Paraoxonase 1 Polymorphisms Research Articles

A multiplex PCR-based DNA assay for the detection of paraoxonase gene cluster polymorphisms

Paraoxonase (PON) is a high-density lipoprotein (HDL) associated protein which is supposed to protect low-density lipoprotein (LDL) against oxidation and to play a role in the development of atherosclerosis. Interindividual variability in serum PON activity is attributable to common variants in components of the PON gene cluster on chromosome 7. We describe experimental conditions that permit the simultaneous determination of three common PON polymorphisms (PON1-192, PON1-55 and PON2-311) that are tightly associated with an increased risk of atherosclerosis. We used a multiplex PCR-based DNA assay using mismatch primers that introduce a unique recognition site for the endonuclease HinfI in the PCR products in case of presence of the R allele of PON 1-192, of the L allele of PON1-55 and of the S allele of PON2-311. The restriction analysis with HinfI allows to identify an electrophoretic band pattern which is specific for the combination of the three polymorphisms. This technique could be applied in the association studies aimed at assessing the role of PON and their polymorphisms in many clinical settings. In a preliminary study on a small population sample from south Italy about 10% of chromosomes exhibited the presumed risk-related haplotype R(192)/L(55)/S(311).

Paraoxonase and atherosclerosis.

There is considerable evidence that the antioxidant activity of high density lipoprotein (HDL) is largely due to the paraoxonase-1 (PON1) located on it. Experiments with transgenic PON1 knockout mice indicate the potential for PON1 to protect against atherogenesis. This protective effect of HDL against low density lipoprotein (LDL) lipid peroxidation is maintained longer than is the protective effect of antioxidant vitamins and could thus be more important. There is evidence that the genetic polymorphisms of PON1 least able to protect LDL against lipid peroxidation are overrepresented in coronary heart disease, particularly in association with diabetes. However, these polymorphisms explain only part of the variation in serum PON1 activity; thus, a more critical test of the hypothesis is likely to be whether low serum PON1 activity is associated with coronary heart disease. Preliminary case-control evidence suggests that this is indeed the case and, thus, that the quest for dietary and pharmacological means of modifying serum PON1 activity may allow the oxidant model of atherosclerosis to be tested in clinical trials.

The BB-paraoxonase genotype is associated with impaired brachial reactivity after acute hypertriglyceridemia in healthy subjects.

The possible relationship between paraoxonase (PON) gene polymorphism and brachial reactivity in healthy adult subjects in the presence of acute hypertriglyceridemia (HT), as a prooxidant factor, was investigated. In 101 healthy subjects the response to flow- induced vasodilatation was measured before and after Intralipid infusion. In the same subjects the A/B PON polymorphism was detected. The frequency was 0.545 for AA genotype, 0.356 for the AB genotype, and 0.099 for the BB genotype. At baseline all genotype groups had a similar increase in brachial artery diameter and flow. After Intralipid infusion, subjects sharing the BB genotype had a significant decrease vs. baseline values in changes in brachial artery diameter (P for trend < 0.001 vs. the other genotypes), but not in flow. In a subgroup of 55 subjects distributed among the 3 PON genotypes the same study protocol was repeated by buccal nitroglycerine administration to study the endothelium-independent vasodilatation. Again, subjects with the BB genotype had the worse vasodilation (P for trend < 0.001). Furthermore, subjects sharing the BB genotype had the lowest endothelium-independent and -dependent changes in diameter (P for trend < 0.001 vs. the other genotypes) independently of gender ratio, basal plasma triglycerides concentrations, and changes in plasma triglycerides concentrations. In conclusion, our study demonstrates that transient HT decreases vascular reactivity more in subjects with the PON BB genotype than in those with the other PON genotypes.

