RESUMO
Genetic testing is crucial in inherited arrhythmogenic channelopathies; however, the clinical interpretation of genetic variants remains challenging. Incomplete penetrance, oligogenic, polygenic or multifactorial forms of channelopathies further complicate variant interpretation. We identified the KCNQ1/p.D446E variant in 2/63 patients with long QT syndrome, 30-fold more frequent than in public databases. We thus characterized the biophysical phenotypes of wildtype and mutant IKs co-expressing these alleles with the ß-subunit minK in HEK293 cells. KCNQ1 p.446E homozygosity significantly shifted IKs voltage dependence to hyperpolarizing potentials in basal conditions (gain of function) but failed to shift voltage dependence to hyperpolarizing potentials (loss of function) in the presence of 8Br-cAMP, a protein kinase A activator. Basal IKs activation kinetics did not differ among genotypes, but in response to 8Br-cAMP, IKs 446 E/E (homozygous) activation kinetics were slower at the most positive potentials. Protein modeling predicted a slower transition of the 446E Kv7.1 tetrameric channel to the stabilized open state. In conclusion, biophysical and modelling evidence shows that the KCNQ1 p.D446E variant has complex functional consequences including both gain and loss of function, suggesting a contribution to the pathogenesis of arrhythmogenic phenotypes as a functional risk allele.
Assuntos
Arritmias Cardíacas , Canalopatias , Canal de Potássio KCNQ1 , Humanos , Alelos , Arritmias Cardíacas/genética , Proteínas Quinases Dependentes de AMP Cíclico , Células HEK293 , Canal de Potássio KCNQ1/genética , FenótipoRESUMO
INTRODUCTION AND OBJECTIVES: Metabolic-associated fatty liver disease (MAFLD) is defined by steatosis in more than 5% of hepatocytes without other liver diseases. Patients with this disease can progress to multiple stages like liver fibrosis, cirrhosis, and hepatocellular carcinoma. miRNAs are single-stranded molecules that regulate metabolic homeostasis; their differential expression postulates them as potential circulating biomarkers for MAFLD. Previous research reported that hsa-miR-140-5p, hsa-miR-148-5p, and hsa-miR-122-3p have a differential expression in patients with MAFLD. This study aimed to investigate the correlation between liver hsa-miR-140-5p, hsa-miR-148-5p, and hsa-miR-122-3p and serum biomarkers CK-18, APOB, IL-6, IL-32, and TNF-α in patients with MAFLD compared with control patients. MATERIALS AND METHODS: A cross-sectional study was carried out with 16 patients of both sexes, aged between 18-60 years, to determine the association between the levels of hsa-miR-140-5p, hsa-miR-148-5p, and hsa-miR-122-3p with MAFLD in liver biopsies of patients who underwent laparoscopic cholecystectomy. RESULTS: Twelve patients presented MAFLD, four without hepatic steatosis. Circulating levels of CK-18 showed a significant difference in patients with MAFLD, and a strong correlation was found between hsa-miR-122-3p, hsa-miR-140-5p, and hsa-miR-148b-5p versus the CAP value. CONCLUSION: There is a correlation between elevated tissue expression of hsa-miR-122-3p, hsa-miR-140-5p, and hsa-miR-148b-3p with plasma levels of CK-18 in patients with simple steatosis compared with patients without the disease.
