Hironosuke Sakamoto

Regulated expression of the BTEB2 transcription factor in vascular smooth muscle cells: analysis of developmental and pathological expression profiles shows implications as a predictive factor for restenosis.

BackgroundWe have previously shown BTEB2, a Krüppel-like zinc finger transcription factor, to regulate expression of the SMemb/NMHC-B gene, which has been implicated in phenotypic modulation of smooth muscle cells (SMCs). The present study was done to assess the developmental and pathological expression profiles of BTEB2 and to further evaluate the clinical relevance of BTEB2 expression in human coronary artery disease. Methods and ResultsImmunohistochemistry showed developmentally regulated expression of BTEB2 with abundant expression in fetal but not in adult aortic SMCs of humans and rabbits. In balloon-injured aortas, predominant expression of BTEB2 was seen in neointimal SMCs. Atherectomy specimens obtained from primary and restenotic lesions showed predominant expression of BTEB2 to stellate SMCs. The incidence of restenosis in primary lesions was significantly higher in lesions containing BTEB2-positive cells than in lesions without (55.6% versus 25.0%, P =0.01). ConclusionsThe present study shows that BTEB2 expression is developmentally and pathologically regulated. BTEB2 is preferentially expressed in dedifferentiated or activated SMCs. Examination of human coronary artery specimens suggests that primary lesions containing BTEB2-positive cells are associated with higher risk of restenosis than BTEB2-negative lesions. These results suggest that BTEB2 can serve as a molecular marker for phenotypic modulation of vascular SMCs.

Exaggerated vascular response due to endothelial dysfunction and role of the renin-angiotensin system at early stage of renal hypertension in rats.

We investigated whether endothelial dysfunction might contribute to the exaggerated vasoconstriction that was induced by the administration of norepinephrine at the early stage of one-kidney, one-clip renal hypertension (1K1C) in rats. We also studied the role of the renin-angiotension system in this phenomenon. Male Wistar rats were killed 48 hours after the induction of renal artery stenosis or sham operation, and ring preparations of the thoracic aorta were obtained. The isometric contraction and relaxation of aortic strips produced by norepinephrine and acetylcholine, respectively, were recorded with a force-displacement transducer. The aorta of 1K1C rats showed a significantly (P < .05) exaggerated contractile response to norepinephrine as compared with that of control rats. Rubbing the endothelium and treatment with methylene blue or NG-monomethyl L-arginine acetate augmented the contractile responses to norepinephrine to a greater extent in control rats than in 1K1C rats; therefore, the responses of the groups did not differ significantly. In the second experiment, rats received 0.05% captopril, 0.02% enalapril, or 0.02% nicardipine in the drinking water for 1 day before and for 48 hours after the induction of renal artery stenosis or sham operation. The increased contractile responses of the aorta to norepinephrine in 1K1C rats were normalized to the level of the control rats by treatment with either captopril or enalapril but not with nicardipine. These results suggest that the endothelial dysfunction may contribute to the exaggerated norepinephrine-induced vasoconstriction observed in the 1K1C rats and that angiotensin I-converting enzyme inhibitors can restore the endothelial function.

Antihypertensive agents prevent nephrosclerosis and left ventricular hypertrophy induced in rats by prolonged inhibition of nitric oxide synthesis.

We investigated the ability of the angiotensin converting enzyme (ACE) inhibitor imidapril hydrochloride, and of the calcium channel blocker amlodipine besilate, to prevent nephrosclerosis and left ventricular hypertrophy (LVH) in rats with hypertension induced by chronic inhibition of nitric oxide (NO). Male Wistar rats were given distilled water (control), NG-nitro-L-arginine methyl ester (L-NAME) 500 mg/L, L-NAME plus imidapril 10 mg/L or 100 mg/L, or L-NAME plus amlodipine 50 mg/L or 100 mg/L in the drinking water (n = 10-12). We then collected 24-h urine samples at 2, 4, and 6 weeks, obtained blood samples at 6 weeks, and histologically examined the kidney and heart. L-NAME markedly reduced the levels of NO metabolites in serum and urine while increasing the tail-cuff blood pressure, the urinary albumin level (1.90 +/- 0.65 v 0.05 +/- 0.02 mg/day/100 g in control), and the area of the left ventricular wall (83.3 +/- 3.0 v 69.8 +/- 1.8 mm2 in control). Nephrosclerosis and myocardial interstitial fibrosis were documented histologically. The plasma renin activity was significantly higher in rats treated with L-NAME than in the control rats. The concomitant administration of imidapril (10 mg/L) with L-NAME completely normalized the tail-cuff pressure, the LVH (70.8 +/- 1.8 mm2), the albuminuria (0.05 +/- 0.01 mg/day/100 g), and the histologic changes. Amlodipine (50 mg/L) also ameliorated the L-NAME-induced effects, but to a lesser extent. Thus, the chronic inhibition of NO synthesis in rats produced nephrosclerosis and LVH that were effectively prevented by giving imidapril at a dose lower than that of amlodipine. We conclude that ACE inhibitors can prevent nephrosclerosis and LVH even in the presence of a reduction in NO production, implying that in rats the inhibition of the renin-angiotensin system is more effective than the blockade of calcium channels in preventing hypertensive tissue injury.