Rabbits possess a serum paraoxonase polymorphism similar to the human Q192R

Serum paraoxonase (PON1) is a high-density lipoprotein (HDL)-associated enzyme that hydrolyses aromatic esters, organophosphates and lactones and can protect low-density lipoprotein (LDL) against oxidation. These properties are influenced by a well-characterized polymorphism (Q192R) in human PON1. We now report the identification and characterization of a phenotypically similar, but genetically distinct polymorphism in rabbit PON1. This polymorphism in rabbits was detected by phenotyping sera obtained from 16 inbred rabbit strains and 20 outbred New Zealand White rabbits by paraoxonase/arylesterase activity. The genetic basis of the rabbit polymorphism was determined by DNA sequencing and found to reside in a region distinct from the human Q192R and M55L polymorphisms. Three variant nucleotides within exon 4 (corresponding to P82S, K93E and S1O1G) were found to segregate with the observed rabbit PON1 phenotypes (rPON1A and rPON1B). The rPON1A and rPON1B proteins were purified and compared to the two human isoforms (192Q and 192R). The human and rabbit PON1s displayed similar characteristics with respect to physical properties and substrate specificity. However, rPON1A and rPON1B hydrolysed a variety of substrates at different rates. The rPON1A was also at least three-fold more efficient at protecting LDL from oxidation than rPON1B. Our characterization of a rabbit PON1 polymorphism provides useful insights into important functional residues in PON1. In addition, due to the observed similarities between the rabbit and human polymorphisms, the rabbit may serve as a good model to examine the effect of human PON1 polymorphisms in disease development.

Polymorphisms in the human paraoxonase (PON1) promoter.

Paraoxonase (PON1) is a protein component of high-density lipoprotein (HDL) particles that protects against oxidative damage to both low-density lipoprotein and HDL and detoxifies organophosphorus pesticides and nerve agents. A wide range of expression levels of PON1 among individuals has been observed. We examined the promoter region of PON1 for genetic factors that might affect PON1 activity levels. We conducted a deletion analysis of the PON1 promoter region in transient transfection assays and found that cell-type specific promoter elements for liver and kidney are present in the first 200bp upstream of the coding sequence. Sequence analysis of DNA from a BAC clone and a YAC clone identified five polymorphisms in the first 1000 bases upstream of the coding region at positions -108, -126, -162, -832 and -909. Additionally, the promoter sequences of two individuals expressing high levels of PON1 and two individuals expressing low levels of PON1 were analysed. The two polymorphisms at -126 and -832 had no apparent effect on expression level in the reporter gene assay. The polymorphisms at position -909, -162 (a potential NF-I transcription factor binding site) and -108 (a potential SP1 binding site) each have approximately a two-fold effect on expression level. The expression level effects of the three polymorphisms appear not to be strictly additive and may depend on context effects.

Paraoxonase Polymorphisms and the Risk of Arterial Ischemic Stroke among Young Adults

84 Despite extensive investigation, the etiology of arterial ischemic stroke (AIS) in the young remains unknown in one-third of patients. Serum paraoxonase (PON) is an HDL-associated esterase that hydrolyzes products of lipid peroxidation and prevents the oxidation of LDL, thus exerting anti-oxidant and anti-atherogenic effects. PON levels have been shown to be significantly reduced in patients with coronary artery disease. Two common polymorphisms in the PON gene, the 55 Leu (L allele)→ Met (M allele) and the 192 Gln (A allele)→ Arg (B allele) substitutions, influence PON activity in a gene-dose dependent manner. As the association of these polymorphisms with cardiovascular disease and stroke is controversial, we investigated their role in a young population with AIS. We carried out a cross-sectional study including 138 patients (84 women, 54 men; mean age: 32.9 years) with a first AIS before the age of 45 years and 102 age-matched controls. The diagnosis of AIS was based on clinical and neuroimaging findings. Fragments of the coding region of the PON gene encompassing the 192 and 55 polymorphisms were analyzed by PCR amplification and posterior restriction endonuclease digestion. Among patients, 17.4% were found to be homozygous and 46.4% heterozygous for the B allele at position 192, compared to 8.8% and 38.3% respectively among controls. This yielded a significantly higher allele frequency of the 192 B allele among patients than controls: 0.406 vs. 0.279, P=0.0056, OR=1.76 (CI=1.17–2.65). Regarding the PON 55 L allele, there was no significant difference between patients and controls (P=0.59). The presence of hyperlipidemia, diabetes mellitus and smoking, factors known to influence PON levels, were analyzed in the patient and control groups and did not significantly alter the difference in allele frequencies. These findings suggest that the PON 192 B allele is associated with an increased risk of AIS among young adults. However, studies including a larger number of patients and analyzing the effect of PON on atherosclerosis are necessary to better understand the role of these observations in the development of AIS in the young.

