[Show abstract][Hide abstract] ABSTRACT: OBJECTIVE: Atherosclerotic lesions are associated with the accumulation of reactive aldehydes derived from oxidized lipids. Although inhibition of aldehyde metabolism has been shown to exacerbate atherosclerosis and enhance the accumulation of aldehyde-modified proteins in atherosclerotic plaques, no therapeutic interventions have been devised to prevent aldehyde accumulation in atherosclerotic lesions.Approach and Results-We examined the efficacy of carnosine, a naturally occurring β-alanyl-histidine dipeptide, in preventing aldehyde toxicity and atherogenesis in apolipoprotein E-null mice. In vitro, carnosine reacted rapidly with lipid peroxidation-derived unsaturated aldehydes. Gas chromatography mass-spectrometry analysis showed that carnosine inhibits the formation of free aldehydes 4-hydroxynonenal and malonaldialdehyde in Cu(2+)-oxidized low-density lipoprotein. Preloading bone marrow-derived macrophages with cell-permeable carnosine analogs reduced 4-hydroxynonenal-induced apoptosis. Oral supplementation with octyl-D-carnosine decreased atherosclerotic lesion formation in aortic valves of apolipoprotein E-null mice and attenuated the accumulation of protein-acrolein, protein-4-hydroxyhexenal, and protein-4-hydroxynonenal adducts in atherosclerotic lesions, whereas urinary excretion of aldehydes as carnosine conjugates was increased. CONCLUSIONS: The results of this study suggest that carnosine inhibits atherogenesis by facilitating aldehyde removal from atherosclerotic lesions. Endogenous levels of carnosine may be important determinants of atherosclerotic lesion formation, and treatment with carnosine or related peptides could be a useful therapy for the prevention or the treatment of atherosclerosis.
Arteriosclerosis Thrombosis and Vascular Biology 04/2013; · 6.34 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Lipid peroxidation products, such as 4-hydroxy-trans-2-nonenal (HNE), cause endothelial activation, and they increase the adhesion of the endothelium to circulating leukocytes. Nevertheless, the mechanisms underlying these effects remain unclear. We observed that in HNE-treated human umbilical vein endothelial cells, some of the protein-HNE adducts colocalize with the endoplasmic reticulum (ER) and that HNE forms covalent adducts with several ER chaperones that assist in protein folding. We also found that at concentrations that did not induce apoptosis or necrosis, HNE activated the unfolded protein response, leading to an increase in XBP-1 splicing, phosphorylation of protein kinase-like ER kinase and eukaryotic translation initiation factor 2α, and the induction of ATF3 and ATF4. This increase in eukaryotic translation initiation factor 2α phosphorylation was prevented by transfection with protein kinase-like ER kinase siRNA. Treatment with HNE increased the expression of the ER chaperones, GRP78 and HERP. Exposure to HNE led to a depletion of reduced glutathione and an increase in the production of reactive oxygen species (ROS); however, glutathione depletion and ROS production by tert-butyl-hydroperoxide did not trigger the unfolded protein response. Pretreatment with a chemical chaperone, phenylbutyric acid, or adenoviral transfection with ATF6 attenuated HNE-induced monocyte adhesion and IL-8 induction. Moreover, phenylbutyric acid and taurine-conjugated ursodeoxycholic acid attenuated HNE-induced leukocyte rolling and their firm adhesion to the endothelium in rat cremaster muscle. These data suggest that endothelial activation by HNE is mediated in part by ER stress, induced by mechanisms independent of ROS production or glutathione depletion. The induction of ER stress may be a significant cause of vascular inflammation induced by products of oxidized lipids.
