Growth Factor Midkine Aggravates Pulmonary Arterial Hypertension via Surface Nucleolin.

Pulmonary arterial hypertension (PAH) is a progressive fatal disease caused by pulmonary arterial remodeling. Midkine regulates cell proliferation and migration, and it is induced by hypoxia, but its roles in pulmonary arterial remodeling remain unclear. Serum midkine levels were significantly increased in PAH patients compared with control patients. Midkine expression was increased in lungs and sera of hypoxia-induced PAH mice. Hypoxia-induced pulmonary arterial remodeling and right ventricular hypertrophy were attenuated in midkine-knockout mice. Midkine-induced proliferation and migration of pulmonary arterial smooth muscle cells (PASMC) and epidermal growth factor receptor (EGFR) signaling were significantly increased under hypoxia, which also induced cell-surface translocation of nucleolin. Nucleolin siRNA treatment suppressed midkine-induced EGFR activation in vitro, and nucleolin inhibitor AS1411 suppressed proliferation and migration of PASMC induced by midkine. Furthermore, AS1411 significantly prevented the development of PAH in Sugen hypoxia rat model. Midkine plays a crucial role in PAH development through interaction with surface nucleolin. These data define a role for midkine in PAH development and suggest midkine-nucleolin-EGFR axis as a novel therapeutic target for PAH.

Therapeutic inhibition of microRNA-34a ameliorates aortic valve calcification via modulation of Notch1-Runx2 signalling.

AIMS: Calcific aortic valve stenosis (CAVS) is the most common valvular heart disease and is increased with elderly population. However, effective drug therapy has not been established yet. This study aimed to investigate the role of microRNAs (miRs) in the development of CAVS. METHODS AND RESULTS: We measured the expression of 10 miRs, which were reportedly involved in calcification by using human aortic valve tissue from patients who underwent aortic valve replacement with CAVS or aortic regurgitation (AR) and porcine aortic valve interstitial cells (AVICs) after treatment with osteogenic induction medium. We investigated whether a specific miR-inhibitor can suppress aortic valve calcification in wire injury CAVS mice model. Expression of miR-23a, miR-34a, miR-34c, miR-133a, miR-146a, and miR-155 was increased, and expression of miR-27a and miR-204 was decreased in valve tissues from CAVS compared with those from AR. Expression of Notch1 was decreased, and expression of Runt-related transcription factor 2 (Runx2) was increased in patients with CAVS compared with those with AR. We selected miR-34a among increased miRs in porcine AVICs after osteogenic treatment, which was consistent with results from patients with CAVS. MiR-34a increased calcium deposition in AVICs compared with miR-control. Notch1 expression was decreased, and Runx2 expression was increased in miR-34a transfected AVICs compared with that in miR-control. Conversely, inhibition of miR-34a significantly attenuated these calcification signals in AVICs compared with miR-control. RNA pull-down assay revealed that miR-34a directly targeted Notch1 expression by binding to Notch1 mRNA 3' untranslated region. In wire injury CAVS mice, locked nucleic acid miR-34a inhibitor suppressed aortic velocity, calcium deposition of aortic valves, and cardiac hypertrophy, which were involved in decreased Runx2 and increased Notch1 expressions. CONCLUSION: miR-34a plays an important role in the development of CAVS via Notch1-Runx2 signalling pathway. Inhibition of miR-34a may be the therapeutic target for CAVS.

Impact of Iron Deficiency on Peripheral Artery Disease After Endovascular Therapy.

Background: Despite advances in endovascular therapy (EVT), peripheral artery disease (PAD) is a public health problem associated with high cardiovascular mortality. Iron deficiency (ID) is associated with poor clinical outcome in patients with heart disease, but whether ID is associated with the severity and clinical outcome of PAD remains unclear. Methods and Results: A total of 449 patients with PAD who received EVT and who had iron and red blood cell measurement were enrolled. ID was defined as transferrin saturation (TSAT) <20%, based on a previous report. TSAT and hemoglobin decreased with deteriorating Fontaine class. During a median follow-up period of 1,064 days, 71 major adverse cardiovascular and leg events (MACLE) and 47 major adverse cardiovascular events (MACE) were noted. All patients were divided into 2 groups based on the presence of ID. On Kaplan-Meier analysis, patients with ID had higher rates of MACE and MACLE than those without. On multivariate Cox proportional hazard regression analysis, TSAT and hemoglobin were independently associated with MACLE. Addition of TSAT to the known risk factors significantly improved the net reclassification index and integrated discrimination improvement. Conclusions: ID, as assessed by TSAT, was associated with the severity and clinical outcome of PAD, indicating that it could be a therapeutic target.

