Catecolaminas plaquetárias e LDL oxidada em indivíduos com apneia obstrutiva do sono
Arquivos
Data
2014
Autores
Feres, Marcia Cristina 
Orientadores
Poyares, Dalva Lucia Rollemberg
Tipo
Tese de doutorado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
Introdução: A Apneia Obstrutiva do Sono (AOS) e um importante fator de risco para doencas cardiovasculares. Varios mecanismos fisiopatologicos tem sido sugeridos para esclarecer a associacao da AOS com a alta incidencia de morbidade e mortalidade cardiovascular. Entre as doencas cardiovasculares mais prevalentes que aparecem em associacao com a AOS, observa-se a hipertensao (HAS), que abrange 50% de prevalencia no mundo e a aterosclerose, que ocupa um relevante percentual nesse contexto. Um dos mecanismos propostos e o envolvimento do sistema nervoso autonomo, inicialmente durante o sono e gradualmente acometendo o periodo de vigilia, levando a alteracoes da pressao sanguinea. Tais alteracoes envolvem aumento da atividade do sistema nervoso simpatico deflagrado pelos eventos respiratorios associados a elevacao dos niveis de catecolaminas. Esse incremento das catecolaminas, juntamente com a hipoxemia, pode ter atuacao indireta no endotelio favorecendo o mecanismo de inflamacao e geracao de substancias reativas do oxigenio e oxidacao das lipoproteinas LDL-C produzindo a LDL oxidada, facilitando a instalacao da aterosclerose. Alem disso, as catecolaminas estao envolvidas na sindrome metabolica, podendo alterar o perfil lipidico de tais pacientes. Entretanto, as metodologias de avaliacao das catecolaminas ainda sao muito controversas devido a sua rapida flutuacao nos fluidos organicos e a interferencias de varios fatores de confusao. Objetivo: Este estudo objetivou utilizar metodologias de quantificacao de niveis de catecolaminas em plaquetas, plasma e urina de 24 horas, em pacientes com AOS com e sem hipertensao arterial, e avaliar um dos mais relevantes parametros lipidicos envolvidos nos mecanismos de aterosclerose, que e o LDL colesterol direto e LDL oxidada em pacientes com AOS associadas ou nao a hipertensao ou dislipidemias. Por fim, avaliar o impacto do tratamento da AOS com CPAP sobre os niveis de LDL oxidada nos pacientes com AOS severa apos um ano de tratamento. Metodos e Casuistica: Foram desenvolvidos 3 estudos (1, 2 e 3). O Estudo 1 foi dividido em dois experimentos. No Experimento 1 para realizar a padronizacao e validacao do metodo de catecolaminas plaquetarias, foram convidados 30 voluntarios que nao estavam fazendo nenhum tratamento de Saúde ou tomando medicamento e aceitaram em participar do estudo. Obteve-se 10 mL de sangue de cada voluntario para validar a metodologia de catecolaminas plaquetarias com comparacao do metodo de radio imunoensaio (RIE) e cromatografia liquida de alta pressao (HPLC). No Experimento 2, apos a validacao, foram selecionados 154 pacientes do Instituto do Sono de São Paulo, que passaram por orientacao sobre dieta hipocalorica com restricao a xantinas, cafeinas, exercicios fisicos, fumo e alcool previamente a coleta de fluidos. Tais pacientes foram alocados em quatro grupos: Hipertensao com AOS, Hipertensao sem AOS, somente AOS sem hipertensao e Grupo controle. Tambem houve uma analise de um subgrupo randomizado para tratamento com CPAP e Sham-CPAP. Os individuos passaram por avaliacoes clinicas, antropometricas, testes cardiopulmonares, polissonografia (PSG) e exames laboratoriais (perfil lipidico, testes de funcao renal e hepatica, glicemia, hemograma, velocidade de hemossedimentacao). Nos empregamos metodos de RIE para determinacao dos niveis de catecolaminas plasmaticas e urinarias alem do metodo de RIE ultrassensivel para os niveis plaquetarios. No Estudo 2 foram selecionados 99 pacientes do mesmo ambulatorio e foram alocados em 3 grupos GI- AOS com comorbidades (HAS e DIS) , GII u AOS e GIII controle. As dosagens do perfil lipidico e de outros parametros laboratoriais foram realizadas por metodos automatizados em equipamentos validados, e a LDL-colesterol direta determinada por reagente com eliminacao da catalase e para LDL oxidada, metodo de enzimaimuno ensaio (EIA). E para o Estudo 3 foram randomizados 72 pacientes com AOS severa e tratados com CPAP e Sham-CPAP por um ano de tratamento. No final do estudo apenas 45 pacientes chegaram a um ano de tratamento. Parametros antropometricos, perfil lipidico, e LDL oxidada foram avaliados nos periodos basais, 