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CCAAT/enhancer-binding proteins (C/EBPs) upregulate transcription of various inflammatory cytokines and acute phase proteins, such as interleukin (IL)-1β, IL-6, tumor necrosis factor-α, and cyclooxygenase-2. Recent studies have demonstrated that peroxisome proliferator-activated receptor (PPAR)-γ is present in atherosclerotic lesions, and negatively regulates expression of these genes. Interestingly, PPAR-γ gene promoter has tandem repeats of C/EBP-binding motif, and C/EBP-δ plays a pivotal role in transactivation of PPAR-γ gene. It has been well known that the interaction between C/EBPs and PPAR-γ plays a central role in maintaining adipocyte differentiation and glucometabolism; however, the relationship between PPAR-γ and C/EBPs in the vessel wall remains unclear. In the present study, we showed that a high level of C/EBP-δ expression induced by inflammation positively regulated transcription and protein expression of PPAR-γ in vascular smooth muscle cells (VSMCs). On the other hand, PPAR-γ ligands troglitazone, pioglitazone, and 15-deoxy-Δ 12,14 -prostaglandin J 2 inhibited IL-1β-induced IL-6 expression at a transcriptional revel in VSMCs. Functional promoter analysis revealed that PPAR-γ ligands inhibited IL-1β-induced transactivation of IL-6 gene via suppression of not only nuclear factor-κB but also C/EBP-DNA binding. Moreover, PPAR-γ ligands suppressed protein expression and transcription of C/EBP-δ through dephosphorylation of signal transducer and activator of transcription 3. These findings strongly suggest that C/EBP-δ is negatively autoregulated via transactivation of PPAR-γ. This feedback mechanism probably downregulates transcription of inflammatory cytokines and acute phase proteins, and modulates inflammatory responses in the early process of atherosclerosis.

Background— In vitro studies suggest that angiotensin II type 1 and type 2 (AT 1 and AT 2 ) receptors exert opposite effects in terms of vasoconstriction, natriuresis, and cell growth, but the role of these receptors in cardiovascular remodeling in vivo is still an enigma. In this study, we tested the hypothesis that AT 2 exerts an antiproliferative effect by inducing apoptosis, thereby antagonizing AT 1a in vascular remodeling. Methods and Results— Vascular injury was induced by polyethylene cuff placement around the left femoral artery of AT 1a -null (AT 1a KO), AT 2 -null (AT 2 KO), and wild-type mice. Neointimal formation as well as DNA synthesis in vascular smooth muscle cells (VSMC) after vascular injury was exaggerated in AT 2 KO mice, but they were both suppressed in AT 1a KO mice compared with those in wild-type mice. In contrast, the number of apoptotic cells in the injured artery in VSMC was significantly increased in AT 1a KO mice but decreased in AT 2 KO mice. Reverse transcriptase–polymerase chain reaction analysis revealed that the expression of bax mRNA was attenuated in AT 2 KO mice. On the other hand, the expression of bcl-2 and bcl-x L mRNA was enhanced in AT 2 KO mice but attenuated in AT 1a KO mice. Immunohistochemical staining with antibody to the bcl-2 protein family supported these results. Conclusions— Our results suggest that AT 2 exerts antiproliferative effects and proapoptotic changes in VSMC by counteracting AT 1a in the process of neointimal formation after vascular injury.

Background The Fas-Fas ligand (FasL) system is involved in apoptosis in many types of cells. Recently, the expression of FasL on endothelial cells was reported. FasL is cleaved by a metalloproteinase and released in serum as soluble FasL (sFasL). Vasoactive substances, including metalloproteinase, are modulated by endothelial dysfunction. Advanced atherosclerosis and impaired endothelial function are seen in hypertensive patients. The inflammatory response has an important role in the development of atherosclerosis, whereas C-reactive protein (CRP) is associated with the presence and severity of atherosclerosis. Objective To measure the intima-media thickness of the common carotid artery and evaluate the relationship between atherosclerosis and serum sFasL concentrations in hypertensive patients. Patients and main outcome measures Forty-seven patients with hypertension participated in the study. The intima-media thickness of the common carotid artery was evaluated by ultrasound imaging. Serum concentrations of sFasL were measured by enzyme-linked immunosorbent assay. Results Intima-media thickness correlated positively with age (r = 0.362, P = 0.012) and sFasL concentrations (r =0.332, P = 0.022), and negatively with creatinine clearance (r = -0.399, P = 0.0055). A general linear model analysis with atherosclerotic risk factors and sFasL revealed that age, sFasL, high-density lipoprotein-cholesterol and systolic blood pressure were significantly associated with intima-media thickness. Furthermore, we demonstrated that serum sFasL is directly associated with CRP concentration (r = 0.316, P = 0.030). Conclusions These results indicated that serum sFasL concentration is associated with atherosclerosis and inflammatory disease, in patients with hypertension.

