Search

Treatment failure due to variability in the pharmacologic response to drugs has more commonly been attributed to non-genetic factors than to prototypic pharmacogenetic mechanisms.1 Treatment failure on low-dose aspirin taken to prevent arterial thrombotic events, including myocardial infarction and stroke, is no exception. A closer look at aspirin’s human pharmacology reveals many potential sources of variability that are rooted in platelet biology, from aspirin insensitive mechanisms of activation to accelerated turnover.

The prostanoids, which include the prostaglandins (PGs) and thromboxane (Tx)A2, are a family of locally acting lipid mediators. Two isozymes, cyclooxygenase (COX)-1 and COX-2, of which only COX-1 is expressed in mature platelets, form an unstable intermediate product, PGH2, from arachidonic acid released from membrane phospholipids. At least nine PG synthases metabolize PGH2 further to five biologically active prostanoids, which each act through one or more specific G-protein coupled receptor.

Mantles of silt- and sand-size particles, paleosols and fluvial deposits preserve valuable information on Holocene environmental change. These archives were used to reconstruct the landscape history in the upper Orkhon Valley close to the former capitals of the Uighurs (Kharbalgasin Tuur) and the Mongolian Empire (Karakorum) near the recent town of Kharkhorin, Central Mongolia. A holistic approach involving the use of high spatial resolution geomorphological mapping, sedimentological and geochemical analysis, palynology, and geochronology shows several phases of landscape activity and stability in the region. This includes phases of fluvial erosion, aeolian sedimentation, and soil formation. By using luminescence and radiocarbon dating, phases of landscape change, indicated by soil formation, occurred mainly at around 6.5–6 ka. Pollen data and a weakly humic horizon at around 1.0 ka probably indicate enhanced moisture supply in the region and a reduction of human activity between the time of the reigns of the Uighurs (8th–9th century AD) and the Mongols (1220–1388 AD). Since 3 ka, especially within the last two millennia of Historical Time (300 B.C.–present), a more intensified human occupation in the Upper Orkhon Valley occurred in this region. This included a more densely grazing of cattle to supply the growing population demands of the Uighurs and Mongols. This overgrazing caused an increase in erosion and the formation and deepening of fluvial gullies, together with soil deflation and subsequent deposition of aeolian sediments. Human activity, in addition to climate, has been dominant in driving landscape evolution of this region since the late Holocene.

Suppression of prostacyclin (PGI 2 ) biosynthesis may explain the increased incidence of myocardial infarction and stroke which has been observed in placebo controlled trials of cyclooxygenase (COX)-2 inhibitors. Herein, we examine if COX-2–derived PGI 2 might condition the response of the vasculature to sustained physiologic stress in experimental models that retain endothelial integrity. Deletion of the PGI 2 receptor (IP) or suppression of PGI 2 with the selective COX-2 inhibitor, nimesulide, both augment intimal hyperplasia while preserving luminal geometry in mouse models of transplant arteriosclerosis or flow-induced vascular remodeling. Moreover, nimesulide or IP deletion augments the reduction in blood flow caused by common carotid artery ligation in wild-type mice. Generation of both thromboxane (Tx)A 2 and the isoprostane, 8, 12 – iso iPF 2α -VI, are increased in the setting of flow reduction and the latter increases further on administration of nimesulide. Deletion of the TxA 2 receptor (TP) reduces the hyperplastic response to nimesulide and carotid ligation, despite further augmentation of TP ligand production. Suppression of COX-2–derived PGI 2 or deletion of IP profoundly influences the architectural response of the vasculature to hemodynamic stress. Mechanism based vascular remodeling may interact with a predisposition to hypertension and atherosclerosis in contributing to the gradual transformation of cardiovascular risk during extended periods of treatment with selective inhibitors of COX-2.