Assuntos
Queratina-18 , MicroRNAs , Hepatopatia Gordurosa não Alcoólica , Adolescente , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adulto Jovem , Biomarcadores , Estudos Transversais , Queratina-18/genética , Fígado/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Hepatopatia Gordurosa não Alcoólica/genéticaRESUMO
ABSTRACT Background: The inflammatory response in gout disease is induced by the activation of NLR family pyrin domain-containing 3 (NLPR3) signaling pathway mediated by IL-1β release. Objective: The objective of the study was to determine the association between single nucleotide polymorphisms (SNPs) within NLRP3 inflammasome genes and gout susceptibility. Methods: Mexican patients with gout from the National Rehabilitation Institute and General Hospital of Mexico were enrolled. A healthy control group was also included. We analyzed the frequency and allelic distribution of eight SNPs from seven different genes within the NLRP3 inflammasome signaling pathway: TLR4 rs2149356, CD14 rs2569190, NLRP3 rs3806268, NLRP3 rs10754558, CARD8 rs2043211, IL-1β rs1143623, P2RX7 rs3751142, and PPARGC1B rs45520937 SNPs. Results: We found that the SNP rs45520937 of PPARGC1B was associated with the risk of developing gout when it was analyzed using the dominant model (Odds ratio [OR] = 2.30; 95% confidence interval [CI]: 1.09-4.86; p = 0.030), and it is proposed that the adaptor molecule CD14 rs2569190 polymorphism could be associated with a lower risk of gout under an additive model (OR= 0.41;95% CI: 0.16-1.05; p = 0.064). No significant associations were identified for the remaining SNPs. Conclusion: Our findings suggest that the PPARGC1B rs45520937 SNP is associated with gout susceptibility.
RESUMO
Background: The inflammatory response in gout disease is induced by the activation of NLR family pyrin domain-containing 3 (NLPR3) signaling pathway mediated by IL-1ß release. Objective: The objective of the study was to determine the association between single nucleotide polymorphisms (SNPs) within NLRP3 inflammasome genes and gout susceptibility. Methods: Mexican patients with gout from the National Rehabilitation Institute and General Hospital of Mexico were enrolled. A healthy control group was also included. We analyzed the frequency and allelic distribution of eight SNPs from seven different genes within the NLRP3 inflammasome signaling pathway: TLR4 rs2149356, CD14 rs2569190, NLRP3 rs3806268, NLRP3 rs10754558, CARD8 rs2043211, IL-1ß rs1143623, P2RX7 rs3751142, and PPARGC1B rs45520937 SNPs. Results: We found that the SNP rs45520937 of PPARGC1B was associated with the risk of developing gout when it was analyzed using the dominant model (Odds ratio [OR] = 2.30; 95% confidence interval [CI]: 1.09-4.86; p = 0.030), and it is proposed that the adaptor molecule CD14 rs2569190 polymorphism could be associated with a lower risk of gout under an additive model (OR= 0.41;95% CI: 0.16-1.05; p = 0.064). No significant associations were identified for the remaining SNPs. Conclusion: Our findings suggest that the PPARGC1B rs45520937 SNP is associated with gout susceptibility.
Assuntos
Gota , Inflamassomos , Proteínas Adaptadoras de Sinalização CARD/genética , Proteínas Adaptadoras de Sinalização CARD/metabolismo , Predisposição Genética para Doença , Genótipo , Gota/genética , Humanos , Inflamassomos/genética , Inflamassomos/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Polimorfismo de Nucleotídeo Único , Proteínas de Ligação a RNA/genéticaRESUMO
Objective: Low HDL-C (high-density lipoprotein cholesterol) is the most frequent dyslipidemia in Mexicans, but few studies have examined the underlying genetic basis. Our purpose was to identify genetic variants associated with HDL-C levels and cardiovascular risk in the Mexican population. Approach and Results: A genome-wide association studies for HDL-C levels in 2335 Mexicans, identified four loci associated with genome-wide significance: CETP, ABCA1, LIPC, and SIDT2. The SIDT2 missense Val636Ile variant was associated with HDL-C levels and was replicated in 3 independent cohorts (P=5.9×10−18 in the conjoint analysis). The SIDT2/Val636Ile variant is more frequent in Native American and derived populations than in other ethnic groups. This variant was also associated with increased ApoA1 and glycerophospholipid serum levels, decreased LDL-C (low-density lipoprotein cholesterol) and ApoB levels, and a lower risk of premature CAD. Because SIDT2 was previously identified as a protein involved in sterol transport, we tested whether the SIDT2/Ile636 protein affected this function using an in vitro site-directed mutagenesis approach. The SIDT2/Ile636 protein showed increased uptake of the cholesterol analog dehydroergosterol, suggesting this variant affects function. Finally, liver transcriptome data from humans and the Hybrid Mouse Diversity Panel are consistent with the involvement of SIDT2 in lipid and lipoprotein metabolism. Conclusions: This is the first genome-wide association study for HDL-C levels seeking associations with coronary artery disease in the Mexican population. Our findings provide new insight into the genetic architecture of HDL-C and highlight SIDT2 as a new player in cholesterol and lipoprotein metabolism in humans.
