Search tips
Search criteria

Results 1-25 (808746)

Clipboard (0)

Related Articles

1.  Paraoxonase inhibits high-density lipoprotein oxidation and preserves its functions. A possible peroxidative role for paraoxonase. 
Journal of Clinical Investigation  1998;101(8):1581-1590.
HDL levels are inversely related to the risk of developing atherosclerosis. In serum, paraoxonase (PON) is associated with HDL, and was shown to inhibit LDL oxidation. Whether PON also protects HDL from oxidation is unknown, and was determined in the present study. In humans, we found serum HDL PON activity and HDL susceptibility to oxidation to be inversely correlated (r2 = 0.77, n = 15). Supplementing human HDL with purified PON inhibited copper-induced HDL oxidation in a concentration-dependent manner. Adding PON to HDL prolonged the oxidation lag phase and reduced HDL peroxide and aldehyde formation by up to 95%. This inhibitory effect was most pronounced when PON was added before oxidation initiation. When purified PON was added to whole serum, essentially all of it became HDL-associated. The PON-enriched HDL was more resistant to copper ion-induced oxidation than was control HDL. Compared with control HDL, HDL from PON-treated serum showed a 66% prolongation in the lag phase of its oxidation, and up to a 40% reduction in peroxide and aldehyde content. In contrast, in the presence of various PON inhibitors, HDL oxidation induced by either copper ions or by a free radical generating system was markedly enhanced. As PON inhibited HDL oxidation, two major functions of HDL were assessed: macrophage cholesterol efflux, and LDL protection from oxidation. Compared with oxidized untreated HDL, oxidized PON-treated HDL caused a 45% increase in cellular cholesterol efflux from J-774 A.1 macrophages. Both HDL-associated PON and purified PON were potent inhibitors of LDL oxidation. Searching for a possible mechanism for PON-induced inhibition of HDL oxidation revealed PON (2 paraoxonase U/ml)-mediated hydrolysis of lipid peroxides (by 19%) and of cholesteryl linoleate hydroperoxides (by 90%) in oxidized HDL. HDL-associated PON, as well as purified PON, were also able to substantially hydrolyze (up to 25%) hydrogen peroxide (H2O2), a major reactive oxygen species produced under oxidative stress during atherogenesis. Finally, we analyzed serum PON activity in the atherosclerotic apolipoprotein E-deficient mice during aging and development of atherosclerotic lesions. With age, serum lipid peroxidation and lesion size increased, whereas serum PON activity decreased. We thus conclude that HDL-associated PON possesses peroxidase-like activity that can contribute to the protective effect of PON against lipoprotein oxidation. The presence of PON in HDL may thus be a major contributor to the antiatherogenicity of this lipoprotein.
PMCID: PMC508738  PMID: 9541487
2.  Macrophage Paraoxonase 2 regulates calcium homeostasis and cell survival under Endoplasmic Reticulum stress conditions and is sufficient to prevent the development of aggravated atherosclerosis in Paraoxonase 2 deficiency/apoE−/− mice on a Western diet 
Molecular genetics and metabolism  2012;107(3):416-427.
Paraoxonase 2 deficiency (PON2-def) alters mitochondrial function and exacerbates the development of atherosclerosis in mice. PON2 overexpression protects against ER stress in cell culture. In this paper, we examined the role of PON2 in the unexplored link between ER stress and mitochondrial dysfunction and tested whether restoration of PON2 in macrophages is sufficient to reduce aggravated atherosclerosis in PON2-def/apoE−/− mice on a Western diet. ER stress response genes, intracellular calcium levels, and apoptotic nuclei were significantly elevated in PON2-def/apoE−/− macrophages compared to apoE−/− macrophages in response to ER stressors, but not at the basal level. In contrast, PON2-def/apoE−/− macrophages exhibited greater mitochondrial stress at the basal level, which was further worsened in response to ER stressors. There was no difference in ER stress response genes and apoptotic nuclei between apoE−/− and PON2-def/apoE−/− macrophages when pretreated with xestospongin (which blocks the release of calcium from ER) suggesting that PON2 modulates cell survival and ER stress by maintaining calcium homeostasis. Treatment with a mitochondrial calcium uptake inhibitor, RU360, attenuated ER stressor mediated mitochondrial dysfunction in PON2-def/apoE−/− macrophages. CHOP expression (ER stress marker) and apoptotic nuclei were significantly higher in aortic lesions of PON2-def/apoE−/− mice compared to apoE−/− mice fed a Western diet. Restoration of PON2 in macrophage reduced ER stress, mitochondrial dysfunction and apoptosis in response to ER stressors. Furthermore, restoration of PON2 in macrophages reduced lesional apoptosis and atherosclerosis in PON2-def/apoE−/− mice on a Western diet. Our data suggest that macrophage PON2 modulates mechanisms that link ER stress, mitochondrial dysfunction and the development of atherosclerosis.
PMCID: PMC3483415  PMID: 22864055
Paraoxonase 2; Mitochondria; Endoplasmic reticulum; Macrophages; Oxidative stress
3.  Impaired hepatic insulin signalling in PON2-deficient mice: a novel role for the PON2/apoE axis on the macrophage inflammatory response 
The Biochemical journal  2011;436(1):91-100.
Hepatic glucose metabolism is strongly influenced by oxidative stress and pro-inflammatory stimuli. PON2 (paraoxonase 2), an enzyme with undefined antioxidant properties, protects against atherosclerosis. PON2-deficient (PON2-def) mice have elevated hepatic oxidative stress coupled with an exacerbated inflammatory response from PON2-deficient macrophages. In the present paper, we demonstrate that PON2 deficiency is associated with inhibitory insulin-mediated phosphorylation of hepatic IRS-1 (insulin receptor substrate-1). Unexpectedly, we observed a marked improvement in the hepatic IRS-1 phosphorylation state in PON2-def/apoE (apolipoprotein E)−/− mice, relative to apoE−/− mice. Factors secreted from activated macrophage cultures derived from PON2-def and PON2-def/apoE−/− mice are sufficient to modulate insulin signalling in cultured hepatocytes in a manner similar to that observed in vivo. We show that the protective effect on insulin signalling in PON2-def/apoE−/− mice is directly associated with altered production of macrophage proinflammatory mediators, but not elevated intracellular oxidative stress levels. We further present evidence that modulation of the macrophage inflammatory response in PON2-def/apoE−/− mice is mediated by a shift in the balance of NO and ONOO− (peroxynitrite) formation. Our results demonstrate that PON2 plays an important role in hepatic insulin signalling and underscores the influence of macrophage-mediated inflammatory response on hepatic insulin sensitivity.
