Paraoxonase (PON 1) as a biomarker of susceptibility for organophosphate toxicity

Paraoxonase (PON1) is an A-esterase capable of hydrolysing the active metabolites (oxons) of a number of organophosphorus (OP) insecticides such as parathion, diazinon and chlorpyrifos. PON1 activity is highest in liver and plasma, and among animal species significant differences exist, with birds and rabbits displaying very low and high activity, respectively. Human PON1 has two polymorphisms in the coding region (Q192R and L55M) and five polymorphisms in the promoter region. The Q192R polymorphism imparts different catalytic activity toward some OP substrates, while the polymorphism at position -108 (C/T) is the major contributor to differences in the level of PON1 expression. Animal studies have shown that PON1 is an important determinant of OP toxicity, with animal species with a low PON1 activity having an increased sensitivity to OPs. Administration of exogenous PON1 to rats or mice protects them from the toxicity of OPs. PON1 knockout mice display a high sensitivity to the toxicity of diazoxon and chlorpyrifos oxon, but not paraoxon. In vitro assayed catalytic efficiencies of purified PON₁₉₂ isoforms for hydrolysis of specific oxon substrates accurately predict the degree of in vivo protection afforded by each isoform. Low PON1 activity may also contribute to the higher sensitivity of newborns to OP toxicity.     

Genetic variability in the cytochrome P450-paraoxonase 1 (PON1) pathway for detoxication of organophosphorus compounds

Detoxication of organophosphorus (OP) compounds is affected by genetic and environmental modulation of a number of enzymes involved in the process. For organophosphorothioate insecticides, different P450 isozymes and variants carry out two reactions that have quite different consequences; (1) they bioactivate their parent compounds to highly toxic oxon forms that are many times more toxic than the parent compounds, and (2) concurrently, they dearylate the parent OP compounds, generating much less toxic metabolites. The ratios at which these different P450s carry out bioactivation versus dearylation differ among the P450 isozymes. The detoxication of the oxon forms of diazinon and chlorpyrifos is achieved by hydrolysis to the respective aromatic alcohols and diethyl phosphates primarily by paraoxonase 1 (PON1), a plasma enzyme tightly associated with high-density lipoprotein particles and also found in liver. Stoichiometric binding to other targets also contributes to the detoxication of these oxons. PON1 is polymorphically distributed in human populations with an amino acid substitution (Gln/Arg) at position 192 of this 354-amino acid protein (the initiator Met residue is cleaved on maturation) that determines the catalytic efficiency of hydrolysis of some substrates. In addition to the variable catalytic efficiency determined by the position 192 amino acid, protein levels of PON1 vary by as much as 15-fold among individuals with the same PON1(192) genotype (Q/Q; Q/R; R/R). The generation of PON1 null mice and transgenic mice, expressing each of the human PON1(192) alloforms in place of mouse PON1, has allowed for the examination of the physiological function of the PON1(192) alloforms in OP detoxication. Sensitivity to diazoxon exposure is primarily determined by the plasma level of PON1, whereas for chlorpyrifos oxon exposure, both the plasma PON1 level and the position 192 amino acid are important--PON1(R192) is more efficient in inactivating chlorpyrifos oxon than is PON1(Q192). The availability of PON1 null mice provides an opportunity to examine the contribution of other enzymes in the OP detoxication pathways without PON1 interference.     