Assuntos
HDL-Colesterol/sangue , Doença da Artéria Coronariana/genética , Hiperlipoproteinemia Tipo II/genética , Proteínas de Transporte de Nucleotídeos/genética , Polimorfismo de Nucleotídeo Único , Adulto , Idade de Início , Animais , Biomarcadores/sangue , Estudos de Casos e Controles , Criança , Doença da Artéria Coronariana/sangue , Doença da Artéria Coronariana/diagnóstico , Doença da Artéria Coronariana/epidemiologia , Modelos Animais de Doenças , Feminino , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Células HEK293 , Fatores de Risco de Doenças Cardíacas , Humanos , Hiperlipoproteinemia Tipo II/sangue , Hiperlipoproteinemia Tipo II/diagnóstico , Hiperlipoproteinemia Tipo II/epidemiologia , Masculino , Análise da Randomização Mendeliana , México/epidemiologia , Camundongos , Pessoa de Meia-Idade , Proteínas de Transporte de Nucleotídeos/metabolismo , Fenótipo , Medição de RiscoRESUMO
Cholesterol homeostasis is essential in normal physiology of all cells. One of several proteins involved in cholesterol homeostasis is the ATP-binding cassette transporter A1 (ABCA1), a transmembrane protein widely expressed in many tissues. One of its main functions is the efflux of intracellular free cholesterol and phospholipids across the plasma membrane to combine with apolipoproteins, mainly apolipoprotein A-I (Apo A-I), forming nascent high-density lipoprotein-cholesterol (HDL-C) particles, the first step of reverse cholesterol transport (RCT). In addition, ABCA1 regulates cholesterol and phospholipid content in the plasma membrane affecting lipid rafts, microparticle (MP) formation and cell signaling. Thus, it is not surprising that impaired ABCA1 function and altered cholesterol homeostasis may affect many different organs and is involved in the pathophysiology of a broad array of diseases. This review describes evidence obtained from animal models, human studies and genetic variation explaining how ABCA1 is involved in dyslipidemia, coronary heart disease (CHD), type 2 diabetes (T2D), thrombosis, neurological disorders, age-related macular degeneration (AMD), glaucoma, viral infections and in cancer progression.
Assuntos
Transportador 1 de Cassete de Ligação de ATP/metabolismo , Colesterol/metabolismo , Transportador 1 de Cassete de Ligação de ATP/deficiência , Transportador 1 de Cassete de Ligação de ATP/genética , Envelhecimento/genética , Envelhecimento/metabolismo , Animais , Doenças Transmissíveis/etiologia , Doença das Coronárias/etiologia , Diabetes Mellitus Tipo 2/etiologia , Dislipidemias/etiologia , Dislipidemias/metabolismo , Oftalmopatias/etiologia , Variação Genética , Humanos , Resistência à Insulina , Lipídeos/sangue , Hepatopatias/etiologia , Malária/etiologia , MicroRNAs/genética , Modelos Biológicos , Mutação , Neoplasias/etiologia , Doenças do Sistema Nervoso/etiologia , Doença de Tangier/etiologiaRESUMO
Osteoarthritis (OA) is the gradual loss of articular cartilage and decrease in subchondral space. One of the risk factors Exposure to cadmium (Cd) through tobacco smoke has been identified as a major OA risk factor. There are no reports addressing the role of Cd in OA progression at the molecular level. Our findings revealed that Cd can promote the activation of metalloproteinases (MMP1, MMP3, MMP9 y MMP13), affecting the expression of COL2A1 and ACAN, and decreasing the presence of glycosaminoglycans and proteoglycans through an inflammatory response related to IL-1ß y a IL-6, as well as oxidative by producing ROS like O2-⢠and H2O2. In conclusion, our findings suggest a cytotoxic role of Cd in the articular cartilage, which could affect OA development.
