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Background: The International Committee of Medical Journal Editors (ICMJE) has published Recommendations for the Conduct, Reporting, Editing, and Publication of Scholarly Work in Medical Journals. These provide a global standard for writing and editing medical articles, including research integrity. However, no study has examined the research integrity-related content of Japanese medical journals' Instructions for Authors. We therefore compared research integrity content in ICMJE member journals with those in the English- and Japanese-language journals of the Japanese Association of Medical Sciences (JAMS)., Materials and Methods: This was a descriptive literature study. We obtained Instructions for Authors from English- and Japanese-language journals listed on the JAMS website and the ICMJE member journals listed on the ICMJE website as of September 1, 2021. We compared the presence of 20 topics (19 in the ICMJE Recommendations plus compliance with ICMJE) in the Instructions for Authors, and analyzed the content of the conflict of interest disclosure., Results: We evaluated 12 ICMJE member journals, and 82 English-language and 99 Japanese-language subcommittee journals. The median number of topics covered was 10.5 for ICMJE member journals, 10 for English-language journals, and three for Japanese-language journals. Compliance with ICMJE was mentioned by 10 (83%) ICMJE member journals, 75 (91%) English-language journals, and 29 (29%) Japanese-language journals. The ICMJE Conflicts of Interest Disclosure Form was requested by seven (64%) ICMJE member journals, 15 (18%) English-language journals, and one (1%) Japanese-language journal., Conclusions: Although the topics in the JAMS English-language journals resembled those in the ICMJE member journals, the median value of ICMJE-related topic inclusion was approximately one-third lower in JAMS Japanese-language journals than in ICMJE member journals. It is hoped that Japanese-language journals whose conflict of interest disclosure policies differ from ICMJE standards will adopt international standards to deter misconduct and ensure publication quality., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: KI received research grants from The Nippon Foundation – Osaka University Project for Infectious Disease Prevention, the Watanabe Memorial Foundation for the Advancement of New Technology, (R2-524), the Inamori Foundation, and the Japan Society for the Promotion of Science (Grant-in-Aid for Early-Career Scientists, 23K12845); and received honoraria from Mimir, Inc., outside the submitted work. YN reports donation from Datack, outside the submitted work. HO reports research grants from I&H Co., Ltd., KRAFT Inc., donation from Neo Plus Pharma Co., Ltd., and YUYAMA Co., Ltd. TN reports research grants from I&H Co., Ltd., Cocokarafine Co., Ltd., and Konica Minolta Inc.; consulting fees from Otsuka Pharmaceutical; honoraria from Pfizer Japan, Merck Sharp & Dohme, Chugai Pharmaceutical, Takeda Pharmaceutical, Janssen Pharmaceutical K.K., Boehringer Ingelheim, Eli Lilly Japan K.K., Maruho, Mitsubishi Tanabe Pharma, Novartis Pharma, Allergan Japan, Novo Nordisk Pharma, Toa Eiyo, Dentsu, and GlaxoSmithKline, Abbott; Stock options from Bon Bon Inc.; and donations from CancerScan and YUYAMA Co., Ltd. All other authors declare no support from any organization for the submitted work; no financial relationships with any organizations that might have an interest in the submitted work in the previous five years; no other relationships or activities that could appear to have influenced the submitted work. This does not alter our adherence to PLOS ONE policies on sharing data and materials., (Copyright: © 2024 Koizumi 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.)

Aortic and valvular calcification are well-known risk factors for cardio-cerebrovascular events in patients undergoing hemodialysis. We investigated the clinical impact of an angulated aorto-septal angle as a result of aortic elongation due to aortic calcification on cardio-cerebrovascular outcomes in patients undergoing hemodialysis. We investigated 306 patients (mean age 65.4 years, 68% male) who underwent pre-scheduled routine echocardiography between April and September 2018. The angle between the anterior wall of the aorta and the ventricular septal surface (ASA) was quantified. We determined aortic and mitral valve calcification scores based on calcified cardiac changes; the aortic and mitral valve scores ranged between 0-9 and 0-6, respectively. The primary endpoint was a composite including cardio-cerebrovascular events and cardio-cerebrovascular death. The mean duration of dialysis among the patients in this analysis was 9.6 years. The primary endpoint was observed in 54 patients during the observational period (median 1095 days). Multivariable Cox proportional hazards analyses identified left ventricular ejection fraction (per 10% increase: hazard ratio [HR] 0.67; 95% confidential interval [CI] 0.53-0.84, P = 0.001), left ventricular mass index (per 10 g/m2 increase: HR 1.14; 95% CI 1.05-1.24, P = 0.001), ASA (per 10 degree increase: HR 0.69; 95% CI 0.54-0.88; P = 0.003), and aortic valve calcification score (HR 1.15; 95% CI 1.04-1.26, P = 0.005) as independent determinants of the primary endpoint. Kaplan-Meier analysis showed a higher incidence of the primary endpoint in patients with ASA <119.4 degrees than those with ASA ≥119.4 degrees (Log-rank P < 0.001). An angulated aorto-septal angle is an independent risk factor for cardio-cerebrovascular events and cardio-cerebrovascular death in patients undergoing hemodialysis., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Nakayama 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.)

Aim: Patients with severe heart failure undergo highly invasive and advanced therapies with uncertain treatment outcomes. For these patients, shared decision-making is necessary. To date, the nursing perspective of the decision-making process for patients facing difficulties and how nurses can support patients in this process have not been fully elucidated. This study aimed to clarify the perceptions of critical care nurses regarding situations with patients with severe heart failure that require difficult decision-making, and their role in supporting these patients., Methods: Individual semi-structured interviews were conducted with 10 certified nurse specialists in critical care nursing at nine hospitals in Japan. A qualitative inductive method was used and the derived relationships among the themes were visually structured and represented., Results: The nurses' perceptions on patients' difficult situations in decision-making were identified as follows: painful decisions under uncertainties; tense relationships; wavering emotions during decision-making; difficulties in coping with worsening medical conditions; patients' wishes that are difficult to realize or estimate; and difficulties in transitioning from advanced medical care. Critical care nurses' roles were summarized into six themes and performed collaboratively within the nursing team. Of these, the search for meaning and value was fundamental. Two positions underpin the role of critical care nurses. The first aims to provide direct support and includes partnerships and rights advocacy. The second aims to provide a holistic perspective to enable necessary adjustments, as indicated by situation assessments and mediation. By crossing various boundaries, co-creating, and forming a good circular relationship in the search for meaning and values, the possibility of expanding treatment and recuperation options may be considered., Conclusions: Patients with severe heart failure have difficulty participating in shared decision-making. Critical care nurses should collaborate within the nursing team to improve interprofessional shared decision-making by providing decisional support to patients that focuses on values and meaning., Competing Interests: We have read the journal’s policy and the authors of this manuscript have the following competing interests: TN received research grants from I&H Co., Ltd., Cocokarafine Co., Ltd., and Konica Minolta Inc.; consulting fees from Otsuka Pharmaceutical; honoraria from Pfizer Japan INC., MSD K.K., Chugai Pharmaceutical Co., Takeda Pharmaceutical Co., Janssen Pharmaceutical K.K., Boehringer Ingelheim International GmbH, Eli Lilly Japan K.K., Maruho Co., Ltd., Mitsubishi Tanabe Pharma Co, Novartis Pharma K.K., Allergan Japan K.K., Maruho Co., Ltd., Novo Nordisk Pharma Ltd., TOA EIYO Ltd., Dentsu co., ONO PHARMACEUTICAL CO., LTD., GSK plc, Alexion Pharmaceuticals, Inc., and Cannon Medical Systems Co.; stock options from Bon Bon Inc.; donations from CancerScan and YUYAMA co. The other authors declare that they have no conflicts of interest to disclose. This does not alter our adherence to PLOS ONE policies on sharing data and materials, (Copyright: © 2023 Inagaki 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.)

Calorie restriction (CR) suppresses not only spontaneous but also chemical- and radiation-induced carcinogenesis. Our previous study revealed that the cancer-preventive effect of CR is tissue dependent and that CR does not effectively prevent the development of thymic lymphoma (TL). We investigated the association between CR and the genomic alterations of resulting TLs to clarify the underlying resistance mechanism. TLs were obtained from previous and new experiments, in which B6C3F1 mice were exposed to radiation at 1 week of age and fed with a CR or standard (non-CR) diet from 7 weeks throughout their lifetimes. All available TLs were used for analysis of genomic DNA. In contrast to the TLs of the non-CR group, those of the CR group displayed suppression of copy-neutral loss of heterozygosity (LOH) involving relevant tumor suppressor genes (Cdkn2a, Ikzf1, Trp53, Pten), an event regarded as cell division-associated. However, CR did not affect interstitial deletions of those genes, which were observed in both groups. In addition, CR affected the mechanism of Ikzf1 inactivation in TLs: the non-CR group exhibited copy-neutral LOH with duplicated inactive alleles, whereas the CR group showed expression of dominant-negative isoforms accompanying a point mutation or an intragenic deletion. These results suggest that, even though CR reduces cell division-related genomic rearrangements by suppressing cell proliferation, tumors arise via diverse carcinogenic pathways including inactivation of tumor suppressors via interstitial deletions and other mutations. These findings provide a molecular basis for improved prevention strategies that overcome the CR resistance of lymphomagenesis., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Nakayama 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.)