Paraoxonase activity against nerve gases measured by capillary electrophoresis and characterization of human serum paraoxonase (PON1) polymorphism in the coding region (Q192R)

An analytical method for determining paraoxonase activity against sarin, soman and VX was established. We used capillary electrophoresis to measure directly the hydrolysis products: alkyl methylphosphonates. After enzymatic reaction of human serum paraoxonase (PON1) with nerve gas, substrate was removed with dichloromethane, and alkyl methylphoshphonates were quantified by capillary electrophoresis of reversed osmotic flow using cationic detergent and sorbic acid. This method was applied to the characterization of human serum PON1 polymorphism for nerve gas hydrolytic activity in the coding region (Q192R). PON1-192 and PON1-55 genotypes were determined by their gel electrophoretic fragmentation pattern with restriction enzymes after polymerase chain reaction (PCR) of blood leukocyte genomic DNA. Frequencies of genotypes among 63 members of our institutes with PON1-192 and PON1-55 were 9.5% (¹⁹²QQ), 30.1% (¹⁹²QR) and 44.4% (¹⁹²RR), and 82.5% (⁵⁵LL), 17.5% (⁵⁵LM) and 0% (⁵⁵MM), respectively. ¹⁹²Q and ¹⁹²R enzymes were purified from the respective genotype human plasma, using blue agarose affinity chromatography and diethyl amino ethane (DEAE) anion exchange chromatography. V_max and K_m were measured using Lineweaver-Burk plots for hydrolytic activities against sarin, soman and VX at pH 7.4 and 25 °C. For sarin and soman, the V_max for ¹⁹²Q PON1 were 3.5- and 1.5-fold higher than those for ¹⁹²R PON1; and k_cat/K_m for ¹⁹²Q PON1 were 1.3- and 2.8-fold higher than those for ¹⁹²R PON1. For VX, there was little difference in V_max and k_cat/K_m between ¹⁹²Q and ¹⁹²R PON1, and VX hydrolyzing activity was significantly lower than those for sarin and soman. PON1 hydrolyzed sarin and soman more effectively than paraoxon.     

Paraoxonase 1 Polymorphisms Within a Mississippi USA Population as Possible Biomarkers of Enzyme Activities Associated With Disease Susceptibility

Paraoxonase (PON1) hydrolyzes paraoxon (PO) and diazoxon (DZO), active metabolites of insecticides parathion and diazinon. The PON1 gene has single nucleotide polymorphisms (SNPs) including a codon 192 arginine (R) to glutamine (Q) and methionine (M) to leucine (L) at codon 55. Hydrolysis of PO (POase), DZO (DZOase), dihydrocoumarin (lactonase), and phenyl acetate (arylesterase) were evaluated for associations with race, gender, age, and PON1 55/192 SNP genotypes. Variables were analyzed both individually and in combination. QQ individuals had higher lactonase (p < 0.001) than RR individuals. This might partially explain why predominantly RR African Americans have higher rates of coronary disease than predominantly QQ Caucasians. Significant (p < 0.001) differences in arylesterase were seen among genotypes with QQ and MM lowest whereas RR and LL were highest. This opposes the prevailing belief that arylesterase is unaffected by genotype and suggests that this activity cannot be used to quantify PON1 protein.     

Paraoxonase (PON)1 Q192R functional genotypes and PON1 Q192R genotype by smoking interactions are risk factors for the metabolic syndrome,but not overweight or obesity

Abstract

Background

The metabolic syndrome (MetS) is a complex of multiple risk factors that contribute to the onset of cardiovascular disorder, including lowered levels of high-density lipoprotein (HDL) and abdominal obesity. Smoking, mood disorders, and oxidative stress are associated with the MetS. Paraoxonase (PON)1 is an antioxidant bound to HDL, that is under genetic control by functional polymorphisms in the PON1 Q192R coding sequence.

Aims and methods

This study aimed to delineate the associations of the MetS with plasma PON1 activity, PON1 Q192R genotypes, smoking, and mood disorders (major depression and bipolar disorder), while adjusting for HDL cholesterol, body mass index, age, gender, and sociodemographic data. We measured plasma PON1 activity and serum HDL cholesterol and determined PON1 Q192R genotypes through functional analysis in 335 subjects, consisting of 97 with and 238 without MetS. The severity of nicotine dependence was measured using the Fagerström Nicotine Dependence Scale.