PMCID: PMC3743554  PMID: 21361875
apolipoprotein E (apoE); insulin receptor substrate-1 (IRS-1); liver; macrophage; paraoxonase; paraoxonase 2 (PON2)
4.  The Human Paraoxonase Gene Cluster As a Target in the Treatment of Atherosclerosis 
Antioxidants & Redox Signaling  2012;16(6):597-632.
The paraoxonase (PON) gene cluster contains three adjacent gene members, PON1, PON2, and PON3. Originating from the same fungus lactonase precursor, all of the three PON genes share high sequence identity and a similar β propeller protein structure. PON1 and PON3 are primarily expressed in the liver and secreted into the serum upon expression, whereas PON2 is ubiquitously expressed and remains inside the cell. Each PON member has high catalytic activity toward corresponding artificial organophosphate, and all exhibit activities to lactones. Therefore, all three members of the family are regarded as lactonases. Under physiological conditions, they act to degrade metabolites of polyunsaturated fatty acids and homocysteine (Hcy) thiolactone, among other compounds. By detoxifying both oxidized low-density lipoprotein and Hcy thiolactone, PONs protect against atherosclerosis and coronary artery diseases, as has been illustrated by many types of in vitro and in vivo experimental evidence. Clinical observations focusing on gene polymorphisms also indicate that PON1, PON2, and PON3 are protective against coronary artery disease. Many other conditions, such as diabetes, metabolic syndrome, and aging, have been shown to relate to PONs. The abundance and/or activity of PONs can be regulated by lipoproteins and their metabolites, biological macromolecules, pharmacological treatments, dietary factors, and lifestyle. In conclusion, both previous results and ongoing studies provide evidence, making the PON cluster a prospective target for the treatment of atherosclerosis. Antioxid. Redox Signal. 16, 597–632.
I. Introduction
II. The PON Family
A. Evolution of the PON genes
B. Structure of the PONs
1. Primary structure
2. Three-dimensional structure
C. Tissue and cellular distribution of PONs
D. Substrates of PONs
1. Chemical substrates
2. Physiological substrates
III. PONs Protect Against Atherosclerosis and Coronary Heart Disease by Decreasing the Toxicity of LDL
A. Atherosclerosis-related CVDs do great harm to human health
B. OxLDL is one of the most important risk factors for atherogenesis
C. PON1 contributes to the anti-atherosclerotic functions of HDL
D. PON activity decreases atherosclerosis
1. PON1 and its protection against atherosclerosis
2. PON2 and its protection against atherosclerosis
3. PON3 and atherosclerosis
4. Roles of the PON gene cluster in atherosclerosis
IV. PONs Protect Against Hcy Toxicity and Associated Atherogenesis
A. Hcy is an important risk factor in atherosclerosis
B. PONs protect against Hcy toxicity
V. PONs Polymorphisms and Coronary Artery Disease
A. PON1 polymorphisms
1. PON1 polymorphisms and PON1 activity
2. PON1 polymorphisms and plasma lipoprotein levels
3. PON1 polymorphisms and CHD
B. PON2 polymorphisms
1. PON2 polymorphisms
2. PON2 polymorphisms and plasma lipoprotein levels
3. PON2 polymorphisms and CHD
VI. Other Diseases Related to PONs
A. PONs' relations to diabetes mellitus and corresponding complications
B. PONs' associations with metabolic syndrome and obesity
C. PONs and aging
D. Other diseases related to PONs
VII. Regulation of PONs
A. Transcriptional regulation of PONs
B. Lipoproteins and their metabolic products regulate PONs expression
C. Biological macromolecules modulate PONs expression
D. Pharmacological modulators of PONs
1. Statins
2. Fibrates
3. Other cardiovascular drugs
4. Diabetic drugs
5. Other drugs
E. Dietary factors and lifestyle factors
1. Vitamin C and vitamin E
2. Natural plant extracts
3. Dietary polyphenol compounds
4. Dietary flavonoids
5. Dietary lipids
6. Alcohol
7. Cigarettes
8. Fasting
VIII. PONs Mimetics
IX. Conclusions and Future Directions
PMCID: PMC3270057  PMID: 21867409
5.  Myeloperoxidase, paraoxonase-1, and HDL form a functional ternary complex 
The Journal of Clinical Investigation  2013;123(9):3815-3828.
Myeloperoxidase (MPO) and paraoxonase 1 (PON1) are high-density lipoprotein–associated (HDL-associated) proteins mechanistically linked to inflammation, oxidant stress, and atherosclerosis. MPO is a source of ROS during inflammation and can oxidize apolipoprotein A1 (APOA1) of HDL, impairing its atheroprotective functions. In contrast, PON1 fosters systemic antioxidant effects and promotes some of the atheroprotective properties attributed to HDL. Here, we demonstrate that MPO, PON1, and HDL bind to one another, forming a ternary complex, wherein PON1 partially inhibits MPO activity, while MPO inactivates PON1. MPO oxidizes PON1 on tyrosine 71 (Tyr71), a modified residue found in human atheroma that is critical for HDL binding and PON1 function. Acute inflammation model studies with transgenic and knockout mice for either PON1 or MPO confirmed that MPO and PON1 reciprocally modulate each other’s function in vivo. Further structure and function studies identified critical contact sites between APOA1 within HDL, PON1, and MPO, and proteomics studies of HDL recovered from acute coronary syndrome (ACS) subjects revealed enhanced chlorotyrosine content, site-specific PON1 methionine oxidation, and reduced PON1 activity. HDL thus serves as a scaffold upon which MPO and PON1 interact during inflammation, whereupon PON1 binding partially inhibits MPO activity, and MPO promotes site-specific oxidative modification and impairment of PON1 and APOA1 function.
PMCID: PMC3754253  PMID: 23908111
6.  Repeated Developmental Exposure of Mice to Chlorpyrifos Oxon Is Associated with Paraoxonase 1 (PON1)-Modulated Effects on Cerebellar Gene Expression 
Toxicological Sciences  2011;123(1):155-169.