Melanoma shows highly aggressive behavior (i.e., local invasion and metastasis). Matrix metalloprotease-3 (MMP-3), a zinc-dependent endopeptidase, degrades several extracellular substrates and contributes to local invasion by creating a microenvironment suitable for tumor development. Here, we report that interleukin-1β (IL-1β) triggers the MMP-3 expression in canine melanoma cells. The activity of MMP-3 in the culture supernatant was increased in IL-1β-treated melanoma cells. IL-1β time- and dose-dependently provoked the mRNA expression of MMP-3. IL-1β induced the migration of melanoma cells; however, this migration was attenuated by UK356618, an MMP-3 inhibitor. When the cells were treated with the nuclear factor-κB (NF-κB) inhibitor TPCA-1, the inhibition of MMP-3 expression was observed. In IL-1β-treated cells, the phosphorylation both of p65/RelA and p105 was detected, indicating NF-κB pathway activation. In p65/RelA-depleted melanoma cells, IL-1β-mediated mRNA expression of MMP-3 was inhibited, whereas this reduction was not observed in p105-depleted cells. These findings suggest that MMP-3 expression in melanoma cells is regulated through IL-1β-mediated p65/RelA activation, which is involved in melanoma cell migration., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2022 Nunomura 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.)

Thrombolytic therapy in the treatment of cardiogenic acute cerebral embolism caused by coagulated blood carries the risk of hemorrhagic complications, and there is a need to develop safer and more reliable treatment methods. Laser thrombolysis therapy, which utilizes the difference in energy absorption between the thrombus and the arterial wall, has shown promise as a new treatment method because it can selectively act only on the thrombus. It has not been applied clinically, however, and one of the main reasons for this is that its underlying mechanism has not been elucidated. We developed a pulse laser thrombolysis system for treating cerebral blood vessels that consists of a diode-pumped solid-state neodymium-yttrium aluminum garnet laser, which has excellent stability and maintainability and is suitable for clinical applications coupled to a small-diameter optical fiber. Moreover, we analyzed the mechanisms that occur during pulsed laser irradiation of transparent glass tubes and gelatin phantoms. We found that bubbles form as a thermal effect in addition to ablation of the pulsed laser irradiation. Furthermore, we detected no shock waves or water jets associated with the bubbles. We analyzed the bubbles' dynamics and growth rate, and their effect on a rabbit blood clot phantom. We concluded that the bubbles generated by the laser irradiation physically cut the thrombus and thereby had a thrombectomy effect. We believe that this study will clarify the mechanism of laser thrombolysis therapy and contribute greatly to the realization of its clinical application., Competing Interests: A patent application on the laser thrombolysis therapy has been filed by Hamamatsu Photonics K. K., Hamamatsu University School of Medicine and Teiji Nakayama. WIPO patent application names and number; HO, TN, KU, DY, and TT; Catheter, examination system and thrombus removing device, WIPO Serial Number 2007/049632 A1. There are no further patents, products in development or marketed products to declare. This does not alter the authors’ adherence to PLOS One’s policies on sharing data and materials.

Chronic kidney disease (CKD) and atrial fibrillation are both major burdens on the health care system worldwide. Several observational studies have reported clinical associations between CKD and atrial fibrillation; however, causal relationships between these conditions remain to be elucidated due to possible bias by confounders and reverse causations. Here, we conducted bidirectional two-sample Mendelian randomization analyses using publicly available summary statistics of genome-wide association studies (the CKDGen consortium and the UK Biobank) to investigate causal associations between CKD and atrial fibrillation/flutter in the European population. Our study suggested a causal effect of the risk of atrial fibrillation/flutter on the decrease in serum creatinine-based estimated glomerular filtration rate (eGFR) and revealed a causal effect of the risk of atrial fibrillation/flutter on the risk of CKD (odds ratio, 9.39 per doubling odds ratio of atrial fibrillation/flutter; 95% coefficient interval, 2.39-37.0; P = 0.001), while the causal effect of the decrease in eGFR on the risk of atrial fibrillation/flutter was unlikely. However, careful interpretation and further studies are warranted, as the underlying mechanisms remain unknown. Further, our sample size was relatively small and selection bias was possible., Competing Interests: The authors have declared that no competing interests exist.

While IFNγ is a well-known cytokine that actively promotes the type I immune response, it is also known to suppress the type II response by inhibiting the differentiation and proliferation of Th2 cells. However, the mechanism by which IFNγ suppresses Th2 cell proliferation is still not fully understood. We found that IFNγ decreases the expression of growth factor independent-1 transcriptional repressor (GFI1) in Th2 cells, resulting in the inhibition of Th2 cell proliferation. The deletion of the Gfi1 gene in Th2 cells results in the failure of their proliferation, accompanied by an impaired cell cycle progression. In contrast, the enforced expression of GFI1 restores the defective Th2 cell proliferation, even in the presence of IFNγ. These results demonstrate that GFI1 is a key molecule in the IFNγ-mediated inhibition of Th2 cell proliferation., Competing Interests: The authors declare no conflicts of interest in association with the present study.

In autoimmune diseases, fibroblasts produce and secrete various cytokines and act as sentinel immune cells during inflammatory states. However, the contribution of sentinel immune cells (i.e. dermal fibroblasts) in autoimmune diseases of the skin, such as atopic dermatitis, has been obscure. The pro-inflammatory cytokine interleukin 1β (IL-1β) induces the expression of chemokines, such as interleukin 8 (IL-8), in autoimmune diseases of the skin. IL-8 induces the activation and recruitment of innate immune cells such as neutrophils to the site of inflammation. IL-1β-mediated induction of IL-8 expression is important for the pathogenesis of autoimmune diseases; however, the intracellular singling remains to be understood. To elucidate the mechanism of the onset of autoimmune diseases, we established a model for IL-1β-induced dermatitis and investigated MAPK signaling pathways in IL-1β-induced IL-8 expression. We also identified that a MAP3K Tpl2 acts as an upstream modulator of IL-1β-induced ERK1/2 activation in dermal fibroblasts. We observed an increase in the expression of IL-8 mRNA and protein in cells treated with IL-1β. ERK1/2 inhibitors significantly reduced IL-1β-induced IL-8 expression, whereas the inhibitor for p38 MAPK or JNK had no effect. IL-1β induced ERK1/2 phosphorylation, which was attenuated in the presence of an ERK1/2 inhibitor. IL-1β failed to induce IL-8 expression in cells transfected with siRNA for ERK1, or ERK2. Notably, a Tpl2 inhibitor reduced IL-1β-induced IL-8 expression and ERK1/2 phosphorylation. We confirmed that the silencing of Tpl2 in siRNA-transfected fibroblasts prevented both in IL-1β-induced IL-8 expression and ERK1/2 phosphorylation. Taken together, our data indicate the importance of Tpl2 in the modulation of ERK1/2 signaling involved in the IL-1β-induced development of autoimmune diseases affecting the dermal tissue, such as atopic dermatitis., Competing Interests: The authors have declared that no competing interests exist.

Since 2012, studies in mice, rats, and humans have suggested that abnormalities in purinergic signaling may be a final common pathway for many genetic and environmental causes of autism spectrum disorder (ASD). The current study in mice was conducted to characterize the bioenergetic, metabolomic, breathomic, and behavioral features of acute hyperpurinergia triggered by systemic injection of the purinergic agonist and danger signal, extracellular ATP (eATP). Responses were studied in C57BL/6J mice in the maternal immune activation (MIA) model and controls. Basal metabolic rates and locomotor activity were measured in CLAMS cages. Plasma metabolomics measured 401 metabolites. Breathomics measured 98 volatile organic compounds. Intraperitoneal eATP dropped basal metabolic rate measured by whole body oxygen consumption by 74% ± 6% (mean ± SEM) and rectal temperature by 6.2˚ ± 0.3˚C in 30 minutes. Over 200 metabolites from 37 different biochemical pathways where changed. Breathomics showed an increase in exhaled carbon monoxide, dimethylsulfide, and isoprene. Metabolomics revealed an acute increase in lactate, citrate, purines, urea, dopamine, eicosanoids, microbiome metabolites, oxidized glutathione, thiamine, niacinamide, and pyridoxic acid, and decreased folate-methylation-1-carbon intermediates, amino acids, short and medium chain acyl-carnitines, phospholipids, ceramides, sphingomyelins, cholesterol, bile acids, and vitamin D similar to some children with ASD. MIA animals were hypersensitive to postnatal exposure to eATP or poly(IC), which produced a rebound increase in body temperature that lasted several weeks before returning to baseline. Acute hyperpurinergia produced metabolic and behavioral changes in mice. The behaviors and metabolic changes produced by ATP injection were associated with mitochondrial functional changes that were profound but reversible., Competing Interests: The authors have declared that no competing interests exist.

