Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 22
Filtrar
1.
Int J Mol Sci ; 24(4)2023 Feb 11.
Artigo em Inglês | MEDLINE | ID: mdl-36835060

RESUMO

Controlling post-prandial hyperglycemia and hyperlipidemia, particularly by regulating the activity of digestive enzymes, allows managing type 2 diabetes and obesity. The aim of this study was to assess the effects of TOTUM-63, a formulation of five plant extracts (Olea europaea L., Cynara scolymus L., Chrysanthellum indicum subsp. afroamericanum B.L.Turner, Vaccinium myrtillus L., and Piper nigrum L.), on enzymes involved in carbohydrate and lipid absorption. First, in vitro inhibition assays were performed by targeting three enzymes: α-glucosidase, α-amylase, and lipase. Then, kinetic studies and binding affinity determinations by fluorescence spectrum changes and microscale thermophoresis were performed. The in vitro assays showed that TOTUM-63 inhibited all three digestive enzymes, particularly α-glucosidase (IC50 of 13.1 µg/mL). Mechanistic studies on α-glucosidase inhibition by TOTUM-63 and molecular interaction experiments indicated a mixed (full) inhibition mechanism, and higher affinity for α-glucosidase than acarbose, the reference α-glucosidase inhibitor. Lastly, in vivo data using leptin receptor-deficient (db/db) mice, a model of obesity and type 2 diabetes, indicated that TOTUM-63 might prevent the increase in fasting glycemia and glycated hemoglobin (HbA1c) levels over time, compared with the untreated group. These results show that TOTUM-63 is a promising new approach for type 2 diabetes management via α-glucosidase inhibition.


Assuntos
Diabetes Mellitus Tipo 2 , Inibidores de Glicosídeo Hidrolases , Extratos Vegetais , alfa-Glucosidases , Animais , Camundongos , alfa-Amilases/metabolismo , alfa-Glucosidases/metabolismo , Diabetes Mellitus Tipo 2/tratamento farmacológico , Inibidores de Glicosídeo Hidrolases/farmacologia , Hipoglicemiantes/farmacologia , Cinética , Lipase/metabolismo , Obesidade , Extratos Vegetais/farmacologia
2.
J Lipid Res ; 58(1): 81-91, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27884961

RESUMO

Obesity is a component of the metabolic syndrome, mechanistically linked to diabetes, fatty liver disease, and cardiovascular disease. Proteins that regulate the metabolic fate of intracellular lipid droplets are potential therapeutic candidates to treat obesity and its related consequences. CIDEC (cell death-inducing DFFA-like effector C), also known in mice as Fsp27 (fat-specific protein 27), is a lipid droplet-associated protein that prevents lipid mobilization and promotes intracellular lipid storage. The consequences of complete loss of FSP27 on hepatic metabolism and on insulin resistance are controversial, as both healthy and deleterious lipodystrophic phenotypes have been reported in Fsp27-/- mice. To test whether therapeutic silencing of Fsp27 might be useful to improve obesity, fatty liver, and glycemic control, we used antisense oligonucleotides (ASOs) in both nutritional (high-fat diet) and genetic (leptin-deficient ob/ob) mouse models of obesity, hyperglycemia, and hepatosteatosis. We show that partial silencing Fsp27 in either model results in the robust decrease in visceral fat, improved insulin sensitivity and whole-body glycemic control, and tissue-specific changes in transcripts controlling lipid oxidation and synthesis. These data suggest that partial reduction of FSP27 activity (e.g., using ASOs) might be exploited therapeutically in insulin-resistant obese or overweight patients.


Assuntos
Diabetes Mellitus/terapia , Fígado Gorduroso/terapia , Obesidade/terapia , Oligonucleotídeos Antissenso/administração & dosagem , Proteínas/genética , Animais , Diabetes Mellitus/genética , Diabetes Mellitus/metabolismo , Dieta Hiperlipídica , Modelos Animais de Doenças , Fígado Gorduroso/genética , Fígado Gorduroso/metabolismo , Humanos , Resistência à Insulina/genética , Gotículas Lipídicas/metabolismo , Gotículas Lipídicas/patologia , Metabolismo dos Lipídeos/genética , Fígado/metabolismo , Fígado/patologia , Camundongos , Camundongos Obesos , Obesidade/genética , Oligonucleotídeos Antissenso/genética , Proteínas/antagonistas & inibidores
3.
J Lipid Res ; 58(2): 317-324, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-28007964

