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Aims Excess sodium causes the development of cardiovascular diseases in conjunction with enhancing renin-angiotensin-aldosterone system (RAAS). Natriuretic peptides are sodium regulators and prevent pathological cardiac alterations by counteracting RAAS. However, it is unknown whether natriuretic peptides inhibit the sodium effect in adverse cardiac alterations. Here, we investigated whether excess salt intake could exacerbate cardiac remodeling in mice with impaired natriuretic peptide signaling. Materials and methods Mice lacking the gene encoding the natriuretic peptide receptor, guanylyl cyclase-A (GC-A), and wild-type mice were administered with either a vehicle substance or a subpressor dose of aldosterone (100 ng/kg/min), alongside low salt (0.001% NaCl), normal salt (0.6% NaCl), or high salt diets (6.0% NaCl) for four weeks. Mice were then sacrificed and the hearts were evaluated by histology and RT-PCR. Key findings Salt load did not induce cardiac changes in vehicle and aldosterone groups in wild-type mice. On the other hand, cardiac hypertrophy and interstitial fibrosis were significantly exacerbated in a salt dependent manner in GC-A knockout (KO) mice administered aldosterone, and were associated with enhanced gene expression relevant to hypertrophy, fibrosis, and oxidative stress conditions. Of note, excess salt intake increased the expression of Sgk1, serum and glucocorticoid responsive kinase-1, in aldosterone-administered GC-A KO mice. These molecular changes were not observed in wild-type mice. Significance The results of the present study demonstrate that excess salt intake induced cardiac remodeling in conjunction with aldosterone administration in GC-A KO mice, indicating that GC-A signaling attenuated the deleterious salt effect in aldosterone-induced cardiac remodeling.

Soluble fms-like tyrosine kinase-1 (sFlt-1), an endogenous inhibitor of vascular endothelial growth factor and placental growth factor, is involved in the pathogenesis of cardiovascular disease. However, the significance of sFlt-1 in heart failure has not been fully elucidated. We found that sFlt-1 is decreased in renal failure and serves as a key molecule in atherosclerosis. In this study, we aimed to investigate the role of the decreased sFlt-1 production in heart failure, using sFlt-1 knockout mice. sFlt-1 knockout mice and wild-type mice were subjected to transverse aortic constriction and evaluated after 7 days. The sFlt-1 knockout mice had significantly higher mortality (52% versus 15%; P =0.0002) attributable to heart failure and showed greater cardiac hypertrophy (heart weight to body weight ratio, 8.95±0.45 mg/g in sFlt-1 knockout mice versus 6.60±0.32 mg/g in wild-type mice; P

Coronary artery disease and cardiac morphology and function were evaluated in 51 patients with hypertrophic cardiomyopathy (HCM), without typical chest pain, using cardiac computed tomography (CT). This study investigated the prevalence of coronary artery disease, the indicators of obstructive coronary stenosis, and the magnitude of left ventricular (LV) hypertrophy. The patients’ mean coronary artery calcium score was 198.8 ± 312.0 and was positively correlated with the number of coronary risk factors (r = 0.32; P < 0.05). Of the 51 patients with HCM, 42 (82.4 %) had some degree of stenosis and 8 (15.7 %) had obstructive stenosis. Noncalcified and mixed plaques were detected in 14 (27.5 %) and 11 (21.6 %) patients, respectively. Multivariate logistic regression revealed that diabetes was an independent indicator of the presence of obstructive stenosis in HCM patients. Multivariate linear regression revealed that low estimated glomerular filtration rates and high triglyceride concentrations were independent indicators of higher LV mass indexes. In conclusion, cardiac CT revealed that coronary artery disease was common among patients with HCM. The presence of obstructive coronary stenosis and the magnitude of LV hypertrophy were related to the presence of diabetes, triglyceride levels, and estimated glomerular filtration rate.

Aims Dilated cardiomyopathy (DCM) is characterized by ventricular dilation associated with systolic dysfunction, which could be caused by mutations in lamina/C gene ( LMNA ). LMNA -linked DCM is severe in males in both human patients and a knock-in mouse model carrying a homozygous p.H222P mutation ( Lmna H222P/H222P). The aim of this study was to investigate the molecular mechanisms underlying the gender difference of LMNA -linked DCM. Methods and results A whole-exome analysis of a multiplex family with DCM exhibiting the gender difference revealed a DCM-linked LMNA mutation, p.R225X. Immunohistochemical analyses of neonatal rat cardiomyocytes expressing mutant LMNA constructs and heart samples from the LMNA -linked DCM patients and Lmna H222P/H222P mice demonstrated a nuclear accumulation of androgen receptor (AR) and its co-activators, serum response factor, and four-and-a-half LIM protein-2. Role of sex hormones in the gender difference was investigated in vivo using the Lmna H222P/H222P mice, where male and female mice were castrated and ovariectomized, respectively, or treated with testosterone or an antagonist of AR. Examination of the mice by echocardiography, followed by the analyses of histological changes and gene/protein expression profiles in the hearts, confirmed the involvement of testicular hormone in the disease progression and enhanced cardiac remodelling in the Lmna H222P/H222P mice. Conclusion These observations indicated that nuclear accumulation of AR was associated with the gender difference in LMNA -linked DCM.

Among members of the TGFβ superfamily, bone morphogenetic protein (BMP) 9 and BMP10 regulate vascular endothelium differentiation and morphogenesis by activating the specific receptor complex, which consists of ALK1 (or ACVRL1), BMPR2, and endoglin. Mutations in ACVRL1, BMPR2, or ENG are associated with hereditary hemorrhagic telangiectasia and pulmonary arterial hypertension in humans [1, 2]. We previously identified TMEM100 as a downstream target gene of BMP9/BMP10-ALK1 signaling pathway [3]. TMEM100 is a novel intracellular protein with two putative transmembrane domains, and its amino acid sequence is highly conserved from fish to humans.