c-Src and Hydrogen Peroxide Mediate Transforming Growth Factor-β1–Induced Smooth Muscle Cell–Gene Expression in 10T1/2 Cells

Objective— Transforming growth factor-β1 (TGF-β1) controls the expression of numerous genes, including smooth muscle cell (SMC)–specific genes and extracellular matrix protein genes. Here we investigated whether c-Src plays a role in TGF-β1 signaling in mouse embryonic fibroblast C3H10T1/2 cells. Methods and Results— TGF-β1 induction of the SMC contractile protein SM22α gene expression was inhibited by PP1 (an inhibitor of Src family kinases) or by C-terminal Src kinase (a negative regulator of c-Src). Induction of SM22α by TGF-β1 was markedly attenuated in SYF cells (c-Src−, Yes−, and Fyn−) compared with Src++ cells (c-Src++, Yes−, and Fyn−). PP1 also inhibited the TGF-β1–induced expression of serum response factor (SRF), a transcription factor regulating the SMC marker gene expression. Confocal immunofluorescence analysis showed that TGF-β1 stimulates production of hydrogen peroxide. Antioxidants such as catalase or NAD(P)H oxidase inhibitors such as apocynin inhibited the TGF-β1–induced expression of SM22α. Furthermore, we demonstrate that TGF-β1 induction of the plasminogen activator inhibitor-1 (PAI-1) gene, which is known to be dependent on Smad but not on SRF, is inhibited by PP1 and apocynin. Conclusion— Our results suggest that TGF-β1 activates c-Src and generates hydrogen peroxide through NAD(P)H oxidase, and these signaling pathways lead to the activation of specific sets of genes, including SM22α and PAI-1.

Myocardial Infarction in Myxoma Patients with Normal Coronary Arteries Case Reports

Three cases of patients with cardiac myxoma who had attacks of acute myocardial infarc tion are presented. Cineangiographic study showed normal coronary arteries. Immunohistochemical and serologic examination revealed that both interleukin-6 and interleukin-8 were secreted in cardiac myxoma. The authors discuss the relation between these cytokines and myocardial infarction with normal coronary arteries.

Visceral fat obesity contributes to the tortuosity of the thoracic aorta on chest radiograph in poststroke Japanese patients.

Tortuosity of the thoracic aorta on chest radiographs is characteristic of atherosclerotic disease. Aging and hypertension are associated with the tortuosity, but little is known about the influence of other atherosclerotic risk factors on this abnormality. The purpose of this study was to examine which atherosclerotic risk factors are determinants for tortuosity of the thoracic aorta. Forty-five poststroke Japanese patients (31 men and 14 women, age range 41-78 years and mean 60.5 ±8.6) were studied. The distance factor, ie, the ratio of meandering vessel length to the straight-line distance between its end points, was used to measure arterial tortuosity. The hospital records were reviewed for clinical and biochemical variables. Tortuosity of the thoracic aorta had a significant positive relationship with body mass index (BMI) (r =0.397, p<0.01), waist circumference (r =0.360, p<0.05), and the cardiothoracic ratio (CTR) (r =0.526, p<0.001), and a significant negative relationship with ankle-brachial pressure index (ABPI) (r =-0.360, p<0.05). Stepwise regression analysis showed that waist circumference and CTR were independently correlated with increased tortuosity, whereas ABPI was negatively correlated with it. These results suggest that visceral fat obesity is a novel contributor to tortuosity of the thoracic aorta, which may be as a shortening of the distance between aortic tethering points due to elevation of the diaphragm by excessive intraabdominal fat and as a consequence of aortic elongation due to arteriosclerosis caused by obesity-related metabolic disorders.

Effects of Conjugated Equine Estrogenand Medroxyprogesterone Acetate on Lipoprotein(a) and Other Lipoproteins in Japanese Postmenopausal Women with and without Dyslipidemia

Background/Aim: The cardiovascular effects of postmenopausal hormone replacement are controversially discussed. We investigated the effects of 12 months of treatment with conjugated equine estrogen and medroxyprogesterone acetate on lipoprotein(a) [Lp(a)] and other lipoproteins in Japanese postmenopausal women (PMW) with and without dyslipidemia. Methods: Forty-three normolipidemic and 17 dyslipidemic PMW [total cholesterol (TC) ≧220 mg/dl or triglyceride (TG) ≧150 mg/dl] received conjugated equine estrogen (0.625 mg) plus medroxyprogesterone acetate (2.5 mg) daily for 12 months, and the results were compared with those of 26 normolipidemic and 14 dyslipidemic subjects declining this treatment as controls. The fasting serum levels of Lp(a), TC, TG, high-density lipoprotein cholesterol, low- density lipoprotein cholesterol, apolipoprotein (Apo) AI, Apo AII, Apo B, Apo CII, and Apo E were measured in each subject at baseline and 12 months after this treatment initiation. Results: The treatment decreased Lp(a) similarly in normolipidemic and dyslipidemic PMW and decreased TC, low-density lipoprotein cholesterol, Apo CII, and Apo E and increased high-density lipoprotein cholesterol, Apo AI, and Apo AII in both groups. The therapy also significantly increased TG in normolipidemic but not dyslipidemic subjects. In controls, the levels of Lp(a) and other lipoproteins were unaltered. Conclusions: In PMW with or without dyslipidemia, improvement in Lp(a) and other lipoproteins may represent cardiovascular benefits of hormone replacement therapy. However, an elevation of the TG levels seen with the therapy warrants caution, especially in PMW without dyslipidemia.