Growth arrest and DNA damage inducible gene 153 ( gadd153 ) is expressed at very low levels in growing cells but is markedly induced in response to cellular stresses, including glucose deprivation, exposure to genotoxic agents, and other growth-arresting situations. Forced expression of GADD153 can induce cell cycle arrest and/or apoptosis in many types of cells. Recently, we reported that GADD153 was induced in vascular smooth muscle cells (VSMCs) in neointimal lesions of balloon-injured carotid arteries. To investigate the underlying molecular mechanisms of gadd153 gene expression in VSMCs, we isolated and characterized a promoter region of the rat gadd153 gene. Sequence alignments of this region revealed 1 TATA-like sequence and several well-known cis elements. The 5′-deletion analysis for this region showed that a domain spanning −447 through −368 drastically reduced the promoter activity to almost equal levels of promoterless control. Because this domain contained a consensus sequence for the nuclear factor 1 family of proteins (NF1), DNA-binding studies were performed by use of 2 types of NF1 consensus probes. Both probes were specifically shifted by nuclear extracts from proliferating VSMCs and were supershifted by antiserum against CCAAT transcription factor/NF1. In addition, promoter activity of a mutant luciferase vector, which was generated by a point mutation at the NF1 binding motif of the gadd153 gene, was 14-fold higher than that of a wild-type one. These results suggest that gadd153 gene expression in VSMCs is negatively regulated by an NF1-binding motif, and NF1 may act as an antiapoptotic factor by continuously suppressing gadd153 gene expression in growing VSMCs.

Background and Purpose —Mechanical stresses on the arterial wall participate in the pathogenesis of atherosclerosis as local factors. The relationships between local mechanical forces and risk factors for atherosclerosis were investigated. Methods —Mechanical forces on the arterial wall were evaluated in the carotid artery in 117 patients with risk factors for atherosclerosis including hypertension, dyslipidemia, diabetes mellitus, and smoking, as well as in 20 age- and sex-matched normal controls. Circumferential wall tension and shear stress were evaluated with Laplace’s law and a poiseuillean parabolic model of velocity distribution. Circumferential wall strain was also evaluated as carotid mechanical force. Results —Mechanical forces in subjects with risk factors were characterized by low wall shear stress, high circumferential wall tension, and reduced strain. Systolic blood pressure was significantly negatively associated with shear stress and circumferential wall strain. HDL cholesterol showed a significant positive correlation with shear stress and a negative correlation with wall tension. Fasting blood glucose was significantly associated with shear stress, while smoking showed a negative correlation with shear stress and a positive correlation with wall tension. Accumulation of risk factors was associated with further deterioration of mechanical forces. Furthermore, stepwise regression analysis showed that the number of risk factors was significantly associated with mechanical forces independently of carotid intima-media thickness. Conclusions —These findings suggest that risk factors for atherosclerosis were associated with alteration of mechanical forces. Consequent alteration in mechanical forces could be an underlying local mechanism for the progression of atherosclerosis.

