Supplementary Material for: Loss of Heterozygosity of PTEN (Encoding Phosphate and Tensin Homolog) Associated with Elevated HER2 Expression Is an Adverse Prognostic Indicator in Gastric Cancer

<b><i>Objective:</i></b> PTEN (the encoding phosphate and tensin homolog) is a well-known cancer suppressor gene and its mutation and loss of heterozygosity (LOH) occurs in various types of carcinomas. This study aimed to examine the association between LOH of PTEN and prognosis in HER2-expressing and nonexpressing gastric cancer patients. <b><i>Methods:</i></b> Fresh-frozen tumor samples of 221 gastric cancer patients with a primary diagnosis of gastric carcinoma were examined for LOH of PTEN. The results were compared with pathological parameters and the HER2 status. To elucidate the role of LOH of PTEN, the activation of the PI3K/AKT pathway was examined immunohistochemically using a phosphorylation-specific antibody. <b><i>Results:</i></b> LOH of PTEN was observed in 20% of the patients (39 of 195 cases). LOH of PTEN was associated with vascular involvement (25 of 39 cases; p = 0.0083), equivocal to positive staining for HER2 (p = 0.0080), and phospho-Akt expression (p = 0.0067). Patients with HER2-expressing gastric cancer with LOH of PTEN had a significantly worse prognosis (p = 0.0050). <b><i>Conclusions:</i></b> Although HER2 expression itself was not a prognostic factor, the combination of HER2 expression and LOH of PTEN exacerbates the malignant potential of gastric cancer through its proliferative function. © 2014 S. Karger AG, Base

Supplementary Material for: Expression Profiling of MicroRNAs in the Inner Ear of Elderly People by Real-Time PCR Quantification

The molecular mechanisms underlying age-related hearing loss are unknown, and currently, there is no treatment for this condition. Recent studies have shown that microRNAs (miRNAs) and age-related diseases are intimately linked, suggesting that some miRNAs may present attractive therapeutic targets. In this study, we obtained 8 human temporal bones from 8 elderly subjects at brain autopsy in order to investigate the expression profile of miRNAs in the inner ear with miRNA arrays. A mean of 478 different miRNAs were expressed in the samples, of which 348 were commonly expressed in all 8 samples. Of these, levels of 16 miRNAs significantly differed between young elderly and old elderly subjects. miRNAs, which play important roles in inner ear development, were detected in all samples, i.e., in both young and old elderly subjects, whether with or without hearing loss. Our results suggest that these miRNAs play important roles not only in development, but also in the maintenance of inner ear homeostasis

Supplementary Material for: The Effects of All-Trans Retinoic Acid on the Induction of Oral Tolerance in a Murine Model of Bronchial Asthma

<b><i>Background:</i></b> Active suppression induced by regulatory T (Treg) cells is reported to be one of the mechanisms involved in oral tolerance. All-trans retinoic acid (ATRA) has been reported to affect Treg cell differentiation. The present study examined the effects of ATRA on the induction of oral tolerance in a murine model of bronchial asthma. <b><i>Methods:</i></b> BALB/c mice were sensitized to and challenged with ovalbumin (OVA) through feeding followed by OVA challenges. In some study groups ATRA was orally administered concomitantly with OVA feeding either in the presence or absence of the retinoic acid receptor antagonist LE135. Lung CD4⁺ T cells were isolated from mice exposed to ATRA and/or OVA, and transferred to control mice. Airway hyperresponsiveness (AHR), cell counts and cytokine levels in bronchoalveolar lavage (BAL) fluid, and lung histology were assessed. <b><i>Results:</i></b> Concomitant administration of ATRA with OVA ameliorated AHR, airway eosinophilia, elevation of cytokines in BAL fluid and goblet cell metaplasia. The proportion of Treg cells in the lungs was increased in mice treated with OVA and ATRA, as compared to those treated with OVA only. Transfer of lung CD4⁺ T cells from mice treated with OVA and ATRA induced suppression of AHR and airway inflammation. LE135 completely reversed the effects of ATRA on AHR, airway allergic inflammation and the number of Treg cells in the lungs. <b><i>Conclusion:</i></b> These data suggested that oral administration of ATRA with OVA had the potential to enhance oral tolerance in this murine model of bronchial asthma. These effects were mediated, at least in part, by Treg cell expansion

Supplementary Material for: Clinicopathological Characteristics, Surgery and Survival Outcomes of Patients with Duodenal Gastrointestinal Stromal Tumors

