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Since obstructive sleep apnea (OSA) is known as a common cause of secondary hypertension, it is necessary to consider OSA a differential diagnosis in treating patients with hypertension. Apnea during sleep causes temporary and repeated hypoxia and subsequent sympathetic nerve activation. Furthermore, dysfunction of endothelial cells due to hypoxia is considered another mechanism leading to enhanced stiffness of the elastic artery and eventually, elevation of blood pressure (BP). As a result, OSA induces a nighttime or morning surge in BP, and long-standing severe OSA also causes daytime hypertension. Therefore, treatment of OSA may be important for BP control, especially in severe OSA cases. For the treatment of OSA, continuous positive airway pressure (CPAP) is a major treatment option, though alternative devices may be useful in CPAP-intolerant cases.

Since obstructive sleep apnea (OSA) is known as a common cause of secondary hypertension, it is necessary to consider OSA a differential diagnosis in treating patients with hypertension. Apnea during sleep causes temporary and repeated hypoxia and subsequent sympathetic nerve activation. Furthermore, dysfunction of endothelial cells due to hypoxia is considered another mechanism leading to enhanced stiffness of the elastic artery and eventually, elevation of blood pressure (BP). As a result, OSA induces a nighttime or morning surge in BP, and long-standing severe OSA also causes daytime hypertension. Therefore, treatment of OSA may be important for BP control, especially in severe OSA cases. For the treatment of OSA, continuous positive airway pressure (CPAP) is a major treatment option, though alternative devices may be useful in CPAP-intolerant cases.

Gold nano-rods, rod-shaped gold nanoparticles, act as contrast agents for in vivo bioimaging, drug delivery vehicles and thermal converters for photothermal therapy. Pro-inflammatory cytokines play critical roles in the development of heart failure. We examined the delivery of GNRs into the failing heart of a transgenic (TG) mouse model of inflammatory cardiomyopathy with the cardiac-specific overexpression of TNF-α. We modified GNRs with polyethylene glycol (PEG) to avoid cytotoxicity and reduce the rapid clearance of nanoparticles from blood. PEG-modified GNRs (4.5 mM as gold atoms, 200 μL) were administered intravenously to TG (n = 7) and wild-type (WT) mice (n = 5). These were killed 24 h later, and the heart, lung, liver, kidney and spleen were excised. A quantitative analysis of gold was performed using inductively coupled plasma mass or optical emission spectrometry. The amount of gold (ng) in the TG heart (3.24 ± 1.56 ng/mg heart weight) was significantly greater than that in the WT heart (1.01 ± 0.19; p

We herein report a case of concurrent cardiac sarcoidosis and large-vessel aortitis detected by 18F-fluorodeoxyglucose positron emission tomography (FDG-PET) and followed up during immunosuppressive therapy. After high-dose prednisolone administration (1 mg/kg), serial FDG-PET showed that almost all of the abnormal FDG uptake in the heart and extracardiac region, including the abdominal to bilateral iliac arteries, had been disappeared. During the tapering of prednisolone, additive methotrexate therapy was needed to treat the recurrence of cardiac sarcoidosis. FDG-PET is a useful tool for detecting cardiac sarcoidosis concomitant with large-vessel aortitis and monitoring the effectiveness of immunosuppressive therapy.

Sleep-disordered breathing (SDB) is often accompanied with the chronic heart failure (CHF). Hypoxemia due to pulmonary congestion from CHF and concurrent SDB might synergistically impair endothelial function and worsen the prognosis. However, the main factors affecting deterioration of endothelial function are unknown and whether the influence of hypoxemia differs in SDB patients with and without CHF remains unclear. Fifty-three patients (CHF group, n = 23; non-CHF group, n = 30) underwent polysomnography to evaluate their SDB and flow-mediated vasodilation (FMD) measurements to assess arterial endothelial function. We examined the relationships between FMD and SDB-related parameters, including our original index of accumulated hypoxemia by SDB throughout one-night sleep: the time desaturation summation index (TDS), calculated as follows: (100% − averaged arterial oxygen saturation during sleep) × total sleep time. The mean age in the CHF and non-CHF groups was 59.0 ± 13.5 and 57.7 ± 11.4 years, respectively. Although the FMD in the 2 groups were not significantly different, well-known adverse factors for FMD such as serum lipid profiles, blood pressure levels, and conventional indices of SDB were worse in the non-CHF group. Only the TDS was not significantly different between 2 groups and associated with FMD as shown by the univariate analysis (CHF: p

Aim: Dipeptidyl peptidase-4 (DPP-4) inhibitors lower blood glucose levels through inhibition of incretin degradation, which stimulates insulin secretion. Recent studies reported that DPP-4 inhibitors suppressed atherogenesis in apolipoprotein E-knockout (ApoEKO) mice. In this study, we investigated whether teneligliptin, a DPP-4 inhibitor, affects the development of abdominal aortic aneurysms (AAA) in ApoEKO mice. Methods: ApoEKO mice were fed a high-fat diet (HFD) and infused with angiotensin (Ang) II by osmotic mini pumps for 4 weeks to induce AAA with (DPP-4i group) or without (control group) teneligliptin administered orally from 1 week before HFD and Ang II infusion to the end of the experiment. Confluent rat vascular smooth muscle cells (VSMCs) were serum-starved for 48 hours, then incubated with or without teneligliptin for another 24 hours and stimulated with Ang II. Results: Treatment with teneligliptin significantly reduced the AAA formation rate (30.7% vs. 71.4% vs. control, P < 0.05), aortic dilatation (1.32 ± 0.09 mm vs. 1.76 ± 0.18 mm in the control, P < 0.05) and severity score (0.75 ± 0.28 vs. 1.91 ± 0.4 in the control, P < 0.05). Elastin degradation grade was also attenuated in DPP-4i group (2.83 ± 0.17 vs. 3.45 ± 0.16 in the control, P < 0.05). The number of macrophages infiltrating into the abdominal aorta was decreased in the DPP-4i group (51.8 ± 29.8/section vs. 219.5 ± 78.5/section in the control, P < 0.05). Teneligliptin attenuated Ang II-induced phosphorylation of extracellular signal-regulated kinase (ERK) and Akt, and mRNA expression of monocyte chemoattractant protein-1 in VSMCs. Conclusion: Treatment with teneligliptin suppressed AAA formation in ApoEKO mice with HFD and Ang II infusion. Suppression of macrophage infiltration by teneligliptin may be involved in the inhibition of AAA formation.