Chronic ethanol consumption is a risk factor for colorectal cancer, and ethanol-induced reactive oxygen species have been suggested to play important roles in the pathogenesis of ethanol-related colorectal cancer (ER-CRC). In this study, the effects of 10-week chronic administration of ethanol on the colonic levels of oxidative stress and advance glycation end product (AGE) levels, as well as fecal microbiota structures, were examined in a mouse model. Chronic oral administration of ethanol in mice (1.0 mL of 1.5% or 5.0% ethanol (v/v) per day per mouse, up to 10 weeks) resulted in the elevation of colonic levels of oxidative stress markers (such as 8-hydroxy-2'-deoxyguanosine and 4-hydroxynonenal) compared to control mice, and this was consistently accompanied by elevated levels of inflammation-associated cytokines and immune cells (Th17 and macrophages) and a decreased level of regulatory T (Treg) cells to produce colonic lesions. It also resulted in an alteration of mouse fecal microbiota structures, reminiscent of the alterations observed in human inflammatory bowel disease, and this appeared to be consistent with the proposed sustained generation of oxidative stress in the colonic environment during chronic ethanol consumption. Moreover, the first experimental evidence that chronic ethanol administration results in elevated levels of advanced glycation end products (AGEs) and their receptors (RAGE) in the colonic tissues in mice is also shown, implying enhanced RAGE-mediated signaling with chronic ethanol administration. The RAGE-mediated signaling pathway has thus far been implicated as a link between the accumulation of AGEs and the development of many types of chronic colitis and cancers. Thus, enhancement of this pathway likely exacerbates the ethanol-induced inflammatory states of colonic tissues and might at least partly contribute to the pathogenesis of ER-CRC., Competing Interests: The authors have declared that no competing interests exist.

The rate of glucose uptake dramatically increases in cancer cells even in the presence of oxygen and fully functioning mitochondria. Cancer cells produce ATP by glycolysis rather than oxidative phosphorylation under aerobic conditions, a process termed as the "Warburg effect." In the present study, we treated canine melanoma cells with the glucose analog 2-deoxy-D-glucose (2-DG) and investigated its effect on cell growth. 2-DG attenuated cell growth in a time- and dose-dependent manner. Cell growth was also inhibited following treatment with the glucose transporter (GLUT) inhibitor WZB-117. The treatment of 2-DG and WZB-117 attenuated the glucose consumption, lactate secretion and glucose uptake of the cells. The mRNA expression of the subtypes of GLUT was examined and GLUT1 and GLUT3 were found to be expressed in melanoma cells. The growth, glucose consumption and lactate secretion of melanoma cells transfected with siRNAs of specific for GLUT1 and GLUT3 was suppressed. These findings suggest that glucose uptake via GLUT1 and GLUT3 plays a crucial role for the growth of canine melanoma cells., Competing Interests: The authors have declared that no competing interests exist.

Backgrounds: An estimated 2.5 million Afghans are living in the Iran and almost half of them are young women at the childbearing ages. Although the evidence indicates lower rates of antenatal care and higher incidence of pregnancy complications in Afghan compared to Iranian women, the underlying reasons are not well defined. Therefore, in the present study, we aimed to explore the prevalence and associated sociodemographic factors of adverse pregnancy outcomes and examine the impact of intimate partner violence, food insecurity, poor mental health, and housing issues on pregnancy outcome in Afghan women living in Iran., Methods: In July 2019, we enrolled 424 Afghan women aged 18-44 years old using the time-location sampling at three community health centers in the south region of Tehran province. The data was collected through face to face interviews using the researcher-developed questionnaire. Using bivariate and multivariate analysis, the impact of poor antenatal care, intimate partner violence, food insecurity, and poor mental health was assessed on the incidence of adverse pregnancy outcome., Results: More than half (56.6%) of Afghan women reported at least one pregnancy complication in their recent pregnancy. The results showed that undocumented, illiterate, and unemployed Afghan women with lower socioeconomic status are more likely to experience adverse pregnancy outcomes. Furthermore, we observed lower prevalence of adverse pregnancy outcomes among documented immigrants with health insurance compared to those with no health insurance. It is also been found that the food insecurity [Adjusted OR = 3.35, 95% CI (1.34-8.36)], poor antenatal care [Adjusted OR = 10.50, 95% CI (5.40-20.39)], intimate partner violence [Adjusted OR = 2.72, 95% CI (1.10-6.77)], and poor mental health [Adjusted OR = 4.77, 95% CI (2.54-8.94)] could adversely impact the pregnancy outcome and we observed higher incidence of adverse outcomes among those suffering from these situations., Conclusion: To our knowledge, this is the first study that explored the prevalence and associated factors of adverse pregnancy outcomes and the impact of intimate partner violence, food insecurity, poor mental health on pregnancy outcome among Afghan women in Iran. Enhancing the psychosocial support and empowering Afghan women through expanding the social network and safety net should be a priority for the central government and international parties. Psychological counseling should be incorporated into routine maternity care for Afghan refugees. Access to free antenatal care is a right for all Afghan women and it should be facilitated by universal health insurance for all Afghans regardless of their legal status., Competing Interests: The authors have declared that no competing interests exist.

Macrophages play an indispensable role in both innate and acquired immunity, while the persistence of activated macrophages can sometimes be harmful to the host, resulting in multi-organ damage. Macrophages develop from monocytes in the circulation. However, little is known about the organ affinity of macrophages in the normal state. Using in vivo imaging with XenoLight DiR®, we observed that macrophages showed strong affinity for the liver, spleen and lung, and weak affinity for the gut and bone marrow, but little or no affinity for the kidney and skin. We also found that administered macrophages were still alive 168 hours after injection. On the other hand, treatment with clodronate liposomes, which are readily taken up by macrophages via phagocytosis, strongly reduced the number of macrophages in the liver, spleen and lung., Competing Interests: The authors have declared that no competing interests exist.

Some findings on the association between glaucoma and statins in the Asian population have been reported. We conducted a retrospective cohort study using health insurance claims data maintained by the JMDC Inc., which comprises data on about three million individuals representing 2.4% of the Japanese population. The association between the potency of statins and open-angle glaucoma in Japanese working-age population was examined using a commercially available health insurance claims and enrollment database. We identified 117,036 patients with a prescription of statins between January 1, 2005 and March 31, 2014; 59,535 patients were selected as new statin users. Of these, 49,671 (83%) patients without glaucoma who were prescribed statins for the first time were part of the primary analysis. New users of statin were defined as those with a prescription of statin at the beginning of the study, but without a prescription six months earlier. The cohort comprised 29,435 (59%) and 20,236 (41%) patients with a prescription of high-potency statin (atorvastatin and rosuvastatin) and low-potency statin (pravastatin, fluvastatin, pitavastatin, and simvastatin), respectively. Using Cox proportional hazards regression analysis, hazard ratios (HRs) were estimated for glaucoma adjusted for baseline characteristics. Although some baseline characteristics were not similar between the high-potency and low-potency statin groups, the standardized difference for all covariates was less than 0.1. No associations were found between high-potency statin use and glaucoma (adjusted HR = 1.08; 95% confidence interval, 0.93-1.24) in the primary analyses, using the risk for glaucoma in the low-potency statin group as reference. The risk of glaucoma with individual statin use was not significantly different from that with pravastatin. No significant association was found between high-potency statins and the increased risk of glaucoma in Japanese working-age population. Further studies are needed to examine the association between statins and glaucoma in the elderly population., Competing Interests: The authors have declared that no competing interests exist.

In this study, we synthesized findings from qualitative studies to identify the key factors that influence child marriage. We used a meta-ethnographic approach coupled with thematic synthesis. We searched literature from nine databases, which were in English language, covering areas in public health, psychology, and social science between 2008 and 2018. Twelve studies were included in the synthesis. We identified six main themes: human insecurity and conflict; legal issues; family values and circumstances; religious beliefs; individual circumstances, beliefs, and knowledge; and social norms. Our findings highlight the impact of human insecurity and conflict, as well as legal issues. In spite of global progress scaling up legislation against child marriage, the legal framework is insufficiently enforced in many settings. Most of the included studies were from the Middle East, Africa, and South Asia. Studies from other parts of the world such as Latin America and Southeast Asia, which have the highest rates of child marriage, are needed., Competing Interests: The authors have declared that no competing interests exist.

