Resumo Fundamento Tem sido demonstrado que um aumento dos níveis séricos de PON1 é protetor contra vários distúrbios. Foi relatado que vários polimorfismos de nucleotídeo único (SNPs, single nucleotide polymorphisms ) do gene PON1 estão associados a níveis e atividade de proteínas enzimáticas séricas. Objetivos Investigar a associação de SNPs do PON1 e atividade da paraoxonase sérica com a doença arterial coronariana (DAC). Métodos Foram estudados 601 pacientes não relacionados submetidos à angiografia coronária, incluindo aqueles com estenose >50% (N=266) e aqueles com estenose <30% (N=335). Os SNPs rs662 e rs840560 do gene da paraoxonase foram determinados utilizando o método ARMS-PCR e o SNP rs705379 foi genotipado utilizando análise de PCR-RFLP. A atividade da paraoxonase sérica foi medida utilizando paraoxon como substrato. O valor de p<0,05 foi considerado significante. Resultados A atividade da paraoxonase sérica não foi significativamente diferente entre os grupos de estudo. Após ajuste para idade, sexo, hipertensão, diabetes mellitus e dislipidemia, o genótipo GG e o modelo codominante de rs662 foram positivamente associados a uma angiografia positiva (respectivamente, OR = 2,424, IC 95% [1,123-5,233], p <0,05, OR = 1,663, IC 95% [1,086-2,547]). A atividade da paraoxonase sérica foi significativamente maior no alelo G e variante GG do polimorfismo rs662, alelo A e variante AA de rs854560 e alelo C e variante CC de rs705379. A análise de haplótipos mostrou que o haplótipo ATC foi significativamente mais prevalente no grupo com angiografia negativa. A análise entre os grupos indicou que o alelo A de rs662 foi significativamente associado à menor atividade da paraoxonase no grupo com angiografia positiva (p=0,019). Conclusões A presença do alelo G do polimorfismo de nucleotídeo único rs662 está independentemente associada ao aumento do risco de DAC.

Abstract Background It has been shown that increased serum PON1 levels are protective against several disorders. Several single nucleotide polymorphisms (SNPs) of the PON1 gene have been reported to be associated with serum enzyme protein levels and activity. Objective To investigate the association of SNPs of PON1 and serum paraoxonase activity with coronary artery disease (CAD). Methods A total of 601 unrelated patients who underwent coronary angiography including those who had >50% stenosis (N=266) and those with <30% stenosis (N=335) were studied. The Paraoxonase gene rs662 and rs840560 SNPs were determined using the ARMS-PCR method and the rs705379 SNP was genotyped using PCR-RFLP analysis. Serum paraoxonase activity was measured using paraoxon as a substrate. A p value of p<0.05 was considered as significant. Results Serum paraoxonase activity was not significantly different between the study groups. After adjustment for age, sex, hypertension, diabetes mellitus and dyslipidemia, the GG genotype and co-dominant model of rs662 was positively associated with a positive angiogram (respectively, OR=2.424, 95%CI [1.123-5.233], p<0.05, OR=1.663, 95%CI [1.086-2.547]). Serum paraoxonase activity was significantly higher in the G allele and GG variant of rs662, A allele and AA variant of rs854560 and C allele and CC variant of rs705379. The haplotype analysis has shown that the ATC haplotype was significantly more prevalent among the angiogram negative group. The analysis between groups indicated that the A allele of rs662 was significantly associated with lower paraoxonase activity in the positive angiogram group (p=0.019). Conclusions The presence of the G allele of the rs662 single nucleotide polymorphism is independently associated to increased risk of CAD.

Resumo Objetivo O objetivo deste estudo foi examinar a relação entre os polimorfismos PON1 e perda recorrente de gravidez PRG. Métodos Em um estudo transversal, foramcoletadas amostras de sangue de 100 mulheres. O DNA foi extraído e os genótipos PON1 foram determinados por amplificação por PCR. Resultados Com relação ao PON1 L55M, a frequência do alelo mutado (M) foi encontrada em 70,5% no PRG e em 53,5% nos controles; o alelo M foi significativamente associado a um risco aumentado de PRG (odds radio ajustado [ORadj] =2,07; intervalo de confiança [IC] 95%; p<0,001). No entanto, em relação ao PON1 Q192R, a frequência do alelo mutado R foi encontrada em 28,5% no PRG e em 33% nos controles. O alelo R não mostrou qualquer risco para PRG (ORadj 0,81; IC 95; p=0,329). Conclusão O presente estudo sugere que há um efeito do polimorfismo genético sobre PRG e fornece evidências adicionais que se combinam com as informações crescentes sobre asmaneiras pelas quais certos genótipos PON1 podemafetar o desenvolvimento do feto no útero.

