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BackgroundWhole-exome sequencing (WES) is a strong diagnostic tool for foetal structural anomalies, but the causative gene for more than half the anomalies have not been identified. Therefore, improving the diagnostic yield based on WES data is essential.MethodsFirst, 138 foetuses with structural anomalies were assessed using conventional WES and copy number variation (CNV) analyses. For undiagnosed cases, we employed a three-step approach for diagnosis. We re-evaluated 1) candidate variants using a loss-of-function observed/expected upper bound fraction (LOEUF) score, 2) all variants of disease-causing genes for clinically diagnosed cases using spliceAI, and 3) the rare variants in all low LOEUF scored genes (< 0.35) using spliceAI.ResultsWe identified molecular diagnoses in 53 of 138 cases (38.4%) using conventional WES and CNV. For undiagnosed cases, for the first step, we diagnosed two X-linked recessive diseases. For the second step, we diagnosed Meckel-Gruber syndrome by detecting likely pathogenic intron variant inTMEM67. In the third step, we identified ade novohemizygous pathogenic variant in one severe hydrops fetalis male, which caused aberrant splicing inCASK. We found a novel phenotype, hydrops fetalis, in CASK-related X-linked dominant disorder. Moreover, we revealed that the LOEUF score of X-linked disease-causing genes was significantly lower than that of autosomal genes among all OMIM-registered genes.ConclusionWe showed that the evaluation of variants, including introns of WES data, in combination with the LOEUF score, could improve the WES diagnostic yield and be useful for evaluation of variants, especially on chromosome X.What is already known on this topic?Molecular genetic diagnosis of foetal structural anomalies using WES is being increasingly implemented. However, more than half of the cases cannot be diagnosed. There is signigicant potential to increase the diagnostic yield by re-analysing WES data.What this study addsIn the present study, we focused on loss-of-function observed/expected upper bound fraction (LOEUF) scores to quantify genetic intolerance, and additional intron analysis for undiagnosed cases using conventional WES data. These approaches enabled the appropriate evaluation of candidate variants and detected overlooked candidate variants on intron. We diagnosed two X-linked recessive disorder cases (Hardikar syndrome and Ritscher-Schinzel syndrome), by re-evaluating candidate variants using the LOEUF score. We also diagnosed one Meckel-Gruber syndrome case caused by an intronic pathogenic variant, that had been overlooked by the conventional method. Moreover, evaluating all variants, including introns with low LOEUF score genes (2,971 genes) that could cause haploinsufficiency helped us find a pathogenic intronic variant onCASKin one hydrops fetalis case, which revealed that CASK-related X-linked dominant disorder could cause hydrops fetalis as severe phenotypes. Finally, the LOEUF score of X-linked genes was significantly lower than that of autosomal genes among all OMIM-registered genes, which meant that gene evaluation using the LOEUF score was helpful, for genetic diagnosis, especially for genes on chromosome X.How this study might affect research, practice, or policyThe evaluation of variants, including introns, in combination with the LOEUF score is expected to contribute to the improvement of the diagnostic yield in WES. These approaches are easy and convenient to implement. The LOEUF score might be useful for evaluation of variants, especially in chromosome X.

Milk lipids are an important energy source for infants, but the composition of milk lipids has not yet been clarified in detail. In this study, we analyzed free fatty acids and their metabolites in milk from humans and cows. In comparison to cow milk, human milk showed a higher content of free fatty acids including polyunsaturated fatty acids, especially ω-3 fatty acids and their metabolites. Polyunsaturated fatty acids were enriched at an early period of lactation, while saturated fatty acids did not change significantly over the period. Moreover, human milk contained high levels of ω-3 fatty acid metabolites, particularly 18-hydroxyeicosapentaenoic acid, an eicosapentaenoic acid-derived metabolite with anti-inflammatory activity. In comparison with human normal milk, thromboxane B

Background Systemic sclerosis (SSc; scleroderma) is an autoimmune connective tissue disease that affects the skin and subcutaneous tissue, in addition to the internal organs of the whole body. Onset in childhood is uncommon; however, both patients with childhood-onset and adult-onset SSc are positive for anti-nuclear antibodies (ANAs).Detection of SSc-related anti-nuclear antibodies is often useful for predicting clinical features, disease course, and outcomes. Case presentation A 5-year-old Japanese female manifested gradually progressive abnormal gait disturbance, regression of motor development, Raynaud’s phenomenon, and the shiny appearance of the skin of the face and extremities at age 2. On admission, she presented a mask-like appearance, loss of wrinkles and skin folds, puffy fingers, moderate diffuse scleroderma (18/51 of the modified Rodnan total skin thickness score), and contracture in the ankle and proximal interphalangeal joints. Grossly visible capillary hemorrhage on nail fold and severe abnormal capillaroscopy findings including bleeding, giant loop and disappearance of capillaryconsistent with the late phase in SSc. A skin biopsy showed fibrous thickening of the dermis, entrapment of an eccrine sweat glands, and thickened fiber. Chest high-resolution computed tomographic scanning demonstrated patchy areas of ill-defined air-space opacity and consolidation predominantly involving the posterior basilar aspects of the lower lobes presenting withinterstitial lung disease. Positive ANA (1:160 nucleolar and homogeneous nuclear staining by indirect fluorescent antibody technique) and double-seropositive for anti-Th/To and anti-PM-Scl antibodies were identified. She was diagnosed with diffuse cutaneous SSc based on the Pediatric Rheumatology European Society/American College of Rheumatology/European League Against Rheumatism Provisional Classification Criteria for Juvenile Systemic Sclerosis and was successfully treated with immunosuppressive agents, including methylprednisolone pulses and intravenous cyclophosphamide. Conclusions We experienced the first case of juvenile SSc with anti-PM-Scl and anti-Th/To antibodies. ILD was identified as a typical feature of patients with these autoantibodies; however, diffuse cutaneous SSc and joint contraction were uncharacteristically associated. The case showed unexpected clinical findings though the existence of SSc-related autoantibodies aids in determining possible organ involvement and to estimate the children’s outcome.