Microarray analysis was used to examine effects of repeated postnatal exposure to chlorpyrifos oxon (CPO) on gene expression in the cerebellum of genetically modified mice. The high-density lipoprotein-associated enzyme paraoxonase 1 (PON1) plays a significant role in the detoxication of CPO, which is present in exposures and generated from chlorpyrifos (CPF) in vivo following exposure. Two factors are important in modulating toxicity of CPO, the Q192R PON1 polymorphism and PON1 plasma level, which is low at birth and increases throughout postnatal development. Mice used in these studies included wild type (PON1+/+), PON1 knockout (PON1−/−), and two transgenic lines (tgHuPON1Q192, tgHuPON1R192) expressing either human PON1Q192 or PON1R192 on the PON1−/− background. PON1R192 hydrolyzes CPO more efficiently than PON1Q192. All four genotypes exposed to CPO (0.35 or 0.50 mg/kg/day) daily from postnatal day (PND) 4 to PND 21 showed significant differences in gene expression on PND 22 compared with controls. Pathway analysis and Gene Set Analysis revealed multiple pathways and gene sets significantly affected by CPO exposure, including genes involved in mitochondrial dysfunction, oxidative stress, neurotransmission, and nervous system development. Comparison between genotypes revealed specific genes, gene sets, and pathways differentially affected between tgHuPON1Q192 and tgHuPON1R192 mice and between PON1−/− and PON1+/+ mice following CPO exposure. Repeated CPO exposure also resulted in a dose-related decrease in brain acetylcholinesterase activity during postnatal development in PON1−/− and tgHuPON1Q192 mice but not in PON1+/+ or tgHuPON1R192 mice. These findings indicate that PON1 status plays a critical role in modulating the effects of neonatal CPO exposure in the developing brain.
PMCID: PMC3164442  PMID: 21673326
chlorpyrifos; chlorpyrifos oxon; paraoxonase; organophosphorus insecticides; microarrays; neurotoxicity
7.  Role of PON2 in innate immune response in an acute infection model 
Molecular genetics and metabolism  2013;110(3):362-370.
N-(3-oxododecanoyl)-L-homoserine lactone (3OC12-HSL) is a quorum-sensing molecule produced by gram-negative microbial pathogens such as Pseudomonas aeruginosa (PAO1). 3OC12-HSL is involved in the regulation of bacterial virulence factors and also alters the function of the host immune cells. Others and we have previously shown that paraoxonase 2 (PON2), a member of the paraoxonase gene family expressed in immune cells, hydrolyzes 3OC12-HSL. In this study, we examined i) whether macrophage PON2 participates in 3OC12-HSL hydrolysis, ii) the effect of PON2 deficiency in acute PAO1 infection in mice and iii) the effect of 3OC12-HSL on PON2 deficient (PON2-def) macrophages. When compared to wild type macrophages, both intact cells and membrane-enriched protein lysates obtained from PON2-def macrophages show a marked impairment in their ability to hydrolyze 3OC12-HSL. PON2 expression (message and protein) is not altered in response to 3OC12-HSL in macrophages. 3OC12-HSL treated PON2-def macrophages showed i) an increase in ER stress and oxidative stress, ii) defective phosphatidylinositol 3-kinase (PI3 kinase)/AKT activation, and iii) reduced phagocytosis function. Moreover, the nitration to phosphorylation ratio of Tyr458 in p85 protein, the regulatory subunit of PI3-kinase that has been correlated with the phagocytosis function of macrophages, was increased in PON2-def macrophages. Antioxidant treatment reversed the effects of PON2 deficiency in macrophage phagocytosis function. Furthermore, following administration of 1.6×107CFU of PAO1, bacterial clearance was significantly reduced in the lungs (5.7 fold), liver (2.5 fold), and spleen (14.8 fold) of PON2-def mice when compared to wild type mice. Our results suggest that PON2 plays an important role in innate immune defense against PAO1 infection.
PMCID: PMC3800229  PMID: 23911207
Paraoxonase 2; Pseudomonas aeruginosa; Quorum sensing; Mitochondrial oxidative stress; Endoplasmic reticulum stress
8.  Protectors or Traitors: The Roles of PON2 and PON3 in Atherosclerosis and Cancer 
Journal of Lipids  2012;2012:342806.
Cancer and atherosclerosis are major causes of death in western societies. Deregulated cell death is common to both diseases, with significant contribution of inflammatory processes and oxidative stress. These two form a vicious cycle and regulate cell death pathways in either direction. This raises interest in antioxidative systems. The human enzymes paraoxonase-2 (PON2) and PON3 are intracellular enzymes with established antioxidative effects and protective functions against atherosclerosis. Underlying molecular mechanisms, however, remained elusive until recently. Novel findings revealed that both enzymes locate to mitochondrial membranes where they interact with coenzyme Q10 and diminish oxidative stress. As a result, ROS-triggered mitochondrial apoptosis and cell death are reduced. From a cardiovascular standpoint, this is beneficial given that enhanced loss of vascular cells and macrophage death forms the basis for atherosclerotic plaque development. However, the same function has now been shown to raise chemotherapeutic resistance in several cancer cells. Intriguingly, PON2 as well as PON3 are frequently found upregulated in tumor samples. Here we review studies reporting PON2/PON3 deregulations in cancer, summarize most recent findings on their anti-oxidative and antiapoptotic mechanisms, and discuss how this could be used in putative future therapies to target atherosclerosis and cancer.
PMCID: PMC3361228  PMID: 22666600
9.  PON3 is upregulated in cancer tissues and protects against mitochondrial superoxide-mediated cell death 
Cell Death and Differentiation  2012;19(9):1549-1560.
