Transient increase in atherosclerotic plaque macrophage content following Streptococcus pneumoniae pneumonia in ApoE-deficient mice.

Introduction: Despite epidemiological associations between community acquired pneumonia (CAP) and myocardial infarction, mechanisms that modify cardiovascular disease during CAP are not well defined. In particular, largely due to a lack of relevant experimental models, the effect of pneumonia on atherosclerotic plaques is unclear. We describe the development of a murine model of the commonest cause of CAP, Streptococcus pneumoniae pneumonia, on a background of established atherosclerosis. We go on to use our model to investigate the effects of pneumococcal pneumonia on atherosclerosis. Methods: C57BL/6J and ApoE-/- mice were fed a high fat diet to promote atherosclerotic plaque formation. Mice were then infected with a range of S. pneumoniae serotypes (1, 4 or 14) with the aim of establishing a model to study atherosclerotic plaque evolution after pneumonia and bacteremia. Laser capture microdissection of plaque macrophages enabled transcriptomic analysis. Results: Intratracheal instillation of S. pneumoniae in mice fed a cholate containing diet resulted in low survival rates following infection, suggestive of increased susceptibility to severe infection. Optimization steps resulted in a final model of male ApoE-/- mice fed a Western diet then infected by intranasal instillation of serotype 4 (TIGR4) S. pneumoniae followed by antibiotic administration. This protocol resulted in high rates of bacteremia (88.9%) and survival (88.5%). Pneumonia resulted in increased aortic sinus plaque macrophage content 2 weeks post pneumonia but not at 8 weeks, and no difference in plaque burden or other plaque vulnerability markers were found at either time point. Microarray and qPCR analysis of plaque macrophages identified downregulation of two E3 ubiquitin ligases, Huwe1 and Itch, following pneumonia. Treatment with atorvastatin failed to alter plaque macrophage content or other plaque features. Discussion: Without antibiotics, ApoE-/- mice fed a high fat diet were highly susceptible to mortality following S. pneumoniae infection. The major infection associated change in plaque morphology was an early increase in plaque macrophages. Our results also hint at a role for the ubiquitin proteasome system in the response to pneumococcal infection in the plaque microenvironment.

OxLDL induces the release of IL-1ß from primed human endothelial and smooth muscle cells via different caspase -1-dependent mechanisms.

Atherosclerosis is characterised by abnormal lipid and cell accumulation within arterial layers that leads to disturbed blood flow. Modified cholesterol forms such as oxidised low-density lipoprotein (oxLDL) enter cells altering their phenotype, triggering over-exuberant repair and arterial occlusion, myocardial infarction or stroke. We hypothesised that oxLDL enters vascular wall cells and induces interleukin-1ß (IL-1ß) secretion, potentially via a caspase-1/NLRP3 mechanism. Human coronary artery endothelial cells (HCAEC) and smooth muscle cells (HCASMC), isolated from different donors, were cultured and stimulated (primed) with pro-inflammatory cytokines TNFα and IL-1α (10 ng/mL each, for 48 h), followed by incubation with human oxLDL (10-50 ug/mL) for up to 6 h. Inhibitors of caspase-1 (YVAD), NLRP3 (MCC950) and gasdermin D (disulfiram) were added 1 h before oxLDL. Cell lysates and culture supernatants were collected and analysed for IL-1ß using ELISA. Microscopy imaging showed oxLDL entered stimulated cells and formed particles. OxLDL at 20 and 50 ug/mL induced the maximum release of IL-1ß from stimulated HCASMCs and HCAECs, respectively, compared to control. Inhibition of either NLRP3, caspase-1 or gasdermin D significantly reduced the release of IL-1ß (4-fold, P < 0.0001; 14-fold, P < 0.0001, 1.5-fold, P < 0.0003, respectively) in HCAEC. In contrast, in HCASMCs, only caspase-1 inhibition reduced the release of IL-1ß (2.1-fold, P < 0.0001). HCAECs and HCASMCs elicited the release of IL-1ß in response to the same stimulus via different mechanisms. In HCAECs, released IL-1ß potentially exits via a GSDMD-induced membrane pore. These data suggest that caspase-1 or gasdermin D inhibition is likely to be effective vessel wall cell-specific strategies for the reduction of atherosclerosis.

Carbon Monoxide Releasing Molecule A1 Reduces Myocardial Damage After Acute Myocardial Infarction in a Porcine Model.