1, 3, 6 e 12 meses pos-tratamento. Calculos Estatisticos: Para o Estudo 1 foram aplicados teste de correlacao de Pearson, curva ROC com Odds-ratio e ANCOVA de uma vias foram empregados para tratamento estatistico dos dados. Como analise complementar foi realizados regressao logistica e indices de coeficiente de correlacao inter e intra-ensaiou dos testes plaquetarios. O teste de validacao que antecedeu o experimento seguiu protocolo de validacao com base nos metodos de J. Westgard. No Estudo 2, foi calculado a regressao linear construido baseados nas variaveis independentes AOS, HAS e DIS. E no Estudo 3 calculou-se a ANOVA de medidas repetidas. Resultados: No Estudo 1, e no Experimento 1, a validacao foi satisfatoria com correlacoes (r= 0,93 e r= 0,92) entre os metodos (RIE e HPLC) para catecolaminas plaquetarias, adrenalina e noradrenalina respectivamente. Desse modo, escolhemos o primeiro pela facilidade de execucao e rapidez. No Experimento 2, os niveis de noradrenalina e adrenalina urinarias foram significativamente associadas a condicao de hipertensao com e sem AOS (p=0,00 e p=0,01) enquanto noradrenalina plaquetaria relacionou-se somente a condicao de AOS sem hipertensao (p=0,03). Os resultados do efeito do tratamento com CPAP sobre as catecolaminas plaquetarias observam-se que o subgrupo tratado apos um ano apresentou reducao significativa de noradrenalina plaquetaria e de excrecao de catecolaminas urinaria (p=0,05 e p=0,04). No Estudo 2, o modelo estatistico de regressao logistica mostrou que somente a presenca de DIS foi associada aos niveis de LDL-colesterol direto (p=0, 001). A presenca de comorbidades (AOS e HAS) nao se associou as taxas de LDL-C direta, entretanto os niveis de LDL oxidada apresentaram expressivos efeitos aditivos para AOS (p=0,01, HAS (p=0,03) e DIS (p=0,03). No Estudo 3, o tratamento com CPAP e Sham-CPAP com relacao a circunferencia abdominal apresentou diferenca estatisticamente significante entre o periodo basal e 1 mes apos CPAP no grupo de CPAP (p=0,02). Com relacao a LDL-C, o grupo CPAP apresentou diferencas estatisticamente significante entre os periodos basais, 1, 3 e 12 meses, (p=0,04, p=0,04 e p=0,03) respectivamente. Com relacao as LDL oxidadas nao apresentaram diferencas estatisticamente significantes entre os periodos basais, 6 meses e 12 meses, nos grupos CPAP e Sham-CPAP (p=0, 011 e p=0,09). Conclusoes: O Estudo 1 demonstrou que o metodo de catecolaminas plaquetarias foi sensivel para monitorar a AOS, indicando alteracao da atividade simpatica mesmo em pacientes sem hipertensao. O tratamento com o CPAP foi capaz de reduzir os niveis de catecolaminas plaquetarias e urinaria. No Estudo 2, a AOS pode ocasionar elevacao das LDL oxidadas, independentemente das comorbidades associadas a AOS (HAS e DIS). No entanto, o tratamento efetivo da AOS (Estudo 3) nao foi capaz de alterar os niveis de LDL-oxidada
Introduction: Obstructive sleep apnea (OSA) is a major risk factor for cardiovascular disease. Several pathophysiological mechanisms have been suggested to explain the association of OSA with high incidence of cardiovascular morbidity and mortality. Among the most prevalent cardiovascular diseases that appear in association with OSA, there was hypertension (HYP), which covers 50% prevalence in the world and atherosclerosis, which occupies a percentage relevance in this context. One of the proposed mechanisms is the involvement of the autonomic nervous system, initially during sleep and gradually affecting wakefulness, leading to changes in blood pressure. Such changes involve increased sympathetic nervous system triggered by respiratory events associated with elevated levels of catecholamine activity. This increase of catecholamines, together with hypoxemia may have an indirect role in the mechanism of promoting endothelial inflammation and generation of oxygen reactive substances and oxidation of lipoproteins LDL-C producing oxidized LDL, to facilitate installation of atherosclerosis. In addition, catecholamines are involved in the metabolic syndrome, may alter the lipid profile of these patients. However, the evaluation methodologies of catecholamines are still very controversial because of its rapid fluctuation in body fluids and the interference of confounders. Objective: This study aims to use methods of quantification of catecholamines in platelets, plasma and urine for 24 hours in OSA patients with and