In the present study we sought to determine the effect of CoCl2, an inhibitor of PHD (prolyl hydroxylase domain protein), on the development of AAA (abdominal aortic aneurysm). AAA was induced in C57BL/6 mice by periaortic application of CaCl2 (AAA group). NaCl (0.9%)-treated mice were used as a sham control (SHAM group). Mice were treated with 0.05% CoCl2 in the drinking water (AAA/CoCl2 group). At 1 and 6 weeks after the operation, aortic tissue was excised for further examination. After 6 weeks of CaCl2 treatment, aortic diameter and macrophage infiltration into the aortic adventitia were increased in the AAA group compared with the SHAM group. Treatment with CoCl2 reduced the aneurysmal size and macrophage infiltration compared with the AAA group. Aortic expression of inflammatory cytokines and MCP-1 (monocyte chemoattractant protein-1) and the activities of MMP-9 (matrix metalloproteinase-9) and MMP-2 were enhanced in the AAA group and attenuated in the AAA/CoCl2 group. Expression of cytokines and the activities of MMPs were already increased after 1 week of CaCl2 treatment, but were suppressed by CoCl2 treatment in association with reduced NF-κB (nuclear factor κB) phosphorylation. Treatment with CoCl2 in mice prevented the development of CaCl2-induced AAA in association with reduced inflammation and ECM (extracellular matrix) disruption. The results of the present study suggest that PHD plays a critical role in the development of AAA and that there is a therapeutic potential for PHD inhibitors in the prevention of AAA development. Abbreviations: AAA, abdominal aortic aneurysm; CAT, catalase; DAB, diaminobenzidine; DMOG, dimethyloxaloylglycine; ECM, extracellular matrix; EPO, erythropoietin; EVG, elastica Van Gieson; H/E, haematoxylin and eosin; HIF, hypoxia-inducible factor; HPRT, hypoxanthine phosphoribosyl transferase; IL, interleukin; LPS, lipopolysaccharide; MCP-1, monocyte chemoattractant protein-1; MMP, matrix metalloproteinase; NF-κB, nuclear factor κB; PHD, prolyl hydroxylase domain protein; qRT-PCR, quantitative real-time reverse transcription–PCR; ROS, reactive oxygen species; SOD1, superoxide dismutase 1; TNFα, tumour necrosis factor α; VEGF, vascular endothelial growth factor; VSMC, vascular smooth muscle cell

Hypoxemia is seen in patients with pulmonary hypertension and hypoxic pulmonary vasoconstriction worsens their clinical condition. However, vasoconstriction is not the only aspect through which hypoxia induces the progression to pulmonary hypertension. Hypoxia‐inducible factor‐1α (HIF‐1α) is a transcription factor responding to hypoxic conditions by regulating hundreds of genes involved in angiogenesis, erythropoiesis, inflammation, and proliferation. We sought to determine the contribution of HIF‐1α in myeloid lineage cells to the pulmonary vascular response to chronic exposure to hypoxia. We generated myeloid‐specific HIF‐1α knockout (MyeHIF1KO) mice by using Cre‐lox P system, and exposed them to hypoxic conditions for 3 weeks to induce pulmonary hypertension. Macrophages from MyeHIF1KO and control mice were used for western blotting, RT‐qPCR, chemotaxis assay, and ATP assay. MyeHIF1KO mice exposed to hypoxia for 3 weeks exhibited a significant reduction in the right ventricular systolic pressure accompanied by a decrease in the ratio of the right ventricular weight to left ventricular weight, muscularization of the small pulmonary arteries, and infiltration of macrophages into the lung and right ventricle compared with control mice. HIF‐1α‐deficient peritoneal macrophages showed less migration toward monocyte chemoattractant protein‐1 and a decrease in intracellular ATP levels. These results indicate that HIF‐1α in macrophages contributes to the progression of pulmonary vascular remodeling and pulmonary hypertension induced by chronic exposure to hypoxic conditions. The inhibition of myeloid‐specific HIF‐1α may be a novel therapeutic strategy for the treatment of pulmonary hypertension.

Purpose: The aim of this study was to determine the influence of lesion preparation using the dual wire scoring balloon on stent expansion and long-term outcomes. Methods: Forty-six consecutive de novo lesions treated with a single >2.5 mm drug-eluting stent under intravascular ultrasound guidance, using two implantation strategies, were randomly assigned to: 1) pre-dilation with a non-compliant balloon (NC group; n = 23) or 2) pre-dilation with a dual wire scoring balloon (DS group; n = 23). Results: Although the balloon size and the maximal dilation pressure for pre-dilatation was larger (3.33 ± 0.28 vs 3.09 ± 0.33 mm, p = 0.01) and higher (11.6 ± 3.2 vs 8.6 ± 2.7 atm, p < 0.01) in the NC group than the DS group, there were no significant differences in the stent expansion. Quantitative coronary angiography at the follow-up demonstrated a smaller in-stent late loss (0.71 ± 0.63 mm vs 0.23 ± 0.52 mm, p = 0.03) in the DS group. During the long-term follow-up, there were no significant differences in the major adverse cardiac event rates. Conclusions: Lesion preparation with a dual wire scoring balloon prior to drug-eluting stent implantation might therefore be a more feasible strategy than that using a non-compliant balloon.

Hypoxia-inducible factor (HIF)-1α is a transcription factor that regulates various genes responding to hypoxic conditions. We previously reported that myeloid-specific activation of HIF-1α had protective effects on hypertensive cardiovascular remodelling in mice. However the role of myeloid lineage HIF-1α in the development of abdominal aortic aneurysm (AAA) has not been determined. Myeloid-specific HIF-1α knockout (HIF-1KO) mice were created using a Cre-lox recombination system in the background of apolipoprotein E-deficient (ApoE−/−) mice. HIF-1KO and control mice were fed high-fat diet (HFD) and infused with angiotensin II (Ang II, 1800 ng/kg/min) by an osmotic mini pump for 4 weeks to induce AAA formation. Deletion of HIF-1α increased aortic external diameter (2.47±0.21 mm versus 1.80±0.28 mm in control, P=0.035). AAA formation rate (94.4% in HIF-1KO versus 81.8% in control) was not statistically significant. Elastic lamina degradation grade determined by Elastica van Gieson (EVG) staining was deteriorated in HIF-1KO mice (3.91±0.08 versus 3.25±0.31 in control, P=0.013). The number of infiltrated macrophages into the abdominal aorta was increased in HIF-1KO mice. Expression of tissue inhibitors of metalloproteinases (TIMPs) was suppressed in the aorta and peritoneal macrophages (PMs) from HIF-1KO mice compared with control mice. HIF-1α in myeloid lineage cells may have a protective role against AAA formation induced by Ang II and HFD in ApoE−/− mice.