Journal of Biological Chemistry 01/2012; 287(14):11398-409. · 4.65 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The generation of oxidized phospholipids in lipoproteins has been linked to vascular inflammation in atherosclerotic lesions. Products of phospholipid oxidation increase endothelial activation; however, their effects on macrophages are poorly understood, and it is unclear whether these effects are regulated by the biochemical pathways that metabolize oxidized phospholipids. We found that incubation of 1-palmitoyl-2-(5'-oxo-valeroyl)-sn-glycero-3-phosphocholine (POVPC) with THP-1-derived macrophages upregulated the expression of cytokine genes, including granulocyte/macrophage colony-stimulating factor (GM-CSF), tumor necrosis factor (TNF)-α, monocyte chemotactic protein 1 (MCP-1), interleukin (IL)-1β, IL-6, and IL-8. In these cells, reagent POVPC was either hydrolyzed to lyso-phosphatidylcholine (lyso-PC) or reduced to 1-palmitoyl-2-(5-hydroxy-valeroyl)-sn-glycero-3-phosphocholine (PHVPC). Treatment with the phospholipase A(2) (PLA(2)) inhibitor, pefabloc, decreased POVPC hydrolysis and increased PHVPC accumulation. Pefabloc also increased the induction of cytokine genes in POVPC-treated cells. In contrast, PHVPC accumulation and cytokine production were decreased upon treatment with the aldose reductase (AR) inhibitor, tolrestat. In comparison with POVPC, lyso-PC led to 2- to 3-fold greater and PHVPC 10- to 100-fold greater induction of cytokine genes. POVPC-induced cytokine gene induction was prevented in bone-marrow derived macrophages from AR-null mice. These results indicate that although hydrolysis is the major pathway of metabolism, reduction further increases the proinflammatory responses to POVPC. Thus, vascular inflammation in atherosclerotic lesions is likely to be regulated by metabolism of phospholipid aldehydes in macrophages.
The Journal of Lipid Research 09/2011; 52(12):2209-25. · 4.39 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Environmental triggers of dilated cardiomyopathy are poorly understood. Acute exposure to acrolein, a ubiquitous aldehyde pollutant, impairs cardiac function and cardioprotective responses in mice. Here, we tested the hypothesis that chronic oral exposure to acrolein induces inflammation and cardiomyopathy. C57BL/6 mice were gavage-fed acrolein (1 mg/kg) or water (vehicle) daily for 48 days. The dose was chosen based on estimates of human daily unsaturated aldehyde consumption. Compared with vehicle-fed mice, acrolein-fed mice exhibited significant (P < 0.05) left ventricular (LV) dilatation (LV end-diastolic volume 36 ± 8 vs. 17 ± 5 μl), contractile dysfunction (dP/dt(max) 4,697 ± 1,498 vs. 7,016 ± 1,757 mmHg/s), and impaired relaxation (tau 15.4 ± 4.3 vs. 10.4 ± 2.2 ms). Histological and biochemical evaluation revealed myocardial oxidative stress (membrane-localized protein-4-hydroxy-trans-2-nonenal adducts) and nitrative stress (increased protein-nitrotyrosine) and varying degrees of plasma and myocardial protein-acrolein adduct formation indicative of physical translocation of ingested acrolein to the heart. Acrolein also induced myocyte hypertrophy (~2.2-fold increased myocyte area, P < 0.05), increased apoptosis (~7.5-fold), and disrupted endothelial nitric oxide synthase in the heart. DNA binding studies, immunohistochemistry, and PCR revealed significant (P < 0.05) activation of nuclear factor-κB in acrolein-exposed hearts, along with upregulated gene expression of proinflammatory cytokines tumor necrosis factor-α and interleukin-1β. Long-term oral exposure to acrolein, at an amount within the range of human unsaturated aldehyde intake, induces a phenotype of dilated cardiomyopathy in the mouse. Human exposure to acrolein may have analogous effects and raise consideration of an environmental, aldehyde-mediated basis for heart failure.