Objective To develop a multianalyte assay for the detection of dermatomyositis (DM)-related autoantibodies using immunoprecipitation (IP) combined with immunoblotting (IB). Methods Sera from 116 DM patients were subjected to RNA and protein immunoprecipitation assays as well as commercial enzyme-linked immunosorbent assays (ELISAs) for anti-aminoacyl transfer RNA synthetase, anti-melanoma differentiation antigen 5 (MDA5), anti-Mi-2, anti-transcriptional intermediary factor-1γ (TIF-1γ), and anti-U1 ribonucleoprotein antibodies. The IP/IB assay was developed by immunoprecipitation of autoantigens from HeLa cell extracts using patient sera, followed by immunoblotting with an antibody against Mi-2, TIF-1γ, OJ, nuclear matrix protein (NXP)-2, MDA5, PM/Scl, small ubiquitin-like modifier activating enzyme (SAE), or Ku. A multianalyte assay was designed by mixing primary antibodies in the IP/IB assay. Results IP assays identified any DM-related autoantibodies in 100 patients (86%), of which 82% were covered by commercial ELISAs, with a false-positive result in two sera and a false-negative result in one serum. The results obtained from the multianalyte IP/IB assay and ‘gold-standard’ IP assays were concordant in terms of the presence or absence of anti-MDA5, anti-TIF-1γ, anti-OJ, anti-NXP-2, anti-PM/Scl, anti-SAE, anti-Mi-2, and anti-Ku antibodies. Conclusion This multianalyte IP/IB assay combined with commercial ELISAs is an alternative to ‘gold-standard’ IP assays for the detection of DM-related autoantibodies.

Our smartphone application–based analysis of 6 million menstrual cycles among Japanese females found age-dependent and seasonal changes of cycle length and basal body temperature., OBJECTIVE: To evaluate the effects of age and season on menstrual cycle length and basal body temperature (BBT). We also examined the effects of climate on cycle length and BBT, taking into account Japanese geographic and social characteristics. METHODS: In this retrospective cohort study, we analyzed data from 6 million menstrual cycles entered into a smartphone application from 310,000 females from 2016 to 2017. Only those who entered more than 10 cycles in 2 years were included. Generalized estimation equations were used to adjust for confounding factors and for within-person correlations of multiple records. Multiple regression analysis was conducted, with age, external average temperature, precipitation amount, and sunshine hours as confounding factors. RESULTS: The mean menstrual cycle length increased from age 15–23 years, subsequently decreased up to age 45 years, and then increased again. Average follicular phase body temperature showed no significant age-dependent changes, but luteal phase body temperature gradually increased up to 29 years and then stabilized and started to decrease after age 42 years. A significant association between external temperature and body temperature (follicular and luteal phase) was observed, though menstrual cycle length did not show such an association. CONCLUSION: These results, derived from data self-entered into a smartphone application, revealed underrecognized age-dependent and seasonal changes in menstrual cycle length and BBT, which will contribute to a better understanding of female reproductive health in the modern world.

It is important to evaluate the effects of drugs considered to control hemorrhage. Tranexamic acid (TXA) has been shown to reduce the risk of death in bleeding trauma patients. Carbazochrome sodium sulfonate (CSS) is often used in combination with TXA; however, it is unknown whether CSS additionally improves the control of bleeding in trauma patients.The aim of this study was to examine whether CSS reduces blood transfusion and death in addition to TXA by improving the control of bleeding. We retrospectively analyzed medical records of trauma patients from 2011 to 2019. We included patients aged ≥16 years, with significant hemorrhage, and who received TXA within eight hours from injury as per CRASH-2 (Clinical Randomisation of an Antifibrinolytic in Significant Haemorrhage) study. The primary outcome was the total amount of red blood cells (RBC), fresh frozen plasma (FFP), and platelet concentrate (PC) received within the first 24 hours from injury. Secondary outcomes were death in hospital within four weeks after injury, vascular occlusive events, and treatment.During this retrospective evaluation period, 5764 admissions with trauma were registered. A total of 326 cases met the selection criteria: 259 cases who received CSS in addition to TXA (CSS group; n=259) and 67 cases who received only TXA (no-CSS group; n=67). The mortality rate was 6% in the no-CSS group and 15.1% in the CSS group. There was no significant difference in mortality and vascular occlusive events between the two groups. We performed multiple regression analyses, with the amount of blood transfusion for each type as explanatory variables. The administration of CSS was an independent factor for the reduction of RBC transfusion (standard partial regression coefficient -0.1, 95% CI [-3.1 to -0.1], p=0.04), but not for transfusion of FFP or PC. We also performed multiple logistic regression analysis, with death as an explanatory variable. CSS was not an independent factor for any cause of death.CSS decreased RBC transfusion in trauma patients, without increasing the risk of vascular occlusion. However, CSS did not decrease mortality. This study can contribute to managing bleeding with trauma, but further research aimed at clarifying the effect of CSS is needed.

