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1.
Nutr Metab Cardiovasc Dis ; 33(12): 2317-2325, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37805309

RESUMO

AIMS: Refractory hypercholesterolemia (RH), caused primarily by the loss-of-function mutation of LDL receptor (LDLR) gene seen in HoFH and HeFH patients, remains a major risk factor for atherosclerotic cardiovascular disease (ASCVD). Statin and ezetimibe combination therapy lower circulating LDL by 30% in HoFH patients. PCSK9 mAB, being an LDLR-dependent therapy, is not effective in HoFH, but lowers LDL by 25% in HeFH patients. A maximum reduction of 50% was noted in HoFH patients treated with ANGPTL3 mAB, which was not enough to achieve therapeutic goal of LDL. Therefore, new approaches are warranted to offer hopes to individuals intolerant to higher dose statins and not able to achieve recommended LDL level. DATA SYNTHESIS: New approaches to lower LDL include gene therapy and gene editing. AAV-based gene therapy has shown encouraging results in animal models. Using CRISPR/Cas9-mediated genome/base editing, gain of function and loss of function have been successfully done in animal models. Recent progress in the refinement of genome/base editing has overcome the issues of off-target mutagenesis with ∼1% mutagenesis in case of PCSK9 and almost no off-target mutagenesis in inactivating ANGPTL3 in animal models showing 50% reduction in cholesterol. Current approaches using CRISPR-Cas9 genome/base editing targeting LDLR-dependent and LDLR-independent pathways are underway. CONCLUSIONS: The new information on gain of LDLR function and inactivation of ANGPTL3 together with developments in genome/base editing technology to overcome off-target insertion and deletion mutagenesis offer hope to refractory hypercholesterolemic individuals who are at a higher risk of developing ASCVD.


Assuntos
Hipercolesterolemia Familiar Homozigota , Inibidores de Hidroximetilglutaril-CoA Redutases , Hipercolesterolemia , Animais , Humanos , Hipercolesterolemia/diagnóstico , Hipercolesterolemia/genética , Hipercolesterolemia/terapia , Pró-Proteína Convertase 9/genética , Pró-Proteína Convertase 9/uso terapêutico , Edição de Genes , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Proteína 3 Semelhante a Angiopoietina
2.
Nutr Metab Cardiovasc Dis ; 31(8): 2490-2506, 2021 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-34172319

RESUMO

BACKGROUND AND AIMS: Cholesterol and triglycerides are risk factors for developing cardiovascular disease. Therefore, appropriate cells and assays are required to discover and develop dual cholesterol and fatty acid inhibitors. A predictive hyperlipidemic animal model is needed to evaluate mechanism of action of lead molecule for therapeutic indications. METHODS AND RESULTS: Primary hepatocytes from rat, hamster, rabbit, and humans were compared for suitability to screen compounds by de novo lipogenesis (DNL) using14C-acetate. Hyperlipidemic hamsters were used to evaluate efficacy and mode of action. In rat hepatocytes DNL assay, both the central moiety and carbon chain length influenced the potency of lipogenesis inhibition. In hyperlipidemic hamsters, ETC-1002 decreased plasma cholesterol and triglycerides by 41% and 49% at the 30 mg/kg dose. Concomitant decreases in non-esterified fatty acids (-34%) and increases in ketone bodies (20%) were associated with induction of hepatic CPT1-α. Reductions in proatherogenic VLDL-C and LDL-C (-71% and -64%) occurred partly through down-regulation of DGAT2 and up-regulation of LPL and PDK4. Activation of PLIN1 and PDK4 dampened adipogenesis and showed inverse correlation with adipose mass. Hepatic concentrations of cholesteryl ester and TG decreased by 67% and 64%, respectively. Body weight decreased with concomitant decreases in epididymal fat. Plasma and liver concentrations of ETC-1002 agreed with the observed dose-response efficacy. CONCLUSIONS: Taken together, ETC-1002 reduced proatherogenic lipoproteins, hepatic lipids and adipose tissues in hyperlipidemic hamsters via induction of LPL, CPT1-α, PDK4, and PLIN1, and downregulation of DGAT2. These characteristics may be useful in the treatment of fatty livers that causes non-alcoholic steatohepatitis.


Assuntos
Colesterol/biossíntese , Ácidos Dicarboxílicos/farmacologia , Ácidos Graxos/biossíntese , Hepatócitos/efeitos dos fármacos , Hiperlipidemias/tratamento farmacológico , Hipolipemiantes/farmacologia , Lipogênese/efeitos dos fármacos , Animais , Carnitina O-Palmitoiltransferase/metabolismo , Células Cultivadas , Colesterol/sangue , Dieta Hiperlipídica , Modelos Animais de Doenças , Ácidos Graxos/sangue , Ácidos Graxos/farmacologia , Hepatócitos/enzimologia , Humanos , Hiperlipidemias/sangue , Hiperlipidemias/enzimologia , Lipase Lipoproteica/metabolismo , Masculino , Mesocricetus , Perilipina-1/metabolismo , Proteínas Quinases/metabolismo , Coelhos , Ratos Wistar
3.
Mol Cell Biochem ; 473(1-2): 247-262, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32661772