RESUMO

α-Chlorofatty aldehydes are generated from myeloperoxidase-derived HOCl targeting plasmalogens, and are subsequently oxidized to α-chlorofatty acids (α-ClFAs). The catabolic pathway for α-ClFA is initiated by ω-oxidation. Here, we examine PPAR-α activation as a mechanism to increase α-ClFA catabolism. Pretreating both HepG2 cells and primary mouse hepatocytes with the PPAR-α agonist, pirinixic acid (Wy 14643), increased the production of α-chlorodicarboxylic acids (α-ClDCAs) in cells treated with exogenous α-ClFA. Additionally, α-ClDCA production in Wy 14643-pretreated wild-type mouse hepatocytes was accompanied by a reduction in cellular free α-ClFA. The dependence of PPAR-α-accelerated α-ClFA catabolism was further demonstrated by both impaired metabolism in mouse PPAR-α-/- hepatocytes and decreased clearance of plasma α-ClFA in PPAR-α-/- mice. Furthermore, Wy 14643 treatments decreased plasma 2-chlorohexadecanoic acid levels in wild-type mice. Additional studies showed that α-ClFA increases PPAR-α, PPAR-δ, and PPAR-γ activities, as well as mRNA expression of the PPAR-α target genes, CD36, CPT1a, Cyp4a10, and CIDEC. Collectively, these results indicate that PPAR-α accelerates important pathways for the clearance of α-ClFA, and α-ClFA may, in part, accelerate its catabolism by serving as a ligand for PPAR-α.


Assuntos
Hepatócitos/metabolismo , PPAR alfa/genética , Ácidos Palmíticos/metabolismo , Animais , Ácidos Graxos/metabolismo , Hepatócitos/efeitos dos fármacos , Humanos , Metabolismo/genética , Camundongos , Camundongos Knockout , Oxirredução , PPAR alfa/metabolismo , PPAR delta/biossíntese , RNA Mensageiro/biossíntese
4.
Hepatology ; 63(5): 1660-74, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26473496

RESUMO

UNLABELLED: Understanding the hepatic regenerative process has clinical interest as the effectiveness of many treatments for chronic liver diseases is conditioned by efficient liver regeneration. Experimental evidence points to the need for a temporal coordination between cytokines, growth factors, and metabolic signaling pathways to enable successful liver regeneration. One intracellular mediator that acts as a signal integration node for these processes is the serine-threonine kinase Akt/protein kinase B (Akt). To investigate the contribution of Akt during hepatic regeneration, we performed partial hepatectomy in mice lacking Akt1, Akt2, or both isoforms. We found that absence of Akt1 or Akt2 does not influence liver regeneration after partial hepatectomy. However, hepatic-specific Akt1 and Akt2 null mice show impaired liver regeneration and increased mortality. The major abnormal cellular events observed in total Akt-deficient livers were a marked reduction in cell proliferation, cell hypertrophy, glycogenesis, and lipid droplet formation. Most importantly, liver-specific deletion of FoxO1, a transcription factor regulated by Akt, rescued the hepatic regenerative capability in Akt1-deficient and Akt2-deficient mice and normalized the cellular events associated with liver regeneration. CONCLUSION: The Akt-FoxO1 signaling pathway plays an essential role during liver regeneration.


Assuntos
Fatores de Transcrição Forkhead/fisiologia , Regeneração Hepática , Proteínas Proto-Oncogênicas c-akt/fisiologia , Proteínas Quinases Dependentes de 3-Fosfoinositídeo/fisiologia , Animais , Proteína Forkhead Box O1 , Fatores de Transcrição Forkhead/antagonistas & inibidores , Hepatócitos/patologia , Hiperplasia , Metabolismo dos Lipídeos , Masculino , Camundongos , Transdução de Sinais/fisiologia
5.
Hepatology ; 61(4): 1227-38, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25418138

RESUMO

UNLABELLED: The cell death-inducing DNA fragmentation factor alpha-like effector c (CIDEC; also known in rodents as FSP27 or fat-specific protein 27) is a lipid droplet-associated protein that promotes intracellular triglyceride (TAG) storage. CIDEC/Fsp27 is highly expressed in adipose tissue, but undetectable in normal liver. However, its hepatic expression rises during fasting or under genetic or diet-induced hepatosteatosis in both mice and patients. Herein, we demonstrate that CIDEC/Fsp27 is a direct transcriptional target of the nuclear receptor PPARα (peroxisome proliferator-activated receptor alpha) in both mouse and human hepatocytes, and that preventing Fsp27 induction accelerates PPARα-stimulated fatty acid oxidation. We show that adenoviral-mediated silencing of hepatic Fsp27 abolishes fasting-induced liver steatosis in the absence of changes in plasma lipids. Finally, we report that anti-Fsp27 short hairpin RNA and PPARα agonists synergize to ameliorate hepatosteatosis in mice fed a high fat diet. CONCLUSIONS: Together, our data highlight the physiological importance of CIDEC/Fsp27 in TAG homeostasis under both physiological and pathological liver steatosis. Our results also suggest that patients taking fibrates likely have elevated levels of hepatic CIDEC, which may limit the efficient mobilization and catabolism of hepatic TAGs.