SummaryThis report presents the case of a 54-year-old female with Takotsubo cardiomyopathy that recurred 12 years after the first episode. The coronary angiography at the first admission revealed ergonovine-induced coronary vasoconstriction in the left coronary artery, and recurrence recurred after the interruption of vasodilator therapy to prevent vasospasm. In addition, the coronary angiography both in the first and second event demonstrated coronary slow flow phenomenon, which improved after the intracoronary administration of nicorandil. These findings indicate that coronary microvascular constriction plays an important role in the pathophysiology of Takotsubo cardiomyopathy.

Inhibition of prolyl hydroxylase domain-containing protein (PHD) by hypoxia stabilizes hypoxia-inducible factor 1 and increases the expression of target genes, such as vascular endothelial growth factor. Although the systemic renin-angiotensin system is activated by hypoxia, the role of PHD in the regulation of the renin-angiotensin system remains unknown. We examined the effect of PHD inhibition on the expression of angiotensin II type 1 receptor (AT 1 R). Hypoxia, cobalt chloride, and dimethyloxalylglycine, all known to inhibit PHD, reduced AT 1 R expression in vascular smooth muscle cells. Knockdown of PHD2, a major isoform of PHDs, by RNA interference also reduced AT 1 R expression. Cobalt chloride diminished angiotensin II–induced extracellular signal–regulated kinase phosphorylation. Cobalt chloride decreased AT 1 R mRNA through transcriptional and posttranscriptional mechanisms. Oral administration of cobalt chloride (14 mg/kg per day) to C57BL/6J mice receiving angiotensin II infusion (490 ng/kg per minute) for 4 weeks significantly attenuated perivascular fibrosis of the coronary arteries without affecting blood pressure level. These data suggest that PHD inhibition may be beneficial for the treatment of cardiovascular diseases by inhibiting renin-angiotensin system via AT 1 R downregulation.

Background—Local delivery of chemotactic factors represents a novel approach to tissue regeneration. However, successful chemokine protein delivery is challenged by barriers including the rapid diffusion of chemokines and cleavage of chemokines by proteases that are activated in injured tissues. Stromal cell–derived factor-1 (SDF-1) is a well-characterized chemokine for attracting stem cells and thus a strong candidate for promoting regeneration. However, SDF-1 is cleaved by exopeptidases and matrix metalloproteinase-2, generating a neurotoxin implicated in some forms of dementia.Methods and Results—We designed a new chemokine called S-SDF-1(S4V) that is resistant to matrix metalloproteinase-2 and exopeptidase cleavage but retains chemotactic bioactivity, reducing the neurotoxic potential of native SDF-1. To deliver S-SDF-1(S4V), we expressed and purified fusion proteins to tether the chemokine to self-assembling peptides, which form nanofibers and allow local delivery. Intramyocardial delivery of S-SDF-1(S4V) after myocardial infarction recruited CXCR4+/c-Kit+stem cells (46±7 to 119±18 cells per section) and increased capillary density (from 169±42 to 283±27 per 1 mm2). Furthermore, in a randomized, blinded study of 176 rats with myocardial infarction, nanofiber delivery of the protease-resistant S-SDF-1(S4V) improved cardiac function (ejection fraction increased from 34.0±2.5% to 50.7±3.1%), whereas native SDF-1 had no beneficial effects.Conclusions—The combined advances of a new, protease-resistant SDF-1 and nanofiber-mediated delivery promoted recruitment of stem cells and improved cardiac function after myocardial infarction. These data demonstrate that driving chemotaxis of stem cells by local chemokine delivery is a promising new strategy for tissue regeneration.

Background: Macrophage migration to the vessels is important for vascular inflammation and induces vascular degenerative diseases. Macrophages secrete matrix metalloprotainases (MMPs), which activate many cytokines and digest extracellular matrix of aorta. MMPs play an important role in the progression of aortic aneurysm. We previously reported that non-specific prolyl hydroxylase domain protein (PHD) inhibitor, Cobalt chloride (CoCl2), suppressed MMP-2 and -9 expression and attenuated experimental abdominal aortic aneurysm (AAA) formation in mice. In this report we investigated the myeloid specific PHD2 knockout effect on MMPs expression and aneurysm formation. Methods: Myeloid specific PHD2 conditional knockout (MyPHD2KO) mice were generated. Experimental AAA was induced by periaortic application of Calcium chloride (CaCl2) for 6 weeks. In vitro Lipopolysaccharide (LPS, 100ng/ml) was used to induce MMP-2 and MMP-9 expression with peritoneal macrophages. MMPs mRNA, protein expression and protein activity were analyzed by qRT-PCR, Western blot analysis and Zymography, respectively. ELISA-based NF-κB p65 Transcription Factor Assay was used to examine NF-κB p65 binding activity with consensus DNA binding site. Results: CaCl2-induced AAA was suppressed with MyPHD2KO mice (max diameter of aneurysm: 1.03mm±0.14mm in MyPHD2KO group, 1.63mm±0.34mm in Control AAA group, p Conclusion: Deletion of PHD2 in myeloid lineage attenuated MMPs expression by NF-κB inactivation and suppressed AAA formation. PHD2 in macrophage may be a novel target for cardiovascular disease treatment.

Background: Hypoxia-inducible factor-1α (Hif1α) is a transcriptional factor that regulates various genes reacting hypoxic conditions. In atherosclerotic legion, Hif1α is thought to be regulating inflammatory responses. We previously reported that myeloid specific Proryl hydroxylase domain protein2 deletion had protective effect on cardiovascular disease in animal model. The roll of Hif1α of myeloid lineage in inflammatory response is not determined. Methods and Results: Myeloid specific Hif1α knock out mice (MyHif KO) were created using Cre-lox recombination. MyHif KO mice were crossed with Apolipoprotein E knockout (ApoEKO) mice to create double knock out mice (MyHif/ApoE DKO group, n=18). ApoEKO with Hif1α flox/flox (without Cre recombinase) mice were used as control group (n=11). Two groups were fed with high fat diet (HFD) and infused with Angiotensin II (AII, 1800ng/kg/min) by osmotic mini pump for 4 weeks to promote abdominal aortic aneurysm (AAA) formation. The genotype of mice was blinded during experiment. There was no significant difference in survival rate between two groups. MyHif/ApoE DKO increased AAA formation rate (94.4% vs 81.8% in control) and aortic diameter (2.42mm±0.21mm vs 1.89±0.27mm in control). AAA classification reported by Daugherty et al was significantly exaggerated in MyHif/ApoE DKO group (2.75±0.35 vs 1.63±0.46 in control, P=0.03). Elastin degradation grade was checked by Elastica van Gieson staining. MyHif/ApoE DKO also deteriorated the elastin degradation grade (3.91±0.08 vs 3.25±0.31, P=0.01). The number of macrophages which migrated to abdominal aorta was increased in MyHif/ApoE DKO group (565±110.3/section vs 260.5±136.5/section in control, P=0.0507), whereas MyHif/ApoE DKO did not change the glucose level, blood pressure and body weight. Peritoneal macrophages from MyHif/ApoE DKO were significantly activated the migration induced by monocyte chemotactic protein-1 in dose dependent manner. Conclusion: Hif1α of myeloid lineage might have vascular protective effect via suppressing the macrophage activation in the model of AII-induced AAA in mice.

