Hisato Takatsu

Noninvasive quantitative tissue characterization and two-dimensional color-coded map of human atherosclerotic lesions using ultrasound Integrated backscatter comparison between histology and Integrated backscatter images

OBJECTIVES The purpose of the present study was to define clinicopathologically whether integrated backscatter (IB) combined with conventional two-dimensional echo (2DE) can differentiate the tissue characteristics of calcification (CL), fibrosis (FI), lipid pool (LP) with fibrous cap, intimal hyperplasia (IH) and thrombus (TH) and can construct two-dimensional tissue plaque structure in vivo. BACKGROUND It is difficult to characterize the components of plaque using conventional 2DE techniques. METHODS Integrated backscatter values of plaques were measured in the right common carotid and femoral arteries (total 24 segments) both during life and after autopsy in 12 patients (age 68 to 84 years, 10 men and two women). Integrated backscatter values were determined using a 5-12 MHz multifrequency transducer, setting the region of interests (ROIs) (11 x 11 pixels) on the echo tomography of the entire arterial wall (55 +/- 10 ROI/segment) and comparing it with histologic features in the autopsied arterial specimens. RESULTS Corrected IB values obtained before death and at autopsy were significantly correlated (r = 0.93, p < 0.01). Corresponding to the histologic features, corrected IB values on the rectangle ROIs obtained during life were divided into five categories: category 1 (TH) 4 < IB < or = 6; category 2 (media and IH or LP in the intima) 7 < IB < or = 13; category 3 (FI) 13 < IB < or = 18, category 4 (mixed lesion) 18 < IB < or = 27 and category 5 (CL) 28 < IB < or = 33. In category 2, media and intima were differentiated using conventional 2DE. Under the above procedures, color-coded maps constructed with IB-2DE obtained during life precisely reflected the histologic features of media and intima. CONCLUSIONS Integrated backscatter with 2DE represents a useful noninvasive tool for evaluating the tissue structure of human plaque.

Considerable Time From the Onset of Plaque Rupture and/or Thrombi Until the Onset of Acute Myocardial Infarction in Humans Coronary Angiographic Findings Within 1 Week Before the Onset of Infarction

BackgroundIt has been thought that the thrombi and bleeding in plaques that occur after plaque rupture or endothelial damage from vessels with mild stenosis suddenly occlude the lumen and cause acute myocardial infarction (AMI). However, our hypothesis is that thrombi and bleeding may not suddenly occlude the lumen. Methods and ResultsThe study group consisted of 20 patients who had coronary angiograms performed within 1 week (3±3 days) before AMI and 20 control patients who had coronary angiograms performed 6 to 18 months (282±49 days) before AMI. The features of infarct-related coronary segments (IRCS) at 3 days before AMI were the presence of a significant stenosis of >50% (95% in incidence and 71±12% diameter stenosis) and Ambrose’s type II eccentric lesions (plus multiple irregularities), an indicator of plaque rupture and/or thrombi (60% [70%]), and the features at 1 year before AMI were mild stenosis of <50% (95% incidence and 30±18% diameter stenosis) with rare Ambrose’s type II eccentric lesions (plus multiple irregularities) (10% [10%]). The same relation was observed in each of the 4 subgroups with Q-wave infarction, non–Q-wave infarction, preceding effort angina within 1 month before AMI, and no preceding effort angina. ConclusionsThe appearance of marked progression and Ambrose’s type II eccentric lesion on coronary angiograms 3 days before AMI suggests the presence of a considerable time from the onset of plaque rupture and/or thrombi until the onset of AMI. These features may be predictors of AMI. The concept provides new insight into the mechanism and prevention of human AMIs.

Process of progression of coronary artery lesions from mild or moderate stenosis to moderate or severe stenosis: A study based on four serial coronary arteriograms per year.