The alpha/beta-Hydrolases (ABH) are a structural class of proteins that are found widespread in nature and includes enzymes that can catalyze various reactions in different substrates. The catalytic versatility of the ABH fold enzymes, which has been a valuable property in protein engineering applications, is based on a similar acid-base-nucleophile catalytic mechanism. In our research, we are concerned with the structure that surrounds the key units of the catalytic machinery, and we have previously found conserved structural organizations that coordinate the catalytic acid, the catalytic nucleophile and the residues of the oxyanion hole. Here, we explore the architecture that surrounds the catalytic histidine at the active sites of enzymes from 40 ABH fold families, where we have identified six conserved interactions that coordinate the catalytic histidine next to the catalytic acid and the catalytic nucleophile. Specifically, the catalytic nucleophile is coordinated next to the catalytic histidine by two weak hydrogen bonds, while the catalytic acid is directly involved in the coordination of the catalytic histidine through by two weak hydrogen bonds. The imidazole ring of the catalytic histidine is coordinated by a CH-π contact and a hydrophobic interaction. Moreover, the catalytic triad residues are connected with a residue that is located at the core of the active site of ABH fold, which is suggested to be the fourth member of a "structural catalytic tetrad". Besides their role in the stability of the catalytic mechanism, the conserved elements of the catalytic site are actively involved in ligand binding and affect other properties of the catalytic activity, such as substrate specificity, enantioselectivity, pH optimum and thermostability of ABH fold enzymes. These properties are regularly targeted in protein engineering applications, and thus, the identified conserved structural elements can serve as potential modification sites in order to develop ABH fold enzymes with altered activities., Competing Interests: The authors have declared that no competing interests exist.

Matrix metalloproteinases (MMPs) play a pivotal role in tissue remodeling by degrading the extracellular matrix (ECM) components. This mechanism is implicated in a variety of physiological and pathological cellular processes including wound healing. One of the key proteins involved in this process is the proinflammatory cytokine interleukin-1β (IL-1β, which induces the expression of MMP-3 mRNA and the secretion of MMP-3 protein by dermal fibroblasts. In this study, we first investigated the contribution of activating transcription factor 2 (ATF-2) to IL-1β-induced MMP-3 expression in dermal fibroblasts. Our results showed that in cells transfected with ATF-2 siRNA or treated with the ATF-2 inhibitor SBI-0087702, IL-1β-induced MMP-3 mRNA expression was reduced. We also demonstrated that IL-1β stimulates the phosphorylation of ATF-2. These observations suggest that ATF-2 plays an important role in IL-1β-induced MMP-3 expression. Next, we investigated the role of MAPK signaling in ATF-2 activation. In cells treated with the extracellular signal-regulated kinase (ERK) inhibitor FR180240, as well as in cells transfected with ERK1 and ERK2 siRNAs, IL-1β-induced MMP-3 mRNA expression was reduced. In addition, we showed that IL-1β induced the phosphorylation of ERK1/2. These observations suggest that ERK1 and ERK2 are involved in IL-1β-induced MMP-3 expression. However, ERK1 and ERK2 do seem to play different roles. While the ERK inhibitor FR180204 inhibited IL-1β-induced ATF-2 phosphorylation, only in cells transfected with ERK1 siRNA, but not ERK2 siRNA, IL-1β-induced ATF-2 phosphorylation was reduced. These findings suggest that the ERK1/ATF-2 signaling axis contributes to IL-1β-induced MMP-3 expression in dermal fibroblasts., Competing Interests: The authors have declared that no competing interests exist.

Interleukin-6 (IL-6) is a pleiotropic cytokine involved in the regulation of the immune response and inflammation. In this study, we investigated effect of the proinflammatory cytokine interleukin-1β (IL-1β) on IL-6 expression in canine dermal fibroblasts. IL-1β induced IL-6 mRNA expression and protein release in a time- and dose-dependent manner. When cells were treated with inhibitors of mitogen-activated protein kinases (MAPKs), the extracellular signal-regulated kinase (ERK) inhibitor FR180240 inhibited IL-1β-induced IL-6 mRNA expression, but not SP600125 or SKF86002, which are c-Jun N-terminal kinase (JNK) and p38 MAPK inhibitors, respectively. In cells treated with U0126, an inhibitor of MAPK/ERK kinase (MEK), which activates ERK, IL-1β-induced IL-6 mRNA expression was also inhibited. IL-1β stimulated ERK1/2 phosphorylation. In cells transfected with ERK1 and ERK2 isoform siRNAs, IL-1β-induced IL-6 mRNA expression was reduced. These observations suggest that IL-1β induces IL-6 expression via ERK1/2 signaling pathway in canine dermal fibroblasts., Competing Interests: The authors have declared that no competing interests exist.

Following the March 2011 accident at Fukushima Daiichi Nuclear Power Plant, many residents of Fukushima have faced anxieties about the health impacts of radiation exposure. Considering that source of information may influence resident anxiety, this study aimed to elucidate the correlation between the two. In addition, a health literacy query was included to examine a possible relationship between anxiety and health literacy skills. A mail survey was conducted in August 2016 among 2000 residents of Fukushima Prefecture aged 20 to 79 years. Survey items included questions about current health anxieties caused by radiation, trusted sources of information about radiation, and media used to obtain information on radiation. The survey valid response rate was 43.4%. Results of multiple linear regression analysis revealed that anxiety was significantly higher for the groups indicating "trust in citizen groups" and "use of internet sites." Anxiety was significantly lower for the groups indicating "trust in government ministries," "trust in local government," and "use of local broadcast television." Also anxiety was significantly lower for groups with higher health literacy. It was found that the significant relationship to anxiety varies depending on the sources of trust and media used. There is a possibility that this was caused by the difference between the contents of each information and media reports. In preparation for any future nuclear accident, government may consider action to improve the media literacy of residents. In addition, improving health literacy of both the recipient and the sender of information can improve access to information and thereby safeguard the health and well-being of the public., Competing Interests: The authors have declared that no competing interests exist.

Background: Vitamin D has been reported to affect both innate, and acquired immunity with immune cells such as dendritic cells having the vitamin D receptors. The co-occurrence of the high prevalence of allergic diseases and vitamin D deficiency globally documented in recent decades, has prompted a hypothesis on whether there is a reasonable association between them., Objective: To investigate the association between serum vitamin D deficiency and allergic symptoms., Methods: Historical cohort. On a cohort study for the association between desert dust exposure and allergic symptoms in 3,327 pregnant women during spring and fall in 2011-2013 in Japan conducted as an adjunct study to the Japan Environment and Children's Study, we promptly acquired subjects' daily allergic symptom scores by sending a web-based questionnaire to each participant on some days. Of the 29,434 answers provided by 3,327 participating pregnant women, we extracted 13,356 answers from 1,475 pregnant women that were answered within a 3-month period after blood samplings. And we measured 25(OH)D levels on those samples to investigate the association between their vitamin D deficiency (serum 25(OH)D < 20ng/mL) and the occurrence of any allergic symptom (allergic symptom score> 0) within 3 months., Results: Serum 25(OH)D was less than 20ng/mL in 1,233 of 1,745 samples (70.7%). The adjusted odds ratio (aOR) for occurrence of any allergic symptom in deficient cases compared with non-deficient cases was 1.33 (95% CI: 1.07-1.64, p = 0.01). Further, vitamin D deficiency significantly enhanced the risk increase at desert dust events and at pollen exposure (p-values for interaction <0.1)., Conclusion: We confirmed the association between serum vitamin D deficiency and allergic symptoms in Japanese pregnant women., Competing Interests: The authors have declared that no competing interests exist.

Background: As a consequence of indoor occupations and reduced exposure to sunlight, concerns have been raised that vitamin D deficiency is widespread in developed countries. Vitamin D is known to be associated with increased risks of morbidity and mortality in various diseases., Objective: To investigate the serum vitamin D status and its relation with life-style factors in pregnant Japanese women., Methods: Among a cohort for 3,327 pregnant women who participated in an the adjunct study of the Japan Environment and Children's Study during 2011-2013, in which data were obtained on various life-style factors, including both dietary intake of vitamin D and frequency of UV exposure, this study consisted of 1,592 pregnant women, from whom 2,030 serum samples were drawn in Jan, Apr, Jul, and Oct, and the association between serum 25(OH)D level and life-style factors were analyzed using linear mixed models., Results: Serum 25(OH)D levels were less than 20ng/mL in 1,486 of 2,030 samples (73.2%). There was an obvious seasonal change, with serum 25(OH)D levels of less than 20 ng/mL in 89.8% and 47.8% of samples in spring (April) and autumn (October), respectively. Both the frequency spent under sunlight and dietary intake of vitamin D were significantly associated with serum 25(OH)D level. An increase in sunlight exposure of more than 15 min for 1 to 2 days per week in non-winter, or dietary intake of 2 μg/day of vitamin D resulted in an elevation of 1 ng/mL in serum 25(OH)D levels., Conclusion: These findings indicate that vitamin D deficiency is very severe in Japanese pregnant women, especially those rarely exposed to sunlight. The benefits of UV rays should also be informed of when its risk is alerted, and clinicians should propose the adequate UV exposure level., Competing Interests: The authors have declared that no competing interests exist.