<i>Background:</i> Duodenal gastrointestinal stromal tumors (GISTs) are a rare subset of GISTs (3-5%), and their clinicopathological features have not been fully described. The purpose of this retrospective study was to examine these characteristics and compare the operative procedures. <i>Methods:</i> Twenty-five patients with duodenal GIST underwent complete resection, local resection (LR) or pancreaticoduodenectomy (PD) from 1990 to 2014 at our 2 hospitals. We analyzed patient characteristics, treatments, histological examinations, postoperative complications and survival outcomes. <i>Results:</i> Twelve patients (48%) with no symptoms were incidentally diagnosed for unrelated reasons. Sixteen patients (64%) had c-<i>kit</i> mutations while 6 (24%) were wild-type, including 4 with a history of neurofibromatosis type 1. Comparing LR (n = 16) and PD (n = 9), the recurrence-free survival rate was significantly worse for PD. On multivariate analysis, however, tumor size was an independent and significant prognostic factor, but not operative procedure. There was no body weight change with LR, but body weight decreased by 7% with PD.<i>Conclusion:</i> Duodenal GISTs had different characteristic genetic mutations compared to other GISTs. LR for duodenal GISTs appears to be oncologically and nutritionally feasible

Supplementary Material for: The balance of CD8-positive T cells and PD-L1 expression in the myocardium predicts prognosis in lymphocytic fulminant myocarditis

Introduction: The clinical significance and prognostic value of T cell involvement and programmed cell death-1 (PD-1)/programmed cell death ligand-1 (PD-L1) have not been established in lymphocytic fulminant myocarditis (FM). We investigated the prognostic impact of the number of CD4+, CD8+, FoxP3+, and PD-1+ T cells, as well as PD-L1 expression, in cardiomyocytes in lymphocytic FM. Methods: This is a single-center observational cohort study. Myocardial tissue was obtained from 16 consecutive patients at lymphocytic FM onset. The median follow-up was 140 days. Cardiac events were defined as a composite of cardiac death and left ventricular-assist device implantation. CD4, CD8, FoxP3, PD-1, and PD-L1 immunostaining was performed on myocardial specimens. Results: The median age of the patients was 52 years (seven men and nine women). There was no significant difference in the number of CD4+ cells. The number of CD8+ cells and the CD8+/CD4+ T cell ratio were higher in the cardiac event group (Event+) than in the group without cardiac events (Event−) (P = 0.048 and P = 0.022, respectively). The number of FoxP3+ T cells was higher in the Event+ group (P = 0.049). Although there was no difference in the number of PD-1+ cells, cardiomyocyte PD-L1 expression was higher in the Event+ group (P = 0.112). Event-free survival was worse in the group with a high CD8+ cell count (P = 0.012) and high PD-L1 expression (P = 0.049). When divided into three groups based on the number of CD8+ cells and PD-L1 expression (CD8highPD-L1high [n = 8], CD8lowPD-L1high [n = 1], and CD8lowPD-L1low [n = 7]), the CD8highPD-L1high group demonstrated the worst event-free survival, while the CD8lowPD-L1high group had a favorable prognosis without cardiac events (P = 0.041). Conclusion: High myocardial expression of CD8+ T cells and PD-L1 may predict a poor prognosis in lymphocytic FM

Supplementary Material for: The balance of CD8-positive T cells and PD-L1 expression in the myocardium predicts prognosis in lymphocytic fulminant myocarditis

Introduction: The clinical significance and prognostic value of T cell involvement and programmed cell death-1 (PD-1)/programmed cell death ligand-1 (PD-L1) have not been established in lymphocytic fulminant myocarditis (FM). We investigated the prognostic impact of the number of CD4+, CD8+, FoxP3+, and PD-1+ T cells, as well as PD-L1 expression, in cardiomyocytes in lymphocytic FM. Methods: This is a single-center observational cohort study. Myocardial tissue was obtained from 16 consecutive patients at lymphocytic FM onset. The median follow-up was 140 days. Cardiac events were defined as a composite of cardiac death and left ventricular-assist device implantation. CD4, CD8, FoxP3, PD-1, and PD-L1 immunostaining was performed on myocardial specimens. Results: The median age of the patients was 52 years (seven men and nine women). There was no significant difference in the number of CD4+ cells. The number of CD8+ cells and the CD8+/CD4+ T cell ratio were higher in the cardiac event group (Event+) than in the group without cardiac events (Event−) (P = 0.048 and P = 0.022, respectively). The number of FoxP3+ T cells was higher in the Event+ group (P = 0.049). Although there was no difference in the number of PD-1+ cells, cardiomyocyte PD-L1 expression was higher in the Event+ group (P = 0.112). Event-free survival was worse in the group with a high CD8+ cell count (P = 0.012) and high PD-L1 expression (P = 0.049). When divided into three groups based on the number of CD8+ cells and PD-L1 expression (CD8highPD-L1high [n = 8], CD8lowPD-L1high [n = 1], and CD8lowPD-L1low [n = 7]), the CD8highPD-L1high group demonstrated the worst event-free survival, while the CD8lowPD-L1high group had a favorable prognosis without cardiac events (P = 0.041). Conclusion: High myocardial expression of CD8+ T cells and PD-L1 may predict a poor prognosis in lymphocytic FM