Malnutrition and sarcopenia often coexist in rehabilitation patients, although they are often overlooked and undertreated in clinical practice. This cross-sectional study aimed to clarify the prevalence of the coexistence of malnutrition and sarcopenia (Co-MS) and its associated factors in convalescent rehabilitation wards in Japan. Consecutive patients aged ≥ 65 years in convalescent rehabilitation wards between November 2018 and October 2020 were included. Malnutrition and sarcopenia were determined by the Global Leadership Initiative on Malnutrition (GLIM) criteria and the Asian Working Group for Sarcopenia (AWGS 2019) criteria, respectively. Patients who presented both with malnutrition and sarcopenia were classified as Co-MS. Potentially associated factors included age, sex, days from onset to admission of rehabilitation wards, reason for admission, pre-morbid functional dependency, comorbidity, activities of daily living, swallowing ability, and oral function and hygiene. The prevalence of malnutrition, sarcopenia, and Co-MS was calculated. Binary logistic regression analyses were performed to compute odds ratios (ORs) and the 95% confidence interval (CI) of possible associated factors for each condition. Overall, 601 patients were eligible for the analysis (median 80 years old, 355 female patients, 70% cerebrovascular disease). Co-MS, malnutrition, and sarcopenia were found in 23.5%, 29.0%, and 62.4% of the enrolled patients, respectively. After adjustment, onset–admission interval (OR = 1.04, 95% CI = 1.02 to 1.06), hospital-associated deconditioning (OR = 4.62, 95% CI = 1.13 to 18.8), and swallowing ability (Food Intake LEVEL Scale) (OR = 0.83, 95% CI = 0.73 to 0.93) were identified as independent explanatory factors of Co-MS. In conclusion, Co-MS was prevalent in geriatric rehabilitation patients, thus, healthcare professionals should be aware of the associated factors to detect the geriatric rehabilitation patients who are at risk of both malnutrition and sarcopenia, and to provide appropriate treatments.

This study aimed to investigate the association between muscle strength and adjusted appendicular skeletal muscle mass (ASM) in patients who have had strokes with the Functional Independence Measure (FIM) and the probability of being discharged.A retrospective cohort study was conducted for older patients who have had strokes admitted to convalescent rehabilitation wards between January 2017 and October 2020. Hand-grip strength (HGS) was used to assess muscle strength. ASM was measured with a bioelectrical impedance analysis, and then divided by height-squared, body weight, body mass index (BMI), body fat mass (BFM), and body fat percentage (BFP) to calculate the adjusted ASM. The primary outcome was FIM at the time of discharge, and the secondary outcome was the probability of being discharged to their home. Multivariate analyses were conducted to adjust for confounding effects.The data of 699 participants (female: 47%; median age, 79 y) were analyzed. HGS was independently associated with FIM at the time of discharge in men (partial regression coefficient [B] = 0.482; 95% confidence interval [CI], 0.225-0.740) and women (B = 0.664; 95% CI, 0.263-1.065) and also was independently associated with being discharged to their home in men (odds ratio [OR]: 1.070; 95% CI, 1.030-1.100) and women (OR: 1.070; 95% CI, 1.000-1.130). Conversely, none of the adjusted ASM indices were associated with the outcomes. The cutoff value of HGS for discharge to home was 15.1 kg for men and 9.5 kg for women.In patients who have had strokes, HGS independently predicted FIM at the time of discharge and the probability of being discharged to their home. The adjusted ASM methods had less predictive value for functional and discharge outcomes.

Maternal glucocorticoid exposure increases the risk of preterm delivery; however, the association between glucocorticoids and preterm premature rupture of membranes (pPROM)-a direct cause of preterm delivery-has rarely been investigated.To examine this association, we evaluated the clinical data of patients with systemic lupus erythematosus (SLE). Mechanism analysis was performed in both human amnion-derived mesenchymal cells (as a model for fetal membranes) and the amnion from SLE patients. We characterized the effects of glucocorticoids on the amnion in both models through comprehensive gene expression profiling and by electric cell-substrate impedance sensing in the mesenchymal cells.The average glucocorticoid dose in cases with pPROM (13.3 mg/day, n = 10) was significantly higher than in those without pPROM (8.5 mg/day, n = 65; P 0.01) among pregnant patients with well-controlled SLE (SLEDAI4, n = 75); however, we did not observe a statistically significant difference in it between cases with or without chorioamnionitis. Glucocorticoid-treated human amnion mesenchymal cells showed decreased electric resistance between cells, indicating increased permeability. Differentially expressed genes upon glucocorticoid treatment were significantly enriched with cell adhesion-related genes. Among them, ITGA8 was strikingly induced in both the amnion mesenchymal cells and in amnion derived from patients with SLE.We observed an association between glucocorticoids and pPROM with non-infectious etiology. Our findings indicate that glucocorticoids increase amnion permeability and modulate cell-adhesion related genes. ITGA8 represents a primary molecule that triggers pPROM through fibrotic remodeling and preventing resealing of the rupture site in fetal amnion.

Aim To clarify the correlation between malnutrition, muscle mass and oral status, and swallowing function recovery in stroke patients receiving enteral nutrition. Methods Patients with stroke and dysphagia receiving any amount of enteral nutrition in rehabilitation wards from 2012 to 2016 were eligible for inclusion in this retrospective study. On admission, body composition by bioimpedance analysis, malnutrition confirmed by the European Society for Clinical Nutrition and Metabolism criteria, oral status, functional independence measure and demographic data were collected. Characteristics were compared between "oral intake alone" and "artificial nutrition" groups based on the discharge status. Kaplan-Meier methods and the Cox proportional hazards model were used to determine explanatory factors for the probability of full oral intake. Results Among 174 patients, 113 were analysed (55 women; median age, 77 years). Overall, 61% and 39% were classified as "oral intake alone" and "artificial nutrition," respectively. Days from onset to admission to rehabilitation wards and motor Functional Independence Measure were higher in the "oral intake alone" group. Kaplan-Meier analysis demonstrated that patients with lower muscle mass exhibited lower probability of full oral intake (P = .009). The Cox proportional hazards model suggested that lower muscle mass (hazard ratio, 0.493; 95% CI, 0.286-0.850) and poor oral hygiene (hazard ratio, 0.573; 95% CI, 0.333-0.987) were independently correlated with "oral intake alone" status. Malnutrition and other oral status are not related to achieving full oral intake. Conclusions Skeletal muscle mass and oral hygiene are independently correlated with full oral intake among stroke patients receiving enteral nutrition during the rehabilitation phase.