Background— Antagonism or deletion of the receptor (the TP) for the cyclooxygenase (COX) product thromboxane (Tx)A 2 , retards atherogenesis in apolipoprotein E knockout (ApoE KO) mice. Although inhibition or deletion of COX-1 retards atherogenesis in ApoE and LDL receptor (LDLR) KOs, the role of COX-2 in atherogenesis remains controversial. Other products of COX-2, such as prostaglandin (PG) I 2 and PGE 2 , may both promote inflammation and restrain the effects of TxA 2 . Thus, combination with a TP antagonist might reveal an antiinflammatory effect of a COX-2 inhibitor in this disease. We addressed this issue and the role of TxA 2 in the promotion and regression of diffuse, established atherosclerosis in Apobec-1/LDLR double KOs (DKOs). Methods and Results— TP antagonism with S18886, but not combined inhibition of COX-1 and COX-2 with indomethacin or selective inhibition of COX-2 with Merck Frosst (MF) tricyclic, retards significantly atherogenesis in DKOs. Although indomethacin depressed urinary excretion of major metabolites of both TxA 2 , 2,3-dinor TxB 2 (Tx-M), and PGI 2 , 2,3-dinor 6-keto PGF 1α (PGI-M), only PGI-M was depressed by the COX-2 inhibitor. None of the treatments modified significantly the increase in lipid peroxidation during atherogenesis, reflected by urinary 8,12-iso-iPF 2α -VI. Combination with the COX-2 inhibitor failed to augment the impact of TP antagonism alone on lesion area. Rather, analysis of plaque morphology reflected changes consistent with destabilization of the lesion coincident with augmented formation of TxA 2 . Despite a marked effect on disease progression, TP antagonism failed to induce regression of established atherosclerotic disease in this model. Conclusions— TP antagonism is more effective than combined inhibition of COX-1 and COX-2 in retarding atherogenesis in Apobec-1/LDLR DKO mice, which perhaps reflects activation of the receptor by multiple ligands during disease initiation and early progression. Despite early intervention, selective inhibition of COX-2, alone or in combination with a TP antagonist, failed to modify disease progression but may undermine plaque stability when combined with the antagonist. TP antagonism failed to induce regression of established atherosclerotic disease. TP ligands, including COX-1 (but not COX-2)–derived TxA 2 , promote initiation and early progression of atherogenesis in Apobec-1/LDLR DKOs but appear unimportant in the maintenance of established disease.

Selective inhibitors of cyclooxygenase (COX)-2, the coxibs, were developed to inhibit inflammatory prostaglandins derived from COX-2, while sparing gastroprotective prostaglandins primarily formed by COX-1. However, COX-2-derived prostaglandins mediate not only pain and inflammation but also affect vascular function, the regulation of hemostasis/ thrombosis, and blood pressure control. All coxibs depress COX-2-dependent prostacyclin (PGI2) biosynthesis without effective suppression of platelet COX-1-derived thromboxane (Tx) A2, unlike aspirin or traditional nonsteroidal anti-inflammatory drugs, which inhibit both COX-1 and COX-2. The actions of PGI2 oppose mediators, which stimulate platelets, elevate blood pressure, and accelerate atherogenesis, including TxA2. Indeed, structurally distinct inhibitors of COX-2 have increased the likelihood of hypertension, myocardial infarction and stroke in controlled clinical trials. The detection of these events in patients is related to the duration of exposure and to their baseline risk of cardiovascular disease. Thus, coxibs should be withheld from patients with preexisting cardiovascular risk factors, and exposed patients at low cardiovascular baseline risk should be monitored for changes in their risk factor profile, such as increases in arterial blood pressure.

Female gender affords relative protection from cardiovascular disease until the menopause. We report that estrogen acts on estrogen receptor subtype alpha to up-regulate the production of atheroprotective prostacyclin, PGI 2 , by activation of cyclooxygenase 2 (COX-2). This mechanism restrained both oxidant stress and platelet activation that contribute to atherogenesis in female mice. Deletion of the PGI 2 receptor removed the atheroprotective effect of estrogen in ovariectomized female mice. This suggests that chronic treatment of patients with selective inhibitors of COX-2 could undermine protection from cardiovascular disease in premenopausal females.