Assuntos
Cádmio/toxicidade , Cartilagem Articular/efeitos dos fármacos , Substâncias Perigosas/toxicidade , Osteoartrite , Animais , Humanos , Interleucina-1beta , MetaloproteasesRESUMO
Non-alcoholic fatty liver disease (NAFLD) encompasses a broad spectrum of histopathological changes ranging from non-inflammatory intracellular fat deposition to non-alcoholic steatohepatitis (NASH), which may progress into hepatic fibrosis, cirrhosis, or hepatocellular carcinoma. Recent data suggest that impaired hepatic cholesterol homeostasis and its accumulation are relevant to the pathogenesis of NAFLD/NASH. Despite a vital physiological function of cholesterol, mitochondrial dysfunction is an important consequence of dietary-induced hypercholesterolemia and was, subsequently, linked to many pathophysiological conditions. The aim in the current study was to evaluate the morphological and molecular changes of cholesterol overload in mouse liver and particularly, in mitochondria, induced by a high-cholesterol (HC) diet for one month. Histopathological studies revealed microvesicular hepatic steatosis and significantly elevated levels of liver cholesterol and triglycerides leading to impaired liver synthesis. Further, high levels of oxidative stress could be determined in liver tissue as well as primary hepatocyte culture. Transcriptomic changes induced by the HC diet involved disruption in key pathways related to cell death and oxidative stress as well as upregulation of genes related to glutathione homeostasis. Impaired liver function could be associated with a decrease in mitochondrial membrane potential and ATP content and significant alterations in mitochondrial dynamics. We demonstrate that cholesterol overload in the liver leads to mitochondrial changes which may render damaged hepatocytes proliferative and resistant to cell death whereby perpetuating liver damage.
Assuntos
Apoptose , Colesterol na Dieta , Dieta Hiperlipídica , Hepatócitos/patologia , Fígado/patologia , Mitocôndrias Hepáticas/patologia , Dinâmica Mitocondrial , Hepatopatia Gordurosa não Alcoólica/patologia , Animais , Apoptose/genética , Proliferação de Células , Células Cultivadas , Modelos Animais de Doenças , Regulação da Expressão Gênica , Hepatócitos/metabolismo , Fígado/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Mitocôndrias Hepáticas/metabolismo , Dinâmica Mitocondrial/genética , Hepatopatia Gordurosa não Alcoólica/genética , Hepatopatia Gordurosa não Alcoólica/metabolismo , Estresse Oxidativo , Fatores de Tempo , TranscriptomaRESUMO
Metabolic factors are the major risk of non-alcoholic fatty liver disease, although other factors may contribute steatosis. Cadmium exposure produces histopathological and molecular changes in liver, which are consistent with steatosis. In the present study, we describe the effect of low cadmium acute treatment on hepatocytes obtained from mice fed with a high cholesterol diet. Our data suggest that hepatocytes with cholesterol overload promote an adaptive response against cadmium-induced acute toxicity by up-regulating anti-apoptotic proteins, managing ROS overproduction, increasing GSH synthesis and MT-II content to avoid protein oxidation. Cadmium treatment increases lipid content in cholesterol-fed mice hepatocytes because of an impaired autophagy process. Our data suggest an essential function of macroautophagy in the regulation of lipid storage induced by Cd on hepatocytes, that implies that alterations in this pathway may be a mechanism that aggravates hepatic steatosis.
