RESUMO
Low-density lipoprotein (LDL) is considered to be a risk factor for atherosclerosis. In the presence of hyperglycemia, LDL undergoes glycoxidative modification and this glycoxidized (glycox) LDL promotes atherosclerosis in type 2 diabetic (T2D) individuals. Moreover, because of its cholesterol content, LDL contributes to aldosterone biosynthesis, which is modulated by angiotensin II (AngII) and has been implicated in cardiovascular complications of T2D. However, the molecular mechanism of the crosstalk between glycoxLDL, AngII, and aldosterone has not been explained clearly. Therefore, this study has been aimed to investigate the impact of in vitro modified glycoxLDL on aldosterone release in an AngII-sensitized adrenocortical carcinoma cell line (NCI H295R). Native LDL (natLDL), isolated from healthy volunteers by sequential density gradient ultracentrifugation, was subjected to d-glucose (200 mmol/l), for glycoxidative modification, at 37 °C for 6 days. The AngII-sensitized H295R cells were treated with natLDL and glycoxLDL for 24 h and the supernatant was used for aldosterone measurement. The treated cells were utilized for protein isolation and mRNA quantification. Compared to natLDL, glycoxLDL produced a significantly greater effect on aldosterone release from AngII-sensitized cells. The treatment with specific pharmacological inhibitors suggests that modified LDL recruits ERK1/2 and janus kinase-2 for transcriptional regulation of aldosterone synthase. Moreover, glycoxLDL modulates aldosterone release via cAMP-dependent protein kinase A (PKA) pathway. However, glycoxLDL induces ERK phosphorylation independent of PKA activation and this novel mechanism could be targeted for therapeutic trials. In conclusion, this in vitro study emphasizes a possible causal relationship between LDL glycoxidative modification, AngII-sensitization, and adrenocortical steroid hormone release.
Assuntos
Córtex Suprarrenal/metabolismo , Aldosterona/metabolismo , Angiotensina II/farmacologia , Lipoproteínas LDL/metabolismo , Córtex Suprarrenal/efeitos dos fármacos , Linhagem Celular , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Citocromo P-450 CYP11B2/genética , Ativação Enzimática/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Glicosilação/efeitos dos fármacos , Humanos , Janus Quinase 2/metabolismo , Oxirredução/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Esteroides/biossíntese , Transcrição Gênica/efeitos dos fármacosRESUMO
We prospectively evaluated whether an effective 12-month uric acid-lowering therapy (ULT) with the available xanthine oxidase (XO) inhibitors allopurinol and febuxostat in patients with chronic tophaceous gout has an impact on oxidative stress and/or vascular function. Patients with chronic tophaceous gout who did not receive active ULT were included. After clinical evaluation, serum uric acid levels (SUA) and markers of oxidative stress were measured, and carotid-femoral pulse wave velocity (cfPWV) was assessed. Patients were then treated with allopurinol (n = 9) or with febuxostat (n = 8) to target a SUA level ≤ 360 µmol/L. After 1 year treatment, the SUA levels, markers of oxidative stress and the cfPWV were measured again. Baseline characteristics of both groups showed no significant differences except a higher prevalence of moderate impairment of renal function (estimated glomerular filtration rate <60 ml/min) in the febuxostat group. Uric acid lowering with either inhibitors of XO resulted in almost equally effective reduction in SUA levels. The both treatment groups did not differ in their baseline cfPWV (allopurinol group: 14.1 ± 3.4 m/s, febuxostat group: 13.7 ± 2.7 m/s, p = 0.80). However, after 1 year of therapy, we observed a significant cfPWV increase in the allopurinol group (16.8 ± 4.3 m/s, p = 0.001 as compared to baseline), but not in the febuxostat patients (13.3 ± 2.3 m/s, p = 0.55). Both febuxostat and allopurinol effectively lower SUA levels in patients with severe gout. However, we observed that febuxostat also appeared to be beneficial in preventing further arterial stiffening. Since cardiovascular events are an important issue in treating patients with gout, this unexpected finding may have important implications and should be further investigated in randomized controlled trials.