RESUMO

Reverse cholesterol transport (RCT) and transintestinal cholesterol efflux (TICE) are two important pathways for body cholesterol elimination. We studied these pathways in an animal model of diabetes and obesity (ob/ob) where HDL function is compromised as a result of hyperglycemia, low-grade inflammation and oxidative stress. Co-treatment of ob/ob mice with PPAR-α (fenofibrate) and LXR (T0901317) agonists increased fecal cholesterol by 12-fold; PPAR-α and LXR agonists individually showed 2.6- and 4.0-fold fecal cholesterol excretion, respectively. We investigated the mechanism of synergistic efficacy of PPAR-α and LXR agonists in fecal cholesterol excretion. LXR agonist and the combination of PPAR-α and LXR agonists had greater HDL-C elevation. Ex vivo cholesterol efflux showed correlation with the fecal cholesterol excretion but was not sufficient to explain 12-fold increases in the fecal cholesterol in the co-treated mice. Therefore, we examined TICE to explain the 12-fold increases in the fecal cholesterol. A strong positive correlation of fecal cholesterol with ATP binding cassette transporter G5 (ABCG5) and G8 and a negative correlation with NPC1L1 was observed. ABCG5, G8 and NPC1L1 are involved in intestinal cholesterol absorption. The extent of influence of PPAR-α and LXR agonists on RCT and TICE was distinctly different. PPAR-α agonist increased fecal cholesterol primarily by influencing TICE, while LXR agonist influenced fecal cholesterol excretion via both RCT and TICE mechanisms. Synergistic efficacy on fecal cholesterol excretion following co-treatment with PPAR-α and LXR agonists occurred through a combination of RCT, TICE, and the key enzyme in bile synthesis, cholesterol 7-α hydroxylase (cyp7a1). These results suggest that cholesterol efflux, biliary cholesterol excretion, and TICE collectively contributed to the 12-fold increases in the fecal cholesterol excretion in ob/ob mice co-treated with PPAR-α and LXR agonists.


Assuntos
Membro 5 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Membro 8 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Colesterol/metabolismo , Fezes , Fenofibrato/farmacologia , Hidrocarbonetos Fluorados/farmacologia , Lipoproteínas/metabolismo , Receptores X do Fígado , Proteínas de Membrana Transportadoras/metabolismo , PPAR alfa , Sulfonamidas/farmacologia , Animais , Sinergismo Farmacológico , Fenofibrato/agonistas , Hidrocarbonetos Fluorados/agonistas , Receptores X do Fígado/agonistas , Receptores X do Fígado/metabolismo , Masculino , Camundongos , Camundongos Obesos , PPAR alfa/agonistas , PPAR alfa/metabolismo , Sulfonamidas/agonistas
4.
Mol Cell Biochem ; 440(1-2): 167-187, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28828539

RESUMO

Coronary artery disease, the leading cause of death in the developed and developing countries, is prevalent in diabetes mellitus with 68% cardiovascular disease (CVD)-related mortality. Epidemiological studies suggested inverse correlation between HDL and CVD occurrence. Therefore, low HDL concentration observed in diabetic patients compared to non-diabetic individuals was thought to be one of the primary causes of increased risks of CVD. Efforts to raise HDL level via CETP inhibitors, Torcetrapib and Dalcetrapib, turned out to be disappointing in outcome studies despite substantial increases in HDL-C, suggesting that factors beyond HDL concentration may be responsible for the increased risks of CVD. Therefore, recent studies have focused more on HDL function than on HDL levels. The metabolic environment in diabetes mellitus condition such as hyperglycemia-induced advanced glycation end products, oxidative stress, and inflammation promote HDL dysfunction leading to greater risks of CVD. This review discusses dysfunctional HDL as one of the mechanisms of increased CVD risks in diabetes mellitus through adversely affecting components that support HDL function in cholesterol efflux and LDL oxidation. The dampening of reverse cholesterol transport, a key process that removes cholesterol from lipid-laden macrophages in the arterial wall, leads to increased risks of CVD in diabetic patients. Therapeutic approaches to keep diabetes under control may benefit patients from developing CVD.


Assuntos
Doença da Artéria Coronariana/sangue , Complicações do Diabetes/sangue , Diabetes Mellitus/sangue , Lipoproteínas HDL/sangue , Animais , Proteínas de Transferência de Ésteres de Colesterol/metabolismo , Doença da Artéria Coronariana/tratamento farmacológico , Doença da Artéria Coronariana/etiologia , Doença da Artéria Coronariana/patologia , Complicações do Diabetes/tratamento farmacológico , Complicações do Diabetes/patologia , Diabetes Mellitus/tratamento farmacológico , Diabetes Mellitus/patologia , Humanos , Lipoproteínas LDL/sangue , Oxirredução
5.
Mol Cell Biochem ; 449(1-2): 167-183, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29644527

RESUMO

Inflammation plays a key role in setting the stage leading to atherosclerosis progression, and high-sensitivity C-reactive protein (CRP) has been recognized as a predictor of cardiovascular risk. As a monotherapy and in combination with statins, gemcabene markedly reduced CRP in humans. Present investigation was undertaken to understand the mechanism of CRP reduction. In human hepatoma cells, gemcabene inhibited IL-6 plus IL-1ß-induced CRP production in a concentration-dependent manner, reaching 70% inhibition at 2 mM. In TNF-α-stimulated primary human coronary artery endothelial cells, both CRP and IL-6 productions were reduced by 70% at 2 mM gemcabene concentration. To investigate the mechanism of gemcabene-mediated reduction of CRP, transfection studies were performed with human CRP regulatory sequences in luciferase/ß-gal system that showed 25-fold increase in IL-6- and IL-6 plus IL-1ß-stimulated CRP transcription. Luciferase activity was reduced by 50% by gemcabene, suggesting transcriptional down-regulation of CRP. Site-directed mutagenesis of human CRP promoter revealed that the overlapping downstream C/EBP and NF-κB binding sites are important for gemcabene-mediated CRP transcription. Gel shift assays identified the transcription factor that binds to the downstream CRP promoter as C/EBP-δ. In conclusion, gemcabene decreases CRP by C/EBP-δ and NF-κB-mediated transcriptional mechanism and suppresses IL-6 and IL-1ß-induced CRP production.