Assuntos
Cloreto de Cádmio/toxicidade , Fígado Gorduroso/etiologia , Hepatócitos/efeitos dos fármacos , Hiperlipidemias/etiologia , Animais , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Colesterol/administração & dosagem , Dieta/efeitos adversos , Fígado Gorduroso/induzido quimicamente , Fígado Gorduroso/patologia , Hepatócitos/patologia , Hiperlipidemias/induzido quimicamente , Hiperlipidemias/patologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Camundongos , Estresse Oxidativo/efeitos dos fármacos , Distribuição AleatóriaRESUMO
Nonalcoholic steatohepatitis is one of the leading causes of liver disease. Dietary factors determine the clinical presentation of steatohepatitis and can influence the progression of related diseases. Cholesterol has emerged as a critical player in the disease and hence consumption of cholesterol-enriched diets can lead to a progressive form of the disease. The aim was to investigate the impact of liver cholesterol overload on the progression of the obstructive cholestasis in mice subjected to bile duct ligation surgery. Mice were fed with a high cholesterol diet for two days and then were subjected to surgery procedure; histological, biochemical, and molecular analyses were conducted to address the effect of cholesterol in liver damage. Mice under the diet were more susceptible to damage. Results show that cholesterol fed mice exhibited increased apoptosis and oxidative stress as well as reduction in cell proliferation. Mortality following surgery was higher in HC fed mice. Liver cholesterol impairs the repair of liver during obstructive cholestasis and aggravates the disease with early fatal consequences; these effects were strongly associated with oxidative stress.
Assuntos
Colestase/etiologia , Colesterol na Dieta/toxicidade , Fígado/patologia , Estresse Oxidativo/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Ductos Biliares/cirurgia , Bilirrubina/análise , Caspase 3/metabolismo , Colestase/patologia , Colesterol/análise , Fígado Gorduroso/etiologia , Glutationa/análise , Imuno-Histoquímica , Icterícia/etiologia , Antígeno Ki-67/metabolismo , Fígado/efeitos dos fármacos , Fígado/enzimologia , Testes de Função Hepática , Camundongos , Camundongos Endogâmicos C57BL , Mortalidade Prematura , Espécies Reativas de Oxigênio/metabolismo , Triglicerídeos/análiseRESUMO
Cholesterol overload in the liver has shown toxic effects by inducing the aggravation of nonalcoholic fatty liver disease to steatohepatitis and sensitizing to damage. Although the mechanism of damage is complex, it has been demonstrated that oxidative stress plays a prominent role in the process. In addition, we have proved that hepatocyte growth factor induces an antioxidant response in hepatic cells; in the present work we aimed to figure out the protective effect of this growth factor in hepatocytes overloaded with free cholesterol. Hepatocytes from mice fed with a high-cholesterol diet were treated or not with HGF, reactive oxygen species present in cholesterol overloaded hepatocytes significantly decreased, and this effect was particularly associated with the increase in glutathione and related enzymes, such as γ-gamma glutamyl cysteine synthetase, GSH peroxidase, and GSH-S-transferase. Our data clearly indicate that HGF displays an antioxidant response by inducing the glutathione-related protection system.
