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Resin glycosides are well known as the purgative ingredients, which are characteristic of convolvulaceous plants. Calystegia hederacea Wall. is a perennial herbaceous vine that is widespread throughout India and East Asia. All parts of this plant are used for the treatment of menoxenia, gonorrhea, etc. Alkaline hydrolysis of the crude resin glycoside fraction of the whole plants of C. hederacea yielded four new glycosidic acids, calyhedic acids A, B, C, and D, along with two known glycosidic acids, calysolic acids A and C, and three known organic acids, 2S-methylbutyric, tiglic, and 2R,3R-nilic acids. Their structures were characterized on the basis of spectroscopic data and chemical evidence. Calyhedic acids A, B, and D were penta-, hexa-, and hepta-glycosides of 12S-hydroxyhexadecanoic acid, respectively, and cayhedic acid C was an isomer of calyhedic acid D, in which the 12S-hydroxyhexadecanoyl residue of calyhedic acid D was replaced by a 11S-hydroxyhexadecanoyl residue. Additionally, cytotoxic activity toward HL-60 human promyelocytic leukemia cells of the crude resin glycoside fraction, the glycosidic acid fraction, calyhedic acid A, and calysolic acid A from C. hederacea was evaluated. Furthermore, to clarify the structure-activity relationship of resin glycosides, the activities of six genuine resin glycosides with calysolic acid A or calysolic acid C as the glycosidic acid, which were isolated from C. soldanella, were examined. Among them, the crude resin glycoside fraction and five genuine resin glycosides with macrolactone structures demonstrated clear cytotoxic activities, while the glycosidic acid fraction, calyhedric acid A, calysolic acid A, and a genuine non-macrolactone-type resin glycoside were either inactive or exhibited weaker activity than the tested macrolactone-type resin glycosides.

Background : Biotin is a water-soluble vitamin that plays various biological roles through histone modification, such as immune functions and fetal growth. Mammalian maternal biotin deficiency during gestation induces fetal growth restriction. Preterm infants are known to be marginal biotin deficiency. However, studies on the biotin status of pregnant women under various conditions are lacking. Method : This was a retrospective case control study to analyze serum biotin concentration during pregnancy and cord blood in normal pregnancy, preterm delivery and small-for-gestational-age (SGA). Results : Twenty pregnant women with normal term delivery, 35 with preterm delivery, 24 with SGA, and 10 non-pregnant adult women were enrolled. Serum biotin concentrations of pregnant women remained low from first to third trimester. The levels of serum biotin in cord blood showed a significant positive correlation with gestational age, and that of pregnant women showed a weak positive correlation with gestational age. The maternal serum biotin levels during second and third trimester of SGA group were significantly lower than those of normal term delivery. Conclusion : This study suggests that maternal biotin deficiency during pregnancy might be the risk of preterm labor or fetal growth restriction. Further studies are required to clarify the roles of biotin in perinatal medicine. J. Med. Invest. 67 : 170-173, February, 2020.

Background : In clinical practice, a large proportion of patients with multiple congenital anomalies and/or intellectual disabilities (MCA / ID) lacks a specific diagnosis. Recently, next-generation sequencing (NGS) has become an efficient strategy for genetic diagnosis of patients with MCA/ID., Objective: To review the utility of NGS for the diagnosis of patients with MCA / ID., Method: Patients with MCA/ID were recruited between 2013 and 2017. Molecular diagnosis was performed using NGS-based targeted panel sequencing for 4,813 genes. Promising causative variants underwent confirmation by Sanger sequencing or chromosomal microarray., Results: Eighteen patients with MCA/ID were enrolled in this study. Of them, 8 cases (44%) were diagnosed by targeted panel sequencing. Most of diagnosed patients were able to receive better counseling and more appropriate medical management., Conclusion: NGS-based targeted panel sequencing seems to be an effective testing strategy for diagnosis of patients with MCA/ID. J. Med. Invest. 67 : 246-249, August, 2020.

Reparative macrophages play an important role in cardiac repair post-myocardial infarction (MI). Bone marrow mononuclear cells (BM-MNCs) have been investigated as a donor for cell therapy but with limited clinical success. These cells, however, may be utilized as a source for reparative macrophages. This translational study aimed to establish a robust in vitro protocol to produce functional reparative macrophages from BM-MNCs and to establish pre-clinical evidence of the efficacy of reparative macrophage transplantation for the treatment of MI. Mouse BM-MNCs were treated with M-CSF plus IL-4, IL-10, TGF-β1 or combinations of these in vitro. The concomitant administration of M-CSF and IL-4 produced the highest rate and largest number of CD11b + F4/80 + CD206 + reparative macrophages. Expression and secretion of tissue repair-related factors including IGF-1, TGF-β1, VEGF and IL1-ra were remarkably enhanced in reparative macrophages compared to BM-MNCs. These cells were transplanted in a mouse MI model, resulting in evident improvement in cardiac function recovery, compared to BM-MNC transplantation. Histological studies showed that reparative macrophage transplantation enhanced myocardial tissue repair including augmented microvascular formation, reduced cardiomyocyte hypertrophy and attenuated interstitial fibrosis. Moreover, survival of reparative macrophages in the heart post-transplantation was increased compared to BM-MNCs. Reparative macrophage transplantation also increased host-derived reparative macrophages in part through TGF-β secretion. In conclusion, concomitant M-CSF + IL-4 treatment effectively produced reparative macrophages from BM-MNCs in vitro. Transplantation of produced reparative macrophage achieved a superior therapeutic efficacy, compared to BM-MNC transplantation, through the enhanced quantity and quality of donor cell engraftment. Further development of this advanced cell-based therapy is warranted.

Mesenchymal stromal/stem cell (MSC)-based therapy is a promising approach for the treatment of heart failure. However, current MSC-delivery methods result in poor donor cell engraftment, limiting the therapeutic efficacy. To address this issue, we introduce here a novel technique, epicardial placement of bi-layered, adhesive dressings incorporating MSCs (MSC-dressing), which can be easily fabricated from a fibrin sealant film and MSC suspension at the site of treatment. The inner layer of the MSC dressing, an MSC-fibrin complex, promptly and firmly adheres to the heart surface without sutures or extra glues. We revealed that fibrin improves the potential of integrated MSCs through amplifying their tissue-repair abilities and activating the Akt/PI3K self-protection pathway. Outer collagen-sheets protect the MSC-fibrin complex from abrasion by surrounding tissues and also facilitates easy handling. As such, the MSC-dressing technique not only improves initial retention and subsequent maintenance of donor MSCs but also augment MSC's reparative functions. As a result, this technique results in enhanced cardiac function recovery with improved myocardial tissue repair in a rat ischemic cardiomyopathy model, compared to the current method. Dose-dependent therapeutic effects by this therapy is also exhibited. This user-friendly, highly-effective bioengineering technique will contribute to future success of MSC-based therapy., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The amount of water drainage from Kushiro Coal Mine in Hokkaido, Japan decreased on the day of all M ≥ 7.5 earthquakes with epicenters within 300 km of the mine during the monitoring period and increased after these earthquakes. This is a valuable finding which would give us a clue to understand pre- and post-seismic rockmass behaviors and contribute for progress in earthquake prediction in future.

