores Depuradores Classe B/antagonistas & inibidores , Receptores Depuradores Classe B/genética , Succinimidas/farmacologia , Tiossemicarbazonas/farmacologia
16.
FEBS Lett ; 594(1): 31-42, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31423582

RESUMO

Late endosomal/lysosomal adaptor and MAPK and mTOR activator (LAMTOR/Ragulator) is a scaffold protein complex that anchors and regulates multiprotein signaling units on late endosomes/lysosomes. To identify LAMTOR-modulated endolysosomal proteins, primary macrophages were derived from bone marrow of conditional knockout mice carrying a specific deletion of LAMTOR2 in the monocyte/macrophage cell lineage. Affymetrix-based transcriptomic analysis and quantitative iTRAQ-based organelle proteomic analysis of endosomes derived from macrophages were performed. Further analyses showed that LAMTOR could be a novel regulator of foam cell differentiation. The lipid droplet formation phenotype observed in macrophages was additionally confirmed in MEFs, where lipidomic analysis identified cholesterol esters as specifically downregulated in LAMTOR2 knockout cells. The data obtained indicate a function of LAMTOR2 in lipid metabolism.


Assuntos
Diferenciação Celular , Células Espumosas/metabolismo , Metabolismo dos Lipídeos , Macrófagos/metabolismo , Proteínas/metabolismo , Animais , Células Cultivadas , Ésteres do Colesterol/metabolismo , Células Espumosas/citologia , Gotículas Lipídicas/metabolismo , Macrófagos/citologia , Camundongos , Proteínas/genética , Transcriptoma
17.
Ann Anat ; 227: 151420, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31563571

RESUMO

BACKGROUND: Cholesterol esters (CEs) accumulate in the cells of the adrenal cortex and are used for the synthesis of steroid hormones. The full molecular pathways involved in mediating the accumulation of CEs within the adrenal cortex are yet to be elucidated. Tissue non-specific alkaline phosphatase (TNAP) is needed for intracellular lipid accumulation of triglycerides in adipocytes and is also expressed in the cortical cells of the adrenal gland. Therefore we aimed to determine if TNAP is needed for the accumulation of CEs within the murine Y1 adrenal cortex cell line. METHODS: Y1 cells were induced to accumulate lipids. Lipid accumulation and TNAP activity and expression were determined throughout intracellular lipid accumulation. The location of TNAP within the cell was determined through immunohistochemical analysis. Lipid accumulation in the cells was associated with a rise in TNAP activity and TNAP was localised to lipid droplets within the Y1 cells. Inhibition of TNAP with a specific inhibitor (levamisole) resulted in the cessation of CE accumulation. DISCUSSION AND CONCLUSIONS: These data demonstrate that TNAP plays a role in the control of lipid accumulation in this adrenal cortex cell line. Therefore, in both triglyceride and CE storing cell types TNAP would seem to be essential for intra-cellular lipid storage.


Assuntos
Córtex Suprarrenal/metabolismo , Fosfatase Alcalina/metabolismo , Ésteres do Colesterol/metabolismo , Córtex Suprarrenal/citologia , Córtex Suprarrenal/efeitos dos fármacos , Fosfatase Alcalina/análise , Fosfatase Alcalina/antagonistas & inibidores , Fosfatase Alcalina/genética , Animais , Compostos Azo , Linhagem Celular , Corantes , Corantes Fluorescentes , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Levamisol/farmacologia , Lipídeos/química , Camundongos , Oxazinas , Regulação para Cima
18.
J Lipid Res ; 60(11): 1868-1879, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31511396

RESUMO

Cholesteryl ester transfer protein (CETP) regulates intravascular lipoprotein metabolism. In vitro studies indicate that ApoF alters CETP function by inhibiting its activity with LDL. To explore in vivo the complexities driving ApoF's effects on CETP, we developed a siRNA-based hamster model of ApoF knockdown. In both male and female hamsters on chow- or fat-fed diets, we measured lipoprotein levels and composition, determined CETP-mediated transfer of cholesteryl esters (CEs) between lipoproteins, and quantified reverse cholesterol transport (RCT). We found that apoF knockdown in chow-fed hamsters had no effect on lipoprotein levels or composition, but these ApoF-deficient lipoproteins supported 50-100% higher LDL CETP activity in vitro. ApoF knockdown in fat-fed male hamsters created a phenotype in which endogenous CETP-mediated CE transfer from HDL to LDL increased up to 2-fold, LDL cholesterol increased 40%, HDL declined 25%, LDL and HDL lipid compositions were altered, and hepatic LDLR gene expression was decreased. Diet-induced hypercholesterolemia obscured this phenotype on occasion. In fat-fed female hamsters, ApoF knockdown caused similar but smaller changes in plasma CETP activity and LDL cholesterol. Notably, ApoF knockdown impaired HDL RCT in fat-fed hamsters but increased sterol excretion in chow-fed animals. These in vivo data validate in vitro findings that ApoF regulates lipid transfer to LDL. The consequences of ApoF knockdown on lipoproteins and sterol excretion depend on the underlying lipid status. By minimizing the transfer of HDL-derived CE to LDL, ApoF helps control LDL cholesterol levels when LDL clearance mechanisms are limiting.