[Show abstract][Hide abstract] ABSTRACT: The metabolism of α,β-unsaturated aldehydes, e.g., 4-hydroxynonenal, involves oxidation to carboxylic acids, reduction to alcohols, and glutathionylation to eventually form mercapturide conjugates. Recently, we demonstrated that P450s can oxidize aldehydes to carboxylic acids, a reaction previously thought to involve aldehyde dehydrogenase. When recombinant cytochrome P450 3A4 was incubated with 4-hydroxynonenal, O(2), and NADPH, several products were produced, including 1,4-dihydroxynonene (DHN), 4-hydroxy-2-nonenoic acid (HNA), and an unknown metabolite. Several P450s catalyzed the reduction reaction in the order (human) P450 2B6 ≅ P450 3A4 > P450 1A2 > P450 2J2 > (mouse) P450 2c29. Other P450s did not catalyze the reduction reaction (human P450 2E1 and rabbit P450 2B4). Metabolism by isolated rat hepatocytes showed that HNA formation was inhibited by cyanamide, while DHN formation was not affected. Troleandomycin increased HNA production 1.6-fold while inhibiting DHN formation, suggesting that P450 3A11 is a major enzyme involved in rat hepatic clearance of 4-HNE. A fluorescent assay was developed using 9-anthracenealdehyde to measure both reactions. Feeding mice a diet containing t-butylated hydroxyanisole increased the level of both activities with hepatic microsomal fractions but not proportionally. Miconazole (0.5 mM) was a potent inhibitor of these microsomal reduction reactions, while phenytoin and α-naphthoflavone (both at 0.5 mM) were partial inhibitors, suggesting the role of multiple P450 enzymes. The oxidative metabolism of these aldehydes was inhibited >90% in an Ar or CO atmosphere, while the reductive reactions were not greatly affected. These results suggest that P450s are significant catalysts of the reduction of α,β-unsaturated aldehydes in the liver.
Chemical Research in Toxicology 08/2011; 24(8):1223-30. · 3.67 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Aldehydes are ubiquitous natural constituents of foods, water and beverages. Dietary intake represents the greatest source of exposure to acrolein and related aldehydes. Oral acrolein induces dyslipidemia acutely and chronically increases atherosclerosis in mice, yet the mechanisms are unknown. Because lipid synthesis and trafficking are largely under hepatic control, we examined hepatic genes in murine models of acute and chronic oral acrolein exposure.
Changes in hepatic gene expression were examined using a Steroltalk microarray. Acute acrolein feeding modified plasma and hepatic proteins and increased plasma triglycerides within 15 min. By 6 h, acrolein altered hepatic gene expression including Insig1, Insig2 and Hmgcr genes and stimulated an acute-phase response (APR) with up-regulation of serum amyloid A genes (Saa) and systemic hypoalbuminemia. To test if decreased HMG-CoA reductase activity could modify acrolein-induced dyslipidemia or the APR, mice were pretreated with simvastatin. Statin treatment, however, did not alter acrolein-induced dyslipidemia or hypoalbuminemia associated with an APR. Few hepatic genes were dysregulated by chronic acrolein feeding in apoE-null mice. These studies confirmed that acute acrolein exposure altered expression of hepatic genes involved with lipid synthesis and trafficking and APR, and thus, indicated a hepatic locus of acrolein-induced dyslipidemia and APR that was independent of HMG CoA-reductase.
Dietary intake of acrolein could contribute to cardiovascular disease risk by disturbing hepatic function.
[Show abstract][Hide abstract] ABSTRACT: This protocol describes the procedure for measuring the relative rates of metabolism of the α,β-unsaturated aldehydes 9-anthracene aldehyde (9-AA) and 4-hydroxy-trans-2-nonenal (4-HNE). More specifically, these assays measure the aldehyde reduction reactions of cytochrome P450s (CYPs). They can be performed using liver microsomal or other tissue fractions, spherosome preparations of recombinant CYPs, or recombinant CYPs from other sources. The method for reduction of 9-AA (a model α,β-unsaturated aldehyde) by CYPs was adapted from an assay for 9-anthracene oxidation published by Marini et al. (2003). For reduction of the endogenous aldehyde 4-HNE, the substrate was incubated with CYP in the presence of oxygen and NADPH, and the metabolites were separated by HPLC, using an adaptation of the method by Srivastava et al. (2010). For both 9-AA and 4-HNE, the first step involves incubation of the substrate with the CYP in an appropriate medium. This is followed by quantification of metabolites through by spectrofluorometry (9-AA) or HPLC coupled with a radiometric assay (4-HNE). Metabolite identification can be achieved by HPLC GC/MS analysis. Inhibitors of cytochrome P450 can be utilized to show the role of the hemoprotein or other enzymes in these reduction reactions. The reduction of CYPs is not inhibited by either anaerobiosis or inclusion of CO in the gaseous phase of the reaction mixture. These characteristics are similar to those reported for some cytochrome P450-catalyzed azo reduction reactions.