Physical inactivity gives rise to numerous diseases and organismal dysfunctions, particularly those related to aging. Musculoskeletal disorders including muscle atrophy, which can result from a sedentary lifestyle, aggravate locomotive malfunction and evoke a vicious circle leading to severe functional disruptions of vital organs such as the brain and cardiovascular system. Although the significance of physical activity is evident, molecular mechanisms behind its beneficial effects are poorly understood. Here, we show that massage-like mechanical interventions modulate immobilization-induced pro-inflammatory responses of macrophages in situ and alleviate muscle atrophy. Local cyclical compression (LCC) on mouse calves, which generates intramuscular pressure waves with amplitude of 50 mmHg, partially restores the myofiber thickness and contracting forces of calf muscles that are decreased by hindlimb immobilization. LCC tempers the increase in the number of cells expressing pro-inflammatory proteins, tumor necrosis factor-α and monocyte chemoattractant protein-1 (MCP-1), including macrophages in situ The reversing effect of LCC on immobilization-induced thinning of myofibers is almost completely nullified when macrophages recruited from circulating blood are depleted by administration of clodronate liposomes. Furthermore, application of pulsatile fluid shear stress, but not hydrostatic pressure, reduces the expression of MCP-1 in macrophages in vitro Together with the LCC-induced movement of intramuscular interstitial fluid detected by µCT analysis, these results suggest that mechanical modulation of macrophage function is involved in physical inactivity-induced muscle atrophy and inflammation. Our findings uncover the implication of mechanosensory function of macrophages in disuse muscle atrophy, thereby opening a new path to develop a novel therapeutic strategy utilizing mechanical interventions., (© 2018 The Author(s).)

Transplantation of mesenchymal stromal cells (MSCs) is a promising new therapy for heart failure. However, the current cell delivery routes result in poor donor cell engraftment. We therefore explored the role of fibrin glue (FG)-aided, instant epicardial placement to enhance the efficacy of MSC-based therapy in a rat ischemic cardiomyopathy model. We identified a feasible and reproducible method to instantly produce a FG-MSC complex directly on the heart surface. This complex exhibited prompt, firm adhesion to the heart, markedly improving initial retention of donor MSCs compared to intramyocardial injection. In addition, maintenance of retained MSCs was enhanced using this method, together contributing the increased donor cell presence. Such increased donor cell quantity using the FG-aided technique led to further improved cardiac function in association with augmented histological myocardial repair, which correlated with upregulation of tissue repair-related genes. We identified that the epicardial layer was eliminated shortly after FG-aided epicardial placement of MSCs, facilitating permeation of the donor MSC's secretome into the myocardium enabling myocardial repair. These data indicate that FG-aided, on-site, instant epicardial placement enhances MSC engraftment, promoting the efficacy of MSC-based therapy for heart failure. Further development of this accessible, advanced MSC-therapy is justified.

Aortic valve regurgitation (AR) is a critical complication during circulatory support with a left ventricular assist device (LVAD). The time-course of AR and related factors, including outflow graft anastomosis site design, were investigated. Twenty-three patients who had continuous-flow LVAD implantation and were supported for more than 6 months were investigated. AR grade (none, 0; trivial, 0.5; mild, 1; mild-moderate, 1.5; moderate, 2; moderate-severe, 2.5; severe, 3) and aortic valve opening were evaluated with echocardiography. Computed tomography was performed to all the patients postoperatively. The angle of the outflow graft to the aorta (O-A angle, parallel 0; tangent 90°, 0-180°), aortic diameter at the anastomosis site, sino-tubular junction (STJ) diameter, distance between the STJ and the anastomosis site, and distance between the anastomosis site and the brachiocephalic artery were measured. The patients' age was 38 ± 11 years. Support duration was 686 ± 354 days. Mean AR grade after continuous-flow LVAD implantation was increased to around mild and was maintained thereafter. No patient needed any intervention to the aortic valve. The aortic valves of 82.6% of patients were closed continuously. The O-A angle (83 ± 14) was positively correlated with maximum AR grade (p = 0.0095). The O-A angle was significantly smaller in patients with maximum AR grade of 1 or less (77 ± 9°) than in those with 1.5 or greater (94 ± 15°, p = 0.021). The other CT measurements had no correlation with AR grade. In conclusion, the O-A angle was correlated with AR grade progression. The O-A angle appears to be one of the important factors related to AR under continuous-flow LVAD support.

Transplantation of mesenchymal stromal cells (MSCs) is an emerging therapy for the treatment of heart failure. However, the delivery method of MSC is currently suboptimal. The use of self-assembling peptide hydrogels, including PuraMatrix ® (PM; 3-D Matrix, Ltd), has been reported for clinical hemostasis and in research models. This study demonstrates the feasibility and efficacy of an advanced approach for MSC-therapy, that is coating of the epicardium with the instantly-produced PM hydrogel incorporating MSCs (epicardial PM-MSC therapy). We optimized the conditions/procedure to produce "instant" 2PM-MSC complexes. After spreading on the epicardium by easy pipetting, the PM-MSC complex promptly and stably adhere to the beating heart. Of note, this treatment achieved more extensive improvement of cardiac function, with greater initial retention and survival of donor MSCs, compared to intramyocardial MSC injection in rat heart failure models. This enhanced efficacy was underpinned by amplified myocardial upregulation of a group of tissue repair-related genes, which led to enhanced repair of the damaged myocardium, i.e. augmented microvascular formation and reduced interstitial fibrosis. These data suggest a potential for epicardial PM-MSC therapy to be a widely-adopted treatment of heart failure. This approach may also be useful for treating diseases in other organs than the heart., (Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Recent research has shown that reparative (alternatively activated or M2) macrophages play a role in repair of damaged tissues, including the infarcted hearts. Administration of IL-4 is known to augment M2 macrophages. This translational study thus aimed to investigate whether IL-4 administration is useful for the treatment of myocardial infarction. Long-acting IL-4 complex (IL-4c; recombinant IL-4 mixed with anti-IL-4 monoclonal antibody as a stabilizer) was administered after coronary artery ligation in mice. It was observed that IL-4c administration increased accumulation of CD206 + F4/80 + M2-like macrophages predominantly in the injured myocardium, compared to the control. Sorted cardiac M2-like macrophages highly expressed wide-ranging tissue repair-related genes. Indeed, IL-4c administration enhanced cardiac function in association with reduced infarct size and enhanced tissue repair (strengthened connective tissue formation, improved microvascular formation and attenuated cardiomyocyte hypertrophy). Experiments using Trib1 -/- mice that had a depleted ability to develop M2 macrophages and other in-vitro studies supported that these IL-4-mediated effects were induced via M2-like macrophages. On the other hand, when administered at Day 28 post-MI, the effects of IL-4c were diminished, suggesting a time-frame for IL-4 treatment to be effective. These data represent proof-of-concept of efficacy of IL-4 treatment for acute myocardial infarction, encouraging its further development.