Takeshi Takami1, Shigeru Yamano2, Sadanori Okada3, Mio Sakuma4, Takeshi Morimoto4, Hiroshi Hashimoto5, Satoshi Somekawa3, Yoshihiko Saito31Department of Internal Medicine, Clinic Jingumae, Kashihara, Japan; 2Department of Internal Medicine, Nara Rehabilitation Center, Nara, Japan; 3First Department of Internal Medicine, Nara Medical University, Kashihara, Japan; 4Center for General Internal Medicine and Emergency Care, Kinki University School of Medicine, Osaka, Japan; 5Department of Neurosurgery, Hashimoto Clinic, Kashihara, JapanPurpose: Blood pressure (BP), age, and reduced renal function are major risk factors for white-matter lesions (WMLs) in the general population. However, it remains unclear whether or not the BP itself or other parameters related to the BP are associated with WMLs in hypertensive patients with well-controlled BP. We investigated the relationships of the presence of WMLs with the central systolic BP (cSBP) and estimated glomerular filtration rate (eGFR) in treated hypertensive patients.Method: We studied 185 hypertensive patients with median duration of hypertension, 10.0 years, whose BP is controlled to SBP and diastolic BP (DBP) of 139 ± 17 and 79 ± 10 mmHg, respectively. We measured cSBP and brain magnetic resonance imaging (MRI) was examined within 2 weeks after last BP and biological measurements.Results: Patients with higher-grade WMLs, as assessed by the presence of Scheltens deep white-matter hyperintensity (SDWMH) in the frontal (grade 0–2 vs 3–6) and parietal areas (grade 0–2 vs 3–6) where small arteries are affected at earlier stage of hypertension, as well as that of Fazekas deep white-matter hyperintensity (FDWMH) (grade 2–3 vs 0–1) and Fazekas periventricular hyperintensity (FPVH) (grade 1–3 vs 0) were older, had higher serum creatinine levels, a longer duration of hypertension, and lower eGFR values. The grade of the WMLs was not associated with either the cSBP or the brachial SBP. In logistic regression analyses after adjustment for age, sex, cSBP, and hypertension duration, showed significant association between eGFR and WMLs. The patients with lower eGFR (

Purpose. To evaluate the influence of effective energy on computed tomography (CT) number in monoenergetic images (MEIs). Methods. Three bottle phantoms filled with water, oil, and a contrast agent were scanned at 100 and 140 kVp tube energy with a dual-source CT scanner. Cardiac dual-energy CT data was collected from 17 patients. CT numbers were measured in the 3 phantom materials and in the left ventricular cavity, myocardium, pericardial fat, and vertebral bone in MEIs from 40 to 190 keV. Results. In the phantoms, the mean CT number increased in oil whereas it decreased in the contrast agent as the energy level increased (P < 0.001). In clinical subjects, the mean CT numbers for the left ventricular cavity, myocardium, and vertebral bone were highest in the 40 keV images (P < 0.001) and decreased as the energy level increased. In contrast, the CT number for pericardial fat was lowest in the 40 keV images (P < 0.001) and increased with increasing energy. Conclusions. The influence of effective energy on CT number varies with material and tissue type in monoenergetic cardiac imaging, which could evaluate tissue characteristics through assessment of the changes in CT number associated with effective energies.

Aims Ablation of right-sided accessory pathways (APs) is sometimes challenging because several anatomical features of the tricuspid annulus (TA) and surrounding structures differ from those of the mitral annulus. This study investigated the electrophysiological characteristics and efficacy of a non-contact mapping (NCM) system for catheter ablation of right-sided APs. Methods and results We examined nine APs in six consecutive patients who underwent catheter ablation of right-sided APs with NCM. In Case 6, we compared NCM with contact activation mapping. Three of six patients had two APs, and one of these had previously failed ablation. We observed atrial activation during sinus rhythm or atrial pacing using a multiple-electrode array (MEA) deployed in the right atrium near the TA. Non-contact mapping identified the AP location as a peri-TA breakout point that appeared prior to or simultaneously with the delta wave onset in all APs. In Case 6 we confirmed that the peri-TA breakout identified by NCM corresponded to the earliest ventricular activation identified by contact mapping. We successfully ablated nine APs by radiofrequency (RF) energy application to the breakout sites, while one AP located just above the pole of the MEA required additional conventionally guided mapping and ablation. The mean RF duration was 189.8 ± 119.0 s. After 33.2 ± 9.4 months of follow-up, one para-hisian AP and one right lateral AP recurred, but these were successfully ablated in a second procedure using NCM. Conclusion Non-contact mapping was able to identify the location of right-sided APs accurately and quickly.

Dual-energy computed tomography (DE-CT) uses polyenergetic X-rays at 100- and 140-kVp tube energy, and generates 120-kVp composite images that are referred to as polyenergetic images (PEIs). Moreover, DE-CT can produce monoenergetic images (MEIs) at any effective energy level. We evaluated whether the image quality of coronary angiography is improved by optimizing the energy levels of DE-CT. We retrospectively evaluated data sets obtained from 24 consecutive patients using cardiac DE-CT at 100- and 140-kVp tube energy with a dual-source scanner. Signal-to-noise ratios (SNRs) were evaluated in the left ascending coronary artery in PEIs, and in MEIs reconstructed at 40, 50, 60, 70, 80, 90, 100, 130, 160 and 190 keV. Energy levels of 100, 120 and 140 kVp generated the highest SNRs in PEIs from 10, 12 and 2 patients, respectively, at 60, 70 and 80 keV in MEIs from 2, 10 and 10 patients, respectively, and at 90 and 100 keV in those from one patient each. Optimization of the energy level for each patient increased the SNR by 16.6% in PEIs (P < 0.0001) and by 18.2% in MEIs (P < 0.05), compared with 120-kVp composite images. The image quality of coronary angiography using DE-CT can be improved by optimizing the energy level for individual patients.

Background: Clinical studies using invasive modalities have reported that statin therapy stabilizes coronary plaque vulnerability. The serial changes of lipid-rich coronary plaques (LRCPs) during rosuvastatin treatment were evaluated non-invasively in patients with acute coronary syndrome (ACS) using dual-source computed tomography (DSCT). Methods and Results: A total of 11 consecutive ACS patients, and 13 LRCPs were serially evaluated on DSCT before and 24 weeks after rosuvastatin treatment. Compared with the baseline, there was no change in post-treatment minimal lumen diameter, lumen volume, or longitudinal length of LRCPs. By contrast, the ratio of lipid core volume to plaque volume significantly decreased from 48.0±9.9% to 43.7±10.6% (P=0.04), and plaque volume decreased from 144.5±85.5mm3 to 119.8±78.0mm3 (P=0.07). The remodeling index of target LRCPs significantly decreased from 1.16±0.10 to 1.06±0.12 (P=0.02). Percent reduction of plaque volume was significantly greater in patients with a lower ratio of low-density lipoprotein to high-density lipoprotein (L/H ratio ≤1.5) at follow-up than patients with higher L/H ratio (>1.5; median -31.7% vs. -6.8%, P=0.03). Conclusions: Rosuvastatin therapy reduced the volume of lipid cores and LRCPs and increased the CT attenuation value of LRCPs. DSCT is an effective modality for the non-invasive evaluation of LRCPs in patients with ACS. (Circ J 2011; 75: 2621-2627)