Assuntos
Alopurinol/uso terapêutico , Supressores da Gota/uso terapêutico , Gota/tratamento farmacológico , Estresse Oxidativo/efeitos dos fármacos , Análise de Onda de Pulso , Tiazóis/uso terapêutico , Ácido Úrico/sangue , Rigidez Vascular/efeitos dos fármacos , Idoso , Alopurinol/efeitos adversos , Biomarcadores/sangue , Doença Crônica , Inibidores Enzimáticos/efeitos adversos , Inibidores Enzimáticos/uso terapêutico , Febuxostat , Alemanha , Taxa de Filtração Glomerular/efeitos dos fármacos , Gota/sangue , Gota/diagnóstico , Gota/fisiopatologia , Supressores da Gota/efeitos adversos , Humanos , Mediadores da Inflamação/sangue , Rim/efeitos dos fármacos , Rim/fisiopatologia , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Índice de Gravidade de Doença , Tiazóis/efeitos adversos , Fatores de Tempo , Resultado do Tratamento , Xantina Oxidase/antagonistas & inibidores , Xantina Oxidase/metabolismoRESUMO
An increased prevalence of diabetes mellitus (DM) has been reported in patients with primary aldosteronism (PA). DM is associated with abnormal structure and metabolism of circulating lipoproteins, which normally serve as a major source of cholesterol for adrenocortical steroidogenesis. The present study has been designed to investigate the effect of diabetically modified lipoproteins on adrenocortical aldosterone synthesis. Lipoproteins (VLDL, LDL, HDL) isolated from healthy volunteers, were subjected to oxidation or glycoxidation in the presence of sodium hypochlorite (3 mmol/l) or glucose (200 mmol/l), and aldosterone synthesis in human adrenocortical cells (H295R) was examined. Native and glycoxidized VLDL had greatest stimulatory effect on aldosterone production by 15-fold and 14-fold, respectively. At the molecular level, these VLDL produced maximum increases in Cyp11B2 mRNA level up to 17-fold. Experiments with the highly selective scavenger receptor class B type I (SR-BI) inhibitor BLT-1 revealed that cholesterol uptake from native and glycoxidized HDL and VLDL for hormone production is considerably mediated by SR-BI. Western blot analysis of extracellular signal-regulated kinase (ERK 1/2) phosphorylation and experiments with the MEK inhibitor U0126 indicated a specific mechanistic role of the ERK cascade in lipoprotein-mediated steroid hormone release. In summary, diabetic dyslipidemia and modification of circulating lipoproteins may promote adrenocortical aldosterone synthesis.
Assuntos
Córtex Suprarrenal/metabolismo , Aldosterona/biossíntese , Complicações do Diabetes/metabolismo , Hiperaldosteronismo/metabolismo , Lipoproteínas/metabolismo , Linhagem Celular Tumoral , Citocromo P-450 CYP11B2/genética , Citocromo P-450 CYP11B2/metabolismo , Complicações do Diabetes/enzimologia , Complicações do Diabetes/genética , Humanos , Hiperaldosteronismo/complicações , Hiperaldosteronismo/enzimologia , Hiperaldosteronismo/genética , OxirreduçãoRESUMO
In highly industrialized countries hyperuricemia is one of the most common metabolic disorders. High uric acid blood levels may lead to the manifestation of gout owing to the precipitation of urate crystals in connective tissue, the skeletal system and kidneys. A primary reduction of renal uric acid excretion can be detected in more than 90% of all cases of hyperuricemia. Despite the identification of several uric acid transporting proteins their pathogenetic role for the induction of primary reduced renal uric acid excretion has not yet been verified. As a result of a case-control study on individuals with normal and reduced renal uric acid excretion, an association of polymorphisms in the human urate transporter 1 gene (hURAT1) with primary reduced urate excretion has been demonstrated for the first time. The hURAT1 gene is an organic anion transporter (SLC22A12), which is preferentially expressed in the apical membrane of proximal renal tubule cells. Functioning as an antiporter, hURAT1 mediates the uptake of urate from the lumen into proximal tubule cells in exchange for organic and inorganic anions. Loss-of-function mutations in the hURAT1 gene are a cause of hereditary renal hypouricemia. The precisely regulated hURAT1 is a candidate gene for hyperuricemia and an important target for the development and optimization of new diagnostic approaches and pharmacological interventions of primary reduced renal uric acid excretion.