Current protocols in toxicology 05/2011; Chapter 4:Unit4.37.
[Show abstract][Hide abstract] ABSTRACT: Acrolein is a dietary aldehyde that is present in high concentrations in alcoholic beverages and foods including cheese, donuts and coffee. It is also abundant in tobacco smoke, automobile exhaust and industrial waste and is generated in vivo during inflammation and oxidative stress.
The goal of this study was to examine the effects of dietary acrolein on atherosclerosis.
Eight-week-old male apoE-null mice were gavage-fed acrolein (2.5mg/kg/day) for 8 weeks. Atherosclerotic lesion formation and composition and plasma lipids and platelet factor 4 (PF4) levels were measured. Effects of acrolein and PF4 on endothelial cell function was measured in vitro.
Acrolein feeding increased the concentration of cholesterol in the plasma. NMR analysis of the lipoproteins showed that acrolein feeding increased the abundance of small and medium VLDL particles. Acrolein feeding also increased atherosclerotic lesion formation in the aortic valve and the aortic arch. Immunohistochemical analysis showed increased macrophage accumulation in the lesions of acrolein-fed mice. Plasma PF4 levels and accumulation of PF4 in atherosclerotic lesions was increased in the acrolein-fed mice. Incubation of endothelial cells with the plasma of acrolein-fed mice augmented transmigration of monocytic cells, which was abolished by anti-PF4 antibody treatment.
Dietary exposure to acrolein exacerbates atherosclerosis in apoE-null mice. Consumption of foods and beverages rich in unsaturated aldehydes such as acrolein may be a contributing factor to the progression of atherosclerotic lesions.
[Show abstract][Hide abstract] ABSTRACT: Oxidative stress-induced inflammation is a major contributor to several disease conditions including sepsis, carcinogenesis and metastasis, diabetic complications, allergic asthma, uveitis and after cataract surgery posterior capsular opacification. Since reactive oxygen species (ROS)-mediated activation of redox-sensitive transcription factors and subsequent expression of inflammatory cytokines, chemokines and growth factors are characteristics of inflammatory disorders, we envisioned that by blocking the molecular signals of ROS that activate redox-sensitive transcription factors, various inflammatory diseases could be ameliorated. We have indeed demonstrated that ROS-induced lipid peroxidation-derived lipid aldehydes such as 4-hydroxy-trans-2-nonenal (HNE) and their glutathione-conjugates (e.g. GS-HNE) are efficiently reduced by aldose reductase to corresponding alcohols which mediate the inflammatory signals. Our results showed that inhibition of aldose reductase (AKR1B1) significantly prevented the inflammatory signals induced by cytokines, growth factors, endotoxins, high glucose, allergens and auto-immune reactions in cellular as well as animal models. We have demonstrated that AKR1B1 inhibitor, fidarestat, significantly prevents tumor necrosis factor-alpha (TNF-α)-, growth factors-, lipopolysachharide (LPS)-, and environmental allergens-induced inflammatory signals that cause various inflammatory diseases. In animal models of inflammatory diseases such as diabetes, cardiovascular, uveitis, asthma, and cancer (colon, breast, prostate and lung) and metastasis, inhibition of AKR1B1 significantly ameliorated the disease. Our results from various cellular and animal models representing a number of inflammatory conditions suggest that ROS-induced inflammatory response could be reduced by inhibition of AKR1B1, thereby decreasing the progression of the disease and if the therapy is initiated early, the disease could be eliminated. Since fidarestat has already undergone phase III clinical trial for diabetic neuropathy and found to be safe, though clinically not very effective, our results indicate that it can be developed for the therapy of a number of inflammation-related diseases. Our results thus offer a novel therapeutic approach to treat a wide array of inflammatory diseases.