The cardiovascular safety of nonsteroidal antiinflammatory drugs (NSAIDs) may be influenced by interactions with antiplatelet doses of aspirin. We sought to quantitate precisely the propensity of commonly consumed NSAIDs—ibuprofen, naproxen, and celecoxib—to cause a drug-drug interaction with aspirin in vivo by measuring the target engagement of aspirin directly by MS. We developed a novel assay of cyclooxygenase-1 (COX-1) acetylation in platelets isolated from volunteers who were administered aspirin and used conventional and microfluidic assays to evaluate platelet function. Although ibuprofen, naproxen, and celecoxib all had the potential to compete with the access of aspirin to the substrate binding channel of COX-1 in vitro, exposure of volunteers to a single therapeutic dose of each NSAID followed by 325 mg aspirin revealed a potent drug-drug interaction between ibuprofen and aspirin and between naproxen and aspirin but not between celecoxib and aspirin. The imprecision of estimates of aspirin consumption and the differential impact on the ability of aspirin to inactivate platelet COX-1 will confound head-to-head comparisons of distinct NSAIDs in ongoing clinical studies designed to measure their cardiovascular risk.

We thank the authors of the letters for their interest in our publication.1 True aspirin drug resistance would be expected to be (1) specific for aspirin, (2) reflected by incomplete inhibition of the molecular target of aspirin, and (3) a relatively stable phenotype that is reproducible when assessed on multiple occasions. It is this last point …

Aspirin prevents heart attack and stroke by irreversibly acetylating serine-529 in human platelet cyclooxygenase-1 (COX-1) and thereby inhibiting the biosynthesis of thromboxane (Tx). We hypothesized that precise quantification of COX-1 acetylation would allow the direct detection of drug-drug interactions between reversible COX inhibitors, nonsteroidal anti-inflammatory drugs (NSAIDs), and aspirin, which may abrogate its cardio-protective effect. We developed a stable isotope dilution liquid chromatography multiple reaction monitoring/mass spectrometry (LC-MRM/MS) method for measurement of platelet COX-1 acetylation. Serine-529 acetylation reflected faithfully aspirin pharmacology determined with conventional markers (serum TxB2 formation, urinary excretion of 11-dehydro-TxB2, and arachidonic acid induced platelet aggregation), but showed lower technical (CV% 3.7±2.7) and inter-individual variability (CV% 3.7). Oral administration of 325 mg of plain aspirin resulted in complete acetylation of platelet COX-1 within 2 hours and sustained maximal acetylation (71 ± 3% of total COX-1 amount) for up to two days in health volunteers (n=8). Recovery from acetylation, which reflects platelet turn-over, occurred at a rate of 7.2 ± 0.2 % per day. We compared the potential of five NSAIDs to block aspirin from acetylating platelet COX-1 in healthy volunteers (n=7 for each NSAID). Single doses of naproxen (500 mg), ibuprofen (600 mg), and diclofenac (100 mg) but not celecoxib (200 mg) and acetaminophen (1000 mg) prevented complete COX-1 acetylation by 325 mg aspirin administered 2 hours after the NSAID. The competition of NSAIDs with aspirin was conditioned by NSAID pharmacokinetics, potency, and COX isoform selectivity, suggesting variability in the likelihood of the drug-drug interaction between compounds and between patients. Monitoring platelet COX-1 acetylation may have utility in the identification of drug interactions that may limit the cardio-protective effect of low dose aspirin.