Increased plasma xanthine oxidoreductase activity deteriorates coronary artery spasm.

Increased reactive oxygen species (ROS) contributes to the development of endothelial dysfunction, which is involved in coronary artery spasm (CAS). Xanthine oxidoreductase (XOR) plays a pivotal role in producing both uric acid and ROS. However, the association between plasma XOR activity and CAS has not been elucidated. The aim of this study was to investigate whether plasma XOR activity is associated with CAS. We measured XOR activity in 104 patients suspected for CAS, who presented without significant coronary artery stenosis and underwent intracoronary acetylcholine provocation tests. CAS was provoked in 44 patients and they had significantly higher XOR activity as compared with those without CAS. The patients were divided into three groups based on the XOR activity. The prevalence rate of CAS was increased with increasing XOR activity. A multivariate logistic regression analysis showed that the 3rd tertile group exhibited a higher incidence of CAS as compared with the 1st tertile group [odds ratio (OR) 6.9, P = 0.001) and the 2nd tertile group (OR 3.2, P = 0.033) after adjustment for conventional CAS risk factors, respectively. The C index was significantly improved by the addition of XOR activity to the baseline model based on CAS risk factors. Furthermore, the 3rd tertile group had the highest incidence of severe spasm defined as total obstruction, flow-limiting stenosis, diffuse spasm, multivessel spasm, and/or lethal arrhythmia. This is a first report to elucidate the association of plasma XOR activity with CAS. Increased plasma XOR activity is significantly associated with CAS.

Decreased Psoas Muscle Computed Tomography Value Predicts Poor Outcome in Peripheral Artery Disease.

BACKGROUND: The prognosis of peripheral artery disease (PAD) and comorbid sarcopenia is poor. Some reports indicate that the computed tomography (CT) value of skeletal muscle, which reflects intramuscular fat deposition as well as skeletal muscle mass, is considered a marker of sarcopenia. However, it remains unclear if skeletal muscle area and CT value are associated with poor outcomes in patients with PAD. Methods and Results: Psoas muscle area and CT value were measured by manual trace at the level of the third lumbar vertebral body in 327 consecutive patients with PAD undergoing endovascular therapy (EVT). The endpoint was major adverse cardiovascular and limb events (MACLE). There were 60 MACLE during the follow-up period. Patients with MACLE had lower mean psoas muscle CT value than those without. However, there was no significant difference in total psoas muscle area between patients with and without MACLE. Kaplan-Meier analysis demonstrated that the lowest tertile of psoas muscle CT value was associated with the highest risk of MACLE. Multivariate Cox hazard analysis revealed that psoas muscle CT value was associated with MACLE after adjustment for Fontaine class, previous ischemic heart disease, prevalence of diabetes mellitus, brain natriuretic peptide, and serum albumin. CONCLUSIONS: Psoas muscle CT value is a feasible predictor of MACLE in patients with PAD.

Intra-articular administration of EP2 enhances the articular cartilage repair in a rabbit model.

We have reported the usefulness of chondrocyte sheets on articular cartilage repair in animal experiments. Here, we investigated the regenerative effects of EP2 signalling with or without chondrocyte sheets. Forty-five rabbits were used, with six rabbits in each of the six groups and nine rabbits for chondrocytes and synovial cells harvesting to fabricate triple-layered chondrocyte sheets: osteochondral defect only (control, Group A), EP2 agonist (Group B), EP2 antagonist (Group C), chondrocyte sheets (Group D), EP2 agonist and chondrocyte sheets (Group E), and EP2 antagonist and chondrocyte sheets (Group F). After surgery, the weight distribution ratio was measured as an indicator of pain alleviation. Injections of the EP2 agonist or EP2 antagonist were given from 4 weeks after surgery. The rabbits were sacrificed at 12 weeks, and the repaired tissues were evaluated for histology. The weight distribution ratio and International Cartilage Repair Society grading were as follows: Group A: 40.5% ± 0.2%, 14.8 ± 0.5; Group B: 43.4% ± 0.7%, 25.4 ± 0.8; Group C: 38.7% ± 0.7%, 13.7 ± 0.3; Group D: 48.6% ± 0.6%, 40.2 ± 0.5; Group E: 49.1% ± 0.3%, 40.5 ± 0.4; and Group F; 46.8% ± 0.4%, 38.7 ± 0.5. Significant differences in histology and pain alleviation were observed between groups except between Groups A and C, between Groups D and E, and between Groups D and F. These findings show that the intra-articular administration of an EP2 agonist achieved pain alleviation and tissue repair. However, no synergistic effect with chondrocyte sheets was observed.

