Motoki Matsuki

Nerve growth factor stimulates regeneration of perivascular nerve, and induces the maturation of microvessels around the injured artery.

An immature vasa vasorum in the adventitia of arteries has been implicated in induction of the formation of unstable atherosclerotic plaques. Normalization/maturation of the vasa vasorum may be an attractive therapeutic approach for arteriosclerotic diseases. Nerve growth factor (NGF) is a pleotropic molecule with angiogenic activity in addition to neural growth effects. However, whether NGF affects the formation of microvessels in addition to innervation during pathological angiogenesis is unclear. In the present study, we show a new role for NGF in neovessels around injured arterial walls using a novel in vivo angiogenesis assay. The vasa vasorum around arterial walls was induced to grow using wire-mediated mouse femoral arterial injury. When collagen-coated tube (CCT) was placed beside the injured artery for 7-14 days, microvessels grew two-dimensionally in a thin layer on the CCT (CCT-membrane) in accordance with the development of the vasa vasorum. The perivascular nerve was found at not only arterioles but also capillaries in the CCT-membrane. Biodegradable hydrogels containing VEGF and NGF were applied around the injured artery/CCT. VEGF significantly increased the total length and instability of microvessels within the CCT-membrane. In contrast, NGF induced regeneration of the peripheral nerve around the microvessels and induced the maturation and stabilization of microvessels. In an ex vivo nerve-free angiogenesis assay, although NGF potentially stimulated vascular sprouting from aorta tissues, no effects of NGF on vascular maturation were observed. These data demonstrated that NGF had potent angiogenic effects on the microvessels around the injured artery, and especially induced the maturation/stabilization of microvessels in accordance with the regeneration of perivascular nerves.

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.

Malnutrition, renal dysfunction and left ventricular hypertrophy synergistically increase the long-term incidence of cardiovascular events

Although malnutrition indicates an unfavorable prognosis in some clinical settings, the synergistic impact of nutritional state, renal dysfunction and left ventricular hypertrophy (LVH) on cardiovascular events is unknown. Among 338 patients aged 40–80 years who underwent echocardiographic evaluation between 2003 and 2005, 161 patients who were followed for >7 years were recruited. Malnutrition was defined as a geriatric nutritional risk index (GNRI) of ⩽96. The mean patient age was 63.5±9.2 years; the mean estimated glomerular filtration rate (eGFR) was 72.9±18.7 ml min−1 per 1.73 m2; the mean LV mass index was 114±33 g m−2; and the mean GNRI was 100.4±6.0. Among the patients, 25% (n=40) had an eGFR of <60 ml min−1 per 1.73 m2, 29% (n=46) exhibited chronic kidney disease (CKD) and 37% (n=59) had LVH. During the follow-up period (median: 96 months), cardiovascular events were observed in 15 patients (9%). Kaplan–Meier curves showed a significantly higher incidence of cardiovascular events in patients with an eGFR of <60 ml min−1 per 1.73 m2 (log-rank P=0.007), a GNRI of ⩽96 (P=0.003) or LVH (P=0.010). In a Cox regression analysis, eGFR, LVH and GNRI were independent determinants of cardiovascular event incidence after adjusting for age, gender and the presence of hypertension and diabetes. Furthermore, the combination of LVH and lower GNRI was significantly associated with a higher rate of cardiovascular events not only in all patients but also in patients with CKD. In conclusion, malnutrition, low eGFR and LVH were independent determinants of cardiovascular event incidence; they synergistically increased rates of these events in the long term. The evaluation and management of LVH progression and the improvement of nutritional status are critical for preventing cardiovascular complications even in non-dialysis patients.