This cross-sectional study investigated the proportion of patients’ recovery from sarcopenia status and the relationship between improvement in sarcopenia (IS) and function and discharge outcome in hospitalized patients with stroke. This study included patients with stroke, aged 65 years or more, with a diagnosis of sarcopenia, who were admitted to a convalescent rehabilitation ward. Sarcopenia was diagnosed according to the Asian Working Group for Sarcopenia 2019 criteria. Patients were divided according to the presence or absence of sarcopenia at discharge: IS group and non-improvement in sarcopenia (NIS) group. Among the 227 participants (mean age: 80.5 years, 125 females), 30% (69/227) of the patients were in the IS group, while 70% (158/227) were in the NIS group. The IS group showed a higher Functional Independence Measure (FIM) than the NIS group (median 112 vs. 101, p = 0.003). The results demonstrated that IS was independently associated with higher FIM (partial regression coefficient, 5.378, 95% confidence interval (CI), 0.709–10.047). The IS group had higher odds of home discharge than the NIS group (odds ratio, 2.560, 95% CI, 0.912–7.170). In conclusion, recovery from sarcopenia may be associated with better function in patients with stroke.

Objective To evaluate upstream and downstream regulators leading to macrophage activation and subsequent cytokine storm in patients with anti-melanoma differentiation-associated gene 5 (MDA5) antibody-associated interstitial lung disease (ILD). Methods We conducted an integrated miRNA-mRNA association analysis using circulating monocytes from 3 patients with anti-MDA5-associated ILD and 3 healthy controls and identified disease pathways and a regulator effect network by Ingenuity Pathway Analysis (IPA). The expression of relevant genes and proteins was verified using an independent validation cohort, including 6 patients with anti-MDA5-associated ILD, 5 with anti-aminoacyl tRNA synthetase antibody-associated ILD, and 6 healthy controls. Results IPA identified 26 matched pairs of downregulated miRNA and upregulated mRNAs and revealed that canonical pathways mediated by type I IFN signalling and C-C motif ligand 2 (CCL2) were responsible for the pathogenic process (P < 0.05 for all pathways). The regulatory network model identified IFN-β; Toll-like receptors 3, 7, and 9; and PU.1 as upstream regulators, while the downstream effect of this network converged at the inhibition of viral infection. mRNA and protein expression analysis using validation cohort showed a trend towards the increased expression of relevant molecules identified by IPA in patients with anti-MDA5-associated ILD compared with those with anti-aminoacyl tRNA synthetase antibody-associated ILD or healthy controls. The expression of all relevant genes in monocytes and serum levels of CCL2 and IFN-β declined after treatment in survivors with anti-MDA5-associated ILD. Conclusion An antiviral proinflammatory network orchestrated primarily by activated monocytes/macrophages might be responsible for cytokine storm in anti-MDA5-associated ILD.

Antiphospholipid antibody syndrome (APS) is defined by the presence of clinical symptoms caused by antiphospholipid antibodies. When APS occurs during pregnancy, it is conventionally treated with low-dose aspirin or heparin. In cases refractory to conventional treatment, intravenous immunoglobulin (IvIg) is sometimes added. We present the case of an APS patient with severe thrombocytopenia who experienced a successful pregnancy after treatment that included intravenous rituximab and IvIg. As far as we know, this is the first report demonstrating a positive pregnancy outcome in this context. Physicians may consider prescribing not only IvIg but also rituximab during the first trimester of pregnancy in APS patients with severe obstetrical complications and thrombocytopenia refractory to conventional treatment.

Deep learning has rapidly been filtrating many aspects of human lives. In particular, image recognition by convolutional neural networks has inspired numerous studies in this area. Hardware and software technologies as well as large quantities of data have contributed to the drastic development of the field. However, the application of deep learning is often hindered by the need for big data and the laborious manual annotation thereof. To experience deep learning using the data compiled by us, we collected 2429 constrained headshot images of 277 volunteers. The collection of face photographs is challenging in terms of protecting personal information; we therefore established an online procedure in which both the informed consent and image data could be obtained. We did not collect personal information, but issued agreement numbers to deal with withdrawal requests. Gender and smile labels were manually and subjectively annotated only from the appearances, and final labels were determined by majority among our team members. Rotated, trimmed, resolution-reduced, decolorized, and matrix-formed data were allowed to be publicly released. Moreover, simplified feature vectors for data sciences were released. We performed gender and smile recognition by building convolutional neural networks based on the Inception V3 model with pre-trained ImageNet data to demonstrate the usefulness of our dataset.

Objective: Familial nonautoimmune hyperthyroidism (FNAH) is a rare disease. To date there are few, if any, reports of pregnancies in women with FNAH. Our objective here is to present such a case. Methods: Free thyroxine (free T4), free triiodothyronine (free T3), thyroid-stimulating hormone (TSH), and antibodies related to the thyroid were measured. Fetal thyroid function indicators including thyroid volume and ossification were checked using ultrasound. Thyroid-stimulating hormone receptor (TSHR) gene analyses were performed. Results: The patient was a 30-year-old woman with no past medical history. She was introduced to our hospital in the fifth gestational week for pregnancy care because her family history revealed that her mother had nonautoimmune hyperthyroidism with a TSHR-activating germ-line mutation (Asn406Ser). The serum free T4 was 1.88 ng/dL (normal, 0.62 to 1.19 ng/dL), free T3 was 3.27 pg/mL (normal, 2.55 to 3.88 pg/mL), TSH was 0.02 μIU/mL (normal, 0.007 to 3.619 μIU/mL), and TSHR was negative which were considered to be consistent with mild primary hyperthyroidism. Serum free T4, free T3, and TSH concentrations were monitored every 4 to 6 weeks with a peak free T4 of 2.23 ng/dL noted at gestational week 9. The patient had no signs related to hyperthyroidism throughout pregnancy. The patient delivered a 3,518 g girl at 40 weeks of gestation. Genetic analysis of her TSHR gene showed heterozygous Asn406Ser mutation. The offspring did not show any signs of prenatal hyperthyroidism, and thyroid function at day 6 after delivery revealed a free T4 of 2.41 ng/dL (normal, 1.83 to 2.91 ng/dL) and a TSH of 3.55 μIU/mL (normal, 0.51 to 4.57 μIU/mL). Conclusion: Women with FNAH and mild thyrotoxicosis prior to pregnancy may have continuous hyperthyroidism with additional change due to the series of human chorionic gonadotropin secretion during pregnancy.