Assuntos
Colesterol/toxicidade , Fator de Crescimento de Hepatócito/sangue , Hepatócitos/metabolismo , Hepatócitos/patologia , Estresse Oxidativo/efeitos dos fármacos , Animais , Antioxidantes/metabolismo , Estudos de Casos e Controles , Células Cultivadas , Dieta , Ensaio de Imunoadsorção Enzimática , Glutationa/metabolismo , Hepatócitos/efeitos dos fármacos , Humanos , Fígado/efeitos dos fármacos , Fígado/patologia , Masculino , Camundongos , Hepatopatia Gordurosa não Alcoólica/sangue , Hepatopatia Gordurosa não Alcoólica/patologia , Proteínas Proto-Oncogênicas c-met/sangue , Espécies Reativas de Oxigênio/metabolismoRESUMO
In recent years, there has been a growing interest to explore the responsiveness to injury in steatotic hepatocyte. VL-17A cells, which express ADH and Cyp2E1 overloaded with free fatty acids (1 mM of oleic and palmitic acid 2:1) showed an increased oxidative damaged after 24 h free fatty acids treatment when exposed to ethanol (100 mM) for 48 h as a second injury. An increment in reactive oxygen species, determined by DCFH-DA, protein oxidation, and apoptosis were observed although an increase in main antioxidant proteins such as superoxide dismutase 1 and glutathione peroxidase were observed, but failed in gamma-glutamylcysteine synthetase, suggesting a decreased capacity of synthesis of glutathione compared with cells treated only with free fatty acids or ethanol. The increased oxidative stress and toxicity in lipid overloaded VL-17A cells subjected to ethanol exposure were accompanied by increases in Cyp2E1 protein expression. Our data show that lipid loaded in an in vitro model, VL-17A cells, is more susceptible to cell damage and oxidative stress when treated with ethanol.
Assuntos
Etanol/toxicidade , Ácidos Graxos não Esterificados/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Álcool Desidrogenase/metabolismo , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Citocromo P-450 CYP2E1/genética , Citocromo P-450 CYP2E1/metabolismo , Fluoresceínas/metabolismo , Glutamato-Cisteína Ligase/metabolismo , Glutationa/biossíntese , Células Hep G2 , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Humanos , Espécies Reativas de Oxigênio/metabolismo , Triglicerídeos/metabolismoRESUMO
Alcohol is undoubtedly, the main toxic agent that people consume by recreation and the abuse is associated with liver damage, mainly by the overproduction of reactive oxygen species and the toxic effects of its first metabolite acetaldehyde. It is known that acetaldehyde targets mitochondria inducing redox imbalance and oxidative stress. Mitochondrial superoxide dismutase transforms superoxide radical into hydrogen peroxide, which in addition, is transformed in water by other enzymes. In the present study we demonstrate that acetaldehyde transiently impairs SOD2 activity in HepG2 cells, the decrease in the enzyme activity was associated to a reduction in the protein content, which was rapidly recovered, to basal values, by synthesis de novo in a mechanism mediated by NF-κB and PKC. The SOD2 impairment was not associated with adduct formation. The recovery on SOD2 activity in HepG2 cells can represent survival advantage for cancer cells, the results shown that SOD2 could be considered a therapeutic target in liver cancer.
Assuntos
Acetaldeído/farmacologia , Células Hep G2/efeitos dos fármacos , Superóxido Dismutase/metabolismo , Células Hep G2/metabolismo , Humanos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Terapia de Alvo Molecular , NF-kappa B/metabolismo , Proteína Quinase C/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase-1RESUMO
Redox signaling is emerging as an essential mechanism in the regulation of biological activities of the cell. The HGF/c-Met signaling pathway has been implicated as a key regulator of the cellular redox homeostasis and oxidative stress. We previously demonstrated that genetic deletion of c-Met in hepatocytes disrupts redox homeostasis by a mechanism involving NADPH oxidase. Here, we were focused to address the mechanism of NADPH oxidase regulation by HGF/c-Met signaling in primary mouse hepatocytes and its relevance. HGF induced a biphasic mechanism of NADPH oxidase regulation. The first phase employed the rapid increase in production of ROS as signaling effectors to activate the Nrf2-mediated protective response resulting in up-regulation of the antioxidant proteins, such as NAD(P)H quinone oxidoreductase and γ-glutamylcysteine synthetase. The second phase operated under a prolonged HGF exposure, caused a suppression of the NADPH oxidase components, including NOX2, NOX4, p22 and p67, and was able to abrogate the TGFß-induced ROS production and improve cell viability. In conclusion, HGF/c-Met induces a Nrf2-mediated protective response by a double mechanism driven by NADPH oxidase.