Background: Previously we reported that activation of vagal nerve by acetylcholinesterase inhibitors attenuated atherogenesis via suppression of inflammatory responses. α7 nicotinic acetylcholine receptor (nAchR) plays an important role in vagal nerve activation. We sought to determine whether AR-R17779, a selective agonist of α7 nAchR, affects the development of abdominal aortic aneurysm (AAA). Methods and Results: AAA was induced by topical application of calcium chloride (CaCl2) to abdominal aorta (AAA group). NaCl (0.9%) was substituted for CaCl2 as a sham operation (SHAM group). AR-R17779 was administered in drinking water (AAA/AR-R group). One and 6 weeks after the operation, aortic tissue was excised for histological and molecular analyses. Aortic diameter and macrophage infiltration into the aortic adventitia were increased in AAA group compared with SHAM group at 6 weeks. Treatment with AR-R17779 reduced the diameter of the aorta (0.73±0.10 mm vs. 1.07±0.15 mm in AAA, p Conclusion: Treatment with AR-R17779 prevented the enlargement of abdominal aorta induced by CaCl2 in association with reduced inflammation and extracellular matrix disruption via NF-kB inactivation. These findings suggest a therapeutic potential of α7nAchR activation for prevention of AAA development.

BACKGROUND Sleep-disordered breathing (SDB) is associated with repeated intermittent hypoxemia, and it is known as one of the risk factors for cardiovascular diseases. Previous studies assessing the effects of frequency and depth of hypoxemia on cardiovascular diseases have shown conflicting results. The aim of the current study was to clarify what SDB-related parameters most predict endothelial dysfunction to better understand the pathogenesis of endothelial dysfunction in patients with SDB. METHODS We conducted polysomnography (PSG) and measured flow-mediated vasodilation response (%FMD) in 50 outpatients suspected of SDB. Evaluated indices included: apnea-hypopnea index (AHI), 3% oxygen desaturation index (3%ODI), averaged arterial oxygen saturation (averaged SpO2), lowest arterial oxygen saturation (lowest SpO2), ratio of arterial oxygen saturation

Thyroid hormone has a broad effect on cardiovascular system. 3,3′,5-triiodo- l -thyronine (T3), a biologically active form of thyroid hormone, increases cardiac contractility. T3 causes arterial relaxation and reduction of systemic vascular resistance, resulting in an increase in cardiac output. However, the molecular mechanisms of vascular relaxation by T3 are incompletely characterized. We studied the effect of T3 on the angiotensin (Ang) II type 1 receptor (AT 1 R) expression in vascular smooth muscle cells. T3 dose-dependently decreased expression levels of AT 1 R mRNA, with a peak at 6 hours of stimulation. Binding assay using [ 125 I]Sar 1 -Ile 8 -Ang II revealed that AT 1 R number was decreased by stimulation with T3 without changing the affinity to Ang II. T3 reduced calcium response of vascular smooth muscle cells to Ang II by 26%. AT 1 R promoter activity measured by luciferase assay was reduced by 50% after 9 hours of T3 administration. mRNA stability was also decreased by T3. Real-time quantitative reverse transcription–polymerase chain reaction and Western blot analysis revealed that AT 1 R mRNA and protein were downregulated in the aorta of T3-treated rats. These results suggest that T3 downregulates AT 1 R expression both at transcriptional and posttranscriptional levels, and attenuates biological function of Ang II. Our results suggest that downregulation of AT 1 R gene expression may play an important role for T3-induced vascular relaxation.

Atherosclerosis is a chronic inflammatory disease. It has been appreciated that vagus nerve inhibits macrophage activation via α7 nicotinic acetylcholine receptor (nAChR), termed the cholinergic anti-inflammatory pathway. We explored the effects of AR-R17779, a selective α7nAChR agonist, on atherosclerosis and aneurysm formation in apolipoprotein E (ApoE)-deficient mice. ApoE-deficient mice were fed a high-fat diet (HFD) and angiotensin II (Ang II) was infused by osmotic minipumps from 10-week-old for 4weeks. AR-R17779 was given in drinking water ad libitum. Oil red O staining of the aorta showed that combined loading of HFD and Ang II induced marked atherosclerosis compared with control mice fed a normal chow. Treatment with AR-R17779 significantly reduced atherosclerotic plaque area and improved survival of mice. Treatment with AR-R17779 also suppressed abdominal aortic aneurysm formation. Quantitative RT-PCR of the aorta revealed that mRNA expression levels of interleukin-1β, interleukin-6 and NOX2 were significantly decreased in AR-R17779-treated mice compared with Ang II+HFD mice. AR-R17779 treatment also reduced blood pressure and serum lipid levels. In conclusion, α7nAChR activation attenuates atherogenesis and aortic abdominal aneurysm formation in ApoE-deficient mice possibly through an anti-inflammatory effect and reduction of blood pressure and lipid levels. Pharmacological activation of α7nAChR may have a therapeutic potential against atherosclerotic vascular diseases through multiple mechanisms.

Objective Activation of vagal nerve suppresses inflammatory responses through activation of α7 nicotinic acetylcholine receptor (nAchR). We sought to determine whether AR-R17779, a selective agonist of α7nAchR, affects the development of abdominal aortic aneurysm (AAA). Methods and results AAA was induced by topical application of calcium chloride (CaCl 2 ) to abdominal aorta (AAA group). NaCl (0.9%) was substituted for CaCl 2 as a sham operation (SHAM group). AR-R17779 was administered in drinking water (AAA/AR-R group). One and 6 weeks after the operation, aortic tissue was excised for histological and molecular analyses. Aortic diameter and macrophage infiltration into the aortic adventitia were increased in AAA group compared with SHAM group at 6 weeks. Treatment with AR-R17779 reduced the diameter of the aorta and macrophage infiltration compared with AAA group. Wavy morphology of the elastic lamellae was lost in AAA group while it was preserved in AAA/AR-R group. Expression of inflammatory cytokines and matrix metalloproteinase (MMP) activities were enhanced in AAA group, which was suppressed in AAA/AR-R group. AR-R17779 treatment suppressed CaCl 2 -induced expression of cytokines, activities of MMPs and NF-κB activation at 1 week when aortic dilatation had not developed. Conclusion Treatment with AR-R17779 prevented the enlargement of abdominal aorta induced by CaCl 2 in association with reduced inflammation and extracellular matrix disruption. These findings suggest therapeutic potential of α7nAchR activation for prevention of AAA development.