Background Contrast-enhanced cardiac magnetic resonance imaging (MRI) can depict papillary muscle (PM) necrosis or fibrosis by late enhancement (LE) of PM, but its clinical significance in old myocardial infarction (OMI) has been little understood. Methods Myocardial LE and PM-LE were detected with contrast imaging in 60 patients with OMI caused by a single culprit coronary artery lesion. Left ventricular (LV) morphology and function, mitral valve geometry, and severity of mitral regurgitation were also evaluated by cine imaging. Sphericity index was calculated for the assessment of LV remodeling. Results PM-LE was detected in 32 of 60 (53.3%) OMI patients. Unilateral PM-LE was detected in 22 patients and bilateral PM-LE in 10 patients. Patients with bilateral PM-LE demonstrated more severe LV remodeling and functional mitral regurgitation than those with unilateral or no PM-LE (sphericity index; bilateral PM-LE, 1.60±0.15, unilateral PM-LE, 1.71±0.29, no PM-LE, 1.85±0.27, p ≤0.05) (mitral regurgitation; bilateral PM-LE, 1.10±0.57, unilateral PM-LE, 0.41±0.73, no PM-LE, 0.54±0.84, p ≤0.05). In cases of unilateral PM-LE, posteromedial PM-LE resulting from right coronary artery-related OMI was accompanied by less severe mitral regurgitation, while anterolateral PM-LE resulting from left coronary artery-related OMI was not associated with severity of mitral regurgitation. Conclusions More than half of patients with OMI showed unilateral or bilateral PM-LE, and bilateral PM-LE was closely related to more severe LV remodeling and functional mitral regurgitation.

Echocardiography and magnetic resonance imaging revealed biventricular non-compaction cardiomyopathy with ventricular (VSD) and atrial (ASD) septal defects in an unconscious, 23-year-old hypoxemic man. Doppler echocardiography showed a left-to-right shunt across the VSD and a right-to-left shunt across the ASD. Cardiac catheterization revealed elevated right atrial pressure, although pulmonary pressure was normal. We considered that the atrial right-to-left shunt had induced the hypoxemia, which was related mainly to right ventricular dysfunction in this biventricular non-compaction cardiomyopathy, but it was not related to pulmonary hypertension.

The purpose of this study was to clarify the characteristics of black-blood echo-planar imaging (BB-EPI) in the assessment of infarct-related myocardial edema (IRME), compared with T2-weighted imaging (T2WI). Thirteen acute myocardial infarction (MI) patients after reperfusion and 11 old MI patients underwent BB-EPI and T2WI, excluding those with posterior MI. In acute MI patients, signal intensity ratio (SI ratio) of edema to normal myocardium was measured. Black-blood echo-planar imaging revealed hyperintensity in the same region identified as IRME on T2WI in all acute MI patients, and SI ratio was significantly higher in BB-EPI (2.66 +/- 1.58) than in T2WI (1.44 +/- 0.22) (P0.05). However, BB-EPI showed hyperintensity in posterior wall, where there is no clinical evidence of acute MI, in 2 out of 13 acute MI patients. Both T2WI and BB-EPI detected no IRME in known old infarct area of all old MI patients, but BB-EPI showed hyperintensity in the posterior wall of 4 out of 11 old MI patients. Black-blood echo-planar imaging can depict IRME with sufficient suppression of background and blood flow signals, and with excellent edema-to-normal myocardium contrast resolution. However, BB-EPI sometimes shows an inconsistent signal area with T2WI specifically in posterior wall. The wide practical use of BB-EPI requires the solution to this serious problem.

We present an alternative method for evaluating cardiac fat tissue-dual gradient-echo in-phase and opposed-phase magnetic resonance imaging (IPOP-MRI) with electrocardiographic (ECG) gating. Conventional IPOP-MRI can be used to evaluate small amounts of fat and is widely used for abdominal imaging, but cardiac motion artifacts make its use difficult for cardiac imaging. Using ECG gating prior to IPOP-MRI, we evaluated lipomatous metaplasia after myocardial infarction. The areas of lipomatous metaplasia measured by IPOP-MRI with ECG gating correlated well with those areas on black-blood T(1)-weighted imaging (r=0.82, P

Background— In-stent thrombosis is mainly triggered by adenosine diphosphate (ADP)-dependent platelet aggregation after percutanous coronary stent implantation. Ectonucleoside triphosphate diphosphohydrolase (E-NTPDase) rapidly hydrolyzes ADP to adenosine monophosphate, inhibiting platelet aggregation. We tested the hypothesis that local delivery of human placental E-NTPDase (pE-NTPDase) gene into injured arteries via gene-eluting stent could prevent subacute in-stent thrombosis. Methods and Results— We generated gene-eluting stents by coating bare metal stents with cationic gelatin hydrogel containing pE-NTPDase cDNA (pE-NTPDase stent), and implanted the stents into rabbit femoral arteries (FA) prone to production of platelet-rich thrombi due to repeated balloon injury at 4-week intervals. After the second injury, E-NTPDase gene expression was severely decreased; however, the implantation of pE-NTPDase stent increased E-NTPDase mRNA levels and NTPDase activity to higher level than normal FA. The FAs with pE-NTPDase stents maintained patency in all rabbits ( P Conclusions— Human pE-NTPDase gene transfer via cationic gelatin-coated stents inhibited subacute in-stent thrombosis and suppressed neointimal hyperplasia and inflammation without antiplatelet drugs.

Background: Late stent thrombosis related to delayed neointimal growth is a major concern after drug-eluting stent (DES) implantation. The time course of neointimal growth and risk factors of uncovered stent struts after sirolimus-eluting stent (SES) was studied using optical coherence tomography (OCT). Methods and Results: The 60 patients were enrolled and classified into G1 (follow-up period 25 months, n=15). The time elapsed since SES implantation was associated with a significant increase in mean neointimal area and neointimal thickness, and also with a significant decrease in the number of uncovered stent struts (G1: 14.8%, G2: 11.7%, and G3: 4.1%, P

Background Placental growth factor (PlGF), a homolog of vascular endothelial growth factor, is reported to stimulate angiogenesis and arteriogenesis in pathological conditions. It was recently demonstrated that PlGF is rapidly produced in myocardial tissue during acute myocardial infarction (MI). However, the effects of exogenous PlGF administration on the healing process after MI are not fully understood. The purpose of the present study was to examine whether PlGF treatment has therapeutic potential in MI. Methods and results Recombinant human PlGF (rhPlGF: 10 microg) was administered continuously for 3 days in a mouse model of acute MI. rhPlGF treatment significantly improved survival rate after MI and preserved cardiac function relative to control mice. The numbers of CD31-positive cells and alpha-smooth muscle actin-positive vessels in the infarct area were significantly increased in the rhPlGF group. Endothelial progenitor cells (Flk-1(+)Sca-1(+) cells) were mobilized by rhPlGF into the peripheral circulation. Furthermore, rhPlGF promoted the recruitment of GFP-labeled bone marrow cells to the infarct area, but only a few of those migrating cells differentiated into endothelial cells. Conclusions Exogenous PlGF plays an important role in healing processes by improving cardiac function and stimulating angiogenesis following MI. It can be considered as a new therapeutic molecule.