ABSTRACT: Infarct size is a major determinant of outcomes after acute myocardial infarction (AMI). Carbon monoxide-releasing molecules (CORMs), which deliver nanomolar concentrations of carbon monoxide to tissues, have been shown to reduce infarct size in rodents. We evaluated efficacy and safety of CORM-A1 to reduce infarct size in a clinically relevant porcine model of AMI. We induced AMI in Yorkshire White pigs by inflating a coronary angioplasty balloon to completely occlude the left anterior descending artery for 60 minutes, followed by deflation of the balloon to mimic reperfusion. Fifteen minutes after balloon occlusion, animals were given an infusion of 4.27 mM CORM-A1 (n = 7) or sodium borate control (n = 6) over 60 minutes. Infarct size, cardiac biomarkers, ejection fraction, and hepatic and renal function were compared amongst the groups. Immunohistochemical analyses were performed to compare inflammation, cell proliferation, and apoptosis between the groups. CORM-A1-treated animals had significant reduction in absolute infarct area (158 ± 16 vs. 510 ± 91 mm2, P < 0.001) and infarct area corrected for area at risk (24.8% ± 2.6% vs. 45.2% ± 4.0%, P < 0.0001). Biochemical markers of myocardial injury also tended to be lower and left ventricular function tended to recover better in the CORM-A1 treated group. There was no evidence of hepatic or renal toxicity with the doses used. The cardioprotective effects of CORM-A1 were associated with a significant reduction in cell proliferation and inflammation. CORM-A1 reduces infarct size and improves left ventricular remodeling and function in a porcine model of reperfused MI by a reduction in inflammation. These potential cardioprotective effects of CORMs warrant further translational investigations.

Dietary Docosahexaenoic Acid Reduces Oscillatory Wall Shear Stress, Atherosclerosis, and Hypertension, Most Likely Mediated via an IL-1-Mediated Mechanism.

BACKGROUND: Hypertension is a complex condition and a common cardiovascular risk factor. Dietary docosahexaenoic acid (DHA) modulates atherosclerosis and hypertension, possibly via an inflammatory mechanism. IL-1 (interleukin 1) has an established role in atherosclerosis and inflammation, although whether IL-1 inhibition modulates blood pressure is unclear. METHODS AND RESULTS: Male apoE-/- (apolipoprotein E-null) mice were fed either a high fat diet or a high fat diet plus DHA (300 mg/kg per day) for 12 weeks. Blood pressure and cardiac function were assessed, and effects of DHA on wall shear stress and atherosclerosis were determined. DHA supplementation improved left ventricular function, reduced wall shear stress and oscillatory shear at ostia in the descending aorta, and significantly lowered blood pressure compared with controls (119.5±7 versus 159.7±3 mm Hg, P<0.001, n=4 per group). Analysis of atheroma following DHA feeding in mice demonstrated a 4-fold reduction in lesion burden in distal aortas and in brachiocephalic arteries (P<0.001, n=12 per group). In addition, DHA treatment selectively decreased plaque endothelial IL-1ß (P<0.01). CONCLUSIONS: Our findings revealed that raised blood pressure can be reduced by inhibiting IL-1 indirectly by administration of DHA in the diet through a mechanism that involves a reduction in wall shear stress and local expression of the proinflammatory cytokine IL-1ß.

Selective improvement of pulmonary arterial hypertension with a dual ETA/ETB receptors antagonist in the apolipoprotein E-/- model of PAH and atherosclerosis.

Idiopathic pulmonary arterial hypertension (IPAH) is increasingly diagnosed in elderly patients who also have an increased risk of co-morbid atherosclerosis. Apolipoprotein E-deficient (ApoE-/-) mice develop atherosclerosis with severe PAH when fed a high-fat diet (HFD) and have increased levels of endothelin (ET)-1. ET-1 receptor antagonists (ERAs) are used for the treatment of PAH but less is known about whether ERAs are beneficial in atherosclerosis. We therefore examined whether treatment of HFD-ApoE-/- mice with macitentan, a dual ETA/ETB receptor antagonist, would have any effect on both atherosclerosis and PAH. ApoE-/- mice were fed chow or HFD for eight weeks. After four weeks of HFD, mice were randomized to a four-week treatment of macitentan by food (30 mg/kg/day dual ETA/ETB antagonist), or placebo groups. Echocardiography and closed-chest right heart catheterization were used to determine PAH phenotype and serum samples were collected for cytokine analysis. Thoracic aortas were harvested to assess vascular reactivity using wire myography, and histological analyses were performed on the brachiocephalic artery and aortic root to assess atherosclerotic burden. Macitentan treatment of HFD-fed ApoE-/- mice was associated with a beneficial effect on the PAH phenotype and led to an increase in endothelial-dependent relaxation in thoracic aortae. Macitentan treatment was also associated with a significant reduction in interleukin 6 (IL-6) concentration but there was no significant effect on atherosclerotic burden. Dual blockade of ETA/ETB receptors improves endothelial function and improves experimental PAH but had no significant effect on atherosclerosis.

