Yosuke Murase

Mineralocorticoid Receptor Antagonism Ameliorates Left Ventricular Diastolic Dysfunction and Myocardial Fibrosis in Mildly Symptomatic Patients With Idiopathic Dilated Cardiomyopathy: A Pilot Study

Background— Mineralocorticoid receptor antagonism reduces mortality associated with heart failure by mechanisms that remain unclear. The effects of the mineralocorticoid receptor antagonist spironolactone on left ventricular (LV) function and chamber stiffness associated with myocardial fibrosis were investigated in mildly symptomatic patients with idiopathic dilated cardiomyopathy (DCM). Methods and Results— Twenty-five DCM patients with a New York Heart Association functional class of I or II were examined before and after treatment with spironolactone for 12 months. LV pressures and volumes were measured simultaneously, and LV endomyocardial biopsy specimens were obtained. Serum concentrations of the carboxyl-terminal propeptide (PIP) and carboxyl-terminal telopeptide (CITP) of collagen type I were measured. The patients were divided into 2 groups on the basis of the serum PIP/CITP ratio (≤35, group A, n=12; >35, group B, n=13), an index of myocardial collagen accumulation. LV diastolic chamber stiffness, the collagen volume fraction, and abundance of collagen type I and III mRNAs in biopsy tissue were greater and the LV early diastolic strain rate (tissue Doppler echocardiography) was smaller in group B than in group A at baseline. These differences and the difference in PIP/CITP were greatly reduced after treatment of patients in group B with spironolactone, with treatment having no effect on these parameters in group A. The collagen volume fraction was significantly correlated with PIP/CITP, LV early diastolic strain rate, and LV diastolic chamber stiffness for all patients before and after treatment with spironolactone. Conclusions— Spironolactone ameliorated LV diastolic dysfunction and reduced chamber stiffness in association with regression of myocardial fibrosis in mildly symptomatic patients with DCM. These effects appeared limited, however, to patients with increased myocardial collagen accumulation.

Association of gene polymorphisms with coronary artery disease in low- or high-risk subjects defined by conventional risk factors

OBJECTIVES The aim of the study was to identify genes that confer susceptibility to coronary artery disease (CAD) in low- or high-risk men or women separately and thereby to assess the genetic risk of CAD in such individuals. BACKGROUND The prevention of CAD would be facilitated by the identification of genes that confer susceptibility to this condition independently in low- or high-risk individuals, as defined by conventional risk factors. METHODS The study population comprised 1661 unrelated Japanese individuals, including 1011 patients with CAD and 650 control subjects. Among all study subjects, 601 individuals (high-risk subjects) had hypertension, diabetes mellitus, and hypercholesterolemia, and 1060 individuals (low-risk subjects) had none of these risk factors for CAD. The genotypes for 37 polymorphisms of 31 candidate genes were determined by a fluorescence- or colorimetry-based allele-specific DNA primer-probe assay system. RESULTS Multivariate logistic regression analysis, with adjustment for age, body mass index, and the prevalence of smoking and hyperuricemia, revealed that the -219G-->T polymorphism of the apolipoprotein E gene in low-risk men, the -1171/5A-->6A polymorphism of the stromelysin-1 gene in low-risk women, the 1019C-->T polymorphism of the connexin 37 gene in high-risk men, and the 3932T-->C polymorphism of the apolipoprotein E gene in high-risk women were significantly associated with CAD. A stepwise forward selection procedure revealed that the effects of these polymorphisms on CAD were statistically independent of age or conventional risk factors. CONCLUSIONS Genotyping of these polymorphisms may prove informative for assessment of the genetic risk of CAD in low- or high-risk men or women.