Mineralocorticoid receptor (MR) blockers attenuate cardiac remodeling in experimental models of heart failure, myocardial infarction and pressure-overload, in which the renin-angiotensin-aldosterone system is activated. Mice lacking the gene encoding guanylyl cyclase-A (GC-A), a common receptor for atrial and brain natriuretic peptide (ANP and BNP, respectively), show marked cardiac hypertrophy and fibrosis, which are almost completely inhibited by both genetic and pharmacological blockade of type 1 angiotensin II receptors. However, the effect of eplerenone, a specific MR blocker, on cardiac remodeling in GC-A knockout (GC-A KO) mice remains unknown. Male 12-week-old GC-A KO mice were assigned to control, eplerenone and hydralazine groups (n=6-7/group). Treatment with eplerenone at a dose of 100 mg/kg body weight/d reduced heart weight/body weight ratios, interstitial fibrosis and blood pressure to levels similar to those seen in wild type mice, in association with reduced transcription of atrial natriuretic peptide, brain natriuretic peptide, transforming growth factor-beta1, collagen I and collagen III. Although hydralazine (5 mg/kg body weight/d) exerted a similar effect on blood pressure, it did not inhibit the cardiac remodeling in GC-A KO mice. In conclusion, eplerenone attenuates cardiac remodeling in GC-A KO mice, most likely in a blood pressure-independent manner, which suggests that signaling downstream of MR is involved in the ventricular remodeling of GC-A KO mice.

Vascular endothelial cadherin (VE-cadherin) is expressed on vascular endothelial cells, which are involved in developmental vessel formation. However, it remains elusive how VE-cadherin–expressing cells function in postnatal neovascularization. To trace VE-cadherin–expressing cells, we developed mice expressing either green fluorescent protein or LacZ driven by VE-cadherin promoter using Cre-loxP system. Although VE-cadherin promoter is less active after birth than during embryogenesis in blood vessels, it is reactivated on cardiac ischemia. Both types of reporter-positive cells are found in the vasculature and in the infarcted myocardium. Those found in the vasculature were pre-existing endothelial cells and incorporated endothelial progenitor cells derived from extracardiac tissue. In addition to the vasculature, VE-cadherin promoter-activated cells were positive for CD45 in the bone marrow cells of the infarcted mice. VE-cadherin promoter–reactivated CD45-positive leukocytes were also found in the infarcted area. In addition, VE-cadherin promoter was activated in the bone marrow vessels of the infarcted mice. Collectively, our findings reveal a new ischemia-induced neovascularization mechanism involving VE-cadherin; the re-expressed VE-cadherin–mediated cell adhesion between cells may be involved not only in homing of bone marrow–derived cells to ischemic area but also mobilization from bone marrow.

Objectives Our aim was to investigate cardiac expression of placental growth factor (PlGF) and its clinical significance in patients with acute myocardial infarction (AMI). Background Placental growth factor is known to stimulate wound healing by activating mononuclear cells and inducing angiogenesis. The clinical significance of PlGF in AMI is not yet known. Methods Fifty-five AMI patients and 43 control subjects participated in the study. Peripheral blood sampling was performed on days 1, 3, and 7 after AMI. Blood was also sampled from the coronary artery (CAos) and the coronary sinus (CS), before and after acute coronary recanalization. Cardiac expression of PlGF was analyzed in a mouse AMI model. Results In AMI patients, peripheral plasma PlGF levels on day 3 were significantly higher than in control subjects. Plasma PlGF levels just after recanalization were significantly higher in the CS than the CAos, which indicates cardiac production and release of PlGF. Peripheral plasma levels of PlGF on day 3 were negatively correlated with the acute phase left ventricular ejection fraction (LVEF), positively correlated with both acute phase peak peripheral monocyte counts and chronic phase changes in LVEF. Placental growth factor messenger ribonucleic acid expression was 26.6-fold greater in a mouse AMI model than in sham-operated mice, and PlGF was expressed mainly in endothelial cells within the infarct region. Conclusions Placental growth factor is rapidly produced in infarct myocardium, especially by endothelial cells during the acute phase of myocardial infarction. Placental growth factor might be over-expressed to compensate the acute ischemic damage, and appears to then act to improve LVEF during the chronic phase.

Efonidipine can block both L- and T- type Ca2+ channels. In a previous in vitro study, we clarified that efonidipine dramatically suppresses aldosterone secretion from human adrenocortical tumor cells during angiotensin II (Ang II)- and K+-stimulation, whereas nifedipine, a dominant L-type Ca2+ channel antagonist, does not. This study was conducted to assess the in vivo effects of efonidipine and nilvadipine on the plasma aldosterone concentration. Placebo, 40 mg of efonidipine, or 2 mg of nilvadipine was administered to five healthy male volunteers. Hemodynamic parameters (pulse rate [PR] and blood pressure [BP]), plasma concentrations of neurohormonal factors (plasma renin activity, Ang II, aldosterone, and adrenocorticotropic hormone [ACTH]), and serum concentrations of Na+ and K+ were measured before and 6 h after administration of the agents. All three agents had little effect on PR and BP. Efonidipine and nilvadipine significantly increased plasma renin activity and Ang II. Both had little effect on ACTH, Na+, and K+. The plasma aldosterone concentration was significantly decreased after efonidipine treatment (88.3 +/- 21.3 to 81.6 +/- 24.9 pg/ml, p = 0.0407), whereas it was significantly increased after nilvadipine treatment (66.5 +/- 12.2 to 82.17 +/- 16.6 pg/ml, p = 0.0049). Placebo had little effect on neurohormonal factors. Efonidipine decreased plasma aldosterone concentration despite the increase in plasma renin activity and Ang II, suggesting that T-type Ca2+ channels may also play an essential role in the secretion of aldosterone in healthy human volunteers.