Peroxisome proliferator-activated receptor (PPAR) gamma belongs to the nuclear receptor superfamily of ligand-dependent transcription factors. Recent results have shown that the ligands for nuclear receptors have rapid effects so called "nongenomic" effects, which are observed within minutes after stimulation. We examined whether 15-deoxy-Delta(12,14)-prostaglandin J(2) (15-d-PGJ2) had rapid effects on cultured vascular smooth muscle cells. Phosphorylation of ERK and c-fos mRNA expression were determined by Western and Northern blot analyses, respectively. PPAR gamma agonists 15-d-PGJ2 and thiazolidinediones such as pioglitazone and troglitazone elicited rapid activation of ERK within 15 min and induced c-fos mRNA expression within 30 min, whereas the PPAR alpha agonist bezafibrate failed to activate ERK. 15-d-PGJ2-induced expression of c-fos mRNA was blocked by PD98059 or U0126, two ERK kinase inhibitors, suggesting that the MEK/ERK pathway mediates 15-d-PGJ2-induced c-fos gene expression. Furthermore, pretreatment with wortmannin, an inhibitor of phosphatidylinositol 3 (PI3)-kinase, inhibited 15-d-PGJ2-induced ERK activation and c-fos mRNA expression, suggesting that PI3-kinase is involved in the process. An electrophoretic mobility shift assay showed that 15-d-PGJ2 enhanced AP-1 binding activity to AP-1 consensus sequence in a time-dependent manner. 15-d-PGJ2 increased thymidine incorporation in a PI3-kinase-dependent manner. Taken together, our findings show that 15-d-PGJ2 and thiazolidinediones activate the MEK/ERK pathway through PI3-kinase and lead to c-fos mRNA expression and DNA synthesis. These findings indicate a novel regulatory mechanism of gene expression by 15-d-PGJ2 and thiazolidinediones.

The plasma level of interleukin-6 (IL-6) is elevated in patients with acute coronary syndromes and has prognostic value. Thrombin is a potent mitogen for vascular smooth muscle cells (VSMCs) and plays an important role in the progression of atherosclerosis. We examined the mechanism of thrombin-induced IL-6 expression in VSMCs. Thrombin induced IL-6 mRNA and protein expression in a dose-dependent manner. Pharmacological inhibition of extracellular signal-regulated protein kinase (ERK), p38 mitogen-activated protein kinase (MAPK), or epidermal growth factor receptor (EGF-R) suppressed the thrombin-induced IL-6 expression. Deletion and mutation analysis of the promoter region of the IL-6 gene by using luciferase as a reporter showed that the DNA segment between −228 and −150 bp containing the cAMP response element (CRE) site played a critical role. Thrombin also induced phosphorylation of CRE binding protein (CREB) in an ERK- and a p38 MAPK-dependent manner. Overexpression of the dominant-negative form of CREB inhibited thrombin-induced IL-6 mRNA expression. These results suggest that the CRE site and CREB play an important role in thrombin-induced IL-6 gene expression in VSMCs. Transactivation of EGF-R and activation of ERK and p38 MAPK are involved in this process. CREB may be a novel transcription factor that regulates thrombin-induced gene expression.

Abstract —Plasminogen activator inhibitor type-1 (PAI-1) plays an integral role not only in the regulation of fibrinolytic activity but also in the pathogenesis of atherosclerosis and hypertension. We investigated the signaling pathways of angiotensin II (Ang II) leading to PAI-1 gene expression. Ang II increased the PAI-1 mRNA and protein levels in a time- and dose-dependent manner through the Ang II type 1 receptor in vascular smooth muscle cells. PAI-1 gene promoter activity measured by luciferase assay was significantly increased by Ang II. PAI-1 mRNA stability was also increased by Ang II. Ang II–induced PAI-1 mRNA upregulation was inhibited by BAPTA-AM, genistein, and AG1478, suggesting that intracellular calcium, tyrosine kinase, and epidermal growth factor receptor transactivation are involved. Furthermore, PD98059, an inhibitor of extracellular signal–regulated kinase (ERK) kinase (MEK), almost completely suppressed Ang II–induced PAI-1 upregulation. Adenovirus-mediated overexpression of the dominant-negative form of Rho-kinase or Y27632, a Rho-kinase inhibitor, also completely prevented PAI-1 induction by Ang II without affecting Ang II–induced ERK activation. These data suggest that activation of MEK/ERK and Rho-kinase pathways plays a pivotal role in PAI-1 gene upregulation by Ang II. The Rho-kinase pathway may be a novel target to inhibit Ang II signaling, and its inhibition may be useful in the treatment of hypertension as well as atherosclerosis.

Background Hypertension induces cardiovascular hypertrophy and fibrosis. Infiltrated macrophages are critically involved in this process. We recently reported that inhibition of prolyl hydroxylase domain protein 2 ( PHD 2), which hydroxylates the proline residues of hypoxia‐inducible factor‐α ( HIF ‐α) and thereby induces HIF ‐α degradation, suppressed inflammatory responses in macrophages. We examined whether myeloid‐specific Phd2 deletion affects hypertension‐induced cardiovascular remodeling. Methods and Results Myeloid‐specific PHD 2‐deficient mice (MyPHD2KO) were generated by crossing Phd2 ‐floxed mice with LysM‐Cre transgenic mice, resulting in the accumulation of HIF ‐1α and HIF ‐2α in macrophage. Eight‐ to ten‐week‐old mice were given N G ‐nitro‐L‐arginine methyl ester (L‐ NAME ), a nitric oxide synthase inhibitor, and Angiotensin II (Ang II) infusion. L‐ NAME /Ang II comparably increased systolic blood pressure in control and MyPHD2KO mice. However, MyPHD2KO mice showed less aortic medial and adventitial thickening, and macrophage infiltration. Cardiac interstitial fibrosis and myocyte hypertrophy were also significantly ameliorated in MyPHD2KO mice. Transforming growth factor ‐β and collagen expression were decreased in the aorta and heart from MyPHD2KO mice. Echocardiographic analysis showed that left ventricular hypertrophy and reduced ejection fraction induced by L‐ NAME /Ang II treatment in control mice were not observed in MyPHD2KO mice. Administration of digoxin that inhibits HIF ‐α synthesis to L‐ NAME /Ang II‐treated MyPHD2KO mice reversed these beneficial features. Conclusions Phd2 deletion in myeloid lineage attenuates hypertensive cardiovascular hypertrophy and fibrosis, which may be mediated by decreased inflammation‐ and fibrosis‐associated gene expression in macrophages. PHD 2 in myeloid lineage plays a critical role in hypertensive cardiovascular remodeling.