Role of Animal Models in Coronary Stenting.

Coronary angioplasty initially employed balloon dilatation only. This technique revolutionized the treatment of coronary artery disease, although outcomes were compromised by acute vessel closure, late constrictive remodeling, and restenosis due to neointimal proliferation. These processes were studied in animal models, which contributed to understanding the biology of endovascular arterial injury. Coronary stents overcome acute recoil, with improvements in the design and metallurgy since then, leading to the development of drug-eluting stents and bioresorbable scaffolds. These devices now undergo computer modeling and benchtop and animal testing before evaluation in clinical trials. Animal models, including rabbit, sheep, dog and pig are available, all with individual benefits and limitations. In smaller mammals, such as mouse and rabbit, the target for stenting is generally the aorta; whereas in larger animals, such as the pig, it is generally the coronary artery. The pig coronary stenting model is a gold-standard for evaluating safety; but insights into biomechanical properties, the biology of stenting, and efficacy in controlling neointimal proliferation can also be gained. Intra-coronary imaging modalities such as intravascular ultrasound and optical coherence tomography allow precise serial evaluation in vivo, and recent developments in genetically modified animal models of atherosclerosis provide realistic test beds for future stents and scaffolds.

Neutrophil elastase promotes interleukin-1ß secretion from human coronary endothelium.

The endothelium is critically involved in the pathogenesis of atherosclerosis by producing pro-inflammatory mediators, including IL-1ß. Coronary arteries from patients with ischemic heart disease express large amounts of IL-1ß in the endothelium. However, the mechanism by which endothelial cells (ECs) release IL-1ß remains to be elucidated. We investigated neutrophil elastase (NE), a potent serine protease detected in vulnerable areas of human carotid plaques, as a potential "trigger" for IL-1ß processing and release. This study tested the hypothesis that NE potentiates the processing and release of IL-1ß from human coronary endothelium. We found that NE cleaves the pro-isoform of IL-1ß in ECs and causes significant secretion of bioactive IL-1ß via extracellular vesicles. This release was attenuated significantly by inhibition of neutrophil elastase but not caspase-1. Transient increases in intracellular Ca(2+) levels were observed prior to secretion. Inside ECs, and after NE treatment only, IL-1ß was detected within LAMP-1-positive multivesicular bodies. The released vesicles contained bioactive IL-1ß. In vivo, in experimental atherosclerosis, NE was detected in mature atherosclerotic plaques, predominantly in the endothelium, alongside IL-1ß. This study reveals a novel mechanistic link between NE expression in atherosclerotic plaques and concomitant pro-inflammatory bioactive IL-1ß secretion from ECs. This could reveal additional potential anti-IL-1ß therapeutic targets and provide further insights into the inflammatory process by which vascular disease develops.

Streptococcus pneumoniae worsens cerebral ischemia via interleukin 1 and platelet glycoprotein Ibα.

OBJECTIVE: Bacterial infection contributes to diverse noninfectious diseases and worsens outcome after stroke. Streptococcus pneumoniae, the most common infection in patients at risk of stroke, is a major cause of prolonged hospitalization and death of stroke patients, but how infection impacts clinical outcome is not known. METHODS: We induced sustained pulmonary infection by a human S. pneumoniae isolate in naive and comorbid rodents to investigate the effect of infection on vascular and inflammatory responses prior to and after cerebral ischemia. RESULTS: S. pneumoniae infection triggered atherogenesis, led to systemic induction of interleukin (IL) 1, and profoundly exacerbated (50-90%) ischemic brain injury in rats and mice, a response that was more severe in combination with old age and atherosclerosis. Systemic blockade of IL-1 with IL-1 receptor antagonist (IL-1Ra) fully reversed infection-induced exacerbation of brain injury and functional impairment caused by cerebral ischemia. We show that infection-induced systemic inflammation mediates its effects via increasing platelet activation and microvascular coagulation in the brain after cerebral ischemia, as confirmed by reduced brain injury in response to blockade of platelet glycoprotein (GP) Ibα. IL-1 and platelet-mediated signals converge on microglia, as both IL-1Ra and GPIbα blockade reversed the production of IL-1α by microglia in response to cerebral ischemia in infected animals. INTERPRETATION: S. pneumoniae infection augments atherosclerosis and exacerbates ischemic brain injury via IL-1 and platelet-mediated systemic inflammation. These mechanisms may contribute to diverse cardio- and cerebrovascular pathologies in humans.