We describe the immunologic findings and the long-term clinical course of 43-year-old woman with classic dermatomyositis and associated rapidly progressive and relapsing course of interstitial lung...

Aim Hospital selection for patients with drug overdose (DOD) is a critical issue. In Tokyo, the 50‐tablet rule has been widely utilized by paramedics for triaging patients with DOD given that it shortens the triage time. However, studies have not investigated the utility of such a rule in local cities. The present study aimed to identify prognostic pre‐ and in‐hospital factors among patients with DOD and determine whether the 50‐tablet rule benefits local cities. Methods Clinical records of patients with DOD admitted at the University of Tsukuba Hospital (Tsukuba, Japan) between 2013 and 2017 were retrospectively reviewed. Results A total of 93 patients with DOD were enrolled in this study. Multivariate analysis showed that creatine kinase and C‐reactive protein values and the total number of ingested pills were in‐hospital risk factors that significantly prolonged intensive care unit (ICU) and hospital stay. Moreover, Glasgow Coma Scale score on admission and the total number of ingested pills were identified as significant pre‐hospital risk factors for prolonged ICU and hospital stay. Setting the total number of ingested pills to 50 tablets did not significantly influence ICU and hospital stay. Conclusion The total number of ingested pills and creatine kinase and C‐reactive protein values were identified as predictive factors for prolonged ICU and hospital stay in patients with DOD after admission. Moreover, pre‐hospital risk factors included Glasgow Coma Scale score and the total number of ingested pills. However, the 50‐tablet rule was determined to be a poor cut‐off value for patients with DOD. We presented a subset of our findings at the 46th annual meeting of the Japanese Society for Acute Medicine (Yokohama, Japan) on 20 November 2018., The total number of ingested pills and creatine kinase and C‐reactive protein values were identified as predictive factors for prolonged intensive care unit and hospital stay in patients with drug overdose after admission. Moreover, pre‐hospital risk factors included Glasgow Coma Scale score and the total number of ingested pills. However, the 50‐tablet rule was determined to be a poor cut‐off value for patients with drug overdose.

The presence of anti-aminoacyl tRNA synthetase (ARS) or anti-melanoma differential-associated gene 5 (MDA5) is strongly related to interstitial lung disease (ILD) in patients with dermatomyositis (DM). Several studies suggest a potential relationship between ILD and anti-small ubiquitin-like modifier activating enzyme (SAE) antibody in DM patients, but detailed clinical characteristics of anti-SAE-associated ILD still remain unknown. We have experienced 2 cases who were positive for anti-SAE antibody, who presented with ILD in the context of clinically amyopathic DM. These 2 patients had the following common ILD characteristics: an insidious course with preserved pulmonary function; a limited extent of pulmonary lesions with subpleural peripheral-dominant small ground glass opacity/consolidation on high-resolution computed tomography; and a favorable treatment response. These findings suggest that anti-SAE-associated ILD is unique in terms of clinical and imaging features and differs from ILD associated with anti-ARS or anti-MDA5 antibody.

BackgroundAttention facilitates gating of information from the sending brain area to receiving areas, with this being achieved by dynamical change in effective connectivity between cortices. However, it is difficult to assess effective connectivity, which refers to the causal influence of one cortical area on another.ObjectiveThe aim of this study is to directly probe the effective connectivity between cortical areas, which is modulated by covertly shifted attention, excluding the thalamic influence.MethodsTranscranial magnetic stimulation (TMS) was used to directly perturb the right retinotopic visual cortex in task-relevant (TR) and task-irrelevant (TIR) networks, and the impact of this was tracked to other areas by concurrent use of electroencephalography (EEG).ResultsKey finding was that TMS to the TR hemisphere led to a stronger evoked potential than did stimulation to the TIR hemisphere. Moreover, stronger beta- and gamma-band effective connectivities assessed as lagged phase synchronizations between stimulated areas and other areas were observed when TMS was delivered to the TR area. These effects were more enhanced when they preceded more prominent alpha lateralization, which is known to be associated with gating information.ConclusionOur results indicate that attention-regulated cortical excitability and feedforward dynamical effective connectivity can be probed by direct cortical stimulation to the TR or TIR area, thereby bypassing thalamic gating. These results bear out the idea that TMS-EEG could help to characterize changes in the functional architecture of brain networks that are required for rapid adaptation to the environment or due to brain diseases.HighlightsAttention-regulated cortical excitability and effective connectivity were probed by TMS-EEG.Stronger TMS evoked potentials were observed in the task-relevant visual hemisphere.Strong effective connectivity between task-relevant visual cortex and other areas were found.