BACKGROUND The process of progression in coronary artery disease is unknown. METHODS AND RESULTS The subjects were 36 patients with 36 objective vessels with clinically significant progression of coronary artery disease (>/=15% per year) in whom 4 serial coronary arteriograms (CAGs) were performed at intervals of approximately 4 months in a 1-year period. The degree of progression of percent stenosis between each of 2 serial CAGs was classified as marked (M: >/=15%), slight (S: 5% to 14%), and no progression (N: <5%). From the pattern of progression, the 36 vessels were classified as 14 type 1 vessels with marked progression (N-->N-->M in 13 vessels and S-->S-->M in 1 vessel) and 22 type 2 vessels without marked progression (S-->S-->S in 18 vessels, N-->S-->S in 4). Percent stenosis at the first, second, third, and final CAGs was 44+/-14%, 46+/-13%, 46+/-13%, and 88+/-10% (P<0.05 versus first CAG) in type 1 vessels and 44+/-11%, 50+/-9%, 59+/-9%, and 67+/-9% in type 2 vessels (P<0.05 for second, third, and final CAGs versus first CAG). Type 1 vessels featured the sudden appearance of severe stenosis due to marked progression, angina pectoris, or myocardial infarction (71%) and Ambrose type II eccentric lesions indicating plaque rupture or thrombi (57%). Type 2 vessels featured continuous slight progression of stenosis with smooth vessel walls; angina pectoris (14%) occurred when the percent stenosis reached a severe level. An increase in serum C-reactive protein was observed only in the type 2 vessel group, which suggests a relation between continuous slight progression and inflammatory change. CONCLUSIONS Two types of stenosis progression provide a new insight into the mechanism of coronary artery disease.

Tumor Necrosis Factor-α (TNF-α) Plays a Protective Role in Acute Viral Myocarditis in Mice A Study Using Mice Lacking TNF-α

BACKGROUND It has been reported that tumor necrosis factor-alpha (TNF-alpha) is expressed in the heart with viral myocarditis and that its expression aggravates the condition. The pathophysiological effects of TNF-alpha on viral myocarditis, however, have not been fully elucidated. METHODS AND RESULTS To investigate the role of TNF-alpha in the progression of viral myocarditis, we used TNF-alpha gene-deficient mice (TNF-alpha(-/-)) and induced acute myocarditis by infection with encephalomyocarditis virus (EMCV). The survival rate of TNF-alpha(-/-) mice after EMCV infection was significantly lower than that of TNF-alpha(+/+) mice (0% versus 67% on day 14). Injection of recombinant human TNF-alpha (0.2 to 4.0 microg/mouse IV) improved the survival of TNF-alpha(-/-) mice in a dose-dependent manner, indicating that TNF-alpha is essential for protection against viral myocarditis. The levels of viral titer and viral genomic RNA of EMCV in the myocardium were significantly higher in TNF-alpha(-/-) than in TNF-alpha(+/+) mice. Histopathological examination showed that the inflammatory changes of the myocardium were less marked in TNF-alpha(-/-) than in TNF-alpha(+/+) mice. Immunohistochemical analysis revealed that the levels of immunoreactivity of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 in the myocardium were decreased in TNF-alpha(-/-) mice compared with TNF-alpha(+/+) mice. CONCLUSIONS These observations suggested that TNF-alpha is necessary for adhesion molecule expression and to recruit leukocytes to inflammatory sites, and thus, the lack of this cytokine resulted in failure of elimination of infectious agents. We concluded that TNF-alpha plays a protective role in the acute stage of viral myocarditis.Background—It has been reported that tumor necrosis factor-α (TNF-α) is expressed in the heart with viral myocarditis and that its expression aggravates the condition. The pathophysiological effects of TNF-α on viral myocarditis, however, have not been fully elucidated. Methods and Results—To investigate the role of TNF-α in the progression of viral myocarditis, we used TNF-α gene–deficient mice (TNF-α−/−) and induced acute myocarditis by infection with encephalomyocarditis virus (EMCV). The survival rate of TNF-α−/− mice after EMCV infection was significantly lower than that of TNF-α+/+ mice (0% versus 67% on day 14). Injection of recombinant human TNF-α (0.2 to 4.0 μg/mouse IV) improved the survival of TNF-α−/− mice in a dose-dependent manner, indicating that TNF-α is essential for protection against viral myocarditis. The levels of viral titer and viral genomic RNA of EMCV in the myocardium were significantly higher in TNF-α−/− than in TNF-α+/+ mice. Histopathological examination showed that the inflam...