Oligodendrocyte progenitor cells (OPCs) undergo marked morphological changes to become mature oligodendrocytes, but the metabolic resources for this process have not been fully elucidated. Although lactate, a metabolic derivative of glycogen, has been reported to be consumed in oligodendrocytes as a metabolite, and to ameliorate hypomyelination induced by low glucose conditions, it is not clear about the direct contribution of lactate to cell cycling and differentiation of OPCs, and the source of lactate for remyelination. Therefore, we evaluated the effect of 1,4-dideoxy-1,4-imino-d-arabinitol (DAB), an inhibitor of the glycogen catabolic enzyme glycogen phosphorylase, in a mouse cuprizone model. Cuprizone induced demyelination in the corpus callosum and remyelination occurred after cuprizone treatment ceased. This remyelination was inhibited by the administration of DAB. To further examine whether lactate affects proliferation or differentiation of OPCs, we cultured mouse primary OPC-rich cells and analyzed the effect of lactate. Lactate rescued the slowed cell cycling induced by 0.4 mM glucose, as assessed by the BrdU-positive cell ratio. Lactate also promoted OPC differentiation detected by monitoring the mature oligodendrocyte marker myelin basic protein, in the presence of both 36.6 mM and 0.4 mM glucose. Furthermore, these lactate-mediated effects were suppressed by the reported monocarboxylate transporter inhibitor, α-cyano-4-hydroxy-cinnamate. These results suggest that lactate directly promotes the cell cycling rate and differentiation of OPCs, and that glycogen, one of the sources of lactate, contributes to remyelination in vivo. J. Cell. Physiol. 232: 986-995, 2017. © 2016 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc., (© 2016 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc.)

Spasticity is a frequent chronic complication in individuals with spinal cord injury (SCI). However, the severity of spasticity varies in patients with SCI. Therefore, an evaluation method is needed to determine the severity of spasticity. We used a contusive SCI model that is suitable for clinical translation. In this study, we examined the feasibility of the swimming test and an EMG for evaluating spasticity in a contusive SCI rat model. Sprague-Dawley rats received an injury at the 8th thoracic vertebra. Swimming tests were performed 3 to 6 weeks after SCI induction. We placed the SCI rats into spasticity-strong or spasticity-weak groups based on the frequency of spastic behavior during the swimming test. Subsequently, we recorded the Hoffman reflex (H-reflex) and examined the immunoreactivity of serotonin (5-HT) and its receptor (5-HT2A) in the spinal tissues of the SCI rats. The spasticity-strong group had significantly decreased rate-dependent depression of the H-reflex compared to the spasticity-weak group. The area of 5-HT2A receptor immunoreactivity was significantly increased in the spasticity-strong group. Thus, both electrophysiological and histological evaluations indicate that the spasticity-strong group presented with a more severe upper motor neuron syndrome. We also observed the groups in their cages for 20 hours. Our results suggest that the swimming test provides an accurate evaluation of spasticity in this contusive SCI model. We believe that the swimming test is an effective method for evaluating spastic behaviors and developing treatments targeting spasticity after SCI., Competing Interests: The authors have declared that no competing interests exist.

Pancreatic islets are heterogeneous clusters mainly composed of α and β cells, and these clusters range in diameter from 50 to several hundred micrometers. Native small islets are known to have a higher insulin secretion ability in vitro and to provide better transplantation outcomes when compared with large islets. In this study, we prepared microengineered pseudo-islets from dispersed rat islet cells using precisely-fabricated agarose gel-based microwells with different diameters (100, 300, or 500 μm) to investigate the function and survival of islet cell aggregates with well-controlled sizes. We observed that dead cells were rarely present in the small pseudo-islets with an average diameter of ∼60 μm prepared using 100 μm microwells. In contrast, we observed more dead cells in the larger pseudo-islets prepared using 300 and 500 μm microwells. The relative amount of hypoxic cells was significantly low in the small pseudo-islets whereas a hypoxic condition was present in the core region of the larger pseudo-islets. In addition, we found that the small-sized pseudo-islets reconstituted the in vivo-tissue like arrangement of the α and β cells, and restored the high insulin secretory capacity in response to high glucose. These results clearly suggest that precise size control of pseudo-islets is essential for maintaining islet cell function and survival in vitro. The small-sized pseudo-islets may be advantageous for providing a better therapeutic approach for treating type 1 diabetes mellitus via islet reorganization and transplantation.

Background: Transplantation of allogeneic mesenchymal stromal cells (MSCs) is a promising treatment for heart failure. We have shown that epicardial placement of cell sheets markedly increases donor cell survival and augments therapeutic effects compared with the current methods. Although immune rejection of intramyocardially injected allogeneic MSCs have been suggested, allogeneic MSCs transplanted on the heart surface (virtual space) may undergo different courses. This study aimed to elucidate immunologic response against epicardially placed allogeneic MSCs, rejection or acceptance of these cells, and their therapeutic effects for heart failure., Methods and Results: At 4 weeks after coronary artery ligation, Lewis rats underwent epicardial placement of MSC sheets from syngeneic Lewis or allogeneic Fischer 344 rats or sham treatment. At days 3 and 10 after treatment, similar ratios (≈50% and 30%, respectively) of grafted MSCs survived on the heart surface in both MSC sheet groups. By day 28, survival of syngeneic MSCs was substantially reduced (8.9%); survival of allogeneic MSCs was more extensively reduced (0.2%), suggesting allorejection. Correspondingly, allogeneic MSCs were found to have evoked an immunologic response, albeit low level, as characterized by accumulation of CD4(+) T cells and upregulation of interleukin 6. Despite this alloimmune response, the allogeneic MSC sheet achieved myocardial upregulation of reparative factors, enhanced repair of the failing myocardium, and improved cardiac function to the equivalent degree observed for the syngeneic MSC sheet., Conclusions: Allogeneic MSCs placed on the heart surface evoked an immunologic response; however, this allowed sufficient early phase donor cell survival to induce equivalent therapeutic benefits to syngeneic MSCs. Further development of this approach toward clinical application is warranted., (© 2016 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.)

Human macrophages play important roles in tumor promotion and are called tumor-associated macrophages (TAMs). We previously demonstrated that human esophageal squamous cell carcinomas (ESCCs) contain TAMs and that these TAMs tend to have tumor-supporting features. Here we exposed human macrophages to conditioned media of TE-series human ESCC cell lines (TECMs) to generate an ESCC extracellular stimuli-influenced TAM model. CD14(+) peripheral blood monocytes (PBMos) from healthy donors were treated with M-CSF and with additional IL-4 or TECM exposure. Morphological changes of the cells and the induction of CD163/CD204 proteins were detected in the TECM-exposed model TAMs by immunofluorescence. A software-assisted immunofluorescent cell image analysis showed increased CD163/CD204 positivity in the model TAMs and a weak to moderate positive correlation between the cytoplasmic area and the sum fluorescent intensity of CD204. Morphological changes of the cells were significantly reflected by several cytomorphometric parameters. PBMos were elongated with M-CSF treatment, then enlarged with TECM exposure. The cytoplasmic aspect ratio was decreased by M-CSF treatment and slightly increased by TECM exposure. The nuclear-cytoplasmic ratio decreased during the whole process of cell differentiation. This system is useful for quantitative assessments of TAM-like morphological changes of macrophages and the induction of CD163/CD204 in a model ESCC microenvironment., (© 2016 Japanese Society of Pathology and John Wiley & Sons Australia, Ltd .)