Paraoxonase Polymorphisms and the Risk of Arterial Ischemic Stroke among Young Adults

84 Despite extensive investigation, the etiology of arterial ischemic stroke (AIS) in the young remains unknown in one-third of patients. Serum paraoxonase (PON) is an HDL-associated esterase that hydrolyzes products of lipid peroxidation and prevents the oxidation of LDL, thus exerting anti-oxidant and anti-atherogenic effects. PON levels have been shown to be significantly reduced in patients with coronary artery disease. Two common polymorphisms in the PON gene, the 55 Leu (L allele)→ Met (M allele) and the 192 Gln (A allele)→ Arg (B allele) substitutions, influence PON activity in a gene-dose dependent manner. As the association of these polymorphisms with cardiovascular disease and stroke is controversial, we investigated their role in a young population with AIS. We carried out a cross-sectional study including 138 patients (84 women, 54 men; mean age: 32.9 years) with a first AIS before the age of 45 years and 102 age-matched controls. The diagnosis of AIS was based on clinical and neuroimaging findings. Fragments of the coding region of the PON gene encompassing the 192 and 55 polymorphisms were analyzed by PCR amplification and posterior restriction endonuclease digestion. Among patients, 17.4% were found to be homozygous and 46.4% heterozygous for the B allele at position 192, compared to 8.8% and 38.3% respectively among controls. This yielded a significantly higher allele frequency of the 192 B allele among patients than controls: 0.406 vs. 0.279, P=0.0056, OR=1.76 (CI=1.17–2.65). Regarding the PON 55 L allele, there was no significant difference between patients and controls (P=0.59). The presence of hyperlipidemia, diabetes mellitus and smoking, factors known to influence PON levels, were analyzed in the patient and control groups and did not significantly alter the difference in allele frequencies. These findings suggest that the PON 192 B allele is associated with an increased risk of AIS among young adults. However, studies including a larger number of patients and analyzing the effect of PON on atherosclerosis are necessary to better understand the role of these observations in the development of AIS in the young.

Polymorphisms of human paraoxonase 1 gene (PON1) and susceptibility to diabetic nephropathy in type I diabetes mellitus.

Oxidative stress is a putative mechanism in the development of diabetic nephropathy. Paraoxonase gene 1 is an HDL-bound enzyme that protects tissues against oxidative damage. Three common polymorphisms of paraoxonase gene 1, T-107C in the promoter, Leu54Met and Gln192Arg, that modify paraoxonase activity have been associated with cardiovascular disease. This study aimed to find whether these polymorphisms also contribute to the development of diabetic nephropathy. The association between diabetic nephropathy and these three polymorphisms was examined in a case-control study. For this purpose, genomic DNA was collected from 188 patients with Type I (insulin-dependent) diabetes mellitus and diabetic nephropathy and from 179 unrelated patients with Type I diabetes but without diabetic nephropathy despite the duration of diabetes of 15 or more years. The genotype and allele frequencies for each of the three polymorphisms (T-107C, Leu54Met and Gln192Arg) were similar in cases and control subjects. The three polymorphisms in paraoxonase gene 1 that have been associated with serum levels of paraoxonase are not associated with diabetic nephropathy. We show that this genetically determined component of the antioxidant capacity of HDL does not play a critical part in the development of diabetic nephropathy.