Abstract —Platelet-derived growth factor-α receptor (PDGF-αR) expression is markedly elevated in cultured vascular smooth muscle cells (VSMCs) from spontaneously hypertensive rats (SHR) when compared with normotensive rat strains, Sprague-Dawley, Wistar, and Wistar-Kyoto rats (WKY). This “almost-all-or-none” type of differential expression strongly suggests that PDGF-αR or its transcription-regulating mechanisms or factors are significantly related to genetic hypertension. To evaluate the role of PDGF-αR in vascular remodeling and hypertension, we have investigated the underlying molecular mechanism. We have recently shown that the regulatory domain responsible for this difference is localized to the PDGF-αR promoter region between –246 and –139, which contains an enhancer core sequence specific for CCAAT-enhancer binding proteins (C/EBPs). We defined the roles of this element for hypertensive strain-specific PDGF-αR gene transcription. DNA-protein binding studies by competition in electromobility shift and supershift assays revealed that 2 members, C/EBP-β and C/EBP-δ, are mainly responsible for DNA-protein complex formation; the former acts as a transcriptional repressor and the latter as an activator of the PDGF-αR gene, respectively. Western or Northern blot analyses supported evidence for high expression of C/EBP-δ seen only in SHR-derived VSMCs. Furthermore, forced expression of C/EBP-δ transactivated the transcriptional efficiency of the PDGF-αR gene even in WKY-derived VSMCs, whereas that of C/EBP-β had an opposite effect in SHR-derived VSMCs. These findings indicate that differential expression of members of the C/EBP family, mainly C/EBP-δ and possibly C/EBP-β, are responsible for the strain-specific gene transcription of PDGF-αR in VSMCs.

BACKGROUND Apoptotic cell death is the major form of myocardial damage produced by coronary ischemic events. OBJECTIVE To assess whether circulating levels of soluble Fas (sFas), an inhibitor of apoptosis, and sFas ligand, an inducer of apoptosis, in patients with coronary artery disease are greater than normal. METHODS Forty-seven patients [acute myocardial infarction (AMI) in 17, old myocardial infarction (OMI) in 15, stable angina in 15] and 10 normal control subjects participated in this study. Serum levels of sFas and sFas ligand in all patients were measured, and cardiac catheterizations were performed. RESULTS Serum levels of sFas were greater than normal only in patients with AMI (4.6 +/- 1.6 ng/ml); the levels were significantly higher than those in patients with OMI (2.1 +/- 0.6 ng/ml) and stable angina (2.2 +/- 0.5 ng/ml), and in normal subjects (2.0 +/- 0.6 ng/ml; P < 0.0001). However, there was no difference among serum levels of sFas ligand for all groups. For patients with AMI, there was no significant correlation between serum levels of sFas and peak levels both of plasma creatine phosphokinase and of plasma myosin light chain type I as clinical indexes of infarct size. However, there were significant correlations between serum levels of sFas and both pulmonary artery wedge pressure (r = 0.767, P = 0.0003) and left ventricular end-diastolic pressure (r = 0.629, P = 0.03). CONCLUSIONS Circulating sFas increases in concentration in relation to the severity of hemodynamic conditions in patients with AMI, but it is independent from size of infarct. Therefore, circulating sFas could play an important role as the marker of pathophysiologic conditions associated with cardiomyocyte apoptosis in AMI.

Abstract —Platelet-derived growth factor (PDGF) and its receptors are widely expressed in several tissues in the stage of cellular growth and development. In adulthood, PDGF β-receptor (PDGFβR) is mainly detected in pathological conditions such as atherosclerotic lesions and injured vascular wall. The purpose of the present study was to elucidate the underlying mechanism of PDGFβR gene expression under pathological conditions in vascular smooth muscle cells (VSMC) and to identify the important cis elements responsible for tissue-specific gene transcription. Gel mobility shift assay and supershift assay indicated that the CCAAT motif located at −67 (C67) was mainly interacted with NF-YC, and this element drove the basal promoter activity of the gene as a putative promoter. On the other hand, another important sequence essential for the basal transcription was found at a 30-bp region (R30) spanning −150 to −121. To test whether R30 actually regulates the tissue-specific transcription of PDGFβR gene, electromobility shift pattern was compared between VSMC and hepatoma cell line (HTC). We obtained the result that DNA-protein complex seen only in nuclear extracts from HTC suppressed the promoter activity in HTC in a tissue-specific manner. Furthermore, cis element decoy transfection experiments for C67 and R30 also revealed that both elements were functionally important in mRNA expression of PDGFβR in VSMC. From these results, we concluded that the basal activity of PDGFβR gene expression was transactivated by the interaction or coordination of both C67 and R30, and the latter one mainly controlled the tissue-specific gene expression in VSMC.