To achieve malignancy, cancer cells convert numerous signaling pathways, with evasion from cell death being a characteristic hallmark. The cell death machinery represents an anti-cancer target demanding constant identification of tumor-specific signaling molecules. Control of mitochondrial radical formation, particularly superoxide interconnects cell death signals with appropriate mechanistic execution. Superoxide is potentially damaging, but also triggers mitochondrial cytochrome c release. While paraoxonase (PON) enzymes are known to protect against cardiovascular diseases, recent data revealed that PON2 attenuated mitochondrial radical formation and execution of cell death. Another family member, PON3, is poorly investigated. Using various cell culture systems and knockout mice, here we addressed its potential role in cancer. PON3 is found overexpressed in various human tumors and diminishes mitochondrial superoxide formation. It directly interacts with coenzyme Q10 and presumably acts by sequestering ubisemiquinone, leading to enhanced cell death resistance. Localized to the endoplasmic reticulum (ER) and mitochondria, PON3 abrogates apoptosis in response to DNA damage or intrinsic but not extrinsic stimulation. Moreover, PON3 impaired ER stress-induced apoptotic MAPK signaling and CHOP induction. Therefore, our study reveals the mechanism underlying PON3's anti-oxidative effect and demonstrates a previously unanticipated function in tumor cell development. We suggest PONs represent a novel class of enzymes crucially controlling mitochondrial radical generation and cell death.
PMCID: PMC3422478  PMID: 22441669
paraoxonase; apoptosis; endoplasmic reticulum stress; mitochondria; oxidative stress; cancer
10.  Regulation of Hepatic Paraoxonase-1 Expression 
Journal of Lipids  2012;2012:684010.
Serum paraoxonase-1 (PON1) is a member of the paraoxonases family (PON1, PON2, and PON3). PON1 is synthesized and secreted by the liver, and in circulation it is associated with HDL. PON1 has antioxidative properties, which are associated with the enzyme's capability to decrease oxidative stress in atherosclerotic lesions and to attenuate atherosclerosis development. Epidemiological evidence demonstrates that low PON1 activity is associated with increased risk of cardiovascular events and cardiovascular disease and is an independent risk factor for coronary artery disease. Therefore, pharmacological modulation of PON1 activity or PON1 gene expression could constitute a useful approach for preventing atherosclerosis. A primary determinant of serum PON1 levels is the availability of the enzyme for release by the liver, the principal site of PON1 production. Together with the enzyme secretion rate, enzymatic turnover, and protein stability, the level of PON1 gene expression is a major determinant of PON1 status. This paper summarizes recent progress in understanding the regulation of PON1 expression in hepatocytes.
PMCID: PMC3324161  PMID: 22548179
11.  Decreased Obesity and Atherosclerosis in Human Paraoxonase 3 Transgenic Mice 
Circulation research  2007;100(8):1200-1207.
Paraoxonase 3 (PON3) is a member of the PON family, which includes PON1, PON2, and PON3. Recently, PON3 was shown to prevent the oxidation of low-density lipoprotein in vitro. To test the role of PON3 in atherosclerosis and related traits, 2 independent lines of human PON3 transgenic (Tg) mice on the C57BL/6J (B6) background were constructed. Human PON3 mRNA was detected in various tissues, including liver, lung, kidney, brain, adipose, and aorta, of both lines of Tg mice. The human PON3 mRNA levels in the livers of PON3 Tg mice were 4- to 7-fold higher as compared with the endogenous mouse Pon3 mRNA levels. Human PON3 protein and activity were detected in the livers of Tg mice as well. No significant differences in plasma total, high-density lipoprotein, and very-low-density lipoprotein/low-density lipoprotein cholesterol and triglyceride and glucose levels were observed between the PON3 Tg and non-Tg mice. Interestingly, atherosclerotic lesion areas were significantly smaller in both lines of male PON3 Tg mice as compared with the male non-Tg littermates on B6 background fed an atherogenic diet. When bred onto the low-density lipoprotein receptor knockout mouse background, the male PON3 Tg mice also exhibited decreased atherosclerotic lesion areas and decreased expression of monocyte chemoattractant protein-1 in the aorta as compared with the male non-Tg littermates. In addition, decreased adiposity and lower circulating leptin levels were observed in both lines of male PON3 Tg mice as compared with the male non-Tg mice. In an F2 cross, adipose Pon3 mRNA levels inversely correlated with adiposity and related traits. Our study demonstrates that elevated PON3 expression significantly decreases atherosclerotic lesion formation and adiposity in male mice. PON3 may play an important role in protection against obesity and atherosclerosis.
PMCID: PMC3740095  PMID: 17379834
atherosclerosis; obesity; genetics
12.  Temporal and Tissue-Specific Patterns of Pon3 Expression in Mouse: In situ Hybridization Analysis 
PON3 is a member of the paraoxonase gene family that includes PON1 and PON2. For example, PON3 and PON1 share approximately 60% identity at the amino acid level. Recent studies have demonstrated that PON3 is present in human and rabbit HDL but not in mouse HDL. Mouse PON3 appears to be cell-associated and is expressed in a wide range of tissues such as liver, adipose, macrophage, and the artery wall. In vitro studies have shown that PON3 can prevent LDL oxidation and destroy bacterial quorum-sensing molecules. Previous studies also showed that human PON3 transgenic mice were protected from obesity and atherosclerosis in both the C57BL/6J wild-type and LDLR knockout genetic background. Administration of adenovirus expressing the human PON3 gene into apoE −/− mice also decreased atherosclerotic lesion formation. In order to further understand the functions of PON3 in physiology and disease, we performed in situ hybridization analysis to examine Pon3 gene expression patterns in newborn and adult mice, in various tissues, including atherosclerotic lesions of apoE −/− mice. Our results show relatively high levels of Pon3 mRNA labeling in the adrenal gland, submaxillary gland, lung, liver, adipose, pancreas, large intestine, and other tissues of newborn mice. In the adult mouse, Pon3 mRNA levels were much lower in the corresponding tissues as mentioned above for the newborn mouse. Sections of the aortic root from the hearts of both wild-type and apoE −/− mice displayed moderate levels of Pon3 mRNA labeling. Pon3 mRNA was also detected in the atherosclerotic lesion areas at the aortic root of apoE −/− hearts. Our data revealed that mouse Pon3 is expressed in a wide range of tissues, and that its expression is temporally controlled.
PMCID: PMC3018859  PMID: 20221872
PON3; In situ hybridization; Developmental regulation; Mouse
13.  Beyond reduction of atherosclerosis: PON2 provides apoptosis resistance and stabilizes tumor cells 
Cell Death & Disease  2011;2(1):e112-.