We clarify clinical characteristics of patients with immune checkpoint inhibitor (ICI)-induced myositis. In 13 of 15 cases with ICI-induced myositis, the type of malignancy was melanoma. Eight, 4, and 3 patients received anti-PD-1 alone, anti-CTLA4 alone, and a combination of those, respectively. The mean period to the onset of ICI-induced myositis from the initiation of ICI was 4 weeks. Myocarditis was a complication in five patients. Seven of the patients died. The causes of death were myocarditis in three patients, respiratory muscle paralysis in two patients, and cancer progression in two patients. In patients without myocarditis or respiratory muscle paralysis, the prognosis for myositis was favorable with normalization of the CK levels occurring upon the cessation of ICI and the administration of immunosuppressive agents. Myocarditis and respiratory muscle paralysis are the major causes of death as immune-related adverse events in patients with ICI-induced myositis.

Despite the widespread use of transcranial magnetic stimulation (TMS), knowledge of its neurophysiological mode of action is still incomplete. Recently, TMS has been proposed to synchronise neural oscillators, and to thereby increase the detectability of corresponding oscillations at the population level. As oscillations in the human brain are known to interact within nested hierarchies via phase-amplitude coupling, TMS might also be able to increase the macroscopic detectability of such coupling. In a concurrent TMS-electroencephalography study, we therefore examined the technique’s influence on theta-gamma, alpha-gamma and beta-gamma phase-amplitude coupling by delivering single-pulse TMS (sTMS) and repetitive TMS (rTMS) over the left motor cortex and right visual cortex of healthy participants. The rTMS pulse trains were of 5 Hz, 11 Hz and 23 Hz for the three coupling variations, respectively. Relative to sham stimulation, all conditions showed transient but significant increases in phase-amplitude coupling at the stimulation site. In addition, we observed enhanced coupling over various other cortical sites, with a more extensive propagation during rTMS than during sTMS. By indicating that scalp-recorded phase-amplitude coupling can be effectively probed with TMS, these findings open the door to the technique’s application in manipulative dissections of such coupling during human cognition and behaviour in healthy and pathological conditions.

Background:Anti-Ro/SS-A (anti-SS-A) antibody is one of myositis-associated antibodies, and is found in patients with anti-synthetase antibodies1. Anti-SS-A antibody targets the complex consisting of Ro52 and Ro60 proteins coupled with cytoplasmic non-coding Y-RNAs. Autoantibody against Ro52 uncoupled with Y-RNAs (anti-uncoupled Ro52) is also present in patients with a variety of connective tissue diseases, including myositis2. However, the majority of previous studies used enzyme-linked immunoassay (EIA) for detection of anti-Ro52 antibodies, resulting in failure to discriminate between anti-Ro52 antibodies coupled and uncoupled with Y-RNAs. The prevalence and clinical significance of anti-uncoupled Ro52 antibodies still remain unclear in patients with anti-synthetase antibodies.Objectives:To elucidate clinical relevance of anti-uncoupled Ro52 antibodies in a cohort of anti-synthetase syndrome employing RNA immunoprecipitation assay (RNA-IP) in combination with EIA.Methods:This is a single-center, cross-sectional study involving 80 patients positive for anti-synthetase antibodies by RNA-IPP. Complete clinical information was obtained from a medical chart review. Serum samples were obtained at first visit and stored at -20°C until use. Anti-SS-A and anti-SS-B antibodies were detected by RNA-IP, and anti-Ro52 and anti-Ro60 antibodies were measured by commercial EIA kits (ORGENTIC, Mainz, Germany). Autoantibodies that immunoprecipitated Y-RNAs regardless of results of anti-anti-Ro52 or anti-Ro60 EIAs were regarded as anti-SS-A antibody, while antibodies that did not immunoprecipitate Y-RNAs but reacted with anti-Ro52 antibodies by EIA were regarded as anti-uncoupled Ro52 antibody. Student’s t-test, Mann-Whitney’s U test, and Fisher’s exact test were employed to compare the clinical features between each group. Cumulative survival rates were compared using log-rank test.Results:In our cohort of 80 patients with anti-synthetase antibody, mean age at diagnosis was 61 ± 12 years, and 76% were female. Clinical diagnosis was classic dermatomyositis (cDM) in 11, clinically amyopathic dermatomyositis (CADM) in 21, polymyositis (PM) in 11, systemic sclerosis (SSc) in 3, myositis-SSc overlap in 5, interstitial lung disease (ILD) alone in 29, and unclassified in 3. The antigenic specificity of anti-synthetase antibodies included Jo-1 in 19, PL-7 in 12, PL-12 in 9, EJ in 21, OJ in 4, and KS in 16. Anti-SS-A anti-SS-B antibodies were found in 14 (17%) and 2 (2.5%) patients, respectively. The presence of anti-Ro60 and anti-SS-A antibodies was almost concordant (P < 0.0001), although the presence of anti-Ro52 and anti-SS-A antibodies was not correlated (P = 0.8). This was primarily because of high prevalence (40%) of autoantibodies to uncoupled Ro52. Interestingly, prevalence of anti-uncoupled Ro52 antibodies was different among antigenic specificities of anti-synthetase antibodies: high in Jo-1 (58%) and EJ (55%), and low in PL-7 (8%) and OJ (0%). Gottron’s sign/papule was more frequent in patients with anti-uncoupled Ro52 than in those without (61% versus 28%, P = 0.005), resulting in clinical diagnosis of cDM or CADM more common in patients with anti-uncoupled Ro52 than in those without (59% versus 26%; P = 0.003). The prevalence and extent of ILD tended to be greater in anti-uncoupled Ro52-positive versus negative patients, but difference did not reach statistical significance. There were no differences in cumulative survival rates between patients stratified by the presence or absence of anti-uncoupled Ro52 antibodies.Conclusion:Autoantibodies to uncoupled Ro52 were commonly found in patients with anti-synthetase antibodies. Anti-Ro52 positivity might be useful for subclassifying anti-synthetase syndrome.References:[1]McHugh NJ et al.Nat Rev Rheumatol. 2018;14(5): 290-302.[2]Schulte-Pelkum J et al.Autoimmun Rev. 2009;8(7): 632-7.Disclosure of Interests:Akira Yoshida: None declared, Yuka Okazaki: None declared, Takahisa Gono Speakers bureau: Astellas, and Medical and Biological Laboratories, Masataka Kuwana Grant/research support from: Acetelion, Consultant of: Acetelion, Bayer, Chugai, Corbus Pharmaceuticals, CSL Behring and Reata Pharmaceuticals. He was a member of the SENSCIS trial Steering Committee (Boehringer Ingelheim)