Abstract Objective The aim of the present study was to examine the relation between the PON1 polymorphisms and recurrent pregnancy loss (RPL). Methods In a cross-sectional study, blood samples were collected from 100 females. DNA was extracted and PON1 genotypes were determined by polymerase chain reaction (PCR) amplification. Results Regarding PON1 L55M, the mutated allele (M) frequency was found in 70.5% in RPL and in 53.5% in controls; theMallele was significantly associated with an increased risk of RPL (adjusted odds ratio [ORadj]=2.07; 95% confidence interval [CI]; p<0.001). However, regarding PON1 Q192R, the R mutated allele frequency was found in 28.5% in RPL and in 33% in controls. The R allele did not show any risk for RPL (ORadj 0.81; 95%CI; p=0.329). Conclusion The present study suggests that there is an effect of genetic polymorphism on RPL and provides additional evidence that combines with the growing information about the ways in which certain PON1 genotypes can affect the development of the fetus in the uterus.

Resumo Objetivo: A enzima cardioprotetora Paraoxonase 1 (PON1) sofre importante influência de polimorfismos genéticos e fatores nutricionais. O objetivo deste estudo foi investigar a influência da alimentação, do estado nutricional e do polimorfismo C(-107)T sobre a atividade arilesterase da PON1 em crianças. Métodos: Estudo transversal com 97 crianças entre 5 e 8 anos, de ambos os sexos, de um ambulatório de pediatria no sul do Brasil. Realizou-se questionário sociodemográfico, de comportamento e de consumo alimentar, medidas antropométricas e coleta de sangue em laboratório. A atividade arilesterase da PON1 foi mensurada pela extinção de fenol (U/mL), realizada extração do DNA e análise do polimorfismo PON1 C(-107)T. O equilíbrio de Hardy-Weinberg foi testado com qui-quadrado e usada regressão linear para estimar a atividade da PON1 segundo quatro modelos de ajuste, erro aceitável de 5%. Resultados: Na amostra o sexo masculino representou 50,5%, 39,2% tinham 6 anos, 54,5% eram eutróficos e 51,5% tinha atividade da PON1 inferior à mediana (90,0;15-30 U/ml). A frequência dos genótipos foi 54,6% (53/97), 31,0% (30/97) e 14,4% (14/97), respectivamente, para CT, CC e TT, estiveram em equilíbrio de Hardy-Weinberg (p = 0,22). Na análise de regressão o modelo que incluiu variáveis sociodemográficas, de frequência do consumo de frutas, verduras, legumes, laticínios e feijões estimou uma variabilidade de 14,8% na atividade da PON1 combinada ao polimorfismo PON1 C(-107)T. Conclusões: Na infância uma alimentação de boa qualidade, com maior participação de alimentos saudáveis foi importante para predizer a atividade da enzima cardioprotetora PON1 combinada ao polimorfismo C(-107)T do gene da PON1.

Abstract Objective: The cardioprotective enzyme paraoxonase-1 (PON1) suffers an important influence from genetic polymorphisms and nutritional factors. The aim of this study was to investigate the influence of diet, nutritional status, and the C(-107)T polymorphism on PON1 arylesterase activity in children. Methods: This was a cross-sectional study with 97 children, aged between 5 and 8 years, of both genders, from a pediatric outpatient clinic in southern Brazil. A sociodemographic, behavioral, and food consumption questionnaire was applied, and anthropometric measurements and laboratory blood samples were taken. PON1 arylesterase activity was measured by phenol extinction (U/mL), and DNA extraction and analysis of the PON1 C(-107)T polymorphism were performed. The Hardy-Weinberg equilibrium was tested with the chi-squared test and linear regression was used to estimate PON1 activity according to four adjustment models, with an acceptable error of 5%. Results: In the sample, the male gender accounted for 50.5%, 39.2% were 6 years of age, 54.5% had normal weight, and 51.5% had PON1 activity below the median (90.0, 15-30 U/mL). Genotype frequency was 54.6% (53/97), 31.0% (30/97), and 14.4% (14/97), respectively, for CT, CC, and TT, consistent with the Hardy-Weinberg equilibrium (p = 0.22). In the regression analysis, the model that included sociodemographic variables as well as frequency of consumption of fruits, vegetables, legumes, dairy products, and beans estimated a variability of 14.8% in PON1 activity combined with the PON1 C(-107)T polymorphism. Conclusions: During childhood, a good-quality diet with greater inclusion of healthy foods was important to predict the activity of the cardioprotective enzyme PON1 combined with the C(-107)T polymorphism of the PON1 gene.