Major contributors to atherosclerosis are oxidative damage and endoplasmic reticulum (ER) stress-induced apoptosis; both of which can be diminished by the anti-oxidative protein paraoxonase-2 (PON2). ER stress is also relevant to cancer and associated with anti-cancer treatment resistance. Hence, we addressed, for the first time, whether PON2 contributes to tumorigenesis and apoptotic escape. Intriguingly, we found that several human tumors upregulated PON2 and such overexpression provided resistance to different chemotherapeutics (imatinib, doxorubicine, staurosporine, or actinomycin) in cell culture models. This was reversed after PON2 knock-down. Remarkably, just deficiency of PON2 caused apoptosis of selective tumor cells per se, demonstrating a previously unanticipated oncogenic function. We found a dual mechanistic role. During ER stress, high PON2 levels lowered redox-triggered induction of pro-apoptotic CHOP particularly via the JNK pathway, which prevented mitochondrial cell death signaling. Apart from CHOP, PON2 also diminished intrinsic apoptosis as it prevented mitochondrial superoxide formation, cardiolipin peroxidation, cytochrome c release, and caspase activation. Ligand-stimulated apoptosis by TRAIL or TNFα remained unchanged. Finally, PON2 knock-down caused vast reactive oxygen species formation and stimulated JNK-triggered CHOP expression, but inhibition of JNK signaling did not prevent cell death, demonstrating the pleiotropic, dominating anti-oxidative effect of PON2. Therefore, targeting redox balance is powerful to induce selective tumor cell death and proposes PON2 as new putative anti-tumor candidate.
PMCID: PMC3077287  PMID: 21368884
paraoxonase; apoptosis; endoplasmic reticulum stress; mitochondria; oxidative stress; CHOP
14.  The Role of ‘Paraoxonase-1 Activity’ as an Antioxidant in Coronary Artery Diseases 
Background and Objectives: A majority of the Coronary Artery Diseases (CAD) result from complications of atherosclerosis. There is a growing body of evidence which has revealed that the reduced activity of the HDL-associated enzyme, paraoxonase1 (PON1), is predictive of vascular disease in humans, which include the results from prospective studies. The mechanisms by which PON1 activity influences risk of vascular disease continue to be evaluated. It is generally thought that PON1 contributes to the antioxidant, and thus, to the antiatherogenic properties of High Density Lipoproteins (HDL). Depleted antioxidant levels could be a risk factor for coronary artery disease. Hence, this study was done to evaluate PON1, as antioxidant, in CAD patients.
Methods: This study was done to determine serum levels of PON1 activity in 50 controls and in 60 clinically and ECG proven CAD cases and to compare PON1 activity with total cholesterol and triglycerides.
Results: Serum levels of PON1 activity (p<0.001) were significantly lower in CAD cases than in controls. Serum total cholesterol (p< 0.001) and triglyceride (p< 0.001) levels were significantly higher in CAD cases than in controls. There was a negative correlation between PON 1 activity and total cholesterol and triglycerides. The negative correlation between PON1 activity and total cholesterol was significant (p<0.05).
Interpretation and Conclusion: From our present study, we can conclude that PON1 can exert a protective effect on HDL by preventing its oxidative damage. Further, a decreased PON 1 activity may be a risk factor for CAD, which is likely to be explained by derangement of PON 1 activity towards lipid peroxidation. This study suggested that serum antioxidant activity of PON1 was an important factor which provided protection from oxidative stress and lipid peroxidation in CAD. Thus, evaluating the effects of PON 1 for CAD patients may be promising in the treatment and prognosis of CAD.
PMCID: PMC3749616  PMID: 23998046
Antioxidants; Coronary artery disease; Oxidative stress; Paraoxonase1
15.  Expression of Human Paraoxonase 1 Decreases Superoxide Levels and Alters Bacterial Colonization in the Gut of Drosophila melanogaster 
PLoS ONE  2012;7(8):e43777.
Paraoxonases (PON) are a family of proteins (PON1, 2 and 3) with multiple enzymatic activities. PON1 interferes with homoserine lactone-mediated quorum sensing in bacteria and with reactive oxygen species (ROS) in humans and mice. PON1 gene mutations have been linked to multiple traits, including aging, and diseases of the cardiovascular, nervous and gastrointestinal system. The overlapping enzymatic activities in the PON family members and high linkage disequilibrium rates within their polymorphisms confound animal and human studies of PON1 function. In contrast, arthropods such as Drosophila melanogaster have no PON homologs, resulting in an ideal model to study interactions between PON genotype and host phenotypes. We hypothesized that expression of PON1 in D. melanogaster would alter ROS. We found that PON1 alters expression of multiple oxidative stress genes and decreases superoxide anion levels in normal and germ-free D. melanogaster. We also found differences in the composition of the gut microbiota, with a remarkable increase in levels of Lactobacillus plantarum and associated changes in expression of antimicrobial and cuticle-related genes. PON1 expression directly decreased superoxide anion levels and altered bacterial colonization of the gut and its gene expression profile, highlighting the complex nature of the interaction between host genotype and gut microbiota. We speculate that the interaction between some genotypes and human diseases may be mediated by the presence of certain gut bacteria that can induce specific immune responses in the gut and other host tissues.
PMCID: PMC3431398  PMID: 22952763
16.  Paraoxonase 2 decreases renal reactive oxygen species production, lowers blood pressure, and mediates dopamine D2 receptor-induced inhibition of NADPH oxidase 
Free radical biology & medicine  2012;53(3):437-446.
The dopamine D2 receptor (D2R) regulates renal reactive oxygen species (ROS) production and impaired D2R function results in ROS-dependent hypertension. Paraoxonase 2 (PON2), which belongs to the paraoxonase gene family, is expressed in various tissues, acting to protect against cellular oxidative stress. We hypothesized that PON2 may be involved in preventing excessive renal ROS production and thus may contribute to maintenance of normal blood pressure. Moreover, the D2R may decrease ROS production, in part, through regulation of PON2.