We present the rare case of a 51-year-old man with acute myocardial infarction (AMI) caused by left main coronary artery (LMCA) occlusion due to intra-aortic thrombus. He arrived at our hospital with sudden left precordial oppression. Transcatheter coronary angiography for suspected AMI detected no coronary lesions. After 10 minutes, noncontrast computed tomography (CT) confirmed significantly high transmural attenuation areas suggesting myocardial infarction in the septal, anterior and lateral wall. Contrast-enhanced CT and transesophageal echocardiography detected a small mass near the ostium of the LMCA in the coronary sinus of Valsalva. We thus considered that the mass occluded the LMCA, which led to AMI. A stent was implanted to prevent the mass from sealing the LMCA ostium, but the patient died of sepsis. A post-mortem examination clarified that the mass was a red thrombus attached to an erosive and atherosclerotic aortic wall.

Endothelial-mesenchymal transformation (EndMT) is essential for endocardial cushion formation during cardiac morphogenesis. We recently identified Tmem100 as an endothelial gene indispensable for vascular development. In this study, we further investigated its roles for EndMT during atrioventricular canal (AVC) cushion formation.Tmem100 was expressed in AVC endocardial cells, and Tmem100 null embryos showed severe EndMT defect in the AVC cushions. While calcineurin-dependent suppression of vascular endothelial growth factor (VEGF) expression in the AVC myocardium is important for EndMT, significant up-regulation of Vegfa expression was observed in Tmem100 null heart. EndMT impaired in Tmem100 null AVC explants was partially but significantly restored by the expression of constitutively-active calcineurin A, suggesting dysregulation of myocardial calcineurin-VEGF signaling in Tmem100 null heart. Moreover, Tmem100 null endocardial cells in explant culture did not show EndMT in response to the treatment with myocardium-derived growth factors, transforming growth factor β2 and bone morphogenetic protein 2, indicating involvement of an additional endocardial-specific abnormality in the mechanism of EndMT defect. The lack of NFATc1 nuclear translocation in endocardial cells of Tmem100 null embryos suggests impairment of endocardial calcium signaling.The Tmem100 deficiency causes EndMT defect during AVC cushion formation possibly via disturbance of multiple calcium-related signaling events.

Werner's syndrome is an autosomal recessive disorder characterized by premature aging [1]. Werner's syndrome patients have genetic disorders such as mutations in the WRN gene that cause rapid replicative senescence, chromosomal instability and sensitivity to various DNA damaging agents [2]. Severe toxicity following induction chemotherapy in a patient with Werner's syndrome has been reported previously [3]. Patients with Werner's syndrome develop a variety of benign and malignant tumors, predominantly of mesenchymal origin. The most common tumor type is sarcoma [4]. Despite the high incidence of sarcoma, there is little information regarding optimal treatment for these patients. Ifosfamide is used in the treatment of various cancers. Common side effects are encephalopathy, myelosuppression, hemorrhagic cystitis, acute renal failure, Fanconi's syndrome, and interstitial pneumonia. Diabetes insipidus (DI) is a condition characterized by excessive thirst and excretion of large amounts of severely diluted urine, with

Background: Benefit of low-dose aspirin for primary prevention of cardiovascular events in diabetes remains controversial. The American Diabetes Association (ADA), the American Heart Association (AHA), and the American College of Cardiology Foundation (ACCF) recommend aspirin for high-risk diabetic patients: older patients with additional cardiovascular risk factors. We evaluated aspirin’s benefit in Japanese diabetic patients stratified by cardiovascular risk. Methods and Results: In the JPAD trial, we enrolled 2,539 Japanese patients with type 2 diabetes and no history of cardiovascular disease. We randomly assigned them to aspirin (81–100mg daily) or no aspirin groups. The median follow-up period was 4.4 years. We stratified the patients into high-risk or low-risk groups, according to the US recommendation: age (older; younger) and coexisting cardiovascular risk factors. The risk factors included smoking, hypertension, dyslipidemia, family history of coronary artery disease, and proteinuria. Most of the patients were classified into the high-risk group, consisting of older patients with risk factors (n=1,804). The incidence of cardiovascular events was higher in this group, but aspirin did not reduce cardiovascular events (hazard ratio [HR], 0.83; 95% confidence interval [CI]: 0.58–1.17). In the low-risk group, consisting of older patients without risk factors and younger patients (n=728), aspirin did not reduce cardiovascular events (HR, 0.55; 95% CI: 0.23–1.21). These results were unchanged after adjusting for potential confounding factors. Conclusions: Low-dose aspirin is not beneficial in Japanese diabetic patients at high risk. (Circ J 2013; 77: 3023–3028)

Background: Accumulating evidence suggests that hematopoiesis, especially erythropoiesis, is disturbed in heart failure (HF) for many reasons. Low hemoglobin and red blood cell distribution width have emerged as prognostic indicators of HF independent of classic predictors. The prognostic implication of mean corpuscular volume (MCV) in HF, however, is unknown. In this context, we investigated the relationship between MCV and prognosis of acute decompensated HF (ADHF). Methods and Results: This retrospective cohort study consisted of 458 consecutive patients with ADHF who had emergency admission to hospital. Patients were divided into 2 groups: MCV ≤100fl (non-macrocytic group, n=400); and MCV >100fl (macrocytic group, n=58). The relationship between MCV and all-cause death was tested using Cox proportional hazard models, adjusting for other predictors. Mean patient age was 72.4 years and mean MCV was 93.0±7.1fl. Hemoglobin was significantly lower in the macrocytic group than the non-macrocytic group. During the mean follow-up of 20.8 months, a total of 173 deaths (37.9%) occurred. Kaplan-Meier analysis showed that all-cause death was significantly higher in the macrocytic group (log-rank P

Background Despite marked improvements in treatment strategies for heart failure (HF), the mortality rate of elderly patients with HF is still high. Detailed causes of death have not been fully understood. Methods and Results We studied 459 consecutive patients with acute decompensated HF (ADHF) emergently admitted to our hospital from 2007 to 2011. Patients were divided into 2 groups

OBJECTIVE To investigate the predictive values of placental growth factor (PlGF) and its endogenous antagonist, soluble fms-like tyrosine kinase-1 (sFlt-1), for the long-term prognosis of patients with stable coronary artery disease (CAD). Both PlGF and sFlt-1 play important roles in the pathological mechanisms of atherosclerosis. We recently demonstrated that the plasma levels of these molecules are correlated with the severity of coronary atherosclerosis. METHODS We enrolled 464 patients with stable CAD who consecutively underwent coronary angiography. Baseline blood samples were collected from the femoral artery immediately before coronary angiography (after the administration of 20 units of heparin), and the plasma levels of PlGF and sFlt-1 were measured. A Cox proportional hazard regression analysis was performed to evaluate the relationship between these parameters and the occurrence of all-cause death (ACD) and total cardiovascular events (TCVE) during a median follow-up of 3.3 years. RESULTS A total of 31 ACDs and 51 TCVEs occurred. Patients with higher PlGF/sFlt-1 ratios (>4.22×10(-2)) had a significantly higher risk of both ACD and TCVE than patients with lower ratios (