The paraoxonase Leu–Met54 and Gln–Arg191 gene polymorphisms are not associated with the risk of coronary heart disease

Background: Evidence has been presented that gene polymorphisms (PON54 L/M, PON191 Q/R) of paraoxonase are risk factors of coronary heart disease. Results: We determined both PON genotypes in 535 male individuals who were free of vascular disease and in 2249 male subjects who underwent coronary angiography, and analysed the relation of both gene variations to CAD and MI. Both gene polymorphisms were in linkage disequilibrium ( P<0.0001). Coronary artery disease: the PON54 gene polymorphism was not associated with an increased risk of CAD. In the total sample and also in younger subjects, an association of the PON191 gene variation with the risk of CAD was not detected when the control group of individuals without coronary heart disease was compared with patients with at least one diseased vessel (verified by coronary angiography). In individuals younger than 62 years, a moderate increase in the relative risk of CAD associated with the PON191 R allele (1.45 (1.08–1.95); P=0.015) were found, when subjects without vessel disease (verified by coronary angiography) were compared with CAD patients. Myocardial infarction: an association of the PON54 gene variation with MI was not detected when the control group of individuals without coronary heart disease were compared with patients with at least one MI. A marginal increase in the risk of MI associated with the PON54 LL genotype (OR 1.27 (1.05–1.51); P=0.011) were detected when patients without MI but with coronary angiography were compared with MI positive patients. Subgroup analyses of low- and high-risk populations did not reveal any association of both PON gene polymorphisms with CAD or MI. Conclusion: The present findings do not strengthen the hypothesis that the paraoxonase gene polymorphisms are independently associated with coronary heart disease indicating that these gene variations are of little usefulness as genetic markers of cardiovascular disease.

Paraoxonase polymorphisms, pesticide exposure and Parkinson's disease in a Caucasian population.

Parkinson's disease (PD) has been associated with exposure to pesticides and oxidative injury. The involvement of paraoxonase in both pesticide metabolism and lipid peroxidation suggests that it may play a role in the pathogenesis of PD. We examined the frequency of polymorphic alleles of the PON1 and PON2 genes in a sample of caucasian subjects with PD. The frequency distribution of these genotypes did not differ significantly between patients and controls, including those who had reported exposure to pesticides.

The human paraoxonase Gln-Arg192 (Q/R) polymorphism in Turkish patients with coronary artery disease

It has been suggested that a Q/R (Gln192Arg) polymorphism of paraoxonase (PON) might be associated with the predisposition to coronary artery disease (CAD). Therefore, we studied the human paraoxonase gene (PON1) polymorphism in Turkish patients with CAD by polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP). This polymorphism was determined in 96 CAD patients and in 105 control subjects. The frequencies of the QQ, QR, and RR genotypes were found as 36.5, 52.0, and 11.5% in CAD patients and 48.6, 41.0, and 10.4% in control subjects, respectively. The QR genotype was the most common in the patient group, whereas the QQ genotype was more frequent in individuals without CAD. Frequency of the R allele was higher among CAD patients compared to controls (38.5% versus 31%). However, neither the genotype nor the allele distribution of the Gln92Arg polymorphism of PON1 was statistically significantly different between the two groups ( P>0.05). Although both systolic and diastolic blood pressure levels were slightly higher in patients with the QQ genotype, there was no differences in regard to age, sex, serum triglyceride, total cholesterol or high-density lipoprotein cholesterol among CAD patients with different PON1 Gln192Arg genotypes. In summary, our results suggest that no association exists between the Gln192Arg polymorphism of paraoxonase and CAD in Turkish patients.

Apolipoprotein E levels in patients with probable Alzheimer S disease are affected by paraoxonase polymorphisms

Apolipoprotein E levels in patients with probable Alzheimer S disease are affected by paraoxonase polymorphisms

Paraoxonase polymorphism (Gln192Arg) as a determinant of the response of human coronary arteries to serotonin.