Resveratrol (3,5,4'-trihydroxystilbene), a polyphenol abundant in red wine, is known to extend the life span of diverse species. On the contrary, it was reported that angiotensin (Ang) II enhances senescence of vascular smooth muscle cells (VSMCs). We, therefore, examined whether resveratrol attenuates Ang II-induced senescence of VSMC. Senescence-associated β-galactosidase (SA β-gal) assay showed that Ang II induced senescence of VSMC. The Ang II-induced senescence was inhibited by losartan, an Ang II type 1 receptor (AT1R) antagonist but not by PD123319, Ang II type 2 receptor antagonist, indicating that AT1R is responsible for the induction of senescence. Resveratrol suppressed Ang II-induced senescence of VSMC in a dose-dependent manner. In addition, resveratrol suppressed Ang II-induced induction of p53 and its downstream target gene p21, both of which play an important role in the induction of senescence. Resveratrol suppressed senescence of VSMC possibly through inhibition of AT1R-dependent induction of p53/p21. Suppression of p53 induction may be involved in the longevity by resveratrol.

Microvascular dysfunction has been proposed as the most likely mechanism of the coronary slow flow phenomenon (CSFP).To determine the effects of isosorbide dinitrate and nicorandil on the CSFP.Changes in thrombolysis in myocardial infarction (TIMI) frame count following the intracoronary administration of isosorbide dinitrate and nicorandil were assessed in 11 patients with the CSFP.After the administration of isosorbide dinitrate, the median TIMI frame count decreased to 32 (range 20-60) [p = 0.003], which was lower than that of the control [43 (29-73)]. The count decreased further to 25 (12-34) [p = 0.041] after the administration of nicorandil. The count after the subsequent administration of contrast medium was increased to 32 (20-49) [p = 0.03].These angiographic findings indicate that the intracoronary administration of nicorandil is superior to isosorbide dinitrate with regard to improving the CSFP. These findings suggest that microvascular spasm is the main factor in the pathogenesis of the CSFP.

Background— Cardiac progenitor cells (CPCs) possess the insulin-like growth factor-1 (IGF-1)-IGF-1 receptor system, and IGF-1 can be tethered to self-assembling peptide nanofibers (NF-IGF-1), leading to prolonged release of this growth factor to the myocardium. Therefore, we tested whether local injection of clonogenic CPCs and NF-IGF-1 potentiates the activation and differentiation of delivered and resident CPCs enhancing cardiac repair after infarction. Methods and Results— Myocardial infarction was induced in rats, and untreated infarcts and infarcts treated with CPCs or NF-IGF-1 only and CPCs and NF-IGF-1 together were analyzed. With respect to infarcts exposed to CPCs or NF-IGF-1 alone, combination therapy resulted in a greater increase in the ratio of left ventricular mass to chamber volume and a better preservation of +dP/dt, −dP/dt, ejection fraction, and diastolic wall stress. Myocardial regeneration was detected in all treated infarcts, but the number of newly formed myocytes with combination therapy was 32% and 230% higher than with CPCs and NF-IGF-1, respectively. Corresponding differences in the volume of regenerated myocytes were 48% and 115%. Similarly, the length density of newly formed coronary arterioles with both CPCs and NF-IGF-1 was 73% and 83% greater than with CPCs and NF-IGF-1 alone, respectively. Importantly, activation of resident CPCs by paracrine effects contributed to cardiomyogenesis and vasculogenesis. Collectively, CPCs and NF-IGF-1 therapy reduced infarct size more than CPCs and NF-IGF-1 alone. Conclusions— The addition of nanofiber-mediated IGF-1 delivery to CPC therapy improved in part the recovery of myocardial structure and function after infarction.

Insulin-like growth factor-1 (IGF-1) is a small protein that promotes cell survival and growth, often acting over long distances. Although for decades IGF-1 has been considered to have therapeutic potential, systemic side effects of IGF-1 are significant, and local delivery of IGF-1 for tissue repair has been a long-standing challenge. In this study, we designed and purified a novel protein, heparin-binding IGF-1 (Xp-HB-IGF-1), which is a fusion protein of native IGF-1 with the heparin-binding domain of heparin-binding epidermal growth factor-like growth factor. Xp-HB-IGF-1 bound selectively to heparin as well as the cell surfaces of 3T3 fibroblasts, neonatal cardiac myocytes and differentiating ES cells. Xp-HB-IGF-1 activated the IGF-1 receptor and Akt with identical kinetics and dose response, indicating no compromise of biological activity due to the heparin-binding domain. Because cartilage is a proteoglycan-rich environment and IGF-1 is a known stimulus for chondrocyte biosynthesis, we then studied the effectiveness of Xp-HB-IGF-1 in cartilage. Xp-HB-IGF-1 was selectively retained by cartilage explants and led to sustained chondrocyte proteoglycan biosynthesis compared to IGF-1. These data show that the strategy of engineering a "long-distance" growth factor like IGF-1 for local delivery may be useful for tissue repair and minimizing systemic effects.