Resumen Introducción: La paraoxonasa-1 (PON1) sérica asociada a las HDL presenta dos polimorfismos comunes en las posiciones 192 y 55. Estos polimorfismos se consideran determinantes para la capacidad de las HDL de proteger a las LDL de su modificación oxidativa. En este contexto, el genotipo de PON1 se ha asociado con enfermedades cerebrovasculares, que incluyen el infarto cerebral. Objetivo: Determinar las frecuencias alélicas y genotípicas de PON1-L55M y PON1- Q192R, así como las actividades enzimáticas de PON1 en sujetos con y sin infarto cerebral aterotrombótico. Métodos: Se incluyeron 28 personas con infarto cerebral aterotrombótico y 29 sin infarto. Las genotipificaciones se realizaron mediante PCR-RFLP y las fenotipificaciones mediante la medición de las actividades paraoxonasa y arilesterasa en suero. Resultados: Para el polimorfismo Q192R, las frecuencias alélicas (Q/R) fueron 0.46/0.54 y 0.48/0.52 (p= 0.843) para el grupo control y el grupo con infarto, respectivamente. Mientras que para el polimorfismo L55M, las frecuencias alélicas (L/M) fueron 0.81/0.19 para el grupo control y 0.78/0.22 para el grupo con infarto (p= 0.610). Los niveles de actividad paraoxonasa no fueron significativamente diferentes entre los grupos control y con infarto (450 vs. 348 Ul/mL, p= 0.093). Mientras que los niveles de actividad arilesterasa fueron significativamente diferentes entre los grupos estudiados (90 vs. 70 Ul/mL, p= 0.001), sin embargo, al ajustarla por regresión lineal múltiple, dejo de ser significativa. Conclusión: Los polimorfismos Q192R y L55M, y la actividad paraoxonasa de la PON1 no son factores de riesgo para el infarto cerebral aterotrombótico en este estudio.

Abstract Background: The serum paraoxonase-1 (PON1) associated to HDL presents two common polymorphisms in the positions 192 and 55. These polymorphisms are considered determinant of the capacity of HDL to protect LDL from their oxidative modification. In this context, the PON1 genotype has been associated with cardiovascular diseases, including stroke. Objective: To determine the allelic and genotypic frequencies of PON1 L55M and Q192R as well as the enzymatic activities of PON1 in subjects with and without atherothrombotic stroke. Methods: There were included 28 people with atherothrombotic stroke and 29 without stroke. The genotyping was carried out by PCR-RFLP and the phenotyping by measurement of the activities of paraoxonase and arylesterase in serum. Results: For the polymorphism Q192R, the allelic frequencies (Q/R) were 0.46/0.54 and 0.48/0.52 (p= 0.843) for the control group and the group with stroke, respectively. While for the polymorphism L55M, the allelic frequencies (L/M) were 0.81/0.19 for the control group, and 0.78/0.22 for the group with stroke (p= 0.610). The activity levels of paraoxonase were not significantly different between the control and stroke groups (450 vs. 348 UI/mL, p= 0.093) While the activity levels of arylesterase were significantly different between the studied groups (90 vs. 70 UI/mL, p= 0.001); however, upon adjustment by multiple linear regression, it was not longer significant. Conclusion: The polymorphisms Q192R and L55M, and the paraoxonase activity of PON1 are not risk factors for atherothrombotic stroke according to the results of this study.