Results

PON1 Q192R functional genotypes and PON1 Q192R genotypes by smoking interactions were associated with the MetS. The QQ and QR genotypes were protective against MetS while smoking increased metabolic risk in QQ carriers only. There were no significant associations between PON1 Q192R genotypes and smoking by genotype interactions and obesity or overweight, while body mass index significantly increased MetS risk. Smoking and especially severe nicotine dependence are significantly associated with the MetS although these effects were no longer significant after considering the effects of the smoking by PON1 Q192R genotype interaction. The MetS was not associated with mood disorders, major depression or bipolar disorder.

Discussion

PON1 Q192R genotypes and genotypes by smoking interactions are risk factors for the MetS that together with lowered HDL and increased body mass and age contribute to the MetS.     

The Arg192 isoform of paraoxonase with low sarin-hydrolyzing activity is dominant in the Japanese

The high-density-lipoprotein-associated enzyme paraoxonase, which has a role in the detoxification of organophosphorus compounds, is known to be polymorphic in humans. The Arg₁₉₂ isoform of paraoxonase hydrolyzes paraoxon more rapidly than the Gln₁₉₂ isoform. However, with respect to the hydrolysis of toxic nerve agents, such as diazoxon, soman, and sarin, the Arg₁₉₂ isoform displays a lower activity than the other isoform. To evaluate the possibility that the genetic polymorphism was involved in the aggravated extent of human injury in the sarin gas poisoning incident in the Tokyo subway in March 1995, we investigated the prevalance of this polymorphism in the Japanese population. We found that the Arg₁₉₂ allele is more common in the Japanese (allele frequency: 0.66) than in people of other races (ranging 0.24–0.31). In the Japanese, 135 out of the 326 subjects (41.4%) investigated were homozygous for the Arg₁₉₂ allele, which shows a very low hydrolysis activity for sarin. Thus, there seems to be a racial difference in vulnerability to toxic nerve agents, such as sarin. The dominance of the Arg₁₉₂ allele in the Japanese population probably worsened the tragedy of March 1995 in the Tokyo subway. Received: 5 April 1997 / Accepted: 10 June 1997     

Paraoxonase1, its Q192R polymorphism and HDL-cholesterol in relation to intensive cardiac care unit stay in ischemic heart disease

AIMS AND OBJECTIVES:

The present study was evaluated the atheroprotective potential of paraoxonase1 (PON1) and its Q192R polymorphism, to determine whether this polymorphism, which is responsible for differential PON1 activity plays any role in the pathogenesis, severity and extent of coronary artery disease (CAD).

MATERIALS AND METHODS:

This hospital-based cross-sectional study investigated 60 diagnosed cases of CAD and 60 age and gender matched controls. All were assessed for serum PON1 activity, PON1 Q192R polymorphism and for classical cardiovascular risk factors. Individual serum phenotyping for PON1 Q192R polymorphism was determined by double substrate hydrolysis assay. Severity of CAD was assessed by the length of intensive cardiac care unit (ICCU) stay.

RESULTS:

Serum PON1 activity is significantly reduced in cases of CAD (92.6 ± 31.13 IU/L when compared with controls (105.26 ± 32.53 IU/L). Furthermore, serum arylesterase activity is reduced in CAD patients (90.31 ± 23.26 kU) when compared with the control subjects (101.61 ± 28.68 kU). Serum PON1 and arylesterase activities are significantly negatively correlated with the length of ICCU stay (r = −393 and r = −374 respectively). There is no significant difference in the occurrence of CAD and length of ICCU stay among the PON1 phenotypes (P = 0.92). Logistic regression analysis after adjustment of established risk factors revealed no significant association between CAD risk and PON1 Q192R polymorphism (odds ratios: 1.179 [95% confidence intervals: 0.507-2.744], P = 0.702).

SUMMARY AND CONCLUSIONS:

The current study demonstrates that the activity of the PON1 enzyme may be more important factor than the PON1 Q192R polymorphism in the severity and extent of CAD.     