Expulsion of parasitic gastrointestinal nematodes requires diverse effector mechanisms coordinated by a Th2-type response. The evolutionarily conserved JmjC protein; Myc Induced Nuclear Antigen (Mina) has been shown to repress IL4, a key Th2 cytokine, suggesting Mina may negatively regulate nematode expulsion. Here we report that expulsion of the parasitic nematode Trichuris muris was indeed accelerated in Mina deficient mice. Unexpectedly, this was associated not with an elevated Th2- but rather an impaired Th1-type response. Further reciprocal bone marrow chimera and conditional KO experiments demonstrated that retarded parasite expulsion and a normal Th1-type response both required Mina in intestinal epithelial cells (IECs). Transcriptional profiling experiments in IECs revealed anti-microbial α-defensin peptides to be the major target of Mina-dependent retention of worms in infected mice. In vitro exposure to recombinant α-defensin peptides caused cytotoxic damage to whipworms. These results identify a latent IEC-intrinsic anthelmintic pathway actively constrained by Mina and point to α-defensins as important effectors that together with Mina may be attractive therapeutic targets for the control of nematode infection., Competing Interests: The authors have declared that no competing interests exist.

Although mutations in several genes have been reported in pulmonary arterial hypertension (PAH), most of PAH cases do not carry these mutations. This study aimed to identify a novel cause of PAH. To determine the disease-causing variants, direct sequencing and multiplex ligation-dependent probe amplification were performed to analyze 18 families with multiple affected family members with PAH. In one of the 18 families with PAH, no disease-causing variants were found in any of BMPR2, ACVRL1, ENG, SMAD1/4/8, BMPR1B, NOTCH3, CAV1, or KCNK3. In this family, a female proband and her paternal aunt developed PAH in their childhood. Whole-exome next-generation sequencing was performed in the 2 PAH patients and the proband's healthy mother, and a BRCA1-associated protein (BRAP) gene variant, p.Arg554Leu, was identified in the 2 family members with PAH, but not in the proband's mother without PAH. Functional analyses were performed using human pulmonary arterial smooth muscle cells (hPASMCs). Knockdown of BRAP via small interfering RNA in hPASMCs induced p53 signaling pathway activation and decreased cell proliferation. Overexpression of either wild-type BRAP or p.Arg554Leu-BRAP cDNA constructs caused cell death confounding these studies, however we observed higher levels of p53 signaling inactivation and hPASMC proliferation in cells expressing p.Arg554Leu-BRAP compared to wild-type BRAP. In addition, p.Arg554Leu-BRAP induced decreased apoptosis of hPASMCs compared with wild-type BRAP. In conclusion, we have identified a novel variant of BRAP in a Japanese family with PAH and our results suggest it could have a gain-of-function. This study sheds light on new mechanism of PAH pathogenesis., Competing Interests: The authors have declared that no competing interests exist.

Various cell-based therapeutic strategies have been investigated for vascular and tissue regeneration after ischemic stroke. We have developed a novel cell population, called regeneration-associated cells (RACs), by quality- and quantity-controlled culture of unfractionated mononuclear cells. RACs were trans-arterially injected into 10-week-old syngeneic male mice at 1, 3, 5 or 7 days after permanent middle cerebral artery occlusion (MCAO) to determine the optimal timing for administration in terms of outcome at day 21. Next, we examined the effects of RACs injection at day 1 after MCAO on neurological deficits, infarct volume, and mediators of vascular regeneration and anti-inflammation at days 7 and 21. Infarct volume at day 21 was significantly reduced by transplantation of RACs at day 1 or 3. RACs injected at day 1 reduced the infarct volume at day 7 and 21. Angiogenesis and anti-inflammatory mediators, VEGF and IL-10, were increased at day 7, and VEGF was still upregulated at day 21. We also observed significantly enhanced ink perfusion in vivo, tube formation in vitro, and definitive endothelial progenitor cell colonies in colony assay. These results suggest that RAC transplantation in MCAO models promoted significant recovery of neural tissues through intensified anti-inflammatory and angiogenic effects., Competing Interests: The authors have declared that no competing interests exist.

Inflammatory and microenvironmental factors produced by cancer cells are thought to directly or indirectly promote cancer cell growth. Prostaglandins, including prostaglandin E2, have key roles as a microenvironment factor in influencing the development of tumors, and are produced by the rate limiting enzyme cyclooxygenase 2 (COX-2). In this study, we used canine melanoma cells treated with the proinflammatory cytokine interleukin 1β (IL-1β) and investigated the transcriptional factor nuclear factor-κB (NF-κB) signaling in IL-1β-induced COX-2 expression. IL-1β induced prostaglandin E2 release and COX-2 mRNA expression in a time- and dose-dependent manner. In the cells treated with the NF-κB inhibitors BAY11-7082 and TPC-1, IL-1β-mediated prostaglandin E2 release and COX-2 mRNA expression were inhibited. IL-1β also provoked phosphorylation of p65/RelA and p105/NF-κB1, which are members of the NF-κB families. The IL-1β-induced phosphorylation of p65 and p105 was attenuated in the presence of both NF-κB inhibitors. In melanoma cells transfected with siRNA of p65 or p105, IL-1β-mediated COX-2 mRNA expression was inhibited. These findings suggest that canonical activation of NF-κB signaling plays a crucial role for inflammatory states in melanoma cells., Competing Interests: The authors have declared that no competing interests exist.

The fornicata (fornicates) is a eukaryotic group known to consist of free-living and parasitic organisms. Genome datasets of two model fornicate parasites Giardia intestinalis and Spironucleus salmonicida are well annotated, so far. The nuclear genomes of G. intestinalis assemblages and S. salmonicida are small in terms of the genome size and simple in genome structure. However, an ancestral genomic structure and gene contents, from which genomes of the fornicate parasites have evolved, remains to be clarified. In order to understand genome evolution in fornicates, here, we present the draft genome sequence of a free-living fornicate, Kipferlia bialata, the divergence of which is earlier than those of the fornicate parasites, and compare it to the genomes of G. intestinalis and S. salmonicida. Our data show that the number of protein genes and introns in K. bialata genome are the most abundant in the genomes of three fornicates, reflecting an ancestral state of fornicate genome evolution. Evasion mechanisms of host immunity found in G. intestinalis and S. salmonicida are absent in the K. bialata genome, suggesting that the two parasites acquired the complex membrane surface proteins on the line leading to the common ancestor of G. intestinalis and S. salmonicida after the divergence from K. bialata. Furthermore, the mitochondrion related organelles (MROs) of K. bialata possess more complex suites of metabolic pathways than those in Giardia and in Spironucleus. In sum, our results unveil the process of reductive evolution which shaped the current genomes in two model fornicate parasites G. intestinalis and S. salmonicida.

Background: Helicobacter pylori infection is strongly associated with gastric cancer occurrence. However, it is unclear whether eradication therapy reduces the risk of gastric cancer occurrence. We evaluated whether H. pylori eradication reduces the risk of primary gastric cancer by using both risk ratio (RR) and risk difference (RD)., Methods: Searches of PubMed, EMBASE, Google scholar, the Cochrane Library, and the Japan Medical Abstracts Society as well as those registered in databases of the Cochrane Central Register of Controlled Trials, metaRegister of Controlled Trials, ClinicalTrials.gov, controlled-trials.com, UMIN-CTR, JMACCT-CTR, and JAPIC-CTI between January 1965 and March 2017, supplemented with manual screening. Randomized controlled trials (RCTs) in which eradication therapy were implemented for the interventional group but not for the control group, and assessed the subsequent occurrence of primary gastric cancer as the main outcome. Two authors independently reviewed articles and extracted data. Integrated results for all data were presented as RR and RD., Results: Seven studies met inclusion criteria. The reductions in risk of primary gastric cancer occurrence in terms of overall RR and RD were 0.67 (95% CI: 0.48 to 0.95) and -0.00 ([95% CI: -0.01 to 0.00]; number needed to treat: 125.5 [95% CI: 70.0 to 800.9]), respectively., Conclusions: The effectiveness of H. pylori eradication therapy in suppressing the occurrence of primary gastric cancer was significant and comparable to that of previous studies in terms of the estimated RR. However, the estimated RD was slight and not statistically significant.

Tumor necrosis factor α (TNF-α) induces the expression and secretion of interleukin 8 (IL-8), which contributes to synovitis in rheumatoid arthritis (RA). To elucidate the mechanism of the onset of RA, we used synovial fibroblasts without autoimmune inflammatory diseases and investigated MAPK signaling pathways in TNF-α-induced IL-8 expression. Synovial fibroblasts isolated from healthy dogs were characterized by flow cytometry, which were positive for the fibroblast markers CD29, CD44, and CD90 but negative for the hematopoietic cell markers CD14, CD34, CD45, and HLA-DR. TNF-α stimulated the secretion and mRNA expression of IL-8 in a time- and dose-dependent manner. ERK and JNK inhibitors attenuated TNF-α-induced IL-8 expression and secretion. TNF-α induced the phosphorylation of ERK1/2 and JNK1/2. TNF-α-induced IL-8 expression was attenuated both in ERK2- and JNK1-knockdown cells. TNF-α-induced ERK1/2 or JNK1/2 was observed in ERK2- or JNK1-knockdown cells, respectively, showing that there is no crosstalk between ERK2 and JNK1 pathways. These observations indicate that the individual activation of ERK2 and JNK1 pathways contributes to TNF-α-induced IL-8 expression in synovial fibroblasts, which appears to be involved in the progress in RA.