without hypertension, and evaluate one of the most important mechanisms involved in lipid parameters of atherosclerosis, which is direct LDL cholesterol and oxidized LDL in patients with OSA or not associated with hypertension or dyslipidemia. Finally, evaluate the impact of treatment of OSA with CPAP on levels of oxidized LDL in patients with severe OSA after one year of treatment.This study aimed to use methods of quantification of catecholamines in platelets, plasma and urine in 24 hours in OSA patients with and without hypertension and lipid important to evaluate a parameter that is involved in atherosclerosis direct LDL cholesterol and oxidized LDL OSA patients associated or not with Dyslipidemia or Hypertension as well as assess the impact of CPAP treatment on oxidized LDL in patients with severe OSA after one year of treatment. methods and Patients: 3 studies (1, 2 and 3) were developed. Study 1 was divided into two experiments. In Experiment 1 to make the standardization and validation of the method of platelet catecholamines, 30 volunteers who were not making any health treatment or taking medication and agreed to participate were asked. There was obtained 10 ml of blood from each volunteer to validate the methodology platelet catecholamines compared with the method of radioimmunoassay (RIA) and high pressure liquid chromatography (HPLC). In Experiment 2, after validation, we selected 154 patients of the Sleep Institute of São Paulo, which passed through guidance on calorie-restricted diet to xanthine, cafeínas, exercise, smoking and alcohol prior to collection of fluids. These patients were divided into four groups: hypertension with OSA, hypertension without OSA, only OSA and control group without hypertension. Also there was an analysis of a sub-group randomized to treatment with CPAP and CPAP-Sham. The subjects underwent clinical assessments, anthropometric, cardiopulmonary exercise testing, polysomnography (PSG) and laboratory tests (lipid profile, renal and liver function tests, blood glucose, blood count, erythrocyte sedimentation rate). We employ RIE methods for determination of plasma levels of urinary catecholamines and besides RIE ultrasensitive method for platelet levels. In Study 2 were selected 99 patients from the same clinic and were divided into 3 groups GI-AOS with comorbidities (hypertension and DIS), GII - GIII OSA and control. The measurements of the lipid profile and other laboratory parameres were performed by automated equipment in validated methods, and direct LDL-cholesterol determined by reaction with elimination of catalase and oxidized LDL, enzimaimuno method (EIA). And for the Study 3 were randomized 72 patients with severe OSA and treated with CPAP and Sham - CPAP for a year of treatment. At the end of the study, only 45 patients reached one year of treatment. Anthropometric parameters, blood lipids, and oxidized LDL were assessed at baseline periods, 1, 3, 6 and 12 months post-treatment. Statistical Calculations: For Study 1 were applied Pearson correlation test, ROC curve with Odds-ratio and one-way ANCOVA were used for statistical treatment of data. As a complementary analysis, we performed logistic regression coefficients and indices of correlation and inter intraensaios of platelet testing. The validation test prior to the experiment was the validation protocol based on methods J. Westgard. In Study 2, a linear regression built based on Indepents main measurements to HYP, DYS and OSA was calculated. And in Study 3 was calculated by ANOVA for repeated measures. Results: In Study 1, and in Experiment 1, the validation was satisfactory with correlations (r=0.93 and r=0.92) between methods (RIA and HPLC) for platelet catecholamines, epinephrine and norepinephrine respectively. Thus, we chose the first for ease of implementation and speed. In Experiment 2, the levels of urinary norepinephrine and epinephrine were significantly associated with the condition of hypertension with and without OSA (p=0.00 and p=0.01) while noradrenaline platelet was related only to the condition of OSA without hypertension (p=0.03). The results of the effect of CPAP treatment on platelet catecholamines, it is observed that the group treated after one year has decreased platelet noradrenaline and urinary catecholamine excretion (p=0.05 and p=0.04). In Study 2, the statistical logistic regression model showed that only the presence of