[Show abstract][Hide abstract] ABSTRACT: Diabetes results in enhanced chemical modification of proteins by advanced lipoxidation end products (ALEs) and advanced glycation end products (AGEs) precursors. These modifications have been linked to the development of several secondary diabetic complications. Our previous studies showed that aldose reductase (AR; AKR1B3) catalyzes the reduction of ALEs and AGEs precursors; however, the in vivo significance of this metabolic pathway during diabetes and obesity has not been fully assessed. Therefore we examined the role of AR in regulating ALEs and AGEs formation in murine models of diet-induced obesity and streptozotocin-induced diabetes. In comparison with wild-type (WT) and AR-null mice fed normal chow, mice fed a high-fat (HF) diet (42% kcal fat) showed increased accumulation of AGEs and protein-acrolein adducts in the plasma. AGEs and acrolein adducts were also increased in the epididymal fat of WT and AR-null mice fed a HF diet. Deletion of AR increased the accumulation of 4-hydroxy-trans-2-nonenal (HNE) protein adduct in the plasma and increased the expression of the AGE receptor (RAGE) in HF fed mice. No change in AGEs formation was observed in the kidneys of HF-fed mice. In comparison, renal tissue from AR-null mice treated with streptozotocin showed greater AGE accumulation than streptozotocin-treated WT mice. These data indicated that AR regulated the accumulation of lipid peroxidation derived aldehydes and AGEs under conditions of severe, but not mild, hyperglycemia and that deletion of AR increased RAGE-induction via mechanisms that were independent of AGEs accumulation.
[Show abstract][Hide abstract] ABSTRACT: Acrolein is a common air pollutant that is present in high concentrations in wood, cotton, and tobacco smoke, automobile exhaust and industrial waste and emissions. Exposure to acrolein containing environmental pollutants such as tobacco smoke and automobile exhaust has been linked to the activation of the coagulation and hemostasis pathways and thereby to the predisposition of thrombotic events in human. To examine the effects of acrolein on platelets, adult male C57Bl/6 mice were subjected acute (5ppm for 6h) or sub-chronic (1ppm, 6h/day for 4days) acrolein inhalation exposures. The acute exposure to acrolein did not cause pulmonary inflammation and oxidative stress, dyslipidemia or induce liver damage or muscle injury. Platelet GSH levels in acrolein-exposed mice were comparable to controls, but acrolein-exposure increased the abundance of protein-acrolein adducts in platelets. Platelets isolated from mice, exposed to both acute and sub-chronic acrolein levels, showed increased ADP-induced platelet aggregation. Exposure to acrolein also led to an increase in the indices of platelet activation such as the formation of platelet-leukocyte aggregates in the blood, plasma PF4 levels, and increased platelet-fibrinogen binding. The bleeding time was decreased in acrolein exposed mice. Plasma levels of PF4 were also increased in mice exposed to environmental tobacco smoke. Similar to inhalation exposure, acrolein feeding to mice also increased platelet activation and established a pro-thrombotic state in mice. Together, our data suggest that acrolein is an important contributing factor to the pro-thrombotic risk in human exposure to pollutants such as tobacco smoke or automobile exhaust, or through dietary consumption.
Toxicology and Applied Pharmacology 10/2010; 248(2):100-10. · 3.98 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Heme oxygenase-1 (HO-1) is an inducible stress-response protein that imparts antioxidant and antiapoptotic effects. However, its pathophysiological role in cardiac remodeling and chronic heart failure (HF) is unknown. We hypothesized that induction of HO-1 in HF alleviates pathological remodeling.
Adult male nontransgenic and myocyte-restricted HO-1 transgenic mice underwent either sham operation or coronary ligation to induce HF. Four weeks after ligation, nontransgenic HF mice exhibited postinfarction left ventricular (LV) remodeling and dysfunction, hypertrophy, fibrosis, oxidative stress, apoptosis, and reduced capillary density, associated with a 2-fold increase in HO-1 expression in noninfarcted myocardium. Compared with nontransgenic mice, HO-1 transgenic HF mice exhibited significantly (P<0.05) improved postinfarction survival (94% versus 57%) and less LV dilatation (end-diastolic volume, 46+/-8 versus 85+/-32 microL), mechanical dysfunction (ejection fraction, 65+/-9% versus 49+/-16%), hypertrophy (LV/tibia length 4.4+/-0.4 versus 5.2+/-0.6 mg/mm), interstitial fibrosis (11.2+/-3.1% versus 18.5+/-3.5%), and oxidative stress (3-fold reduction in tissue malondialdehyde). Moreover, myocyte-specific HO-1 overexpression in HF promoted tissue neovascularization and ameliorated myocardial p53 expression (2-fold reduction) and apoptosis. In isolated mitochondria, mitochondrial permeability transition was inhibited by HO-1 in a carbon monoxide (CO)-dependent manner and was recapitulated by the CO donor tricarbonylchloro(glycinato)ruthenium(II) (CORM-3). HO-1-derived CO also prevented H2O2-induced cardiomyocyte apoptosis and cell death. Finally, in vivo treatment with CORM-3 alleviated postinfarction LV remodeling, p53 expression, and apoptosis.