Apoptosis and Overexpression of Bax Protein and bax mRNA in Smooth Muscle Cells Within Intimal Hyperplasia of Human Radial Arteries Analysis With Arteriovenous Fistulas Used for Hemodialysis

There is a type of arteriosclerosis with remodeling of middle-size arteries in which intimal hyperplasia of smooth muscle cells (SMCs) plays the main role, and there are few macrophages, T lymphocytes, and foam cells. It is unknown whether apoptosis and the expression of Bax, an inducer of apoptosis, are increased according to the progression of this type of human arteriosclerosis, which is different from so-called atherosclerosis. Bax heterodimerizes with Bcl-2, an inhibitor of apoptosis, and the ratio of Bax to Bcl-2 determines cellular apoptosis or survival. Thus, we investigated apoptosis and the expressions of Bax, bax mRNA, and Bcl-2 in human arteriovenous (AV) fistulas used for hemodialysis, a representative of arteriosclerosis of the aforementioned type. The material was 20 radial arteries obtained from 20 patients with chronic renal failure undergoing AV shunt surgery. SMCs, macrophages, and T lymphocytes were immunohistochemically identified at the light microscopic (LM) level. Apoptosis was detected by in situ terminal deoxynucleotidyl transferase (TdT)-mediated digoxigenin-dUTP nick end labeling (TUNEL) at both the LM and electron microscopic (EM) level. Cell proliferating activity was estimated by proliferating cell nuclear antigen (PCNA). Bax and Bcl-2 were detected by immunohistochemistry and Western blot analysis. Expression of bax mRNA was detected by in situ hybridization. LM TUNEL-positive cells in both the intima and media were significantly increased according to the percent stenosis of the vessels. EM analysis revealed that ultrastructures of apoptotic SMCs were seen in both synthetic and contractile phenotypes. Their frequency of occurrence in the intima and media were greater in those vessels with >50% stenosis than in those with <50% stenosis (5.2+/-0.7% versus 1.0+/-0.3% in the intima and 2. 1+/-0.5% versus 0.2+/-0.1% in the media). The proportion of apoptotic SMCs with ruptured plasma membranes was greater than that of apoptotic SMCs with intact membranes in the intima of the former (4.1+/-0.6% versus 1.1+/-0.1%). Only those SMCs with apoptotic ultrastructures had TUNEL-positive nuclei with moderate or marked accumulation of immunogold particles at the EM level. However, ultrastructures of oncosis (primary necrosis) were not observed. Immunohistochemical analyses showed significant positive correlations between percent stenosis of vessels and the percentage of either PCNA-positive intimal cells or Bax-positive areas in the intima and media. Bcl-2-positive cells were not observed in the intima but mainly in the outer media. The percentage of Bcl-2-positive medial cells was definitely decreased at an early stage after formation of the AV fistula but did not change according to the duration of hemodialysis or the progression of arteriosclerosis. Western blot analysis of Bax or Bcl-2 and in situ hybridization of bax mRNA confirmed the immunohistochemical data. Thus, regulation of cellularity in intimal hyperplasia of SMCs in human arteriosclerosis with remodeling is mediated by proliferation and apoptosis but not oncosis. The apoptosis is probably induced by an increase in the Bax to Bcl-2 ratio.