Objectives: Ideal alternatives for replacing native arteries, which have biocompatibility such as growth potential, anti-thrombogenesis and durability, have yet to be discovered. We previously demonstrated the utility of tissue-engineered vascular autografts; however, the use of these autografts is limited to low-pressure conditions. The aim of this study was to create the tissue-engineered arterial patch (TEAP) that could be used in high-pressure systems, and to evaluate the maturation in this regenerative tissue., Methods: We developed a new biodegradable polymer scaffold, which is composed of a co-polymer of epsilon-caprolactone and lactide acid [P(CL/LA)] and a poly-L-lactide acid (PLLA). To obtain mechanical strength, we modified PLLA that is degraded by hydrolysis for 1-2 years in contrast to polyglycolic acid in our low-pressure study previously. We implanted an oval-shaped patch (30 × 15 mm) of this polymer without cell seeding into the descending aorta of 12 dogs, and followed the animals for 1, 3 and 6 months (n = 4 in each group). The cell proliferation in the patch was evaluated with histological and immunohistochemical methods. Additionally, the expression of vascular endothelial growth factor (VEGF) and smooth muscle myosin heavy chain (smMHC) mRNA in the patches was determined with reverse transcriptase-polymerase chain reaction., Results: Macroscopically, there was no incidence of rupture or aneurysmal formation on the patch. The luminal surface of the TEAP was covered with a single layer of endothelial cells stained with vWF immunohistochemically at 1 month after implantation. αSMA-positive cells that indicated smooth muscle cells and collagen fibres were observed in the patch and they increased over time. The VEGF mRNA expression in the patch at 1 month was significantly higher than that of native arterial tissue (1 month; 0.124 ± 0.017 ng/µl, native; 0.009 ± 0.003 ng/µl, P < 0.05). The smMHC mRNA expression gradually increased, and reached ∼ 60% of that of the native artery at 6 months (6 months: 0.351 ± 0.028 ng/µl, native: 0.540 ± 0.027 ng/µl)., Conclusions: We demonstrated the maturation of endothelial and smooth muscle cells in TEAP, suggesting that this biodegradable polymer scaffold could be used as an alternative vascular material even in high-pressure systems., (© The Author 2015. Published by Oxford University Press on behalf of the European Association for Cardio-Thoracic Surgery. All rights reserved.)

We provided a left ventricular assist device (LVAD) for a 22-year-old man with congenital L-transposition of the great arteries after anatomic repair at the age of 7 years. He was hospitalized for progressive low-output syndrome caused by intractable biventricular failure. He received LVAD in his morphologic left ventricle with a concomitant pulmonary valve replacement. After the surgery, critical multiorgan failure with severe right heart failure occurred. It took three postoperative months to normalize all organ function following improvement of morphologic right ventricular function. He has remained stable with LVAD support for 1.5 years., (Copyright © 2014 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

The algorithm for the physiological control provided by left ventricular assist devices (LVADs) has been controversial. In particular, little is known about the physiological control algorithm (such as for achieving physiological circadian rhythms) in continuous-flow LVADs. To investigate the existence of circadian variation, we retrospectively evaluated the LVAD flow-correlated motor current of patients supported by continuous-flow LVADs. The motor current and the pump speed were collected from the external controller every 10 min after device implantation, and the data were divided for every 30-day period, which began on midnight on the first post-operative day. The subjects were 18 patients (mean age 37.7, mean body surface area 1.71 m(2) at the time of operation) with dilated cardiomyopathy or dilated phase of hypertrophic cardiomyopathy. As of August 1, 2013, the patients' median support duration was 889 days. The mean calculated dominant period of motor current variation was 24.0 h and the mean amplitude was 11.7 mA for the entire duration. The amplitude of the motor current circadian variation tended to be increased until around the fifth month. The motor current had a tendency to be relatively low during the night time and high during the day time. A significant difference was found between the night-time and day-time mean motor current for the entire duration (p < 0.05). In conclusion, the circadian variation of the motor current could be observed over long term in patients with fixed rotation speed continuous-flow LVAD support.

Background: The EVAHEART left ventricular assist device was approved in 2010 by the Japanese Pharmaceuticals and Medical Devices Agency (PMDA) for bridge to heart transplantation (BTT). However, its effectiveness has not been evaluated since approval. In this study we evaluated the EVAHEART device in a commercial setting in Japan., Methods: Ninety-six consecutive patients enrolled in the Japanese Registry for Mechanically Assisted Circulatory Support (J-MACS), who were listed for transplant or likely to be listed and who received an EVAHEART device, were enrolled from 2011 to 2013 at 14 Japanese centers. Patients' survival rates, adverse events and quality-of-life data were obtained from the J-MACS Registry., Results: Patients' median age was 43 years (85% male). The Interagency Registry for Mechanically Assisted Circulatory Support profiles revealed 12 patients in Level 1, 45 in Level 2, 37 in Level 3 and 1 in Level 4. The mean support duration was 384.7 days, with a cumulative duration of 101.2 years. The Kaplan-Meier survival rate during support was 93.4% at 6 months, 87.4% at 1 year and 87.4% at 2 years. Seventy-seven patients (80.2%) currently remain on support, 7 received a transplant and 10 died during support. Major adverse events included drive-line infection (14.6%) and neurologic events such as ischemic stroke (17.7%), hemorrhage (13.5%), transient ischemic attack (3.1%), pump thrombosis (1%) and hemolysis (1%). There was no gastrointestinal (GI) bleeding or right heart failure requiring right ventricular assist device (RVAD). There was no pump exchange due to mechanical failure., Conclusions: The EVAHEART device provides safe, reliable and long-term circulatory support with improved survival in commercial settings of BTT in Japan, where the transplant waiting period is much longer. Incidences of GI bleeding, hemolysis, right ventricular failure, device thrombosis and mechanical failure were extremely rare in patients on EVAHEART devices., (Copyright © 2014 International Society for Heart and Lung Transplantation. Published by Elsevier Inc. All rights reserved.)

Thrombosis was detected in the right ventricle 7 days after a bidirectional Glenn operation in a 1-year-old boy with pulmonary atresia with intact ventricular septum (PAIVS) and immediately before pulmonary valvuloplasty in a 21-day-old boy with PAIVS. The risk of thrombosis and the blood clotting cascade should be assessed by blood tests, and surveillance echocardiography should be frequently applied in order to diagnose thrombosis early in such patients.