Background Clinical trials have shown that treatment of patients with type 2 diabetes with pioglitazone, a peroxisome proliferator-activated receptor (PPAR)γ agonist, reduces cardiovascular events. However, the effect of PPARγ agonists on endoplasmic reticulum (ER) stress that plays an important role in the progression of atherosclerosis has not been determined. We sought to determine the effect of PPARγ agonists on ER stress induced by palmitate, the most abundant saturated fatty acid in the serum. Methods and Results Protein expression of ER stress marker was evaluated by Western blot analysis and stearoyl-CoA desaturase1 (SCD-1) mRNA expression was evaluated by qRT-PCR. Macrophage apoptosis was detected by flowcytometry. Pioglitazone and rosiglitazone reduced palmitate-induced phosphorylation of PERK, a marker of ER stress, in RAW264.7, a murine macrophage cell line. Pioglitazone also suppressed palmitate-induced apoptosis in association with inhibition of CHOP expression, JNK phosphorylation and cleavage of caspase-3. These effects of pioglitazone were reversed by GW9662, a PPARγ antagonist, indicating that PPARγ is involved in this process. PPARγ agonists increased expression of SCD-1 that introduces a double bond on the acyl chain of long-chain fatty acid. 4-(2-Chlorophenoxy)-N-(3-(3-methylcarbamoyl)phenyl)piperidine-1-carboxamide, an inhibitor of SCD-1, abolished the anti-ER stress and anti-apoptotic effects of pioglitazone. These results suggest that PPARγ agonists attenuate palmitate-induced ER stress and apoptosis through SCD-1 induction. Up-regulation of SCD-1 may contribute to the reduction of cardiovascular events by treatment with PPARγ agonists.

Prostaglandin F(2alpha) (PGF(2alpha)) is a vasoactive factor that causes constriction and hypertrophy of vascular smooth muscle cells (VSMCs). However, the mechanism of PGF(2alpha)-induced hypertrophy is largely unknown. Cyclic AMP-response element (CRE)-binding protein (CREB), the best characterized stimulus-induced transcription factor, activates transcription of target genes with CRE and promotes cell growth. We examined the role of CREB in PGF(2alpha)-induced hypertrophy of VSMCs. PGF(2alpha) induced phosphorylation of CREB at serine 133, which is a critical marker of activation, after 5-10min of stimulation in a dose-dependent manner. Pharmacological inhibition of extracellular signal-regulated protein kinase and p38 mitogen-activated protein kinase (p38-MAPK) suppressed PGF(2alpha)-induced CREB phosphorylation. Inhibition of epidermal growth factor receptor (EGFR) and mitogen- and stress-activated protein kinase-1 also suppressed PGF(2alpha)-induced CREB phosphorylation. Overexpression of dominant-negative form of CREB (AdCREB M1), of which serine 133 was replaced with alanine, inhibited PGF(2alpha)-induced c-fos mRNA expression as well as hypertrophy of VSMCs [hypertrophy index (microg/10(4)cell); control 8.13, PGF(2alpha) 9.85, AdCREB M1 7.91, and AdCREB M1+PGF(2alpha) 8.43]. These results suggest that PGF(2alpha) activated CRE-dependent gene transcription through EGFR transactivation, and the CREB pathway plays a critical role in PGF(2alpha)-induced hypertrophy of VSMCs.

Proliferation of vascular smooth muscle cells (VSMCs) is induced by various mitogens through activation of extracellular signal-regulated protein kinase (ERK) pathway. We recently reported that peroxisome proliferator-activated receptor (PPAR)gamma activators such as 15-deoxy-Delta12,14-prostaglandin J2 (15-d-PGJ2) and thiazolidinediones (TZDs) activated MEK/ERK pathway through phosphatidylinositol 3-kinase (PI3-K) and induced proliferation of VSMCs. However, the precise mechanisms of PPARgamma activators-induced activation of PI3-K/ERK pathway have not been determined. We examined whether transactivation of growth factor receptor is involved in this process. Stimulation of VSMCs with 15-d-PGJ2 or TZDs for 15 min induced phosphorylation of ERK1/2 and Akt. 15-d-PGJ2- or TZDs-induced phosphorylation of ERK1/2 and Akt was inhibited by AG1478, an inhibitor of epidermal growth factor receptor (EGF-R) as well as AG1295, an inhibitor of platelet derived growth factor receptor (PDGF-R). 15-d-PGJ2-induced phosphorylation of both EGF-R and PDGF-R. GM6001, a matrix metalloproteinase inhibitor, and PP2, a Src family protein kinase inhibitor, suppressed 15-d-PGJ2- and TZDs-induced phosphorylation of EGF-R and PDGFbeta-R as well as activation of ERK1/2 and Akt. PDGFbeta-R was co-immunoprecipitated with EGF-R, regardless of the presence or absence of 15-d-PGJ2. These data suggest that 15-d-PGJ2 and TZDs activate PI3-K/ERK pathway through Src family kinase- and matrix metalloproteinase-dependent transactivation of EGF-R and PDGF-R. Both receptors seemed to associate constitutively. This novel signaling mechanisms may contribute to diverse biological functions of PPARgamma activators.

Background—The balance between apoptosis and proliferation of vascular smooth muscle cells (VSMCs) is believed to contribute to the vascular remodeling process. Cyclic AMP response element–binding protein (CREB) is a critical transcription factor for the survival of neuronal cells and T lymphocytes. However, the role of CREB in blood vessels is incompletely characterized.Methods and Results—Nuclear staining with Hoechst 33258 or propidium iodine showed an increase in apoptotic cells with activation of caspase-3 in VSMCs infected with adenovirus expressing the dominant-negative form of CREB (AdCREBM1). Basal expression of Bcl-2 and Bcl-2 promoter activity were decreased by infection with AdCREBM1. Immunohistochemistry revealed that CREB was mainly induced and activated in the neointimal α-smooth muscle actin–positive cells of rat carotid artery after balloon injury. Infection with AdCREBM1 suppressed neointimal formation (intima-media ratio) by 33.8% after 14 days of injury, which was accompanied by an increase in apoptosis as indicated by terminal deoxynucleotidyl transferase–mediated dUTP nick end-labeling–positive cells and a decrease in bromodeoxyuridine incorporation.Conclusions—These results suggest that CRE-dependent gene transcription might play an important role in the survival and proliferation of VSMCs. CREB might be a novel transcription factor mediating the vascular remodeling process and a potential therapeutic target for atherosclerotic disease.

Thrombin is a potent mitogen for vascular smooth muscle cells (VSMCs) and plays an important role in the progression of atherosclerosis. Although recent reports have suggested that cAMP response element-binding protein (CREB) is necessary for the survival of neuronal cells, the role of CREB in VSMC proliferation is not determined. We examined the role of CREB in thrombin-induced VSMC proliferation and the effect of thrombin on phosphorylation of CREB at Ser133, which is a critical marker for activation by Western blot analysis. Thrombin induced phosphorylation of CREB in a dose-dependent manner. An oligopeptide, SFLLRN, which activates the thrombin receptor, also induced the phosphorylation of CREB. Inhibition of extracellular signal-regulated protein kinase or inhibition of p38 mitogen-activated protein kinase suppressed the thrombin-induced CREB phosphorylation. Inhibition of the epidermal growth factor receptor by AG1478 also inhibited the thrombin-induced CREB phosphorylation. Overexpression of the dominant-negative form of CREB inhibited thrombin-induced c- fos mRNA expression and incorporation of [ 3 H]thymidine and [ 3 H]leucine. These results suggest that CREB-dependent gene transcription plays a critical role in thrombin-induced proliferation and hypertrophy of VSMCs. Transactivation of the epidermal growth factor receptor and 2 mitogen-activated protein kinase pathways are involved in this process. CREB may be a novel transcription factor mediating the vascular remodeling process induced by thrombin.