N-Methyl-1-Deoxynojirimycin (MOR-14), an α-Glucosidase Inhibitor, Markedly Reduced Infarct Size in Rabbit Hearts

BACKGROUND N-methyl-1-deoxynojirimycin (MOR-14), an alpha-glucosidase inhibitor, reduces the glycogenolytic rate by inhibiting the alpha-1,6-glucosidase of glycogen-debranching enzyme in the liver, in addition to possessing an antihyperglycemic action by blocking alpha-1,4-glucosidase in the intestine. Because the reduction of the glycogenolytic rate may be one of the mechanisms of myocardial protection in ischemic preconditioning, the compounds inhibiting myocardial alpha-1,6-glucosidase may be protective against ischemic damage. Thus, we investigated whether MOR-14 could inhibit alpha-1,6-glucosidase and reduce the infarct size in rabbit hearts without collateral circulation. METHODS AND RESULTS MOR-14 dose-dependently decreased the alpha-1,6-glucosidase activity in rabbit heart extract. A tracer study demonstrated the myocardial uptake of a considerable amount of MOR-14 sufficient to fully inhibit alpha-1,6-glucosidase. To assess the infarct size-reducing effect of MOR-14, 54 rabbits were subjected to 30-minute coronary occlusion followed by 48-hour reperfusion. Preischemic treatment with 25, 50, and 100 mg/kg of MOR-14 dose-dependently reduced the infarct size (to 26+/-4%, 19+/-3%, and 14+/-2% of the area at risk, respectively), compared with the saline control (45+/-5%) without altering the blood pressure or heart rate. Another 40 rabbits given 100 mg of MOR-14 or saline 10 minutes before ischemia were euthanized at 10 or 30 minutes of ischemia for biochemical analysis. MOR-14 decreased the alpha-1,6-glucosidase activity to approximately 20% in vivo, reduced the glycogen breakdown, and attenuated the lactate accumulation at both 10 and 30 minutes of ischemia. CONCLUSIONS Preischemic treatment with MOR-14 preserved glycogen, attenuated the accumulation of lactate, and reduced the myocardial infarct size by 69%. This cardioprotective effect was independent of changes of blood pressure and heart rate or regional blood flow. It may be associated with alpha-1,6-glucosidase inhibition, because MOR-14 markedly decreased the alpha-1,6-glucosidase activity in the heart.

Renoprotective Effect of the Addition of Losartan to Ongoing Treatment with an Angiotensin Converting Enzyme Inhibitor in Type-2 Diabetic Patients with Nephropathy

Angiotensin converting enzyme inhibitors (ACE-Is) and angiotensin II receptor blockers (ARBs) are frequently used for the treatment for glomerulonephritis and diabetic nephropathy because of their albuminuria- or proteinuria-reducing effects. To many patients who are nonresponsive to monotherapy with these agents, combination therapy appears to be a good treatment option. In the present study, we examined the effects of the addition of an ARB (losartan) followed by titration upon addition and at 3 and 6 months (n=14) and the addition of an ACE-I followed by titration upon addition and at 3 and 6 months (n=20) to the drug regimen treatment protocol in type 2 diabetic patients with nephropathy for whom more than 3-month administration of an ACE-I or the combination of an ACE-I plus a conventional antihypertensive was ineffective to achieve a blood pressure (BP) of 130/80 mmHg and to reduce urinary albumin to <30 mg/day. During the 12-month treatment, addition of losartan or addition of an ACE-I to the treatment protocol reduced systolic blood pressure (SBP) by 10% and 12%, diastolic blood pressure (DBP) by 7% and 4%, and urinary albumin excretion by 38% and 20% of the baseline value, respectively. However, the effects on both BP and urinary albumin were not significantly different between the two therapies. In conclusion, addition of losartan or an ACE-I to an ongoing treatment with an ACE-I, or addition of an ACE-I to ongoing treatment with a conventional antihypertensive were equally effective at reducing the urinary albumin excretion and BP, and provided renal protection in patients with type-2 diabetic nephropathy.

Sympathetic tone assessed by washout of iodine 125-labeled metaiodobenzylguanidine from the murine left ventricle-influence of immobilization stress and inhibition of the renin-angiotensin system