DIS was associated with levels of direct LDL-cholesterol (p=0.001). Comorbidities (OSA and hypertension) was not associated with rates of LDL-C directly, however the levels of oxidized LDL showed significant additive effects for OSA (p=0.01, SAH (p=0.03) and DIS (p=0.03). In Study 3, treatment with CPAP and CPAP-Sham regarding abadominal circumference showed a statistically significant difference between baseline and 1 month after CPAP in CPAP group (p=0.02). With respect to LDL-C, the CPAP group showed statistically significant differences entrte periodsos baseline, 1, 3 and 12 months (p=0.04, p=0.04 and p=0.03) respectively. Regarding the oxidized LDL showed no statistically significant differences betweenthe baseline periods, 6 months and 12 months in groups Sham-CPAP and CPAP (p=0.011 and p=0.09). Conclusions: Study 1 demonstrated that the method of platelet catecholamines was sensitive to monitor the OSA, indicating alteration of sympathetic activity in patients without hypertension. The CPAP treatment was able to reduce the levels of urinary catecholamines, and platelet. In Study 2, OSA can cause elevation of oxidized LDL, regardless of comorbidities associated with OSA (SAH and DIS). However, effective treatment of OSA (Study 3) was unable to alter the levels of oxidized LDL.
Introduction: Obstructive sleep apnea (OSA) is a major risk factor for cardiovascular disease. Several pathophysiological mechanisms have been suggested to explain the association of OSA with high incidence of cardiovascular morbidity and mortality. Among the most prevalent cardiovascular diseases that appear in association with OSA, there was hypertension (HYP), which covers 50% prevalence in the world and atherosclerosis, which occupies a percentage relevance in this context. One of the proposed mechanisms is the involvement of the autonomic nervous system, initially during sleep and gradually affecting wakefulness, leading to changes in blood pressure. Such changes involve increased sympathetic nervous system triggered by respiratory events associated with elevated levels of catecholamine activity. This increase of catecholamines, together with hypoxemia may have an indirect role in the mechanism of promoting endothelial inflammation and generation of oxygen reactive substances and oxidation of lipoproteins LDL-C producing oxidized LDL, to facilitate installation of atherosclerosis. In addition, catecholamines are involved in the metabolic syndrome, may alter the lipid profile of these patients. However, the evaluation methodologies of catecholamines are still very controversial because of its rapid fluctuation in body fluids and the interference of confounders. Objective: This study aims to use methods of quantification of catecholamines in platelets, plasma and urine for 24 hours in OSA patients with and without hypertension, and evaluate one of the most important mechanisms involved in lipid parameters of atherosclerosis, which is direct LDL cholesterol and oxidized LDL in patients with OSA or not associated with hypertension or dyslipidemia. Finally, evaluate the impact of treatment of OSA with CPAP on levels of oxidized LDL in patients with severe OSA after one year of treatment.This study aimed to use methods of quantification of catecholamines in platelets, plasma and urine in 24 hours in OSA patients with and without hypertension and lipid important to evaluate a parameter that is involved in atherosclerosis direct LDL cholesterol and oxidized LDL OSA patients associated or not with Dyslipidemia or Hypertension as well as assess the impact of CPAP treatment on oxidized LDL in patients with severe OSA after one year of treatment. methods and Patients: 3 studies (1, 2 and 3) were developed. Study 1 was divided into two experiments. In Experiment 1 to make the standardization and validation of the method of platelet catecholamines, 30 volunteers who were not making any health treatment or taking medication and agreed to participate were asked. There was obtained 10 ml of blood from each volunteer to validate the methodology platelet catecholamines compared with the method of radioimmunoassay (RIA) and high pressure liquid chromatography (HPLC). In Experiment 2, after validation, we selected 154 patients of the Sleep Institute of São Paulo, which passed through guidance on calorie-restricted diet to xanthine, cafeínas, exercise, smoking and alcohol prior to collection