Lack of association of polymorphisms of the lymphotoxin α gene with myocardial infarction in Japanese

Vascular inflammation plays an important role in the development of myocardial infarction (MI). Lymphotoxin α (LTA) is a cytokine with multiple functions in regulation of the immune system and inflammatory reactions. The aim of this study was to examine whether polymorphisms of the LTA gene are associated with the risk of MI in Japanese men and women. A case-control association study was performed for the 252A→G and 804C→A polymorphisms of the LTA gene and the prevalence of MI. The study population comprised 3,689 unrelated Japanese individuals (2,486 men, 1,203 women), including 1891 patients with MI (1,493 men, 398 women) and 1798 control subjects (993 men, 805 women). Among the control subjects 257 individuals (108 men, 149 women) who had none of the conventional risk factors for coronary artery disease (CAD) were defined as low-risk controls. Genotypes for the two polymorphisms were determined with a fluorescence-based allele-specific DNA primer assay system. Among all study subjects the 252A→G and 804C→A polymorphisms exhibited linkage disequilibrium. No association of either polymorphism with MI was detected in men or in women in comparisons with total control or low-risk control subjects. However, each of the two polymorphisms was associated with the prevalence of type 2 diabetes mellitus both in men with MI and in those without MI in a recessive genetic model. No association was detected between the polymorphisms and other conventional risk factors for CAD. The LTA gene thus does not appear to be a susceptibility locus for MI in Japanese men or women, although it might affect susceptibility to type 2 diabetes in Japanese men.

Effects of Extracellular Matrix on Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells into Smooth Muscle Cell Lineage: Utility for Cardiovascular Tissue Engineering

Background: Bone marrow-derived mesenchymal stem cells (MSCs) can differentiate into various types of cell, and the extracellular matrix (ECM) is acknowledged to be important for the regulation of cell functions. In this study, we demonstrated the effects of ECMs on the differentiation of human bone marrow-derived MSCs into a smooth muscle cell (SMC) lineage. Methods: Human MSCs (hMSCs) were cultured on dishes coated with 3 types of ECM including laminin (LM), collagen type IV (Col-IV) and fibronectin for 7 days, and simultaneously cultured on a noncoated dish as a control. Cell numbers of these cultured hMSCs were counted, and their expression of SMC-specific genes and proteins was evaluated. hMSCs were then seeded on LM-coated biodegradable sheets and implanted into rat subcutaneous space. After 2 weeks of implantation, these tissues were evaluated. Results: The number of hMSCs was significantly increased by culture on Col-IV-coated dishes. The expression of SMC-specific genes and proteins (α-smooth muscle actin, ASMA; h1-calponin, CALP) in hMSC was significantly upregulated from culture on LM-coated dishes. LM-coated sheets showed a significantly increased expression of ASMA and CALP protein in vivo. Moreover, a fully differentiated marker (SM2) was expressed in the in vivo implanted hMSCs in the course of 2 weeks on the LM-coated sheet. Conclusion: These results suggest that the signal transduction of the cell-matrix interaction for the differentiation of hMSCs into SMCs was activated when cultured with LM. LM-coated materials may thus be useful for cardiovascular tissue engineering.

Decellularized ureter for tissue-engineered small-caliber vascular graft

Previous attempts to create small-caliber vascular prostheses have been limited. The aim of this study was to generate tissue-engineered small-diameter vascular grafts using decellularized ureters (DUs). Canine ureters were decellularized using one of four different chemical agents [Triton-X 100 (Tx), deoxycholate (DCA), trypsin, or sodium dodecyl sulfate (SDS)] and the histology, residual DNA contents, and immunogenicity of the resulting DUs were compared. The mechanical properties of the DUs were evaluated in terms of water permeability, burst strength, tensile strength, and compliance. Cultured canine endothelial cells (ECs) and myofibroblasts were seeded onto DUs and evaluated histologically. Canine carotid arteries were replaced with the EC-seeded DUs (n = 4). As controls, nonseeded DUs (n = 5) and PTFE prostheses (n = 4) were also used to replace carotid arteries. The degree of decelularization and the maintenance of the matrix were best in the Tx-treated DUs. Tx-treated and DCA-treated DUs had lower remnant DNA contents and immunogenicity than the others. The burst strength of the DUs was more than 500 mmHg and the maximum tensile strength of the DUs was not different to that of native ureters. DU compliance was similar to that of native carotid artery. The cell seeding test resulted in monolayered ECs and multilayered α-smooth muscle actin-positive cells on the DUs. The animal implantation model showed that the EC-seeded DUs were patent for at least 6 months after the operation, whereas the nonseeded DUs and PTFE grafts become occluded within a week. These results suggest that tissue-engineered DUs may be a potential alternative conduit for bypass surgery.