Indications that paraoxonase-1 contributes to plasma high density lipoprotein levels in familial hypercholesterolemia

HDL-associated paraoxonase type 1 (PON1) can protect LDL and HDL against oxidative modification in vitro and therefore may protect against cardiovascular disease. We investigated the effects of PON1 levels, activity, and genetic variation on high density lipoprotein-cholesterol (HDL-C) levels, circulating oxidized LDL (OxLDL), subclinical inflammation [high-sensitive C-reactive protein (Hs-CRP)], and carotid atherosclerosis. PON1 genotypes (L55M, Q192R, -107C/T, -162A/G, -824G/A, and -907G/C) were determined in 302 patients with familial hypercholesterolemia. PON1 activity was monitored by the hydrolysis rate of paraoxon, diazoxon, and phenyl acetate. PON1 levels, OxLDL, and Hs-CRP were determined using an immunoassay. The genetic variants of PON1 that were associated with high levels and activity of the enzyme were associated with higher HDL-C levels (P values for trend: 0.008, 0.020, 0.042, and 0.037 for L55M, Q192R, -107C/T, and -907G/C, respectively). In addition to the PON1 genotype, there was also a positive correlation between PON1 levels and activity and HDL-C (PON1 levels: r = 0.37, P < 0.001; paraoxonase activity: r = 0.23, P = 0.01; diazoxonase activity: r = 0.29, P < 0.001; arylesterase activity: r = 0.19, P = 0.03). Our observations support the hypothesis that both PON1 levels and activity preserve HDL-C in plasma.     

Differential hydrolysis of homocysteine thiolactone by purified human serum (192)Q and (192)R PON1 isoenzymes

Human serum paraoxonase 1 (PON1) is a HDL-associated enzyme that catalyzes the hydrolysis of a variety of aromatic carboxylic acid esters and several organophosphates. Recently it has been suggested that a physiological substrate of serum PON1 is homocysteine thiolactone which is a putative risk factor in atherosclerosis. In this study, human (192)Q and (192)R PON1 isoenzymes were purified from the respective phenotype human serum, using a protocol consisting of ammonium sulfate precipitation and four chromatography steps: gel filtration, ion-exchange, non-specific affinity, and a second ion-exchange. Using paraoxon as substrate, overall purification fold was found as 742 for (192)R PON1 and 590 for (192)Q PON1. The final purified enzymes were shown as single protein bands close to 45kDa on SDS-PAGE and confirmed by Western blot. Substrate kinetics were studied with phenyl acetate, paraoxon and homocysteine thiolactone. Both PON1 isoenzymes showed mixed type inhibition with phenyl acetate. K(m) values of (192)Q and (192)R PON1 for homocysteine thiolactone were 23.5mM and 22.6mM respectively. For (192)R PON1, the V(max) was 2.5-fold and k(cat)/K(m) was 2.6-fold higher than those for (192)Q PON1 when homocysteine thiolactone is used as substrate. The present data suggest that defining (192)Q and (192)R PON1 isoforms could be a good predictor and prognostic marker in the cardiovascular risk assessment.     

Serum paraoxonase level and paraoxonase polymorphism in patients with acromegaly

Background: Acromegalic patients have increased cardiometabolic risk factors due to an elevation of growth hormone (GH) levels. Human serum paraoxonase (PON), a high-density lipoprotein (HDL)-related enzyme, is one of the major bioscavengers and decreases the oxidation of low-density lipoprotein (LDL), a key regulator in the pathogenesis of atherosclerosis. In this study, we investigated a potential relationship between serum PON levels or PON polymorphisms and acromegaly.

Methods: A total of 48 acromegalic patients and 44 healthy controls were included in this study. Serum GH levels, insulin-like growth factor-1 levels and lipid profiles were measured. Serum PON levels, as well as PON 1 L55M and Q192R gene polymorphisms, were examined.

