Berenice Barajas

Ambient Particulate Pollutants in the Ultrafine Range Promote Early Atherosclerosis and Systemic Oxidative Stress

Air pollution is associated with significant adverse health effects, including increased cardiovascular morbidity and mortality. Exposure to particulate matter with an aerodynamic diameter of <2.5 &mgr;m (PM2.5) increases ischemic cardiovascular events and promotes atherosclerosis. Moreover, there is increasing evidence that the smallest pollutant particles pose the greatest danger because of their high content of organic chemicals and prooxidative potential. To test this hypothesis, we compared the proatherogenic effects of ambient particles of <0.18 &mgr;m (ultrafine particles) with particles of <2.5 &mgr;m in genetically susceptible (apolipoprotein E–deficient) mice. These animals were exposed to concentrated ultrafine particles, concentrated particles of <2.5 &mgr;m, or filtered air in a mobile animal facility close to a Los Angeles freeway. Ultrafine particle–exposed mice exhibited significantly larger early atherosclerotic lesions than mice exposed to PM2.5 or filtered air. Exposure to ultrafine particles also resulted in an inhibition of the antiinflammatory capacity of plasma high-density lipoprotein and greater systemic oxidative stress as evidenced by a significant increase in hepatic malondialdehyde levels and upregulation of Nrf2-regulated antioxidant genes. We conclude that ultrafine particles concentrate the proatherogenic effects of ambient PM and may constitute a significant cardiovascular risk factor.

Heme Oxygenase-1 Expression in Macrophages Plays a Beneficial Role in Atherosclerosis

Heme oxygenase (HO-1) is the rate-limiting enzyme in the catabolism of heme, which leads to the generation of biliverdin, iron, and carbon monoxide. It has been shown to have important antioxidant and antiinflammatory properties that result in a vascular antiatherogenic effect. To determine whether HO-1 expression in macrophages constitutes a significant component of the protective role in atherosclerosis, we evaluated the effect of decreased or absent HO-1 expression in peritoneal macrophages on oxidative stress and inflammation in vitro, and the effect of complete deficiency of HO-1 expression in macrophages in atherosclerotic lesion formation in vivo. We found that compared with HO-1+/+ controls, peritoneal macrophages from HO-1−/− and HO-1+/− mice exhibited (1) increased reactive oxygen species (ROS) generation, (2) increased proinflammatory cytokines such as monocyte chemotactic protein 1 (MCP-1) and interleukin 6 (IL-6), and (3) increased foam cell formation when treated with oxLDL, attributable in part to increased expression of scavenger receptor A (SR-A). Bone marrow transplantation experiments performed in lethally irradiated LDL-R null female mice, reconstituted with bone marrow from HO-1−/− versus HO-1+/+ mice, revealed that HO-1−/− reconstituted animals exhibited atherosclerotic lesions with a greater macrophage content as evaluated by immunohistochemistry and planimetric assessment. We conclude that HO-1 expression in macrophages constitutes an important component of the antiatherogenic effect by increasing antioxidant protection and decreasing the inflammatory component of atherosclerotic lesions.

Air-pollutant chemicals and oxidized lipids exhibit genome-wide synergistic effects on endothelial cells

BackgroundAmbient air pollution is associated with increased cardiovascular morbidity and mortality. We have found that exposure to ambient ultrafine particulate matter, highly enriched in redox cycling organic chemicals, promotes atherosclerosis in mice. We hypothesize that these pro-oxidative chemicals could synergize with oxidized lipid components generated in low-density lipoprotein particles to enhance vascular inflammation and atherosclerosis.ResultsWe have used human microvascular endothelial cells (HMEC) to study the combined effects of a model air pollutant, diesel exhaust particles (DEP), and oxidized 1-palmitoyl-2-arachidonyl-sn-glycero-3-phosphorylcholine (ox-PAPC) on genome-wide gene expression. We treated the cells in triplicate wells with an organic DEP extract, ox-PAPC at various concentrations, or combinations of both for 4 hours. Gene-expression profiling showed that both the DEP extract and ox-PAPC co-regulated a large number of genes. Using network analysis to identify coexpressed gene modules, we found three modules that were most highly enriched in genes that were differentially regulated by the stimuli. These modules were also enriched in synergistically co-regulated genes and pathways relevant to vascular inflammation. We validated this synergy in vivo by demonstrating that hypercholesterolemic mice exposed to ambient ultrafine particles exhibited significant upregulation of the module genes in the liver.ConclusionDiesel exhaust particles and oxidized phospholipids synergistically affect the expression profile of several gene modules that correspond to pathways relevant to vascular inflammatory processes such as atherosclerosis.

