Search

Purpose: To investigate walking ability in Japanese patients with Rett syndrome (RTT)., Methods: Walking ability was assessed in 100 female Japanese patients with RTT using univariate and multivariate analysis in all age groups, and in patients over 10 years of age. We analyzed walking ability and confounding factors including prenatal-perinatal histories, developmental milestones, somatic and head growth, anthropometric data, body mass index, age of loss of purposeful hand use, age at onset of stereotypic hand movement, history of autistic behavior, age at regression, presence or absence of seizures, and the results of MECP2 genetic examination from the Japanese Rett syndrome database., Results: Univariate analysis revealed that acquisition of walking in all age groups was significantly correlated with the acquisition of meaningful words, microcephaly, and crawling (P < 0.0001, P = 0.005, P < 0.0001, respectively). Univariate analysis revealed that walking ability over 10 years of age was significantly correlated with acquisition of meaningful words, microcephaly, and body mass index (P < 0,0001, P = 0.005, P = 0.0018, respectively). MECP2 mutations R306C, R133C, and R294X were significantly associated with different acquisition of crawling (P = 0.004) and walking (P = 0.01). Multivariate analysis revealed that only acquisition of meaningful words was significantly correlated with walking ability over 10 years of age. This trend excluded the genetic effects of R306C, R133C, and R294X., Conclusions: Meaningful word acquisition was robustly associated with walking ability over 10 years. Prognosis of walking ability may be predicted by the acquisition of meaningful words. This information is potentially useful for early intervention and the planning of comprehensive treatment for young children with RTT., (Copyright © 2020 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.)

Cryptosporidium spp. are enteric protozoan parasites that infect a wide range of hosts including humans, and domestic and wild animals. The aim of this study was to molecularly characterize the Cryptosporidium spp. found in calf faeces in Japan. A total of 80 pre-weaned beef and dairy calves' diarrhoeic faecal specimens were collected from nine different prefectures in Japan. A nested polymerase chain reaction targeting the small subunit 18S rRNA and GP60 genes were used to detect the Cryptosporidium genotypes and subtypes. 83.8% (67 out of 80) of the specimens were positive for Cryptosporidium spp.; Cryptosporidium was found in both beef and dairy calves. Cryptosporidium parvum was the predominant species, detected in 77.5% (31/40) of beef calves and 80% (32/40) of dairy calves. Cryptosporidium bovis was also detected, 5.0% (2/40) of dairy calves, and C. ryanae was also found 2.5% (1/40) of dairy calves. One mixed-species infection, 2.5% (1/40) was detected in a beef calf having C. parvum, and C. ryanae. We detected the most common subtype of C. parvum (i.e., IIaA15G2R1), as well as other subtypes (i.e., IIaA14G3R1, IIaA14G2R1, and IIaA13G1R1) that have not previously been detected in calves in Japan. Our results demonstrate the widespread diversity of Cryptosporidium infection in calves in Japan., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The role of sphingosine 1-phosphate (S1P) and its sphingosine-1-phosphate receptors (S1PRs) in non-alcoholic steatohepatitis (NASH) is unclear. We aimed to analyze the role of S1P/S1PRs in a Melanocortin-4 receptor (Mc4r)-deficient NASH murine model using FTY720, the functional antagonist of S1PR1, S1PR3, S1PR4, and S1PR5, and JTE-013, the antagonist of S1PR2. We observed that, compared to that in the control, the mRNA of S1pr1 tended to decrease, whereas those of S1pr2 and S1pr3 significantly increased in Mc4r-knockout (KO) mice subjected to a Western diet (WD). While the fat area did not differ, fibrosis progression differed significantly between control mice and mice in which liver S1PRs were blocked. Lipidomic and metabolomic analysis of liver tissues showed that JTE-013-administered mice showed elevation of S-adenosyl-l-methionine level, which can induce aberrant methylation due to reduction in glycine N-methyltransferase (GNMT) and elevation in diacylglycerol (DG) and triacylglycerol (TG) levels, leading to increased susceptibility to hepatocellular carcinoma (HCC). These phenotypes are similar to those of Gnmt-KO mice, suggesting that blocking the S1P/S1PR2 axis triggers aberrant methylation, which may increase DG and TG, and hepatocarcinogenesis. Our observations that the S1P/S1PR2 axis averts HCC occurrence may assist in HCC prevention in NASH., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Immune-mediated thrombotic thrombocytopenic purpura (iTTP) is a rare autoimmune disorder caused by neutralizing anti-ADAMTS13 autoantibodies. In white individuals, HLA allele DRB1*11 is a predisposing factor for iTTP, whereas DRB1*04 is a protective factor. However, the role of HLA in Asians is unclear. In this study, we analyzed 10 HLA loci using next-generation sequencing in 52 Japanese patients with iTTP, and the allele frequency in the iTTP group was compared with that in a Japanese control group. We identified the following HLA alleles as predisposing factors for iTTP in the Japanese population: DRB1*08:03 (odds ratio [OR], 3.06; corrected P [Pc] = .005), DRB3/4/5*blank (OR, 2.3; Pc = .007), DQA1*01:03 (OR, 2.25; Pc = .006), and DQB1*06:01 (OR,: 2.41; Pc = .003). The estimated haplotype consisting of these 4 alleles was significantly more frequent in the iTTP group than in the control group (30.8% vs 6.0%; Pc < .001). DRB1*15:01 and DRB5*01:01 were weak protective factors for iTTP (OR, 0.23; Pc = .076; and OR, 0.23, Pc = .034, respectively). On the other hand, DRB1*11 and DRB1*04 were not associated with iTTP in the Japanese. These findings indicated that predisposing and protective factors for iTTP differ between Japanese and white individuals. HLA-DR molecules encoded by DRB1*08:03 and DRB1*11:01 have different peptide-binding motifs, but interestingly, bound to the shared ADAMTS13 peptide in an in silico prediction model., (© 2020 by The American Society of Hematology.)

Steryl glucosides (SGs) and acylated steryl glucosides (ASGs) are phytochemicals found in plant-based foods and are known as bioactive compounds with potential health benefits. These include anti-inflammatory properties, anti-diabetic effects, and modulation of immunoregulatory functions as well as having cholesterol lowering effects. In this study, three major SGs, i.e., glucosides of β-sitosterol, stigmasterol, and campesterol, were synthesized and used as standards for measurement of their contents in rice bran (RB)-based fermented food (FBRA) utilizing Aspergillus oryzae and raw material (RM). The compounds were quantified using liquid chromatography/electrospray ionization-tandem mass spectrometry. It was found that β-sitosteryl glucoside was most abundant among the analyzed glucosides in both samples, and the contents of each SG in FBRA decreased about 35% from those of RM. In contrast to SGs, the contents of ASGs in FBRA increased 1.5-fold during the fermentation process as evidenced by an alkaline hydrolysis. The present results suggest that the FBRA might have greater beneficial effects than the RM, since ASGs have shown to have more potent cholesterol lowering effects and stronger anti-diabetic properties than SGs., (Copyright © 2020. Published by Elsevier Inc.)

