Yukihiro Saito

Fragmented QRS is associated with torsades de pointes in patients with acquired long QT syndrome

BACKGROUNDnAcquired long QT syndrome (LQTS) is a disease due to a secondary repolarization abnormality induced by various predisposing factors. In contrast to congenital LQTS, risk factors that produce acquired LQTS include organic heart diseases that often exhibit depolarization abnormality. Although various repolarization parameters have been evaluated in acquired LQTS, the existence of depolarization abnormality in association with torsades de pointes (TdP) has not been reported.nnnOBJECTIVEnThe purpose of this study was to evaluate both repolarization (QT components) and depolarization parameters (fragmented QRS [fQRS]) in acquired LQTS patients with markedly prolonged QT interval.nnnMETHODSnSeventy patients with acquired severe QT prolongation (QTc ≥ 550 ms) were studied. Thirty-two patients had syncope or TdP (syncope group). Thirty-eight patients did not have any symptoms (asymptomatic group). The existence of fQRS and QT components (QT, QTc, Tpe [interval between peak and end of T wave] intervals, and U-wave voltage) was analyzed.nnnRESULTSnThe syncope group had more frequent fQRS (81%) than did the asymptomatic group (21%, P < .01) and the incidence of fQRS was not different before and after removal of predisposing factors. The incidence of organic heart disease was not different between the two groups. No differences in QTc interval were noted between the syncope and asymptomatic groups, although the syncope group had longer QT and Tpe intervals and higher U wave than the asymptomatic group (P < .01).nnnCONCLUSIONnAcquired predisposing factors promoted repolarization abnormality (especially prolongation of QT and Tpe intervals), and the existence of fQRS had an important role in the development of TdP in patients with acquired LQTS.

epoprostenol sodium for treatment of pulmonary arterial hypertension

The release of endogenous prostacyclin (PGI2) is depressed in patients with pulmonary arterial hypertension (PAH). PGI2 replacement therapy by epoprostenol infusion is one of the best treatments available for PAH. Here, we provide an overview of the current clinical data for epoprostenol. Epoprostenol treatment improves symptoms, exercise capacity, and hemodynamics, and is the only treatment that has been shown to reduce mortality in patients with idiopathic PAH (IPAH) in randomized clinical trials. We have reported that high-dose epoprostenol therapy (>40 ng/kg/min) also results in marked hemodynamic improvement in some patients with IPAH. High-dose epoprostenol has a pro-apoptotic effect on PAH-PASMCs via the IP receptor and upregulation of Fas ligand (FasL) in vitro. However, long-term intravenous administration of epoprostenol is sometimes associated with catheter-related infections and leads to considerable inconvenience for the patient. In the future, the development of new routes of administration or the development of powerful PGI2 analogs, IP-receptor agonists, and gene and cell-based therapy enhancing PGI2 production with new routes of administration is required.

Beta-Blockers and Oxidative Stress in Patients with Heart Failure

Oxidative stress has been implicated in the pathogenesis of heart failure. Reactive oxygen species (ROS) are produced in the failing myocardium, and ROS cause hypertrophy, apoptosis/cell death and intracellular Ca2+ overload in cardiac myocytes. ROS also cause damage to lipid cell membranes in the process of lipid peroxidation. In this process, several aldehydes, including 4-hydroxy-2-nonenal (HNE), are generated and the amount of HNE is increased in the human failing myocardium. HNE exacerbates the formation of ROS, especially H2O2 and ·OH, in cardiomyocytes and subsequently ROS cause intracellular Ca2+ overload. Treatment with beta-blockers such as metoprolol, carvedilol and bisoprolol reduces the levels of oxidative stress, together with amelioration of heart failure. This reduction could be caused by several possible mechanisms. First, the beta-blocking effect is important, because catecholamines such as isoproterenol and norepinephrine induce oxidative stress in the myocardium. Second, anti-ischemic effects and negative chronotropic effects are also important. Furthermore, direct antioxidative effects of carvedilol contribute to the reduction of oxidative stress. Carvedilol inhibited HNE-induced intracellular Ca2+ overload. Beta-blocker therapy is a useful antioxidative therapy in patients with heart failure.