Ventricular wall stress and silent myocardial damage are associated with pulse pressure in the general population.

Pulse pressure (PP) is a risk factor for cardiovascular diseases and is associated with increased afterload and myocardial oxygen demand. Brain natriuretic peptide (BNP) and heart-type fatty acid-binding protein (H-FABP) are known as biomarkers indicating ventricular wall stress and silent myocardial damage. However, the association between PP and ventricular wall stress and silent myocardial damage in the general population is unclear. The authors enrolled 3504 patients who participated in a community-based annual health check. Serum levels of BNP and H-FABP were measured as markers of ventricular wall stress and silent myocardial damage. Patients were divided into four groups according to the quartiles of PP. Patients in the highest PP group showed higher serum BNP and H-FABP levels than that of the other groups. Multivariate logistic analysis showed that high PP was independently associated with ventricular wall stress and silent myocardial damage on the basis of BNP and H-FABP levels. Compared with systolic blood pressure, diastolic blood pressure, and mean blood pressure, PP was superior in predicting ventricular wall stress and silent myocardial damage evaluated according to BNP and H-FABP levels, which was reflected by the receiver operating characteristic analysis. Screening of healthy patients revealed that high PP was related to high BNP and H-FABP levels, suggesting that an asymptomatic general population with high PP may be exposed to ventricular wall stress and myocardial damage and might be susceptible to silent heart failure.

Circulating Surfactant Protein-D Is Associated With Clinical Outcomes in Peripheral Artery Disease Patients Following Endovascular Therapy.

BACKGROUND: Peripheral artery disease (PAD) is a risk factor for the development of cardiovascular disease and death. Surfactant protein-D (SP-D) is a 43-kDa protein secreted from type II pneumocytes in the lungs. Recent studies have demonstrated that circulating SP-D plays a key role in the development of atherosclerosis and is related to clinical outcomes in patients with ischemic heart disease. However, it remains unclear whether circulating SP-D is associated with clinical outcomes in patients with PAD.Methods and Results:We enrolled 364 patients with PAD who underwent endovascular therapy. We measured serum levels of SP-D and Krebs von den Lungen-6 (KL-6). During a median follow-up period of 974 days, there were 69 major adverse cardiovascular and leg events (MACLE), including 48 major adverse cardiovascular events (MACE). Kaplan-Meier analysis demonstrated that patients with high SP-D (≥110 ng/mL) had higher rates of MACE and MACLE than those with low SP-D. Multivariate Cox proportional hazard regression analysis demonstrated that SP-D, but not KL-6, was an independent predictor of MACE and MACLE. The addition of SP-D to known risk factors significantly improved the C index and net reclassification index. The circulating SP-D level was affected by sex, diabetes mellitus, and cilostazol prescription. CONCLUSIONS: Circulating SP-D was associated with clinical outcomes in patients with PAD, suggesting that it may be a new therapeutic target in these patients.

The impact of non-alcoholic fatty liver disease fibrosis score on cardiac prognosis in patients with chronic heart failure.

Liver abnormalities have a strong impact on clinical outcomes in patients with heart failure (HF), and are known as cardio-hepatic syndrome. The non-alcoholic fatty liver disease (NAFLD) fibrosis score (NFS) has been developed to identify liver fibrosis in patients with NAFLD. It remains to be determined whether NFS is associated with cardiovascular prognosis in patients with chronic heart failure (CHF). We calculated NFS in 516 patients with CHF admitted to our hospital. The clinical endpoints were deaths due to progressive HF, myocardial infarction, stroke, and sudden cardiac death, and rehospitalization for worsening HF. There were 173 cardiovascular events noted during a median follow-up of 464 days. Patients with cardiovascular events showed a higher NFS as compared with those without. We divided the patients into four groups according to quartiles of NFS. The proportion of New York Heart Association functional class III/IV and serum brain natriuretic peptide levels were increased with increasing NFS. Kaplan-Meier analysis revealed that cardiovascular event rate was increased with increasing NFS in patients with CHF. In multivariate Cox proportional hazards analysis, NFS was independently associated with cardiovascular events after adjustment for confounding factors. Elevated NFS was associated with unfavorable outcomes in patients with CHF. Liver fibrosis assessed by NFS may provide valuable prognostic information in patients with CHF.