Background: Recessive dystrophic epidermolysis bullosa (RDEB) is a monogenic skin blistering disorder caused by mutations in the type VII collagen gene. A combination of biological technologies, including induced pluripotent stem cells (iPSCs) and several gene-editing tools, allows us to develop gene and cell therapies for such inherited diseases. However, the methodologies for gene and cell therapies must be continuously innovated for safe clinical use., Objective: In this study, we used the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology to correct the pathogenic mutation in RDEB-specific iPSCs, and the piggyBac transposon system so that no residual gene fragments remained in the genome of iPSCs after correcting the mutation., Methods: For homologous recombination (HR)-based gene editing using CRISPR/Cas9, we designed guide RNA and template DNA including homologous sequences with drug-mediated selection cassette flanked by inverted repeat sequences of the transposon. HR reaction using CRISPR/Cas9 was induced in RDEB-specific iPSCs, and mutation-corrected iPSCs (MC-iPSCs) was obtained. Consequently, the selection cassette in the genome of MC-iPSCs was removed by transposase expression., Results: After CRISPR/Cas9-induced gene editing, we confirmed that the pathogenic mutation in RDEB-specific iPSCs was properly corrected. In addition, MC-iPSCs had no genetic footprint after removing the selection cassette by transposon system, and maintained their "stemness". When differentiating MC-iPSCs into keratinocytes, the expression of type VII collagen was restored., Conclusions: Our study demonstrated one of the safer approaches to establish gene and cell therapies for skin hereditary disorders for future clinical use., Competing Interests: Declaration of Competing Interest The authors have no conflict of interest to declare., (Copyright © 2020 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.)

Introduction: Melanocortin 4 receptor-deficient (MC4R-KO) mice fed a high-fat diet (HFD) develop liver pathology similar to human nonalcoholic steatohepatitis (NASH). However, although liver histology and blood biochemistry have been reported, hepatic function has not been evaluated. In the present study, we evaluated hepatic function in MC4R-KO mice fed an HFD using dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) with gadolinium‑ethoxybenzyl‑diethylenetriamine pentaacetic acid (Gd-EOB-DTPA)., Materials and Methods: Wild type (WT) mice and MC4R-KO mice were fed a standard diet (SD) or an HFD for 20 weeks. The hepatic signal intensity was obtained from DCE-MRI images, and relative enhancement (RE), the time to maximum RE (T max ), and the half-life of RE elimination (T 1/2 ) were calculated. Histopathological analysis was then performed., Results: Histological analysis with nonalcoholic fatty liver disease activity score (NAS) revealed that MC4R-KO mice fed an HFD achieved the NAS of 5. There was moderate fibrosis in MC4R-KO mice fed an HFD. DCE-MRI with Gd-EOB-DTPA showed that T max and T 1/2 were significantly longer in MC4R-KO mice fed an HFD compared with wild type (WT) mice (T max , WT, 3.9 ± 0.4 min; MC4R-KO, 7.4 ± 1.5 min; T 1/2 , WT, 23.7 ± 1.9 min; MC4R-KO, 62.5 ± 18.5 min). T max and T 1/2 were significantly correlated with histopathologic score (steatosis vs. Tmax, rho = 0.48, P = 0.04; steatosis vs. T 1/2 , rho = 0.50, P = 0.03; inflammation vs. Tmax, rho = 0.55, P = 0.02; inflammation vs. T 1/2 , rho = 0.61, P < 0.01; ballooning vs. T 1/2 , rho = 0.51, P = 0.03;fibrosis vs T max , rho = 0.72, P < 0.01; fibrosis vs T 1/2 , rho = 0.75, P < 0.01)., Conclusions: MC4R-KO mice fed an HFD developed obesity and NASH. The liver kinetics of Gd-EOB-DTPA were significantly different in MC4R-KO mice fed an HFD from WT mice, and correlated with the histopathologic score. These results suggest that MC4R-KO mice fed an HFD mimic the hepatic pathology and liver function of human NASH, and therefore might be useful for the study of hepatic dysfunction during the fibrotic stage of NASH., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Lymphatic filariasis (LF) has been declared eliminated in Sri Lanka in September 2016. To maintain elimination status, a surveillance system to detect hidden endemic foci or LF resurgence is of highest priority. In this paper, we have reported an investigation of LF transmission in Trincomalee district where a surveillance program was not carried out due to 30 years of civil unrest. Proposed surveillance system included, measurement of anti-filarial IgG4 in urine of schoolchildren in areas where LF transmission could exist and assessment of circulating filarial antigen (CFA) and microfilaria (mf) in all urine antibody positive schoolchildren, their family members and 10-15 neighbours of each urine antibody positive household. Spatial distribution of the anti-filarial antibody titers in urine in a high antibody suspected area was analyzed using GPS logger data. Among 2301 school children from 11 schools studied, 41 (1.8%) urine antibody positives were found. The antibody positive rates of the schools ranged between 0 and 4.0%. Nine of the 630 (1.4%) examined became positive for CFA but were negative for mf. Although there were no mf positives, positive CFA and antibody results indicated the existence of Wuchereria bancrofti in Trincomalee. Highest antibody titres in an area correlated with the prevalences of urine antibodies and CFA. Spatial analysis showed LF transmission foci. Therefore, a combination of the non-invasive methods, urine ELISA and GPS mapping, will be a new effective surveillance system to identify hidden LF transmission foci., (Copyright © 2018. Published by Elsevier B.V.)

Glycolysis, the classic pathway for producing energy, has been known to be involved in neural development. Notch signaling also contributes to neural development and regulation of glycolysis in various tissues. However, the role of Notch signaling in glycolysis-related gene regulation during neural development is poorly understood. Here, we analyzed mRNA expression patterns and levels of glucose transporters (GLUT) as well as rate-limiting enzymes in glycolysis using zebrafish mib1 ta52b mutants, in which Notch signaling was deficient at the early embryonic and larval stages. Our results indicated that in neural tissues, Notch signaling positively regulates glut1a and glut3a expression and negatively regulates hk2 expression at the larval stage but may not regulate them during early embryonic stages. Therefore, these results suggest that Notch signaling regulates glycolysis-related gene expression in a context-dependent manner in neural tissues at different developmental stages., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The nonstructural protein NSs of severe fever with thrombocytopenia syndrome phlebovirus blocks type I interferon (IFN)-stimulated JAK-STAT signaling. However, there is continuing controversy as to whether NSs targets STAT1 or STAT2 or both for this blockade. The present study was designed to gain a further understanding of the blockade mechanism. Immunoprecipitation experiments revealed a stronger interaction of NSs with STAT2 than with any other component constituting the JAK-STAT pathway. Expression of NSs resulted in the formation of cytoplasmic inclusion bodies (IBs), and affected cytoplasmic distribution of STAT2. STAT2 was relocated to NSs-induced IBs. Consequently, NSs inhibited IFN-α-stimulated tyrosine phosphorylation and nuclear translocation of STAT2. These inhibitory effects as well as the signaling blockade activity were not observed in NSs mutant proteins lacking the STAT2-binding ability. In contrast, NSs affected neither subcellular distribution nor phosphorylation of STAT1 in response to IFN-α and IFN-γ, demonstrating that NSs has little physical and functional interactions with STAT1. Taken together, these results suggest that NSs sequesters STAT2 into NSs-induced IBs, thereby blocking type I IFN JAK-STAT signaling., (Copyright © 2018 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.)