Enhancement of Spontaneous Activity by HCN4 Overexpression in Mouse Embryonic Stem Cell-Derived Cardiomyocytes - A Possible Biological Pacemaker.

Background Establishment of a biological pacemaker is expected to solve the persisting problems of a mechanical pacemaker including the problems of battery life and electromagnetic interference. Enhancement of the funny current (I f) flowing through hyperpolarization-activated cyclic nucleotide-gated (HCN) channels and attenuation of the inward rectifier K+ current (I K1) flowing through inward rectifier potassium (Kir) channels are essential for generation of a biological pacemaker. Therefore, we generated HCN4-overexpressing mouse embryonic stem cells (mESCs) and induced cardiomyocytes that originally show poor I K1 currents, and we investigated whether the HCN4-overexpressing mESC-derived cardiomyocytes (mESC-CMs) function as a biological pacemaker in vitro. Methods and Results The rabbit Hcn4 gene was transfected into mESCs, and stable clones were selected. mESC-CMs were generated via embryoid bodies and purified under serum/glucose-free and lactate-supplemented conditions. Approximately 90% of the purified cells were troponin I-positive by immunostaining. In mESC-CMs, expression level of the Kcnj2 gene encoding Kir2.1, which is essential for generation of I K1 currents that are responsible for stabilizing the resting membrane potential, was lower than that in an adult mouse ventricle. HCN4-overexpressing mESC-CMs expressed about a 3-times higher level of the Hcn4 gene than did non-overexpressing mESC-CMs. Expression of the Cacna1h gene, which encodes T-type calcium channel and generates diastolic depolarization in the sinoatrial node, was also confirmed. Additionally, genes required for impulse conduction including Connexin40, Connexin43, and Connexin45 genes, which encode connexins forming gap junctions, and the Scn5a gene, which encodes sodium channels, are expressed in the cells. HCN4-overexpressing mESC-CMs showed significantly larger I f currents and more rapid spontaneous beating than did non-overexpressing mESC-CMs. The beating rate of HCN4-overexpressing mESC-CMs responded to ivabradine, an I f inhibitor, and to isoproterenol, a beta-adrenergic receptor agonist. Co-culture of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) with aggregates composed of mESC-CMs resulted in synchronized contraction of the cells. The beating rate of hiPSC-CMs co-cultured with aggregates of HCN4-overexpressing mESC-CMs was significantly higher than that of non-treated hiPSC-CMs and that of hiPSC-CMs co-cultured with aggregates of non-overexpressing mESC-CMs. Conclusions We generated HCN4-overexpresssing mESC-CMs expressing genes required for impulse conduction, showing rapid spontaneous beating, responding to an I f inhibitor and beta-adrenergic receptor agonist, and having pacing ability in an in vitro co-culture system with other excitable cells. The results indicated that these cells could be applied to a biological pacemaker.

Immortalized multipotent pericytes derived from the vasa vasorum in the injured vasculature. A cellular tool for studies of vascular remodeling and regeneration.

Adventitial microvessels, vasa vasorum in the vessel walls, have an active role in the vascular remodeling, although its mechanisms are still unclear. It has been reported that microvascular pericytes (PCs) possess mesenchymal plasticity. Therefore, microvessels would serve as a systemic reservoir of stem cells and contribute to the tissues remodeling. However, most aspects of the biology of multipotent PCs (mPCs), in particular of pathological microvessels are still obscure because of the lack of appropriate methods to detect and isolate these cells. In order to examine the characteristics of mPCs, we established immortalized cells residing in adventitial capillary growing at the injured vascular walls. We recently developed in vivo angiogenesis to observe adventitial microvessels using collagen-coated tube (CCT), which also can be used as an adventitial microvessel-rich tissue. By using the CCT, CD146- or NG2-positive cells were isolated from the adventitial microvessels in the injured arteries of mice harboring a temperature-sensitive SV40 T-antigen gene. Several capillary-derived endothelial cells (cECs) and PCs (cPCs) cell lines were established. cECs and cPCs maintain a number of key endothelial and PC features. Co-incubation of cPCs with cECs formed capillary-like structure in Matrigel. Three out of six cPC lines, termed capillary mPCs demonstrated both mesenchymal stem cell- and neuronal stem cell-like phenotypes, differentiating effectively into adipocytes, osteoblasts, as well as schwann cells. mPCs differentiated to ECs and PCs, and formed capillary-like structure on their own. Transplanted DsRed-expressing mPCs were resident in the capillary and muscle fibers and promoted angiogenesis and myogenesis in damaged skeletal muscle. Adventitial mPCs possess transdifferentiation potential with unique phenotypes, including the reconstitution of capillary-like structures. Their phenotype would contribute to the pathological angiogenesis associated with vascular remodeling. These cell lines also provide a reproducible cellular tool for high-throughput studies on angiogenesis, vascular remodeling, and regeneration as well.