Background: Secure distal anastomosis and reliable brain protection are indispensable for successful total arch replacement (TAR). In 2002, we introduced a modified elephant trunk technique, a novel approach to distal anastomosis, and employed antegrade selective cerebral perfusion. We retrospectively analyzed 107 consecutive patients to evaluate the efficacy of this technique for TAR with antegrade selective cerebral perfusion., Methods: Since 2002 we have employed moderate hypothermic circulatory arrest, selective antegrade cerebral perfusion, and open distal anastomosis with a modified elephant trunk technique in TAR. Between February 2002 and September 2011, 107 TARs were performed in 88 males and 19 females (age, 33 to 88 years; mean, 70.9±9.5 years). Etiologies of cases were as follows: 89 true aneurysm due to atherosclerosis; 5 infectious aneurysm; 1 aortic dilation with bicuspid aortic valve; 12 aortic dissection, including 1 of acute aortic dissection case; and 2 Marfan syndrome. Concomitant procedures included 19 coronary artery bypass grafting (CABG) cases, 2 aortic valve replacement cases, 1 mitral valve plasty case, 1 Bentall procedure case, and 1 case of Bentall with CABG., Results: The operative mortality within 30 days was 0.9% (1 of 107), and overall hospital mortality was 1.9% (2 of 107). Temporary and permanent neurologic dysfunction occurred in 5 patients each (4.7%). The Kaplan-Meier survival analysis revealed a 5-year survival rate of 91.8%., Conclusions: The modified elephant trunk technique using selective antegrade cerebral perfusion provided secure distal anastomosis and demonstrated excellent results, with low operative mortality and few neurologic complications., (Copyright © 2014 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

In Japan, Japanese cedar pollen dispersal is one of the major causes of pollinosis. Sydowia japonica is an ascomycetous fungus that grows exclusively on the male strobili of Japanese cedar, suggesting a possible mechanism for controlling pollen dispersal. To evaluate this possibility, eleven isolates of S. japonica were collected from around Japan and used as an inoculum to male strobili of Japanese cedar. The treatment demonstrated that the fungus infected only the pollen and prevented pollen dispersal. The fungus did not cause any additional symptoms to other parts of Japanese cedar, such as needles, stems, and buds. All S. japonica isolates collected around Japan could serve to control pollen dispersal. Periodic observation of the fungal pathogenesis with stereomicroscope and scanning electron microscope showed that hyphal fragments and conidia of S. japonica germinated on the surface of male strobili, and the germ tube entered pollen sacs through opening microsporophylls. Within the pollen sacs, the hyphae penetrated pollen gradually, such that all pollen was infected by the fungus by approximately one month before the pollen dispersal season. The infected pollen was destroyed due to the fungal infection and was never released. Our data suggests a novel approach of preventing pollen dispersal using pollen-specific fungal infection.

Novel 4-substituted 2-pyridin-2-ylamides were developed using in-silico ligand-based drug design (LBDD) in an attempt to identify inhibitors of SH2-containing 5'-inositol phosphatase 2 (SHIP2), which is implicated in insulin-resistant type 2 diabetes. Among the compounds synthesized, N-[4-(4-chlorobenzyloxy)pyridin-2-yl]-2-(2,6-difluorophenyl)- acetamide (CPDA, 4a) was identified as a potent SHIP2 inhibitor. CPDA was found to enhance in vitro insulin signaling through the Akt pathway more efficiently than the previously reported SHIP2 inhibitor AS1949490, and ameliorated abnormal glucose metabolism in diabetic (db/db) mice., (Copyright © 2013 Elsevier Masson SAS. All rights reserved.)

We report a case of a 59-year-old woman who presented with hypovolemic shock and compensated acidosis (preoperative arterial blood gases: pH 7.3, P(CO2) 31.9 mmHg, Pa(O2) 112.3 mmHg, base excess -9.8, Hb 6.4 g x dl(-1)) due to perforated descending colon, necessitating emergency surgery. Tracheal intubation had been performed preoperatively. Prior to induction of anesthesia, blood pressure was 106/74 mmHg, heart rate 119 beats x min(-1), and Sp(O2) 100% breathing room air. Anesthesia was induced with remifentanil influsion at a rate of 0.05 mg x kg(-1) x min(-1), sevoflurane 1% and rocuronium bromide 30mg, and was maintained with oxygen, air, remifentanil and sevoflurane. For a critical hypovolemia, in accordance to the guidelines for intraoperative critical hemorrhage and the Japanese practical guidelines for blood components therapy, we started to transfuse incompatible red cell (O+) since the identification of blood typing was suspended. The duration of surgery was 104 min, with an intraoperative total bleeding of 125 ml. Four units of total blood transfusion and 3,050 ml of infusion of Ringer's acetate solution were administered. The patient was transferred to ICU with tracheal intubation. No adverse reactions associated with blood type incompatibility were recognized.

A rare case requiring replacement of an intact Smeloff-Cutter ball prosthesis in the mitral position 40 years after implantation is presented. The Smeloff-Cutter ball valve prosthesis was designed to have two open cages. It has two potential advantages: a relatively large, effective orifice area and its self-washing effect that prevents thrombus formation. There have been only a few reports of survivors with ball valve prostheses in place for more than three decades especially in the mitral position. This is a valuable report describing the long-term durability of a Smeloff-Cutter ball valve prosthesis in the mitral position.

Background: In Japan, malignant hyperthermia susceptibility (MHS) is diagnosed by detecting an enhanced rate of CICR from the sarcoplasmic reticulum in skinned skeletal muscle fibers. We usually consider the CICR test should be completed within 48 hours of muscle excision. We measured the viability period of the CICR test in the following four treatment groups., Methods: Muscle bundles prepared from a rabbit were dissected and stored in muscle relaxant solution at -30 degrees C, 4 degrees C, 20 degrees C, and 37 degrees C. The CICR rate was measured in a part of the biopsied muscle using chemically skinned fibers that had been prepared according to Endo's method., Results: With muscle bundles stored at -30 degrees C and 37 degrees C, the rate of CICR could not be measured. Function related CICR of the muscle stored at 4 degrees C and 20 degrees C lasted for four days., Conclusions: This results showed that rate of CICR can be measured for at least 4 days when skeletal muscle bundles of rabbits are stored both at 4 degrees C and at 20 degrees C. Further studies are expected including the optimum conditions on stored temperature and stored days in the human muscle.