D2R co-localized with PON2 in the brush border of mouse renal proximal tubules. Renal PON2 protein was decreased (-33%±6%) in D2-/- relative to D2+/+ mice. The renal subcapsular infusion of PON2 siRNA decreased PON2 protein expression (-55%), increased renal oxidative stress (2.2-fold), associated with increased renal NADPH oxidase expression (Nox1: 1.9-fold; Nox2: 2.9-fold; and Nox4: 1.6-fold) and activity (1.9-fold), and elevated arterial blood pressure (systolic: 134±5 vs. 93±6 mmHg; diastolic: 97±4 vs. 65±7 mmHg; mean: 113±4 vs. 75±7 mmHg). To determine the relevance of the PON2 and D2R interaction in humans, we studied human renal proximal tubule cells. Both D2R and PON2 were found in non-lipid and lipid rafts and physically interacted with each other. Treatment of these cells with the D2R/D3R agonist quinpirole (1μM, 24h) decreased ROS production (-35%±6%), associated with decreased NADPH oxidase activity (-32%±3%) and expression of Nox2 (-41%±7%) and Nox4 (-47%±8%) protein, and increased expression of PON2 mRNA (2.1-fold) and protein (1.6-fold) at 24h. Silencing PON2 (siRNA, 10nM, 48 h) not only partially prevented the quinpiroleinduced decrease in ROS production by 36%, but also increased basal ROS production (1.3-fold) which was associated with an increase in NADPH oxidase activity (1.4-fold) and expression of Nox2 (2.1-fold) and Nox4 (1.8-fold) protein. Inhibition of NADPH oxidase with diphenylene iodonium (10 μM/30 min) inhibited the increase in ROS production caused by PON2 silencing.
Our results suggest that renal PON2 is involved in the inhibition of renal NADPH oxidase activity and ROS production and contributes to the maintenance of normal blood pressure. PON2 is positively regulated by D2R and may, in part, mediate the inhibitory effect of renal D2R on NADPH oxidase activity and ROS production.
PMCID: PMC3408834  PMID: 22634053
Paraoxonase 2; Dopamine D2 receptor; Reactive oxygen species; NADPH oxidase; Hypertension
17.  Mildly oxidized LDL induces an increased apolipoprotein J/paraoxonase ratio. 
Journal of Clinical Investigation  1997;99(8):2005-2019.
We have examined the effects of mildly oxidized LDL and atherosclerosis on the levels of two proteins associated with HDL; apolipoprotein J (apoJ), and paraoxonase (PON). On an atherogenic diet, PON activity decreased by 52%, and apoJ levels increased 2.8-fold in fatty streak susceptible mice, C57BL/6J (BL/6), but not in fatty streak resistant mice, C3H/HeJ (C3H). Plasma PON activity was also significantly decreased, and apoJ levels were markedly increased in apolipoprotein E knockout mice on the chow diet, resulting in a 9.2-fold increase in the apoJ/PON ratio as compared to controls. Furthermore, a dramatic increase in the apoJ/PON ratio (over 100-fold) was observed in LDL receptor knockout mice when they were fed a 0.15%-cholesterol-enriched diet. Injection of mildly oxidized LDL (but not native LDL) into BL/6 mice (but not in C3H mice) on a chow diet resulted in a 59% decrease in PON activity (P < 0.01) and a 3.6-fold increase in apoJ levels (P < 0.01). When an acute phase reaction was induced in rabbits, or the rabbits were placed on an atherogenic diet, hepatic mRNA for apoJ was increased by 2.7-fold and 2.8-fold, respectively. Treatment of HepG2 cells in culture with mildly oxidized LDL (but not native LDL) resulted in reduced mRNA levels for PON (3.0-fold decrease) and increased mRNA levels for apoJ (2.0-fold increase). In normolipidemic patients with angiographically documented coronary artery disease who did not have diabetes and were not on lipid-lowering medication (n = 14), the total cholesterol/HDL cholesterol ratio was 3.1+/-0.9 as compared to 2.9+/-0.4 in the controls (n = 19). This difference was not statistically significant. In contrast, the apoJ/PON ratio was 3.0+/-0.4 in the patients compared to 0.72+/-0.2 in the controls (P < 0.009). In a subset of these normolipidemic patients (n = 5), the PON activity was low (48+/-6.6 versus 98+/-17 U/ml for controls; P < 0.009), despite similar normal HDL levels, and the HDL from these patients failed to protect against LDL oxidation in co-cultures of human artery wall cells. We conclude that: (a) mildly oxidized LDL can induce an increased apoJ/PON ratio, and (b) the apoJ/PON ratio may prove to be a better predictor of atherosclerosis than the total cholesterol/HDL cholesterol ratio.
PMCID: PMC508026  PMID: 9109446
18.  Autoimmune-Mediated Reduction of High-Density Lipoprotein-Cholesterol and Paraoxonase-1 Activity in SLE-Prone gld Mice 
Arthritis and rheumatism  2011;63(1):201-211.
To characterize modifications of high-density lipoprotein (HDL) in autoimmune gld (generalized lymphoproliferative disorder) mice that may be relevant to premature atherosclerosis in systemic lupus erythematosus and assess their relationship to specific aspects of autoimmune disease.
HDL-cholesterol (HDL-C), apolipoprotein-A1 (ApoA1), paraoxonase-1 (PON1) activity, hepatic gene expression and HDL biogenesis were measured in ageing female gld and wild-type (WT) congenic mice. Autoantibodies, lymphoid organs and cytokines were analyzed by enzyme-linked immunosorbent assay, flow cytometry and multiplex assay respectively.
Plasma HDL-C, HDL-ApoA1 and HDL-associated PON1 activity were reduced in ageing gld mice in association with the development of autoimmunity independently of changes in hepatic ApoA1 and PON1 expression or HDL biogenesis. Hepatic induction of the acute phase reactant, serum amyloid A-1, resulted in its incorporation onto HDL in gld mice. Deletion of the lipid-sensitive receptor, G2A, in gld mice (G2A-/-gld) attenuated reductions in HDL-C and PON1 activity without altering hepatic ApoA1 and PON1 expression, HDL biogenesis or levels of acute phase pro-inflammatory cytokines. Plasma anti-ApoA1 autoantibodies were elevated in ageing gld mice commensurate with detectable increases in ApoA1 immune complexes. Autoantibodies were lower in ageing G2A-/-gld mice compared to gld mice and anti-ApoA1 autoantibody levels were significantly related to HDL-C concentration (r=-0.645, p<0.00004) and PON1 activity (r=-0.555, p<0.0007) amongst autoimmune gld and G2A-/-gld mice.