OBJECTIVE: To understand the regulatory mechanism of platelet-derived growth factor beta-receptor gene expression. METHODS: A 1.7 kb genomic fragment was obtained from a rat genomic library. After we had determined an entire sequence of this fragment, transcription start sites were determined both by primer extension analysis and by riboprobe mapping. We performed a functional promoter assay by using a dual-luciferase reporter system. Progressive 5'-deletions of the fragment and site-directed mutagenesis for the CCAAT motif located at -67 or -94 were used for the assay, and their promoter activities in vascular smooth muscle cells were assessed. Gel-mobility shift analysis was also performed for the CCAAT motif at -67. Effects of the upstream sequence spanning -310 through -120 on heterologous gene promoters were also investigated. RESULTS: Multiple transcription start sites were observed in the 5'-flanking region, and the 1.7 kb sequence was actually active as a functional promoter in vascular smooth muscle cells. Two important sequences responsible for the basal transcriptional activity were identified by the functional promoter assay. One was the CCAAT motif at -67 which acts as a promoter itself, and the other was the upstream region spanning -310 through -210 which positively regulates the basal promoter activity. CONCLUSION: The basal promoter activity of the rat platelet-derived growth factor beta-receptor gene is mainly regulated by the interaction or coordination of two sequences, the CCAAT motif and the upstream control element.

Objectives To clarify whether the functional changes during the transition from compensatory myocardial hypertrophy to failure are associated with changes in sarcoplasmic reticulum gene expression. Methods We examined the gene expression of sarcoplasmic reticulum proteins [sarcoplasmic reticulum Ca 2+ -ATPase (SERCA), phospholamban, calsequestrin and ryanodine receptor] in Dahl salt-sensitive (Dahl-S) rats fed a high-salt (8%) diet from the age of 6 weeks. In-vivo contractile functioning was evaluated using echocardiography, and gene expression of sarcoplasmic reticulum proteins in the left ventricle was analyzed by Northern blotting for each stage of left ventricular hypertrophy. Results SERCA messenger RNA (mRNA) levels in Dahl-S rats with compensatory hypertrophy did not change significantly, whereas phospholamban mRNA levels were increased by 61% (P

To elucidate whether there is a difference in the progression of target-organ damage between primary aldosteronism and essential hypertension, we compared left ventricular hypertrophy and extracardiac target-organ damage in 23 patients with primary aldosteronism and 116 patients with essential hypertension. The severity of hypertensive retinopathy and the renal involvement in primary aldosteronism were subclinical and similar to those in essential hypertension without left ventricular hypertrophy but significantly milder than those in essential hypertension with left ventricular hypertrophy. There was a strongly significant correlation between the degree of left ventricular mass index and the severity of hypertensive retinopathy and renal involvement independent of office blood pressure in essential hypertension. In contrast, left ventricular hypertrophy markedly progressed despite the mild extracardiac target-organ damage in primary aldosteronism. Left ventricular end-diastolic dimension index in primary aldosteronism (3.16±0.50 cm/m 2 ) was significantly larger than in essential hypertension without (2.87±0.23) and with (2.88±0.22) left ventricular hypertrophy. On the other hand, there was no difference in extracardiac target-organ damage between 13 primary aldosteronism patients with eccentric left ventricular hypertrophy and the 26 essential hypertensive patients with eccentric left ventricular hypertrophy. The results suggest that predominantly volume load, be it due to aldosteronism or other mechanisms, resulting in eccentric left ventricular hypertrophy is less likely to cause extracardiac target-organ damage than hemodynamic or nonhemodynamic mechanisms resulting in concentric left ventricular hypertrophy.

Abstract Autonomic nervous function was evaluated by means of power spectral analysis of heart rate variability in hospitalized dipper (n=31) and non-dipper (n=31) essential hypertensive subjects. Twenty-four–hour blood pressure (BP) measurement was performed by the cuff-oscillometric method to evaluate the nocturnal decrease of BP. The non-dipper subjects were defined as those whose nocturnal decrease of systolic BP was

A74-year-old woman was admitted for evaluation of chest pain. She had fallen and hit her head 4 days before her admission and had lost consciousness. Her chest pain had begun immediately after this event. An ECG on admission revealed abnormal Q waves and ST-segment elevation (Figure 1A⇓). Her plasma creatine phosphokinase level was significantly elevated (386 IU/L), with an increase in MB isozyme level on admission. …