Background: Non-restorative sleep (NRS) was suggested to be associated with cardiovascular outcomes. However, causative factors for NRS have not been fully elucidated. This study aimed to clarify factors and their relationships with NRS to better understand the clinical and epidemiological implications of NRS and to develop a score that can objectively evaluate NRS status., Methods: Study subjects consisted of 9,788 community residents (age 53.6 ± 13.4 y). Subjective NRS as well as possible clinical and lifestyle factors for NRS were investigated by questionnaires. Other clinical parameters were obtained from personal records of information obtained at the baseline examination., Results: A total of 3,261 participants complained of NRS. Factors independently associated with subjective NRS were younger age (odds ratio = 1.43), use of a hypnotic drug (2.04), irregular sleep schedule (2.02), short sleep duration (<5 h, 11.7; 5-6 h, 4.81; 6-7 h, 2.40), frequent sleepiness (2.33), routine stress (4.63), no habitual exercise (1.61), nocturia symptoms (1.43), symptoms of gastroesophageal reflux disease (1.44), and depression (1.46) (all P <0.001). The NRS score comprised of these 10 factors was linearly associated with the frequency of subjective NRS (Ptrend <0.001). Frequency of individuals with a high NRS score was greater in women (52.3%) than in men (42.1%, P<0.001), while no clear association was observed with common risk factors for cardiovascular diseases., Conclusions: NRS was a phenomenon representing various clinical and lifestyle features. Careful attention should be paid to individuals with a high NRS score who might be at risk for mental fatigue and have unfavorable lifestyle factors.

Respiratory syncytial virus (RSV) infections occur every year worldwide. Most infants are infected with RSV by one year of age and are reinfected because immune responses after the first infection are too weak to protect against subsequent infections. In the present study, immune responses against RSV were investigated in order to obtain a better understanding of repetitive RSV infections in cotton rats. No detectable neutralizing antibody (NT) was developed after the first infection, and the second infection was not prevented. The results of histological examinations revealed severe inflammation, viral antigens were detected around bronchial epithelial cells, and infectious viruses were recovered from lung homogenates. Following the second infection neutralizing antibodies were significantly elevated, and CD8+ cells were activated in response to RSV-F253-265. No viral antigens was detected thereafter in lung tissues and infectious viruses were not recovered. Similar results were obtained in the present study using the subgroups A and B. These results support the induction of humoral and cellular immune responses following repetitive infections with RSV; however, these responses were insufficient to eliminate viruses in the first and second infections.

Removal of pathogenic organisms from reprocessed surgical instruments is essential to prevent iatrogenic infections. Some bacteria can make persistent biofilms on medical devices. Contamination of non-disposable equipment with prions also represents a serious risk to surgical patients. Efficient disinfection of prions from endoscopes and other instruments such as high-resolution cameras remains problematic because these instruments do not tolerate aggressive chemical or heat treatments. Herein, we develop a new washing system that uses both the alkaline and acidic water produced by electrolysis. Electrolyzed acidic water, containing HCl and HOCl as active substances, has been reported to be an effective disinfectant. A 0.15% NaCl solution was electrolyzed and used immediately to wash bio-contaminated stainless steel model systems with alkaline water (pH 11.9) with sonication, and then with acidic water (pH 2.7) without sonication. Two bacterial species (Staphylococcus aureus and Pseudomonas aeruginosa) and a fungus (Candida albicans) were effectively removed or inactivated by the washing process. In addition, this process effectively removed or inactivated prions from the stainless steel surfaces. This washing system will be potentially useful for the disinfection of clinical devices such as neuroendoscopes because electrolyzed water is gentle to both patients and equipment and is environmentally sound.

Background: Accumulating evidence has shown that methionine- and choline-deficient high fat (MCD+HF) diet induces the development of nonalcoholic fatty liver disease (NAFLD), in which elevated reactive oxygen species play a crucial role. We have reported that peroxiredoxin 4 (PRDX4), a unique secretory member of the PRDX antioxidant family, protects against NAFLD progression. However, the detailed mechanism and potential effects on the intestinal function still remain unclear., Methods & Results: Two weeks after feeding mice a MCD+HF diet, the livers of human PRDX4 transgenic (Tg) mice exhibited significant suppression in the development of NAFLD compared with wild-type (WT) mice. The serum thiobarbituric acid reactive substances levels were significantly lower in Tg mice. In contrast, the Tg small intestine with PRDX4 overexpression showed more suppressed shortening of total length and villi height, and more accumulation of lipid in the jejunum, along with lower levels of dihydroethidium binding. The enterocytes exhibited fewer apoptotic but more proliferating cells, and inflammation was reduced in the mucosa. Furthermore, the small intestine of Tg mice had significantly higher expression of cholesterol absorption-regulatory factors, including liver X receptor-α, but lower expression of microsomal triglyceride-transfer protein., Conclusion: Our present data provide the first evidence of the beneficial effects of PRDX4 on intestinal function in the reduction of the severity of NAFLD, by ameliorating oxidative stress-induced local and systemic injury. We can suggest that both liver and intestine are spared, to some degree, by the antioxidant properties of PRDX4.

An inactivated Japanese encephalitis virus (JEV) vaccine, which induces neutralizing antibodies, has been used for many years in Japan. In the present study, the JEV prM-E protein gene was cloned, inserted at the P/M junction of measles AIK-C cDNA, and an infectious virus was recovered. The JEV E protein was expressed in B95a cells infected with the recombinant virus. Cotton rats were inoculated with recombinant virus. Measles PA antibodies were detected three weeks after immunization. Neutralizing antibodies against JEV developed one week after inoculation, and EIA antibodies were detected three weeks after immunization. The measles AIK-C-based recombinant virus simultaneously induced measles and JEV immune responses, and may be a candidate for infant vaccines. Therefore, the present strategy of recombinant viruses based on a measles vaccine vector would be applicable to the platform for vaccine development.

Bone marrow stromal cells (BMSCs) are considered as candidates for regenerative therapy and a useful model for studying neuronal differentiation. The role of basic fibroblast growth factor (bFGF) in neuronal differentiation has been previously studied; however, the signaling pathway involved in this process remains poorly understood. In this study, we investigated the signaling pathway in the bFGF-induced neuronal differentiation of canine BMSCs. bFGF induced the mRNA expression of the neuron marker, microtubule associated protein-2 (MAP2) and the neuron-like morphological change in canine BMSCs. In the presence of inhibitors of fibroblast growth factor receptors (FGFR), phosphatidylinositol 3-kinase (PI3K) and Akt, i.e., SU5402, LY294002, and MK2206, respectively, bFGF failed to induce the MAP2 mRNA expression and the neuron-like morphological change. bFGF induced Akt phosphorylation, but it was attenuated by the FGFR inhibitor SU5402 and the PI3K inhibitor LY294002. In canine BMSCs, expression of FGFR-1 and FGFR-2 was confirmed, but only FGFR-2 activation was detected by cross-linking and immunoprecipitation analysis. Small interfering RNA-mediated knockdown of FGFR-2 in canine BMSCs resulted in the attenuation of bFGF-induced Akt phosphorylation. These results suggest that the FGFR-2/PI3K/Akt signaling pathway is involved in the bFGF-induced neuronal differentiation of canine BMSCs.

Introduction: Association of knee and low back pain with sleep disturbance is poorly understood. We aimed to clarify the independent and combined effects of these orthopedic symptoms on sleep in a large-scale general population., Methods: Cross-sectional data about sleep and knee/low back pain were collected for 9,611 community residents (53±14 years old) by a structured questionnaire. Sleep duration less than 6 h/d was defined as short sleep. Sleep quality and the presence of knee and low back pain were evaluated by dichotomous questions. Subjects who complained about knee or low back pains were graded by tertiles of a numerical response scale (NRS) score and a Roland-Morris disability questionnaire (RDQ) score respectively. Multivariate regression analyses were performed to determine the correlates of short sleep duration and poor sleep quality., Results: Frequency of participants who complained of the orthopedic symptoms was as follows; knee pain, 29.0%; low back pain, 42.0% and both knee and low back pain 17.6%. Both knee and low back pain were significantly and independently associated with short sleep duration (knee pain: odds ratio (OR) = 1.19, p<0.01; low back pain: OR = 1.13, p = 0.01) and poor sleep quality (knee pain: OR = 1.22, p<0.01; low back pain; OR = 1.57, p<0.01). The group in the highest tertile of the NRS or RDQ score had the highest risk for short sleep duration and poor sleep quality except for the relationship between the highest tertile of the RDQ score and short sleep duration.(the highest tertile of the NRS: OR for short sleep duration = 1.31, p<0.01; OR for poor sleep quality = 1.47, p<0.01; the highest tertile of the RDQ: OR for short sleep duration = 1.11, p = 0.12; OR for poor sleep quality = 1.81, p<0.01) Further, coincident knee and low back pain raised the odds ratios for short sleep duration (either of knee or low back pain: OR = 1.10, p = 0.06; both knee and low back pain: OR = 1.40, p<0.01) and poor sleep quality (either of knee or low back pain: OR = 1.61, p<0.01; both knee and low back pain: OR = 2.17, p<0.01)., Conclusion: Knee and low back pains were independently associated with short sleep duration and poor sleep quality. Further, they additively increased the correlation with these sleep problems in the general population.