Autoantibodies against ApoA1 contribute to reducing HDL-C and PON1 activity in autoimmune gld mice independently of hepatic HDL biogenesis, suggesting that functional impairment and premature clearance of HDL immune complexes may be principal mechanisms involved.
PMCID: PMC3032585  PMID: 20882670
19.  In vivo administration of BL-3050: highly stable engineered PON1-HDL complexes 
Serum paraoxonase (PON1) is a high density lipoprotein (HDL)-associated enzyme involved in organophosphate (OP) degradation and prevention of atherosclerosis. PON1 comprises a potential candidate for in vivo therapeutics, as an anti-atherogenic agent, and for detoxification of pesticides and nerve agents. Because human PON1 exhibits limited stability, engineered, recombinant PON1 (rePON1) variants that were designed for higher reactivity, solubility, stability, and bacterial expression, are candidates for treatment. This work addresses the feasibility of in vivo administration of rePON1, and its HDL complex, as a potentially therapeutic agent dubbed BL-3050.
For stability studies we applied different challenges related to the in vivo disfunctionalization of HDL and PON1 and tested for inactivation of PON1's activity. We applied acute, repetitive administrations of BL-3050 in mice to assess its toxicity and adverse immune responses. The in vivo efficacy of recombinant PON1 and BL-3050 were tested with an animal model of chlorpyrifos-oxon poisoning.
Inactivation studies show significantly improved in vitro lifespan of the engineered rePON1 relative to human PON1. Significant sequence changes relative to human PON1 might hamper the in vivo applicability of BL-3050 due to adverse immune responses. However, we observed no toxic effects in mice subjected to repetitive administration of BL-3050, suggesting that BL-3050 could be safely used. To further evaluate the activity of BL-3050 in vivo, we applied an animal model that mimics human organophosphate poisoning. In these studies, a significant advantages of rePON1 and BL-3050 (>87.5% survival versus <37.5% in the control groups) was observed. Furthermore, BL-3050 and rePON1 were superior to the conventional treatment of atropine-2-PAM as a prophylactic treatment for OP poisoning.
In vitro and in vivo data described here demonstrate the potential advantages of rePON1 and BL-3050 for treatment of OP toxicity and chronic cardiovascular diseases like atherosclerosis. The in vivo data also suggest that rePON1 and BL-3050 are stable and safe, and could be used for acute, and possibly repeated treatments, with no adverse effects.
PMCID: PMC2785756  PMID: 19922610
20.  Paraoxonase 2 (PON2) in the mouse central nervous system: a neuroprotective role? 
Toxicology and applied pharmacology  2011;256(3):369-378.
The aim of this study was to characterize the expression of paraoxonase 2 (PON2) in mouse brain and to assess its antioxidant properties. PON2 levels were highest in lung, intestine, heart and liver, and lower in brain; in all tissues, PON2 expression was higher in female than in male mice. PON2 knockout [PON2-/-] mice did not express any PON2, as expected. In brain, the highest levels of PON2 were found in the substantia nigra, the nucleus accumbens and the striatum, with lower levels in the cerebral cortex, hippocampus, cerebellum and brainstem. A similar regional distribution of PON2 activity (measured by dihydrocoumarin hydrolysis) was also found. PON3 was not detected in any brain area, while PON1 was expressed at very low levels, and did not show any regional difference. PON2 levels were higher in astrocytes than in neurons isolated from all brain regions, and were highest in cells from the striatum. PON2 activity and mRNA levels followed a similar pattern. Brain PON2 levels were highest around birth, and gradually declined. Subcellular distribution experiments indicated that PON2 is primarily expressed in microsomes and in mitochondria. The toxicity in neurons and astrocytes of agents known to cause oxidative stress (DMNQ and H2O2) was higher in cells from PON2-/- mice than in the same cells from wild-type mice, despite similar glutathione levels. These results indicate that PON2 is expressed in brain, and that higher levels are found in dopaminergic regions such as the striatum, suggesting that this enzyme may provide protection against oxidative stress-mediated neurotoxicity.
PMCID: PMC3155737  PMID: 21354197
Paraoxonase 2; PON2; striatum; antioxidant
21.  Modulation of paraoxonases during infectious diseases and its potential impact on atherosclerosis 
The paraoxonase (PON) gene family includes three members, PON1, PON2 and PON3, aligned in tandem on chromosome 7 in humans and on chromosome 6 in mice. All PON proteins share considerable structural homology and have the capacity to protect cells from oxidative stress; therefore, they have been implicated in the pathogenesis of several inflammatory diseases, particularly atherosclerosis. The major goal of this review is to highlight the modulation of each of the PONs by infective (bacterial, viral and parasitic) agents, which may shed a light on the interaction between infectious diseases and PONs activities in order to effectively reduce the risk of developing atherosclerosis.
PMCID: PMC3457911  PMID: 22824324
Atherosclerosis; Paraoxonases; Oxidative stress; Infectious diseases
22.  Human PON1, a biomarker of risk of disease and exposure 
Chemico-biological interactions  2010;187(1-3):355-361.
Human paraoxonase 1 (PON1) is a high-density lipoprotein (HDL)-associated serum enzyme that exhibits a broad substrate specificity. In addition to protecting against exposure to some organophosphorus (OP) pesticides by hydrolyzing their toxic oxon metabolites, PON1 is important in protecting against vascular disease by metabolizing oxidized lipids. Recently, PON1 has also been shown to play a role in inactivating the quorum sensing factor N-(3-oxododecanoyl)-L-homoserine lactone (3OC12-HSL) of Pseudomonas aeruginosa. Native, untagged engineered recombinant human PON1 (rHuPON1) expressed in E. coli and purified by conventional column chromatographic purification is stable, active, and capable of protecting PON1 knockout mice (PON1-/-) from exposure to high levels of the OP compound diazoxon. The bacterially-derived rHuPON1 can be produced in large quantities and lacks the glycosylation of eukaryotic systems that can produce immunogenic complications when inappropriately glycosylated recombinant proteins are used as therapeutics. Previous studies have shown that the determination of PON1 status, which reveals both PON1192 functional genotype and serum enzyme activity level, is required for a meaningful evaluation of PON1’s role in risk of disease or exposure. We have developed a new two-substrate assay/analysis protocol that provides PON1 status without use of toxic OP substrates, allowing for use of this protocol in non-specialized laboratories. Factors were also determined for inter-converting rates of hydrolysis of different substrates. PON1 status also plays an important role in revealing changes in HDL-associated PON1 activities in male patients with Parkinson disease (PD). Immunolocalization studies of PONs 1, 2 and 3 in nearly all mouse tissues suggests that the functions of PONs 1 and 3 extend beyond the plasma and the HDL particle.