Neural oscillations are ubiquitous throughout the cortex, but the frequency of oscillations differs from area to area. To elucidate the mechanistic architectures establishing various rhythmic activities, we tested whether spontaneous neural oscillations in different cortical modules can be entrained by direct perturbation with distinct frequencies of transcranial magnetic stimulation (TMS). While recording the electroencephalogram (EEG), we applied single-pulse TMS (sTMS) and repetitive TMS (rTMS) at 5, 11, and 23 Hz to motor or visual cortex. To assess entrainment, defined as phase locking of intrinsic oscillations to periodic external force , we examined local and global modulation of the phase-locking factor (PLF). sTMS triggered transient phase locking in a wide frequency band with distinct PLF peaks at 21 Hz in the motor cortex and 8 Hz in the visual cortex. With TMS pulse trains of 11 Hz over visual cortex and 23 Hz over motor cortex, phase locking was progressively enhanced at the stimulation frequency and lasted for a few cycles after the stimulation terminated. Moreover, such local entrainment propagated to other cortical regions, suggesting that rTMS entrained intrinsic neural oscillations locally and globally via network nodes. Because the entrainment was frequency-specific for each target site, these frequencies may correspond to the natural frequency of each cortical module and of the global networks. rTMS enables direct manipulation of the brain and is thus useful for investigating the causal roles of synchronous neural oscillations and synchrony in brain functions, and for the treatment of clinical symptoms associated with impaired oscillations and synchrony.Significance StatementWe provide the first evidence for area- and frequency-specific entrainment by frequency-tuned repetitive transcranial magnetic stimulation (rTMS), and the propagation of this entrainment to other areas. Our results indicate that rTMS at the natural frequency of each cortical system is particularly effective for entraining oscillatory phase. Moreover, local entrainment led to global entrainment in functionally coupled areas. The ability to control brain rhythms in the intact human brain is highly beneficial for studying the causal roles of rhythmic activity in brain function. Moreover, this modulatory technique has the potential to treat patients with impaired rhythmic networks in disorders such as schizophrenia and stroke.

Objectives: To investigate the systemic sclerosis–related phenotype in fos-related antigen-1 transgenic mice and its underlying mechanisms. Methods: Lung and skin sections of constitutive fos-related antigen-1 transgenic mice and wild-type mice were examined by tissue staining and immunohistochemistry. The tricuspid regurgitation pressure gradient was measured by transthoracic echocardiography with a Doppler technique. To assess the impact of fos-related antigen-1 expression on macrophage function, bone marrow–derived mononuclear cells were derived from mice that expressed fos-related antigen-1 under the control of doxycycline and wild-type littermates. These bone marrow–derived mononuclear cells were induced to differentiate into macrophages with or without doxycycline, and analyzed for gene and protein expression. Finally, lung explants obtained from systemic sclerosis patients and control donors were subjected to immunohistochemistry. Results: The lungs of fos-related antigen-1 transgenic mice showed excessive fibrosis of the interstitium and thickening of vessel walls, with narrowing lumen, in an age-dependent manner. The tricuspid regurgitation pressure gradient was significantly elevated in fos-related antigen-1 transgenic versus control mice. Increased dermal thickness and the loss of subdermal adipose tissue were also observed in the fos-related antigen-1 transgenic mice. These changes were preceded by a perivascular infiltration of mononuclear cells, predominantly consisting of alternatively activated or M2 macrophages. Overexpressing fos-related antigen-1 in bone marrow–derived mononuclear cell cultures increased the expression of M2-related genes, such as Il10, Alox15, and Arg1. Finally, fos-related antigen-1-expressing M2 macrophages were increased in the lung tissues of systemic sclerosis patients. Conclusions: The fos-related antigen-1 transgenic mouse serves as a genetic model of systemic sclerosis that recapitulates the major vascular and fibrotic manifestations of the lungs and skin in systemic sclerosis patients. M2 polarization mediated by the up-regulation of fos-related antigen-1 may play a critical role in the development of systemic sclerosis.

Background Concomitant malignancy is one of the prognostic factors in patients with myositis,1 but clinical parameters for mortality still remain unknown in patients with cancer-associated myositis (CAM). Objectives Initial predictors for mortality were examined using a multicenter cohort of CAM patients. Methods This retrospective study enrolled 67 consecutive patients diagnosed as having CAM in 3 referral hospitals between 1995 and 2017. Clinical data at diagnosis of myositis as well as treatment regimens and outcomes of myositis and malignancy were collected by review of medical charts. Myositis-specific autoantibodies (MSAs) were comprehensively detected using RNA immunoprecipitation (IP), enzyme-linked immunosorbent assay, and IP-immunoblotting. We initially conducted a univariate analysis to select variables that were different between survivors and dead cases. In multivariate analysis, the Cox proportional hazard model with backward selection method (p>0.20) was employed to identify factors independently associated with mortality. Explanatory variables were chosen based on the following three models. The Model 1 included age at diagnosis of myositis, gender and candidate variables (p Results The median age at diagnosis of myositis was 63 years, and 62% were female. MSAs were detected in 47 patients: anti-TIF1-γ in 27, anti-ARS in 6, anti-MDA5 in 5, anti-Mi-2 in 3, anti-NXP2 in 3, anti-SAE in 2, and anti-SRP in 1. During the median observation period of 2 years, 19 (28%) of 67 CAM patients were dead due to tumour in 16, ILD in 1, and an unknown cause in 2. The univariate analysis identified significant poor prognostic factors (p Conclusions Male and progression of malignancy at diagnosis of myositis were identified as predictors of survivals in CAM patients. Reference [1] Callen JP. Dermatomyositis. Lancet2000;355:53–7. Disclosure of Interest H. Kadota: None declared, T. Gono: None declared, Y. Yamaguchi: None declared, E. Watanabe: None declared, Y. Okazaki: None declared, Y. Nakajima: None declared, S. Kobayashi: None declared, Y. Shirai: None declared, M. Takeno: None declared, C. Terai: None declared, M. Kuwana Grant/research support from: Astellas, Speakers bureau: Astellas and Japan Blood Products Organisation

