Supplementary Material for: The Role of Proteasome Inhibitor MG132 in 2,4-Dinitrofluorobenzene-Induced Atopic Dermatitis in NC/Nga Mice

<b><i>Background:</i></b> Although immunosuppressants for therapy of atopic dermatitis (AD) are still being sought, proteasome inhibitors are also potential candidates for the treatment of AD. Proteasome inhibitors exert various effects by blocking proteasomal degradation and help regulate processes such as apoptosis induction via caspase-9, cell cycle progression via cyclins, NF-κB inactivation via IκB, and downregulation of antigen cross-presentation. The cells targeted by proteasome inhibitors are therefore activated cells undergoing proliferation or differentiation, and antigen-presenting cells carrying out protein degradation. <b><i>Objectives:</i></b> This study investigated the therapeutic effects and side effects of a proteasome inhibitor, MG132, on the treatment of AD. <b><i>Methods:</i></b> AD-like disease in NC/Nga mice housed under specific pathogen-free conditions was induced by repeated application of 2,4-dinitrofluorobenzene (DNFB). Disease progression was evaluated by inflammation score, histopathology, and serum IgE level, and the effects of systemic MG132 administration were investigated. The percentages and absolute numbers for each population of Th1, Th2, and Th17 cells in the axillary lymph nodes were analyzed by flow cytometry. <b><i>Results:</i></b> DNFB application increased the expression of a unique major histocompatibility complex class I mutant molecule D/L^dm7 in dendritic cells (DCs), and increased Th1 and Th17 cells in NC/Nga mice. In vivo MG132 administration to NC/Nga mice with DNFB-induced dermatitis reduced Th17 cells but maintained the level of Th1 cells, resulting in the alleviation of dermatitis lesions by decreasing both serum IgE hyperproduction and mast cell migration. To understand the mechanisms maintaining Th1 cell levels following in vivo MG132-administration, we focused on the role of proteasomes regulating D/L^dm7 expression. Interestingly, 20S proteasome activity was higher in NC/Nga DCs than in BALB/c DCs. In vitro MG132 administration partially increased D/L^dm7 expression in a dose-dependent manner during DC maturation, and induced IFN-γ production from autoreactive CD8+ T cells but not from CD4+ T cells following coculturing with D/L^dm7-upregulated DCs. <b><i>Conclusion:</i></b> Although MG132 administration temporarily alleviated AD pathogenesis in NC/Nga mice, prolonged MG132 treatment may result in immunopathogenesis leading to chronic AD due to its side effect of maintaining Th1 levels via autoreactive CD8+ T cells

Supplementary Material for: Social Function and Frontopolar Activation during a Cognitive Task in Patients with Bipolar Disorder

<b><i>Background:</i></b> It is important to understand the neural basis of functional impairments in patients with bipolar disorder (BD) in order to be able to address the recovery. Recently, neurocognitive impairment emerged as a predictor of psychosocial function. A number of functional brain imaging studies have shown that social function is associated with activation of the prefrontal cortex during a cognitive task in healthy adults, and in patients with major depressive disorder and schizophrenia. However, few studies have been conducted in patients with BD. <b><i>Methods:</i></b> We performed multichannel near-infrared spectroscopy (NIRS) imaging to investigate the activation of the prefrontal cortex during a verbal fluency task (VFT). We also used the Social Adaptation Self-Evaluation Scale (SASS) to assess social functioning in patients with BD. Thirty-three depressed patients with BD and 65 age-, gender- and task performance-matched healthy controls (HCs) participated in this study. <b><i>Results:</i></b> Depressed patients with BD showed reduced activation in the broader bilateral prefrontal cortex during the VFT compared to HCs. Moreover, a significant positive correlation was observed between the total SASS scores and right prefrontal activation in patients with BD. In the SASS subscores, the interest and motivation factor was also positively correlated with frontopolar activation. <b><i>Conclusions:</i></b> These results suggest an association between social function and prefrontal activation in depressed patients with BD. The present study provides evidence that NIRS imaging could be helpful in understanding the neural basis of social function

Supplementary Material for: Risk Factors and Clinical Features in Cashew Nut Oral Food Challenges