We designed an automated device to study active avoidance learning abilities of zebrafish. Open source tools were used for the device control, statistical computing, and graphic outputs of data. Using the system, we developed active avoidance tests to examine the effects of trial spacing and aging on learning. Seven-month-old fish showed stronger avoidance behavior as measured by color preference index with discrete spaced training as compared to successive spaced training. Fifteen-month-old fish showed a similar trend, but with reduced cognitive abilities compared with 7-month-old fish. Further, in 7-month-old fish, an increase in learning ability during trials was observed with discrete, but not successive, spaced training. In contrast, 15-month-old fish did not show increase in learning ability during trials. Therefore, these data suggest that discrete spacing is more effective for learning than successive spacing, with the zebrafish active avoidance paradigm, and that the time course analysis of active avoidance using discrete spaced training is useful to detect age-related learning impairment., (Copyright © 2017 Elsevier Ireland Ltd and Japan Neuroscience Society. All rights reserved.)

Metabolic syndrome is a newly identified risk factor for hepatocellular carcinoma (HCC); however, tumor-specific biomarkers still remain unclear. We performed cross-species analysis to compare gene signatures of HCC from human patients and melanocortin 4 receptor-knockout mice, which develop HCC with obesity, insulin resistance, and dyslipidemia. Unsupervised hierarchical clustering and principle component analysis of 746 differentially expressed orthologous genes classified HCC of 152 human patients and melanocortin 4 receptor-knockout mice into two distinct subgroups, one of which included mouse HCC and was causatively associated with metabolic risk factors. Nine genes commonly overexpressed in human and mouse metabolic disease-associated HCC were identified; fatty acid binding protein 4 (FABP4) was remarkably enriched in intratumoral activated hepatic stellate cells (HSCs). Subclones constitutively expressing FABP4 were established from a human HSC cell line in which expression levels of inflammatory chemokines, including IL-1A and IL-6, were up-regulated through NF-κB nuclear translocation, resulting in recruitment of macrophages. An immunohistochemical validation study of 106 additional human HCC samples indicated that FABP4-positive HSCs were distributed in tumors of 38 cases, and the FABP4-high group consisted of patients with nonviral and nonalcoholic HCC (P = 0.027) and with multiple metabolic risk factors (P < 0.001) compared with the FABP4-low group. Thus, FABP4 overexpression in HSCs may contribute to hepatocarcinogenesis in patients with metabolic risk factors by modulation of inflammatory pathways., (Copyright © 2018 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Alleviating morbidity due to lymphatic filariasis (LF)-especially in elderly patients who are rather ignorant-is presently the biggest challenge for the national filariasis campaign. We introduced two follow-up schemes and compared each other to address three key programmatic issues (1) locating patients, (2) educating patients, family members on practice of lymphoedema self-care (3) well sustained daily self-care. Hundred and seven lymphoedema patients were introduced to the new Community Home Based Care (CHBC) programme as a part of MMDP programme at their homes. Twenty seven of 107 patients were selected by purposive sampling and followed-up under two schemes, 14 in Daily follow-up (DFU) scheme and 13 in Monthly follow-up (MFU) scheme. Impact was assessed using a KAP score, number of entry lesions (EL) and number of ADL episodes, limb volume, its appearance, changes in the quality of life and gained benefits. Visiting patients in their homes to introduce lymphoedema care programme was a success. KAP scores of the more important activities on lymphoedema care were significantly higher in DFU scheme. Number of patients (51.9%; 14/27) who had EL/s at baseline reduced significantly to 18.5% (5/27) at one year follow-up. The mean numbers of ADL episodes/year reduced significantly in both schemes. Six photographs of 27 showed obvious improvement in lymphoedema and its grade. Mean volume of lymphoedema reduced significantly in both schemes at one year no significant difference between schemes. Benefit score at one year revealed that the patients in DFU scheme received significantly higher amount of benefits compared to MFU scheme. In conclusion daily instruction has significantly motivated the patient and his/her family bringing a new hope., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Objective: Arima syndrome (AS) is a rare disease and its clinical features mimic those of Joubert syndrome or Joubert syndrome-related diseases (JSRD). Recently, we clarified the AS diagnostic criteria and its severe phenotype. However, genetic evidence of AS remains unknown. We explored causative genes of AS and compared the clinical and genetic features of AS with the other JSRD., Patients and Methods: We performed genetic analyses of 4 AS patients of 3 families with combination of whole-exome sequencing and Sanger sequencing. Furthermore, we studied cell biology with the cultured fibroblasts of 3 AS patients., Results: All patients had a specific homozygous variant (c.6012-12T>A, p.Arg2004Serfs*7) or compound heterozygous variants (c.1711+1G>A; c.6012-12T>A, p.Gly570Aspfs*19;Arg2004Serfs*7) in centrosomal protein 290 kDa (CEP290) gene. These unique variants lead to abnormal splicing and premature termination. Morphological analysis of cultured fibroblasts from AS patients revealed a marked decrease of the CEP290-positive cell number with significantly longer cilium and naked and protruded ciliary axoneme without ciliary membrane into the cytoplasm., Conclusion: AS resulted in cilia dysfunction from centrosome disruption. The unique variant of CEP290 could be strongly linked to AS pathology. Here, we provided AS specific genetic evidence, which steers the structure and functions of centrosome that is responsible for normal ciliogenesis. This is the first report that has demonstrated the molecular basis of Arima syndrome., (Copyright © 2017 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.)

Induced pluripotent stem (iPS) cells have the ability to differentiate into multiple cell types in the body and have an unlimited growth potential. However, iPS cell-derived melanocytes produced by existing protocols have significant limitations in developing novel strategies for regenerative medicine and cell therapies of pigmentation disorders in humans because they involve culture in media containing fetal bovine serum and nonphysiological agents. In this study, we established an in vitro approach to generate iPS cell-derived human melanocytes that have higher proliferation rates and increased melanin production compared with melanocytes prepared by previously reported approaches. Importantly, our iPS cell-derived human melanocytes are prepared in fetal bovine serum-free culture conditions that do not contain any nonphysiological agents. We designed two original methods, transferring black colonies by pipette and recovering black cell pellets from centrifuged medium, and numerous human iPS cell-derived melanocytes proliferated in gelatinous dishes coated with Matrigel after 12 days. We also succeeded in inducing melanin pigmentation in the nude mouse skin in vivo using those human iPS cell-derived melanocytes. We propose that this method using iPS cells established from T cells in the blood of normal human volunteers could be applied clinically to develop regenerative medicine and cell therapies for various forms of human pigmentation disorders., (Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.)

The increasing frequency of fire due to drainage of tropical peatland has become a major environmental problem in Southeast Asia. To clarify the effects of changes in land use on carbon dioxide emissions, we measured oxidative peat decomposition (PD) at different stages of disturbance at three sites in Central Kalimantan, Indonesia: an undrained peat swamp forest (UF), a heavily drained peat swamp forest (DF), and a drained and burned ex-forest (DB). PD exhibited seasonality, being less in the wet season and greater in the dry season. From February 2014 to December 2015, mean PD (±SE) were 1.90±0.19, 2.30±0.33, and 1.97±0.25μmolm -2 s -1 at UF, DF, and DB, respectively. The groundwater level (GWL) was a major controlling factor of PD at all sites. At UF and DF, PD and GWL showed significant quadratic relationships. At DB, PD and GWL showed significant positive and negative relationships during the dry and wet seasons, respectively. Using these relationships, we estimated annual PD from GWL data for 2014 and 2015 as 698 and 745gCm -2 yr -1 at UF (mean GWL: -0.23 and -0.39m), 775 and 825gCm -2 yr -1 at DF (-0.55 and -0.59m), and 646 and 748gCm -2 yr -1 at DB (-0.22 and -0.62m), respectively. The annual PD was significantly higher in DF than in UF or DB, in both years. Despite the very dry conditions, the annual PD values at these sites were much lower than those reported for tropical peat at plantations (e.g., oil palm, rubber, and acacia). The differences in the relationship between PD and GWL indicate that separate estimations are required for each type of land. Moreover, our results suggest that PD can be enhanced by drainage both in forests and at burned sites., (Copyright © 2017 Elsevier B.V. All rights reserved.)