Theaflavins are polyphenols found in black tea, whose physiological activities are not well understood. This study on mice evaluated the influence of a single oral administration of theaflavins on energy metabolism by monitoring the initial metabolic changess in skeletal muscle and brown adipose tissue (BAT). Oxygen consumption (VO2) and energy expenditure (EE) were increased significantly in mice treated with theaflavin rich fraction (TF) compared with the group administered vehicle alone. There was no difference in locomotor activity. Fasting mice were euthanized under anesthesia before and 2 and 5, 20-hr after treatment with TF or vehicle. The mRNA levels of uncoupling protein-1 (UCP-1) and peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) in BAT were increased significantly 2-hr after administration ofTF. The levels of UCP-3 and PGC-1α in the gastrocnemius muscle were increased significantly 2 and 5-hr after administration of TF. The concentration of phosphorylated AMP-activated protein kinase (AMPK) 1α was also increased significantly in the gastrocnemius 2 and 5-hr after treatment with TF. These results indicate that TF significantly enhances systemic energy expenditure, as evidenced by an increase in expression of metabolic genes.

Background: Birth outcomes, such as preterm birth, low birth weight (LBW), and small for gestational age (SGA), are crucial indicators of child development and health., Purpose: To evaluate whether home visits from public health nurses for high-risk pregnant women prevent adverse birth outcomes., Methods: In this quasi-experimental cohort study in Kyoto city, Japan, high-risk pregnant women were defined as teenage girls (range 14-19 years old), women with a twin pregnancy, women who registered their pregnancy late, had a physical or mental illness, were of single marital status, non-Japanese women who were not fluent in Japanese, or elderly primiparas. We collected data from all high-risk pregnant women at pregnancy registration interviews held at a public health centers between 1 July 2011 and 30 June 2012, as well as birth outcomes when delivered from the Maternal and Child Health Handbook (N = 964), which is a record of prenatal check-ups, delivery, child development and vaccinations. Of these women, 622 women were selected based on the home-visit program propensity score-matched sample (pair of N = 311) and included in the analysis. Data were analyzed between January and June 2014., Results: In the propensity score-matched sample, women who received the home-visit program had lower odds of preterm birth (odds ratio [OR], 0.62; 95% confidence interval [CI], 0.39 to 0.98) and showed a 0.55-week difference in gestational age (95% CI: 0.18 to 0.92) compared to the matched controlled sample. Although the program did not prevent LBW and SGA, children born to mothers who received the program showed an increase in birth weight by 107.8 g (95% CI: 27.0 to 188.5)., Conclusion: Home visits by public health nurses for high-risk pregnant women in Japan might be effective in preventing preterm birth, but not SGA.

Salinity represents a major abiotic stress factor that can adversely limit the production, quality and geographical distribution of crops. In this study we focused on dedifferentiated calli with fundamental cell functions, the salt tolerance of which had not been previously examined. The experimental approach was based on activation tagging without regeneration of plants for the identification of salt-tolerant mutants of Arabidopsis. Among 62,000 transformed calli that were screened, 18 potential mutants resistant to 150 mM NaCl were obtained. Thermal asymmetric interlaced (TAIL)-PCR was performed to determine the location of T-DNA integration in the genome. In one line, referred to as salt tolerant callus 1 (stc1), expression of a gene [At4g39800: myo-inositol-1-P-synthase 1 (MIPS1)] was considerably enhanced in calli. Plants regenerated from calli showed tolerance to salt in germination and subsequent growth. Retransformation of wild-type Arabidopsis with MIPS1 conferred salt tolerance, indicating that MIPS1 is the causal gene. The over-expression of MIPS1 increased the content of total inositol. The involvement of MIPS1 in salt tolerance through the fundamental cell growth has been proved in Arabidopsis.

Background: Attitudes of healthcare professionals regarding spontaneous reporting of adverse drug reactions (ADRs) in Japan are not well known, and Japan's unique system of surveillance, called early post-marketing phase vigilance (EPPV), may affect these reporting attitudes. Our objectives were to describe potential effects of EPPV and to test whether ADR seriousness, prominence, and frequency are related to changes in reporting over time., Methods: A manufacturer's database of spontaneous ADR reports was used to extract data from individual case safety reports for 5 drugs subject to EPPV. The trend of reporting and the time lag between ADR onset and reporting to the manufacturer were examined. The following indices for ADRs occurring with each drug were calculated and analyzed to assess reporting trends: Serious:Non-serious ratio, High prominence:Low prominence ratio, and High frequency:Low frequency ratio., Results: For all 5 drugs, the time lag between ADR onset and reporting to the manufacturer was shorter in the EPPV period than in the post-EPPV period. All drugs showed higher Serious:Non-serious ratios in the post-EPPV period. No specific patterns were observed for the High prominence:Low prominence ratio. The High frequency:Low frequency ratio for peginterferon alpha-2a and sevelamer hydrochloride decreased steadily throughout the study period., Conclusions: Healthcare professionals may be more likely to report serious ADRs than to report non-serious ADRs, but the effect of event prominence on reporting trends is still unclear. Factors associated with ADR reporting attitude in Japan might be different from those in other countries because of EPPV and the involvement of medical representatives in the spontaneous reporting process. Pharmacovigilance specialists should therefore be cautious when comparing data between different time periods or different countries. Further studies are needed to elucidate the underlying mechanism of spontaneous ADR reporting in Japan.

A strong causal link exists between psychological stress and insulin resistance as well with hypertension. Meanwhile, stress-related responses play critical roles in glucose metabolism in hypertensive patients. As clinical trials suggest that angiotensin-receptor blocker delays the onset of diabetes in hypertensive patients, we investigated the effects of irbesartan on stress-induced adipose tissue inflammation and insulin resistance. C57BL/6J mice were subjected to 2-week intermittent restraint stress and orally treated with vehicle, 3 and 10 mg/kg/day irbesartan. The plasma concentrations of lipid and proinflammatory cytokines [Monocyte Chemoattractant Protein-1 (MCP-1), tumor necrosis factor-α, and interleukin-6] were assessed with enzyme-linked immunosorbent assay. Monocyte/macrophage accumulation in inguinal white adipose tissue (WAT) was observed with CD11b-positive cell counts and mRNA expressions of CD68 and F4/80 using immunohistochemistry and RT-PCR methods respectively. The mRNA levels of angiotensinogen, proinflammatory cytokines shown above, and adiponectin in WAT were also assessed with RT-PCR method. Glucose metabolism was assessed by glucose tolerance tests (GTTs) and insulin tolerance tests, and mRNA expression of insulin receptor substrate-1 (IRS-1) and glucose transporter 4 (GLUT4) in WAT. Restraint stress increased monocyte accumulation, plasma free fatty acids, expression of angiotensinogen and proinflammatory cytokines including MCP-1, and reduced adiponectin. Irbesartan reduced stress-induced monocyte accumulation in WAT in a dose dependent manner. Irbesartan treatment also suppressed induction of adipose angiotensinogen and proinflammatory cytokines in WAT and blood, and reversed changes in adiponectin expression. Notably, irbesartan suppressed stress-induced reduction in adipose tissue weight and free fatty acid release, and improved insulin tolerance with restoration of IRS-1 and GLUT4 mRNA expressions in WAT. The results indicate that irbesartan improves stress-induced adipose tissue inflammation and insulin resistance. Our results suggests that irbesartan treatment exerts additive benefits for glucose metabolism in hypertensive patients with mental stress.

Background and Aim: We previously identified an anti-inflammatory compound, zonarol, a hydroquinone isolated from the brown algae Dictyopteris undulata as a marine natural product. To ascertain the in vivo functions of zonarol, we examined the pharmacological effects of zonarol administration on dextran sulfate sodium (DSS)-induced inflammation in a mouse model of ulcerative colitis (UC). Our goal is to establish a safe and effective cure for inflammatory bowel disease (IBD) using zonarol., Methods and Results: We subjected Slc:ICR mice to the administration of 2% DSS in drinking water for 14 days. At the same time, 5-aminosalicylic acid (5-ASA) at a dose of 50 mg/kg (positive control) and zonarol at doses of 10 and 20 mg/kg, were given orally once a day. DSS-treated animals developed symptoms similar to those of human UC, such as severe bloody diarrhea, which were evaluated by the disease activity index (DAI). Treatment with 20 mg/kg of zonarol, as well as 5-ASA, significantly suppressed the DAI score, and also led to a reduced colonic ulcer length and/or mucosal inflammatory infiltration by various immune cells, especially macrophages. Zonarol treatment significantly reduced the expression of pro-inflammatory signaling molecules, and prevented the apoptosis of intestinal epithelial cells. Finally, zonarol protected against in vitro lipopolysaccharide (LPS)-induced activation in the RAW264.7 mouse macrophage cell line., Conclusions: This is the first report that a marine bioproduct protects against experimental UC via the inhibition of both inflammation and apoptosis, very similar to the standard-of-care sulfasalazine, a well-known prodrug that releases 5-ASA. We believe that the oral administration of zonarol might offer a better treatment for human IBDs than 5-ASA, or may be useful as an alternative/additive therapeutic strategy against UC, without any evidence of side effects.