Assuntos
Artrite Gotosa/genética , Hiperuricemia/genética , Transportadores de Ânions Orgânicos/genética , Proteínas de Transporte de Cátions Orgânicos/genética , Ácido Úrico/urina , Artrite Gotosa/diagnóstico , Artrite Gotosa/urina , Marcadores Genéticos/genética , Variação Genética , Humanos , Hiperuricemia/diagnóstico , Hiperuricemia/urina , Túbulos Renais Proximais/metabolismo , Polimorfismo Genético/genéticaRESUMO
AIMS/HYPOTHESIS: Glycoxidised LDL has been implicated in the pathogenesis of atherosclerosis, a major complication of diabetes. Since atherogenesis may occur at an early stage of diabetes, we investigated whether circulating LDL isolated from subjects with IGT (n = 20) showed an increased glycoxidation status and explored the proatherogenic effects of LDL samples on macrophages. SUBJECTS AND METHODS: We investigated LDL modifications using GC-MS. Murine macrophages were incubated with LDL samples for 1 h, and then mRNA expression rates of the scavenger receptors CD36 and scavenger receptor class B type 1 (SCARB1, formerly known as SR-BI) and transcription factor peroxisome proliferator-activator receptor gamma (PPARgamma) were quantified by real-time RT-PCR. RESULTS: The GC-MS experiments revealed that oxidative modifications of proline, arginine, lysine and tyrosine residues in apolipoprotein B100 were three- to fivefold higher in LDL samples from IGT subjects compared with those from NGT subjects (n = 20). Moreover, LDL glycoxidation estimated by both Nepsilon-(carboxymethyl)lysine (CML) and Nepsilon-(carboxyethyl)lysine (CEL) residues was increased more than ninefold in LDL from IGT subjects compared with samples from NGT subjects. Compared with NGT LDL, IGT LDL elicited a significantly higher CD36 (p < 0.05) and PPARG (p < 0.05) gene expression, whereas SCARB1 mRNA expression was not affected. CONCLUSIONS/INTERPRETATION: These data suggest that IGT is associated with increased glycoxidation of circulating LDL, which might contribute to the conversion of macrophages into a proatherogenic phenotype.
Assuntos
Antígenos CD36/genética , Regulação da Expressão Gênica , Intolerância à Glucose/sangue , Lipoproteínas LDL/sangue , Macrófagos/fisiologia , PPAR gama/genética , Adulto , Antígenos CD/genética , Glicemia/metabolismo , Feminino , Humanos , Lipoproteínas LDL/isolamento & purificação , Masculino , Pessoa de Meia-Idade , Valores de ReferênciaRESUMO
Alterations in both vasculature and renin-angiotensin-aldosterone system are a consistent finding in the metabolic syndrome. Adrenal tissue is highly vascularized and encounters blood flow, exceeding by far the volume expected for its size. Endothelial cells in the adrenal vasculature are therefore a major cellular component of adrenocortical tissue. The aim of the study was to analyze the cellular interaction between endothelial and steroid producing cells, focusing on endothelial cell-factor-mediated activation of aldosterone synthesis. The interaction between human endothelial (HUVECs) cell-conditioned medium and human adrenocortical (NCI-H295R) cells IN VITRO induced a significant surge in aldosterone secretion. The endothelial cell-conditioned medium together with angiotensin II and forskolin also potentiated aldosterone release by 1.5-fold and 2.6-fold, respectively, while preincubation of NCI-H295R cells for 24 h with endothelial cell-conditioned medium enhanced and sensitized the response of NCI-H295R to subsequent angiotensin II and forskolin stimuli by 2.5-fold and 2.2-fold, respectively. The increase in aldosterone release after preincubation with endothelial cell-conditioned medium was sensitive to cycloheximide and KN-93. Cellular conditioning with endothelial-cell factors exerts a hitherto unknown paracrine regulation of aldosterone production in human adrenocortical cells. This interaction may contribute to altered basal aldosterone release and have a role in patients with hypertension.