PMCID: PMC3035622  PMID: 20338154
paraoxonase 1; Parkinson disease; organophosphate; therapy for OP poisoning; chlorpyrifos/chlorpyrifos oxon; diazinon/diazoxon
23.  Paraoxonase: Its antiatherogenic role in chronic renal failure 
Indian Journal of Nephrology  2010;20(1):9-14.
Paraoxonase (PON) is an aryldialkylphosphatase, which reversibly binds and hydrolyzes organophosphates. The PON family has three members (PON1, PON2 and PON3); they share structural properties and enzymatic activities. PON1 is shown to reside over high density lipoprotein (HDL) and has both antioxidant and antiatherogenic functions. Function of PON2 and PON3 are speculative and still under research. Several methodologies were developed over the years to determine the activity and mass of PON1, of which spectrophotometer-based methods using certain chemicals as substrate predominate. Several studies have shown decreased levels of PON1 in chronic renal failure (CRF) patients, particularly those on hemodialysis. The role of PON1 in development of cardiovascular disease has drawn considerable attention in recent years. Several authors have shown decreased levels of HDL and PON1 activity in CRF patients on hemodialysis and reported this to be a risk factor in the development of CVD. Enhancement or maintenance of the PON1 activity may prevent development of CVDs and its consequences in patients on hemodialysis.
PMCID: PMC2878404  PMID: 20535264
Atherosclerosis; chronic renal failure; hemodialysis; high density lipoprotein; low density lipoprotein oxidation; paraoxonase
24.  Both Paraoxonase-1 Genotype and Activity Do Not Predict the Risk of Future Coronary Artery Disease; the EPIC-Norfolk Prospective Population Study 
PLoS ONE  2009;4(8):e6809.
Paraoxonase-1 (PON1) is an antioxidant enzyme, that resides on high-density lipoprotein (HDL). PON1-activity, is heavily influenced by the PON1-Q192R polymorphism. PON1 is considered to protect against atherosclerosis, but it is unclear whether this relation is independent of its carrier, HDL. In order to evaluate the atheroprotective potential of PON1, we assessed the relationships among PON1-genotype, PON1-activity and risk of future coronary artery disease (CAD), in a large prospective case-control study.
Methodology/Principal Findings
Cases (n = 1138) were apparently healthy men and women aged 45–79 years who developed fatal or nonfatal CAD during a mean follow-up of 6 years. Controls (n = 2237) were matched by age, sex and enrollment time. PON1-activity was similar in cases and controls (60.7±45.3 versus 62.6±45.8 U/L, p = 0.3) and correlated with HDL-cholesterol levels (r = 0.16, p<0.0001). The PON1-Q192R polymorphism had a profound impact on PON1-activity, but did not predict CAD risk (Odds Ratio [OR] per R allele 0.98[0.84–1.15], p = 0.8). Using conditional logistic regression, quartiles of PON1-activity showed a modest inverse relation with CAD risk (OR for the highest versus the lowest quartile 0.77[0.63–0.95], p = 0.01; p-trend = 0.06). PON1-activity adjusted for Q192R polymorphism correlated better with HDL-cholesterol (r = 0.26, p<0.0001) and more linearly predicted CAD risk (0.79[0.64–0.98], p = 0.03; p-trend = 0.008). However, these relationships were abolished after adjustment for HDL (particles-cholesterol-size) and apolipoproteinA-I (0.94[0.74–1.18], p-trend = 0.3).
This study, shows that PON1-activity inversely relates to CAD risk, but not independent of HDL, due to its close association with the HDL-particle. These data strongly suggest that a low PON1-activity is not a causal factor in atherogenesis.
PMCID: PMC2728540  PMID: 19710913
25.  Quercetin & Ethanol Attenuate the Progression of Atherosclerotic Plaques with Concomitant Up Regulation of Paraoxonase1 (PON1) Gene Expression and PON1 Activity in LDLR-/- Mice 
Since moderate wine drinking is atheroprotective it is clinically relevant to elucidate its possible mechanism/s of action/s. Our objective is to demonstrate the potential benefits of the wine components, quercetin and ethanol, on the development of aortic plaques with parallel changes in antiatherogenic factors.
Methods and Results
The effects of quercetin and ethanol on the development of aortic atherosclerotic lesions, liver PON1 gene expression, and serum PON1 activity were measured in LDLR-/- mice on an atherogenic diet for 4-weeks and 8-weeks. Depending on the duration and dosage of these modulators, 12.5-25 mg/dl quercetin (12.5Q-25Q) and 18-25% ethanol, the magnitude of decreases in aortic lesions caused by moderate ethanol and quercetin ranged from 20-70% (p<0.05-p<0.001) based on ultrasound biomicroscopy (UBM) analyses, and from 18-61% (p<0.05-p<0.001) based on morphometric analyses. The composite plot of all the UBM and morphometric data showed significant correlation between these two methods (p = 0.0001, Pearson r = 0.79 for 4-weeks treatment; p = 0.000004, Pearson r = 0.84 for 8-weeks treatment). Concomitantly, 4-week treatments with 12.5Q and 18% ethanol up regulated liver PON1 mRNA by 41% (p<0.05) and 37% (p<0.05), respectively, accompanied by 92% (p<0.001) and 61% (p<0.001) increases in serum PON1 activity, respectively. The corresponding values after 8-weeks treatment with 12.5Q and 18% ethanol were 23% (p<0.05) and 40% (p<0.02) with respect to the up regulation of liver PON1 mRNA expression while the stimulations of serum PON1 activity were 75% (p<0.001) and 90% (p<0.001), respectively.
Based on these findings, we conclude that quercetin and moderate ethanol significantly inhibit the progression of atherosclerosis by up regulating the hepatic expression of the antiatherogenic gene, PON1, with concomitant increased serum PON1 activity.
PMCID: PMC2929280  PMID: 20586760
atherosclerosis; paraoxonase; LDLR-/- mouse; ethanol; quercetin

Results 1-25 (808746)