Patients with chronic kidney disease (CKD) die of cardiovascular diseases for unknown reasons. Blood vessel formation in plaques and its relationship with plaque stability could be involved with signaling through the Flt-1 receptor and its ligands, vascular endothelial growth factor, and the closely related placental growth factor (PlGF). Flt-1 also exists as a circulating regulatory splice variant short-inhibitory form (sFlt-1) that serves as a decoy receptor, thereby inactivating PlGF. Heparin releases sFlt-1 by displacing the sFlt-1 heparin-binding site from heparin sulfate proteoglycans. Heparin could provide diagnostic inference or could also induce an antiangiogenic state. In the present study, postheparin sFlt-1 levels were lower in CKD patients than in control subjects. More importantly, sFlt-1 levels were inversely related to atherosclerosis in CKD patients, and this correlation was more robust after heparin injection, as verified by subsequent cardiovascular events. Knockout of apolipoprotein E (ApoE) and/or sFlt-1 showed that the absence of sFlt-1 worsened atherogenesis in ApoE-deficient mice. Thus, the relationship between atherosclerosis and PlGF signaling, as regulated by sFlt-1, underscores the underappreciated role of heparin in sFlt-1 release. These clinical and experimental data suggest that novel avenues into CKD-dependent atherosclerosis and its detection are warranted.

BackgroundBrain natriuretic peptide (BNP) and amino-terminal proBNP (NT-proBNP) are useful biomarkers for diagnosis and prediction of prognosis. Both of these peptides are elevated in patients with chronic kidney disease (CKD), but there is no evidence as to which peptide is the more suitable biomarker in patients with severe renal dysfunction.Methods and resultsThis retrospective cohort study evaluated patients with cardiovascular diseases (64.9±11.7 years, mean±SD). The end points were all-cause death and a composite end point of all-cause death, nonfatal myocardial infarction, nonfatal stroke, hospitalization for severe heart failure, and initiation of hemodialysis. Baseline plasma BNP and NT-proBNP levels, expressed as log-transformed data, were closely correlated in patients with CKD stages 1–3 (n=998) (r2=0.870, p

Members of the transforming growth factor-β superfamily play essential roles in various aspects of embryonic development and physiological organ function. Among them, bone morphogenetic protein (BMP) 9 and BMP10 regulate embryonic vascular development by activating their endothelial receptor ALK1 (activin receptor-like kinase 1, also called Acvrl1). ALK1-mediated intracellular signaling is implicated in the etiologies of human diseases, but their downstream functional proteins are largely unknown. In this study, we identified Tmem100 , a gene encoding a previously uncharacterized intracellular transmembrane protein, to be an embryonic endothelium-enriched gene activated by BMP9 and BMP10 through the ALK1 receptor. Tmem100 null mice showed embryonic lethality due to impaired differentiation of arterial endothelium and defects of vascular morphogenesis, which phenocopied most of the vascular abnormalities observed with the Acvrl1/Alk1 deficiency. The activity of Notch- and Akt-mediated signaling, which is essential for vascular development, was down-regulated in Tmem100 null mice. Cre -mediated deletion of Tmem100 in endothelial cells was sufficient to recapitulate the null phenotypes. These data indicated that TMEM100 may play indispensable roles downstream of BMP9/BMP10-ALK1 signaling during endothelial differentiation and vascular morphogenesis.

Aims Morphological characteristics of non-significant coronary plaques (NSCPs) that develop rapid progression have not been fully elucidated. The aim of this study was to clarify the morphological characteristics of NSCPs in patients with coronary artery disease (CAD) using intravascular optical coherence tomography (OCT). Methods and results Fifty-three consecutive CAD patients undergoing percutaneous coronary intervention were enrolled and 69 NSCPs (per cent diameter stenosis

Clinical studies using invasive modalities have reported that statin therapy stabilizes coronary plaque vulnerability. The serial changes of lipid-rich coronary plaques (LRCPs) during rosuvastatin treatment were evaluated non-invasively in patients with acute coronary syndrome (ACS) using dual-source computed tomography (DSCT).A total of 11 consecutive ACS patients, and 13 LRCPs were serially evaluated on DSCT before and 24 weeks after rosuvastatin treatment. Compared with the baseline, there was no change in post-treatment minimal lumen diameter, lumen volume, or longitudinal length of LRCPs. By contrast, the ratio of lipid core volume to plaque volume significantly decreased from 48.0 ± 9.9% to 43.7 ± 10.6% (P=0.04), and plaque volume decreased from 144.5 ± 85.5 mm³ to 119.8 ± 78.0 mm³ (P=0.07). The remodeling index of target LRCPs significantly decreased from 1.16 ± 0.10 to 1.06 ± 0.12 (P=0.02). Percent reduction of plaque volume was significantly greater in patients with a lower ratio of low-density lipoprotein to high-density lipoprotein (L/H ratio ≤ 1.5) at follow-up than patients with higher L/H ratio (1.5; median -31.7% vs. -6.8%, P=0.03).Rosuvastatin therapy reduced the volume of lipid cores and LRCPs and increased the CT attenuation value of LRCPs. DSCT is an effective modality for the non-invasive evaluation of LRCPs in patients with ACS. ).

Background Dual gradient-echo in-phase (IP) and opposed-phase (OP) magnetic resonance imaging (MRI) can evaluate small amounts of fat, and is widely used for imaging abdominal organs. Epicardial fat has recently been revealed as having an important role in the development of heart disease, but IPOP-MRI has not been widely applied in this field, probably because of cardiac motion artifacts. Purpose To demonstrate the feasibility of IPOP-MRI with electrocardiography (ECG)-gating for evaluation of epicardial fat in the normal human heart, compared to black-blood T1-weighted imaging (T1WI). Material and Methods Ten healthy volunteers were studied. Epicardial fat volume was quantitatively evaluated on OP-from-IP subtraction images and T1-weighted images. Results Epicardial fat was clearly visualized on the subtraction images as hyperintense areas that corresponded to regions identified as epicardial fat on the T1-weighted images. Epicardial fat volume correlated well between the two methods (r = 0.93, P Conclusion IPOP-MRI with ECG-gating may be a useful alternative method to black-blood T1WI for evaluation of epicardial fat.