BACKGROUND Microfluidic devices can create hemodynamic conditions for platelet assays. We validated an 8-channel device in a study of interdonor response to acetylsalicylic acid (ASA, aspirin) with whole blood from 28 healthy individuals. METHODS Platelet deposition was assessed before treatment or 24 h after ingestion of 325 mg ASA. Whole blood (plus 100 μmol/L H-d-Phe-Pro-Arg-chloromethylketone to inhibit thrombin) was further treated ex vivo with ASA (0–500 μmol/L) and perfused over fibrillar collagen for 300 s at a venous wall shear rate (200 s−1). RESULTS Ex vivo ASA addition to blood drawn before aspirin ingestion caused a reduction in platelet deposition [half-maximal inhibitory concentration (IC50) approximately 10–20 μmol/L], especially between 150 and 300 s of perfusion, when secondary aggregation mediated by thromboxane was expected. Twenty-seven of 28 individuals displayed smaller deposits (45% mean reduction; range 10%–90%; P < 0.001) from blood obtained 24 h after ASA ingestion (no ASA added ex vivo). In replicate tests, an R value to score secondary aggregation [deposition rate from 150 to 300 s normalized by rate from 60 to 150 s] showed R < 1 in only 2 of 28 individuals without ASA ingestion, with R > 1 in only 3 of 28 individuals after 500 μmol/L ASA addition ex vivo. At 24 h after ASA ingestion, 21 of 28 individuals displayed poor secondary aggregation (R < 1) without ex vivo ASA addition, whereas the 7 individuals with residual secondary aggregation (R > 1) displayed insensitivity to ex vivo ASA addition. Platelet deposition was not correlated with platelet count. Ex vivo ASA addition caused similar inhibition at venous and arterial wall shear rates. CONCLUSIONS Microfluidic devices quantified platelet deposition after ingestion or ex vivo addition of aspirin.

Background— Low dose aspirin reduces the secondary incidence of myocardial infarction and stroke. Drug resistance to aspirin might result in treatment failure. Despite this concern, no clear definition of aspirin resistance has emerged, and estimates of its incidence have varied remarkably. We aimed to determine the commonality of a mechanistically consistent, stable, and specific phenotype of true pharmacological resistance to aspirin—such as might be explained by genetic causes. Methods and Results— Healthy volunteers (n=400) were screened for their response to a single oral dose of 325-mg immediate release or enteric coated aspirin. Response parameters reflected the activity of the molecular target of aspirin, cyclooxygenase-1. Individuals who appeared aspirin resistant on 1 occasion underwent repeat testing, and if still resistant were exposed to low-dose enteric coated aspirin (81 mg) and clopidogrel (75 mg) for 1 week each. Variable absorption caused a high frequency of apparent resistance to a single dose of 325-mg enteric coated aspirin (up to 49%) but not to immediate release aspirin (0%). All individuals responded to aspirin on repeated exposure, extension of the postdosing interval, or addition of aspirin to their platelets ex vivo. Conclusions— Pharmacological resistance to aspirin is rare; this study failed to identify a single case of true drug resistance. Pseudoresistance, reflecting delayed and reduced drug absorption, complicates enteric coated but not immediate release aspirin administration. Clinical Trial Registration— URL: http://www.clinicaltrials.gov . Unique identifier: NCT00948987.

Mit dem Standard Nr. 4 des DIIR wurde ein Rahmenwerk zur Prüfung von Projekten geschaffen. Der Leitfaden bietet Ihnen, basierend auf dem Standard Nr. 4 und internationalen Projektmanagement-Methoden wie PMBoK, PRINCE2 und ICB, einen konkreten und praxistauglichen Überblick zu: Prüfgebieten und -feldern, Prüfungsplanung und Ablauf einer projektbegleitenden Revision, Dokumentation der Prüfung sowie Best-Practice-Methoden in der Projektrevision. Die Darstellung des Audit Universe erleichtert Ihnen zudem die Projektrevision in der Praxis und hilft, projektinhärente Risiken für das Unternehmen zu reduzieren. Der Leitfaden ist branchenübergreifend für alle Arten von Projekten geeignet.