The effect of the renin inhibitor ES-1005 or captopril on the expression of the kidney renin gene was investigated in sodium-depleted marmosets. We measured the level of kidney renin messenger RNA (mRNA) after continuous administration of ES-1005 (48 mg/kg per day) or captopril (2 mg/kg per day) intraperitoneally, via an osmotic mini-pump, for one week. The level of kidney renin mRNA was measured by densitometric Northern blot analysis using an alpha-32P-labelled human renin cDNA fragment as the hybridization probe. Captopril treatment markedly increased plasma renin activity and the level of kidney renin mRNA by 4.7-fold and 6.3-fold, respectively. ES-1005 treatment completely inhibited plasma renin activity and significantly decreased the level of kidney renin mRNA (46% of the normal control P less than 0.01). However, plasma immunoreactive renin concentration was significantly increased by the treatment with ES-1005 (P less than 0.05). These results suggest that the treatment with the renin inhibitor ES-1005 for one week has a paradoxical effect on kidney renin gene expression and renin release from the kidney in sodium-depleted marmosets.

To the Editor: The first major septal branch of the left anterior descending coronary artery seems to be closely related to disorder of the conduction system. Blood supply to the anterosuperior fascicle of the left bundle branch originates exclusively from the septal branches.1 During myocardial ischemic attack due to stenosis of the proximal left anterior descending coronary artery, from the ostium of the left coronary artery to just before the first major septal branch, left-axis deviation often appears.2 3 4 Very recently, we reported5 that transient leftward QRS-axis shift during treadmill exercise testing or PTCA was a highly specific marker of proximal left anterior descending coronary artery disease. However, no one has confirmed that …

P115 Mechanical stresses to the arterial wall were evaluated in the carotid artery in 117 patients with risk factors for atherosclerosis including hypertension, dyslipidemia, and diabetes mellitus. Circumferential wall tension and shear stress were evaluated using Laplace’s law and Poiseuillean‘s parabolic model of flow velocity distribution. Arterial strain was also evaluated as the circumferential wall stretch. Wall tension was significantly related to BMI (r=0.40, p

Objective : This study was undertaken to clarify whether the myocardial dysfunction observed in the hypertensive heart is an intrinsic property of the myocyte or not. Materials and methods : We investigated left ventricular function and contractile function of myocytes from 30-week-old spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY). We also evaluated the effect of angiotensin II on contractile function of myocytes from both rats. Results : The time constant of isovolumic pressure fall was significantly greater in SHR (13.2 ± 0.6 ms) than in WKY (10.3 ± 0.5 ms). The extent of shortening in isolated myocytes was significantly higher in SHR (11.3 ± 0.4%) than in WKY (9.8 ± 0.4%, P

OBJECTIVES: To elucidate an association between left ventricular geometric adaptation to sustained hypertension and preclinical extracardiac target organ damage in essential hypertension. We also studied the clinical significance of neurohumoral factors for cardiovascular structural changes. DESIGN: One hundred and forty patients with essential hypertension were divided into four subgroups, based on left ventricular mass index and relative wall thickness. With respect to extracardiac target organ damage, we measured the funduscopic grade of retinal changes and serum creatinine levels. RESULTS: Among the hypertensive patients, only 19 (14%) had a typical concentric hypertrophy (increase in left ventricular mass index and relative wall thickness). Hypertensive patients with concentric hypertrophy had the most advanced funduscopic abnormalities and the greatest renal involvement, and hypertensive patients without left ventricular hypertrophy had the least extracardiac target organ damage. Plasma renin activity and plasma aldosterone concentration were higher in hypertensive patients with than in those without concentric hypertrophy. In a multiple regression model there was a strongly significant correlation between the degree of left ventricular mass index and the severity of hypertensive retinopathy and renal involvement, independent of office blood pressure. CONCLUSIONS: These results clearly demonstrate that echocardiographically determined left ventricular mass and geometry stratify extracardiac target organ damage in patients with essential hypertension more closely than office blood pressure. The present study also suggests that, in addition to blood pressure load, the renin-angiotensin-aldosterone system appears to play an important role in myocardial hypertrophy and peripheral vascular damage in hypertension.