Assuntos
Fígado Gorduroso/etiologia , PPAR alfa/fisiologia , Proteínas/fisiologia , Animais , Células Cultivadas , Dieta , Jejum , Hepatócitos , Humanos , Fígado , Camundongos
6.
J Hepatol ; 61(2): 358-65, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24768901

RESUMO

BACKGROUND & AIMS: Perilipin-5 (PLIN5) is a member of the perilipin family of lipid droplet (LD)-associated proteins. PLIN5 is expressed in oxidative tissues including the liver, and is critical during LD biogenesis. Studies showed that statins reduce hepatic triglyceride contents in some patients with non-alcoholic fatty liver disease and in rodent models of diet-induced hepatosteatosis. Whether statins alter triglyceride synthesis, storage, and/or utilization within the hepatocyte is unknown, though. Here we tested the hypothesis that statins alter the metabolism of LD in the hepatocyte during physiological conditions, such as fasting-induced steatosis. METHODS: Mice were gavaged with saline or atorvastatin, and the expression of LD-associated genes was determined in fed and fasted animals. The accumulation of triglycerides and LD was studied in mouse or human primary hepatocytes in response to statins, and following knock-down of SREBP2 or PLIN5. RESULTS: We show that statins decrease the levels of PLIN5, but not other LD-associated genes, in both mouse liver and mouse/human primary hepatocytes, which is paralleled by a significant reduction in both intracellular triglycerides and the number of LD. We identify an atypical negative sterol regulatory sequence in the proximal promoter of mouse/human PLIN5 that recruits the transcription factor SREBP2 and confers response to statins. Finally, we show that the statin-dependent reduction of hepatocyte triglyceride contents is mimicked by partial knock-down of PLIN5; conversely, ectopic overexpression of PLIN5 reverts the statin effect. CONCLUSIONS: PLIN5 is a physiological regulator of triglyceride metabolism in the liver, and likely contributes to the pleiotropic effects of statins.


Assuntos
Hepatócitos/metabolismo , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Proteínas Musculares/fisiologia , Triglicerídeos/metabolismo , Animais , Hepatócitos/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intracelular/análise , Gotículas Lipídicas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Musculares/análise , Proteína de Ligação a Elemento Regulador de Esterol 2/fisiologia
7.
Front Cardiovasc Med ; 11: 1342388, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38317864

RESUMO

Introduction: Totum-070 is a combination of five plant extracts enriched in polyphenols to target hypercholesterolemia, one of the main risk factors for cardiovascular diseases. The aim of this study was to investigate the effects of Totum-070 on cholesterol levels in an animal model of diet-induced hypercholesterolemia. Methods: C57BL/6JOlaHsd male mice were fed a Western diet and received Totum-070, or not, by daily gavage (1g/kg and 3g/kg body weight) for 6 weeks. Results: The Western diet induced obesity, fat accumulation, hepatic steatosis and increased plasma cholesterol compared with the control group. All these metabolic perturbations were alleviated by Totum-070 supplementation in a dose-dependent manner. Lipid excretion in feces was higher in mice supplemented with Totum-070, suggesting inhibition of intestinal lipid absorption. Totum-070 also increased the fecal concentration of short chain fatty acids, demonstrating a direct effect on intestinal microbiota. Discussion: The characterization of fecal microbiota by 16S amplicon sequencing showed that Totum-070 supplementation modulated the dysbiosis associated with metabolic disorders. Specifically, Totum-070 increased the relative abundance of Muribaculum (a beneficial bacterium) and reduced that of Lactococcus (a genus positively correlated with increased plasma cholesterol level). Together, these findings indicate that the cholesterol-lowering effect of Totum-070 bioactive molecules could be mediated through multiple actions on the intestine and gut microbiota.

8.
J Lipid Res ; 54(9): 2475-84, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23772048

RESUMO

Class I alcohol dehydrogenases (ADH1s) are the rate-limiting enzymes for ethanol and vitamin A (retinol) metabolism in the liver. Because previous studies have shown that human ADH1 enzymes may participate in bile acid metabolism, we investigated whether the bile acid-activated nuclear receptor farnesoid X receptor (FXR) regulates ADH1 genes. In human hepatocytes, both the endogenous FXR ligand chenodeoxycholic acid and synthetic FXR-specific agonist GW4064 increased ADH1 mRNA, protein, and activity. Moreover, overexpression of a constitutively active form of FXR induced ADH1A and ADH1B expression, whereas silencing of FXR abolished the effects of FXR agonists on ADH1 expression and activity. Transient transfection studies and electrophoretic mobility shift assays revealed functional FXR response elements in the ADH1A and ADH1B proximal promoters, thus indicating that both genes are direct targets of FXR. These findings provide the first evidence for direct connection of bile acid signaling and alcohol metabolism.


Assuntos
Álcool Desidrogenase/metabolismo , Ácido Quenodesoxicólico/farmacologia , Álcool Desidrogenase/genética , Álcoois/metabolismo , Animais , Sequência de Bases , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Células Hep G2 , Hepatócitos/citologia , Hepatócitos/efeitos dos fármacos , Humanos , Isoxazóis/farmacologia , Ligantes , Masculino , Camundongos , Regiões Promotoras Genéticas/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Citoplasmáticos e Nucleares/agonistas , Receptores Citoplasmáticos e Nucleares/metabolismo , Sequências Repetitivas de Ácido Nucleico/genética , Elementos de Resposta/genética , Transdução de Sinais/efeitos dos fármacos
9.
Biochem Biophys Res Commun ; 434(1): 102-9, 2013 Apr 26.
Artigo em Inglês | MEDLINE | ID: mdl-23541942

RESUMO

Here we report that bile acid chenodeoxycholic acid (CDCA) and synthetic farnesoid X receptor (FXR) agonist GW4064 robustly induced tumor suppressor N-Myc downstream regulated gene 2 (NDRG2) expression in human hepatoma cells and primary hepatocytes. Knockdown of FXR abolished the induction by CDCA, whereas overexpression of a constitutively active form of FXR increased NDRG2 expression. A FXR-response element was identified within intronic regions of human and murine genes. Moreover, mice given GW4064 exhibit an increase of Ndrg2 expression in liver and kidney, where both NDRG2 and FXR are enriched. The identification of NDRG2 as a bile acid regulated gene may provide novel knowledge toward the understanding of NDRG2 physiological function and the link between metabolism and cancer.