<p><b><i>Background:</i></b> Cashew nuts (CN) are capable of causing severe allergic reactions. However, little has been reported about the details of CN oral food challenges (OFC). <b><i>Methods:</i></b> CN-specific IgE (sIgE) levels were measured for 1 year in 66 patients who underwent an OFC with >3 g CN for diagnosis or confirmation of tolerance acquisition between June 2006 and August 2014. We retrospectively analyzed the OFC and patient background. <b><i>Results:</i></b> The median (IQR) age of the 66 patients (48 boys/men and 18 girls/women) was 7.0 years (5.7-8.8). Twelve patients (18.2%) had a positive OFC result; 6 of 8 (75%) patients with a history of an immediate reaction to CN failed the OFC. Anaphylaxis was experienced by 5 of these 12 (42%) patients. A history of an immediate reaction to CN and the CN sIgE levels were significantly different for patients with a positive or negative OFC result (<i>p</i> < 0.01). Among patients without a previous immediate reaction to CN, the 95% positive predictive value (PPV) for the CN sIgE level for a positive OFC result was 66.1 kU_A/L. <b><i>Conclusions:</i></b> A history of an immediate reaction to CN and high CN sIgE were risk factors for a positive OFC result. The number of positive OFC results was relatively low, but there was a high probability of anaphylaxis. We should consider the indication of OFC carefully for patients with a history of immediate reactions to CN and avoid OFC for patients without such a history whose CN sIgE values are >66.1 kU_A/L (95% PPV).</p

Supplementary Material for: Colorectal Cancer with BRAF D594G Mutation Is Not Associated with Microsatellite Instability or Poor Prognosis

<i>Objective:</i><i>BRAF</i> D594G mutations in colorectal cancer patients are not clearly understood. We retrospectively investigated the clinicopathological features of colorectal cancers with <i>BRAF</i> D594G mutations. <i>Methods:</i> We selected 908 colorectal cancer patients who underwent surgical resection from January 2008 to January 2013, and assessed <i>BRAF</i>, <i>KRAS</i>, microsatellite instability, and CpG island methylator phenotype (CIMP). <i>Results:</i> We detected <i>BRAF</i> D594G in 7 patients and <i>BRAF</i> V600E in 45 patients. The clinicopathological features of cancers with <i>BRAF</i> D594G mutation were similar to those with <i>BRAF</i> wild-type, but differed from those with <i>BRAF</i> V600E mutations. Regarding microsatellite instability status, 44.4% of cases with <i>BRAF</i>V600E mutations exhibited high microsatellite instability, compared to 14.3% of those with <i>BRAF</i> D594G mutations and 4.4% of those with <i>BRAF</i> wild-type. There were no CIMP-positive tumors in cancers with <i>BRAF</i> D594G mutations, whereas 67.8% of tumors with <i>BRAF</i> V600E mutations were CIMP-positive. In stage IV cancers, the survival rates of patients at 2 years were 8.5, 50.0, and 68.2% in the <i>BRAF</i> V600E mutation, <i>BRAF</i> D594G mutation, and <i>BRAF</i> wild-type groups, respectively.<i>Conclusion:</i> Colorectal cancers with <i>BRAF</i> D594G mutations exhibit similar clinicopathological features, microsatellite instability status, and prognosis as those with <i>BRAF</i> wild-type

Erratum: Reduced Abundance of Butyrate-Producing Bacteria Species in the Fecal Microbial Community in Crohn's Disease

<b><i>Background:</i></b> The global alteration of the gut microbial community (dysbiosis) plays an important role in the pathogenesis of inflammatory bowel diseases (IBDs). However, bacterial species that characterize dysbiosis in IBD remain unclear. In this study, we assessed the alteration of the fecal microbiota profile in patients with Crohn's disease (CD) using 16S rRNA sequencing. <b><i>Summary:</i></b> Fecal samples from 10 inactive CD patients and 10 healthy individuals were subjected to 16S rRNA sequencing. The V3-V4 hypervariable regions of 16S rRNA were sequenced by the Illumina MiSeq™II system. The average of 62,201 reads per CD sample was significantly lower than the average of 73,716 reads per control sample. The genera <i>Bacteroides</i>, <i>Eubacterium</i>, <i>Faecalibacterium</i> and <i>Ruminococcus</i> significantly decreased in CD patients as compared to healthy controls. In contrast, the genera <i>Actinomyces</i> and <i>Bifidobacterium</i> significantly increased in CD patients. At the species level, butyrate-producing bacterial species, such as <i>Blautia faecis</i>, <i>Roseburia inulinivorans</i>, <i>Ruminococcus torques</i>, <i>Clostridium lavalense</i>, <i>Bacteroides uniformis</i> and <i>Faecalibacterium prausnitzii</i> were significantly reduced in CD patients as compared to healthy individuals (p < 0.05). These results of 16S rRNA sequencing were confirmed in additional CD patients (n = 68) and in healthy controls (n = 46) using quantitative PCR. The abundance of <i>Roseburia inulinivorans</i> and <i>Ruminococcus torques</i> was significantly lower in C-reactive protein (CRP)-positive CD patients as compared to CRP-negative CD patients (p < 0.05). <b><i>Key Message:</i></b> The dysbiosis of CD patients is characterized by reduced abundance of multiple butyrate-producing bacteria species