Assuntos
Apoproteínas/deficiência , Apoproteínas/genética , Ésteres do Colesterol/metabolismo , LDL-Colesterol/metabolismo , Dieta , Técnicas de Silenciamento de Genes , Animais , Proteínas de Transferência de Ésteres de Colesterol/metabolismo , Cricetinae , Feminino , Regulação da Expressão Gênica/genética , Células HEK293 , Humanos , Fígado/metabolismo , Masculino , RNA Interferente Pequeno/genética
19.
Sci Rep ; 9(1): 11642, 2019 08 12.
Artigo em Inglês | MEDLINE | ID: mdl-31406145

RESUMO

Amyotrophic lateral sclerosis (ALS) is characterized by progressive loss of upper and lower motor neurons leading to muscle paralysis and death. While a link between dysregulated lipid metabolism and ALS has been proposed, lipidome alterations involved in disease progression are still understudied. Using a rodent model of ALS overexpressing mutant human Cu/Zn-superoxide dismutase gene (SOD1-G93A), we performed a comparative lipidomic analysis in motor cortex and spinal cord tissues of SOD1-G93A and WT rats at asymptomatic (~70 days) and symptomatic stages (~120 days). Interestingly, lipidome alterations in motor cortex were mostly related to age than ALS. In contrast, drastic changes were observed in spinal cord of SOD1-G93A 120d group, including decreased levels of cardiolipin and a 6-fold increase in several cholesteryl esters linked to polyunsaturated fatty acids. Consistent with previous studies, our findings suggest abnormal mitochondria in motor neurons and lipid droplets accumulation in aberrant astrocytes. Although the mechanism leading to cholesteryl esters accumulation remains to be established, we postulate a hypothetical model based on neuroprotection of polyunsaturated fatty acids into lipid droplets in response to increased oxidative stress. Implicated in the pathology of other neurodegenerative diseases, cholesteryl esters appear as attractive targets for further investigations.


Assuntos
Esclerose Amiotrófica Lateral/patologia , Metabolismo dos Lipídeos/genética , Neurônios Motores/metabolismo , Medula Espinal/patologia , Superóxido Dismutase-1/genética , Envelhecimento/metabolismo , Esclerose Amiotrófica Lateral/genética , Esclerose Amiotrófica Lateral/metabolismo , Animais , Cardiolipinas/análise , Cardiolipinas/metabolismo , Ésteres do Colesterol/análise , Ésteres do Colesterol/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Ácidos Graxos Insaturados/análise , Ácidos Graxos Insaturados/metabolismo , Feminino , Humanos , Gotículas Lipídicas/patologia , Lipidômica , Masculino , Espectrometria de Massas , Córtex Motor/metabolismo , Neurônios Motores/química , Mutação , Estresse Oxidativo/genética , Ratos , Ratos Transgênicos , Medula Espinal/química , Medula Espinal/citologia , Medula Espinal/metabolismo , Superóxido Dismutase-1/metabolismo
20.
World J Gastroenterol ; 25(30): 4172-4180, 2019 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-31435171

RESUMO

Lysosomal acid lipase (LAL) plays a key role in intracellular lipid metabolism. Reduced LAL activity promotes increased multi-organ lysosomal cholesterol ester storage, as observed in two recessive autosomal genetic diseases, Wolman disease and Cholesterol ester storage disease. Severe liver steatosis and accelerated liver fibrosis are common features in patients with genetic LAL deficiency. By contrast, few reliable data are available on the modulation of LAL activity in vivo and on the epigenetic and metabolic factors capable of regulating its activity in subjects without homozygous mutations of the Lipase A gene. In the last few years, a less severe and non-genetic reduction of LAL activity was reported in children and adults with non-alcoholic fatty liver disease (NAFLD), suggesting a possible role of LAL reduction in the pathogenesis and progression of the disease. Patients with NAFLD show a significant, progressive reduction of LAL activity from simple steatosis to non-alcoholic steatohepatitis and cryptogenic cirrhosis. Among cirrhosis of different etiologies, those with cryptogenic cirrhosis show the most significant reductions of LAL activity. These findings suggest that the modulation of LAL activity may become a possible new therapeutic target for patients with more advanced forms of NAFLD. Moreover, the measurement of LAL activity may represent a possible new marker of disease severity in this clinical setting.


Assuntos
Cirrose Hepática/patologia , Hepatopatia Gordurosa não Alcoólica/diagnóstico , Esterol Esterase/deficiência , Doença de Wolman/patologia , Biomarcadores/sangue , Biomarcadores/metabolismo , Ésteres do Colesterol/metabolismo , Progressão da Doença , Teste em Amostras de Sangue Seco , Terapia de Reposição de Enzimas/métodos , Humanos , Metabolismo dos Lipídeos/genética , Fígado/patologia , Cirrose Hepática/prevenção & controle , Testes de Função Hepática/métodos , Lisossomos/metabolismo , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/patologia , Índice de Gravidade de Doença , Esterol Esterase/sangue , Esterol Esterase/genética , Esterol Esterase/uso terapêutico , Triglicerídeos/metabolismo , Doença de Wolman/tratamento farmacológico , Doença de Wolman/genética
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