Impact of Objective Malnutrition Status on the Clinical Outcomes in Patients With Peripheral Artery Disease Following Endovascular Therapy.

BACKGROUND: Peripheral artery disease (PAD) is an athero-occlusive disease and a known risk factor for cardiovascular events. The controlling nutritional status (CONUT) score and geriatric nutritional risk index (GNRI) are objective tools for evaluating malnutrition and are reportedly associated with poor clinical outcomes in patients with fatal diseases. However, the effect of malnutrition on the clinical outcomes in patients with PAD remains unclear.Methods and Results:We enrolled 357 patients with PAD who underwent endovascular therapy. Malnutrition was diagnosed by CONUT score and GNRI as in previous reports. During a median follow-up period of 1,071 days, there were 67 major adverse cardiovascular and leg events (MACLEs). The CONUT score- and GNRI-based malnutrition statuses were identified in 56% and 46% of the patients, respectively. Proportion of malnutrition increased with advancing Fontaine class. The multivariate Cox proportional hazard regression analysis demonstrated that both the CONUT score- and GNRI-based malnutrition status was an independent predictor of MACLEs. The Kaplan-Meier analysis demonstrated that the MACLE ratio increased with deteriorating malnutrition. Finally, the addition of the CONUT score or GNRI to the known risk factors significantly improved the net reclassification index and integrated discrimination index. CONCLUSIONS: Malnutrition was common and closely associated with the clinical outcomes in patients with PAD, indicating that it is a novel therapeutic target in the management of these patients.

Impact of restrictive lung disorder on cardiovascular mortality in a general population: The Yamagata (Takahata) study.

BACKGROUND: Obstructive lung disorder (OLD) is known to be associated with cardiovascular disease. However, the impact of restrictive lung disorder (RLD) on cardiovascular mortality has not been fully investigated in the apparently healthy general population. OBJECTIVES: To clarify whether RLD is associated with cardiovascular mortality in the general population. METHODS AND RESULTS: This community-based cohort study included 3247 subjects who participated in an annual health check in Takahata. We performed spirometry in registered subjects and found that 194 (6%) had RLD, 262 (8%) had OLD, and 73 (2%) had RLD and OLD (Mixed). During a 10-year follow-up, there were 210 deaths, including 57 cardiovascular deaths. Cardiovascular mortality of subjects with RLD was significantly higher than that of subjects with normal lung function. Although the subjects with RLD were younger, comprised fewer smokers, and were more likely to be female than those with OLD, cardiovascular mortality of subjects with RLD was comparable to that of subjects with OLD. Subjects with RLD had a higher prevalence of atrial fibrillation (AF) than those with OLD, and the prevalence of AF was increased with advanced severity of RLD. Multivariate Cox proportional hazard analysis revealed that RLD was an independent predictor of cardiovascular death (hazard ratio 2.61, 95% confidence interval, 1.22-5.21) after adjustment for confounders, but OLD was not. The net reclassification improvement and integrated discrimination improvement were significantly increased by the addition of RLD to conventional cardiovascular risk factors. CONCLUSION: The presence of RLD was associated with cardiovascular mortality in the general population.

Platelet-activated serum might have a therapeutic effect on damaged articular cartilage.

Platelet-activated serum (PAS) was collected from rabbits. This contains high concentrations of growth factors, including vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF)-BB, and transforming growth factor-beta (TGF-ß). PAS was injected into the knee joints of Japanese White rabbits subjected to anterior cruciate ligament transection (ACL-T) to investigate its therapeutic effects on articular cartilage. The effect of Avastin (an anti-VEGF monoclonal antibody) on VEGF expression was also investigated. The levels of VEGF, PDGF-BB, and TGF-ß in PAS, platelet-rich plasma (PRP) and autologous serum from untreated rabbits were analysed by enzyme-linked immunosorbent assays. The samples (n = 24 rabbits) were divided into control (C), PAS (S), Avastin (A) and PAS + Avastin (S + A) treatment groups. Intra-articular injections were administered weekly for 7 weeks after ACL-T, during which the weight distribution ratios of the damaged limbs were evaluated. Histological evaluation was performed 12 weeks after ACL-T using Mankin score. The VEGF, PDGF-BB and TGF-ß expression levels were significantly higher (P < 0.05) in the PAS than in the PRP or autologous serum samples. The weight distribution ratios of damaged limbs improved significantly after ACL-T in all treatment groups (P < 0.05). The proximal medial, distal medial and lateral aspects of joints in the treatment groups showed significant differences in Mankin scores compared with controls (P < 0.05). The damaged limb weight distribution ratios, Mankin scores and articular cartilage structure did not differ significantly among the three treatment groups, which all showed significant improvements in structure compared with controls. Copyright © 2017 John Wiley & Sons, Ltd.