Background-Oxidation of LDL plays a role in endothelial dysfunction. Paraoxonase, an enzyme present on HDL, protects LDL against oxidation. Paraoxonase activity is genetically determined in part, and 3 genotypes have been described with variable enzymatic activity. We hypothesized that the paraoxonase polymorphism might influence endothelial function. Methods and Results-Twenty-seven patients with clinical manifestations of coronary artery disease underwent provocative testing by intracoronary administration of serotonin. None of the coronary arteries studied had significant (>50%) stenosis. Ten patients had the QQ genotype and 17 had the QR genotype. At proximal segments, the mean percentage reduction in lumen diameter in response to serotonin was greater in QQ patients than in QR patients (10(-5) mol/L: P<0.05; 10(-4) mol/L: P<0.006). Similarly, at distal segments, constriction in response to serotonin was greater in QQ patients than in QR patients (10(-6) mol/L: P<0. 03; 10(-5) mol/L: P<0.07). Conclusions-These results suggest a higher synthesis or release of endothelium-derived relaxing factors to counteract the vasoconstrictor effect of serotonin in patients with the R allele. These findings provide evidence that the paraoxonase polymorphism may play a role in the regulation of coronary vasomotor tone.

Promoter polymorphisms of human paraoxonase PON1 gene and serum paraoxonase activities and concentrations.

Paraoxonase (PON) is a serum enzyme with a wide species distribution. It protects lipoproteins from toxic oxidative modifications and is an antiatherogenic mechanism of major potential. Activity levels of PON are major determinants of the protective function; consequently, factors that influence PON levels are of particular relevance. The present study has identified 3 polymorphisms in the promoter region of the human PON1 gene. Cell transfection studies have revealed their variable impact on promoter activity, with up to 2-fold differences in reporter gene expression. Genotyping studies have established that the polymorphisms are frequent in the population, a finding that is consistent with a major impact on PON concentrations. The physiological relevance of the polymorphisms was underlined by showing that they are associated with highly significant differences in serum concentrations and activities of PON. The study thus firmly establishes a genetic basis for variations in serum PON levels and, consequently, serum PON activity. It is consistent with the suggestion that variations in a major antioxidant function of high density lipoprotein are, to an important degree, genetically determined.

Risk of myocardial infarction associated with Gln/Arg 192 polymorphism in the human paraoxonase gene and diabetes mellitus. The REGICOR Investigators.

There is increasing evidence that paraoxonase, an HDL-linked enzyme, exerts its effect by removing lipid-peroxidation products. We have conducted a case-control study in Gerona, Spain, to find out whether paraoxonase1 polymorphism at codon 192 (Q and R alleles) is associated with increased risk of coronary heart disease, and how diabetes mellitus, associated with high oxidative risk, influences such an association. One hundred and fifty six consecutive myocardial infarction patients and 310 age- and sex-matched control subjects were studied. There were no differences in the distribution of genotype and allele frequencies between patients and controls. The odds ratios for diabetes and dyslipaemia in control and patients stratified by genotype group were compared. Whereas dyslipaemic status was significantly related to myocardial infarction in QQ homozygotes and R carriers, diabetes mellitus was significantly associated with myocardial infarction only in R-carrier subjects. In logistic regression analysis, diabetic R carriers demonstrated a more than two and a half-fold increase in myocardial infarction risk compared with non-diabetic R carriers (OR: 2.65, P<0.05). These data indicate that the R allele of the paraoxonase1-192 polymorphism is not an independent risk factor for myocardial infarction in our population. However, the interaction between this polymorphism and diabetes mellitus leads to increased myocardial infarction risk in diabetic patients with the R allele.

Paraoxonase polymorphism and its effect on male reproductive outcomes among Chinese pesticide factory workers

Paraoxonase polymorphism and its effect on male reproductive outcomes among Chinese pesticide factory workers

Paraoxonase polymorphism and its effect on male reproductive outcomes among Chinese pesticide factory workers

Paraoxonase polymorphism and its effect on male reproductive outcomes among Chinese pesticide factory workers

Paraoxonase polymorphism and its effect on male reproductive outcomes among Chinese pesticide factory workers.