Results: No significant differences were found in terms of age, gender, presence of diabetes, serum LDL cholesterol (LDL-C), HDL-C, or triglyceride levels between the case and control groups (P?>?0.05). A statistically significant difference was found in serum PON levels between the cases and controls (P?=?0.007). The median serum PON level was 101?±?63.36?U/l in the case group and 63?±?60.50?U/l in the control group. There was a significant correlation between serum PON levels and IGF-1 levels (P?=?0.004, r?=?0.319); however, no significant differences were found in PON1 L55M and PON Q192R polymorphisms between the patients and controls (P?=?0.607 and P?=?0.308, respectively). In addition, no significant differences were found in serum PON levels in acromegalic patients who were and were not in remission (P?=?0.385), nor between those with PON1 L55M and Q192R polymorphisms (P?=?0.161 and P?=?0.336, respectively).

Conclusions: Elevated serum PON levels were detected in acromegalic patients, independently of their remission status. This suggests protective effects for cardiometabolic risk parameters.     

Paraoxonase 1 R/Q alleles are associated with differential accumulation of saturated versus 20:5n3 fatty acid in human adipose tissue

Serum paraoxonase 1 (PON1) function has been associated with human cardiovascular disease. The projected mechanism postulates interaction of PON1 with lipoproteins and insulin signaling resulting in alterations in lipid homeostasis. Recently, PON2 was shown to directly regulate triglyceride accumulation in macrophages and PON1 was detected in the interstitial space of adipocytes. The aims of the present study were a) to examine the relationship of the PON1 function with serum parameters related to lipid homeostasis, and b) to examine a possible role of PON1 in the regulation of lipid composition in the human adipose tissue. Two important genetic variations with functional impact on PON1 activity in humans are the Q192R and the L55M. The present study evaluated the impact of the Q192R and the L55M polymorphisms in a cross-section of the population on the island of Crete, as regards to PON1 activity, plasma lipids/lipoproteins, parameters of the metabolic syndrome, and the fatty acid composition of the adipose tissue. We detected a significant association of the polymorphisms with blood pressure, fasting blood glucose, triglycerides, apolipoprotein B, serum iron, and homocysteine. Furthermore, a novel function is suggested for PON1 on the fatty acid composition in the adipose tissue through the positive association of the R allele with saturated fatty acid and of the Q allele with 20:5n3 fatty acid deposition.     

A preliminary study of human paraoxonase and PON 1 L/M55–PON 1 Q/R 192 polymorphisms in Turkish patients with coronary artery disease

Paraoxonase 1 (PON 1) is a high‐density lipoprotein (HDL)‐associated enzyme with antioxidant function protecting low‐density lipoprotein (LDL) from oxidation. PON 1 has two amino acid polymorphisms in coding region; L/M 55 and Q/R 192. These polymorphisms modulate paraoxonase activity of the enzyme. PON 1 activity decreases in coronary artery disease (CAD). In the present study, distribution of PON 1 L/M 55 and Q/R 192 polymorphisms and the effect of these polymorphisms on the activities of PON 1, and on the severity of CAD in 277 CAD (+) patient and 92 CAD (?) subjects were examined. PON 1 L/M 55 and Q/R 192 genotypes were determined by PCR, RFLP and agarose gel electrophoresis techniques. Genotype distributions and allele frequencies for PON 1 Q/R 192 polymorphism were not significantly different between controls and CAD (+) patient group (p > 0.05), but in genotype and allele distribution of PON 1 L/M55 polymorphism, there was significantly difference among groups (p < 0.05). Genotype distributions for both polymorphisms were not significantly different between subgroups of single‐vessel disease (SVD), double‐vessel disease (DVD) and triple‐vessel disease (TVD). Serum PON 1 activity was lower in CAD (+) group than in controls and this was also statistically significant (p < 0.001). In both groups, the highest PON activities were detected in LL and RR genotypes. In summary, our results suggest that there is an association between the PON 1 L/M 55 polymorphism of paraoxonase and CAD in Turkish patients but not with PON 1 Q/R 192 polymorphism. However, it is hard to correlate these polymorphisms and severity of CAD. Copyright © 2009 John Wiley & Sons, Ltd.     