Assuntos
Proteína C-Reativa/biossíntese , Proteína delta de Ligação ao Facilitador CCAAT/metabolismo , Caproatos/farmacologia , Regulação para Baixo/efeitos dos fármacos , Transcrição Gênica/efeitos dos fármacos , Linhagem Celular Tumoral , Humanos , Interleucina-1beta/biossíntese , Interleucina-6/biossíntese
6.
J Cardiovasc Pharmacol ; 72(1): 3-10, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29621036

RESUMO

Gemcabene, a late-stage clinical candidate, has shown efficacy for LDL-C, non-HDL cholesterol, apoB, triglycerides, and hsCRP reduction, all risk factors for cardiovascular disease. In rodents, gemcabene showed changes in targets, including apoC-III, apoA-I, peroxisomal enzymes, considered regulated through peroxisome proliferator-activated receptor (PPAR) gene activation, suggesting a PPAR-mediated mechanism of action for the observed hypolipidemic effects observed in rodents and humans. In the current study, the gemcabene agonist activity against PPAR subtypes of human, rat, and mouse were compared with known lipid lowering PPAR activators. Surprisingly, gemcabene showed no or little PPAR-α transactivation compared with reference agonists, which showed concentration-dependent transactivation against human PPAR-α of 2.4- to 30-fold (fenofibric acid), 17-fold (GW590735), and 2.3- to 25-fold (WY-14643). These agents also showed robust transactivation of mouse and rat PPAR-α in a concentration-dependent manner. The known PPAR-δ agonists, GW1516, L165041, and GW0742, showed potent agonist activity against human, mouse, and rat receptors (ranging from 165- to 396-fold). By contrast, gemcabene at the highest concentration tested (300 µM) showed no response in mouse and rat and a marginal response against human PPAR-δ receptors (3.2-fold). For PPAR-γ, gemcabene showed no agonist activity against all 3 species at 100 µM and marginal activity (3.6- to 5-fold) at 300 µM. By contrast, the known agonists, rosiglitazone, indomethacin, and muraglitazar showed strong activation against the mouse, rat, and human PPAR-γ receptors. No clear antagonist activity was observed with gemcabene against any PPAR subtypes for all 3 species over a wide range of concentrations. In summary, the transactivation studies rule out gemcabene as a direct agonist or antagonist of PPAR-α, PPAR-γ, and PPAR-δ receptors of these 3 species. These data suggest that the peroxisomal effects observed in rodents and the lipid regulating effects observed in rodents and humans are not related to a direct activation of PPAR receptors by gemcabene.


Assuntos
Caproatos/farmacologia , Doenças Cardiovasculares/prevenção & controle , Hiperlipidemias/tratamento farmacológico , Hipolipemiantes/farmacologia , Receptores Ativados por Proliferador de Peroxissomo/efeitos dos fármacos , Animais , Doenças Cardiovasculares/sangue , Doenças Cardiovasculares/diagnóstico , Linhagem Celular , Relação Dose-Resposta a Droga , Humanos , Hiperlipidemias/sangue , Hiperlipidemias/diagnóstico , Ligantes , Lipídeos/sangue , Camundongos , Receptores Ativados por Proliferador de Peroxissomo/genética , Receptores Ativados por Proliferador de Peroxissomo/metabolismo , Ratos , Transdução de Sinais/efeitos dos fármacos , Especificidade da Espécie , Transfecção
8.
J Lipid Res ; 54(8): 2095-2108, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23709692

RESUMO

ETC-1002 is an investigational drug currently in Phase 2 development for treatment of dyslipidemia and other cardiometabolic risk factors. In dyslipidemic subjects, ETC-1002 not only reduces plasma LDL cholesterol but also significantly attenuates levels of hsCRP, a clinical biomarker of inflammation. Anti-inflammatory properties of ETC-1002 were further investigated in primary human monocyte-derived macrophages and in in vivo models of inflammation. In cells treated with ETC-1002, increased levels of AMP-activated protein kinase (AMPK) phosphorylation coincided with reduced activity of MAP kinases and decreased production of proinflammatory cytokines and chemokines. AMPK phosphorylation and inhibitory effects of ETC-1002 on soluble mediators of inflammation were significantly abrogated by siRNA-mediated silencing of macrophage liver kinase B1 (LKB1), indicating that ETC-1002 activates AMPK and exerts its anti-inflammatory effects via an LKB1-dependent mechanism. In vivo, ETC-1002 suppressed thioglycollate-induced homing of leukocytes into mouse peritoneal cavity. Similarly, in a mouse model of diet-induced obesity, ETC-1002 restored adipose AMPK activity, reduced JNK phosphorylation, and diminished expression of macrophage-specific marker 4F/80. These data were consistent with decreased epididymal fat-pad mass and interleukin (IL)-6 release by inflamed adipose tissue. Thus, ETC-1002 may provide further clinical benefits for patients with cardiometabolic risk factors by reducing systemic inflammation linked to insulin resistance and vascular complications of metabolic syndrome.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Tecido Adiposo/efeitos dos fármacos , Anti-Inflamatórios não Esteroides/farmacologia , Ácidos Dicarboxílicos/farmacologia , Ácidos Graxos/farmacologia , Leucócitos/efeitos dos fármacos , Macrófagos/enzimologia , Proteínas Serina-Treonina Quinases/metabolismo , Quinases Proteína-Quinases Ativadas por AMP , Tecido Adiposo/citologia , Tecido Adiposo/imunologia , Animais , Relação Dose-Resposta a Droga , Humanos , Inflamação , Leucócitos/citologia , Leucócitos/imunologia , Macrófagos/efeitos dos fármacos , Masculino , Fosforilação , Ratos , Ratos Sprague-Dawley , Relação Estrutura-Atividade
9.
J Lipid Res ; 54(1): 134-51, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23118444