The effects of using vitrified chondrocyte sheets on pain alleviation and articular cartilage repair.

The effect of using vitrified-thawed chondrocyte sheets on articular cartilage repair was examined because the methods for storing chondrocyte sheets are essential for allogeneic chondrocyte sheet transplantation. Six Japanese white rabbits were used as sources of articular chondrocytes and synovial cells. Chondrocytes were harvested from the femur, and synovial cells were harvested from inside the knee joints. After coculture of the chondrocytes with synovial cells, triple-layered chondrocyte sheets were fabricated. Eighteen rabbits were used, with six rabbits in each of three groups: osteochondral defect only (control, group A); chondrocyte sheets (group B); and vitrified-thawed chondrocyte sheets (group C). An osteochondral defect was created on the femur. After transplantation, the weight distribution ratio of the undamaged and damaged limbs was measured as a pain-alleviating effect. The rabbits were euthanized at 12 weeks, and the transplanted tissues were evaluated for histology (Safranin O staining and immunostaining) using the International Cartilage Repair Society grading system. For both evaluations, significant differences were observed between groups A and B, and between groups A and C (p < 0.05). No significant differences were observed between groups B and C. Thus, pain-alleviating effects and tissue repair were achieved using vitrified-thawed chondrocyte sheets. Copyright © 2017 John Wiley & Sons, Ltd.

Association of plasma xanthine oxidoreductase activity with severity and clinical outcome in patients with chronic heart failure.

BACKGROUND: Oxidative stress due to purine degradation is associated with the development of chronic heart failure (CHF). Xanthine oxidoreductase (XOR) is a rate-limiting enzyme of purine degradation that plays a key role in uric acid (UA) production with a resultant increase in reactive oxygen species. However, the relationship between plasma XOR activity and CHF severity and clinical outcome remains unclear. METHODS AND RESULTS: We measured XOR activity in 440 patients with CHF and 44 control subjects. Abnormally high and low XOR activities were identified based on the results for 95% of the control subjects (high and low XOR activities ≥120 and <33pmol/100µL/h, respectively). The prevalence rates of high and low XOR activities increased with advancing New York Heart Association functional class. There were 158 cardiac events during a median follow-up period of 1034days. Multivariate Cox proportional hazard regression analysis showed that both high and low XOR activities were significantly associated with cardiac events in patients with CHF after adjustment for confounding risk factors including serum UA and loop diuretic use. Kaplan-Meier analysis revealed that the cardiac event rate was significantly higher in patients with either high or low XOR activity. The net reclassification index was significantly improved by adding XOR activity to the basic risk factors. CONCLUSIONS: We provide the first evidence of an association of plasma XOR activity with CHF severity and clinical outcome. Plasma XOR activity could be used to identify high-risk CHF patients and could be a therapeutic target for XOR inhibitors.

Association of the Aspartate Aminotransferase to Alanine Aminotransferase Ratio with BNP Level and Cardiovascular Mortality in the General Population: The Yamagata Study 10-Year Follow-Up.

Background. Early identification of high risk subjects for cardiovascular disease in health check-up is still unmet medical need. Cardiovascular disease is characterized by the superior increase in aspartate aminotransferase (AST) to alanine aminotransferase (ALT). However, the association of AST/ALT ratio with brain natriuretic peptide (BNP) levels and cardiovascular mortality remains unclear in the general population. Methods and Results. This longitudinal cohort study included 3,494 Japanese subjects who participated in a community-based health check-up, with a 10-year follow-up. The AST/ALT ratio increased with increasing BNP levels. And multivariate logistic analysis showed that the AST/ALT ratio was significantly associated with a high BNP (≥100 pg/mL). There were 250 all-cause deaths including 79 cardiovascular deaths. Multivariate Cox proportional hazard regression analysis revealed that a high AST/ALT ratio (>90 percentile) was an independent predictor of all-cause and cardiovascular mortality after adjustment for confounding factors. Kaplan-Meier analysis demonstrated that cardiovascular mortality was higher in subjects with a high AST/ALT ratio than in those without. Conclusions. The AST/ALT ratio was associated with an increase in BNP and was predictive of cardiovascular mortality in a general population. Measuring the AST/ALT ratio during routine health check-ups may be a simple and cost-effective marker for cardiovascular mortality.