Assuntos
Ácidos e Sais Biliares/farmacologia , Proteínas/genética , Proteínas Supressoras de Tumor/genética , Proteínas Adaptadoras de Transdução de Sinal , Animais , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Células Hep G2 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas/metabolismo , Receptores Citoplasmáticos e Nucleares/agonistas , Receptores Citoplasmáticos e Nucleares/antagonistas & inibidores , Receptores Citoplasmáticos e Nucleares/genética , Proteínas Supressoras de Tumor/metabolismo , Regulação para Cima/fisiologia
10.
Nutr Res ; 118: 70-84, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-37598559

RESUMO

Global prevalence of obesity and type 2 diabetes are rapidly increasing to pandemic proportions. A novel supplement composed of 5 plant extracts from olive leaf, bilberry, artichoke, chrysanthellum, and black pepper was designed to prevent type 2 diabetes development in people at risk. It was previously shown to improve body weight and glucose control in preclinical rodent models, with these effects being accompanied by increased fecal energy excretion and in vitro inhibition of several digestive enzymes. Thus, we hypothesized that, in mice fed a high-fat diet (HFD), a single dose of this botanical supplementation would decrease the responses to oral fat and carbohydrate tolerance tests, and that chronic supplementation would result in increased fecal triglyceride content. We showed that acute administration in HFD-fed mice (1.452 g/kg body weight) markedly reduced circulating triglycerides following an oral lipid gavage, whereas glycemic responses to various carbohydrate tests were only mildly affected. When incorporated into the food (2.5%) of HFD-fed mice, chronic supplementation prevented body weight gain and improved glucose homeostasis and lipid tolerance. Fecal free fatty acid content, but not triglyceride, was significantly increased in supplemented animals, suggesting reduced lipid absorption in the digestive tract. Congruently, this botanical supplementation downregulated several genes associated with fatty acid transport whose expression was increased by HFD, principally in the jejunum. This study provides novel insights as for the mode of action behind the antiobesity effect of this plant-based supplementation, in HFD-fed mice.


Assuntos
Diabetes Mellitus Tipo 2 , Extratos Vegetais , Humanos , Animais , Camundongos , Dieta Hiperlipídica/efeitos adversos , Polifenóis/farmacologia , Diabetes Mellitus Tipo 2/metabolismo , Fígado/metabolismo , Aumento de Peso , Peso Corporal , Triglicerídeos/metabolismo , Nutrientes , Carboidratos , Camundongos Endogâmicos C57BL
11.
Nutrients ; 15(8)2023 Apr 14.
Artigo em Inglês | MEDLINE | ID: mdl-37111121

RESUMO

TOTUM-070 is a patented polyphenol-rich blend of five different plant extracts showing separately a latent effect on lipid metabolism and potential synergistic properties. In this study, we investigated the health benefit of such a formula. Using a preclinical model of high fat diet, TOTUM-070 (3 g/kg of body weight) limited the HFD-induced hyperlipemia with a reduction in triglyceride (-32% after 6 weeks; -20.3% after 12 weeks) and non-HDL cholesterol levels (-21% after 6 weeks; -38.4% after 12 weeks). To further investigate such a benefit and its underlying mechanisms in humans, we designed an ex vivo clinical approach to collect the circulating bioactives resulting from TOTUM-070 ingestion and to determine their biological activities on human hepatocytes. Human serum was obtained from healthy subjects before and after intake of TOTUM-070 (4995 mg). The presence of circulating metabolites was assessed by UPLC-MS/MS. Serum containing metabolites was further incubated with hepatocytes cultured in a lipotoxic environment (palmitate, 250 µM). RNA sequencing analyses show that lipid metabolism was one of the most impacted processes. Using histologic, proteomic, and enzymatic assays, the effects of human TOTUM-070 bioactives on hepatocyte metabolism were characterized by (1) the inhibition of lipid storage, including both (2) triglycerides (-41%, p < 0.001) and (3) cholesterol (-50%, p < 0.001) intracellular content, (4) a reduced de novo cholesterol synthesis (HMG-CoA reductase activity -44%, p < 0.001), and (5) a lowered fatty acid synthase protein level (p < 0.001). Altogether, these data support the beneficial impact of TOTUM-070 on lipid metabolism and provide new biochemical insights in human mechanisms occurring in liver cells.