RESUMO

ETC-1002 (8-hydroxy-2,2,14,14-tetramethylpentadecanedioic acid) is a novel investigational drug being developed for the treatment of dyslipidemia and other cardio-metabolic risk factors. The hypolipidemic, anti-atherosclerotic, anti-obesity, and glucose-lowering properties of ETC-1002, characterized in preclinical disease models, are believed to be due to dual inhibition of sterol and fatty acid synthesis and enhanced mitochondrial long-chain fatty acid ß-oxidation. However, the molecular mechanism(s) mediating these activities remained undefined. Studies described here show that ETC-1002 free acid activates AMP-activated protein kinase in a Ca(2+)/calmodulin-dependent kinase ß-independent and liver kinase ß 1-dependent manner, without detectable changes in adenylate energy charge. Furthermore, ETC-1002 is shown to rapidly form a CoA thioester in liver, which directly inhibits ATP-citrate lyase. These distinct molecular mechanisms are complementary in their beneficial effects on lipid and carbohydrate metabolism in vitro and in vivo. Consistent with these mechanisms, ETC-1002 treatment reduced circulating proatherogenic lipoproteins, hepatic lipids, and body weight in a hamster model of hyperlipidemia, and it reduced body weight and improved glycemic control in a mouse model of diet-induced obesity. ETC-1002 offers promise as a novel therapeutic approach to improve multiple risk factors associated with metabolic syndrome and benefit patients with cardiovascular disease.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , ATP Citrato (pro-S)-Liase/metabolismo , Metabolismo dos Carboidratos/efeitos dos fármacos , Ácidos Dicarboxílicos/farmacologia , Inibidores Enzimáticos/farmacologia , Ácidos Graxos/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Terapia de Alvo Molecular/métodos , Quinases Proteína-Quinases Ativadas por AMP , ATP Citrato (pro-S)-Liase/antagonistas & inibidores , Animais , Biomarcadores/sangue , Biomarcadores/metabolismo , Cálcio/metabolismo , Cricetinae , Ácidos Dicarboxílicos/química , Ácidos Dicarboxílicos/uso terapêutico , Dieta/efeitos adversos , Dislipidemias/sangue , Dislipidemias/tratamento farmacológico , Dislipidemias/metabolismo , Metabolismo Energético/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/química , Inibidores Enzimáticos/uso terapêutico , Ácidos Graxos/biossíntese , Ácidos Graxos/química , Ácidos Graxos/uso terapêutico , Feminino , Glucagon/metabolismo , Glucose/biossíntese , Células Hep G2 , Humanos , Fígado/citologia , Fígado/efeitos dos fármacos , Fígado/enzimologia , Fígado/metabolismo , Masculino , Camundongos , Obesidade/sangue , Obesidade/tratamento farmacológico , Obesidade/etiologia , Obesidade/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Ratos , Transdução de Sinais/efeitos dos fármacos , Esteróis/biossíntese
10.
Cells ; 12(12)2023 06 16.
Artigo em Inglês | MEDLINE | ID: mdl-37371118

RESUMO

Since the discovery of the LDL receptor in 1973 by Brown and Goldstein as a causative protein in hypercholesterolemia, tremendous amounts of effort have gone into finding ways to manage high LDL cholesterol in familial hypercholesterolemic (HoFH and HeFH) individuals with loss-of-function mutations in the LDL receptor (LDLR) gene. Statins proved to be the first blockbuster drug, helping both HoFH and HeFH individuals by inhibiting the cholesterol synthesis pathway rate-limiting enzyme HMG-CoA reductase and inducing the LDL receptor. However, statins could not achieve the therapeutic goal of LDL. Other therapies targeting LDLR include PCSK9, which lowers LDLR by promoting LDLR degradation. Inducible degrader of LDLR (IDOL) also controls the LDLR protein, but an IDOL-based therapy is yet to be developed. Among the LDLR-independent pathways, such as angiopoietin-like 3 (ANGPTL3), apolipoprotein (apo) B, apoC-III and CETP, only ANGPTL3 offers the advantage of treating both HoFH and HeFH patients and showing relatively better preclinical and clinical efficacy in animal models and hypercholesterolemic individuals, respectively. While loss-of-LDLR-function mutations have been known for decades, gain-of-LDLR-function mutations have recently been identified in some individuals. The new information on gain of LDLR function, together with CRISPR-Cas9 genome/base editing technology to target LDLR and ANGPTL3, offers promise to HoFH and HeFH individuals who are at a higher risk of developing atherosclerotic cardiovascular disease (ASCVD).