Impairment of insulin and IGF-I signaling in the brain is one of the causes of dementia associated with diabetes mellitus and Alzheimer's disease. However, the precise pathological processes are largely unknown. In the present study, we found that SH2-containing inositol 5'-phosphatase 2 (SHIP2), a negative regulator of phosphatidylinositol 3,4,5-trisphosphate-mediated signals, is widely expressed in adult mouse brain. When a dominant-negative mutant of SHIP2 was expressed in cultured neurons, insulin signaling was augmented, indicating physiological significance of endogenous SHIP2 in neurons. Interestingly, SHIP2 mRNA and protein expression levels were significantly increased in the brain of type 2 diabetic db/db mice. To investigate the impact of increased expression of SHIP2 in the brain, we further employed transgenic mice overexpressing SHIP2 and found that increased amounts of SHIP2 induced the disruption of insulin/IGF-I signaling through Akt. Neuroprotective effects of insulin and IGF-I were significantly attenuated in cultured cerebellar granule neurons from SHIP2 transgenic mice. Consistently, terminal deoxynucleotide transferase-mediated dUTP nick end labeling assay demonstrated that the number of apoptosis-positive cells was increased in cerebral cortex of the transgenic mice at an elderly age. Furthermore, SHIP2 transgenic mice exhibited impaired memory performance in the Morris water maze, step-through passive avoidance, and novel-object-recognition tests. Importantly, inhibition of SHIP2 ameliorated the impairment of hippocampal synaptic plasticity and memory formation in db/db mice. These results suggest that SHIP2 is a potent negative regulator of insulin/IGF-I actions in the brain, and excess amounts of SHIP2 may be related, at least in part, to brain dysfunction in insulin resistance with type 2 diabetes.

The preparation of a hybrid of graphene oxide and per-6-amino-beta-cyclodextrin is reported. The hybrid formed a supramolecular graphene oxide polymer network upon addition of guest dimer.

Bisphenol A (BPA) is a monomer used in manufacturing a wide range of chemical products, including epoxy resins and polycarbonate. BPA, an important endocrine disrupting chemical that exerts estrogen-like activities, is detectable at nanomolar levels in human serum worldwide. The pregnancy associated doses of 17beta-estradiol (E2) plus tumor-necrosis factor-alpha (TNF-alpha) induce distorted maturation of human dendritic cells (DCs) that result in an increased capacity to induce T helper (Th) 2 responses. The current study demonstrated that the presence of BPA during DC maturation influences the function of human DCs, thereby polarizing the subsequent Th response. In the presence of TNF-alpha, BPA treatment enhanced the expression of CC chemokine ligand 1 (CCL1) in DCs. In addition, DCs exposed to BPA/TNF-alpha produced higher levels of IL-10 relative to those of IL-12p70 on CD40 ligation, and preferentially induced Th2 deviation. BPA exerts the same effect with E2 at the same dose (0.01-0.1 microM) with regard to DC-mediated Th2 polarization. These findings imply that DCs exposed to BPA will provide one of the initial signals driving the development and perpetuation of Th2-dominated immune response in allergic reactions.

Objective: The development of a tissue-engineered vascular graft with the ability to grow and remodel holds promise for advancing cardiac surgery. In 2001, we began a human trial evaluating these grafts in patients with single ventricle physiology. We report the late clinical and radiologic surveillance of a patient cohort that underwent implantation of tissue-engineered vascular grafts as extracardiac cavopulmonary conduits., Methods: Autologous bone marrow was obtained and the mononuclear cell component was collected. Mononuclear cells were seeded onto a biodegradable scaffold composed of polyglycolic acid and epsilon-caprolactone/L-lactide and implanted as extracardiac cavopulmonary conduits in patients with single ventricle physiology. Patients were followed up by postoperative clinic visits and by telephone. Additionally, ultrasonography, angiography, computed tomography, and magnetic resonance imaging were used for postoperative graft surveillance., Results: Twenty-five grafts were implanted (median patient age, 5.5 years). There was no graft-related mortality (mean follow-up, 5.8 years). There was no evidence of aneurysm formation, graft rupture, graft infection, or ectopic calcification. One patient had a partial mural thrombosis that was successfully treated with warfarin. Four patients had graft stenosis and underwent successful percutaneous angioplasty., Conclusion: Tissue-engineered vascular grafts can be used as conduits in patients with single ventricle physiology. Graft stenosis is the primary mode of graft failure. Further follow-up and investigation for the mechanism of stenosis are warranted., (Published by Mosby, Inc.)

Forest gap dynamics affects light intensity on the forest floor, which in turn may influence defense and survival of tree seedlings. Current-year Fagus crenata seedlings show high mortality under the canopy caused by damping-off. In contrast, they survive pathogen attacks in gaps. However, defense mechanisms against damping-off have not been fully understood. In order to determine the resistance factors that affect mortality in current-year seedlings, we compared seedling survival and chemical and histological characteristics of the hypocotyls of seedlings from closed-stand and forest-edge plots. Damping-off occurred in the current-year seedlings mainly from the end of June to July; survival rate of the seedlings was higher in the forest-edge plot than in the closed-stand plot. By performing an inoculation test on the seedling hypocotyls, we identified Colletotrichum dematium and Cylindrocarpon sp. as the causative pathogens under low illumination only. In the beginning of July, only seedling hypocotyls from the forest-edge plot exhibited periderm formation. From mid-June to July, seedling hypocotyls from the forest-edge plot accumulated approximately twice the amount of total phenols as those accumulated by seedling hypocotyls from the closed-stand plot. The ethyl acetate phase of methanol extracts of hypocotyls showed antifungal activity. We conclude that seedlings from the forest-edge plot may resist pathogenic attack via periderm formation and increased phenol synthesis. Plant defense mechanisms that are controlled by light intensity may be important for promoting seedling regeneration in forest gap dynamics.

CD1d-restricted invariant NKT (iNKT) cells play crucial roles in various types of immune responses, including autoimmune diseases, infectious diseases and tumor surveillance. The mechanisms underlying their adjuvant functions are well understood. Nevertheless, although IL-4 and IL-10 production characterize iNKT cells able to prevent or ameliorate some autoimmune diseases and inflammatory conditions, the precise mechanisms by which iNKT cells exert immune regulatory function remain elusive. This study demonstrates that the activation of human iNKT cells by their specific ligand alpha-galactosylceramide enhances IL-12p70 while inhibiting the IL-23 production by monocyte-derived dendritic cells, and in turn down-regulating the IL-17 production by memory CD4(+) Th cells. The ability of the iNKT cells to regulate the differential production of IL-12p70/IL-23 is mainly mediated by a remarkable hallmark of their function to produce both Th1 and Th2 cytokines. In particular, the down-regulation of IL-23 is markedly associated with a production of IL-4 and IL-10 from iNKT cells. Moreover, Th2 cytokines, such as IL-4 and IL-13 play a crucial role in defining the biased production of IL-12p70/IL-23 by enhancement of IL-12p70 in synergy with IFN-gamma, whereas inhibition of the IFN-gamma-promoted IL-23 production. Collectively, the results suggest that iNKT cells modify the IL-12p70/IL-23 balance to enhance the IL-12p70-induced cell-mediated immunity and suppress the IL-23-dependent inflammatory pathologies. These results may account for the long-appreciated contrasting beneficial and adverse consequence of ligand activation of iNKT cells.