Dobutamine stress testing as a diagnostic tool for evaluation of myocardial contractile reserve in asymptomatic or mildly symptomatic patients with dilated cardiomyopathy.

OBJECTIVES We performed dobutamine stress testing for evaluation of myocardial contractile reserve in asymptomatic or mildly symptomatic patients with dilated cardiomyopathy (DCM). BACKGROUND Catecholamine sensitivity is reduced in failing hearts as a result of myocardial abnormalities in the beta-adrenergic receptor signaling pathway. However, little is known about adrenergic myocardial contractile reserve in asymptomatic or mildly symptomatic patients with DCM. METHODS The maximal first derivative of left ventricular pressure (LV dP/dt(max)) was determined during infusion of dobutamine (10 microg kg(-1) min(-1)) in 46 asymptomatic or mildly symptomatic (New York Heart Association functional class I or II) patients with DCM. The expression of messenger ribonucleic acid (mRNA) for contractile regulatory proteins in endomyocardial biopsy specimens was quantified by reverse transcription and real-time polymerase chain reaction analysis. Plasma norepinephrine levels were measured in all patients and [(123)I]metaiodobenzylguanidine (MIBG) scintigraphy performed. RESULTS Patients were classified into 3 groups based on the percentage increase in LV dP/dt(max) induced by dobutamine (DeltaLV dP/dt(max)) and on LV ejection fraction (LVEF) at baseline: group I (n = 18): DeltaLV dP/dt(max) >100% and LVEF >25%; group IIa (n = 17): DeltaLV dP/dt(max) <or=100% and LVEF > 25%; and group IIb (n = 11): DeltaLV dP/dt(max) <or=100% and LVEF <or=25%. The amounts of beta(1)-adrenergic receptor, sarcoplasmic reticulum Ca(2+)-adenosine triphosphatase, and phospholamban mRNA were significantly smaller in groups IIa and IIb than in group I. The plasma norepinephrine level was increased and the delayed heart/mediastinum count ratio in MIBG scintigraphy was decreased in both groups IIa and IIb. CONCLUSIONS Dobutamine stress testing is a useful diagnostic tool for identifying reduced adrenergic myocardial contractile reserve related to altered myocardial expression of beta(1)-adrenergic receptor, sarcoplasmic reticulum Ca(2+)-adenosine triphosphatase, and phospholamban genes even in asymptomatic or mildly symptomatic patients with DCM.

Evaluation of compliance and stiffness of decellularized tissues as scaffolds for tissue-engineered small caliber vascular grafts using intravascular ultrasound.

This study evaluated the compliance and stiffness of decellularized canine common carotid artery as well as decellularized canine ureter and compared it with that of polytetrafluoroethylene, elastin gel combined with polylactic acid tube, and canine common carotid artery. To calculate the compliance and stiffness, internal diameters and cross-sectional areas were measured according to changes in the intraluminal pressures using intravascular ultrasound in a closed circuit system equipped with a syringe pump. The pressure-area curve, stiffness parameter β, and diameter compliance were evaluated. Canine common carotid artery and decellularized canine common carotid artery, as well as decellularized canine ureter, showed a compliant response, a J-shaped curve. However, the latter evidenced different characteristics in the low pressure range. Although the cross-sectional area of the elastin gel combined with polylactic acid tube showed some changes, it did not present a J-shape curve. Polytetrafluoroethylene exhibited a noncompliant response. The results in this study have shown that the compliance in the decellularized matrices was maintained after cell extraction, which demonstrated the importance of the remaining matrix structure in the mechanical properties of decellularized tissue. A clear difference between the decellularized matrices and synthetic materials was noted in terms of the compliance, even in materials composed of relatively elastic materials.