A social communication intervention to increase validating comments by children with language impairment.

PURPOSE: Four children identified with language impairment (LI) participated in a social communication intervention to increase the production of validating comments, including making positive statements, sharing information, and asking peers questions about themselves. METHOD: A case study design was used. Baseline measures were collected from 3 cooperative learning sessions for each participant. The intervention lasted 10 weeks, with 40 (15-min) sessions for 3 of the children and 20 (30-min) sessions for the remaining child. Each week, participants took part in sessions of group instruction, novel peer play, and review with the clinician. Data monitoring the production of validating comments were taken from the novel peer play interactions. RESULTS: One child produced a notable increase in validating comments during intervention, and 2 other participants produced more modest improvement. All 3 maintained these gains in the follow-up sessions. The remaining participant produced little change from baseline during the intervention. With respect to social outcomes, changes were not noted in peer acceptance and friendship. Teachers reported notable improvement in the sociable behavior of 2 of the children. CONCLUSION: The 4 participants showed varied increases in the production of validating comments. Possible factors influencing the successful application of the intervention are discussed.

Interleukin-1 mediates neuroinflammatory changes associated with diet-induced atherosclerosis.

BACKGROUND: Systemic inflammation contributes to brain pathology in cerebrovascular disease through mechanisms that are poorly understood. METHODS AND RESULTS: Here we show that atherosclerosis, a major systemic inflammatory disease, is associated with severe cerebrovascular inflammation in mice and that this effect is mediated by the proinflammatory cytokine interleukin-1 (IL-1). Apolipoprotein E-deficient mice fed Paigen or Western diets develop vascular inflammation, microglial activation, and leukocyte recruitment in the brain, which are absent in apolipoprotein E-deficient mice crossed with IL-1 type 1 receptor-deficient mice. Systemic neutralization of IL-1ß with an anti-IL-1ß antibody reversed aortic plaque formation (by 34% after a Paigen and 45% after a Western diet) and reduced inflammatory cytokine expression in peripheral organs. Central, lipid accumulation-associated leukocyte infiltration into the choroid plexus was reversed by IL-1ß antibody administration. Animals fed a Western diet showed 57% lower vascular inflammation in the brain than that of mice fed a Paigen diet, and this was reduced further by 24% after IL-1ß antibody administration. CONCLUSIONS: These results indicate that IL-1 is a key driver of systemically mediated cerebrovascular inflammation and that interventions against IL-1ß could be therapeutically useful in atherosclerosis, dementia, or stroke. (J Am Heart Assoc. 2012;1:e002006 doi: 10.1161/JAHA.112.002006.).

Inhibition of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) reverses experimental pulmonary hypertension.

Pulmonary arterial hypertension (PAH) is a life-threatening disease characterized by the progressive narrowing and occlusion of small pulmonary arteries. Current therapies fail to fully reverse this vascular remodeling. Identifying key pathways in disease pathogenesis is therefore required for the development of new-targeted therapeutics. We have previously reported tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) immunoreactivity within pulmonary vascular lesions from patients with idiopathic PAH and animal models. Because TRAIL can induce both endothelial cell apoptosis and smooth muscle cell proliferation in the systemic circulation, we hypothesized that TRAIL is an important mediator in the pathogenesis of PAH. We demonstrate for the first time that TRAIL is a potent stimulus for pulmonary vascular remodeling in human cells and rodent models. Furthermore, antibody blockade or genetic deletion of TRAIL prevents the development of PAH in three independent rodent models. Finally, anti-TRAIL antibody treatment of rodents with established PAH reverses pulmonary vascular remodeling by reducing proliferation and inducing apoptosis, improves hemodynamic indices, and significantly increases survival. These preclinical investigations are the first to demonstrate the importance of TRAIL in PAH pathogenesis and highlight its potential as a novel therapeutic target to direct future translational therapies.

Paigen diet-fed apolipoprotein E knockout mice develop severe pulmonary hypertension in an interleukin-1-dependent manner.