Background: We have recently found that exposure to acute restraint stress suppresses antigen-specific antibody production, including IgE, in a murine model of allergic rhinitis. Although age-related alterations in immune responses are known, it remains unclear whether aging modulates the antibody production under stressful conditions. In this study, we set out to determine the effects of aging on antibody production under acute restraint stress in mice., Methods: Both young and aged CBA/J mice were repeatedly sensitized intranasally with phospholipase A2 (PLA2) without adjuvants. Restraint stress was applied using uniform cylinders once a week for a continuous 8h period, on 5 occasions in total. Blood samples were taken at 0, 20 and 30 days after primary sensitization, and production of PLA2-specific antibodies and levels of IL-4, IFN-gamma, IL-10 and IL-1 beta in sera were determined by ELISA., Results: Repeated intranasal sensitization with PLA2 induced PLA2-specific IgE, IgG1 and IgG2a production in aged mice. We found that exposure to restraint stress significantly inhibited production of PLA2-specific IgE, IgG1 and IgG2a in aged mice. In addition, antibody production under restraint stress decreased significantly in aged mice when compared with young mice. No IL-4, IFN-gamma, IL-10 or IL-1 beta were detected in sera from non-stressed or stressed aged mice., Conclusions: Aging exacerbates the immunosuppressive role of acute restraint stress in antigen-specific antibody production in mice.

Background: Although BCG has been found to impart protection against leprosy in many populations, the utility of repeat or booster BCG vaccinations is still unclear. When a policy of giving a second BCG dose to school children in Brazil was introduced, a trial was conducted to assess its impact against tuberculosis, and a leprosy component was then undertaken in parallel., Objective: to estimate the protection against leprosy imparted by a second dose of BCG given to schoolchildren., Methods and Findings: This is a cluster randomised community trial, with 6 years and 8 months of follow-up., Study Site: City of Manaus, Amazon region, a leprosy-endemic area in Brazil., Participants: 99,770 school children with neonatal BCG (aged 7-14 years at baseline), of whom 42,662 were in the intervention arm (revaccination)., Intervention: BCG given by intradermal injection., Main Outcome: Leprosy (all clinical forms)., Results: The incidence rate ratio of leprosy in the intervention over the control arm within the follow-up, in schoolchildren with neonatal BCG, controlled for potential confounders and adjusted for clustering, was 0.99 (95% confidence interval: 0.68 to 1.45)., Conclusions/significance: There was no evidence of protection conferred by the second dose of BCG vaccination in school children against leprosy during the trial follow-up. These results point to a need to consider the effectiveness of the current policy of BCG vaccination of contacts of leprosy cases in Brazilian Amazon region.

Ryanodine receptor (RyR) is a Ca(2+) channel that mediates Ca(2+) release from intracellular stores. Altered Ca(2+) homeostasis in skeletal muscle which usually occurs as a result of point mutations in type 1 RyR1 (RyR1) is a key molecular event triggering malignant hyperthermia (MH). There are three RyR isoforms, and we herein show, for the first time, that human dendritic cells (DCs) preferentially express RyR1 mRNA among them. The RyR activator, 4-chloro-m-cresol (4CmC), induced Ca(2+) release in DCs, and this response was eliminated by dantrolene, an inhibitor of the RyR1, and was unaffected by xestospongin C, a selective inhibitor of IP(3) receptor. Activation of RyR1 reduced LPS-induced IL-10 production, promoted the expression of HLA-DR and CD86, and thereby exhibited an improved capacity to stimulate allogeneic T cells. These findings demonstrate that RyR1-mediated calcium signaling modifies diverse DC responses and suggest the feasibility of using DC preparations for the diagnosis of MH.

Three Japanese patients demonstrated plasma prekallikrein (PK) deficiency (PKD) after an examination of the proband family line named 'PKD Seki'. A molecular genetic analysis of these PK genes showed homozygous amino acid substitutions Gly104Arg and Asn124Ser in exon 5, which encodes part of the apple domain 2 (A2) of the heavy chain. This is the first case involving substitutions in the heavy chain of the PK gene which affected blood coagulation. Because the apple domains of PK bind to the C-terminal domain (D6(H)) of high-molecular weight kininogen (HMWK), the two substitutions in A2 may therefore be the main cause of PKD Seki. We subsequently investigated the effects of amino acid substitutions in A2 to elucidate the binding activity of PK to HMWK using mutant A2 proteins produced in Escherichia coli. We clearly demonstrated that the Gly104Arg-substitution with the Asn124Ser-substitution in A2 reduce the binding activity of A2 to HMWK. PKD Seki is the first significant case to show the amino acid substitutions in the A2 affecting the binding capacity of PK with HMWK. Our findings therefore suggest that the binding of PK to HMWK may play a crucial role in the first step of blood coagulation.

Objective: This study was undertaken to compare long-term results of various types of surgical repairs for either congenitally corrected transposition of the great arteries or double outlet right ventricle with discordant atrioventricular connections, and to analyze the risk factors that affect early and late mortality and reintervention., Methods: Between January 1972 and September 2005, a total of 189 patients (median age 8.3 years, range 2 months to 47 years old) with congenitally corrected transposition of the great arteries or double outlet right ventricle with discordant atrioventricular connections underwent definitive repairs. The definitive repairs comprised a conventional repair (atrial septal defect, or ventricular septal defect closure with or without pulmonary stenosis release, or isolated tricuspid valve surgery) in 36 patients (group I), conventional Rastelli in 31 patients (group II), double-switch operation (atrial switch plus arterial switch) in 15 patients (group III), atrial switch plus intraventricular rerouting (with or without extracardiac conduits) in 69 patients (group IV), and a Fontan-type repair in 38 patients (group V). The mean follow-up period was 10.1 years. Hospitalization and late mortality and reoperation were indicated as events. Risk factors for these events were analyzed by logistic regression for hospital death and a Cox proportional hazards model for late events., Results: The Kaplan-Meier survival including hospital and late mortality was 62.4% at 32 years in group I, 78.5% at 27 years in group II, 74.5% at 15 years in group III, 80% at 16 years in group IV, and 79.3% at 22 years in group V. The reoperation-free ratio was 64.2% in group I, 76.6% in group II, 84.4% in group III, 89.6% in group IV, and 91.3% in group V. Risk analyses showed that the risk for hospital death was preoperative in patients with more than moderate tricuspid regurgitation and a cardiopulmonary bypass time of more than 240 minutes. A risk for late mortality was the presence of tricuspid regurgitation. Risks for reoperation were preoperative cardiomegaly, preoperative tricuspid regurgitation of more than grade II, ventricular septal defect enlargement, and body weight less than 10 kg. Risks for pacemaker implantation, as indicated by multivariate analysis, were ventricular septal defect enlargement during operation and age less than 3 years., Conclusions: There were no statistical differences between long-term survival rates of patients who underwent conventional surgical repair versus those of patients who underwent anatomic surgical repair. Results of conventional repair were satisfactory except in patients with significant tricuspid regurgitation. Results of anatomic repair were also satisfactory even for patients with significant tricuspid regurgitation, and therefore, anatomic repair should be the procedure of choice for those patients.