Adiponectin acts as a positive indicator of left ventricular diastolic dysfunction in patients with hypertrophic cardiomyopathy.

Background Adiponectin is an adipose-derived plasma protein that exhibits beneficial actions on the heart. Recently, it was shown that adiponectin levels were elevated in patients with systolic heart failure. Objective To investigate the association between adiponectin levels and left ventricular (LV) diastolic function in patients with hypertrophic cardiomyopathy (HCM), characterised by diastolic dysfunction. Methods Twenty-six patients with HCM showing LV ejection fraction of >60% were enrolled. LV pressure half-time (T1/2) was measured as an index of myocardial relaxation. Patients were divided into two groups on the basis of baseline T1/2 (group A: T1/2< 35 ms, group B: T1/2≥ 35 ms). Blood samples were simultaneously collected from the coronary sinus (CS) and aortic root (Ao) as well as the peripheral vein (PV) for measurement of plasma adiponectin levels. Results Plasma adiponectin levels were significantly higher in group B than in group A. Adiponectin levels in the PV were positively correlated with the baseline T1/2 in patients with HCM. The transcardiac gradient of adiponectin as calculated by the Ao−CS difference was significantly higher in group A than in group B. The transcardiac gradient of adiponectin also inversely correlated with the baseline T1/2 and adiponectin levels in PV in patients with HCM. The expression of AdipoR1 but not AdipoR2 in the heart decreased in group B. The baseline T1/2 was negatively associated with AdipoR1 expression in patients with HCM. Conclusions These data document that adiponectin is an indicator of LV diastolic dysfunction in patients with HCM.

Relation of a common variant of the adiponectin gene to serum adiponectin concentration and metabolic traits in an aged Japanese population

Adiponectin is an adipocyte-derived protein that is down-regulated in obesity-linked disorders. Variants of the adiponectin gene (ADIPOQ) have been shown to affect adiponectin level. We have now examined the relation of polymorphisms of ADIPOQ to adiponectin concentration and to metabolic disorders in the Kita-Nagoya Genomic Epidemiology study, a population-based study of elderly Japanese. The genomic region including ADIPOQ was genotyped for 30 single nucleotide polymorphisms in 500 subjects of a screening population with the use of a fluorescence- or colorimetry-based allele-specific DNA primer–probe assay system. Four polymorphisms were then selected for genotyping in an additional 2797 subjects. Serum adiponectin level was negatively associated with metabolic abnormalities after adjustment for age and sex. The minor alleles of the rs1656930, Ile164Thr, and rs9882205 polymorphisms were associated with a low serum adiponectin level. Whereas the minor alleles of rs1656930 and rs9882205 were common (minor allele frequency of 6.2 and 38.5%, respectively), that of Ile164Thr was rare (0.9%). The minor allele of rs1656930 was positively associated with systolic blood pressure and the prevalence of hypertension. The association of rs1656930 with adiponectin level was replicated in an independent population. A subject with the 164Thr/Thr genotype had an extremely low serum adiponectin level (0.6 μg/ml) and the phenotype of metabolic syndrome. Our results suggest that a common variant of ADIPOQ, the minor allele of rs1656930, is associated with hypoadiponectinemia and hypertension. Screening for a common genetic background underlying low adiponectin levels might provide important information for assessment and management of metabolic disorders.

The ratio of adiponectin to homeostasis model assessment of insulin resistance is a powerful index of each component of metabolic syndrome in an aged Japanese population: Results from the KING Study

We evaluated the ratio of adiponectin level to homeostasis model assessment of insulin resistance (A/H ratio) as a risk marker for metabolic syndrome (MetS) and each of its components. The A/H ratio may prove to be a powerful index for evaluation of risk for MetS and each of its components.