HO-1 induction in the failing heart is an important cardioprotective adaptation that opposes pathological LV remodeling, and this effect is mediated, at least in part, by CO-dependent inhibition of mitochondrial permeability transition and apoptosis. Augmentation of HO-1 or its product, CO, may represent a novel therapeutic strategy for ameliorating HF.
[Show abstract][Hide abstract] ABSTRACT: Aldehydes such as acrolein are ubiquitous pollutants present in automobile exhaust, cigarette, wood, and coal smoke. Such aldehydes are also constituents of several food substances and are present in drinking water, irrigation canals, and effluents from manufacturing plants. Oral intake represents the most significant source of exposure to acrolein and related aldehydes. To study the effects of short-term oral exposure to acrolein on lipoprotein levels and metabolism, adult mice were gavage-fed 0.1 to 5 mg acrolein/kg bwt and changes in plasma lipoproteins were assessed. Changes in hepatic gene expression related to lipid metabolism and cytokines were examined by qRT-PCR analysis. Acrolein feeding did not affect body weight, blood urea nitrogen, plasma creatinine, electrolytes, cytokines or liver enzymes, but increased plasma cholesterol and triglycerides. Similar results were obtained with apoE-null mice. Plasma lipoproteins from acrolein-fed mice showed altered electrophoretic mobility on agarose gels. Chromatographic analysis revealed elevated VLDL cholesterol, phospholipids, and triglycerides levels with little change in LDL or HDL. NMR analysis indicated shifts from small to large VLDL and from large to medium-small LDL with no change in the size of HDL particles. Increased plasma VLDL was associated with a significant decrease in post-heparin plasma hepatic lipase activity and a decrease in hepatic expression of hepatic lipase. These observations suggest that oral exposure to acrolein could induce or exacerbate systemic dyslipidemia and thereby contribute to cardiovascular disease risk.
Toxicology and Applied Pharmacology 02/2010; 243(1):1-12. · 3.98 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Oxidation of lipids generates large quantities of highly reactive alpha,beta-unsaturated aldehydes (enals). Enals and their protein adducts accumulate in the tissues of several pathologies. In vitro, low concentrations of enals such as HNE (4-hydroxy trans-2-nonenal) affect cell signaling whereas high concentrations of enals are cytotoxic. Direct conjugation of the C2-C3 double bond of enals with the sulfhydryl group of GSH is a major route for the metabolism and detoxification of enals. Recently, we found that glutathionyl conjugate of HNE (GS-HNE) enhances the peritoneal leukocyte infiltration and stimulates the formation of proinflammatory lipid mediators. Moreover, the reduced form of the glutathione conjugate of HNE (GS-DHN) elicits strong mitogenic signaling in smooth muscle cells. In this chapter we discuss the methods to study the metabolism of enals and the redox signaling properties of glutathionyl conjugates of HNE.
Methods in enzymology 01/2010; 474:297-313. · 1.90 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Atherosclerotic lesion formation is associated with the accumulation of oxidized lipids. Products of lipid oxidation, particularly aldehydes, stimulate cytokine production and enhance monocyte adhesion; however, their contribution to atherosclerotic lesion formation remains unclear.
To test the hypothesis that inhibition of aldehyde removal by aldose reductase (AR), which metabolizes both free and phospholipid aldehydes, exacerbates atherosclerotic lesion formation.