BackgroundBecause it is not metabolized as is norepinephrine (NE), most of the metaiodobenzylguanidine (MIBG) taken up by the heart is considered to be lost subsequently by release concomitant with sympathetic stimulation. Therefore we examined whether the washout of MIBG is influenced by sympathetic tone, which we modulated by using immobilization stress or activation of the renin-angiotensin system (RAS).Methods and ResultsIn 175 male ddY mice, left ventricular radioactivity was counted 30 minutes or 4 hours after injection of 74 kBq (2 μCi) of iodine 125- or iodine 131-labeled MIBG (125I- or 131I-MIBG). The washout rates of MIBG were determined under immobilization stress or under sodium loading or restriction in combination with losartan (10 mg/kg) or cilazapril (1 mg/kg) pretreatment. Immobilization enhanced the washout of 125I-MIBG (80.9% vs 57.9% in the control animals); this was determined to be related to washout from the neuronal compartment, because the nonneuronal component assessed through desipramine pretreatment was not affected. Pretreatment with losartan or cilazapril decreased the facilitation of 125I-MIBG washout in sodium-restricted mice (40.9% and 33.7%, respectively, vs 43.5% in the control animals), but not in sodium-loaded mice.ConclusionsMeasurement of MIBG washout may be feasible for determining the changes in sympathetic tone caused by immobilization stress or activation of the RAS.

Usefulness of resting thallium-201 delayed imaging for detecting myocardial viability in patients with previous myocardial infarction

To test the feasibility of resting thallium-201 (201Tl) initial and delayed scintigraphy for detecting the area of viable myocardium, we performed single photon emission computed tomography (SPECT) in 57 patients with previous myocardial infarction (MI). All had received coronary arteriography (CAG) and left ventriculography (LVG). Initial and delayed myocardial imagings were carried out 10 min and 2 hours, respectively, after the injection of201Tl at rest. Redistribution was judged by visual interpretation and/or the circumferential profile curve, and found in the infarcted or its adjacent area in 40 of the 57 cases (70.2%). A negative washout (net increase of201Tl uptake in delayed image) was detected in 17 of these 40 cases. In 10 of the 57 patients, both exercise and rest-injected201Tl myocardial images were obtained at exercise and rest, and compared visually. The areas of abnormal perfusion were smaller in the resting delayed images than those seen after exercise in 9 of the 10 cases, and were equal in one case. Thus, resting201Tl delayed myocardial scintigraphy appears to reduce the underestimation of the size of the viable myocardium by the usual201Tl images obtained after exercise or by single initial images obtained at rest in patients with previous MI.

Washout of I-123 meta-iodobenzylguanidine for assessing cardiac sympathetic activity with progression of hypertension in Dahl salt-sensitive rats.

BackgroundTo evaluate cardiac sympathetic activity, a simple method should be developed to replace such complex methods as the spillover rate of tritiated norepinephrine (3H-norepinephrine) or microneurography of sympathetic nerve activity. The goal of this study is to evaluate cardiac sympathetic activities by analyzing the washout of I-123 meta-iodobenzylguanidine (123I-MIBG), a radiolabeled norepinephrine analogue, in Dahl salt-sensitive (DS) rats as it relates to the progression of hypertension.Methods and ResultsDahl salt-resistant (DR) rats and DS rats were fed an 8% salt diet starting at age 5 weeks. Marked hypertension and cardiac hypertrophy developed in the DS rats, whereas DR rats remained normotensive. Then the time-activity curves of 123I-MIBG from 15 to 200 minutes were obtained from both DS and DR strains at ages 8, 11, and 13 weeks using dynamic scintigraphic analysis. We also examined the nonneuronal washout of 123I-MIBG using dynamic scintigraphic studies in desipramine pretreated normal rats. In the preliminary study with desipramine pretreatment, the majority of the nonneuronal 123I-MIBG washout occurred by 90 minutes after injection. Therefore the late-phase washout in the control rats was found to reflect the neuronal washout. We then applied exponential curve fitting to the time activity curves acquired in the 90- to 200-minute period after 123I-MIBG injection in both DR and DS rats. When we compared the coefficients of these washout curves in the DS and DR rats as an index of cardiac sympathetic activities, the coefficient values remained high during all stages in DS rats, whereas they decreased with age in DR rats.ConclusionMeasurement of late-phase 123I-MIBG washout may be a useful tool for assessing the change in sympathetic activity in the progression of hypertension without the influence of extraneuronal washout of 123I-MIBG and left ventricular hypertrophy.