Assuntos
Córtex Suprarrenal/citologia , Córtex Suprarrenal/efeitos dos fármacos , Angiotensina II/farmacologia , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Aldosterona/biossíntese , Aldosterona/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Células Cultivadas , Colforsina/farmacologia , Meios de Cultivo Condicionados/farmacologia , Humanos , Biossíntese de Proteínas/efeitos dos fármacosRESUMO
Hypercholesterolemia and oxidative stress are major risk factors in atherogenesis. In the last years, lipid apheresis has been established as an effective clinical therapy by lowering not only elevated plasma low-density lipoprotein (LDL) levels but also by reducing the incidence of cardiovascular events. The aim of the present study was to investigate peripheral leukocyte oxidant generation in patients with familial hypercholesterolemia (FH) undergoing regular LDL apheresis. The activity state of leukocytes was estimated prior to, immediately after, and 2 days after LDL apheresis carried out by two distinct techniques: hemoperfusion with the DALI system and heparin-induced extracorporeal LDL precipitation (HELP). Oxidant generating activity was measured by chemiluminescence (CL) in whole blood and isolated polymorphonuclear leukocytes (PMNL). The results of our study show increased baseline respiratory burst activities in FH patients as compared to healthy controls. Apheresis with the HELP system was followed by increases in leukocyte count, zymosan-induced whole blood CL, and plasma PMNL elastase levels. The DALI technique caused no changes in leukocyte count and elastase levels and decreased whole blood CL activity. Two days after lipid removal the observed changes returned to pre-apheresis levels. Leukocyte activity parameters before and after apheresis did not correlate with the corresponding plasma levels of triglycerides, total cholesterol, and LDL cholesterol, suggesting that different handling in the framework of both apheresis techniques rather than lipid profile changes during therapy accounted for leukocyte activity modulation.
Assuntos
Remoção de Componentes Sanguíneos , Hemoperfusão , Heparina/farmacologia , Hiperlipoproteinemia Tipo II/terapia , Leucócitos/metabolismo , Lipoproteínas LDL/isolamento & purificação , Estresse Oxidativo , Adulto , Idoso , Precipitação Química , Feminino , Humanos , Hiperlipoproteinemia Tipo II/metabolismo , Lipoproteínas LDL/sangue , Masculino , Pessoa de Meia-Idade , Explosão RespiratóriaRESUMO
The pretreatment of rats with the substance LPH and LXU caused decrease of lethality in endotoxin shock and suppression of the following changes 6 hours post endotoxin: Increase of plasma ICDH activity, decrease in platelet count, decrease of the hepatic energy state and increase of the ATP content in the lung. The catabolic glucose metabolism during endotoxin shock was not influenced by LPH and LXU administration.
Assuntos
Metabolismo Energético/efeitos dos fármacos , Isocitrato Desidrogenase/metabolismo , Fígado/metabolismo , Pulmão/metabolismo , Choque Séptico/metabolismo , Nucleotídeos de Adenina/metabolismo , Animais , Glicemia/metabolismo , Endotoxinas/farmacologia , Escherichia coli , Fígado/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Masculino , Contagem de Plaquetas , Ratos , Ratos Endogâmicos , Choque Séptico/sangueRESUMO
Metabolites of the energy and carbohydrate metabolism and mitochondrial function in the liver were compared in rats with reversible as well as with irreversible shock. 6 h after induction of shock there was a close correlation between the severity of shock and the energy state of the liver. Only rats with irreversible shock showed a marked deterioration in parameters of the adenylate system, whereas in animals with reversible shock the energy state remained at control levels. Liver glycogen and glucose stores declined similarly in all shocked rats. The capacity of isolated liver mitochondria to produce ATP did not essentially differ in reversible and irreversible shock. Further investigations should consider the intracellular environment in evaluating the mitochondrial function in vivo during endotoxin shock.
Assuntos
Metabolismo Energético/efeitos dos fármacos , Glicólise/efeitos dos fármacos , Mitocôndrias Hepáticas/metabolismo , Choque Séptico/metabolismo , Difosfato de Adenosina/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Biotransformação , Glicemia/metabolismo , Metabolismo dos Carboidratos , Endotoxinas/toxicidade , Isocitrato Desidrogenase/metabolismo , Lactatos/sangue , Masculino , Mitocôndrias Hepáticas/efeitos dos fármacos , Consumo de Oxigênio/efeitos dos fármacos , Contagem de Plaquetas , Ratos , Ratos Endogâmicos , Choque Séptico/sangueRESUMO
Biochemical and electron optical changes in the liver cell were recorded at the experimental brain death in dogs. The investigation results refer to a hypoxic lesion at the beginning of the brain death despite of a strong flow in the hepatic artery and the portal vein. An enlarged shunt flow is discussed to be the cause.--A distinct recovery tendency shows the morphological picture at the state of the brain death despite reducing the hepatic blood-flow and without aimed therapeutic measures. This recovery tendency appears better in the electron microscopical finding than in the behaviour of the cell metabolites.