The 192R/Q polymorphs of serum paraoxonase PON1 differ in HDL binding, lipolactonase stimulation, and cholesterol efflux

Serum paraoxonase (PON1) is a HDL-associated enzyme exhibiting potentially antiatherogenic properties. Here, we examined the common PON1-192R/Q human polymorphism. Despite numerous studies, the effect of this polymorphism on the antiatherogenic potential of PON1 is yet unresolved. Our structural model suggests that amino acid 192 constitutes part of the HDL-anchoring surface and active site of PON1. Based on our findings that PON1 is an interfacially activated lipolactonase that selectively binds HDL carrying apolipoprotein A-I (apoA-I) and is thereby greatly stabilized and catalytically activated, we examined the interaction of the PON1-192 isozymes with reconstituted HDL-apoA-I particles. We found that PON1 position 192 is indeed involved in HDL binding. The PON1-192Q binds HDL with a 3-fold lower affinity than the R isozyme and consequently exhibits significantly reduced stability, lipolactonase activity, and macrophage cholesterol efflux. We also observed the lower affinity and stability of the 192Q versus the 192R isozyme in sera of individuals belonging to the corresponding genotypes. The observed differences in the properties of PON1-192R/Q isozymes provide a basis for further analysis of the contribution of the 192R/Q polymorphism to the susceptibility to atherosclerosis, although other factors, such as the overall levels of PON1, may play a more significant role.     

Differential effects of paraoxonase 1 (PON1) polymorphisms on cancer risk: evidence from 25 published studies

Paraoxonase is an HDL-associated enzyme that plays a preventive role against oxidative stress, which is thought to contribute to cancer development. PON1 activity varies widely among individuals, which is in part related to two common nonsynonymous polymorphisms in the PON1 gene (Q192R and L55M). The polymorphisms in PON1 have been implicated in cancer risk. However, results from the studies to date have been conflicting. To clarify the association, a meta-analysis was performed for 7,073 cases and 9,520 controls from 25 published case–control studies. Odds ratios (ORs) with 95% confidence intervals (CIs) were used to assess the strength of the association. Significant associations between PON1-L55M but not Q192R polymorphism and total cancer were observed from all the comparisons. In stratified analyses, PON1-55M allele was a risk factor for breast cancer. Similarly, increased risk was observed for prostate cancer (OR = 1.18, 95% CI: 1.01–1.36, P _{heterogeneity} = 0.260) and Caucasian population (OR = 1.18, 95% CI: 1.02–1.38, P _{heterogeneity} = 0.1) of the LM genotype, compared with the LL genotype. For PON1-Q192R polymorphism, PON1-192R allele was a decreased risk factor for cancer in the Asian group (RR vs QQ: OR = 0.61, 95% CI: 0.38–0.98, P _{heterogeneity} = 0.268; QR vs QQ: OR = 0.71, 95% CI: 0.52–0.96, P _{heterogeneity} = 0.130; RR + QR vs QQ: OR = 0.71, 95% CI: 0.53–0.95, P _{heterogeneity} = 0.135). Although some modest bias could not be eliminated, this meta-analysis suggests that the PON1-55M allele is a risk factor for the development of cancer, in particular for breast cancer. Future studies with larger sample sizes are warranted to further evaluate these associations.     