Supplementary Material for: The balance of CD8-positive T cells and PD-L1 expression in the myocardium predicts prognosis in lymphocytic fulminant myocarditis

Introduction: The clinical significance and prognostic value of T cell involvement and programmed cell death-1 (PD-1)/programmed cell death ligand-1 (PD-L1) have not been established in lymphocytic fulminant myocarditis (FM). We investigated the prognostic impact of the number of CD4+, CD8+, FoxP3+, and PD-1+ T cells, as well as PD-L1 expression, in cardiomyocytes in lymphocytic FM. Methods: This is a single-center observational cohort study. Myocardial tissue was obtained from 16 consecutive patients at lymphocytic FM onset. The median follow-up was 140 days. Cardiac events were defined as a composite of cardiac death and left ventricular-assist device implantation. CD4, CD8, FoxP3, PD-1, and PD-L1 immunostaining was performed on myocardial specimens. Results: The median age of the patients was 52 years (seven men and nine women). There was no significant difference in the number of CD4+ cells. The number of CD8+ cells and the CD8+/CD4+ T cell ratio were higher in the cardiac event group (Event+) than in the group without cardiac events (Event−) (P = 0.048 and P = 0.022, respectively). The number of FoxP3+ T cells was higher in the Event+ group (P = 0.049). Although there was no difference in the number of PD-1+ cells, cardiomyocyte PD-L1 expression was higher in the Event+ group (P = 0.112). Event-free survival was worse in the group with a high CD8+ cell count (P = 0.012) and high PD-L1 expression (P = 0.049). When divided into three groups based on the number of CD8+ cells and PD-L1 expression (CD8highPD-L1high [n = 8], CD8lowPD-L1high [n = 1], and CD8lowPD-L1low [n = 7]), the CD8highPD-L1high group demonstrated the worst event-free survival, while the CD8lowPD-L1high group had a favorable prognosis without cardiac events (P = 0.041). Conclusion: High myocardial expression of CD8+ T cells and PD-L1 may predict a poor prognosis in lymphocytic FM

Supplementary Material for: Prognostic nutritional index after introduction of atezolizumab with bevacizumab predicts prognosis in advanced hepatocellular carcinoma: A multicenter study

Introduction: Atezolizumab plus bevacizumab (Atez/Bev) is the preferred treatment for advanced hepatocellular carcinoma (HCC). However, biomarkers of therapeutic efficacy have remained unclear. We took a retrospective approach to explore the role of prognostic nutritional index (PNI) for predicting the outcomes of Atez/Bev treatment. Methods: One hundred twenty-five HCC patients were enlisted; these patients received Atez/Bev treatment and underwent dynamic computerized tomography/magnetic resonance imaging to determine the treatment response on at least one occasion between October 2020 and January 2023, and their PNI before treatment and at the beginning of the second cycle (PNI-2c) was evaluated. Results: During the initial evaluation, 2 (2%), 28 (22%), 70 (56%), and 25 (20%) patients exhibited a complete response (CR), partial response (PR), stable disease (SD), and progressive disease (PD), respectively. Patients with non-PD tended to have higher PNI at baseline and PNI-2c than those with PD (p = 0.245 and 0.122, respectively), with optimal baseline PNI and PNI-2c cut-off values of 42.6 and 40.4, respectively. PNI at baseline could not be used to predict overall survival (OS) or progression-free survival (PFS). However, PNI-2c predicted OS and PFS (PNI-2c ≥ 40.4 vs. < 40.4: 25.3 vs. 16.2 months, P = 0.008 for OS; 12.7 vs. 8.4 months, P = 0.036 for PFS). A multivariate analysis showed a significant association between PNI-2c and OS. Conclusions: PNI-2c is a predictor of prognosis in HCC patients treated with Atez/Bev therapy