Objective: Therapeutic options for Parkinson's disease mainly consist of L-dopa and dopamine agonists. However, in Japan, the product labeling of the ergot dopamine agonists, cabergoline and pergolide, was revised in April 2007 due to the risk of developing cardiac valvulopathy. Here, we describe the prescribing trends of anti-Parkinson drugs from 2005 through 2010 in Japan, and examined whether these trends changed after the drug safety measures in 2007., Methods and Patients: We used medical claim data from January 2005 to December 2010 for Parkinson's disease patients older than 30 years who were prescribed anti-Parkinson drugs. We calculated the proportion of patients prescribed each drug for each year, and compared the proportions of first-line drugs prescribed before and after April 2007. We also examined the prescription variations of cabergoline/pergolide users one year before or after April 2007., Results: L-dopa was the most frequently prescribed drug for Parkinson's disease (2005, 58%; 2010, 51%). The proportion of patients prescribed ergot dopamine agonists markedly decreased and non-ergot dopamine agonists increased after 2007. Among first-line drugs, the proportion of non-ergot agents increased after April 2007. Among 54 cabergoline/pergolide users, 24 (44%) discontinued these drugs, nine of whom switched to non-ergot agents., Conclusion: L-dopa was the mainstay of Parkinson's disease treatment between 2005 and 2010 in Japan. There was a decrease in ergot agents and an increase in non-ergot agents prescribed after the regulatory actions in 2007.

Macrophage infiltration of skin GVHD lesions correlates directly with disease severity, but the mechanisms underlying this relationship remain unclear and GVHD with many macrophages is a therapeutic challenge. Here, we characterize the macrophages involved in GVHD and report that dexamethasone palmitate (DP), a liposteroid, can ameliorate such GVHD by inhibiting macrophage functions. We found that host-derived macrophages could exacerbate GVHD in a mouse model through expression of higher levels of pro-inflammatory TNF-α and IFN-γ, and lower levels of anti-inflammatory IL-10 than resident macrophages in mice without GVHD. DP significantly decreased the viability and migration capacity of primary mouse macrophages compared to conventional dexamethasone in vitro. DP treatment on day 7 and day 14 decreased macrophage number, and attenuated GVHD score and subsequent mortality in a murine model. This is the first study to provide evidence that therapy for GVHD should be changed on the basis of infiltrating cell type.

Objectives: This study aims to clarify the cause of background coloration in the epithelia between each dilated intra papillary capillary loop in esophago-pharyngeal squamous cell carcinoma., Design: This is a single center retrospective study including 124 patients with 160 lesions who underwent esophagogastroduodenoscopy in Nagasaki University Hospital from September 2007 to March 2012; a detailed comparison between endoscopic images and pathology was performed. Immunohistological assessment using anti-human hemoglobin antibody (anti-Hb Ab) was performed to verify the presence of hemoglobin (Hb) component in the cancer cells. Real-time polymerase chain reaction (RT-PCR) and in situ hybridization (ISH) on Hb-β mRNA were performed to assess the production of Hb component within the cancer cells., Results: A strong positivity for anti-Hb Ab was observed in the squamous cell carcinoma area, whereas non-cancerous mucosa showed no immunopositivity for Hb. The concordance rate between anti-Hb Ab immunoreactivity and the presence of BC was as high as 80.9%. The amount of Hb-β mRNA expression was three times higher in cancer tissues compared with the surrounding non-cancerous mucosa. ISH images showed that the expression exclusively occurred in cancer cells, indicating that Hb is probably produced within cancer cells., Conclusions: The background coloration observed is partly due to an extravascular component of Hb. RT-PCR and ISH analyses indicate that Hb is produced within cancer cells.

In repetitive measurements of flow-mediated dilatation (FMD), the duration of the interval between measurements remains controversial. In this pilot study, we conducted three sequential measurements of low-flow-mediated constriction (L-FMC), FMD and flow-mediated total dilation (FMTD; L-FMC+ FMD) at baseline and intervals of 15 and 60 min in 30 healthy males. FMD15, L-FMC15, and FMTD15 were significantly lower than the respective first measurements, but all indices showed full recovery at 60 min in all subjects. The baseline diameter was slightly increased at 15 min and restored at 60 min, but the maximum diameter, and the baseline and reactive flow velocity unchanged. We examined the relationship between recovery rate of FMTD at 15 min (FMTD-R) and cardio-ankle vascular index (CAVI). Univariate analysis showed moderate correlation between FMTD-R, and CAVI and L-FMC0. Patients were divided according to FMTD-R value; the low-FMTD-R group [below the median value (-26.2%)] included a significantly higher proportion of smokers and higher CAVI values than the high-FMTD-R group. The reproducibility of FMTD and FMTD-R was evaluated in another group of 25 healthy subjects. The range of variation across measurements was 1.1% for FMTD and 4.6% for FMTD-R; with intraclass correlation coefficients of 0.93 and 0.95, respectively. The present study demonstrated blunted recovery of FMD within 15 min, suggesting the need for selection of a more adequate interval between measurements to avoid underestimation of FMD in subsequent measurements. The findings demonstrated the reproducibility of FMTD-R and FMTD measurements, and that FMTD-R might be involved in arterial stiffness and early vascular impairment in the healthy subjects.

Background: Several studies have reported osteomyelitis of the jaw (OMJ) as a side effect of bisphosphonates (BPs), and the risk of oral BPs has been recently clarified. However, other systemic risk factors of OMJ remain unclear. Importantly, the possibility of risk classification based on the clinical characteristics of patients has not been explored. Here, we clarified risk factors of OMJ and evaluate the predictive accuracy of risk indices in osteoporosis patients., Methods: We performed sub-analysis using a database developed for a retrospective cohort study in patients taking medications for osteoporosis at Kyoto University Hospital. Risk indices for OMJ were constructed using logistic regression analysis, and odds ratios (OR) for OMJ cases and 95% confidence intervals (CI) were estimated. Potential risk factors included in the statistical analysis were age; sex; diabetes; use of oral BPs, corticosteroids, cancer chemotherapy, antirheumatic drugs, and biologic agents; and their interactions. Risk indices were calculated by the sum of potential risk factors of an individual patient multiplied by the regression coefficients. The discriminatory power of the risk indices was assessed by receiver operating characteristic (ROC) analysis., Results: In analysis of all patients, oral BPs (OR: 4.98, 95% CIs: 1.94-12.75), age (OR: 1.28, 95% CI: 1.06-1.60) and sex-chemotherapy interaction (OR: 11.70, 95% CI: 1.46-93.64) were significant risk factors of OMJ. Areas under the ROC curves of these risk indices provided moderate sensitivity or specificity regardless of group (0.683 to 0.718)., Conclusions: Our data suggest that oral BP use, age, and sex-chemotherapy are predictors of OMJ in osteoporosis patients. The risk indices are moderately high, and allow the prediction of OMJ incidence.

Background: Poor retention in the care of patients with human immunodeficiency virus (HIV) is associated with adverse patient outcomes such as antiretroviral therapy failure and death. Therefore, appropriate case management is required for better patient retention; however, which intervention in case management is important has not been fully investigated. Meanwhile, in Japan, each local government is required to organize mental health services for patients with HIV so that a case manager at an HIV care facility can utilize them, but little is known about the association between implementation of the services and loss to follow-up. Therefore, we investigated that by a nested case-control study., Methods: The target population consisted of all patients with HIV who visited Osaka National Hospital, the largest HIV care facility in western Japan, between 2000 and 2010. Loss to follow-up was defined as not returning for follow-up care more than 1 year after the last visit. Independent variables included patient demographics, characteristics of the disease and treatment, and whether the patients have received mental health services. For each case, three controls were randomly selected and matched., Results: Of the 1620 eligible patients, 88 loss to follow-up cases were identified and 264 controls were matched. Multivariate-adjusted conditional logistic regression revealed that loss to follow-up was less frequent among patients who had received mental health services implemented by their case managers (adjusted odds ratio [95% confidence interval] 0.35 [0.16-0.76]). Loss to follow-up also occurred more frequently in patients who did not receive antiretroviral therapy (adjusted odds ratio [95% confidence interval], 7.51 [3.34-16.9]), who were under 30 years old (2.74 [1.36-5.50]), or who were without jobs (3.38 [1.58-7.23])., Conclusion: Mental health service implementation by case managers has a significant impact on patient retention.