Delivery of Imatinib-Incorporated Nanoparticles into Lungs Suppresses the Development of Monocrotaline-Induced Pulmonary Arterial Hypertension

Platelet-derived growth factor (PDGF) is implicated in the pathogenesis of pulmonary arterial hypertension (PAH). Imatinib, a PDGF-receptor tyrosine kinase inhibitor, improved hemodynamics, but serious side effects and drug discontinuation are common when treating PAH. A drug delivery system using nanoparticles (NPs) enables the reduction of side effects while maintaining the effects of the drug. We examined the efficacy of imatinib-incorporated NPs (Ima-NPs) in a rat model and in human PAH-pulmonary arterial smooth muscle cells (PASMCs). Rats received a single intratracheal administration of PBS, FITC-NPs, or Ima-NPs immediately after monocrotaline injection. Three weeks after monocrotaline injection, intratracheal administration of Ima-NPs suppressed the development of pulmonary hypertension, small pulmonary artery remodeling, and right ventricular hypertrophy in the rat model of monocrotaline-induced PAH. We also examined the effects of imatinib and Ima-NPs on PDGF-induced proliferation of human PAH-PASMCs by (3)H-thymidine incorporation. Imatinib and Ima-NPs significantly inhibited proliferation after 24 hours of treatment. Ima-NPs significantly inhibited proliferation compared with imatinib at 24 hours after removal of these drugs. Delivery of Ima-NPs into lungs suppressed the development of MCT-induced PAH by sustained antiproliferative effects on PAS-MCs.

Pioglitazone prevents the endothelial dysfunction induced by ischemia and reperfusion in healthy subjects

Background: No study has investigated whether pioglitazone (an agonist of peroxisome proliferator-activated receptor gamma) protects against ischemia and reperfusion (IR)–induced endothelial dysfunction in humans. Methods and Results: In the first crossover study, 20 volunteers were randomized to 1 week of pioglitazone (30 mg/d, postoperatively) or control (no treatment). In the second single-arm study, 15 volunteers received pioglitazone and the cyclooxygenase-2 inhibitor meloxicam for 1 week. On day 7, endothelium-dependent flow-mediated dilation (FMD) of the distal brachial artery was measured before and after IR (15 minutes of ischemia followed by 15 minutes of reperfusion in the proximal upper arm). Pre-IR brachial-artery diameter and FMD were similar across the 2 sessions (control, pioglitazone) in protocol 1 and between the 2 protocols. IR significantly blunted FMD after no treatment (pre-IR FMD: 10.2% ± 2.6%; post-IR FMD: 3.5% ± 1.9%, P < 0.01) but not after pioglitazone administration (pre-IR FMD: 9.7% ± 2.5%; post-IR FMD: 8.8% ± 2.9%, P = 0.11). This protective effect was accompanied by an increase in serum levels of the antioxidant enzyme extracellular superoxide dismutase and was not affected by concomitant administration of the cyclooxygenase-2 inhibitor meloxicam (P = 0.10). Conclusions: In humans, pioglitazone provides potent protection against IR-induced endothelial dysfunction.