Late gadolinium enhancement on cardiac magnetic resonance represents the depolarizing and repolarizing electrically damaged foci causing malignant ventricular arrhythmia in hypertrophic cardiomyopathy

BACKGROUND The presence of a myocardial scar detected by late gadolinium enhancement (LGE) on cardiac magnetic resonance (CMR) has been described as a predictor of all-cause mortality in hypertrophic cardiomyopathy (HCM). However, the detailed spatial relationship between LGE site and electrical abnormality is unclear in high-risk HCM with malignant arrhythmia. OBJECTIVE The purpose of this study was to elucidate the detailed relationship between the site on CMR imaging and the electrically damaged site, a potential origin of ventricular arrhythmias in patients with HCM. METHODS Fifty consecutive HCM patients underwent contrast-enhanced CMR. Of those patients, 18 patients with ventricular tachycardia underwent electrophysiology study including endocardial mapping of the left ventricle (LV). The LGE area was calculated at 12 different LV sites: anterior, lateral, posterior, and septal segments of the basal, middle, and apical portions. At each LV site, the bipolar electrogram, effective refractory period (ERP), and monophasic action potential were recorded. RESULTS LGE-positive segments demonstrated a significantly lower amplitude (4.0 ± 2.8 mV vs 7.3 ± 3.6 mV; P < .001), longer duration (54.7 ± 17.8 vs 40.6 ± 7.8 ms; P < .001), longer ERP (320 ± 42 ms vs 284 ± 37 ms; P = .001), and longer monophasic action potential duration measured at 90% repolarization (321 ± 19 ms vs 283 ± 25 ms; P < .001) than did LGE-negative segments. The LGE area negatively correlated with the amplitude (r = -0.59; P < .001) and positively correlated with the duration (r = 0.64; P < .001), ERP (r = 0.44; P < .001), and action potential duration measured at 90% repolarization (r = 0.63; P < .001). All the observed VTs originated from LGE-positive segments. CONCLUSION The spatial distribution of LGE significantly correlates with depolarizing and repolarizing electrical damage in high-risk HCM with malignant ventricular arrhythmia.

Ninjurin1 Is a Novel Factor to Regulate Angiogenesis Through the Function of Pericytes

BACKGROUND Capillary pericytes (cPCs), the mural cells of microvessels, play an important role in the formation and maintenance of microvessels; however, little is known about the mechanisms of how cPCs regulate angiogenesis. To identify factors that modulate cPC function, genes whose levels were altered in cPCs during neovessel formation were identified through a microarray screen. METHODS AND RESULTS Ninjurin1 (nerve injury-induced protein, Ninj1) was selected as a candidate factor for angiogenesis regulation. Ninj1 was expressed in capillary cells including endothelial cells (cECs) and was expressed at a higher level in cPCs. Hypoxia induced the gene expression of Ninj1 in addition of vascular endothelial growth factor (VEGF) in cPCs. When cPCs were co-incubated with a thoracic aorta in a three-dimensional Matrigel system, the length of the EC-tubes sprouting from the aorta was increased. Small interfering RNA-mediated downregulation of Ninj1 in cPCs enhanced these cPCs-mediated angiogenic effects, whereas overexpression of Ninj1 attenuated their effects. The production of angiogenic growth factors, such as VEGF and angiopoietin 1, by cPCs was enhanced by the downregulation of Ninj1, and reduced by the overexpression of Ninj1. CONCLUSIONS Ninj1 is a novel regulator for the angiogenic effect of PCs. Specifically, Ninj1 negatively regulates the formation of neovessels, that is, the EC-tube, by reducing the trophic effects of cPCs.