Assuntos
Metabolismo dos Lipídeos , Polifenóis , Humanos , Polifenóis/farmacologia , Polifenóis/metabolismo , Cromatografia Líquida , Proteômica , Espectrometria de Massas em Tandem , Hepatócitos , Colesterol , Triglicerídeos , Dieta Hiperlipídica , Fígado/metabolismo
12.
Nutrients ; 15(24)2023 Dec 09.
Artigo em Inglês | MEDLINE | ID: mdl-38140315

RESUMO

Atherosclerotic cardiovascular disease is the leading cause of mortality worldwide, and hypercholesterolemia is a central risk factor for atherosclerosis. This study evaluated the effects of Totum-070, a plant-based polyphenol-rich supplement, in hamsters with high-fat diet (HFD)-induced dyslipidemia. The molecular mechanisms of action were explored using human Caco2 enterocytes. Totum-070 supplementation reduced the total cholesterol (-41%), non-HDL cholesterol (-47%), and triglycerides (-46%) in a dose-dependent manner, compared with HFD. HFD-induced hepatic steatosis was also significantly decreased by Totum-070, an effect associated with the reduction in various lipid and inflammatory gene expression. Upon challenging with olive oil gavage, the post-prandial triglyceride levels were strongly reduced. The sterol excretion in the feces was increased in the HFD-Totum-070 groups compared with the HFD group and associated with reduction of intestinal cholesterol absorption. These effects were confirmed in the Caco2 cells, where incubation with Totum-070 inhibited cholesterol uptake and apolipoprotein B secretion. Furthermore, a microbiota composition analysis revealed a strong effect of Totum-070 on the alpha and beta diversity of bacterial species and a significant decrease in the Firmicutes to Bacteroidetes ratio. Altogether, our findings indicate that Totum-070 lowers hypercholesterolemia by reducing intestinal cholesterol absorption, suggesting that its use as dietary supplement may be explored as a new preventive strategy for cardiovascular diseases.


Assuntos
Aterosclerose , Hipercolesterolemia , Hiperlipidemias , Cricetinae , Animais , Humanos , Hipercolesterolemia/etiologia , Extratos Vegetais/farmacologia , Extratos Vegetais/metabolismo , Dieta Hiperlipídica/efeitos adversos , Polifenóis/farmacologia , Polifenóis/metabolismo , Células CACO-2 , Mesocricetus , Colesterol/metabolismo , Hiperlipidemias/metabolismo , Triglicerídeos/metabolismo , Aterosclerose/etiologia , Aterosclerose/prevenção & controle , Aterosclerose/metabolismo , Fígado/metabolismo
13.
Arterioscler Thromb Vasc Biol ; 29(12): 2191-7, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19762784

RESUMO

OBJECTIVE: Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a central player in the regulation of cholesterol homeostasis, increasing the low-density lipoprotein (LDL) receptor degradation. Our study aimed at exploring the pathogenic consequences in vivo and in vitro of a PCSK9 prodomain mutation found in a family with hypobetalipoproteinemia (FHBL). METHODS AND RESULTS: A white 49-year-old diabetic man had profound FBHL (LDLC: 16 mg/dL) whereas his daughter and sister displayed a milder phenotype (LDLC 44 mg/dL and 57 mg/dL, respectively), all otherwise healthy with a normal liver function. A monoallelic PCSK9 double-mutant R104C/V114A cosegregated with FBHL, with no mutation found at other FHBL-causing loci. A dose-effect was also found in FBHL relatives for plasma APOB and PCSK9 (very-low to undetectable in proband, approximately 50% decreased in sister and daughter) and LDL catabolic rate (256% and 88% increased in proband and daughter). Transient transfection in hepatocytes showed severely impaired processing and secretion of the double mutant which acted as a dominant negative over secretion of wild-type PCSK9. CONCLUSIONS: These results show that heterozygous PCSK9 missense mutations may associate with profound hypobetalipoproteinemia and constitute the first direct evidence in human that decrease of plasma LDLC concentrations associated to PCSK9 LOF mutations are attributable to an increased clearance rate of LDL.


Assuntos
LDL-Colesterol/sangue , Hipobetalipoproteinemias/enzimologia , Hipobetalipoproteinemias/genética , Mutação de Sentido Incorreto , Serina Endopeptidases/sangue , Serina Endopeptidases/genética , Adulto , Substituição de Aminoácidos , Apolipoproteínas B/sangue , Feminino , Genes Dominantes , Hepatócitos/enzimologia , Heterozigoto , Humanos , Hipobetalipoproteinemia Familiar por Apolipoproteína B/sangue , Hipobetalipoproteinemia Familiar por Apolipoproteína B/enzimologia , Hipobetalipoproteinemia Familiar por Apolipoproteína B/genética , Hipobetalipoproteinemias/sangue , Cinética , Masculino , Pessoa de Meia-Idade , Linhagem , Pró-Proteína Convertase 9 , Pró-Proteína Convertases , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Serina Endopeptidases/deficiência , Transfecção
14.
Arterioscler Thromb Vasc Biol ; 29(5): 684-90, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-19265033

