Prevalence of plasmid-mediated AmpC β-lactamase-producing Escherichia coli and spread of the ST131 clone among extended-spectrum β-lactamase-producing E. coli in Japan.

In 2010, a total of 1327 clinical Escherichia coli isolates from five hospitals in the Kyoto and Shiga regions of Japan were analysed by PCR. The prevalences of plasmid-mediated AmpC β-lactamase (pAmpC)-producers, extended-spectrum β-lactamase (ESBL)-producers and co-producers of pAmpC and ESBL were 1.7%, 9.7% and 0.3%, respectively. Less than one-half of the pAmpC-producers were reported to be resistant to third-generation cephalosporins, cephamycins and β-lactam/β-lactam inhibitors using the old 2009 Clinical and Laboratory Standards Institute (CLSI) breakpoints. CMY-2 was the most prevalent pAmpC type (95%), and CTX-M-14 (38%), CTX-M-15 (26%) and CTX-M-27 (19%) were the most prevalent ESBL types. The worldwide O25b-ST131-B2 clone accounted for 11% of pAmpC-producers and 41% of ESBL-producers. The O25b-ST131-B2 clone was characterised by a CTX-M-27- or CTX-M-15-type ESBL and ciprofloxacin-non-susceptibility with quadruple mutations in the quinolone resistance-determining regions (S83L and D87N in GyrA and S80I and E84V in ParC). A significant proportion of pAmpC-producers and the O25b-ST131-B2 clone were found in Japan by a recent regional surveillance programme

Economic costs for outpatient treatment of eating disorders in Japan

Abstract Background Few studies have examined the economic costs of outpatient care for eating disorders in Japan. This study aimed to clarify the reimbursement for outpatient treatment of eating disorders and compare the costs between the departments of Psychosomatic Medicine and Psychiatry in Japan. Method A multicenter, prospective, observational study of patients with an eating disorder was conducted in the Psychosomatic Medicine departments of three centers and the Psychiatry departments of another three centers in Japan. We analyzed medical reimbursement for an outpatient revisit, time of clinical interviews, and the treatment outcome measured by the Eating Disorder Examination Questionnaire (EDE-Q) global scores and body mass index (BMI) at 3 months. Multivariate linear regression models were performed to adjust for covariates. Results This study included 188 patients in the Psychosomatic Medicine departments and 68 in the Psychiatry departments. The average reimbursement cost for an outpatient revisit was 4670 yen. Even after controlling for covariates, the Psychosomatic Medicine departments had lower reimbursement points per minute of interviews than the Psychiatry departments (coefficient = − 23.86; 95% confidence interval = − 32.09 to − 15.63; P < 0.001). In contrast, EDE-Q global scores and BMI at 3 months were not significantly different between these departments. Conclusions This study clarifies the economic costs of treating outpatients with eating disorders in Japan. The medical reimbursement points per interview minute were lower in Psychosomatic Medicine departments than in Psychiatry departments, while there were no apparent differences in the treatment outcomes. Addressing this issue is necessary to provide an adequate healthcare system for patients with eating disorders in Japan

Augmentation of antitumor immunity by fusions of ethanol-treated tumor cells and dendritic cells stimulated via dual TLRs through TGF-β1 blockade and IL-12p70 production.

The therapeutic efficacy of fusion cell (FC)-based cancer vaccine generated with whole tumor cells and dendritic cells (DCs) requires the improved immunogenicity of both cells. Treatment of whole tumor cells with ethanol resulted in blockade of immune-suppressive soluble factors such as transforming growth factor (TGF)-β1, vascular endothelial growth factor, and IL-10 without decreased expression of major histocompatibility complex (MHC) class I and the MUC1 tumor-associated antigen. Moreover, the ethanol-treated tumor cells expressed "eat-me" signals such as calreticulin (CRT) on the cell surface and released immunostimulatory factors such as heat shock protein (HSP)90α and high-mobility group box 1 (HMGB1). A dual stimulation of protein-bound polysaccharides isolated from Coriolus versicolor (TLR2 agonist) and penicillin-inactivated Streptococcus pyogenes (TLR4 agonist) led human monocyte-derived DCs to produce HSP90α and multiple cytokines such as IL-12p70 and IL-10. Interestingly, incorporating ethanol-treated tumor cells and TLRs-stimulated DCs during the fusion process promoted fusion efficiency and up-regulated MHC class II molecules on a per fusion basis. Moreover, fusions of ethanol-treated tumor cells and dual TLRs-stimulated DCs (E-tumor/FCs) inhibited the production of multiple immune-suppressive soluble factors including TGF-β1 and up-regulated the production of IL-12p70 and HSP90α. Most importantly, E-tumor/FCs activated T cells capable of producing high levels of IFN-γ, resulting in augmented MUC1-specific CTL induction. Collectively, our results illustrate the synergy between ethanol-treated whole tumor cells and dual TLRs-stimulated DCs in inducing augmented CTL responses in vitro by FC preparations. The alternative system is simple and may provide a platform for adoptive immunotherapy

Combined TLR2/4-Activated Dendritic/Tumor Cell Fusions Induce Augmented Cytotoxic T Lymphocytes