Apoptosis‐Resistant Cardiac Progenitor Cells Modified With Apurinic/Apyrimidinic Endonuclease/Redox Factor 1 Gene Overexpression Regulate Cardiac Repair After Myocardial Infarction

Overcoming the insufficient survival of cell grafts is an essential objective in cell‐based therapy. Apurinic/apyrimidinic endonuclease/redox factor 1 (APE1) promotes cell survival and may enhance the therapeutic effect of engrafted cells. The aim of this study is to determine whether APE1 overexpression in cardiac progenitor cells (CPCs) could ameliorate the efficiency of cell‐based therapy. CPCs isolated from 8‐ to 10‐week‐old C57BL/6 mouse hearts were infected with retrovirus harboring APE1‐DsRed (APE1‐CPC) or a DsRed control (control‐CPC). Oxidative stress‐induced apoptosis was then assessed in APE1‐CPCs, control‐CPCs, and neonatal rat ventricular myocytes (NRVMs) cocultured with these CPCs. This analysis revealed that APE1 overexpression inhibited CPC apoptosis with activation of transforming growth factor β‐activated kinase 1 (TAK1) and nuclear factor (NF)‐κB. In the coculture model, NRVM apoptosis was inhibited to a greater extent in the presence of APE1‐CPCs compared with control‐CPCs. Moreover, the number of surviving DsRed‐positive CPC grafts was significantly higher 7 days after the transplant of APE1‐CPCs into a mouse myocardial infarction model, and the left ventricular ejection fraction showed greater improvement with attenuation of fibrosis 28 days after the transplant of APE1‐CPCs compared with control‐CPCs. Additionally, fewer inflammatory macrophages and a higher percentage of cardiac α‐sarcomeric actinin‐positive CPC‐grafts were observed in mice injected with APE1‐CPCs compared with control‐CPCs after 7 days. In conclusion, antiapoptotic APE1‐CPC graft, which increased TAK1‐NF‐κB pathway activation, survived effectively in the ischemic heart, restored cardiac function, and reduced cardiac inflammation and fibrosis. APE1 overexpression in CPCs may serve as a novel strategy to improve cardiac cell therapy.

Impact of metabolic disturbances and malnutrition-inflammation on 6-year mortality in Japanese patients undergoing hemodialysis.

Metabolic syndrome confers an increased risk of cardiovascular disease (CVD) in the general population. The relationship between adiponectins, and clinical outcomes in patients undergoing hemodialysis remains controversial. We investigated whether adiponectins, biomarkers of inflammation, nutrition status and clinical features predict the mortality of patients undergoing hemodialysis for 6 years. We measured baseline plasma total and high‐molecular‐weight (HMW) adiponectins, tumor necrosis factor (TNF)‐α, serum high sensitivity C‐reactive protein (hsCRP), and clinical characteristics including visceral fat area (VFA) and the Geriatric Nutritional Risk Index (GNRI) in 133 patients undergoing chronic hemodialysis. Forty‐one of the 133 patients died during follow‐up. The deceased patients were significantly older, had more prior CVD and diabetes, higher TNF‐α and hsCRP levels but lower GNRI. VFA, and total and HMW adiponectin did not significantly differ between the two groups. TNF‐α and hsCRP levels and GNRI score were significant for predicting all‐cause and cardiovascular mortality in receiver operating curve analyses. When stratified by a GNRI score of 96, Cox proportional hazards analyses identified TNF‐α as a significant predictor of all‐cause mortality (hazard ratio [HR] 1.23; P = 0.038) and hsCRP as a significant predictor of all‐cause and cardiovascular mortality (HR, 2.32, P = 0.003; HR 2.30, P = 0.012, respectively) after adjusting for age, sex, diabetes mellitus, and prior CVD, only in malnourished patients. These results demonstrate that malnutrition and the inflammatory markers TNF‐α and hsCRP, but not metabolic markers, including VFA and adiponectins have a significant impact on 6‐year all‐cause and cardiovascular mortality in Japanese patients undergoing hemodialysis.