The 12/15-lipoxygenase (LO) cascade governs the generation of 12-hydroperoxy-eicosatetraenoic acid (HPETE) and 15-HPETE from arachidonic acid. The 5-LO pathway plays a fundamental role in the biosynthesis of leukotrienes, essential inflammatory lipid mediators. Cyclooxygenase (COX)-1 and -2 biosynthetic pathways are responsible for prostaglandin and thromboxane formation. Experimental investigations in animal models using 12/15-LO deficient mice, 12/15-LO or 15-LO transgenic mice, or pharmacological 15-LO inhibition have all demonstrated the essential role of 12/15-LO in atherogenesis. The underlying mechanisms are linked to low-density lipoprotein oxidation, pro-inflammatory Th1 cytokine production and enhanced monocyte-endothelial cell interaction. Human genetic studies as well as disruption of the 5-LO gene in mouse models of hyperlipidemia revealed that 5-LO and 5-LO-activating protein are associated with risks of human cardiovascular disease, and that this cascade plays an important role in aortic aneurysm pathogenesis through leukotriene-mediated inflammatory chemokine production. COX-1 plays an active role in atherogenesis via thromboxane A(2), while COX-2-derived prostaglandin (PGI(2)) protects against atherosclerosis in murine models. Recent data demonstrated that selective inhibition of COX-2 augments the risk of cardiovascular events in patients. Selective inhibition or blockade of selective components in these two enzymatic pathways through systemic drug delivery or medical device approaches (e.g., drug-eluting stents) may have therapeutic benefit against certain cardiovascular diseases.

Isoprostanes (iPs) are prostaglandin (PG) isomers generated by free radical-catalyzed peroxidation of polyunsaturated fatty acids (PUFAs). Urinary F(2)-iPs, PGF(2alpha) isomers derived from arachidonic acid (AA) are used as indices of lipid peroxidation in vivo. We now report the characterization of two major F(3)-iPs, 5-epi-8,12-iso-iPF(3alpha)-VI and 8,12-iso-iPF(3alpha)-VI, derived from the omega-3 fatty acid, eicosapentaenoic acid (EPA). Although the potential therapeutic benefits of EPA receive much attention, a shift toward a diet rich in omega-3 PUFAs may also predispose to enhanced lipid peroxidation. Urinary 5-epi-8,12-iso-iPF(3alpha)-VI and 8,12-iso-iPF(3alpha)-VI are highly correlated and unaltered by cyclooxygenase inhibition in humans. Fish oil dose-dependently elevates urinary F(3)-iPs in mice and a shift in dietary omega-3/omega-6 PUFAs is reflected by an increasing slope [m] of the line relating urinary 8, 12-iso-iPF(3alpha)-VI and 8,12-iso-iPF(2alpha)-VI. Administration of bacterial lipopolysaccharide evokes a reversible increase in both urinary 8,12-iso-iPF(3alpha)-VI and 8,12-iso-iPF(2alpha)-VI in humans on an ad lib diet. However, while excretion of the iPs is highly correlated (R(2) median = 0.8), [m] varies by an order of magnitude, reflecting marked inter-individual variability in the relative peroxidation of omega-3 versus omega-6 substrates. Clustered analysis of F(2)- and F(3)-iPs refines assessment of the oxidant stress response to an inflammatory stimulus in vivo by integrating variability in dietary intake of omega-3/omega-6 PUFAs.