<div><p>Induction of antitumor immunity by dendritic cell (DC)-tumor fusion cells (DC/tumor) can be modulated by their activation status. In this study, to address optimal status of DC/tumor to induce efficient antigen-specific cytotoxic T lymphocytes (CTLs), we have created various types of DC/tumor: 1) un-activated DC/tumor; 2) penicillin-killed <i>Streptococcus pyogenes</i> (OK-432; TLR4 agonist)-activated DC/tumor; 3) protein-bound polysaccharides isolated from <i>Coriolus versicolor</i> (PSK; TLR2 agonist)-activated DC/tumor; and 4) Combined OK-432- and PSK-activated DC/tumor. Moreover, we assessed the effects of TGF-β1 derived from DC/tumor on the induction of MUC1-specific CTLs. Combined TLR2- and TLR4-activated DC/tumor overcame immune-suppressive effect of TGF-β1 in comparison to those single activated or un-activated DC/tumor as demonstrated by: 1) up-regulation of MHC class II and CD86 expression on DC/tumor; 2) increased fusion efficiency; 3) increased production of fusions derived IL-12p70; 4) activation of CD4⁺ and CD8⁺ T cells that produce high levels of IFN-γ; 5) augmented induction of CTL activity specific for MUC1; and 6) superior efficacy in inhibiting CD4⁺CD25⁺Foxp3⁺ T cell generation. However, DC/tumor-derived TGF-β1 reduced the efficacy of DC/tumor vaccine <i>in vitro</i>. Incorporating combined TLRs-activation and TGF-β1-blockade of DC/tumor may enhance the effectiveness of DC/tumor-based cancer vaccines and have the potential applicability to the field of adoptive immunotherapy.</p> </div

Phenotypic and functional characterization of DCs.

<p>(A) Imm-DCs, OK-DCs, PSK-DCs, and OPK-DCs were analyzed by flow cytometry for expression of the indicated antigens (n = 5). Unfilled histogram profile indicates the isotype control, and solid histogram indicates the specific antibody. (B) MFI of the indicated molecules expressed by DCs (Imm-DCs, OK-DCs, PSK-DCs, and OPK-DCs) was analyzed (n = 5). (C) Production of IL-12p70 or HSP90α by DCs (Imm-DCs, OK-DCs, PSK-DCs, and OPK-DCs at 1×10⁵/mL) (n = 5) was analyzed by ELISA. The results are expressed as the mean ± SD. ***<i>P</i><0.001; **<i>P</i><0.01; *<i>P</i><0.05.</p

Phenotypic characterization of DC/tumor.

<p>(A) Eight types of DC/tumor (Imm/Mock, OK/Mock, PSK/Mock, OPK/Mock, Imm/TGF-β, OK/TGF-β, PSK/TGF-β, and OPK/TGF-β) (n = 4) were stained with FITC-conjugated mAbs against MUC1 and PE-conjugated mAbs against HLA-DR or CD86 and analyzed by two-color flow cytometry. The numbers of events in each region are shown. (B) Percentage of cells positive for both MUC1 and HLA-DR or CD86 in eight types of DC/tumor (Imm/Mock, OK/Mock, PSK/Mock, OPK/Mock, Imm/TGF-β, OK/TGF-β, PSK/TGF-β, and OPK/TGF-β) (n = 4) were analyzed. (C) Eight types of DC/tumor (Imm/Mock, OK/Mock, PSK/Mock, OPK/Mock, Imm/TGF-β, OK/TGF-β, PSK/TGF-β, and OPK/TGF-β) (n = 4) were stained with FITC-conjugated mAbs against MUC1 and PE-conjugated mAbs against HLA-DR or CD86 and analyzed by FACS, where the fused cells were identified as MUC1⁺HLA-DR⁺ or MUC1⁺CD86⁺. MFI of MUC1⁺HLA-DR⁺ or MUC1⁺CD86⁺ populations was analyzed. The results are expressed as the mean ± SD. ***<i>P</i><0.001; **<i>P</i><0.01; *<i>P</i><0.05.</p

Induction of MUC1-specific CTL responses against PANC-1 cells by DC/tumor.

<p>(A) T cells (n = 3) were stimulated with the same number of fused cells that coexpressed both MUC1 and HLA-DR in eight types of DC/tumor in the same set of experiments. Stimulated T cells were incubated with PKH-26-labeled PANC-1 cells at a ratio of 80∶1 for cytotoxicity assays. (B) T cells (n = 3) stimulated by OPK/Mock or OPK/TGF-β were cocultured with PKH-26-labeled PANC-1 cells at the indicated E:T ratios. Percentage of cytotoxicity (mean ± SD) was determined by flow cytometry CTL assays. (C) T cells (HLA-A2⁺) (n = 5) stimulated by eight types of DC/tumor (HLA-A2⁺) (Imm/Mock, OK/Mock, PSK/Mock, OPK/Mock, Imm/TGF-β, OK/TGF-β, PSK/TGF-β, and OPK/TGF-β) were stained with FITC-conjugated mAbs against CD8 and PE-conjugated pentamer against HLA-A2/MUC1. The numbers of events in gated CD8⁺ T cells are shown. (D) The percentage of CD8⁺ T cells reactive to MUC1 among the whole CD8⁺ T cell population (n = 5) was shown as a percentage of the double-positive population (pentamer⁺CD8⁺) in the total CD8⁺ T cells. The results are expressed as the mean ± SD. ***<i>P</i><0.001; **<i>P</i><0.01; *<i>P</i><0.05.</p