Recent studies suggest a crucial role of reactive oxygen species (ROS) for the signaling of angiotensin (Ang) II through Ang II type 1 receptor (AT 1 -R). However, the role of ROS in the regulation of AT 1 -R expression has not been explored. In this study, we examined the effect of an antioxidant on the homologous downregulation of AT 1 -R by Ang II. Ang II (10 −6 mol/L) decreased AT 1 -R mRNA with a peak suppression at 6 hours of stimulation in rat aortic vascular smooth muscle cells. Preincubation of vascular smooth muscle cells with N -acetylcysteine (NAC), a potent antioxidant, almost completely inhibited the Ang II–induced downregulation of AT 1 -R mRNA. The effect of NAC was due to stabilization of the AT 1 -R mRNA that was destabilized by Ang II. The Ang II–induced AT 1 -R mRNA downregulation was also blocked by PD98059, an extracellular signal–regulated protein kinase (ERK) kinase inhibitor. Ang II–induced ERK activation was inhibited by NAC as well as by PD98059. Exogenous H 2 O 2 also suppressed AT 1 -R mRNA. These results suggest that the production of ROS and the activation of ERK are critical for the downregulation of AT 1 -R mRNA. The generation of ROS through stimulation of AT 1 -R not only mediates signaling of Ang II but also may play a crucial role in the adaptation process of AT 1 -R to the sustained stimulation of Ang II.

Background —Peroxisome proliferator-activated receptor γ (PPARγ) activators, such as troglitazone (Tro), not only improve insulin resistance but also suppress the neointimal formation after balloon injury. However, the precise mechanisms have not been determined. Angiotensin II (Ang II) plays crucial roles in the pathogenesis of atherosclerosis, hypertension, and neointimal formation after angioplasty. We examined the effect of PPARγ activators on the expression of Ang II type 1 receptor (AT 1 -R) in cultured vascular smooth muscle cells (VSMCs). Methods and Results —AT 1 -R mRNA and AT 1 -R protein levels were determined by Northern blot analysis and radioligand binding assay, respectively. Natural PPARγ ligand 15-deoxy-Δ 12,14 -prostaglandin J 2 , as well as Tro, reduced the AT 1 -R mRNA expression and the AT 1 -R protein level. The PPARγ activators also reduced the calcium response of VSMCs to Ang II. PPARγ activators suppressed the AT 1 -R promoter activity measured by luciferase assay but did not affect the AT 1 -R mRNA stability, suggesting that the suppression occurs at the transcriptional level. Conclusions —PPARγ activators reduced the AT 1 -R expression and calcium response to Ang II in VSMCs. Downregulation of AT 1 -R may contribute to the inhibition of neointimal formation by PPARγ activators.

P157 Plasminogen activator inhibitor-1 (PAI-1) plays an integral role not only in the regulation of plasminogen activity and fibrinolytic system but also in the pathogenesis of atherosclerosis and hypertension. Because angiotensin II (Ang II) is also involved in these processes, we investigated its role in the intracellular signaling cascade leading to PAI-1 gene expression in vascular smooth muscle cells (VSMC). Ang II increased the PAI-1 mRNA and protein levels through Ang II type 1 receptor. Although PAI-1 mRNA stability was not increased by Ang II, PAI-1 gene promoter activity, which was measured by luciferase assay, was significantly increased by Ang II. This process did not require de novo protein synthesis. BAPTA-AM, genistein and AG1478 completely inhibited the Ang II-induced PAI-1 mRNA upregulation, suggesting that intracellular calcium, tyrosine kinase and epidermal growth factor (EGF) receptor transactivation were involved in this process. However, inhibition of protein kinase C (PKC) by calphostin C, GF109203, or prolonged exposure to PMA failed to abolish the Ang II-induced PAI-1 upregulation, suggesting PKC pathway was not involved. PD98059 suppressed Ang II-induced PAI-1 upregulation, whereas SB203580 did not, suggesting that MEK/ERK1/2 pathway rather than p38 MAP kinase pathway was crucial in this process. Furthermore, adenovirus-mediated expression of dominant negative form of Rho kinase or Rho kinase inhibitor Y27632 also completely suppressed PAI-1 induction by Ang II without affecting Ang II-induced ERK1/2 activation. These data suggest that activation of both MEK/ERK1/2 and Rho kinase pathways will be necessary for the upregulation of PAI-1 gene expression and these two pathways may act synergically to promote PAI-1 gene transcription at least at the downstream of ERK1/2 in VSMC. These findings are important biological and therapeutical implications for the evolution of arterial wall thrombus and the pathogenesis of atherosclerosis by Ang II.

Abstract — 3-Hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase inhibitors, so-called statins, reduce the relative risk of a major coronary event by lowering the serum cholesterol level. In addition, statins may confer beneficial effects by cholesterol-lowering independent mechanisms, which are incompletely characterized. Because angiotensin II (Ang II) plays crucial roles in the pathogenesis of cardiovascular diseases, we examined the effect of statins on the expression of the Ang II type 1 receptor (AT 1 -R) in cultured vascular smooth muscle cells (VSMCs). Cerivastatin and fluvastatin reduced the AT 1 -R mRNA and the AT 1 -R protein levels; however, pravastatin lacked this effect. Cerivastatin and fluvastatin suppressed the AT 1 -R promoter activity measured by luciferase assay but did not affect AT 1 -R mRNA stability, suggesting that the suppression occurs at the transcriptional level. Coincubation of VSMCs with mevalonate or geranylgeranyl pyrophosphate but not with farnesyl pyrophosphate reversed the cerivastatin-induced AT 1 -R downregulation. Overexpression of dominant-negative Rho A also suppressed AT 1 -R mRNA expression. Treatment with cerivastatin for 24 hours reduced the calcium response of VSMCs to Ang II. Taken together, statins downregulate AT 1 -R expression through a mevalonate-dependent, geranylgeranyl pyrophosphate-dependent, and Rho A-dependent manner and attenuate the biological function of Ang II. Downregulation of AT 1 -R may contribute to the cholesterol-independent beneficial effect of statins on the cardiovascular system.