RESUMO

OBJECTIVES: Proprotein convertase subtilisin kexin type 9 (PCSK9) is a natural inhibitor of the low-density lipoprotein receptor, and its deficiency in humans results in low plasma LDL-cholesterol and protection against cardiovascular disease. We explored whether PCSK9 expression impacts postprandial triglyceridemia, another important cardiovascular risk factor. METHODS AND RESULTS: Real-time PCR and confocal microscopy were used to show that PCSK9 is expressed throughout the entire small intestine and in human enterocytes. On olive oil gavage, PCSK9-deficient mice showed a dramatically decreased postprandial triglyceridemia compared with their wild-type littermates. Lymph analysis revealed that intestinal TG output is not quantitatively modified by PCSK9 deletion. However, PCSK9-/- mice present with a significant reduction of lymphatic apoB secretion compared to PCSK9+/+ mice. Modulating PCSK9 expression in polarized CaCo-2 cells confirmed the relationship between PCSK9 and apoB secretion; PCSK9-/- mice consistently secrete larger TG-rich lipoprotein than wild-type littermates. Finally, kinetic studies showed that PCSK9-deficient mice have an increased ability to clear chylomicrons compared to wild-type littermates. CONCLUSION: These findings indicate that in addition to its effect on LDL-cholesterol, PCSK9 deficiency might protect against cardiovascular disease by reducing postprandial triglyceridemia.


Assuntos
Apolipoproteínas B/metabolismo , Enterócitos/metabolismo , Intestino Delgado/metabolismo , Serina Endopeptidases/metabolismo , Triglicerídeos/metabolismo , Animais , Células CACO-2 , Quilomícrons/metabolismo , Duodeno/metabolismo , Células Caliciformes/metabolismo , Humanos , Íleo/metabolismo , Camundongos , Camundongos Knockout , Microscopia Confocal , Pró-Proteína Convertase 9 , Pró-Proteína Convertases , Triglicerídeos/sangue
15.
Med Sci (Paris) ; 26(4): 385-90, 2010 Apr.
Artigo em Francês | MEDLINE | ID: mdl-20412743

RESUMO

Type 2 diabetes (T2D) frequently occurs in the context of a dysregulation of plasma lipoproteins with an increased triglyceride content in pancreatic beta cells, leading to lipotoxicity and subsequent cell death. More recently, accumulating data suggest that cholesterol homeostasis is a major regulator of beta cell function. Intra-cellular cholesterol accumulation leads to islet dysfunction and impaired insulin secretion. The role of essential cholesterol modulators like the ATP-binding cassette transporter A1 or the LDL receptor has emerged in regulating insulin secretion in beta cells. Intracellular cholesterol impacts both the beta-cell membrane organization in microdomains as well as the dynamic regulation of glucose-induced insulin secretion. There is also evidence suggesting that the different lipoprotein classes have varying effects on beta cell apoptosis and proliferation. Here we review the impact of cholesterol metabolism on islet function and its potential relationship to T2D.


Assuntos
Colesterol/metabolismo , Ilhotas Pancreáticas/metabolismo , Transportador 1 de Cassete de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/fisiologia , Animais , Apoptose , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/etiologia , Diabetes Mellitus Tipo 2/metabolismo , Humanos , Hiperlipoproteinemia Tipo II/complicações , Hiperlipoproteinemia Tipo II/fisiopatologia , Insulina/metabolismo , Resistência à Insulina/fisiologia , Secreção de Insulina , Ilhotas Pancreáticas/patologia , Lipídeos de Membrana/metabolismo , Síndrome Metabólica/metabolismo , Camundongos , Modelos Biológicos , Ensaios Clínicos Controlados Aleatórios como Assunto , Ratos , Receptores de LDL/fisiologia , Proteína de Ligação a Elemento Regulador de Esterol 2/fisiologia
16.
Biochem Biophys Res Commun ; 390(4): 1288-93, 2009 Dec 25.
Artigo em Inglês | MEDLINE | ID: mdl-19878649

RESUMO

PCSK9 (Proprotein Convertase Subtilisin Kexin type 9) is a proprotein convertase that plays a key role in cholesterol homeostasis by decreasing hepatic low-density lipoprotein receptor (LDLR) protein expression. Here, we investigated the expression and the function of PCSK9 in pancreatic islets. Immunohistochemistry analysis showed that PCSK9 co-localized specifically with somatostatin in human pancreatic delta-cells, with no expression in alpha- and beta-cells. PCSK9 seems not to be secreted by mouse isolated islets maintained in culture. Pcsk9-deficiency led to a 200% increase in LDLR protein content in mouse isolated islets, mainly in beta-cells. Conversely, incubation of islets with recombinant PCSK9 almost abolished LDLR expression. However, Pcsk9-deficiency did not alter cholesterol content nor glucose-stimulated insulin secretion in mouse islets. Finally, invivo glucose tolerance was similar in Pcsk9(+/+) and Pcsk9(-/-) mice under basal conditions and following streptozotocin treatment. These results suggest, at least in mice, that PCSK9 does not alter insulin secretion.