The balance of fetuin-A and osteoprotegerin is independently associated with diastolic dysfunction in hemodialysis patients

Fetuin-A and osteoprotegerin (OPG) are arterial calcification regulators, which are related to cardiovascular survival in hemodialysis patients. We hypothesized that a balance of these calcification regulators might mediate the progression of left ventricular (LV) diastolic dysfunction in hemodialysis patients. We recruited 63 hemodialysis patients and measured their serum fetuin-A, OPG, arterial stiffness, aortic calcification and echocardiographic parameters, including the transmitral early diastolic velocity/tissue Doppler mitral annular early diastolic velocity ratio (E/E′), and analyzed the relationships between these variables. Fetuin-A levels were significantly and negatively correlated with the ankle–brachial pulse wave velocity (baPWV), aortic calcification score (AOCS), left atrial volume index (LAVI), LV mass index (LVMI) and E/E′. OPG levels and the ratio of OPG to fetuin-A levels were significantly and positively correlated with the baPWV, AOCS, LAVI and E/E′. A stepwise multiple regression analysis revealed that E/E′ was independently correlated with fetuin-A levels (β=−0.334, P=0.02), OPG levels (β=0.367, P=0.01) and the ratio of OPG to fetuin-A (β=0.295, P=0.04). Categorizing the patients according to their serum fetuin-A and OPG levels revealed that patients with low fetuin-A and high OPG levels had the highest LAVI, LVMI and E/E′ values after adjusting for potential confounders. Serum fetuin-A levels negatively reflected, whereas OPG levels and the ratio of OPG to fetuin-A positively reflected an increase in vascular and ventricular stiffness, leading to the aggravation of diastolic dysfunction. Therefore, based on our results, the balance of the tissue calcification regulators fetuin-A and OPG could mediate the progression of LV diastolic dysfunction in hemodialysis patients.

An Acute Myocardial Infarction Case that Survived An Out‐of‐Hospital Cardiac Arrest in Which Prominent Ischemic J Waves Were Documented

We describe a case of a myocardial infarction, in which prominent ischemic J waves were documented during recurrent ventricular fibrillation attacks. The patient was referred to our hospital to treat an out‐of hospital cardiac arrest. Although the 12‐lead electrocardiogram obtained just after the first cardioversion did not show any apparent J waves, a J wave‐like steep downsloping type ST‐segment elevation associated with q waves in the inferior leads was documented during multiple episodes of ventricular fibrillation. Our report revealed the appearance of J waves as an important marker for lethal arrhythmias in acute ischemia. (PACE 2012; 35:e27–e30)

Apurinic/apyrimidinic endonucelase 1 maintains adhesion of endothelial progenitor cells and reduces neointima formation

Apurinic/apyrimidinic endonuclease 1 (APE1) is a multifunctional protein that processes DNA-repair function and controls cellular response to oxidative stress. Endothelial progenitor cells (EPCs) are recruited to oxidative stress-rich injured vascular walls and positively contribute to vascular repair and endothelialization. We hypothesized that APE1 functions for EPCs-mediated inhibition of neointima formation in injured vasculature. EPCs isolated from bone marrow cells of C57BL6 mice (12-16 wk old) were able to survive in the presence of hydrogen peroxide (H2O2; up to 1,000 μM) due to the highly expressed reactive oxygen species (ROS) scavengers. However, adhesion capacity of EPCs was significantly inhibited by H2O2 (100 μM) even though an intracellular ROS was retained at small level. An APE1-selective inhibitor or RNA interference-mediated knockdown of endogenous APE1 in EPCs aggravated the H2O2-mediated inhibition of EPCs-adhesion. In contrast, when APE1 was overexpressed in EPCs using an adenovirus harboring the APE1 gene (APE-EPCs), adhesion was significantly improved during oxidative stress. To examine in vivo effects of APE1 in EPCs, APE-EPCs were transplanted via the tail vein after wire-mediated injury of the mouse femoral artery. The number of adherent EPCs at injured vascular walls and the vascular repair effect of EPCs were enhanced in APE-EPCs compared with control EPCs. Among the cellular functions of EPCs, adhesion is especially sensitive to oxidative stress. APE1 enhances in vivo vascular repair effects of EPCs in part through the maintenance of adhesion properties of EPCs. APE1 may be a novel and useful target gene for effective cellular transplantation therapy.