Assuntos
Hipercolesterolemia Familiar Homozigota , Inibidores de Hidroximetilglutaril-CoA Redutases , Hipercolesterolemia , Hiperlipoproteinemia Tipo II , Receptores de LDL , Animais , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Hipercolesterolemia/tratamento farmacológico , Hipercolesterolemia/genética , Hiperlipoproteinemia Tipo II/tratamento farmacológico , Hiperlipoproteinemia Tipo II/genética , Pró-Proteína Convertase 9/genética , Receptores de LDL/genética , Receptores de LDL/metabolismo , Fatores de Risco , Humanos
11.
J Lipid Res ; 53(12): 2490-514, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22798688

RESUMO

The adenosine monophosphate-activated protein kinase (AMPK) is a metabolic sensor of energy metabolism at the cellular as well as whole-body level. It is activated by low energy status that triggers a switch from ATP-consuming anabolic pathways to ATP-producing catabolic pathways. AMPK is involved in a wide range of biological activities that normalizes lipid, glucose, and energy imbalances. These pathways are dysregulated in patients with metabolic syndrome (MetS), which represents a clustering of major cardiovascular risk factors including diabetes, lipid abnormalities, and energy imbalances. Clearly, there is an unmet medical need to find a molecule to treat alarming number of patients with MetS. AMPK, with multifaceted activities in various tissues, has emerged as an attractive drug target to manage lipid and glucose abnormalities and maintain energy homeostasis. A number of AMPK activators have been tested in preclinical models, but many of them have yet to reach to the clinic. This review focuses on the structure-function and role of AMPK in lipid, carbohydrate, and energy metabolism. The mode of action of AMPK activators, mechanism of anti-inflammatory activities, and preclinical and clinical findings as well as future prospects of AMPK as a drug target in treating cardio-metabolic disease are discussed.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Metabolismo dos Carboidratos , Doenças Cardiovasculares/tratamento farmacológico , Ativadores de Enzimas/farmacologia , Ativadores de Enzimas/uso terapêutico , Metabolismo dos Lipídeos , Proteínas Quinases Ativadas por AMP/química , Animais , Doenças Cardiovasculares/enzimologia , Doenças Cardiovasculares/metabolismo , Ativadores de Enzimas/administração & dosagem , Humanos
12.
Biochim Biophys Acta ; 1811(2): 76-83, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21081177

RESUMO

Fenofibrate, a PPAR-α agonist, lowers triglycerides (TG) and raises high-density lipoproteins (HDL-C) in humans. While fenofibrate is very effective in lowering TG, it does not raise HDL-C in humans to the same extent as seen in human apoAI transgenic (hAI-Tg) mice. We studied the mechanism of this discordance using the following compounds as tools: cholic acid that down-regulates human apoAI, and fenofibrate, that elevates hapoAI and HDL-C in hAI-Tg mice. We hypothesized that additional sequences, including apoCIII and AIV genes on chromosome 11, not present in the hapoAI transgene may be responsible for the dampened effect of fibrates on HDL-C seen in humans. For this, hAI-Tg mice with 11kb DNA segment and hapoAI-CIII-AIV-Tg mice with 33kb DNA segment harboring apoCIII and AIV genes were employed. These mice were treated with fenofibrate and cholic acid. Fenofibrate increased apoAI and HDL-C levels, and HDL size in the apoAI-Tg mice via up-regulation of the hapoAI mRNA and increased activity and mRNA of PLTP, respectively. Consistent with earlier findings, cholic acid showed similar effects of lowering HDL-C, and elevating LDL-C in hAI-Tg mice as well as in the hAI-CIII-AIV-Tg mice. Fenofibrate decreased TG and increased HDL size in hAI-CIII-AIV-Tg mice as well, but surprisingly, did not elevate serum levels of hapoAI or hepatic AI mRNA, suggesting that additional sequences not present in the hapoAI transgene (11kb) may be partly responsible for the dampened effect on HDL-C seen in hAI-CIII-AIV-Tg mice. Since hAI-CIII-AIV-Tg mouse mimics fenofibrate effects seen in humans, this transgenic mouse could serve as a better predictive model for screening HDL-C raising compounds.


Assuntos
Apolipoproteína A-I/sangue , Fenofibrato/metabolismo , Hipolipemiantes/metabolismo , Lipoproteínas HDL/sangue , Animais , Apolipoproteína A-I/genética , Antígenos CD36/genética , Antígenos CD36/metabolismo , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Ácido Cólico/metabolismo , Humanos , Lipoproteínas HDL/genética , Camundongos , Camundongos Transgênicos , Triglicerídeos/sangue
13.
Biochem Biophys Res Commun ; 398(4): 665-70, 2010 Aug 06.
Artigo em Inglês | MEDLINE | ID: mdl-20609357