Paraoxonase 1 gene polymorphisms, paraoxonase/arylesterase activities and oxidized low-density lipoprotein levels in patients with migraine

The vascular endothelial dysfunction has been implicated in the pathogenesis of migraine. Oxidized low‐density lipoprotein (ox‐LDL) may impair endothelial function. Paraoxonase‐1 (PON‐1) prevents oxidative modification of LDL cholesterol (LDL‐C). So we investigated serum PON‐1 and arylesterase (ARE) activities, PON‐1 55 L/M and 192Q/R polymorphisms and the serum lipid profile in patients with migraine. Biochemical parameters and PON‐1 polymorphism analyses were assessed in 104 patients with migraine and 86 healthy subjects. Ox‐LDL was detected by ELISA, and polymorphisms were determined using PCR–restriction fragment length polymorphism analysis. Patients with migraine had lower PON‐1 and ARE activities (p < 0·001, for both) and higher ox‐LDL and LDL‐C levels (p < 0·001, for both) and ox‐LDL: LDL‐C ratio (p < 0·005) than the controls. The genotype distribution and the allele frequencies for PON‐1 55 L/M and 192Q/R polymorphisms were not different among the study populations. The results of our current study indicate that migrainous patients have decreased serum PON‐1 and ARE activities and increased serum ox‐LDL levels, which may have a clinical importance in the treatment of migraine. Copyright © 2011 John Wiley & Sons, Ltd.     

Nippostrongylus brasiliensis: Infection decreases plasma butyrylcholinesterase activity in rats

Plasma butyrylcholinesterase (BChE) hydrolyzes ester-containing compounds such as succinylcholine, as well as acting as a scavenger against neurotoxic organophosphates (OPs). We previously found that Nippostrongylus brasiliensis infection makes rats more susceptible to OP toxicity by decreasing serum paraoxonase-1 (PON1) activity. In the present study, we examined the effects of N.brasiliensis infection on acetylcholinesterase (AChE) activity in plasma, red blood cells (RBCs), brain and diaphragm, as well as serum PON1 activity, in rats at day 7 after infection. N.brasiliensis infection significantly decreased plasma BChE and PON1 activities without significantly altering AChE activity in RBCs, brain and diaphragm. These results provide further insight into the unusual deleterious effects of intestinal nematode infections on body homeostasis.     

The determination of Q192R polymorphism of paraoxonase 1 by using non-toxic substrate p-nitrophenylacetate

CONTEXT:

The human serum paraoxonase 1 (PON1) is calcium-dependent esterase and associates with the high density serum lipoproteins. PON1 plays a major role in oxidation of high density lipoprotein and low density lipoprotein and prevention of atherogenesis in coronary heart disease. PON1Q and R allele hydrolyses number of substrates like paraoxon (PO) (diethyl p-nitrophenyl phosphate) and phenylacetate.

AIMS:

The aim of the study is to the determination of Q192R polymorphism of PON1 by using non-toxic substrate p-nitrophenylacetate and compares it with the phenotype determined by using PO as substrate.

MATERIALS AND METHODS:

The study group consists of 60 healthy normal patients. Paraoxonase activity was measured using the procedure described by Eckerson (Reference method) and for phenotyping; the ratio of hydrolysis of PO in the presence of 1 M NaCl (salt-stimulated PON1, SALT) to the hydrolysis of phenylacetate (PA) is calculated. In new method (Haagen et al.) arylesterase activity measured using p-nitrophenylacetate and for phenotyping arylesterase, the ratio of inhibition of enzymatic hydrolysis of p-nitrophenylacetate (substrate) by phenyl acetate to non-inhibited hydrolysis of p-nitrophenylacetate (inhibited arylesterase activity (IA-IA₀)/non-inhibited arylesterase activity (NIA).

RESULTS:

It was found that paraoxonase activity is trimodally distributed in both the methods. There is no significant difference in the distribution of PON1 phenotypes of both reference method and new method being frequencies 0.946 and 0.376 respectively and there was no significant difference for phenotypic polymorphism for an individual by both methods (χ²= 0.15 and P = 0.9262).