NF-E2–Related Factor 2 Promotes Atherosclerosis by Effects on Plasma Lipoproteins and Cholesterol Transport That Overshadow Antioxidant Protection

Objective—To test the hypothesis that NF-E2–related factor 2 (Nrf2) expression plays an antiatherogenic role by its vascular antioxidant and anti-inflammatory properties. Methods and Results—Nrf2 is an important transcription factor that regulates the expression of phase 2 detoxifying enzymes and antioxidant genes. Its expression in vascular cells appears to be an important factor in the protection against vascular oxidative stress and inflammation. We developed Nrf2 heterozygous (HET) and homozygous knockout (KO) mice on an apolipoprotein (apo) E–null background by sequential breeding, resulting in Nrf2−/−, apoE−/− (KO), Nrf2−/+, apoE−/− (HET) and Nrf2+/+, and apoE−/− wild-type littermates. KO mice exhibited decreased levels of antioxidant genes with evidence of increased reactive oxygen species generation compared with wild-type controls. Surprisingly, KO males exhibited 47% and 53% reductions in the degree of aortic atherosclerosis compared with HET or wild-type littermates, respectively. Decreased atherosclerosis in KO mice correlated with lower plasma total cholesterol in a sex-dependent manner. KO mice also had a decreased hepatic cholesterol content and a lower expression of lipogenic genes, suggesting that hepatic lipogenesis could be reduced. In addition, KO mice exhibited atherosclerotic plaques characterized by a lesser macrophage component and decreased foam cell formation in an in vitro lipid-loading assay. This was associated with a lower rate of cholesterol influx, mediated in part by decreased expression of the scavenger receptor CD36. Conclusion—Nrf2 expression unexpectedly promotes atherosclerotic lesion formation in a sex-dependent manner, most likely by a combination of systemic metabolic and local vascular effects.

Nrf2 activation by sulforaphane restores the age-related decrease of TH1 immunity: Role of dendritic cells

BACKGROUND The decrease in cellular immunity with aging is of considerable public health importance. Recent studies suggest that the redox equilibrium of dendritic cells (DCs) is a key factor in maintaining protective cellular immunity and that a disturbance of this homeostatic mechanism could contribute to immune senescence. OBJECTIVES We sought (1) to elucidate the role of DC redox equilibrium in the decrease of contact hypersensitivity (CHS) and T(H)1 immunity during aging and (2) to determine how restoration of glutathione (GSH) levels by the Nrf2-mediated antioxidant defense pathway affects this decrease. METHODS We assessed the effect of Nrf2 deficiency and boosting of GSH levels by the Nrf2 agonist sulforaphane or the thiol precursor N-acetyl cysteine (NAC) on the CHS response to contact antigens in old mice. We studied the effect of SFN and NAC on restoring T(H)1 immunity by treating DCs ex vivo before adoptive transfer and in vivo challenge. RESULTS Aging was associated with a decreased CHS response that was accentuated by Nrf2 deficiency. Systemic SFN treatment reversed this decrease through Nrf2-mediated antioxidant enzyme expression and GSH synthesis. Adoptive transfer of DCs from old animals induced a weakened CHS response in recipient animals. Treatment of DCs from old animals with SFN or NAC ex vivo restored the in vivo challenge response. CONCLUSION SFN and NAC upregulate T(H)1 immunity in aging through a restoration of redox equilibrium.

Nrf2 deficiency in dendritic cells enhances the adjuvant effect of ambient ultrafine particles on allergic sensitization.

Particulate matter (PM) is an important risk factor for asthma. Generation of oxidative stress by PM is a major mechanism of its health effects. Transcription factor nuclear factor (erythroid-derived 2)-like 2 (Nrf2) mediates antioxidant and phase II enzymes and is essential in protecting against oxidative stress and lung inflammation. We have previously shown that ambient ultrafine particles (UFP) could exert a potent adjuvant effect on allergic sensitization to ovalbumin (OVA) in mice. We hypothesized that Nrf2 deficiency in dendritic cells (DC) could enhance the adjuvant potential of UFP on allergic sensitization. We show that the adjuvant effect of intranasally instilled UFP is significantly enhanced in Nrf2 knockout (Nrf2-/-) mice compared with their wild-type (Nrf2+/+) counterparts. Under resting conditions, Nrf2-/- DC displayed an intrinsic predilection to a T helper 2-favoring cytokine profile characterized by a low level of IL-12p70 and a high level of IL-6 as compared to Nrf2+/+ DC. Adoptive transfer of OVA/UFP-treated Nrf2-/- DC provoked a more severe allergic inflammation in the lung than Nrf2+/+ DC in the same treatment group. We conclude that Nrf2 deficiency in DC may promote a constitutive immune-polarizing cytokine milieu, which we propose may have contributed to the augmented adjuvant effect of UFP on allergic sensitization.