Berenice Barajas

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 microm (PM(2.5)) 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 microm (ultrafine particles) with particles of <2.5 microm in genetically susceptible (apolipoprotein E-deficient) mice. These animals were exposed to concentrated ultrafine particles, concentrated particles of <2.5 microm, 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 PM(2.5) 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 (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.

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. CONCLUSIONS: Nrf2 expression unexpectedly promotes atherosclerotic lesion formation in a sex-dependent manner, most likely by a combination of systemic metabolic and local vascular effects.

BACKGROUND: Ambient 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. RESULTS: We 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. CONCLUSION: Diesel 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.