Background— Renal dysfunction is commonly accompanied by a worsening of atherosclerosis; however, the underlying molecular mechanism is not fully understood. We examined the role played by soluble fms-like tyrosine kinase-1 (sFlt-1), an endogenous antagonist of the proatherogenic cytokine placental growth factor (PlGF), in the worsening of atherosclerosis in patients with renal dysfunction and in an animal model of renal failure. Methods and Results— In this study, 329 patients who received cardiac catheterization and 76 patients who underwent renal biopsy were enrolled. Both plasma sFlt-1 levels and renal sFlt-1 mRNA expression were positively correlated with estimated glomerular filtration rate ( P P Conclusions— The present study demonstrates that a reduction in the circulating levels of sFlt-1 is associated with the worsening of atherosclerosis that accompanies renal dysfunction.

Placental growth factor and vascular endothelial growth factor increase angiogenesis and promote healing after acute myocardial infarction (MI), but the significance of soluble Fms-like tyrosine kinase-1 (sFlt-1), an antagonist of placental growth factor and vascular endothelial growth factor, in the setting of acute MI has not been elucidated. The development of acute heart failure in the immediate period after MI is a dreaded complication, but there are no useful biomarkers that identify patients at risk of acute heart failure. We wished to investigate the clinical significance of circulating sFlt-1 during acute MI. We enrolled 174 patients with acute MI, and arterial blood sampling was performed. Plasma levels of sFlt-1 were measured by enzyme-linked immunosorbent assay and their relation to clinical parameters was analyzed. Circulating levels of sFlt-1 on admission were significantly increased in patients with acute MI compared to controls (528.1 +/- 290.9 vs 355.7 +/- 205.0 pg/ml, p

Aldosterone synthase (CYP11B2) and 11β-hydroxylase (CYP11B1) regulate aldosterone and cortisol production, respectively. The expression of these enzymes is promoted by calcium influx through Cav3.2, a T-type calcium channel. Neuron-restrictive silencer factor (NRSF) binds to neuron-restrictive silencer element (NRSE) to suppress the transcription of NRSE-containing genes. We found a NRSE-like sequence in human CYP11B2 and CYP11B1 genes as well as the CACNA1H gene of many mammalian species. The CACNA1H gene encodes the α-subunit of Cav3.2. Here we investigated how NRSF/NRSE regulates aldosterone and cortisol synthesis. Inhibition of endogenous NRSF by an adenovirus-expressing dominant-negative NRSF (AD/dnNRSF) increased human CYP11B2 and CYP11B1 mRNA expression, leading to aldosterone and cortisol secretion in human adrenocortical (H295R) cells. In reporter gene experiments, NRSE suppressed luciferase reporters driven by CYP11B2 and CYP11B1 promoters and dnNRSF enhanced them. Moreover, cotransfection of dnNRSF increased luciferase activity of reporter genes after deletion or mutation of NRSE, suggesting that NRSF/NRSE regulates transcription of CYP11B2 and CYP11B1 genes indirectly. AD/dnNRSF augmented mRNA expression of rat CYP11B2 and CYP11B1 genes, neither of which contains a NRSE-like sequence in rat adrenal cells. AD/dnNRSE also significantly increased CACNA1H mRNA in H295R and rat adrenal cells. Efonidipine, a T/L-type calcium channel blocker, significantly suppressed dnNRSF-mediated up-regulation of CYP11B2 and CYP11B1 expression. Moreover, NRSF/NRSE is also involved in angiotensin II- and K+-stimulated augmentation of CYP11B2 and CYP11B1 gene transcription. In conclusion, NRSF/NRSE controls aldosterone and cortisol synthesis by regulating CYP11B2 and CYP11B1 gene transcription mainly through NRSF/NRSE-mediated enhancement of the CACNA1H gene.

Background Coronary spasm (CS) plays an important role in the pathogenesis of many types of ischemic heart disease, but morphological appearance of non-stenotic coronary segments with CS is not fully understood. We evaluate the morphological characteristics of coronary arteries in patients with coronary spastic angina (CSA) using intravascular optical coherence tomography (OCT). Methods We evaluated 37 patients with resting chest pain whose coronary angiograms did not reveal significant stenosis. These patients underwent an acetylcholine (ACh) provocation test. OCT was performed after complete dilatation of coronary arteries, and additionally during ACh-induced CS in four patients. Results Based on the ACh test, 23 patients were diagnosed as having CSA, and the remaining 14 patients without CS were referred to as CS-negative. OCT study revealed that coronary segments with ACh-induced CS had homogeneous intimal thickening, and quantitative analysis showed that CS-positive segments had a significantly greater intima area as compared with corresponding CS-negative segments without lipid or calcium content. By contrast, CS-positive segments had a significantly smaller intima area as compared with CS-negative segments with lipid or calcium deposit. During ACh-induced CS, lumen and total vascular areas were significantly decreased, whereas intima area did not change in comparison with complete vasodilatation. The luminal surface of the intima formed a markedly wavy configuration during CS. Conclusions Coronary artery segments involved in CS are characterized by diffuse intimal thickening without lipid or calcium content. High-resolution coronary OCT imaging could make it possible to analyze the vascular pathophysiology in patients with CS.

Gadolinium-containing magnetic resonance imaging (MRI) contrast agents have been reported to induce nephrogenic systemic fibrosis in patients with renal dysfunction, and therefore their use has been restricted. However, even without contrast agents, MRI can provide much valuable information on the pathophysiology of renal diseases. This report describes the case of a 90-year-old man with a hydronephrotic and dysfunctioning right kidney induced by testicular cancer evaluated by non-contrast MRI. He was referred to our hospital complaining of lower abdominal distention and appetite loss. Physical examination revealed a painless enlargement of the right scrotum and inguinal lymph node. Laboratory analysis revealed renal dysfunction, after which a non-contrast MRI was performed. T1-, T2-, and diffusion-weighted images (T1WI, T2WI, and DWI, respectively) revealed carcinoma of the right testicle with extensive multiple lymphadenopathy. Moreover, a retroperitoneal metastatic tumor was detected adjacent to the right kidney, which appeared to constrict the right urinary duct. An enlarged right kidney with loss of corticomedullary differentiation was observed on T1WI. The right kidney and enlarged renal pelvis were observed as large signal intensity areas on T2WI. DWI showed an increased signal intensity of the right kidney and a decreased apparent diffusion coefficient. These findings clarified that the retroperitoneal metastasis from the right testicular cancer led to hydronephrosis and dysfunctioning of the right kidney. The present case indicates that non-contrast MRI is useful for the evaluation of renal diseases even in elderly patients with renal dysfunction.