A 79-year-old man was scheduled to undergo transurethral resection of the hypertrophied prostate during general anesthesia. Anesthesia was maintained with sevoflurane 0.5-1.0% and nitrous oxide 50% in oxygen mixture. Immediately after using an evacuator to remove small resected pieces of the prostate, end-tidal carbon dioxide pressure (EtCO2) decreased suddenly from 31 mmHg to 18 mmHg. Concurrently, decreases in systolic blood pressure from 110 mmHg to 60 mmHg and oxygen saturation from 100% to 95%, and an increase in arterial-alveolar carbon dioxide tension difference (a-ADCO2) were observed. We initially suspected the onset of pulmonary thrombosis as the cause of these events, ventilated the patient's lungs with 100% oxygen and administered 5,000 units of heparine and vasopressors intravenously. However, transesophageal echocardiography done subsequently revealed the presence of strongly echogenic images compatible with the air in the left cardiac atrium and the contribution of the air to those events. The EtCO2, blood pressure, and oxygen saturation improved in about 20 minutes after the initial decrease of the EtCO2 had been detected. Thereafter, surgical procedure was done uneventfully. In routine anesthetic management of transurethral surgery, it should be considered that the sudden deterioration in vital signs may have been caused by evacuator used.

We synthesized 9alpha-fluoromedroxyprogesterone acetate (FMPA) in order to test whether it is a more potent anti-angiogenic agent than medroxyprogesterone acetate (MPA), which has been widely used as a therapeutic agent for breast and endometrium cancers. FMPA was previously synthesized in 10 steps (total yield: 1%). An efficient synthesis of FMPA has been achieved in 6 steps (total yield: 12%). We examined the anti-tumor effect of FMPA, complexed with dimethyl-beta-cyclodextrin (DM-beta-CyD), on rat mammary carcinomas induced by 7,12-dimethylbenz[a]anthracene (DMBA). FMPA showed great anti-tumor effect on DMBA-induced rat mammary carcinomas.

Ryanodine receptor 1 (RYR1) gene mutations are associated with central core disease (CCD), multiminicore disease (MmD) and malignant hyperthermia (MH), and have been reported to be responsible for 47-67% of patients with CCD and rare cases with MmD. However, to date, the true frequency and distribution of the mutations along the RYR1 gene have not been determined yet, since mutation screening has been limited to three 'hot spots', with particular attention to the C-terminal region. In this study, 27 unrelated Japanese CCD patients were included. Clinical histories and muscle biopsies were carefully reviewed. We sequenced all the 106 exons encoding RYR1 with their flanking exon-intron boundaries, and identified 20 novel and 3 previously reported heterozygous missense mutations in 25 of the 27 CCD patients (93%), which is a much higher mutation detection rate than that perceived previously. Among them, six were located outside the known 'hot spots'. Sixteen of 27 (59%) CCD patients had mutations in the C-terminal 'hot spot'. Three CCD patients had a probable autosomal recessive disease with two heterozygous mutations. Patients with C-terminal mutations had earlier onset and rather consistent muscle pathology characterized by the presence of distinct cores in almost all type 1 fibres, interstitial fibrosis and type 2 fibre deficiency. In contrast, patients with mutations outside the C-terminal region had milder clinical phenotype and harbour more atypical cores in their muscle fibres. We also sequenced two genes encoding RYR1-associated proteins as candidate causative genes for CCD: the 12 kD FK506-binding protein (FKBP12) and the alpha1 subunit of L-type voltage-dependent calcium channel or dihydropyridine receptor (CACNA1S). However, no mutation was found, suggesting that these genes may not, or only rarely, be responsible for CCD. Our results indicate that CCD may be caused by RYR1 mutations in the majority of patients.

Background: Malignant hyperthermia (MH) is a disorder of calcium homeostasis in skeletal muscle triggered by volatile anesthetics or succinylcholine in susceptible persons. More than 100 mutations in the ryanodine receptor type 1 gene (RYR1) have been associated with MH susceptibility, central core disease, or both. RYR1 mutations may account for up to 70% of MH-susceptible cases. The authors aimed to determine the frequency and distribution of RYR1 mutations in the Japanese MH-susceptible population., Methods: The authors selected 58 unrelated Japanese diagnosed as MH-susceptible for having an enhanced Ca-induced Ca release rate from the sarcoplasmic reticulum on chemically skinned muscle fibers. They sequenced the entire RYR1 coding region from genomic DNA. Muscle pathology was also characterized., Results: Seven previously reported and 26 unknown RYR1 potentially pathogenic sequence variations were identified in 33 patients (56.9%). Of these patients, 48% had cores on muscle biopsy. The mutation detection rate was higher in patients with clear enhancement of Ca-induced Ca release rate (72.4%), whereas all patients with central core disease had RYR1 mutations. Six patients harbored potentially causative compound heterozygous sequence variations., Conclusions: Distribution and frequency of RYR1 mutations differed markedly from those of the North American and European MH-susceptible population. Comprehensive screening of the RYR1 gene is recommended for molecular investigations in MH-susceptible individuals, because many mutations are located outside the "hot spots." Based on the observed occurrence of compound heterozygous state, the prevalence of a possibly predisposing phenotype in the Japanese population might be as high as 1 in 2,000 people.

Local anesthetics affect intracellular Ca2+ movement in the myocyte. The use of isomers may help to reveal specific mechanisms of action, such as receptor mediation. In the present study, we used skinned fibers from mammalian skeletal muscle to test whether bupivacaine enantiomers had different effects on Ca2+ release and uptake by the sarcoplasmic reticulum, and on the Ca2+ sensitivity of the contractile system. Ca2+-induced Ca2+ release was enhanced by S-bupivacaine 1 approximately 3 mM, but inhibited by R-bupivacaine 3 mM, remaining unaffected at lower doses. These enantiomers inhibited Ca2+ uptake to different degrees, with R-bupivacaine having a stronger effect. Ca2+ sensitivity of the contractile system was equally enhanced by R- and S-bupivacaine. These findings might help to explain the myoplasmic Ca2+ elevation induced by bupivacaine. The observed stereoselectivity suggests effects on specific proteins, the ryanodine Ry1 receptor and the Ca2+-ATPase pump, rather than non-specific increase in Ca2+ permeability.

Background: Exposure to acute stressors modulates both innate and acquired immune function. However, little is known about whether stress has the potential to modulate the pathogenesis of allergic rhinitis., Objectives: To determine the effects of acute restraint stress on the initiation of allergic rhinitis in a murine model., Methods: CBA/J mice were repeatedly intranasally sensitized with phospholipase A2 (PLA2) from honeybee venom without adjuvant. Restraint stress was applied using uniform cylinders once a week for a continuous 8-hour period, on five occasions in total. Production of PLA2-specific antibodies and degree of nasal and blood eosinophilia were compared between stressed and control mice., Results: Repeated intranasal sensitization with PLA2 induced PLA2-specific IgE and marked eosinophilia in both the nose and blood in CBA/J mice. Exposure to restraint stress significantly inhibited production of PLA2-specific IgE, IgG1 and IgG2a. Conversely, the stress exerted no significant effect on eosinophilia., Conclusions: Exposure to acute restraint stress inhibits antigen-specific antibody production, but not local or systemic eosinophilia. The results of this study suggest that acute stress has the potential to modulate the initiation of allergic rhinitis., (Copyright (c) 2005 S. Karger AG, Basel.)