Smooth Muscle Cell Outgrowth from Coronary Atherectomy Specimens in vitro Is Associated with Less Time to Restenosis and Expression of a Key Transcription Factor KLF5/BTEB2

Atherectomy specimens offer an opportunity to study the biology of coronary artery lesions. We cultured smooth muscle cells (SMCs) from specimens obtained from 24 patients with coronary restenosis after angioplasty to study the relationship between activity of SMCs (in vitro outgrowth) and the time course of restenosis. We also examined expression of a Kruppel-like zinc-finger transcription factor 5 (KLF; also known as BTEB2 and IKLF), which is markedly induced in activated SMCs, in the same specimens. SMC outgrowth was observed in 9 of 24 specimens (37.5%). Restenosis occurred sooner (p < 0.01) in patients whose specimens showed outgrowth compared to those whose specimens showed no outgrowth. Immunostaining for KLF5 was more common in specimens with outgrowth (89 vs. 20%, p < 0.01). These data suggest that the number of activated SMCs in lesions may determine in vitro outgrowth and also affect the time to restenosis.

Dietary Zn deficiency does not influence systemic blood pressure and vascular nitric oxide signaling in normotensive rats.

Because zinc (Zn) is an important component for cell protection against certain oxygen species, it has been suggested that Zn deficiency impairs the potent oxidant defense capacity, which is constitutively provided in the vascular system. However, the influence of dietary Zn deficiency on systemic blood pressure and vascular system is controversial and unclear. We therefore examine the effect of dietary Zn deficiency on systemic blood pressure, a potent superoxide scavenger, aortic Cu/Zn superoxide dismutase (SOD) activity, a most representative synthase of the endothelium-derived relaxing factor, and aortic endothelial nitric oxide synthase (eNOS) expression. Furthermore, the direct effects of intravenous administration of NOS inhibitor, Nω-nitro-l-arginine methyl ester (l-NAME), and a SOD mimetic compound, tempol, in normotensives were tested in Wistar-Kyoto (WKY) rats. A Zn-deficient diet (4 wk) contributed to growth retardation, the decrease in thymus weight, and the lower levels of serum Zn compared with the standard diet group. However, no significant difference in conscious systolic and diastolic blood pressure was found in the Zn-deficiency group. The administration of l-NAME caused an increase in the mean arterial pressure (MAP) levels in the two groups of rats and the involvement of the vasodilator nitric oxide (NO) in the regulation of systemic BP in the normotensive state. On the other hand, administration of the superoxide scavenger, tempol, led to a decrease in MAP levels in the two groups of rats, indicating the participation of the oxygen free radical, superoxide, in the maintenance of the systemic BP in a normotensive state. There were no significant differences between the Zn-deficient diet group and the standard diet group in the normotensive state. eNOS expression and Cu/Zn SOD activity in the aorta were also intact in Zn-deficient normotensive rats. These findings suggest that the 4 wk of Zn deficiency was inadequate to alter systemic blood pressure and focal NO signaling in the normotensive state. Long-term Zn deficiency affects the neuronal, immune, and hematopoietic systems, which contribute to systemic and/or local circulation. However, Zn deficiency alone does not cause hypertension and local vascular dysfunction in the normotensive state.

Prediction of Transition to Chronic Atrial Fibrillation in Patients With Paroxysmal Atrial Fibrillation

To the Editor: We read with great interest the recent report of Abe et al1 demonstrating that P-wave–triggered signal-averaged electrocardiography may be useful to predict the transition to chronic atrial fibrillation in patients with paroxysmal atrial fibrillation. In their study, although sex, age, and presence of organic heart disease were not associated with an increased risk for the transition to chronic atrial fibrillation, patients with the abnormality of a P-wave–triggered signal-averaged ECG had an 11-fold greater risk for chronic atrial fibrillation than those without the abnormality. Echocardiographically, left atrial dimension in patients with chronic atrial fibrillation was larger than that in patients with paroxysmal atrial fibrillation (40.5 versus 36.4 mm, P <0.05), but there were no significant differences in left ventricular dimensions measured at end diastole and end systole or in ejection fraction between the 2 groups. In contrast to the findings by Abe et al, we2 previously reported that congestive heart failure and reduction in left ventricular ejection fraction were predictors of the transition to chronic atrial fibrillation in patients with new-onset atrial fibrillation. To identify predictors of the transition to chronic atrial fibrillation within the first year after onset, we retrospectively reviewed clinical records, standard 12-lead ECGs, and M-mode echocardiograms of 137 patients with new-onset, nonrheumatic atrial fibrillation. One year after onset, 30 (22%) of 137 patients …