RESUMO

To study the mechanism of low levels of full length and truncated apoB in individuals heterozygous for apoB truncation, a non-sense mutation was introduced in one of the three alleles of apob gene of HepG2 cells by homologous recombination. Despite very low levels of apoB-82 (1-2%) in the media, a prominent N-terminal apoB protein of 85 kDa (apoB-15) was secreted that fractionated at d>1.065 in density gradient ultracentrifugation. The mechanism of production of this short protein was studied by (35)S-methionine pulse-chase experiment. Oleate prevented presecretory degradation of apoB-100 in the cell and resulted in increased secretion of newly synthesized apoB-100 with decreases in the apoB-15, suggesting that rescue of pre-secretary intracellular degradation of apoB restricted the production and secretion of apoB-15. Further investigation on the degradation of transmembrane forms of apoB, in the presence and absence of a cysteine protease inhibitor, N-acetyl-leucyl-leucyl-norleucinal (ALLN), showed appearance of detectable levels of newly synthesized apoB-82 in the cell and the media together with increased apoB-100 secretion, and reduction in the secretion of apoB-15. Compared to ER membrane, the levels of apoB were higher in the luminal content, and presence of both oleate and ALLN had additive effect on apoB secretion. These results suggest that the presence of improper folding of apoB during translocation led to the cleavage of both apoB-100 and apoB-82 by ALLN-sensitive protease and generation of 85 kDa N-terminal fragment of apoB.


Assuntos
Apolipoproteínas B/metabolismo , Cisteína Proteases/metabolismo , Hipobetalipoproteinemia Familiar por Apolipoproteína B/metabolismo , Apolipoproteínas B/química , Apolipoproteínas B/genética , Códon sem Sentido , Inibidores de Cisteína Proteinase/farmacologia , Células Hep G2 , Humanos , Hipobetalipoproteinemia Familiar por Apolipoproteína B/genética , Leupeptinas/farmacologia , Dobramento de Proteína , Transporte Proteico
14.
Mol Cell Biochem ; 345(1-2): 197-206, 2010 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-20740305

RESUMO

Cholesterol ester transfer protein (CETP) and apolipoprotein (apo) E are important in peroxisome proliferation activated receptor-α (PPAR-α)-mediated regulation of lipoprotein metabolism. Therefore, popularly used apolipoprotein E knockout mice are not suitable to evaluate PPAR-α agonists. In this study, we aimed to: a) evaluate hamster as a model for insulin resistance, hyperlipidemia and atherosclerosis; and b) investigate the effect of a PPAR-α activator, fenofibrate, in this model. A high fat high cholesterol (HFHC) diet increased serum cholesterol and triglycerides, but inclusion of fenofibrate in the diet decreased cholesterol and proatherogenic lipoproteins, VLDL and LDL, in a time-dependent manner. Concomitantly, serum levels of triglycerides also decreased. These reductions were attributed, in part, to the down-regulation of lipogenic genes and upregulation of lipoprotein lipase. The HFHC diet caused body weight gain and mild insulin resistance, both of which were prevented following the treatments with fenofibrate. Insulin resistance was further investigated in high fructose-fed hamsters. Fenofibrate prevented both hyperinsulinemia and hypertriglyceridemia. The insulin sensitizing activity of fenofibrate appeared to occur via reductions in protein tyrosine phophatase-1B. To determine whether lowering of lipids by fenofibrate treatment contributed to the reduced risks of developing atherosclerosis in hyperlipidemic hamsters, we measured lipid deposition in the aorta. Our results showed that fenofibrate treatment reduced aortic lipid deposition by 70%. These findings suggest that hamster may be an adequate animal model to evaluate the efficacy of lipid lowering, insulin sensitizing and antiatherosclerotic agents. We also show that fenofibrate is an effective antiatherosclerotic agent in hyperlipidemic hamster model.


Assuntos
Aorta/patologia , Aterosclerose/tratamento farmacológico , Fenofibrato/farmacologia , Hiperlipidemias/tratamento farmacológico , Resistência à Insulina , Metabolismo dos Lipídeos/efeitos dos fármacos , Animais , Aterosclerose/prevenção & controle , Colesterol/administração & dosagem , Colesterol/sangue , Cricetinae , Gorduras na Dieta/administração & dosagem , Gorduras na Dieta/sangue , Fenofibrato/uso terapêutico , Hipolipemiantes/farmacologia , Mesocricetus , Modelos Animais , PPAR alfa/agonistas , Aumento de Peso
15.
J Diabetes Metab Disord ; 19(1): 363-371, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32550187

RESUMO

OBJECTIVES: ATP-binding cassette transporter A1 (ABCA1) is a key player in the reverse cholesterol transport (RCT) and HDL biogenesis. Since RCT is compromised as a result of ABCA1 dysfunction in diabetic state, the objective of this study was to investigate the regulation of ABCA1 in a stably transfected 293 cells expressing ABCA1 under the control of cAMP response element. METHODS: To delineate transcriptional and posttranscriptional regulation of ABCA1, 293 cells were stably transfected with the full length ABCA1 cDNA under the control of CMV promoter harboring cAMP response element. cAMP-mediated regulation of ABCA1 and cholesterol efflux were studied in the presence of 8-Br-cAMP and after withdrawal of 8-Br-cAMP. The mechanism of cAMP-mediated transcriptional induction of the ABCA1 gene was studied in protein kinase A (PKA) inhibitors-treated cells. RESULTS: The transfected 293 cells expressed high levels of ABCA1, while non-transfected wild-type 293 cells showed very low levels of ABCA1. Treatments of transfected cells with 8-Br-cAMP increased ABCA1 protein by 10-fold and mRNA by 20-fold. Cholesterol efflux also increased in parallel. Withdrawal of 8-Br-cAMP caused time-dependent rapid diminution of ABCA1 protein and mRNA, suggesting ABCA1 regulation at the transcriptional level. Treatment with PKA inhibitors abolished the cAMP-mediated induction of the ABCA1 mRNA and protein, resulting dampening of ABCA1-dependent cholesterol efflux. CONCLUSIONS: These results demonstrate that transfected cell line mimics cAMP response similar to normal cells with natural ABCA1 promoter and suggest that ABCA1 is a short-lived protein primarily regulated at the transcriptional level to maintain cellular cholesterol homeostasis.