Non-invasive human electroencephalography (EEG) coupled with transcranial magnetic stimulation (TMS) is currently used to measure coarse stimulus-response relationships in brain physiology during behavior. However, with key modifications, the TMS-EEG technique holds even greater promise for monitoring fine-scale neural signatures of human behavior. Here, we demonstrate that a novel TMS-EEG co-registration technique can dynamically monitor individual human variation in perception based solely on EEG resting-state intrinsic effective connectivity probed by TMS-based phase resetting of ongoing activity. We used a bistable stimulus task, where the percept is perceived as either horizontal or vertical apparent motion, to record gamma band interhemispheric integration of information. Fine-grained inter-individual behavioral differences in horizontal motion bias could be measured by tracking resting-state gamma-band effective connectivity from right hMT+ to left hMT+. Thus, our method of triggering intrinsic resting-state effective connectivity in oscillatory dynamics can monitor individual differences in perception via the long-range integration of information. This technique will be useful for the manipulative dissection of individual-scale human cognition mediated by neural dynamics and may also expand neurofeedback approaches.

Of 207 adult patients with idiopathic inflammatory myopathies, detection of autoantibodies by RNA immunoprecipitation showed that 99 patients (48%) were antibody-positive. We divided these 99 into five subgroups: anti-signal recognition particle (SRP), anti-aminoacyl transfer RNA synthetase, anti-Ku, anti-U1RNP, and anti-SSA/B. Younger age at onset, severe weakness, muscle atrophy, elevated creatine kinase, and necrosis in muscle fibers without inflammatory cell infiltration were found significantly more frequently among the patients with anti-SRP antibodies (n=41) compared to the antibody-negative patients (n=108). Autoantibody detection by RNA immunoprecipitation can provide useful information associated with clinical and histological findings.

Objective Defective vasculogenesis is thought to play a role in the pathogenesis of systemic sclerosis (SSc). We undertook this study to explore the in vivo functional capacity of CD34+CD133+CD309+ endothelial progenitor cells (EPCs) in SSc patients. Methods CD133+ cells and EPCs were enumerated by flow cytometry. Immunomagnetically sorted circulating CD133+ cells from 16 patients with SSc and 12 healthy subjects, as well as murine colon carcinoma CT-26 cells, were transplanted beneath the skin of SCID mice. Tumor volume and blood vessel density were measured 10 days later. Human EPC incorporation into the vascular wall was evaluated using tumor sections double stained for mouse CD31 and human CD31. Results The number of CD133+ cells and EPCs was significantly decreased in SSc patients as compared to healthy controls (P = 0.001 and P = 0.02, respectively), while the proportion of EPCs in CD133+ cells was similar between the 2 groups. CT-26 cells produced markedly stronger tumor growth and neovessel formation when transplanted with CD133+ cells from healthy subjects than when transplanted with CD133+ cells from SSc patients (P = 0.001 and P = 0.008, respectively). Tumors from mice that received transplants of CT-26 cells and SSc-derived CD133+ cells formed fewer vessels incorporating human EPC–derived mature endothelial cells than did tumors from mice that received transplants of CT-26 cells and CD133+ cells from healthy control subjects (P = 0.0002). Conclusion We established a system that can be used to evaluate the in vivo neovascularization capacity of freshly isolated EPCs. EPCs contribute to vascularization by incorporating into vessel walls and by differentiating into endothelial cells. These EPC functions are impaired in SSc.

Background. To confi rm the antigen specifi cities of autoantibodies that precipitate 140-kDa (anti-p140) or 155/140-kDa polypeptides (anti-p155/140) previously identifi ed by radioimmunoprecipitation in Korean patients with dermatomyositis (DM) and to look into the relationship between each MSA and clinical features of DM. Methods. Seventeen serum samples of classic DM patients who had been found to have either anti-p140 antibodies ( n 9) or anti-p155/140 ( n 8) antibodies in our previous study were examined using enzyme-linked immunosorbent assay (for anti-MDA5 antibodies) and immunoblotting (for anti-MJ/NXP-2 and anti-TIF-1 γ antibodies). Results. Seven out of nine anti-p140 antibody positive patients were found to have anti-MDA5 antibodies. Two out of nine had anti-MJ/NXP-2 antibodies with no interstitial lung disease (ILD). All eight anti-p155/140 antibody positive patients were found to have anti-TIF-1 γ antibodies. Anti-TIF-1 γ and anti-MDA5 antibodies were simultaneously detected in one patient with antip155/140 antibody, who suff ered HIV infection and non-Hodgkin ’ s lymphoma. The associations between anti-MDA5 antibody and rapidly progressive ILD and between anti-TIF-1 γ antibody and cancer-associated DM were confi rmed to be signifi cant. Conclusions. Although radioimmunoprecipitation still looks to be a good screening tool, confi rmation with antigen-specifi c assays seems mandatory. The associations between anti-MDA5 and rapidly progressive ILD and between anti-TIF-1 γ and cancer-associated DM were confi rmed in Korean patients with DM.