Objective: Paraoxonase 1 (PON1), an esterase associated with high-density lipoproteins (HDL), has several polymorphisms: the PON1 Q192R polymorphism (coding region) is responsible for variations in the paraoxonase activity of the enzyme. The PON1 192R allele is considered a risk factor for developing cardiometabolic disorders in some populations. The aim of the present study is to determine the distribution of the PON1 Q192R polymorphism and its association with the lipid profile and APO A1 in an Andean population. Materials and methods: A descriptive, cross-sectional study involving 79 healthy adults (26 males and 53 females), residents of the district of Junín at 4105 m.a.s.l. The PCR/RFLP technique was used for the analysis of the PON1 Q192R polymorphism. The serum values of the lipid profile and APO A1, and the paraoxonase and arylesterase activities of the enzyme were determined. Results: The distribution of the genotypes for PON1192 was QQ 13.9%, QR 45.6% and RR 40.5%, and the most frequent allele was 192R 63%. The baseline and stimulated paraoxonase activities were different based on their genotypes, while the esterase activity of the enzyme was not influenced by the polymorphism. On the other hand, no association was found between the PON1 Q192R polymorphism and the lipid profile and APO A1. Conclusions: The high prevalence of the PON1 192R allele among the studied population probably indicates an important role in the development of cardiometabolic diseases

Objetivo: La paraoxonasa 1 (PON1), una esterasa asociada a las lipoproteínas de alta densidad (HDL), presenta diversos polimorfismos: el polimorfismo PON1 Q192R (región codificante) es el responsable de las variaciones de la actividad paraoxonasa de la enzima. El alelo PON1 192R es considerado un factor de riesgo para desarrollar trastornos cardiometabólicos en algunas poblaciones. El objetivo del presente estudio es determinar la distribución de los polimorfismos PON1 Q192R, y su asociación con el perfil lipidíco y APO A1 en una población andina. Materiales y métodos: Estudio descriptivo, transversal, en el que participaron 79 personas adultas (26 hombres y 53 mujeres), clínicamente sanas, residentes del distrito de Junín a 4105 msnm. Se empleó la técnica PCR/RFLP para el análisis del sitio polimórfico Q192R del gen PON1, y se determinaron los valores séricos del perfil lipídico y APO A1, y las actividades paraoxonasa y arilesterasa de la enzima. Resultados: La distribución de los genotipos para PON1192 fue: QQ 13,9%, QR 45,6% y RR 40,5%, y el alelo más frecuente fue 192R 63%. Las actividades paraoxonasa basal y estimulada fueron diferentes según sus genotipos, mientras que la actividad esterasa de la enzima no estuvo influenciada por el polimorfismo. Por otra parte, no se encontró asociación entre el polimorfismo PON1 Q192R y el perfil lipídico y APO A1. Conclusiones: La alta prevalencia del alelo PON1 192R en la población estudiada probablemente indique un papel importante en el desarrollo de enfermedades cardiometabólica

Abstract Objectives Oxidative stress and inflammation are important processes in development of atherosclerosis. Paraoxonase 1 (PON1) is a bioscavenger enzyme associated with inflammation and oxidative stress. We evaluate the association of two single nucleotide polymorphisms in PON1 gene, and enzyme activities with lipid profile and glycemia. Methods This case-control study consisted of 126 patients with coronary artery disease (CAD) and 203 healthy controls. PON Q192R and L55M polymorphisms were detected by real-time PCR. Paraoxonase and arylesterase activities were determined spectrophotometrically. Blood glucose, cholesterol, triglycerides, HDL, and LDL were measured. Results PON1 QR192 polymorphism had a major effect on paraoxonase but no effect on arylesterase serum activities. Paraoxonase activity was higher in RR genotype and lowest in QQ genotype. Paraoxonase and arylesterase activities were higher in LL and lower in MM genotypes of PON1 LM55 polymorphism. RQ and LM variants showed intermediate activities between respective homozygous. Elevated concentrations of triglycerides in cases correlate with QQ variant or the presence of M allele. Glucose levels were elevated in cases with QQ variant or with the presence of M allele. Cholesterol and LDL did not show variations in control and cases with any variant of both polymorphisms. HDL is lower in cases with respect to controls independently of genotypes. All differences were significant with p < 0.05. Conclusions Our results confirm the relationship between variations in PON1 activities and lipid metabolism, and showed that genetically programmed low PON1 activities would have certain responsibility in the increase in glycemia and concomitantly the aggravation of atherosclerotic disease.