Hyaluronan (HA) is a key molecule of the extracellular matrix that is thought to be critically involved in both atherosclerosis and restenosis. Recently, it has been demonstrated that the cyclooxygenase (COX) products, prostacyclin and prostaglandin E(2), induce HA synthesis in vitro by transcriptional up-regulation of HA-synthase 2 (HAS2) and HAS1. The relative roles in atherosclerotic and restenotic artery disease of tissue specifically expressed COX-1 and COX-2 are still under debate. Thus, the present study aimed to investigate the effect of COX isoform inhibition on HA-accumulation and regulation of HAS isoform expression in two models of pathologic artery remodelling in vivo. Firstly, ApoE-deficient mice were treated with a prototypic isoform non-selective inhibitor, indomethacin or with a prototypic COX-2 selective inhibitor, rofecoxib, for 8 weeks. Aortic HAS mRNA expression and HA-accumulation in atherosclerotic aortic root lesions were analyzed. Secondly, neointimal hyperplasia was induced by carotid artery ligation in ApoE-deficient mice on a high fat diet and the effects of the COX inhibitors were determined after 4 weeks of treatment. Intimal HA-accumulation was markedly reduced in both models by indomethacin and rofecoxib. This coincided with a strong inhibition of HAS1 mRNA expression in both models and with decreased HAS2 mRNA in the aorta of ApoE-deficient mice. HAS3 was not affected. The repression of HA-accumulation by both COX-2 selective and non-selective COX inhibition implicates COX-2 in the regulation of HA synthesis via stimulation of HAS1 and HAS2 expression in vivo. Modulation of vascular HA-accumulation might play a role in chronic effects of COX inhibitors on the progression of atherosclerosis.

Prostaglandin D(2) (PGD(2)) is a cyclooxygenase (COX) product of arachidonic acid that activates D prostanoid receptors to modulate vascular, platelet, and leukocyte function in vitro. However, little is known about its enzymatic origin or its formation in vivo in cardiovascular or inflammatory disease. 11,15-dioxo-9alpha-hydroxy-2,3,4,5-tetranorprostan-1,20-dioic acid (tetranor PGDM) was identified by mass spectrometry as a metabolite of infused PGD(2) that is detectable in mouse and human urine. Using liquid chromatography-tandem mass spectrometry, tetranor PGDM was much more abundant than the PGD(2) metabolites, 11beta-PGF(2alpha) and 2,3-dinor-11beta-PGF(2alpha), in human urine and was the only endogenous metabolite detectable in mouse urine. Infusion of PGD(2) dose dependently increased urinary tetranor PGDM2,3-dinor-11beta-PGF(2alpha)11beta-PGF(2alpha) in mice. Deletion of either lipocalin-type or hemopoietic PGD synthase enzymes decreased urinary tetranor PGDM. Deletion or knockdown of COX-1, but not deletion of COX-2, decreased urinary tetranor PGDM in mice. Correspondingly, both PGDM and 2,3-dinor-11beta-PGF(2alpha) were suppressed by inhibition of COX-1 and COX-2, but not by selective inhibition of COX-2 in humans. PGD(2) has been implicated in both the development and resolution of inflammation. Administration of bacterial lipopolysaccharide coordinately elevated tetranor PGDM and 2,3-dinor-11beta-PGF(2alpha) in volunteers, coincident with a pyrexial and systemic inflammatory response, but both metabolites fell during the resolution phase. Niacin increased tetranor PGDM and 2,3-dinor-11beta-PGF(2alpha) in humans coincident with facial flushing. Tetranor PGDM is an abundant metabolite in urine that reflects modulated biosynthesis of PGD(2) in humans and mice.

A short-lived lipid mediator, thromboxane (Tx) A2, is synthesized by activated platelets and released as a local signal, which amplifies activation and recruits additional platelets to the site of clot formation1,2 TxA2 is also a potent vasoconstrictor and stimulates mitogenesis, accelerating hemostasis and the proliferative response to vascular injury.3-6 Derived from arachidonic acid (AA) by the sequential action of prostaglandin G/H synthase (PGHS)-1 and thromboxane synthase (TxAS) (Fig. 31-1), TxA2 targets a specific G protein-coupled transmembrane thromboxane receptor (TP) that signals to facilitate dense granule secretion and integrin αIIbβ3 activation in platelets. Patients deficient in TxA2 formation have a mild bleeding disorder,7-16 as have those with a defective response to TxA2, 17-19 such as an arginine60 to leucine mutation of the TP which disrupts receptor-G protein interactions. 20