Assuntos
Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Serina Endopeptidases/biossíntese , Células Secretoras de Somatostatina/enzimologia , Animais , Linhagem Celular , Colesterol/metabolismo , Glucose/metabolismo , Glucose/farmacologia , Humanos , Secreção de Insulina , Células Secretoras de Insulina/efeitos dos fármacos , Camundongos , Pró-Proteína Convertase 9 , Pró-Proteína Convertases , Receptores de LDL/metabolismo , Serina Endopeptidases/genética , Células Secretoras de Somatostatina/efeitos dos fármacos
17.
FEBS Lett ; 582(6): 949-55, 2008 Mar 19.
Artigo em Inglês | MEDLINE | ID: mdl-18298956

RESUMO

The purpose of this study was to determine whether bile acids (BAs) modulate hepatic pro-protein convertase subtilisin/kexin 9 (PCSK9) gene expression. Immortalized human hepatocytes were treated with various BAs. Chenodeoxycholic acid (CDCA) treatment specifically decreased both PCSK9 mRNA and protein contents. Moreover, activation of the BA-activated farnesoid X receptor (FXR) by its synthetic specific agonist GW4064 also decreased PCSK9 expression. Of functional relevance, coadministration of CDCA counteracted the statin-induced PCSK9 expression, leading to a potentiation of LDL receptor activity. This study suggests that a transcriptional repression of PCSK9 by CDCA or FXR agonists may potentiate the hypolipidemic effect of statins.


Assuntos
Ácido Quenodesoxicólico/farmacologia , Proteínas de Ligação a DNA/agonistas , Hepatócitos/efeitos dos fármacos , Receptores Citoplasmáticos e Nucleares/agonistas , Serina Endopeptidases/genética , Fatores de Transcrição/agonistas , Transcrição Gênica/efeitos dos fármacos , Hepatócitos/enzimologia , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Isoxazóis/farmacologia , Pravastatina/farmacologia , Pró-Proteína Convertase 9 , Pró-Proteína Convertases , RNA Mensageiro/antagonistas & inibidores , RNA Mensageiro/metabolismo , Receptores de LDL/agonistas , Receptores de LDL/metabolismo , Serina Endopeptidases/metabolismo
18.
Atherosclerosis ; 243(2): 499-509, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26520906

RESUMO

RATIONALE: Recently, there has been significant interest in the therapeutic administration of miRNA mimics and inhibitors to treat cardiovascular disease. In particular, miR-27b has emerged as a regulatory hub in cholesterol and lipid metabolism and potential therapeutic target for treating atherosclerosis. Despite this, the impact of miR-27b on lipid levels in vivo remains to be determined. As such, here we set out to further characterize the role of miR-27b in regulating cholesterol metabolism in vitro and to determine the effect of miR-27b overexpression and inhibition on circulating and hepatic lipids in mice. METHODS AND RESULTS: Our results identify miR-27b as an important regulator of LDLR activity in human and mouse hepatic cells through direct targeting of LDLR and LDLRAP1. In addition, we report that modulation of miR-27b expression affects ABCA1 protein levels and cellular cholesterol efflux to ApoA1 in human hepatic Huh7 cells. Overexpression of pre-miR-27b in the livers of wild-type mice using AAV8 vectors increased pre-miR-27b levels 50-fold and reduced hepatic ABCA1 and LDLR expression by 50% and 20%, respectively, without changing circulating and hepatic cholesterol and triglycerides. To determine the effect of endogenous miR-27b on circulating lipids, wild-type mice were fed a Western diet for one month and injected with 5 mg/kg of LNA control or LNA anti-miR-27b oligonucleotides. Following two weeks of treatment, the expression of ABCA1 and LDLR were increased by 10-20% in the liver, demonstrating effective inhibition of miR-27b function. Intriguingly, no differences in circulating and hepatic lipids were observed between treatment groups. CONCLUSIONS: The results presented here provide evidence that short-term modulation of miR-27b expression in wild-type mice regulates hepatic LDLR and ABCA1 expression but does not influence plasma and hepatic lipid levels.


Assuntos
Transportador 1 de Cassete de Ligação de ATP/metabolismo , Colesterol/sangue , Dieta Hiperlipídica , Fígado/metabolismo , MicroRNAs/metabolismo , Receptores de LDL/metabolismo , Regiões 3' não Traduzidas , Transportador 1 de Cassete de Ligação de ATP/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Biomarcadores/sangue , Células COS , Chlorocebus aethiops , Biologia Computacional , Bases de Dados Genéticas , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Células Hep G2 , Humanos , Macaca mulatta , Masculino , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Receptores de LDL/genética , Fatores de Tempo , Transfecção , Triglicerídeos/sangue
19.
J Clin Endocrinol Metab ; 98(4): E732-6, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23450051