Resumen Objetivos La enzima paraoxonasa 1 (PON1), está asociada con el estrés oxidativo y la inflamación, procesos importantes en el desarrollo de la aterosclerosis. Evaluamos la asociación de 2 polimorfismos de un solo nucleótido en el gen PON1 y sus actividades enzimáticas con el perfil lipídico y la glucemia. Métodos Estudio caso-control en 126 pacientes con enfermedad coronaria y 203 controles sanos. Los polimorfismos PON Q192R y L55M fueron detectados por PCR en tiempo real y las actividades de paraoxonasa y arilesterasa por espectrofotometría. Se midieron glucemia, colesterol, triglicéridos, HDL y LDL. Resultados El polimorfismo PON1 QR192 afectó la actividad de paraoxonasa pero no la de arilesterasa. La actividad de paraoxonasa fue mayor en el genotipo RR y menor en QQ. Ambas actividades fueron mayores en el genotipo LL y menores en MM del polimorfismo PON1 LM55. Las variantes RQ y LM mostraron actividades intermedias entre los respectivos homocigotos. Concentraciones elevadas de triglicéridos en los casos correlacionaron con la variante QQ o la presencia del alelo M. Los niveles de glucosa fueron elevados en los casos QQ o con la presencia del alelo M. El colesterol y el LDL no variaron ni en los casos ni en los controles con ambos polimorfismos. El HDL fue menor en los casos respecto de los controles, independientemente del genotipo. Conclusiones Los resultados confirman la relación entre las variaciones en las actividades de PON1 y el metabolismo lipídico y mostraron que las bajas actividades de PON1 genéticamente programadas tendrían cierta responsabilidad en el aumento de la glucemia y, concomitantemente, en la agravación de la enfermedad aterosclerótica.

Purpose: To determine the Paraoxonase-1 (PON1) activity as well as its pheno- and genotypes at position 192 in Mexican subjects with diagnosis of coronary heart disease (CHD). Methods: We determined the PON1-192 polymorphism by PCR-RFLP, and serum PON1 activity, using either paraoxon (PONase activity) or phenylacetate (ARE activity) as substrates, in 155 clinically healthy individuals (control group), and 155 patients with at least one myocardial infarction (CHD group). The biochemical A/B phenotype was determined by the ratio of the NaCI 1 M-stimulated PONase activity divided by the ARE activity. Results: We found significantly lower PONase and ARE activities in CHD patients as compared to controls (233.1 ± 102.1 vs 295.8 ± 159.1 nmol/min/mL, and 103.1 ± 33.7 vs 220.2 ± 120.7. micromol/min/mL, respectively, p < 0.05 for both). Alíele and genotype frequencies for PON1-192 were similar in CHD patients and healthy controls. Moreover, in the control group, the PON1-192 Q/R genotype did not matched with the A/B phenotype as has been proposed by other studies. Conclusions: There were important differences in the ARE and PONase activities between Mexican CHD patients and controls, suggesting that PON1 activity could be a good marker of CHD risk, whereas PON 1-192 lacks of value to assess such risk.

Propósito: Determinar la actividad de la enzima paraoxonasa-1 (PON1), y sus feno- y genotipos en posición 192 de la cadena de aminoácidos en sujetos mexicanos con diagnóstico de enfermedad arterial coronaria (EAC). Métodos: El polimorfismo de la PON1-192 se determinó por PCR-RFLP, y la actividad PON1 en suero se estableció utilizando dos substratos, paraoxón (actividad PONasa) o fenilacetato (denominada actividad ARE), en 155 individuos clínicamente sanos (grupo control), y en 155 pacientes con enfermedad cardiovascular que habían sufrido al menos un infarto (grupo EAC). Los sujetos en el estudio fueron clasificados de acuerdo a su fenotipo bioquímico como AA, AB y BB si el cociente de la actividad PONasa estimulada con NaCI 1M dividida por la actividad ARE era < 2, entre 2 y 8, y > 8, respectivamente. Resultados: Las actividades PONasa y ARE fueron significativamente menores en los pacientes con EAC en relación con los sujetos control (233.1 ± 102.1 vs 295.8 ± 159.1 nmol/ min/mL, y 103.1 ± 33.7 vs 220.2 ± 120.7 micro-mol/min/mL, respectivamente, p < 0.05 para ambos). Las frecuencias tanto alélicas como genotípicas de PON1-192 fueron similares en ambos grupos. Además, en nuestro estudio el genotipo PON1-192Q/R no corresponde con el fenotipo bioquímico A/B como se había propuesto anteriormente por otros autores. Conclusiones: Los sujetos mexicanos con EAC presentan actividades PON1 menores que los individuos control, sugiriendo que la actividad PON1, podría ser marcador de riesgo de EAC, mientras que el genotipo PON1-192 carece de valor informativo en la evaluación de dicho riesgo en esta población en particular.