The activation of phosphatidylinositol 3-kinase-AKTs-mammalian target of rapamycin cell signaling pathway leads to cell overgrowth and abnormal migration and results in various types of cortical malformations, such as hemimegalencephaly (HME), focal cortical dysplasia, and tuberous sclerosis complex. However, the pathomechanism underlying abnormal cell migration remains unknown. With the use of fetal mouse brain, we performed causative gene analysis of the resected brain tissues from a patient with HME and investigated the pathogenesis. We obtained a novel somatic mutation of the MTOR gene, having approximately 11% and 7% mutation frequency in the resected brain tissues. Moreover, we revealed that the MTOR mutation resulted in hyperphosphorylation of its downstream molecules, S6 and 4E-binding protein 1, and delayed cell migration on the radial glial fiber and did not affect other cells. We suspect cell-autonomous migration arrest on the radial glial foot by the active MTOR mutation and offer potential explanations for why this may lead to cortical malformations such as HME., (Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Insulin-like growth factor binding protein (IGFBP)-3 regulates IGF bioactivity, induces apoptosis, and inhibits cell growth independent of IGFs, but the functional role of IGFBP3 in the brain is not clear. In the present study, we revealed the effect of IGFBP3 on the brain by characterizing the phenotype of Igfbp3-null mice. Compared with wild-type mice, Igfbp3-null mice had significantly decreased IGF-1 content in the brain but no change in weights of brain and body. In Igfbp3-null mice, the number of dendritic spines was significantly reduced, and the dendritic diameter was thickening. In addition, in Igfbp3-null mice, a decrease in phosphorylated Akt and ERK1/2 significantly reduced PSD-95 expression, and GAD65/67 expression was significantly decreased. These results indicate that IGFBP3 deficiency impairs neuronal structure and signaling. In behavioral studies, Igfbp3-null mice were hyperactive, and a Y-maze alternation test revealed impaired spatial working memory but no anxiety-like behavior. Monoaminergic analysis using high-performance liquid chromatography indicated that Igfbp3-null mice had lower levels of dopamine and serotonin compared with wild-type mice, suggesting an abnormal monoaminergic neurotransmission. In conclusion, our studies found that the deletion of IGFBP3 results in behavioral impairments that are associated with abnormal synaptic function and monoaminergic neurotransmission, which helps to characterize the critical role of IGFBP3 in the brain., (Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

The Posttraumatic Diagnostic Scale (PDS) is a brief, self-report questionnaire developed for the diagnostic screening and assessment of the severity of posttraumatic stress disorder (PTSD); the PDS is based on the criteria outlined in the Diagnostic and Statistical Manual of Mental Disorders (4th edition; DSM-IV). We investigated the validity and reliability of the Japanese version of the PDS in a clinical (n=109) and a non-clinical (n=116) sample, recruited from an outpatient psychiatric facility and a university student population, respectively. The Japanese versions of the PDS and the Clinician-Administered PTSD Scale (CAPS/DSM-IV) were administered to the participants. The Japanese PDS's diagnostic sensitivity and specificity exceeded 90%. The correlation between the severity scores assessed by the Japanese PDS and the CAPS was also high (r=0.92). The findings suggest that the Japanese version of the PDS is useful for diagnostically screening PTSD and assessing symptom severity., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Aims: To clarify the incidence of spontaneous preterm birth (PB) and septic abortion (sab) in Gifu prefecture in Japan., Study Design: This prospective, population-based cohort study was approved by our hospital's Institutional Review Board. All 36 hospitals (100%) in Gifu prefecture offering obstetrical services participated in the study. Patient enrollment criteria were: sab and PB from 22 to <37weeks gestation (WG), excluding for maternal and fetal indications. Pathological examinations before 36 WG and associated factors for both PB and chorioamnionitis (CAM) stage 3 were analyzed by multiple logistic regression analysis judging from minimum daily clinical information in Gifu prefecture., Results: The sab rate per all deliveries was 29/16871 (0.17%) at 16.9±2.9 WG. The total spontaneous PB rate was 615/16871 (3.65%) at 34.5±2.7 WG, with birth weight (BW) 2267±557g. There were 26 (0.15%) PBs from 22+0 to 27+6 WG (weeks+days) at 25.2±1.5 WG, with BW 745±199g. Among 214 pathological examinations, CAM was detected in 80% (sab) and 63% (PB<36 WG), respectively. Funisitis were 14% and 17% respectively. Episodes of serial genital bleeding and/or hematoma at <12 WG were more frequent in sab and earlier PB (<28 WG) associated with CAM stage 3 (odds 1.9, P<0.0001). Combined factors such as bleeding and past history of CAM correlated with earlier delivery at 23.4±5.9 WG (P=0.0032)., Conclusion: In Gifu prefecture, the incidence of sab was 0.17% (per all deliveries) and 3.65% of spontaneous PB. The combined risk of past CAM history and bleeding was associated with earlier delivery among total preterm birth., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

To evaluate the relative contribution of cytochrome P450 (CYP) isoforms responsible for carvedilol (CAR) oxidation, enantioselective metabolism of CAR was investigated in human liver microsomes (HLMs) and recombinant human CYPs by using the substrate depletion assay. CYP2D6 exhibited the highest contribution to the metabolism of R-CAR, followed by CYP3A4, CYP1A2, and CYP2C9, whereas the metabolism of the S-enantiomer was mainly mediated by CYP1A2, followed by CYP2D6 and CYP3A4. In HLMs, metabolism of R- and S-CAR was markedly inhibited by quinidine; R-CAR metabolism (57-61% decrease) was more inhibited than S-CAR metabolism (37-43% decrease), and furafylline and ketoconazole almost equally inhibited metabolism of both enantiomers by 25-32% and 30-50%, respectively. The absence of CYP2D6 in a mixture of five major recombinant CYP isoforms at the approximate ratio as in HLMs resulted in a 42% and 25% decrease in the metabolic activities for R- and S-CAR, respectively. Moreover, the absence of CYP1A2 in the mixture resulted in a 16% and 39% decrease in the metabolic activities for R- and S-CAR, respectively. Our results suggest the stereoselective metabolism of CAR is determined by not only the activity of CYP2D6 but also of CYP1A2 and CYP3A4., (Copyright © 2016 The Japanese Society for the Study of Xenobiotics. Published by Elsevier Ltd. All rights reserved.)

Expanded human T cells from a Japanese healthy male were used to generate integration-free induced pluripotent stem cells (iPSCs) by exogenous expression of four reprogramming factors, OCT3/4, SOX2, cMYC, KLF4, using Sendai virus vector (SeVdp). The authenticity of established iPSC line, WT-iPSC4, was confirmed by the expressions of stem cell markers and the differentiation capability into three germ layer. WT-iPSC4 may be a useful cell resource as a normal control for the comparative study using disease-specific iPSCs., (Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.)

Expanded human T cells from a Japanese female with recessive dystrophic epidermolysis bullosa (RDBE) were used to generate integration-free induced pluripotent stem cells (iPSCs) by exogenous expression of four reprogramming factors, OCT3/4, SOX2, cMYC, KLF4, using Sendai virus vector (SeVdp). The authenticity of established iPSC line, RDEB-iPSC26, was confirmed by the expressions of stem cell markers and the differentiation capability into three germ layer. RDEB-iPSC26 may be a useful cell resource for the establishment of in vitro RDEB modeling and the study for developing gene and cell therapy., (Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.)

The endoplasmic reticulum (ER) is important in various cellular functions, such as secretary and membrane protein biosynthesis, lipid synthesis, and calcium storage. ER stress, including membrane distortion, is associated with many diseases such as Huntington's disease. In particular, nuclear envelope distortion is related to neuronal cell death associated with polyglutamine. However, the mechanism by which polyglutamine causes ER membrane distortion remains unclear. We used electron microscopy, fluorescence protease protection assay, and alkaline treatment to analyze the localization of polyglutamine in cells. We characterized polyglutamine embedded in the ER membrane and noted an effect on morphology, including the dilation of ER luminal space and elongation of ER-mitochondria contact sites, in addition to the distortion of the nuclear envelope. The polyglutamine embedded in the ER membrane was observed at the same time as Bax insertion. These results demonstrated that the ER membrane may be a target of polyglutamine, which triggers cell death through Bax., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: Neither the atrial fibrillation (AF) maintenance mechanism after pulmonary vein isolation (PVI) nor the mechanism of AF termination via stepwise ablation is clearly understood., Methods: Among 226 consecutive AF patients (154 paroxysmal (P-AF) and 72 persistent AF (Per-AF) patients), left atrial endocardial non-contact mapping was performed after PVI in the initial 10 P-AF and 16 Per-AF patients to define the AF maintenance mechanism. Subsequently, effect of stepwise catheter ablation (linear roof lesion and complex fractionated atrial electrogram (CFAE) following PVI) was evaluated in all patients., Results: After PVI, AF was maintained by the activation around isolated PV/mitral annulus, focal discharge and disorganized activations mostly observed over residual CFAE region (pivoting activation, wave break and fusion). CFAE region in P-AF was smaller than Per-AF after PVI (1.6 ± 2.1 vs. 7.7 ± 2.5 cm(2), p<0.0001). The frequency of pivoting activation, wave break and fusion in P-AF were lower than those in Per-AF (1.9 ± 2.0 vs. 11.8 ± 5.0 times/s; p<0.0001, 0.1 ± 0.3 vs. 3.6 ± 2.5 times/s; p<0.001, 5.8 ± 3.6 vs. 9.8 ± 3.2 times/s; p<0.01). AF termination was more frequent in P-AF than Per-AF (94.8% vs 81.9%, p=0.0019). AF termination by PVI alone was more frequent in P-AF than Per-AF (85.6% vs. 18.6%, p<0.0001). However, AF termination via roof line and/or CFAE ablation was less frequent in P-AF than Per-AF (14.4 vs. 81.4%, p<0.0001)., Conclusions: Disorganized activations after PVI, more prominent in Per-AF, were associated with residual CFAE region. Most P-AF was terminated by PVI alone, however additional roof line lesion and CFAE ablation were necessary to terminate Per-AF, consistent with mapping results., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Expanded human T cells from a Japanese male with lymphedema-distichiasis syndrome (LDS) were used to generate integration-free induced pluripotent stem cells (iPSCs) by exogenous expression of four reprogramming factors, OCT3/4, SOX2, cMYC, KLF4, using Sendai virus vector (SeVdp). The authenticity of established iPSC line, LDS-iPSC8, was confirmed by the expression of stem cell markers and the differentiation capability into three germ layers. LDS-iPSC8 may be a useful cell resource for the establishment of in vitro LDS modeling and the study for vascular and lymph vessel development., (Copyright © 2016 Roslin Cells Ltd. Published by Elsevier B.V. All rights reserved.)

Human African trypanosomiasis (HAT) is a disease caused by Kinetoplastid infection. Serological tests are useful for epidemiological surveillance. The aim of this study was to develop a multiplex serological assay for HAT to assess the diagnostic value of selected HAT antigens for sero-epidemiological surveillance. We cloned loci encoding eight antigens from Trypanosoma brucei gambiense, expressed the genes in bacterial systems, and purified the resulting proteins. Antigens were subjected to Luminex multiplex assays using sera from HAT and VL patients to assess the antigens' immunodiagnostic potential. Among T. b. gambiense antigens, the 64-kDa and 65-kDa invariant surface glycoproteins (ISGs) and flagellar calcium binding protein (FCaBP) had high sensitivity for sera from T. b. gambiense patients, yielding AUC values of 0.871, 0.737 and 0.858 respectively in receiver operating characteristics (ROC) analysis. The ISG64, ISG65, and FCaBP antigens were partially cross-reactive to sera from Trypanosoma brucei rhodesiense patients. The GM6 antigen was cross-reactive to sera from T. b. rhodesiense patients as well as to sera from VL patients. Furthermore, heterogeneous antibody responses to each individual HAT antigen were observed. Testing for multiple HAT antigens in the same panel allowed specific and sensitive detection. Our results demonstrate the utility of applying multiplex assays for development and evaluation of HAT antigens for use in sero-epidemiological surveillance., (Copyright © 2015 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.)

Background: Selective cyclooxygenase-2 (COX-2) inhibitors, conventional non-selective nonsteroidal anti-inflammatory drugs (NSAIDs), and acetaminophen have been adopted for the relief of mild to moderate acute and chronic pain. However, it remains unclarified whether the therapeutic differences in pain sensation exist among these agents. The aim of this study was to compare the efficacy of different types of analgesic agents for postoperative acute pain management., Methods: A single-center, randomized, controlled study was performed in consecutive patients who underwent the second-look procedure with removal of internal fixation after anterior cruciate ligament reconstruction or arthroscopic meniscal repair/meniscectomy. Celecoxib (400 mg for the first dose and then 200 mg), loxoprofen (60 mg), or acetaminophen (600 mg) was orally administered from postoperative 3 h. The pain intensity on a 100-mm VAS scale and subjective assessment of therapeutic pain-relief were compared among these three treatment groups until postoperative 2 days. The acquired data were analyzed according to the per-protocol analysis principle., Results: A total of 432 patients were screened, and 160 were enrolled. The VAS score tended to decrease over time in all groups. There was a significant improvement in the pain score both at rest and on movement, and subjective impression in the celecoxib-treated group compared with acetaminophen at postoperative 2 days. On the other hand, loxoprofen resulted in the benefit only in the pain score at rest in comparison with acetaminophen. Any comparisons between celecoxib and loxoprofen showed insignificant differences throughout observations. No adverse effects were confirmed in each group., Conclusions: These obtained findings in our dose setting conditions suggest that celecoxib and loxoprofen treatments were superior to acetaminophen in pain-relief, though the superiority of loxoprofen over acetaminophen was modest. Overall, selective COX-2 inhibitors including conventional NSAIDs seem to have a possible advantage in acute pain management of relatively less invasive surgery., (Copyright © 2015 The Japanese Orthopaedic Association. Published by Elsevier B.V. All rights reserved.)

Cell death abnormal (ced)-3 and ced-4 genes regulate apoptosis to maintain tissue homeostasis in Caenorhabditis elegans. Apoptosome formation and CED-4 translocation drive CED-3 activation. However, the precise role of CED-4 translocation is not yet fully understood. In this study, using a combination of immunoprecipitation and reverse transcription-polymerase chain reaction methods in cells and a glutathione-S-transferase pull down assay in a cell-free system, we show that CED-4 binds ced-3 mRNA. In the presence of ced-3 mRNA, CED-4 protein is enriched in the microsomal fraction and interacts with ribosomal protein L10a in mammalian cells, increasing the levels of CED-3. These results suggest that CED-4 forms a complex with ced-3 mRNA and delivers it to ribosomes for translation., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Background: The aim of our study was to develop a completely scaffold-free, viable, contractile cardiac tissue capable of being grafted into the damaged native heart., Methods: Our technology is based on the fundamental characteristics of the self-assembling nature of cells. We created contractile cardiac spheroids by plating a mixture of rat neonatal ventricular cardiomyocytes, human dermal fibroblasts, and human coronary microartery endothelial cells in ultralow attachment plates. First, the optimal cell ratios for the 3 cell sources were determined. Next, approximately 1 × 10(4) optimal spheroids were fused into a patch-like construct, and the morphologic characteristics and mechanical functions of these patches were evaluated. Finally, the cardiac patches were grafted into the hearts of F344 nude rats, and histologic studies were performed after transplantation., Results: Synchronous beating of the cardiac patch was confirmed electrophysiologically and mechanically. A micronetwork of endothelial cells was also demonstrated in the construct, and the histologic study performed 5 days after transplantation showed the grafts to be viable, with functioning microvascular structures inside the graft tissue., Conclusions: We consider the application of our scaffold-free 3-dimensional tissue engineering technology to cardiac regeneration therapy is feasible and expect that this technology will become a promising tool for the treatment of end-stage heart failure., (Copyright © 2016 International Society for Heart and Lung Transplantation. Published by Elsevier Inc. All rights reserved.)

Fetal hepatic stem/progenitor cells, hepatoblasts, are highly proliferative cells and the source of both hepatocytes and cholangiocytes. In contrast, mature hepatocytes have a low proliferative potency and high metabolic functions. Cell proliferation is regulated by cell cycle-related molecules. However, the correlation between cell cycle regulation and hepatic maturation are still unknown. To address this issue, we revealed that the cell cycle inhibitor p57(Kip2) was expressed in the hepatoblasts and mesenchymal cells of fetal liver in a spatiotemporal manner. In addition, we found that hepatoblasts in p57(Kip2)-/- mice were highly proliferative and had deficient maturation compared with those in wild-type (WT) mice. However, there were no remarkable differences in the expression levels of cell cycle- and bipotency-related genes except for Ccnd2. Furthermore, p57(Kip2)-/- hepatoblasts could differentiate into mature hepatocytes in p57(Kip2)-/- and WT chimeric mice, suggesting that the intrinsic activity of p57(Kip2) does not simply regulate hepatoblast maturation., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The production of amyloid β (Aβ) in the brain from Aβ precursor protein (APP) through γ-secretase is important for the pathogenesis of Alzheimer's disease (AD). Our previous studies have demonstrated that autophagy impairment and endoplasmic reticulum stress increase presenilin 1 expression and enhance γ-secretase activity through the phosphorylation of eukaryotic translation initiation factor 2α (eIF2α) and the translation of activating transcription factor 4 (ATF4). However, the inhibitory molecules for γ-secretase are largely unknown. Here, we demonstrate that the levels of ATF4 expression are increased in the brain of APP23 AD model mice; furthermore, these levels enhanced in the brain of APP23 mice crossed with obese and diabetic db/db (Lepr(db/db)) mice. A polyhydroxylated flavonoid, quercetin, suppressed presenilin 1 expression and Aβ secretion in autophagy-impaired cells by the induction of growth arrest and DNA damaged-inducible gene (GADD) 34, which mediates eIF2α dephosphorylation, leading to decreased ATF4 expression. GADD34 induction was observed in the brain of wild-type mice, and APP23 mice fed quercetin in their diet. After the long-term feeding of quercetin, deterioration in memory assessed by freezing behavior was delayed in APP23 mice. These results indicate that quercetin may reduce eIF2α phosphorylation and ATF4 expression through GADD34 induction in the brain, leading to the improvement of memory in aged mice and the delay of deterioration in memory at the early stage of AD in AD model mice., (Copyright © 2015 Elsevier Inc. All rights reserved.)

We report on the clinical, neuropathological, and genetic findings of a Japanese case with myocerebrohepatopathy spectrum (MCHS) disorder due to polymerase gamma (POLG) mutations. A girl manifested poor sucking and failure to thrive since 4 months of age and had frequent vomiting and developmental regression at 5 months of age. She showed significant hypotonia and hepatomegaly. Laboratory tests showed hepatocellular dysfunction and elevated protein and lactate levels in the cerebrospinal fluid. Her liver function and neurologic condition exacerbated, and she died at 8 months of age. At autopsy, fatty degeneration and fibrosis were observed in the liver. Neuropathological examination revealed white matter-predominant spongy changes with Alzheimer type II glia and loss of myelin. Enzyme activities of the respiratory chain complex I, III, and IV relative to citrate synthase in the muscle were normal in the biopsied muscle tissue, but they were reduced in the liver to 0%, 10%, and 14% of normal values, respectively. In the liver, the copy number of mitochondrial DNA compared to nuclear DNA was reduced to 3.3% of normal values as evaluated by quantitative polymerase chain reaction. Genetic analysis revealed compound heterozygous mutations for POLG (I1185T/A957V). This case represents the differential involvement of multiple organs and phenotype-specific distribution of brain lesions in mitochondrial DNA depletion disorders., (Copyright © 2014 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.)

Background: The dermal papilla is composed of a small clump of mesenchymal cells, called dermal papilla cells (DPCs). DPCs closely interact with epidermal cells to give rise to hair follicles and shafts during hair follicle development and the hair cycle. DPCs are promising cell sources for hair regeneration therapy for alopecia patients. However, once DPCs are put into conventional two-dimensional culture conditions, they quickly lose their capability to produce hair follicles., Objective: We aimed to expand a sufficiently large population of DPCs that retain their hair inductive activity., Methods: Murine DPCs were cultured in the presence of platelet-derived growth factor-AA (PDGF-AA) and fibroblast growth factor 2 (FGF2). Expressions of follicular-related genes were analyzed by real time PCR and hair inductive activity was determined by patch assay and chamber assay in vivo., Results: FGF2 significantly increased the expression of platelet-derived growth factor receptor alpha (PDGFRα) in cultured vibrissal DPCs. PDGF-AA, a ligand of PDGFRα, promoted proliferation of DPCs synergistically when utilized with FGF2 and enhanced the expression of several follicular-related genes in DPCs. Hair reconstitution assays revealed that DPCs treated with both PDGF-AA and FGF-2 were able to maintain their hair inductive activity better than those treated with FGF2 alone., Conclusion: Both cell proliferation and hair inductive activity in murine DPCs are maintained by the synergistic effect of FGF2 and PDGF-AA., (Copyright © 2015 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.)

Stereotyped movement of paired pectoral fins in zebrafish larvae could be considered a simple model with which to investigate the neural basis of behavior. Using a high-speed camera, we explored the repertoire of pectoral fin movements by naturally behaving larvae at 5-6 days post-fertilization. Previously, two types of fin movements were characterized in association with locomotion: 'CRAWLing,' an alternating fin movement associated with slow swimming, and 'TUCKing,' the adduction of both fins associated with fast swimming. We here describe a third mode of fin movement, which we call 'Munch's SCREAM', in which both pectoral fins were flipped anteriorly so that they reached the skin on the sides of the head, thus covering the otic vesicles. This behavior occurred spontaneously and was often associated with a slight regression or a sudden bending and change in body orientation. It could be also induced effectively in the agarose-embedded larvae by tactile stimulation on the skin around the eye and nose, some of which are associated with struggling, in which waves of bending propagate from the tail to the head. Larvae can still CRAWL and perform the SCREAM even when their forebrain and midbrain have been removed, suggesting that the neural circuits involved in the SCREAM are present in the hindbrain and/or spinal cord., (Copyright © 2014 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved.)

The broad diversity of neurons is vital to neuronal functions. During vertebrate development, the spinal cord is a site of sensory and motor tasks coordinated by interneurons and the ongoing neurogenesis. In the spinal cord, V2-interneuron (V2-IN) progenitors (p2) develop into excitatory V2a-INs and inhibitory V2b-INs. The balance of these two types of interneurons requires precise control in the number and timing of their production. Here, using zebrafish embryos with altered Notch signaling, we show that different combinations of Notch ligands and receptors regulate two functions: the maintenance of p2 progenitor cells and the V2a/V2b cell fate decision in V2-IN development. Two ligands, DeltaA and DeltaD, and three receptors, Notch1a, Notch1b, and Notch3 redundantly contribute to p2 progenitor maintenance. On the other hand, DeltaA, DeltaC, and Notch1a mainly contribute to the V2a/V2b cell fate determination. A ubiquitin ligase Mib, which activates Notch ligands, acts in both functions through its activation of DeltaA, DeltaC, and DeltaD. Moreover, p2 progenitor maintenance and V2a/V2b fate determination are not distinct temporal processes, but occur within the same time frame during development. In conclusion, V2-IN cell progenitor proliferation and V2a/V2b cell fate determination involve signaling through different sets of Notch ligand-receptor combinations that occur concurrently during development in zebrafish., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Neonatal hypoxic-ischemic encephalopathy (HIE) remains a serious burden in neonatal care. Hypothermia provides a good outcome in some babies with HIE. Here, we investigated the biological mechanisms of its neuroprotective effect and sought for a new therapeutic target. We made neonatal HIE rats and subjected some of them to hypothermia at 28°C for 3 hours. We pathologically confirmed the efficacy of hypothermia against the neonatal HIE brain. To clarify the molecular mechanism of hypothermia's efficacy, we analyzed mRNA expression, immunoassay, and pathology in the brain with or without HIE and/or hypothermia. We selected from these analyses 12 molecules with possible neuroprotective effects. After identification of lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) as a therapeutic target candidate, we examined the efficacy of an anti-LOX-1 neutralizing antibody in neonatal HIE rats. Administration of an anti-LOX-1 neutralizing antibody reduced infarction area, brain edema, and apoptotic cell death to a degree comparable with hypothermia. Protection from those pathological conditions was considered part of the therapeutic mechanism of hypothermia. The efficacy of administering anti-LOX-1 neutralizing antibody was similar to that of hypothermia. LOX-1 is a promising therapeutic target in neonatal HIE, and the inhibition of LOX-1 may become a novel treatment for babies who have experienced asphyxia., (Copyright © 2014 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Multiple sclerosis (MS) is a chronic demyelinating disease often displaying a relapsing-remitting course of neurological manifestations that is mimicked by experimental autoimmune encephalomyelitis (EAE) in animal models of MS. In particular, NOD mice immunized with myelin oligodendrocyte glycoprotein peptide 35-55 develop chronic relapsing-remitting EAE (CREAE). To elucidate the mechanisms that cause MS relapse, we investigated the histopathology and cytokine production of spleen cells and mRNA expression levels in the central nervous system (CNS) of CREAE mice. During the first attack, inflammatory cell infiltration around small vessels and in the subarachnoid space was observed in the spinal cord. Spleen cell production and mRNA expression in the CNS of several cytokines, including IFN-γ, TNF-α, IL-6, IL-17, and CC chemokine ligand 2 (CCL2), were higher in CREAE mice than in controls. Afterwards, parenchymal infiltration and demyelination were observed histologically in the spinal cord and corresponded with the more severe clinical symptoms of the first and second relapses. IL-17 and CCL2, but not IFN-γ, TNF-α, or IL-6, were also produced by spleen cells during recurrences. Our results suggested that the immune mechanisms in relapses were different from those in the first attack for CREAE. Further investigation of CREAE mechanisms may provide important insights into successful therapies for human relapsing-remitting MS., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Aim: We have never known any epidemiological study of Arima syndrome since it was first described in 1971. To investigate the number of Arima syndrome patients and clarify the clinical differences between Arima syndrome and Joubert syndrome, we performed the first nationwide survey of Arima syndrome, and herein report its results. Furthermore, we revised the diagnostic criteria for Arima syndrome., Methods: As a primary survey, we sent out self-administered questionnaires to most of the Japanese hospitals with a pediatric clinic, and facilities for persons with severe motor and intellectual disabilities, inquiring as to the number of patients having symptoms of Arima syndrome, including severe psychomotor delay, agenesis or hypoplasia of cerebellar vermis, renal dysfunction, visual dysfunction and with or without ptosis-like appearance. Next, as the second survey, we sent out detailed clinical questionnaires to the institutes having patients with two or more typical symptoms., Results: The response rate of the primary survey was 72.7% of hospitals with pediatric clinic, 63.5% of national hospitals and 66.7% of municipal and private facilities. The number of patients with 5 typical symptoms was 13 and that with 2-4 symptoms was 32. The response rate of the secondary survey was 52% (23 patients). After reviewing clinical features of 23 patients, we identified 7 Arima syndrome patients and 16 Joubert syndrome patients. Progressive renal dysfunction was noticed in all Arima syndrome patients, but in 33% of those with Joubert syndrome., Conclusion: It is sometimes difficult to distinguish Arima syndrome from Joubert syndrome. Some clinicians described a patient with Joubert syndrome and its complications of visual dysfunction and renal dysfunction, whose current diagnosis was Arima syndrome. Thus, the diagnosis of the two syndromes may be confused. Here, we revised the diagnostic criteria for Arima syndrome., (Copyright © 2013 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.)

Mast cells (MCs) mature exclusively in peripheral tissues, hampering research into their developmental and functional programs. Here, we employed deep cap analysis of gene expression on skin-derived MCs to generate the most comprehensive view of the human MC transcriptome ever reported. An advantage is that MCs were embedded in the FANTOM5 project, giving the opportunity to contrast their molecular signature against a multitude of human samples. We demonstrate that MCs possess a unique and surprising transcriptional landscape, combining hematopoietic genes with those exclusively active in MCs and genes not previously reported as expressed by MCs (several of them markers of unrelated tissues). We also found functional bone morphogenetic protein receptors transducing activatory signals in MCs. Conversely, several immune-related genes frequently studied in MCs were not expressed or were weakly expressed. Comparing MCs ex vivo with cultured counterparts revealed profound changes in the MC transcriptome in in vitro surroundings. We also determined the promoter usage of MC-expressed genes and identified associated motifs active in the lineage. Befitting their uniqueness, MCs had no close relative in the hematopoietic network (also only distantly related with basophils). This rich data set reveals that our knowledge of human MCs is still limited, but with this resource, novel functional programs of MCs may soon be discovered.

In development, epigenetic mechanisms such as DNA methylation have been suggested to provide a cellular memory to maintain multipotency but also stabilize cell fate decisions and direct lineage restriction. In this study, we set out to characterize changes in DNA methylation and gene expression during granulopoiesis using 4 distinct cell populations ranging from the oligopotent common myeloid progenitor stage to terminally differentiated neutrophils. We observed that differentially methylated sites (DMSs) generally show decreased methylation during granulopoiesis. Methylation appears to change at specific differentiation stages and overlap with changes in transcription and activity of key hematopoietic transcription factors. DMSs were preferentially located in areas distal to CpG islands and shores. Also, DMSs were overrepresented in enhancer elements and enriched in enhancers that become active during differentiation. Overall, this study depicts in detail the epigenetic and transcriptional changes that occur during granulopoiesis and supports the role of DNA methylation as a regulatory mechanism in blood cell differentiation.

Human blood monocytes comprise at least 3 subpopulations that differ in phenotype and function. Here, we present the first in-depth regulome analysis of human classical (CD14(++)CD16(-)), intermediate (CD14(+)CD16(+)), and nonclassical (CD14(dim)CD16(+)) monocytes. Cap analysis of gene expression adapted to Helicos single-molecule sequencing was used to map transcription start sites throughout the genome in all 3 subsets. In addition, global maps of H3K4me1 and H3K27ac deposition were generated for classical and nonclassical monocytes defining enhanceosomes of the 2 major subsets. We identified differential regulatory elements (including promoters and putative enhancers) that were associated with subset-specific motif signatures corresponding to different transcription factor activities and exemplarily validated novel downstream enhancer elements at the CD14 locus. In addition to known subset-specific features, pathway analysis revealed marked differences in metabolic gene signatures. Whereas classical monocytes expressed higher levels of genes involved in carbohydrate metabolism, priming them for anaerobic energy production, nonclassical monocytes expressed higher levels of oxidative pathway components and showed a higher mitochondrial routine activity. Our findings describe promoter/enhancer landscapes and provide novel insights into the specific biology of human monocyte subsets.

CD4(+)CD25(+)FOXP3(+) human regulatory T cells (Tregs) are essential for self-tolerance and immune homeostasis. Here, we describe the promoterome of CD4(+)CD25(high)CD45RA(+) naïve and CD4(+)CD25(high)CD45RA(-) memory Tregs and their CD25(-) conventional T-cell (Tconv) counterparts both before and after in vitro expansion by cap analysis of gene expression (CAGE) adapted to single-molecule sequencing (HeliScopeCAGE). We performed comprehensive comparative digital gene expression analyses and revealed novel transcription start sites, of which several were validated as alternative promoters of known genes. For all in vitro expanded subsets, we additionally generated global maps of poised and active enhancer elements marked by histone H3 lysine 4 monomethylation and histone H3 lysine 27 acetylation, describe their cell type-specific motif signatures, and evaluate the role of candidate transcription factors STAT5, FOXP3, RUNX1, and ETS1 in both Treg- and Tconv-specific enhancer architectures. Network analyses of gene expression data revealed additional candidate transcription factors contributing to cell type specificity and a transcription factor network in Tregs that is dominated by FOXP3 interaction partners and targets. In summary, we provide a comprehensive and easily accessible resource of gene expression and gene regulation in human Treg and Tconv subpopulations.

Hematopoietic differentiation is governed by a complex regulatory program controlling the generation of different lineages of blood cells from multipotent hematopoietic stem cells. The transcriptional program that dictates hematopoietic cell fate and differentiation requires an epigenetic memory function provided by a network of epigenetic factors regulating DNA methylation, posttranslational histone modifications, and chromatin structure. Aberrant interactions between epigenetic factors and transcription factors cause perturbations in the blood cell differentiation program that result in various types of hematopoietic disorders. To elucidate the contributions of different epigenetic factors in human hematopoiesis, high-throughput cap analysis of gene expression was used to build transcription profiles of 199 epigenetic factors in a wide range of blood cells. Our epigenetic transcriptome analysis revealed cell type- (eg, HELLS and ACTL6A), lineage- (eg, MLL), and/or leukemia- (eg, CHD2, CBX8, and EPC1) specific expression of several epigenetic factors. In addition, we show that several epigenetic factors use alternative transcription start sites in different cell types. This analysis could serve as a resource for the scientific community for further characterization of the role of these epigenetic factors in blood development.

Tracheo-innominate artery fistula is fatal unless treated surgically. We describe our surgical approach and results in seven patients. The average patient age was 15.7 years; all patients had prior severe neurological deficits. Three of seven patients were in hemorrhagic shock; control of preoperative bleeding was achieved with tracheostomy tube cuff overinflation. The innominate artery and the trachea were exposed through a collar incision and partial upper sternotomy. The innominate artery was divided at the aortic arch and at the bifurcation, with one exception. Cerebral blood flow was monitored by the blood pressure difference in the bilateral upper extremities and by near-infrared spectroscopy. The tracheal fistula was left adherent to the innominate artery in all but one patient. All patients were discharged without new neurologic deficits or severe morbidity. Overall survival was 84% at 37 months, without any vascular, tracheal, or neurological events., (Crown Copyright © 2014. Published by Mosby, Inc. All rights reserved.)

For the declaration of elimination of lymphatic filariasis, reliable epidemiological data in all parts of a country are required. In Sri Lanka, due to social disturbance, there are 3 provinces whose endemicity has been declared unknown. Further, a recent report revealed an endemic pocket, which is on the border with the district that was not covered by the national elimination program. These facts indicate the necessity of more extensive studies to discover hidden endemic foci. To facilitate such studies, we evaluated 2 methods of Rapid Assessment Procedure (RAP) in Hambantota district, where the filariasis endemicity was low: (1) indirect questioning by mailing a questionnaire to each local leader (IndQ), asking about the presence of clinical cases, and (2) focus group discussion (FGD) by villagers. The information given by people was validated with clinical examination by doctors (CE) and IgG4 ELISA using urine samples. In the results: there was a strong positive correlation between CE and ELISA rates. The hydrocele rates obtained by FGD or IndQ were associated significantly with CE rates. The rates by FGD or Cluster-IndQ ('modified' IndQ) were also associated significantly with ELISA rates. The IndQ was most cost-effective. Based on these findings, we have concluded that screening by IndQ and confirmation by the ELISA would be an effective and practical way in Sri Lanka to locate endemic foci in hitherto unsurveyed districts., (© 2013 Elsevier Ireland Ltd. All rights reserved.)

The endoplasmic reticulum (ER) plays a pivotal role in cellular functions such as the ER stress response. However, the effect of the ER membrane on caspase activation remains unclear. This study reveals that polyglutamine oligomers augmented at ER induce insertion of Bax into the ER membrane, thereby activating caspase-7. In line with the role of ER in cell death induced by polyglutamine expansion, the ER membrane was found to be disrupted and dilated in the brain of a murine model of Huntington's disease. We can conclude that polyglutamine expansion may drive caspase-7 activation by disrupting the ER membrane., (Copyright © 2013 Elsevier Inc. All rights reserved.)