In atherosclerotic lesions of apolipoprotein (apo)E-null mice, AR protein was located in macrophage-rich regions and its abundance increased with lesion progression. Treatment of apoE-null mice with AR inhibitors sorbinil or tolrestat increased early lesion formation but did not affect the formation of advanced lesions. Early lesions of AR(-/-)/apoE(-/-) mice maintained on high-fat diet were significantly larger when compared with age-matched AR(+/+)/apoE(-/-) mice. The increase in lesion area attributable to deletion of the AR gene was seen in both male and female mice. Pharmacological inhibition or genetic ablation of AR also increased the lesion formation in male mice made diabetic by streptozotocin treatment. Lesions in AR(-/-)/apoE(-/-) mice exhibited increased collagen and macrophage content and a decrease in smooth muscle cells. AR(-/-)/apoE(-/-) mice displayed a greater accumulation of the AR substrate 4-hydroxy trans-2-nonenal (HNE) in the plasma and protein-HNE adducts in arterial lesions than AR(+/+)/apoE(-/-) mice.
These observations indicate that AR is upregulated in atherosclerotic lesions and it protects against early stages of atherogenesis by removing toxic aldehydes generated in oxidized lipids.
Circulation Research 10/2009; 105(8):793-802. · 11.86 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Exposure to arsenic-contaminated water has been shown to be associated with cardiovascular disease, especially atherosclerosis. We examined the effect of arsenic exposure on atherosclerotic lesion formation, lesion composition and nature in ApoE-/- mice. Early post-natal exposure (3-week-old mice exposed to 49 ppm arsenic as NaAsO(2) in drinking water for 7 weeks) increased the atherosclerotic lesion formation by 3- to 5-fold in the aortic valve and the aortic arch, without affecting plasma cholesterol. Exposure to arsenic for 13 weeks (3-week-old mice exposed to 1, 4.9 and 49 ppm arsenic as NaAsO(2) in drinking water) increased the lesion formation and macrophage accumulation in a dose-dependent manner. Temporal studies showed that continuous arsenic exposure significantly exacerbated the lesion formation throughout the aortic tree at 16 and 36 weeks of age. Withdrawal of arsenic for 12 weeks after an initial exposure for 21 weeks (to 3-week-old mice) significantly decreased lesion formation as compared with mice continuously exposed to arsenic. Similarly, adult exposure to 49 ppm arsenic for 24 weeks, starting at 12 weeks of age increased lesion formation by 2- to 3.6-fold in the aortic valve, the aortic arch and the abdominal aorta. Lesions of arsenic-exposed mice displayed a 1.8-fold increase in macrophage accumulation whereas smooth muscle cell and T-lymphocyte contents were not changed. Expression of pro-inflammatory chemokine MCP-1 and cytokine IL-6 and markers of oxidative stress, protein-HNE and protein-MDA adducts were markedly increased in lesions of arsenic-exposed mice. Plasma concentrations of MCP-1, IL-6 and MDA were also significantly elevated in arsenic-exposed mice. These data suggest that arsenic exposure increases oxidative stress, inflammation and atherosclerotic lesion formation.
Toxicology and Applied Pharmacology 09/2009; 241(1):90-100. · 3.98 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To examine the role of aldo-keto reductases (AKRs) in the cardiovascular metabolism of the precursors of advanced glycation end products (AGEs).
Steady-state kinetic parameters of AKRs with AGE precursors were determined using recombinant proteins expressed in bacteria. Metabolism of methylglyoxal and AGE accumulation were studied in human umbilical vein endothelial cells (HUVECs) and C57 wild-type, akr1b3 (aldose reductase)-null, cardiospecific-akr1b4 (rat aldose reductase), and akr1b8 (FR-1)-transgenic mice. AGE accumulation and atherosclerotic lesions were studied 12 weeks after streptozotocin treatment of C57, akr1b3-null, and apoE- and akr1b3-apoE-null mice.
Higher levels of AGEs were generated in the cytosol than at the external surface of HUVECs cultured in high glucose, indicating that intracellular metabolism may be an important regulator of AGE accumulation and toxicity. In vitro, AKR 1A and 1B catalyzed the reduction of AGE precursors, whereas AKR1C, AKR6, and AKR7 were relatively ineffective. Highest catalytic efficiency was observed with AKR1B1. Acetol formation in methylglyoxal-treated HUVECs was prevented by the aldose reductase inhibitor sorbinil. Acetol was generated in hearts perfused with methylglyoxal, and its formation was increased in akr1b4- or akr1b8-transgenic mice. Reduction of AGE precursors was diminished in hearts from akr1b3-null mice. Diabetic akr1b3-null mice accumulated more AGEs in the plasma and the heart than wild-type mice, and deletion of akr1b3 increased AGE accumulation and atherosclerotic lesion formation in apoE-null mice.