A multiangle scan technique was used in combination with usual spin echo and cine MR techniques to study a patient with aortic arch dissection. The intimal flap was seen with each technique and the jet into the entry site was observed with cine MR technique. Exact short axis images of aortic arch were obtained only with multiangle scan. Retrograde dissection and involvement of left subclavian artery were detected clearly on the short axis images.

The role of glutathione in the central nervous system in regulating blood pressure (BP) and sympathetic nerve activity (SNA) was investigated in rats. Intracerebroventricular (ICV) injection of glutathione disulfide (GSSG: 1.7-33 nmol) resulted in a dose-dependent increase in BP [delta mean BP: 17 +/- 1 mm Hg (n = 7) for 33-nmol dose] together with a marked increase in SNA [163 +/- 13 to 672 +/- 70 spikes/10 s (n = 7), p less than 0.001]. Intracerebroventricular administration of its reduced form (GSH, 33 nmol) produced a vasodepressor response (delta mean BP: -9 +/- 2 mm Hg) accompanied by a corresponding decrease in SNA [192 +/- 15 to 54 +/- 22 spikes/10 s (n = 6), p less than 0.01]. These responses were not due to a leakage into the systemic circulation, since intravenous injection of GSSG or GSH (33 nmol) did not show any cardiovascular effects. Electrical stimulation of the posterior hypothalamus induced hypertension with a significant decrease of GSSG in the brain stem. The results indicate that GSSG has a stimulatory control over the sympathetic nervous system while GSH has an inhibitory effect on SNA. Glutathione disulfide and GSH may act within the central nervous system to modulate the tone of the sympathetic nervous system.

Dipeptide and tripeptide derivatives containing a statine residue were synthesized as human renin inhibitors. ES-305, bis[(1-naphthyl)methyl]acetyl-histidyl-statine-2(S)-methylbutylami de, was found to be a highly potent human renin inhibitor that is species-specific and enzyme-specific. The replacement of the methylbutylamide of ES-305 with the leucyl-lysinol (ES-1005) showed similar high potency against human renin (Ki value of 2.4 x 10(-9) M) and monkey renin (Ki value of 7.9 x 10(-9) M) as ES-305. ES-1005 competitively inhibited human renin. The compound was about one order of magnitude less potent against pig, dog, and rabbit renins. It had moderate inhibitory potencies against cathepsin D and pepsin (IC50 of cathepsin D and pepsin of 1.6 x 10(-5) and 8.0 x 10(-6) M, respectively). ES-1005, a newly synthesized tripeptide derivative containing statine, is a highly potent inhibitor of not only primate renin but also a wide variety of nonprimate renins.

Angiongensin converting enzyme (ACE) activities in 46 benign prostatic hypertrophy (BPH) tissues from 23 patients were compared with respect to histologic patterns, i.e., cystic, adenomatous and fibromuscular components. There was a good correlation between the enzyme activity and the percentage of cystic component in the tissue. On immunofluorescent study using antiserum against purified human kidney ACE, human prostatic ACE in tissues from BPH was mainly located in tubular epithelial cells and cystic lumens.

The effect of brain glutathione reductase activity on blood pressure regulation was investigated. The intravenous administration of the glutathione reductase inhibitor, nitrofurantoin (0.1-0.3 mg/rat), to seven normotensive Wistar rats caused dose-dependent rises in blood pressure and the heart rate (delta mean blood pressure 17 +/- 1 mmHg; delta heart rate 89 +/- 7 beats/min for 0.3 mg). Rats treated with 0.3 mg nitrofurantoin showed a 50% decrease in glutathione reductase activity with a twofold increase in the ratio of glutathione disulphide to reduced glutathione in the hypothalamus and brainstem, and a 1.5-fold increase in plasma noradrenaline and plasma renin activity compared with controls. These nitrofurantoin-induced effects were totally abolished by pretreatment with a sympathetic ganglion blocker (4 mg pentolinium tartrate, administered subcutaneously) except for the increased ratio of glutathione disulphide to reduced glutathione and the decreased glutathione reductase activity in the brain. These results suggest that the blood pressure elevation caused by inhibition of brain glutathione reductase activity occurs through activation of the sympathetic nervous system and the renin-angiotensin system.