Aim: We previously found, using a mouse model, that activation of proinflammatory cytokines after acute myocardial infarction (AMI) augments neointimal hyperplasia of a remote artery. The present study assessed the progression of luminal narrowing of non-culprit coronary arteries (NCCA) in patients following AMI.Methods: The study group comprised 21 AMI patients successfully treated with bare-metal stents and 16 stable angina (SA) patients treated with sirolimus-eluting stents. Clinical backgrounds were similar for both groups. Quantitative coronary angiography was performed before and after stent implantation and at 6-months of follow-up.Results: We evaluated 126 non-culprit coronary segments (73 in AMI and 53 in SA). The minimum lumen diameter (MLD) (mm) of NCCA decreased significantly from 2.61±0.79 to 2.44±0.71 in the AMI group, but changed only slightly from 2.02±0.56 to 2.02±0.50 in the SA group. The absolute change in the MLD of NCCA was significantly greater (0.17±0.53) in the AMI, than in the SA (0.0070±0.261) group.Conclusion: luminal narrowing of non-culprit coronary segments progressed in AMI patients within 6 months of stent implantation, but progressed only slightly in SA patients.

Coronary spasm (CS) plays an important role in the pathogenesis of many kinds of ischemic heart disease. However, morphological characteristics of coronary artery of CS remain unknown. We evaluated 37 patients with coronary spastic angina (CSA) who underwent diagnostic acetylcholine (ACh) provocation test, and 2 acute coronary syndrome patients suspected to have severe CS. The intravascular optical coherence tomography (OCT) was performed after complete dilatation of coronary artery in all patients and additionally performed during ACh-induced CS in 4 patients. Based on the ACh provocation test, 23 patients who developed CS and ischemic ECG changes were diagnosed as coronary spastic angina (CSA), and other 14 patients without CS were referred as CS-negative patients. CS-negative patients were further divided into 2 sub-groups according to the lipid and/or calcification content in coronary arterial wall by OCT findings. Intravascular OCT revealed most coronary segments with ACh-induced CS had homogenous intima thickening, and quantitative OCT analysis showed that CS-positive segments had significantly larger intima area compared with CS-negative segments without lipid and/or calcification (2.73±0.07 vs. 1.36±0.06 mm 2 , P2 , P

Background: Cardiac MRI permits diagnosis with acute myocardial infarction (AMI) by detection of infarct-related myocardial edema (IRME) on black-blood T2-weighted images (T2WIs), and myocardial scar on delayed-enhanced images (DEs). Diffusion-weighted image (DWI) is an established method for detection of cerebral infarction edema, but few reports are available on detection of IRME. Objective: To detect IRME using DWIs, and compare the contrast-to-noise ratio (CNR) with T2WIs. Methods: Cardiac MRI was performed on a 1.5T scanner (SIEMENS Avanto, Erlangen, Germany) in 12 AMI patients within 14 days after successful reperfusion. T2WIs were acquired using a double inversion-recovery fast-spin echo sequence. DWIs were acquired using combined cardiac and respiratory gated single-shot echo-planar sequence with a b-value of 50 s/mm 2 . DEs were acquired using an inversion-recovery segmented gradient-echo sequence, fifteen minutes after administering 0.15 mmol/kg gadolinium-DTPA. CNR was calculated in the T2WIs and DWIs using the following formula: CNR = (signal intensity[ME]-signal intensity[M]) / SD(B), where ME = myocardial edema, M = myocardium, and SD(B) =standard deviation of background signal intensity. Results: IRME was detected on T2WIs, and myocardial scar on DEs in all patients. DWIs revealed an area of significantly increased signal intensity in the same region identified as IRME on T2WIs in all patients. The CNR of DWIs (78.9 ± 44.5) was significantly higher than those of T2WIs (31.5 ± 23.3) (p = 0.0001). Conclusion: Our findings indicate that low b-value DWI provides better contrast in infarct-related myocardial edema than T2WIs.

Gap junctions (GJs) constituted by neighboring cardiac myocytes are essential for gating ions and small molecules to coordinate cardiac contractions. cAMP is suggested to be a potent stimulus for enhancement of GJ function. However, it remains elusive how cAMP potentiates the GJ of cardiomyocytes. Here we demonstrated that the gating function of GJ is enhanced by the protein kinase A (PKA)–dependent signal, and that the accumulation of connexin43 (Cx43), the most abundant Cx in myocytes, is enhanced by an exchange protein directly activated by cAMP (Epac) (Rap1 activator)–dependent signal. The gating function of GJs was analyzed by microinjected dye transfer method. The accumulation of Cx43 was analyzed by quantitative immunostaining. Using the PKA-specific activator N 6 -benzoyladenosine-3′,5′-cyclic monophosphate (6Bnz) and Epac-specific activator 8-(4-chlorophenylthio)-2′- O -methyladenosine-3′,5′-cyclic monophosphate (8CPT), we could delineate the two important downstream signals of cAMP for enhanced GJ neoformation. Whereas 6Bnz potentiated gating function of GJs with slight accumulation of Cx43 at cell–cell contacts, 8CPT remarkably enhanced the accumulation of Cx43 with a slight effect on gating. We further noticed that adherens junctions (AJs) were maturated by 8CPT, as marked by increased neural-cadherin immunostaining. Because AJ formation precedes the GJ formation, AJ formation accelerated by Epac-Rap1 signal may result in enhanced GJ formation. The involvement of Epac-Rap1 signal in GJ neoformation was further confirmed by evidence that inactivation of Rap1 by overexpression of Rap1GAP1b perturbed the accumulation of Cx43 at cell–cell contacts. Collectively, PKA and Epac cooperatively enhance functional GJ neoformation in cardiomyocytes.

Cyclic AMP (cAMP) is a well-known intracellular signaling molecule improving barrier function in vascular endothelial cells. Here, we delineate a novel cAMP-triggered signal that regulates the barrier function. We found that cAMP-elevating reagents, prostacyclin and forskolin, decreased cell permeability and enhanced vascular endothelial (VE) cadherin-dependent cell adhesion. Although the decreased permeability and the increased VE-cadherin-mediated adhesion by prostacyclin and forskolin were insensitive to a specific inhibitor for cAMP-dependent protein kinase, these effects were mimicked by 8-(4-chlorophenylthio)-2′-O-methyladenosine-3′, 5′-cyclic monophosphate, a specific activator for Epac, which is a novel cAMP-dependent guanine nucleotide exchange factor for Rap1. Thus, we investigated the effect of Rap1 on permeability and the VE-cadherin-mediated cell adhesion by expressing either constitutive active Rap1 or Rap1GAPII. Activation of Rap1 resulted in a decrease in permeability and enhancement of VE-cadherin-dependent cell adhesion, whereas inactivation of Rap1 had the counter effect. Furthermore, prostacyclin and forskolin induced cortical actin rearrangement in a Rap1-dependent manner. In conclusion, cAMP-Epac-Rap1 signaling promotes decreased cell permeability by enhancing VE-cadherin-mediated adhesion lined by the rearranged cortical actin.