Aldose reductase-catalyzed reduction is an important pathway in the endothelial and cardiac metabolism of AGE precursors, and it prevents AGE accumulation and atherosclerotic lesion formation.
[Show abstract][Hide abstract] ABSTRACT: Hemorrhagic cystitis and diffuse inflammation of the bladder, common side effects of cyclophosphamide (CY) treatment, have been linked to the generation of acrolein derived from CY metabolism. Metabolic removal of acrolein involves multiple pathways, which include reduction, oxidation, and conjugation with glutathione. Herein, we tested the hypothesis that glutathione S-transferase P (GSTP), the GST isoform that displays high catalytic efficiency with acrolein, protects against CY-induced urotoxicity by detoxifying acrolein. Treatment of wild-type (WT) and mGstP1/P2 null (GSTP-null) mice with CY caused hemorrhagic cystitis, edema, albumin extravasation, and sloughing of bladder epithelium; however, CY-induced bladder ulcerations of the lamina propria were more numerous and more severe in GSTP-null mice. CY treatment also led to greater accumulation of myeloperoxidase-positive cells and specific protein-acrolein adducts in the bladder of GSTP-null than WT mice. There was no difference in hepatic microsomal production of acrolein from CY or urinary hydroxypropyl mercapturic acid output between WT and GSTP-null mice, but CY induced greater c-Jun NH(2)-terminal kinase (JNK) and c-Jun, but not extracellular signal-regulated kinase or p38, activation in GSTP-null than in WT mice. Pretreatment with mesna (2-mercaptoethane sulfonate sodium) abolished CY toxicity and JNK activation in GSTP-null mice. Taken together, these data support the view that GSTP prevents CY-induced bladder toxicity, in part by detoxifying acrolein. Because polymorphisms in human GSTP gene code for protein variants differing significantly in their catalytic efficiency toward acrolein, it is likely that GSTP polymorphisms influence CY urotoxicity. In addition, pretreatment with dietary or nutrient inducers of GSTP may be of use in minimizing bladder injury in patients undergoing CY therapy.
Journal of Pharmacology and Experimental Therapeutics 09/2009; 331(2):456-69. · 3.89 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Aldose reductase (AR) is a multi-functional AKR (AKR1B1) that catalyzes the reduction of a wide range of endogenous and xenobiotic aldehydes and their glutathione conjugates with high efficiency. Previous studies from our laboratory show that AR protects against myocardial ischemia-reperfusion injury, however, the mechanisms by which it confers cardioprotection remain unknown. Because AR metabolizes aldehydes generated from lipid peroxidation, we tested the hypothesis that it protects against ischemic injury by preventing ER stress induced by excessive accumulation of aldehyde-modified proteins in the ischemic heart. In cell culture experiments, exposure to model lipid peroxidation aldehydes-4-hydroxy-trans-2-nonenal (HNE), 1-palmitoyl-2-oxovaleroyl phosphatidylcholine (POVPC) or acrolein led to an increase in the phosphorylation of ER stress markers PERK and eIF2-alpha and an increase in ATF3. The reduced metabolite of POVPC 1-palmitoyl-2-hydroxyvaleroyl phosphatidylcholine (PHVPC) was unable to stimulate JNK phosphorylation. No increase in phospho-eIF2-alpha, ATF3 or phospho-PERK was observed in cells treated with the reduced HNE metabolite 1,4-dihydroxynonenol (DHN). Lysates prepared from isolated perfused mouse hearts subjected to 15 min of global ischemia followed by 30 min of reperfusion ex vivo showed greater phosphorylation of PERK and eIF2-alpha than hearts subjected to aerobic perfusion alone. Ischemia-induced increases in phospho-PERK and phospho-eIF2-alpha were diminished in the hearts of cardiomyocyte-specific transgenic mice overexpressing the AR transgene. These observations support the notion that by removing aldehydic products of lipid peroxidation, AR decreases ischemia-reperfusion injury by diminishing ER stress.