of fluids. These patients were divided into four groups: hypertension with OSA, hypertension without OSA, only OSA and control group without hypertension. Also there was an analysis of a sub-group randomized to treatment with CPAP and CPAP-Sham. The subjects underwent clinical assessments, anthropometric, cardiopulmonary exercise testing, polysomnography (PSG) and laboratory tests (lipid profile, renal and liver function tests, blood glucose, blood count, erythrocyte sedimentation rate). We employ RIE methods for determination of plasma levels of urinary catecholamines and besides RIE ultrasensitive method for platelet levels. In Study 2 were selected 99 patients from the same clinic and were divided into 3 groups GI-AOS with comorbidities (hypertension and DIS), GII - GIII OSA and control. The measurements of the lipid profile and other laboratory parameres were performed by automated equipment in validated methods, and direct LDL-cholesterol determined by reaction with elimination of catalase and oxidized LDL, enzimaimuno method (EIA). And for the Study 3 were randomized 72 patients with severe OSA and treated with CPAP and Sham - CPAP for a year of treatment. At the end of the study, only 45 patients reached one year of treatment. Anthropometric parameters, blood lipids, and oxidized LDL were assessed at baseline periods, 1, 3, 6 and 12 months post-treatment. Statistical Calculations: For Study 1 were applied Pearson correlation test, ROC curve with Odds-ratio and one-way ANCOVA were used for statistical treatment of data. As a complementary analysis, we performed logistic regression coefficients and indices of correlation and inter intraensaios of platelet testing. The validation test prior to the experiment was the validation protocol based on methods J. Westgard. In Study 2, a linear regression built based on Indepents main measurements to HYP, DYS and OSA was calculated. And in Study 3 was calculated by ANOVA for repeated measures. Results: In Study 1, and in Experiment 1, the validation was satisfactory with correlations (r=0.93 and r=0.92) between methods (RIA and HPLC) for platelet catecholamines, epinephrine and norepinephrine respectively. Thus, we chose the first for ease of implementation and speed. In Experiment 2, the levels of urinary norepinephrine and epinephrine were significantly associated with the condition of hypertension with and without OSA (p=0.00 and p=0.01) while noradrenaline platelet was related only to the condition of OSA without hypertension (p=0.03). The results of the effect of CPAP treatment on platelet catecholamines, it is observed that the group treated after one year has decreased platelet noradrenaline and urinary catecholamine excretion (p=0.05 and p=0.04). In Study 2, the statistical logistic regression model showed that only the presence of DIS was associated with levels of direct LDL-cholesterol (p=0.001). Comorbidities (OSA and hypertension) was not associated with rates of LDL-C directly, however the levels of oxidized LDL showed significant additive effects for OSA (p=0.01, SAH (p=0.03) and DIS (p=0.03). In Study 3, treatment with CPAP and CPAP-Sham regarding abadominal circumference showed a statistically significant difference between baseline and 1 month after CPAP in CPAP group (p=0.02). With respect to LDL-C, the CPAP group showed statistically significant differences entrte periodsos baseline, 1, 3 and 12 months (p=0.04, p=0.04 and p=0.03) respectively. Regarding the oxidized LDL showed no statistically significant differences betweenthe baseline periods, 6 months and 12 months in groups Sham-CPAP and CPAP (p=0.011 and p=0.09). Conclusions: Study 1 demonstrated that the method of platelet catecholamines was sensitive to monitor the OSA, indicating alteration of sympathetic activity in patients without hypertension. The CPAP treatment was able to reduce the levels of urinary catecholamines, and platelet. In Study 2, OSA can cause elevation of oxidized LDL, regardless of comorbidities associated with OSA (SAH and DIS). However, effective treatment of OSA (Study 3) was unable to alter the levels of oxidized LDL.
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FERES, Marcia Cristina. Catecolaminas palquetárias e LDL oxidada em indivíduos com apneia obstrutiva do sono. 2014. 183 f. Tese (Doutorado em Ciências) – Escola Paulista de Medicina, Universidade Federal de São Paulo. São Paulo, 2014.