16.
Phytomedicine ; 52: 117-128, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30599890

RESUMO

BACKGROUND: Traditional therapy using natural products, especially flavonoids and alkaloids have been in practice for a long time. Among flavonoids, curcumin, quercetin, berberine, and epigallocatechin have been studied in greater detail in terms of their anticancer and anti-inflammatory activities. Although many studies focused on the PI3K, MAP kinase and NF-κB pathways, a thorough investigation of modulation of players in the apoptotic and Wnt/ß-catenin signaling pathway by curcumin and quercetin has not been done. Also, only few studies have been carried out on curcumin and quercetin co-treatment studies. HYPOTHESIS/PURPOSE: We hypothesized that the combination of natural products will have synergistic effects and the antiproliferative effect will be attenuated via apoptotic as well as Wnt/ß-catenin signaling pathways. STUDY DESIGN AND METHODS: To test our hypothesis, we compared potency of natural anticancer agents in four cancer cell lines, A549, HCT116, MCF7, and A375 by MTT and colony proliferation assays and investigated mechanism of anticancer activities by analyzing players in apoptotic and Wnt/ß-catenin signaling pathways in A375 cells treated with test agents individually or in combination. RESULTS: Epicatechins, up to 100 µM concentration, did not inhibit cancer cell proliferation, while curcumin inhibited proliferation in A549 and HCT116 cancer cell lines with an IC50 of 3 to 8.5 µM. Quercetin showed stronger inhibition of cell proliferation than berberine. Combination study with two most potent agents, curcumin and quercetin, in 4 cancer cell lines, suggested synergistic effect on cell proliferation with several fold decreases in IC50. Further investigation of the mechanism of action of curcumin and quercetin in melanoma cells, A375, suggested that inhibition of cell proliferation occurred through down-regulation of Wnt/ß-catenin signaling pathway proteins, DVL2, ß-catenin, cyclin D1, Cox2, and Axin2. In addition, both curcumin and quercetin induced apoptosis by down-regulating BCL2 and inducing caspase 3/7 through PARP cleavage. CONCLUSION: These results demonstrate that curcumin and quercetin inhibit cancer cell proliferation synergistically and Wnt/ß-catenin signaling and apoptotic pathways are partly responsible for antiproliferative activities.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Curcumina/farmacologia , Quercetina/farmacologia , Via de Sinalização Wnt/efeitos dos fármacos , Proteína Axina/metabolismo , Linhagem Celular Tumoral , Ciclina D1/metabolismo , Ciclo-Oxigenase 2/metabolismo , Regulação para Baixo , Humanos , beta Catenina/metabolismo
17.
Indian J Biochem Biophys ; 45(6): 410-5, 2008 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19239128

RESUMO

Apolipoprotein (apo) E plays an important role in the whole body cholesterol homeostasis. Recent studies suggest that it may also be involved in the local cholesterol transport in the brain, and influence the pathogenesis of Alzheimer's disease (AD) by interacting with the beta-amyloid protein and brain lipoprotein receptors. Since apoE expression is highest in the brain, next only to the liver and associated with the pathogenesis of AD, we hypothesized that dietary and hormonal intervention, known to regulate hepatic apoE expression may also regulate brain apoE and thereby influence local cholesterol transport. To test this hypothesis, groups of male C57BL mice were fed either regular rodent chow or high fat (HF) and high cholesterol enriched diet for 3 weeks. In a separate study, groups of male mice were administered pharmacological doses of 17-beta estradiol for 5 consecutive days and sacrificed on the 6th day. As expected, HF diet elevated liver apoE mRNA and apoE synthesis. Similar to liver, brain apoE mRNA and synthesis also increased, following HF feeding. Estradiol administration increased liver apoE synthesis without affecting apoE mRNA. Interestingly, estradiol administration also increased the brain apoE synthesis, but without altering the brain apoE mRNA. These studies suggested that dietary cholesterol and estrogen administration elevated the brain apoE by different mechanisms.


Assuntos
Apolipoproteínas E/biossíntese , Encéfalo/metabolismo , Colesterol na Dieta/administração & dosagem , Estrogênios/farmacologia , Animais , Apolipoproteínas E/genética , Transporte Biológico Ativo , Colesterol/biossíntese , Feminino , Regulação da Expressão Gênica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , RNA Mensageiro/biossíntese
18.
J Diabetes Metab Disord ; 17(2): 381-391, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30918873