HCN4-Overexpressing Mouse Embryonic Stem Cell-Derived Cardiomyocytes Generate a New Rapid Rhythm in Rats with Bradycardia

A biological pacemaker is expected to solve the persisting problems of an artificial cardiac pacemaker including short battery life, lead breaks, infection, and electromagnetic interference. We previously reported HCN4 overexpression enhances pacemaking ability of mouse embryonic stem cell-derived cardiomyocytes (mESC-CMs) in vitro. However, the effect of these cells on bradycardia in vivo has remained unclear. Therefore, we transplanted HCN4-overexpressing mESC-CMs into bradycardia model animals and investigated whether they could function as a biological pacemaker. The rabbit Hcn4 gene was transfected into mouse embryonic stem cells and induced HCN4-overexpressing mESC-CMs. Non-cardiomyocytes were removed under serum/glucose-free and lactate-supplemented conditions. Cardiac balls containing 5 × 103 mESC-CMs were made by using the hanging drop method. One hundred cardiac balls were injected into the left ventricular free wall of complete atrioventricular block (CAVB) model rats. Heart beats were evaluated using an implantable telemetry system 7 to 30 days after cell transplantation. The result showed that ectopic ventricular beats that were faster than the intrinsic escape rhythm were often observed in CAVB model rats transplanted with HCN4-overexpressing mESC-CMs. On the other hand, the rats transplanted with non-overexpressing mESC-CMs showed sporadic single premature ventricular contraction but not sustained ectopic ventricular rhythms. These results indicated that HCN4-overexpressing mESC-CMs produce rapid ectopic ventricular rhythms as a biological pacemaker.

Cell-based Biological Pacemakers: Progress and Problems.

The number of permanent pacemaker implantations has been increasing due to the aging of populations worldwide and the increase in the numbers of patients with heart diseases. Commercially available mechanical pacemakers are very useful but still have some problems including short battery life, a risk of infection, the absence of physiological autonomic responsiveness, metal allergy, and electronic interference. A biological pacemaker may resolve these problems and regenerate the cardiac pacemaker. Cell-based therapy and gene therapy have been addressed with the goal of solving the challenges of biological pacemaker. However, the clinical application of a biological pacemaker has not yet been realized. Here we discuss the types of cells that can be used for a biological pacemaker and the problems that remain regarding the clinical applications of cell-based therapy.

Eicosapentaenoic acid prevents arterial calcification in klotho mutant mice

Background The klotho gene was identified as an “aging-suppressor” gene that accelerates arterial calcification when disrupted. Serum and vascular klotho levels are reduced in patients with chronic kidney disease, and the reduced levels are associated with arterial calcification. Intake of eicosapentaenoic acid (EPA), an n-3 fatty acid, reduces the risk of fatal coronary artery disease. However, the effects of EPA on arterial calcification have not been fully elucidated. The aim of this study was to determine the effect of EPA on arterial calcification in klotho mutant mice. Methods and results Four-week-old klotho mutant mice and wild-type (WT) mice were given a diet containing 5% EPA (EPA food, klotho and WT: n = 12, each) or not containing EPA (control food, klotho and WT: n = 12, each) for 4 weeks. Calcium volume scores of thoracic and abdominal aortas assessed by computed tomography were significantly elevated in klotho mice after 4 weeks of control food, but they were not elevated in klotho mice after EPA food or in WT mice. Serum levels of EPA and resolvin E1, an active metabolite of EPA, in EPA food-fed mice were significantly increased compared to those in control food-fed mice. An oxidative stress PCR array followed by quantitative PCR revealed that NADPH oxidase-4 (NOX4), an enzyme that generates superoxide, gene expression was up-regulated in arterial smooth muscle cells (SMCs) of klotho mice. Activity of NOX was also significantly higher in SMCs of klotho mice than in those of WT mice. EPA decreased expression levels of the NOX4 gene and NOX activity. GPR120, a receptor of n-3 fatty acids, gene knockdown by siRNA canceled effects of EPA on NOX4 gene expression and NOX activity in arterial SMCs of klotho mice. Conclusions EPA prevents arterial calcification together with reduction of NOX gene expression and activity via GPR120 in klotho mutant mice.