Inflammatory mechanisms are proposed to play a significant role in the pathogenesis of pulmonary arterial hypertension (PAH). Previous studies have described PAH in fat-fed apolipoprotein E knockout (ApoE(-/-)) mice. We have reported that signaling in interleukin-1-receptor-knockout (IL-1R1(-/-)) mice leads to a reduction in diet-induced systemic atherosclerosis. We subsequently hypothesized that double-null (ApoE(-/-)/IL-1R1(-/-)) mice would show a reduced PAH phenotype compared with that of ApoE(-/-) mice. Male IL-1R1(-/-), ApoE(-/-), and ApoE(-/-)/IL-1R1(-/-) mice were fed regular chow or a high-fat diet (Paigen diet) for 8 weeks before phenotyping for PAH. No abnormal phenotype was observed in the IL-1R1(-/-) mice. Fat-fed ApoE(-/-) mice developed significantly increased right ventricular systolic pressure and substantial pulmonary vascular remodeling. Surprisingly, ApoE(-/-)/IL-1R1(-/-) mice showed an even more severe PAH phenotype. Further molecular investigation revealed the expression of a putative, alternatively primed IL-1R1 transcript expressed within the lungs but not aorta of ApoE(-/-)/IL-1R1(-/-) mice. Treatment of ApoE(-/-) and ApoE(-/-)/IL-1R1(-/-) mice with IL-1-receptor antagonist prevented progression of the PAH phenotype in both strains. Blocking IL-1 signaling may have beneficial effects in treating PAH, and alternative IL-1-receptor signaling in the lung may be important in driving PAH pathogenesis.

Dietary phosphate modulates atherogenesis and insulin resistance in apolipoprotein E knockout mice--brief report.

OBJECTIVE: Epidemiological studies link higher serum phosphate and the phosphatonin fibroblast growth factor 23 with cardiovascular events and atheroma, and they link lower serum phosphate with insulin resistance and the metabolic syndrome. We investigated whether manipulating dietary phosphate influences atherogenesis or insulin sensitivity in mice. METHODS AND RESULTS: Apolipoprotein E knockout mice were fed an atherogenic diet with low (0.2%), standard (0.6%), or high (1.6%) phosphate content. Serum phosphate and fibroblast growth factor 23 significantly increased with increasing dietary phosphate intake, but lipid profile and blood pressure were unaffected. After 20 weeks, mice on the higher phosphate diet had significantly more atheroma at the aortic sinus (42±1.9% versus 30±1.5% for high versus low phosphate, P<0.01). Compared with standard and high-phosphate diet groups, mice on a low-phosphate diet had more adipose tissue and a 4-fold increase in insulin resistance measured by homeostatic model assessment (43.7±9.3 versus 8.9±0.7 for low versus high phosphate, P<0.005). CONCLUSIONS: A high-phosphate diet accelerates atherogenesis in apolipoprotein E(-/-) mice, whereas low phosphate intake induces insulin resistance. These data indicate for the first time that controlling dietary phosphate intake may influence development of both atherosclerosis and the metabolic syndrome.

TRAIL attenuates the development of atherosclerosis in apolipoprotein E deficient mice.

TRAIL (tumour necrosis factor-related apoptosis inducing ligand) is most often reported to induce apoptosis in tumour cells. It is expressed in artery walls but its role and regulation in vascular pathologies is little studied. We aimed to measure the effect of genetic deletion of TRAIL on atherosclerosis in a mouse model. TRAIL was mainly expressed in endothelium, smooth muscle cells and macrophages within plaques. The absence of TRAIL in chow and in fat-fed mice led to greater lesion coverage in aortae (8 weeks, % area ± SEM), n=7-8, 1.24 ± 0.2 (no TRAIL, chow diet) vs. 0.42 ± 0.1, p<0.01 and 3.4 ± 0.8 (no TRAIL, Western diet) vs. 0.94 ± 0.2, p<0.01 and larger, smooth muscle cell rich lesions at aortic roots than control mice (8 weeks, mean lesion area/total cross sectional area ± SEM, n=7-8, 0.17 ± 0.01 (no TRAIL, chow diet) vs. 0.135 ± 0.006, p<0.05 and 0.36 ± 0.03 (no TRAIL, Western diet) vs. 0.23 ± 0.02, p<0.05) particularly at early time points. The larger early lesions appeared to be as a result of increased smooth muscle cells in lesions of TRAIL deficient, pro-atherosclerotic animals. We conclude that TRAIL attenuates plaque size at early stages of atherosclerosis.

A novel mouse model of in situ stenting.