RESUMO

CONTEXT: PCSK9 (proprotein convertase subtilisin kexin type 9) is a secreted protease that modulates cholesterol homeostasis by decreasing low-density lipoprotein receptor expression. Low levels of plasma lipoproteins are related to severity of illness and survival in patients of intensive care units (ICU). OBJECTIVE: The aim of the study was to investigate the regulation of plasma PCSK9 and its association with plasma lipid parameters and clinical markers of severity during critical illness. DESIGN AND PATIENTS: The plasma biobank from the previously published HYPOLYTE prospective study was used to measure PCSK9 concentrations by ELISA at days 0 and 8 in 111 patients admitted to surgical ICU for severe multiple trauma. Patients were randomly assigned to hydrocortisone therapy or placebo. RESULTS: Plasma PCSK9 levels were increased by 2-fold between days 0 and 8 (231 ± 116 vs 481 ± 227 ng/ml; P = .0001). Hydrocortisone therapy did not alter PCSK9 concentrations (451 ± 216 vs 511 ± 239 ng/ml in placebo group; P = .33). PCSK9 was positively associated with low-density lipoprotein-cholesterol (Pearson coefficient, 0.26; P = .007) at day 0, but not at day 8. At day 8, an inverse correlation was found between PCSK9 and high-density lipoprotein-cholesterol (ß = -653; P = .004). Although baseline PCSK9 concentrations were not associated to severity scores, PCSK9 values at day 8 were related to injury severity score (ß = 6.17; P = .0007), length of stay in ICU (ß = 6.14; P = .0001), and duration of both mechanical ventilation (ß = 8.26; P = .0001) and norepinephrine infusion (ß = 18.57; P = .015). CONCLUSIONS: Plasma PCSK9 appears as a late biomarker of illness severity in patients with severe multiple trauma.


Assuntos
Pró-Proteína Convertases/fisiologia , Serina Endopeptidases/fisiologia , Ferimentos e Lesões/diagnóstico , Adulto , Biomarcadores/sangue , LDL-Colesterol/sangue , Estado Terminal/terapia , Feminino , Humanos , Hidrocortisona/administração & dosagem , Hidrocortisona/uso terapêutico , Bombas de Infusão , Unidades de Terapia Intensiva , Masculino , Placebos , Prognóstico , Pró-Proteína Convertase 9 , Pró-Proteína Convertases/sangue , Serina Endopeptidases/sangue , Índice de Gravidade de Doença , Índices de Gravidade do Trauma , Ferimentos e Lesões/sangue , Ferimentos e Lesões/classificação , Ferimentos e Lesões/tratamento farmacológico
20.
Nutr Metab (Lond) ; 10(1): 4, 2013 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-23298392

RESUMO

BACKGROUND: PCSK9 (Proprotein Convertase Subtilisin Kexin type 9) is a circulating protein that promotes hypercholesterolemia by decreasing hepatic LDL receptor protein. Under non interventional conditions, its expression is driven by sterol response element binding protein 2 (SREBP2) and follows a diurnal rhythm synchronous with cholesterol synthesis. Plasma PCSK9 is associated to LDL-C and to a lesser extent plasma triglycerides and insulin resistance. We aimed to verify the effect on plasma PCSK9 concentrations of dietary interventions that affect these parameters. METHODS: We performed nutritional interventions in young healthy male volunteers and offspring of type 2 diabetic (OffT2D) patients that are more prone to develop insulin resistance, including: i) acute post-prandial hyperlipidemic challenge (n=10), ii) 4 days of high-fat (HF) or high-fat/high-protein (HFHP) (n=10), iii) 7 (HFruc1, n=16) or 6 (HFruc2, n=9) days of hypercaloric high-fructose diets. An acute oral fat load was also performed in two patients bearing the R104C-V114A loss-of-function (LOF) PCSK9 mutation. Plasma PCSK9 concentrations were measured by ELISA. For the HFruc1 study, intrahepatocellular (IHCL) and intramyocellular lipids were measured by 1H magnetic resonance spectroscopy. Hepatic and whole-body insulin sensitivity was assessed with a two-step hyperinsulinemic-euglycemic clamp (0.3 and 1.0 mU.kg-1.min-1). FINDINGS: HF and HFHP short-term diets, as well as an acute hyperlipidemic oral load, did not significantly change PCSK9 concentrations. In addition, post-prandial plasma triglyceride excursion was not altered in two carriers of PCSK9 LOF mutation compared with non carriers. In contrast, hypercaloric 7-day HFruc1 diet increased plasma PCSK9 concentrations by 28% (p=0.05) in healthy volunteers and by 34% (p=0.001) in OffT2D patients. In another independent study, 6-day HFruc2 diet increased plasma PCSK9 levels by 93% (p<0.0001) in young healthy male volunteers. Spearman's correlations revealed that plasma PCSK9 concentrations upon 7-day HFruc1 diet were positively associated with plasma triglycerides (r=0.54, p=0.01) and IHCL (r=0.56, p=0.001), and inversely correlated with hepatic (r=0.54, p=0.014) and whole-body (r=-0.59, p=0.0065) insulin sensitivity. CONCLUSIONS: Plasma PCSK9 concentrations vary minimally in response to a short term high-fat diet and they are not accompanied with changes in cholesterolemia upon high-fructose diet. Short-term high-fructose intake increased plasma PCSK9 levels, independent on cholesterol synthesis, suggesting a regulation independent of SREBP-2. Upon this diet, PCSK9 is associated with insulin resistance, hepatic steatosis and plasma triglycerides.

SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA