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We previously suggested that the signal transducer and activator of transcription 1 (STAT1) gene is autoregulated in an interferon (IFN)-dependent manner via a distal regulatory region approximately 5.5-6.2 kb upstream of the murine and human STAT1 promoters (designated 5.5URR). Here, we examined whether this IFN-dependent positive feedback mechanism of the STAT1 gene actually functions in cells. First, we created human embryonic kidney 293 cell mutants lacking the IFN-responsive transcription factor binding sites (IFN-stimulated response element and IFN-gamma-activated sequence) within the 5.5URR and stimulated them with IFN-α/γ. The mutants showed a loss of response to IFN, indicating that the 5.5URR is essential for IFN-induced transcriptional enhancement in STAT1 gene expression. Second, we cloned the full-length 11 kb human STAT1 promoter, including the region upstream of the 5.5URR, from the start codon and linked it to a luciferase gene. Reporter assays showed that IFN-α/γ significantly activated the STAT1 promoter via the 5.5URR. Furthermore, recombinant DNA linking the full-length STAT1 promoter to STAT1 cDNA was introduced into STAT1-deficient cells. In vitro reconstitution experiments showed that IFN-α/γ stimulation increased STAT1 protein levels via the 5.5URR. These results demonstrate that the 5.5URR confers IFN-dependent autoregulation of the STAT1 promoter., (© 2024 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.)

Our investigation to explore cellular alterations related to undernutrition in cancer cells revealed that the protein level of heterogenous nuclear ribonucleoprotein A1 (hnRNP A1) is drastically decreased by serum/glucose starvation. Its loss was reversible, serum/glucose starvation-specific and universal throughout cell types and species. The hnRNP A1 mRNA level and hnRNP A1 mRNA/protein stability were not altered under this condition. CCND1 mRNA, which we newly identified as the binding target of hnRNP A1, was decreased by serum/glucose starvation. Under similar conditions, CCND1 protein was reduced in vitro and in vivo, whereas hnRNP A1 mRNA level and CCND1 mRNA level revealed no correlation in most clinical samples. Functional analyses revealed that CCND1 mRNA stability is certainly dependent on hnRNP A1 protein level and that RNA recognition motif-1 (RRM1) in hnRNP A1 plays a central role in maintaining CCND1 mRNA stability and subsequent protein expression. The injection of RRM1-deleted hnRNP A1-expressing cancer cells in the mouse xenograft model did not form any tumours, and that of hnRNP A1-expressing cancer cells retained CCND1 expression at the lesion adjacent to necrosis with a slight increase in tumour volume. Furthermore, RRM1 deletion caused growth suppression with the induction of apoptosis and autophagy, whereas CCND1 restoration completely recovered it. Our results indicate that serum/glucose starvation triggers entire hnRNP A1 protein loss, and its loss may play a role in CCND1 mRNA destabilization and CCND1-mediated cellular event inhibition, i.e. growth promotion, apoptosis induction and autophagosome formation., (© 2023 Federation of European Biochemical Societies.)

This study aimed to evaluate the effectiveness of acupuncture therapy in preventing emergence agitation (EA) in children. A systematic review and meta-analysis were conducted across multiple locations according to the articles searched. Seven databases, including trial registration sites, were searched. A total of six trials were included involving 489 patients; of them, 244 received acupuncture therapy. Randomized clinical trials (RCTs) evaluating the incidence of EA compared with placebo/sham or standard care in children were included. The primary outcome was the incidence of EA, as evaluated using a specific assessment tool. Data about the incidence rate of EA, heterogeneity, quality of trials and evidence, and adverse events were collected. Additionally, data about patient demographic characteristics, type of anesthesia, duration and onset of acupuncture therapy, EA and pain score, time taken for extubation, and post-anesthesia care unit length of stay were collected. The results indicated that the overall incidence of EA in the acupuncture therapy group and the control group was 23.4% and 39.5%, respectively, with no significant difference (risk ratio, 0.62; 95% confidence interval, 0.26-1.48; I2 = 63%). Subgroup analysis showed a significant difference in the overall incidence of EA in the acupuncture therapy and control groups according to surgery type (high-risk vs. low-risk surgery), suggesting that acupuncture therapy may be effective in reducing EA for patients undergoing high-risk surgery. The quality of evidence was downgraded to "very low" due to the study designs, inconsistency, and possible publication bias. In conclusion, this meta-analysis shows that the currently available RCTs are insufficient to determine the effectiveness of acupuncture therapy in preventing EA in children undergoing general anesthesia., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Mihara et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Limited data are available regarding part-time infectious disease consultations (IDCs) and their importance in tertiary care teaching hospitals in Japan. This is a retrospective review of IDCs from June 2016 to March 2021 and describes IDC services provided by part-time infectious disease specialists once a week for 4 hours, and their impact on the quality of medical care, including antimicrobial stewardship. Data, such as the requesting department, requesting reasons, and final diagnoses, were analyzed. In April 2018, part-time infectious disease specialists launched consultation services and attended an antimicrobial stewardship team conference. Meropenem, tazobactam/piperacillin, and cefepime monthly days of therapy (DOT) were calculated to assess the effect of each intervention; a pre-post analysis was conducted using the Kruskal-Wallis test. Additional quality improvement (QI) projects related to infectious diseases were implemented. There were 237 IDCs during the study period. Consultations were mostly requested by the General Internal Medicine, Emergency Medicine, and Cardiology departments. The most common diagnoses were bone/joint, respiratory, and genitourinary infections. Infectious disease services, even on a part-time basis, achieve good outcomes in patient management, antimicrobial stewardship, and QI projects. DOT/1000 patient-days were reduced for meropenem and cefepime, while it increased for tazobactam/piperacillin. The DOT/1000 patient-days for the 3-antipseudomonal agents significantly decreased during this period. After implementing the QI tetanus vaccination project in the Emergency Room, the number of tetanus toxoid vaccinations per month increased., Competing Interests: The authors have no funding and conflicts of interest to disclose., (Copyright © 2022 the Author(s). Published by Wolters Kluwer Health, Inc.)

Staphylococcus pettenkoferi is a coagulase-negative staphylococci (CoNS) species first isolated in 2002. Human infections caused by S. pettenkoferi are rare. We herein report three cases of S. pettenkoferi bacteremia in a tertiary care hospital in Japan. Staphylococcus pettenkoferi can be a causative pathogen of catheter related blood stream infection including complicated infection, and unknown source of bacteremia. All of the patients presented with fever and shaking chills, and good clinical outcome. Further research is needed to determine the role of this organism as a pathogen and frequency., (Copyright © 2022 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Virus-encoded microRNAs (miRNAs) target viral and host mRNAs to repress protein production from viral and host genes, and regulate viral persistence, cell transformation, and evasion of the immune system. The present study demonstrated that simian virus 40 (SV40)-encoded miRNA miR-S1 targets a cellular miRNA miR-1266 to derepress their respective target proteins, namely, T antigens (Tags) and telomerase reverse transcriptase (TERT). An in silico search for cellular miRNAs to interact with viral miR-S1 yielded nine potential miRNAs, five of which, including miR-1266, were found to interact with miR-S1 in dual-luciferase tests employing reporter plasmids containing the miRNA sequences with miR-S1. Intracellular bindings of miR-1266 to miR-S1 were also verified by the pull-down assay. These miRNAs were recruited into the Ago2-associated RNA-induced silencing complex. Intracellular coexpression of miR-S1 with miR-1266 abrogated the downregulation of TERT and decrease in telomerase activity induced by miR-1266. These effects of miR-S1 were also observed in miR-1266-expressing A549 cells infected with SV40. Moreover, the infected cells contained more Tag, replicated more viral DNA, and released more viral particles than control A549 cells infected with SV40, indicating that miR-S1-induced Tag downregulation was antagonized by miR-1266. Collectively, the present results revealed an interplay of viral and cellular miRNAs to sequester each other from their respective targets. This is a novel mechanism for viruses to manipulate the expression of viral and cellular proteins, contributing to not only viral lytic and latent replication but also cell transformation observed in viral infectious diseases including oncogenesis.

Pain, autonomic distress, and emergence agitation occur commonly in children undergoing general anesthesia. While acupuncture therapy has been reported to effectively reduce such pain and autonomic distress in children, its effect in preventing emergence agitation remains unclear. Therefore, we will conduct a systematic review and meta-analysis with trial sequential analysis to evaluate the effect of acupuncture therapy in preventing emergence agitation in children undergoing general anesthesia. Methods and analysis This protocol was prepared according to the 2015 Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) for Protocols guidelines. We will conduct a search for randomized controlled trials that evaluated the effect of acupuncture therapy in preventing emergence agitation. The following databases will be searched for relevant articles: MEDLINE, CENTRAL, Embase, and Web of Science; four pre-registration sites will be accessed from inception to April 1, 2021. No language restrictions will be applied. Two authors will independently scan and select eligible studies, extract the data, and assess the risk of bias. The incidence of emergence agitation will be combined as a risk ratio with a 95% confidence interval using a random-effect model. The I2 statistics will be used to assess heterogeneity. We will evaluate the quality of the clinical trials using the Cochrane methodology and assess the quality of evidence using the Grading of Recommendation Assessment, Development, and Evaluation approach. If appropriate, a trial sequential analysis will be performed. Expected outcomes This meta-analysis will be the first to evaluate the effect of acupuncture therapy in preventing emergence agitation in children. The findings from this meta-analysis have the potential to reveal pivotal factors that affect the clinical effect of acupuncture therapy, thereby supporting the optimization of acupuncture therapy for emergence agitation. Protocol registration University Hospital Medical Information Network Clinical Trials Registry (UMIN000040775)., Competing Interests: The authors have declared that no competing interests exist.

SV40-encoded microRNA (miRNA), miR-S1, downregulates the large and small T antigens (LTag and STag), which promote viral replication and cellular transformation, thereby presumably impairing LTag and STag functions essential for the viral life cycle. To explore the functional significance of miR-S1-mediated downregulation of LTag and STag as well as the functional roles of miR-S1, we evaluated viral DNA replication and proinflammatory cytokine induction in cells transfected with simian virus 40 (SV40) genome plasmid and its mutated form lacking miR-S1 expression. The SV40 genome encodes two mature miR-S1s, miR-S1-3p and miR-S1-5p, of which miR-S1-3p is the predominantly expressed form. MiR-S1-3p exerted strong repressive effects on a reporter containing full-length sequence complementarity, but only marginal effect on one harboring a sequence complementary to its seed sequence. Consistently, miR-S1-3p downregulated LTag and STag transcripts with complete sequence complementarity through miR-S1-3p-Ago2-mediated mRNA decay. Transfection of SV40 plasmid induced higher DNA replication and lower LTag and STag transcripts in most of the examined cells compared to that miR-S1-deficient SV40 plasmid. However, miR-S1 itself did not affect DNA replication without the downregulation of LTag transcripts. Both LTag and STag induced the expression of tumor necrosis factor α (TNFα) and interleukin (IL)-17F, which was slightly reduced by miR-S1 due to miR-S1-mediated downregulation of LTag and STag. Forced miR-S1 expression did not affect TNFα expression, but increased IL-17F expression. Overall, our findings suggest that miR-S1-3p is a latent modifier of LTag and STag functions, ensuring efficient viral replication and attenuating cytokine expression detrimental to the viral life cycle.

Inhibition of prostaglandin E 2 signaling via EP2/EP4 prostanoid receptors suppresses Insulin-like growth factor (IGF)-1-induced proliferation of pancreatic cancer BxPC-3 cells. To better understand the mechanism of EP2/EP4 signaling for controlling cell proliferation, we performed metabolome analyses in BxPC-3 cells treated with IGF-1 alone or IGF-1 plus EP2/EP4 inhibitors. These analyses revealed increased g-aminobutyric acid and 5-oxoproline production following the addition of EP2/EP4 inhibitors to IGF-1-treated cells. The expression of a 5-oxoproline-catalyzing enzyme, γ-glutamylcyclotransferase (GGCT), was also upregulated by IGF-1 treatment and further enhanced by the addition of EP2/EP4 inhibitors. Knockdown of GGCT expression resulted in the loss of suppressive effects of EP2/EP4 inhibitors on IGF-1-induced BxPC-3 cell proliferation, whereas GGCT overexpression repressed the basal proliferation of BxPC-3 cells but did not affect the suppressive effects of EP2/EP4 inhibitors. To summarize, we propose a role for EP2/EP4 signaling in regulating IGF-1-induced cell proliferation, in which EP2/EP4 signaling represses IGF-1-induced GGCT expression, which mediates and whose amount controls a branch of IGF-1 signaling to promote cell proliferation via extracellular signal-regulated kinase phosphorylation., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Background: Emergence agitation (EA) is a frequent phenomenon in children recovering from general anaesthesia and increases the risk of self-injury. Previously, our group reported that stimulating the heart 7 (HT7) acupuncture point bilaterally using two neuromuscular transmission monitoring devices (NTMs) decreased the incidence of EA. However, bilateral stimulation is a barrier to clinical use because two NTMs are needed for one patient., Objective: The objective of this study was to examine the efficacy of unilateral electrical stimulation of HT7 using an NTM to prevent EA in children., Design: Prospective, double-blinded, randomized clinical trial., Setting: Kanagawa Children's Medical Centre, Yokohama, Japan., Patients: One hundred children (ages 18-96 months) with ASA-PS I or II, who were scheduled to undergo inguinal hernia repair or orchiopexy under sevoflurane anaesthesia., Intervention: Patients were randomly assigned to one of the following two groups: (1) HT7 group: unilateral (right side) stimulation of the HT7 acupuncture point using a single-twitch electrical stimulus (1 Hz, 50 mA) throughout the surgery, and (2) control group: electrodes alone were attached to the HT7 point on the right side; an electrical stimulus was not applied., Main Outcome Measures: The primary outcome was the incidence of EA evaluated using the pediatric anaesthesia emergence delirium (PAED) scale. The secondary outcomes were the incidence of EA evaluated using Aono's scale, the severity of EA, PACU stay duration, and postoperative pain., Results: There was no statistical difference between the incidence of EA in the HT7 and the control group (28.0% and 24.0%, respectively; P > 0.99). The risk ratio was 1.17 (95% confidence interval: 0.60-2.27)., Conclusions: We observed that there was no effect of unilateral single-twitch electrical stimulation to the HT7 on the incidence of EA, contrary to the findings with bilateral HT7 stimulation., Competing Interests: The authors have declared that no competing interests exist.

Reactive arthritis after Group A streptococcal infection (poststreptococcal reactive arthritis: PSRA) that does not meet the Jones criteria for acute rheumatic fever (ARF) has been reported as a new entity for over a decade. In Japan there are few reports of PSRA. We encountered four children with arthritis accompanied with Group A streptococcal infection in our department. We investigated our cases and the recent Japanese literature. Japanese cases of PSRA are frequently accompanied with uveitis and erythema nodosum, and tonsillectomy resolved their symptoms in some cases. There were overlap cases between ARF, juvenile idiopathic arthritis, and PSRA., (Copyright © 2018 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Background and Aims: Tsumura-Suzuki obese diabetic (TSOD) is a good model of metabolic syndrome showing typical lesions found in nonalcoholic fatty liver disease and nonalcoholic steatohepatitis, and develops spontaneous hepatic tumors with a high frequency. Majority of the developing tumors overexpress glutamine synthetase (GS), which is used as a marker of hepatocellular carcinoma (HCC). The aim of this study is to assess the status of expression of metabolism-related genes and the level of bile acids in the TSOD mice-derived tumors and to determine the association with metabolic dysregulation between human HCC and TSOD mice-derived tumors., Methods: GS-positive hepatic tumors or adjacent normal tissues from 71-week-old male TSOD mice were subjected to immunohistochemical staining, quantitative RT-PCR (qRT-PCR), quantitation of cholic acid and taurocholic acid., Results: We found that downregulation of the rate-limiting enzyme for betaine synthesis (BADH), at both mRNA and protein levels in GS-positive TSOD mice-derived tumors. Furthermore, the bile acid receptor FXR and the bile acid excretion pump BSEP (Abcb11) were found to be downregulated, whereas BAAT and Akr1c14, involved in primary bile acid synthesis and bile acid conjugation, were found to be upregulated at mRNA level in GS-positive TSOD mice-derived tumors. BAAT and Akr1c14 were also overexpressed at protein levels. Total cholic acid was found to be increased in GS-positive TSOD mice-derived tumors., Conclusion: Our results strongly support the significance of TSOD mice as a model of spontaneously developing HCC.

Background: Emergence agitation (EA) is a common complication in children after general anesthesia. The goal of this 2-phase study was (1) to develop a predictive model (EA risk scale) for the incidence of EA in children receiving sevoflurane anesthesia by performing a retrospective analysis of data from our previous study (phase 1) and (2) to determine the validity of the EA risk scale in a prospective observational cohort study (phase 2)., Methods: Using data collected from 120 patients in our previous study, logistic regression analysis was used to predict the incidence of EA in phase 1. The optimal combination of the predictors was determined by a stepwise selection procedure using Akaike information criterion. The β-coefficient for the selected predictors was calculated, and scores for predictors determined. The predictive ability of the EA risk scale was assessed by a receiver operating characteristic (ROC) curve, and the area under the ROC curve (c-index) was calculated with a 95% confidence interval (CI). In phase 2, the validity of the EA risk scale was confirmed using another data set of 100 patients (who underwent minor surgery under general anesthesia). The ROC curve, the c-index, the best cutoff point, and the sensitivity and specificity at the point were calculated. In addition, we calculated the gray zone, which ranges between the two points where sensitivity and specificity, respectively, become 90%., Results: In phase 1, the final model of the multivariable logistic regression analysis included the following 4 predictors: age (logarithm odds ratios [OR], -0.38; 95% CI, -0.81 to 0.00), Pediatric Anesthesia Behavior score (logarithm OR, 0.65; 95% CI, -0.09 to 1.40), anesthesia time (logarithm OR, 0.60; 95% CI, -0.18 to 1.19), and operative procedure (logarithm OR, 2.53; 95% CI, 1.30-3.75 for strabismus surgery and logarithm OR, 2.71; 95% CI, 0.99-4.45 for tonsillectomy). The EA risk scale included these 4 predictors and ranged from 1 to 23 points. In phase 2, the incidence of EA was 39%. The c-index of phase 1 was 0.84 (95% CI, 0.74-0.94), and the c-index of phase 2 was 0.81 (95% CI, 0.72-0.89). The best cutoff point for the EA risk scale was 11 (sensitivity = 87% and specificity = 61%). The gray zone ranged from 10 to 13 points, and included 38% of patients., Conclusions: We developed and validated an EA risk scale for children receiving sevoflurane anesthesia. In our validation cohort, this scale has excellent predictive performance (c-index > 0.8). The EA risk scale could be used to predict EA in children and adopt a preventive strategy for those at high risk. This score-based preventive approach should be studied prospectively to assess the safety and efficacy of such a strategy.

Background: Emergence agitation is common in children recovering from general anaesthesia. The prevention of emergence agitation remains an important challenge in the field of paediatric anaesthesia., Objective: We aimed to examine the effectiveness of electrically stimulating the heart 7 (HT7) acupuncture site with a peripheral nerve stimulator (PNS) during surgery, for preventing emergence agitation in paediatric patients recovering from general anaesthesia., Design: A double-blind, randomised, controlled, parallel-group trial., Setting: Kanagawa Children's Medical Centre, Yokohama, Japan., Patients: One hundred and twenty patients aged 18 to 96 months (American Society of Anesthesiologists physical status I or II) undergoing minor elective surgery under general anaesthesia with sevoflurane., Intervention: Patients were randomly assigned to either undergo bilateral stimulation of HT7 with two PNS devices (1 Hz, 50 mA) during surgery (Group HT7) or a control group that did not undergo electrical stimulation of HT7 during surgery., Main Outcome Measures: The primary outcome was the incidence of emergence agitation evaluated in the postanaesthesia care unit (PACU) using the Paediatric Anaesthesia Emergence Delirium scale. The secondary outcomes were the time from operation completion to tracheal extubation, PACU stay duration and postoperative pain scores., Results: The incidence of emergence agitation was significantly lower in the HT7 group compared with the control group (31.7 vs. 56.7%, respectively; P = 0.010). The risk ratio was 0.56 (95% confidence interval 0.36 to 0.86) and the number needed to treat was 4 (95% confidence interval 3 to 13). There were no statistically significant differences between groups in time from operation completion to tracheal extubation, PACU stay duration or postoperative pain., Conclusion: Bilateral electrical stimulation of HT7 using two PNS devices significantly decreases the incidence of emergence agitation., Trial Registration: UMIN Clinical Trial Registry (registry number: UMIN000011704).

We previously identified a distal regulatory element located approximately 5.5-kb upstream of the signal transducer and activator of transcription 1 (STAT1) gene, thereafter designating it as 5.5-kb upstream regulatory region (5.5URR). In this study, we investigated the functional roles of 5.5URR in the transcriptional regulation of STAT1 gene. A chromosome conformation capture assay indicated physical interaction of 5.5URR with the STAT1 core promoter. In luciferase reporter assays, 5.5URR-combined STAT1 core promoter exhibited significant increase in reporter activity enhanced by forced STAT1 expression or interferon (IFN) treatment, but STAT1 core promoter alone did not. The 5.5URR contained IFN-stimulated response element and GAS sites, which bound STAT1 complexes in electrophoretic mobility shift assays. Consistently, chromatin immunoprecipitation (ChIP) assays of HEK293 cells with Halo-tagged STAT1 expression indicated the association of Halo-tagged STAT1 with 5.5URR. ChIP assays with IFN treatment demonstrated that IFNs promoted the recruitment of Halo-tagged STAT1 to 5.5URR. Forced STAT1 expression or IFN treatment increased the expression of endogenous STAT1 and other IFN signaling pathway components, such as STAT2, IRF9 and IRF1, besides IFN-responsive genes. Collectively, the results suggest that 5.5URR may provide a regulatory platform for positive feedback control of STAT1 expression possibly to amplify or sustain the intracellular IFN signals., (© 2015 The Molecular Biology Society of Japan and Wiley Publishing Asia Pty Ltd.)

Single-nucleotide polymorphisms associated with type 2 diabetes (T2D) have been identified in Jazf1, which is also involved in the oncogenesis of endometrial stromal tumors. To understand how Jazf1 variants confer a risk of tumorigenesis and T2D, we explored the functional roles of JAZF1 and searched for JAZF1 target genes in myogenic C2C12 cells. Consistent with an increase of Jazf1 transcripts during myoblast proliferation and their decrease during myogenic differentiation in regenerating skeletal muscle, JAZF1 overexpression promoted cell proliferation, whereas it retarded myogenic differentiation. Examination of myogenic genes revealed that JAZF1 overexpression transcriptionally repressed MEF2C and MRF4 and their downstream genes. AMP deaminase1 (AMPD1) was identified as a candidate for JAZF1 target by gene array analysis. However, promoter assays of Ampd1 demonstrated that mutation of the putative binding site for the TR4/JAZF1 complex did not alleviate the repressive effects of JAZF1 on promoter activity. Instead, JAZF1-mediated repression of Ampd1 occurred through the MEF2-binding site and E-box within the Ampd1 proximal regulatory elements. Consistently, MEF2C and MRF4 expression enhanced Ampd1 promoter activity. AMPD1 overexpression and JAZF1 downregulation impaired AMPK phosphorylation, while JAZF1 overexpression also reduced it. Collectively, these results suggest that aberrant JAZF1 expression contributes to the oncogenesis and T2D pathogenesis., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Background: Acute rheumatic fever (ARF) is an illness caused by group A streptococcus (GAS) infection, and remains the leading cause of acquired heart disease in worldwide. Distinguishing between ARF and septic arthritis may be difficult. This report describes a case of suppurative arthritis overlapping with ARF., Case Presentation: A 4-year-old, previously healthy boy presented with fever and left leg pain. The level of anti-streptolysin O (ASO) was elevated. His throat swab cultures grew GAS, but none were detected in his synovial fluid. Magnetic resonance imaging revealed suspected arthritis and osteomyelitis. The patient was treated for septic arthritis, but was subsequently diagnosed with ARF, after the development of carditis., Conclusion: The clinical and laboratory features of ARF and suppurative arthritis demonstrate substantial overlap. Patients with an elevated ASO should undergo a careful cardiac examination for carditis associated with ARF by an echocardiogram., (Copyright © 2015 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

MicroRNA-1 (miR-1) has recently been suggested to function as a tumor suppressor. Its functional relevance was assessed by exploring structural and tumorigenic properties of lung cancer A549 cells stably transduced with retrovirus containing pre-miR-1. A549 cells overexpressing miR-1 exhibited a significant morphological change from a mesenchymal to an epithelial phenotype characterized by cell polarization and intercellular junctions. The cells showed increased expression of E-cadherin, which colocalized with cortical actin filaments and vinculin to form typical adherens junction at the apical regions of intercellular borders. Additionally, they exhibited occludin-positive tight junctions at similar apical regions. Moreover, their migratory and invasive activities were inhibited, and their sensitivity to doxorubicin was increased slightly compared to control mock-infected cells. These structural and tumorigenic properties induced by miR-1 were associated with the reduced expression of Slug, which was a transcriptional repressor of E-cadherin or an inducer of epithelial-to-mesenchymal transition. Consistently, Slug was identified as a miR-1 target by bioinformatics and a luciferase reporter assay with plasmids containing luciferase-Slug 3'UTR. Collectively, the data presented here suggest that re-expression of miR-1 may be an effective therapy that prevents cancer malignancy by converting cells from a mesenchymal phenotype to an epithelial phenotype via the downregulation of Slug., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Chromatin loops formed between distant regulatory elements and promoters modulate gene expression. We identified a novel distant regulatory element located approximately 120kb downstream of the gls promoter, and examined its regulatory relevance to gls gene expression in C2C12 cells by a chromosome conformation capture assay. The distant element physically interacted with the gls promoter in myoblasts but not in myotubes. Semiquantitative analysis by real-time PCR showed more abundant gls transcripts in myoblasts than in myotubes. These findings suggest that this distant element differentially regulates gls gene expression through dynamic formation and abrogation of a chromatin loop during myogenesis., (Copyright © 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

In skeletal muscles, the sarcolemma is possibly stabilized and protected against contraction-imposed stress by intermediate filaments (IFs) tethered to costameric sarcolemma. Although there is emerging evidence that plectin links IFs to costameres through dystrophin-glycoprotein complexes (DGC), the molecular organization from plectin to costameres still remains unclear. Here, we show that plectin 1, a plectin isoform expressed in skeletal muscle, can interact with beta-synemin, actin and a DGC component, alpha-dystrobrevin, in vitro. Ultrastructurally, beta-synemin molecules appear to be incorporated into costameric dense plaques, where they seem to serve as actin-associated proteins rather than IF proteins. In fact, they can bind actin and alpha-dystrobrevin in vitro. Moreover, in vivo immunoprecipitation analyses demonstrated that beta-synemin- and plectin-immune complexes from lysates of muscle light microsomes contained alpha-dystrobrevin, dystrophin, nonmuscle actin, metavinculin, plectin and beta-synemin. These findings suggest a model in which plectin 1 interacts with DGC and integrin complexes directly, or indirectly through nonmuscle actin and beta-synemin within costameres. The DGC and integrin complexes would cooperate to stabilize and fortify the sarcolemma by linking the basement membrane to IFs through plectin 1, beta-synemin and actin. Besides, the two complexes, together with plectin and IFs, might have their own functions as platforms for distinct signal transduction.

The Japanese population is rapidly aging, thereby causing excess demand for facilities for elderly invalids. It is imperative that social measures and scientific studies be carried out to enable better care of bedridden elderly people. The purpose of the present study was to review the histological changes that occur in disuse atrophy of skeletal muscles, the primary pathophysiology of bedridden invalids, with the object of developing a staging standard to be used by researchers and clinicians. Rat hindlimb suspension was used as an experimental model. Atrophy of the soleus muscle was evaluated qualitatively and quantitatively on immunofluorescence microscopy. The myofibrils decreased significantly in the first 2-3 weeks of disuse atrophy. The earliest morphological change was fan-shaped multistep forking of sarcomeres, which appeared by the first week. This type of muscular lesion, designated here as 'sarcomeric disarray', was first described in the present study. Central-core lesions appeared mainly in slow muscle fibers by the second week. These lesions disappeared by the fourth or fifth week. Nerves remained intact and no inflammation or regeneration occurred up to the fifth week. Methods and criteria were compiled for staging of disuse atrophy based on the present results and a diagnosis kit designed for studies on disuse atrophy of skeletal muscles.

Escherichia coli is used as an indicator microorganism in public health. The conventional way to detect E. coli requires several days to produce a result, because it requires incubation of cells. Therefore a rapid and sensitive detection method is needed. T4e-/GFP phage, characterized by suppression of lysozyme and fusion of GFP (green fluorescent protein) to its SOC (small outer capsid) protein, was constructed, and it was shown to be able to detect E. coli K12 sensitively within several hours. However, because the host range of T4 phage to E. coli present in sewage water and sea water is narrow, this phage cannot be used to detect E. coli in environmental water. Two phages named IP008 and IP052, which have a broad host range to E. coli present in sewage influent, were screened from sewage influent. Mixture of these two phages produced clear plaques on 50% of E. coli screened from sewage influent. To use these phages as a tool for detection of E. coli, gfp was inserted into gene e, which encodes a lytic enzyme, and thus lytic-activity-suppressed phages were constructed (IP008e-/GFP and IP052e-/GFP). However, the fluorescent intensity of E. coli cells infected with IP008e-/GFP and IP052e-/GFP was not enough for visualization of the cell. Therefore, in addition to the insertion of gfp into gene e, fusion of GFP to SOC of IP008e-/GFP and IP052e-/GFP was conducted to produce IP008e-/2xGFP and IP052e-/2xGFP. E. coli cells infected with IP008e-/2xGFP and IP052e-/2xGFP showed much stronger fluorescence intensity than E. coli cells infected by IP008e-/GFP and IP052e-/GFP. It is anticipated that, using these GFP-labeled phages, a broad range of E. coli present in sewage influent water can be detected rapidly.

Background: Skeletal muscles are composed of heterogeneous collections of muscle fiber types, the arrangement of which contributes to a variety of functional capabilities in many muscle types. Furthermore, skeletal muscles can adapt individual myofibers under various circumstances, such as disease and exercise, by changing fiber types. This study was performed to examine the influence of dystrophin deficiency on fiber type composition of skeletal muscles in canine X-linked muscular dystrophy in Japan (CXMDJ), a large animal model for Duchenne muscular dystrophy., Methods: We used tibialis cranialis (TC) muscles and diaphragms of normal dogs and those with CXMDJ at various ages from 1 month to 3 years old. For classification of fiber types, muscle sections were immunostained with antibodies against fast, slow, or developmental myosin heavy chain (MHC), and the number and size of these fibers were analyzed. In addition, MHC isoforms were detected by gel electrophoresis., Results: In comparison with TC muscles of CXMDJ, the number of fibers expressing slow MHC increased markedly and the number of fibers expressing fast MHC decreased with growth in the affected diaphragm. In populations of muscle fibers expressing fast and/or slow MHC(s) but not developmental MHC of CXMDJ muscles, slow MHC fibers were predominant in number and showed selective enlargement. Especially, in CXMDJ diaphragms, the proportions of slow MHC fibers were significantly larger in populations of myofibers with non-expression of developmental MHC. Analyses of MHC isoforms also indicated a marked increase of type I and decrease of type IIA isoforms in the affected diaphragm at ages over 6 months. In addition, expression of developmental (embryonic and/or neonatal) MHC decreased in the CXMDJ diaphragm in adults, in contrast to continuous high-level expression in affected TC muscle., Conclusion: The CXMDJ diaphragm showed marked changes in fiber type composition unlike TC muscles, suggesting that the affected diaphragm may be effectively adapted toward dystrophic stress by switching to predominantly slow fibers. Furthermore, the MHC expression profile in the CXMDJ diaphragm was markedly different from that in mdx mice, indicating that the dystrophic dog is a more appropriate model than a murine one, to investigate the mechanisms of respiratory failure in DMD.

miR-1, miR-133a, and miR-206 are muscle-specific microRNAs expressed in skeletal muscles and have been shown to contribute to muscle development. To gain insight into the pathophysiological roles of these three microRNAs in dystrophin-deficient muscular dystrophy, their expression in the tibialis anterior (TA) muscles of mdx mice and CXMD(J) dogs were evaluated by semiquantitative RT-PCR and in situ hybridization. Their temporal and spatial expression patterns were also analyzed in C2C12 cells during muscle differentiation and in cardiotoxin (CTX)-injured TA muscles to examine how muscle degeneration and regeneration affect their expression. In dystrophic TA muscles of mdx mice, miR-206 expression was significantly elevated as compared to that in control TA muscles of age-matched B10 mice, whereas there were no differences in miR-1 or miR-133a expression between B10 and mdx TA muscles. On in situ hybridization analysis, intense signals for miR-206 probes were localized in newly formed myotubes with centralized nuclei, or regenerating muscle fibers, but not in intact pre-degenerated fibers or numerous small mononucleated cells, possibly proliferating myoblasts and inflammatory infiltrates. Similar increased expression of miR-206 was also found in C2C12 differentiation and CTX-induced regeneration, in which differentiated myotubes or regenerating fibers showed abundant expression of miR-206. However, CXMD(J) TA muscles contained smaller amounts of miR-206, miR-1, and miR-133a than controls. They exhibited more severe and more progressive degenerative alterations than mdx TA muscles. Taken together, these observations indicated that newly formed myotubes showed markedly increased expression of miR-206, which might reflect active regeneration and efficient maturation of skeletal muscle fibers.

Using murine models, we have previously demonstrated that recombinant adeno-associated virus (rAAV)-mediated microdystrophin gene transfer is a promising approach to treatment of Duchenne muscular dystrophy (DMD). To examine further therapeutic effects and the safety issue of rAAV-mediated microdystrophin gene transfer using larger animal models, such as dystrophic dog models, we first investigated transduction efficiency of rAAV in wild-type canine muscle cells, and found that rAAV2 encoding beta-galactosidase effectively transduces canine primary myotubes in vitro. Subsequent rAAV2 transfer into skeletal muscles of normal dogs, however, resulted in low and transient expression of beta-galactosidase together with intense cellular infiltrations in vivo, where cellular and humoral immune responses were remarkably activated. In contrast, rAAV2 expressing no transgene elicited no cellular infiltrations. Co-administration of immunosuppressants, cyclosporine and mycophenolate mofetil could partially improve rAAV2 transduction. Collectively, these results suggest that immune responses against the transgene product caused cellular infiltration and eliminated transduced myofibers in dogs. Furthermore, in vitro interferon-gamma release assay showed that canine splenocytes respond to immunogens or mitogens more susceptibly than murine ones. Our results emphasize the importance to scrutinize the immune responses to AAV vectors in larger animal models before applying rAAV-mediated gene therapy to DMD patients.