AIMS: Animal models of stenting are mostly limited to larger animals or involve substantial abdominal surgery in rodents. We aimed to develop a simple, direct model of murine stenting. METHODS AND RESULTS: We designed a miniature, self-expanding, nitinol wire coil stent that was pre-loaded into a metal stent sheath. This was advanced into the abdominal aorta of the mouse, via femoral access, and the stent deployed. In-stent restenosis was investigated at 1, 3, 7, and 28 days post-stenting. The model was validated by investigation of neointima formation in mice deficient in signalling via the interleukin-1 receptor (IL-1R1), compared with other injury models. Ninety-two per cent of mice undergoing the procedure were successfully stented. All stented vessels were patent. Inflammatory cells were seen in the adventitia and around the stent strut up to 3 days post-stenting. At 3 days, an early neointima was present, building to a mature neointima at 28 days. In mice lacking IL-1R1, the neointima was 64% smaller than that in wild-type controls at the 28-day timepoint, in agreement with other models. CONCLUSION: This is the first description of a successful model of murine in situ stenting, using a stent specifically tailored for use in small thin-walled arteries. The procedure can be undertaken by a single operator without the need for an advanced level of microsurgical skill and is reliable and reproducible. The utility of this model is demonstrated by a reduction in in-stent restenosis in IL-1R1-deficient mice.

Interleukin-1 regulates multiple atherogenic mechanisms in response to fat feeding.

BACKGROUND: Atherosclerosis is an inflammatory process that develops in individuals with known risk factors that include hypertension and hyperlipidaemia, influenced by diet. However, the interplay between diet, inflammatory mechanisms and vascular risk factors requires further research. We hypothesised that interleukin-1 (IL-1) signaling in the vessel wall would raise arterial blood pressure and promote atheroma. METHODOLOGY/PRINCIPAL FINDINGS: Apoe(-/-) and Apoe(-/-)/IL-1R1(-/-) mice were fed high fat diets for 8 weeks, and their blood pressure and atherosclerosis development measured. Apoe(-/-)/IL-R1(-/-) mice had a reduced blood pressure and significantly less atheroma than Apoe(-/-) mice. Selective loss of IL-1 signaling in the vessel wall by bone marrow transplantation also reduced plaque burden (p < 0.05). This was associated with an IL-1 mediated loss of endothelium-dependent relaxation and an increase in vessel wall Nox 4. Inhibition of IL-1 restored endothelium-dependent vasodilatation and reduced levels of arterial oxidative stress. CONCLUSIONS/SIGNIFICANCE: The IL-1 cytokine system links atherogenic environmental stimuli with arterial inflammation, oxidative stress, increased blood pressure and atherosclerosis. This is the first demonstration that inhibition of a single cytokine can block the rise in blood pressure in response to an environmental stimulus. IL-1 inhibition may have profound beneficial effects on atherogenesis in man.

Interleukin-1beta and signaling of interleukin-1 in vascular wall and circulating cells modulates the extent of neointima formation in mice.

Interleukin (IL)-1 is an important mediator of inflammation and cardiovascular disease. Here, we examined the role of IL-1 in arterial neointima formation. Carotid artery neointima was induced by ligation, and arteries were harvested 4 weeks after injury. The neointima/media of mice deficient in the IL-1 signaling receptor (IL-1R1(-/-)) was significantly reduced compared to IL-1R1(+/+) controls (P < 0.01). IL-1R1(+/+) mice receiving subcutaneous IL-1ra also had significantly reduced neointima/media compared with placebo (P < 0.05). IL-1beta(-/-) mice had reduced neointima/media compared to wild-type (P < 0.05), whereas IL-1alpha(-/-) mice were no different from controls. Mice deficient in the P2X(7) receptor (involved in IL-1 release) or caspase-1 (involved in IL-1 activation) did not differ in their response to carotid ligation compared to controls. To examine the site of IL-1 signaling, we generated chimeric mice. IL-1R1(+/+) mice receiving IL-1R1(-/-) marrow and IL-1R1(-/-) mice receiving IL-1R1(+/+) marrow both had significantly reduced neointima/media compared with IL-1R1(+/+) to IL-1R1(+/+) (P < 0.05) but had significantly greater neointima/media than IL-1R1(-/-) to IL-1R1(-/-) controls (P < 0.05). These data confirm the importance of IL-1beta signaling in mediating arterial neointima formation and suggest the involvement of IL-1 signaling in both circulating and arterial wall cells. Furthermore, receptor antagonism may be a better therapeutic target than interruption of IL-1beta processing or release.