The role of macrophage transcription factor MafB in atherosclerotic plaque stability.

BACKGROUND AND AIMS: Macrophage differentiation is associated with the development of atherosclerosis and plaque vulnerability and is regulated by transcription factor MafB. We previously reported that MafB attenuates macrophage apoptosis, which is associated with atherosclerotic plaque instability. The aim of this study was to elucidate the role of MafB in the progression of atherosclerotic plaque. METHODS: We generated macrophage-specific dominant-negative (DN) MafB transgenic mice and intercrossed DN-MafB mice with apolipoprotein E (ApoE) knockout (KO) mice. RESULTS: There was no significant difference in advanced atherosclerotic lesion area between DN-MafB/ApoE KO mice and littermate control ApoE KO mice 9 weeks after high-cholesterol diet. However, DN-MafB/ApoE KO mice showed significantly larger necrotic cores and lower collagen content in atherosclerotic plaques than ApoE KO mice. Although there was no difference in intraplaque macrophage infiltration and efferocytosis, DN-MafB/ApoE KO mice showed significantly more apoptotic macrophages at the plaque edges than did ApoE KO mice. Real-time PCR analysis revealed that peritoneal macrophages of DN-MafB/ApoE KO mice had a greater increase in matrix metalloproteinase-9 and mRNA expression of inflammatory/M1 macrophage markers (tissue necrosis factor-α, interleukin-6, CD11c, and p47phox) after lipopolysaccharide stimulation than those of ApoE KO mice. CONCLUSION: Macrophage-specific inhibition of MafB may destabilize atherosclerotic plaques in advanced lesions.

Assessment of the Safety of Chondrocyte Sheet Implantation for Cartilage Regeneration.

We have previously studied the effects of chondrocyte sheets on the repair and regeneration of articular cartilage by using temperature-responsive culture inserts. On the basis of this work, we succeeded in rapid fabrication of chondrocyte sheets with the use of a coculture method in which inserts were placed between synoviocytes and chondrocytes. Treatment of cartilage defects using layered chondrocyte sheets promotes repair and regeneration; this method is compatible with in vivo osteoarthritis models that reproduce partial-thickness defects. In human stem cell clinical research guidelines, the Ministry of Health, Labour and Welfare (MHLW) approved several applications related to this technology. Indeed, its translation to a clinical setting is already yielding favorable results. In this study, we evaluated the risk of tumorigenesis associated with this treatment and characterized the dynamics of biological processes associated with the posttransplantation cell sheets in vivo. Furthermore, we also confirmed the safety of the procedure by using array comparative genomic hybridization (array CGH) and G-band staining to screen for deleterious genetic aberrations during prolonged subculture of cells. The safety of chondrocytes that were cultured for longer than normal was confirmed by the array CGH and G-band staining results. In addition, tumorigenicity testing confirmed that culture chondrocyte sheets are not tumorigenic. Furthermore, from the evaluation of bioluminescence imaging following implantation of the cell sheets, it was confirmed that the transplanted chondrocytes and synoviocytes remained in the knee joint and did not transfer elsewhere over time. We believe that the technique used in this study is a highly useful method for evaluating the safety of not only chondrocytes but also extensive subculturing in general.

Comorbid renal tubular damage and hypoalbuminemia exacerbate cardiac prognosis in patients with chronic heart failure.