Serum paraoxonase has been associated with the metabolism of organophosphate pesticides in humans. Molecular analysis of the human paraoxonase gene (PON1) has revealed that Arg192 homozygotes have a greater detoxifying capability than Gln192 homozygotes. We examined the effects of PON1 genotypes on male reproductive outcomes and its interaction with exposure to organophosphate pesticides. We studied 60 Chinese pesticide-factory workers and 89 textile-factory workers who were unexposed to pesticides. The respective allele frequencies of Arg192 and Gln192 were 0.62 and 0.38. Pesticide exposure among 36 exposed subjects and 12 unexposed subjects, regardless of gender, was assessed by personal measurement of pesticide residues over an entire 8-hr shift and measurement of urinary p-nitrophenol level over a 24-hr period. We analyzed semen and hormone data collected from male subjects. When the three PON1 genotypes were analyzed separately, a gene dose effect was not detected. We used the unexposed Arg192 homo/heterozygotes as the reference group, and re-analyzed the data. Exposed Arg192 homo/heterozygotes had significantly lower sperm count (chi 2 = 9.01, P < 0.01) and lower percentage of sperm with normal morphology (chi 2 = 4.18, P < 0.05) than the reference group. Both unexposed Gln192 homozygotes (chi 2 = 4.90, P < 0.05) and exposed Arg192 homo/heterozygotes (chi 2 = 10.00, P < 0.01) showed significantly lower sperm concentrations than the reference group. In addition, exposed Arg192 homo/heterozygotes had significantly higher serum LH levels (chi 2 = 7.94, P < 0.01) than the reference group. Because of a small sample size, our findings are highly preliminary. Nevertheless, it calls for further investigation of the interaction between the PON1 genotype and organophosphate pesticide exposure on male reproductive outcomes.

No association of paraoxonase gene polymorphism with atherosclerosis or Alzheimer's disease.

Both AD and paraoxonase (PON) have been reported to be related to lipids and atherosclerosis, suggesting that the PON gene (PON) is a possible genetic risk factor for AD. We found no association of PON polymorphism with severity of atherosclerosis, densities of AD-type, neuropathologic change, or development of AD in 47 AD and 90 nondemented patients. Our study suggests that PON polymorphism does not play a causal role in the development of atherosclerosis or AD.

Paraoxonase polymorphisms are not associated with cardiovascular risk in renal transplant recipients

Paraoxonase (PON1) gene variants have been identified as risk factors for cardiovascular disease (CVD). There are two common PON1 polymorphisms at position 55 (Leu-Met change) and 192 (Gln-Arg change) of the amino acid chain. Leucine at position 55 and arginine at position 192 have been associated with increased cardiovascular risk. The increased prevalence of CVD in renal transplant recipients can be only partly explained by the increased prevalence of conventional risk factors. We therefore investigated PON1 polymorphisms in renal transplant recipients (N = 491) with (N = 103) and without CVD (N = 388) using polymerase chain reaction-restriction fragment length analysis. PON1 polymorphisms and their associated PON1/arylesterase activities were also assessed in a subgroup of patients (N = 165). The genotype distribution and allele frequencies for both polymorphisms were similar in both groups. The frequencies for LL, LM, and MM genotypes for the 55 position in patients with CVD were 0.39, 0.51, and 0.10, respectively, compared with 0.43, 0.43, and 0.14 in patients without CVD (P = 0.31). The distribution for the QQ, QR, and RR genotypes at the 192 position were 0.48, 0.43, and 0.09, respectively, in patients with CVD compared with 0.46, 0.46, and 0.08 in patients without CVD (P = 0.8). There were highly significant differences in serum activities of PON1/arylesterase between genotypes defined by 55 and 192 polymorphisms. Leucine at position 55 and arginine at position 192 were associated with higher activities. These data indicate that there is no association between the PON1 gene variants, conferring higher enzyme activity, and the increased cardiovascular risk in renal transplant recipients.