RESUMO

Energy imbalance resulting from high calorie food intake and insufficient metabolic activity leads to increased body mass index (BMI) and sets the stage for metabolic derangement influencing lipid and carbohydrate metabolism and ultimately leading to insulin resistance, dyslipidemia, and type 2 diabetes. 70% of cardiovascular disease (CVD) deaths occur in patients with diabetes. Environment-induced physiological perturbations trigger epigenetic changes through chromatin modification and leads to type 2 diabetes and progression of nonalcoholic fatty liver disease (NAFLD) and CVD. Thus, in terms of disease progression and pathogenesis, energy homeostasis, metabolic dysregulation, diabetes, fatty liver, and CVD are interlinked. Since advanced glycation end products (AGEs) and low-grade inflammation in type 2 diabetes play definitive roles in the pathogenesis of liver and vascular diseases, a natural checkpoint to prevent diabetes and associated complications appears to be the identification and management of prediabetes together with weight management, since 70% of prediabetic individuals develop diabetes during their life time, and every kg of weight increase is associated with up to 9% increase in diabetes risk. A good proportion of diabetes and obesity population have fatty liver that progresses to non-alcoholic steatohepatitis (NASH) and cirrhosis, and increased risk of hepatocellular carcinoma. Diabetes and NASH both have elevated oxidative stress, impaired cholesterol elimination, and increased inflammation that leads to CVD risk. This review addresses life-style-induced metabolic pathway derangement and how it contributes to epigenetic changes, type 2 diabetes and NASH progression, which collectively lead to increased risk of CVD.

19.
Nutr Metab (Lond) ; 12: 20, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26064179

RESUMO

A large body of evidence suggests that atherosclerosis is an inflammatory disease, in which cytokines and growth factors play a major role in disease progression. The methanolic extracts of Sphaeranthus indicus as well as its active ingredient, 7-hydroxy frullanoide (7-HF), are shown to suppress LPS-induced cytokine production from mononuclear cells, and inhibit the expression of VCAM1, ICAM1 and E-selectin by TNF-α- stimulated HUVECs in a concentration-dependent manner. We tested the hypothesis that the inhibition of cytokines and adhesion molecules should attenuate the progression of atherosclerosis, independent of changes in the lipid profile. Studies were carried out in two animal models: a high fat-fed LDLr(-/-) mouse and a high fat-fed hyperlipidemic hamster. Methanolic extract of S. indicus was dosed to hyperlipidemic LDLr(-/-) at 100 and 300 mg (equivalent to 20 and 60 mg 7-HF)/kg body weight/ day for 8 weeks, and plasma lipids as well as aortic lesion area were quantitated. Hyperlipidemic hamsters were treated with one dose of 200 mg/kg/day. S. indicus extract treatment did not alter the lipid profile in both animal models, but reduced aortic lesion area in LDLr(-/-) mice and hyperlipidemic hamsters by 22 % and 45 %, respectively. Fenofibrate, included as a reference agent, decreased aortic lesions by 26 % in LDLr (-/-) mice and 84 % in hyperlipidemic hamsters, respectively, which was driven by massive reductions in proatherogenic lipoproteins. The lipid-independent anti-atherosclerotic activity of S. indicus was associated with the reductions in the circulating levels of MCP-1, TNF-α, and IL-6 via phosphorylation and degradation of IkB-α that prevents translocation of NF-kB in the nucleus to induce proinflammatory cytokines. Our findings demonstrate that anti-inflammatory agents that lower pro-inflammatory proteins inhibit the progression of atherosclerosis. The methanolic extract of S. inducus, currently being used to treat psoriasis, offer promise to benefit individuals who have high circulating pro-inflammatory cytokines, and predisposed to coronary artery disease.

20.
J Neurol Sci ; 210(1-2): 11-8, 2003 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-12736081

RESUMO

The scavenger receptor class B type I (SR-BI), a receptor for high-density lipoproteins (HDL), facilitates cholesterol delivery to steroidogenic tissues, and brings excess body cholesterol to liver for excretion. Scavenger receptors are also involved in the internalization of aggregates of Alzheimer's disease (AD) amyloid beta-protein, and selective uptake of HDL-associated vitamin E in the brain. Therefore, modulation of the brain SR-BI may affect these processes. The present study examined the expression of SR-BI receptors in murine brain and their regulation by estradiol administration and cholesterol feeding. Liver and brain appeared to express similar SR-BI transcripts. Expression of SR-BI was highest in the adrenals and lowest in the brain. In rats, estradiol administration decreased SR-BI in liver, but increased in adrenals. In mice, estrogen treatment decreased hepatic SR-BI, but interestingly increased the levels of brain SR-BI mRNA. Cholesterol feeding did not alter mouse hepatic SR-BI mRNA, but increased brain SR-BI levels. ATP-binding cassette transporter A1 (ABCA1), involved in cellular cholesterol transport, increased in cholesterol-fed mouse liver, but did not show changes in the brain. These studies suggest that SR-B1 is expressed in the brain and regulated by hormonal and nutritional stimuli, which may influence the pathophysiology of neurological disorders like AD.


Assuntos
Encéfalo/metabolismo , Antígenos CD36/metabolismo , Colesterol na Dieta/farmacologia , Estradiol/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas de Membrana , Receptores Imunológicos , Receptores de Lipoproteínas , Glândulas Suprarrenais/efeitos dos fármacos , Glândulas Suprarrenais/metabolismo , Animais , Northern Blotting/métodos , Western Blotting/métodos , Antígenos CD36/genética , Clonagem Molecular/métodos , Mucosa Intestinal/metabolismo , Intestinos/efeitos dos fármacos , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , RNA Mensageiro/biossíntese , Ratos , Ratos Sprague-Dawley , Receptores Depuradores , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Receptores Depuradores Classe B , Testículo/efeitos dos fármacos , Testículo/metabolismo
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