BACKGROUND: Renal tubular damage (RTD) and hypoalbuminemia are risks for poor prognosis in patients with chronic heart failure (CHF). Renal tubules play a pivotal role in amino acid and albumin reabsorption, which maintain serum albumin levels. The aims of the present study were to (1) examine the association of RTD with hypoalbuminemia, and (2) assess the prognostic importance of comorbid RTD and hypoalbuminemia in patients with CHF. METHODS AND RESULTS: We measured N-acetyl-ß-D-glucosamidase (NAG) levels and the urinary ß2-microglobulin to creatinine ratio (UBCR) in 456 patients with CHF. RTD was defined as UBCR ≥ 300 µg/g or NAG ≥ 14.2 U/g. There were moderate correlations between RTD markers and serum albumin (NAG, r = -0.428, P < 0.0001; UBCR, r = -0.399, P < 0.0001). Multivariate logistic analysis showed that RTD was significantly related to hypoalbuminemia in patients with CHF. There were 134 cardiac events during a median period of 808 days. The comorbidity of RTD and hypoalbuminemia was increased with advancing New York Heart Association functional class. Multivariate Cox proportional hazard regression analysis showed that the presence of RTD and hypoalbuminemia was associated with cardiac events. The net reclassification index was significantly improved by adding RTD and hypoalbuminemia to the basic risk factors. CONCLUSION: Comorbid RTD and hypoalbuminemia are frequently observed and increase the risk for extremely poor outcome in patients with CHF.

High-mobility group box 1-mediated heat shock protein beta 1 expression attenuates mitochondrial dysfunction and apoptosis.

AIMS: Apoptosis of cardiomyocytes is thought to account for doxorubicin cardiotoxicity as it contributes to loss of myocardial tissue and contractile dysfunction. Given that high-mobility group box 1 (HMGB1) is a nuclear DNA-binding protein capable of inhibiting apoptosis, we aimed to clarify the role of HMGB1 in heat shock protein beta 1 (HSPB1) expression during doxorubicin-induced cardiomyopathy. METHODS AND RESULTS: Mitochondrial damage, cardiomyocyte apoptosis, and cardiac dysfunction after doxorubicin administration were significantly attenuated in mice with cardiac-specific overexpression of HMGB1 (HMGB1-Tg) compared with wild type (WT) -mice. HSPB1 levels after doxorubicin administration were significantly higher in HMGB1-Tg mice than in WT mice. Transfection with HMGB1 increased the expression of HSPB1 at both the protein and mRNA levels, and HMGB1 inhibited mitochondrial dysfunction and apoptosis after exposure of cardiomyocytes to doxorubicin. HSPB1 silencing abrogated the inhibitory effect of HMGB1 on cardiomyocyte apoptosis. Doxorubicin increased the binding of HMGB1 to heat shock factor 2 and enhanced heat shock element promoter activity. Moreover, HMGB1 overexpression greatly enhanced heat shock element promoter activity. Silencing of heat shock factor 2 attenuated HMGB1-dependent HSPB1 expression and abrogated the ability of HMGB1 to suppress cleaved caspase-3 accumulation after doxorubicin stimulation. CONCLUSIONS: We report the first in vivo and in vitro evidence that cardiac HMGB1 increases HSPB1 expression and attenuates cardiomyocyte apoptosis associated with doxorubicin-induced cardiomyopathy. Cardiac HMGB1 increases HSPB1 expression in cardiomyocytes in a heat shock factor 2-dependent manner.

Sarcopenia evaluated by fat-free mass index is an important prognostic factor in patients with chronic heart failure.

BACKGROUND AND AIM: Chronic heart failure (CHF) is a major cause of morbidity and mortality, and cardiac cachexia and sarcopenia are serious complications associated with weight loss and increased catabolism. Fat-free mass index (FFMI) is an indicator of resting energy expenditure and is used for the clinical diagnosis of sarcopenia. In the present study, we investigated the impact of sarcopenia, as evaluated by FFMI, on cardiac prognosis in patients with CHF. METHODS AND RESULTS: We calculated FFMI in 267 CHF patients who were prospectively followed until they died due to cardiac event, or until they were re-hospitalized. Fat-free mass (FFM) was estimated by the formula [FFM (kg)=7.38+0.02908×urinary creatinine (mg/day)] and normalized by the square of the patient's height in meters to calculate FFMI. During the follow-up periods, there were 83 cardiac events, including 19 cardiac deaths. FFMI was lower in patients with cardiac events than in those without (17.0 kg/m(2) vs. 17.6 kg/m(2), P=0.045). Multivariate Cox hazard analysis revealed that decreased FFMI was associated with an unfavorable outcome (adjusted hazard ratio 0.68, 95% confidence interval 0.47-0.98). The patients were divided into two groups according to their median FFMI. The Kaplan-Meier analysis revealed that significantly higher cardiac event rate was observed in the low-FFMI group (log-rank test, P=0.017). CONCLUSIONS: Decreased FFMI was associated with an unfavorable prognosis in patients with CHF.