Background & Aims: Variability in response to drugs may influence both efficacy and safety. Cyclooxygenase (COX)-2 inhibitors pose a cardiovascular risk by potentially increasing the likelihood of thrombosis, hypertension, and atherogenesis. Differences between individuals in the response to COX-2 inhibitors would be expected to influence their susceptibility to cardiovascular complications. We examined the variability in degree and selectivity of COX-2 inhibition in humans in response to celecoxib and rofecoxib. Methods: Fifty healthy volunteers received placebo, rofecoxib (25 mg), and celecoxib (200 mg), randomized by order. COX-1 and COX-2 inhibition was determined using ex vivo and in vivo indices of enzymatic activity. A subset of 5 individuals underwent 5 replicate studies to estimate variability in drug response both within and between subjects. Results: Despite the higher COX-2 selectivity of rofecoxib in vitro, the average selectivity attained by 25 mg rofecoxib and 200 mg celecoxib in vivo were not different. However, there was considerable variability at an individual level in the degree of COX-2 inhibition and selectivity attained by both drugs. Approximately one third of the variability was attributable to differences between individuals, suggesting the contribution of genetic sources of variance, such as candidate polymorphisms detected in COX-1 and CYP2C9. Conclusions: The actual degree of selectivity for inhibition of COX-2 achieved by the coxibs relates both to chemical properties of the drug and to factors within an individual that modulate drug response. These sources of variability might be exploited to identify patients uniquely susceptible to benefit or at developing risk of cardiovascular complications.

Before menopause, women are relatively protected against cardiovascular disease. Although estrogen (E 2 ) retards atherogenesis in animal models and improves endothelial function in women with hyperlipidemia, the mechanisms of atheroprotection remain to be defined completely. The prostaglandin PGI 2 inhibits platelet activation, vascular smooth muscle contraction, leukocyte-endothelial cell interactions, and cholesteryl ester hydrolase. PGI 2 analogs limit atherogenesis, and atherosclerotic lesions have a reduced ability to produce PGI 2 . Cyclooxygenase 2 (COX-2), expressed in vascular endothelial cells, catalyzes the formation of prostaglandin endoperoxide from arachidonic acid, and this subsequently is transformed to PgI 2 in endothelial cells by the action of PGI 2 synthase. Selective COX-2 inhibitors retard the excretion of PGI 2 metabolites. These mechanisms may be relevant to atheroprotection in women because E 2 increases the expression of COX-2 in vascular tissues and promotes the in vitro production of PGI 2 . Male mice lacking low-density lipoprotein receptor (LDLR) developed atherosclerosis more rapidly than did females, despite the lack of differences in total plasma cholesterol or high-density lipoprotein cholesterol. Female mice lacking both PGI 2 receptor and the LDLR developed more prominent aortic lesions than did LDLR-knockout females (ie, lacking the gene for the LDL receptor). This effect was not gene dose-dependent and was not apparent in male mice. Further observations indicated that PGI 2 decreases the platelet activation that characterizes early atherogenesis in female mice. It appears that PGI 2 serves as an antioxidant in female LDLR-knockout mice before atherogenesis and at its earliest stage. Adding hydrogen peroxide to cultured mouse aortic smooth muscle cells increased synthesis of PGI 2 and lipid peroxidation. There was no change in the expression of COX-2. Peroxide increased reactive oxygen species, and this effect was augmented by the lack of PGI 2 receptor expression. These observations suggest that the PGI 2 receptor modulates oxidant stress under baseline conditions. Importantly, E 2 stimulated COX-2 expression and PGI 2 formation. These findings suggest that E 2 receptor alpha-mediated COX-2 stimulation of PGI 2 makes a substantial contribution to atheroprotection in female LDLR-knockout mice. It is possible that ongoing treatment of patients with selective COX-2 inhibitors might undermine protection from cardiovascular disease in premenopausal females.