CONCLUSION:

The Q192R polymorphism of PON1 by using non-toxic substrate p-nitrophenylacetate showed trimodal distribution of QQ (homozygous), QR (heterozygous), and RR (homozygous) phenotype and it is comparable with reference method. This method can be used for PON1 phenotype in different pathological and complex disease conditions.     

Paraoxonase 1 (PON1) polymorphisms, haplotypes and activity in predicting cad risk in North-West Indian Punjabis

Background

Human serum paraoxonase-1 (PON1) prevents oxidation of low density lipoprotein cholesterol (LDL-C) and hydrolyzes the oxidized form, therefore preventing the development of atherosclerosis. The polymorphisms of PON1 gene are known to affect the PON1 activity and thereby coronary artery disease (CAD) risk. As studies are lacking in North-West Indian Punjabi''s, a distinct ethnic group with high incidence of CAD, we determined PON1 activity, genotypes and haplotypes in this population and correlated them with the risk of CAD.

Methodology/Principal Findings

350 angiographically proven (≥70% stenosis) CAD patients and 300 healthy controls were investigated. PON1 activity was determined towards paraoxon (Paraoxonase; PONase) and phenylacetate (Arylesterase; AREase) substrates. In addition, genotyping was carried out by using multiplex PCR, allele specific oligonucleotide –PCR and PCR-RFLP methods and haplotyping was determined by PHASE software. The serum PONase and AREase activities were significantly lower in CAD patients as compared to the controls. All studied polymorphisms except L55M had significant effect on PONase activity. However AREase activity was not affected by them. In a logistic regression model, after adjustment for the conventional risk factors for CAD, QR (OR: 2.73 (1.57–4.72)) and RR (OR, 16.24 (6.41–41.14)) genotypes of Q192R polymorphism and GG (OR: 2.07 (1.02–4.21)) genotype of −162A/G polymorphism had significantly higher CAD risk. Haplotypes L-T-G-Q-C (OR: 3.25 (1.72–6.16)) and L-T-G-R-G (OR: 2.82 (1.01–7.80)) were also significantly associated with CAD.

Conclusions

In conclusion this study shows that CAD patients had lower PONase and AREase activities as compared to the controls. The coding Q192R polymorphism, promoter −162A/G polymorphism and L-T-G-Q-C and L-T-G-R-G haplotypes are all independently associated with CAD.     

European versus Asian differences for the associations between paraoxonase‐1 genetic polymorphisms and susceptibility to type 2 diabetes mellitus

Many studies have examined the associations between paraoxonase‐1 (PON1) genetic polymorphisms (Q192R, rs662 and L55M, rs854560) and the susceptibility to type 2 diabetes mellitus (T2DM) across different ethnic populations. However, the evidence for the associations remains inconclusive. In this study, we performed a meta‐analysis to clarify the association of the two PON1 variants with T2DM risk. We carried out a systematic search of PubMed, Embase, CNKI and Wanfang databases for studies published before June 2017. The pooled odds ratios (ORs) for the association and their corresponding 95% confidence intervals (CIs) were calculated by a random‐ or fixed‐effect model. A total of 50 eligible studies, including 34 and 16 studies were identified for the PON1 Q192R (rs662) and L55M (rs854560) polymorphism, respectively. As for the PON1 Q192R polymorphism, the 192R allele was a susceptible factor of T2DM in the South or East Asian population (OR > 1, P < 0.05) but represented a protective factor of T2DM in European population (OR = 0.66, 95% CI = 0.45–0.98) under a heterozygous genetic model. With regard to the PON1 L55M polymorphism, significant protective effects of the 55M allele on T2DM under the heterozygous (OR = 0.77, 95% CI = 0.61–0.97) and dominant (OR = 0.80, 95% CI = 0.65–0.99) genetic models were found in the European population, while no significant associations in the Asian populations under all genetic models (P > 0.05). In summary, by a comprehensive meta‐analysis, our results firmly indicated that distinct effects of PON1 genetic polymorphisms existed in the risk of T2DM across different ethnic backgrounds.