Transcriptome Modifications in Porcine Adipocytes via Toll-Like Receptors Activation

Adipocytes are the most important cell type in adipose tissue playing key roles in immunometabolism. We previously reported that nine members of the Toll-like receptor (TLR) family are expressed in an originally established porcine intramuscular pre-adipocyte (PPI) cell line. However, the ability of TLR ligands to modulate immunometabolic transcriptome modifications in porcine adipocytes has not been elucidated. Herein, we characterized the global transcriptome modifications in porcine intramuscular mature adipocytes (pMA), differentiated from PPI, following stimulation with Pam3csk4, Poly(I:C) or LPS which are ligands for TLR2, TLR3, and TLR4, respectively. Analysis of microarray data identified 530 (218 up, 312 down), 520 (245 up, 275 down), and 525 (239 up, 286 down) differentially expressed genes (DEGs) in pMA following the stimulation with Pam3csk4, Poly(I:C), and LPS, respectively. Gene ontology classification revealed that DEGs are involved in several biological processes including those belonging to immune response and lipid metabolism pathways. Functionally annotated genes were organized into two groups for downstream analysis: immune response related genes (cytokines, chemokines, complement factors, adhesion molecules, and signal transduction), and genes involved with metabolic and endocrine functions (hormones and receptors, growth factors, and lipid biosynthesis). Differential expression analysis revealed that EGR1, NOTCH1, NOS2, TNFAIP3, TRAF3IP1, INSR, CXCR4, PPARA, MAPK10, and C3 are the top 10 commonly altered genes of TLRs induced transcriptional modification of pMA. However, the protein-protein interaction network of DEGs identified EPOR, C3, STAR, CCL2, and SAA2 as the major hub genes, which were also exhibited higher centrality estimates in the Gene-Transcription factor interaction network. Our results provide new insights of transcriptome modifications associated with TLRs activation in porcine adipocytes and identified key regulatory genes that could be used as biomarkers for the evaluation of treatments having immunomodularoty and/or metabolic functional beneficial effects in porcine adipocytes.Fil: Igata, Manami. Tohoku University; JapónFil: Islam, M. Aminul. Tohoku University; Japón. Bangladesh Agricultural University; BangladeshFil: Tada, Asuka. Tohoku University; JapónFil: Takagi, Michihiro. Tohoku University; JapónFil: Humayun Kober, AKM. Tohoku University; Japón. Chittagong Veterinary and Animal Sciences University; BangladeshFil: Albarracín, Leonardo Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; Japón. Universidad Nacional de Tucumán. Facultad de Ciencias Exactas y Tecnología. Departamento de Ciencias de la Computación; ArgentinaFil: Aso, Hisashi. Tohoku University; JapónFil: Ikeda-Ohtsubo, Wakako. Tohoku University; JapónFil: Miyazawa, Kenji. Takanashi Milk Products Co.; JapónFil: Yoda, Kazutoyo. Takanashi Milk Products Co.; JapónFil: He, Fang. Takanashi Milk Products Co.; JapónFil: Takahashi, Hideki. Tohoku University; JapónFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; JapónFil: Kitazawa, Haruki. Tohoku University; Japó

Design and synthesis of a new chromophore, 2-(4-nitrophenyl)benzofuran, for two-photon uncaging using near-IR light

C. K. acknowledges the HPC resources of CINES and of IDRIS under the allocations 2014-[x2014080649] made by GENCI (Grand Equipement National de Calcul Intensif).International audienceA new chromophore, 2-(4-nitrophenyl)benzofuran (NPBF), was designed for two-photon (TP) uncaging using near-IR light. The TP absorption (TPA) cross-sections of the newly designed NPBF chromophore were determined to be 18 GM at 720 nm and 54 GM at 740 nm in DMSO. The TP uncaging reaction of a caged benzoate with the NPBF chromophore quantitatively produced benzoic acid with an efficiency (δu) of ∼5.0 GM at 740 nm. The TP fragmentation of an EGTA unit was observed with δu = 16 GM. This behavior makes the new chromophore a promising TP photoremovable protecting group for physiological studies

Immunobiotic Lactobacillus jensenii TL2937 Alleviates Dextran Sodium Sulfate-Induced Colitis by Differentially Modulating the Transcriptomic Response of Intestinal Epithelial Cells

Immunobiotics have emerged as a promising intervention to alleviate intestinal damage in inflammatory bowel disease (IBD). However, the beneficial properties of immunobiotics are strain dependent and, therefore, each strain has to be evaluated in order to demonstrate its potential application in IBD. Our previous in vitro and in vivo studies demonstrated that Lactobacillus jensenii TL2937 attenuates gut acute inflammatory response triggered by Toll-like receptor 4 activation. However, its effect on colitis has not been evaluated before. In this work, we studied whether the TL2937 strain was able to protect against the development of colitis in a dextran sodium sulfate (DSS)-induced mouse model and we delved into the mechanisms of action by evaluating the effect of the immunobiotic bacteria on the transcriptomic response of DSS-challenged intestinal epithelial cells. L. jensenii TL2937 was administered to adult BALB/c mice before the induction of colitis by the administration of DSS. Colitis and the associated inflammatory response were evaluated for 14 days. Mice fed with L. jensenii TL2937 had lower disease activity index and alterations of colon length when compared to control mice. Reduced myeloperoxidase activity, lower production of pro-inflammatory (TNF-α, IL-1, CXCL1, MCP-1, IL-15, and IL-17), and higher levels of immunoregulatory (IL-10 and IL-27) cytokines were found in the colon of TL2937-treated mice. In addition, the treatment of porcine intestinal epithelial (PIE) cells with L. jensenii TL2937 before the challenge with DSS differentially regulated the activation of the JNK pathway, leading to an increase in epithelial cell integrity and to a differential immunotranscriptomic response. TL2937-treated PIE cells had a significant reduction in the expression of inflammatory cytokines (TNF-α, IL-1α, IL-1β, IL-6, IL-15), chemokines (CCL2, CCL4, CCL8, CXCL4, CXCL5, CXCL9, CXCL10), adhesion molecules (SELE, SELL, EPCAM), and other immune factors (NCF1, NCF2, NOS2, SAA2) when compared to control cells after the challenge with DSS. The findings of this work indicate that (a) L. jensenii TL2937 is able to alleviate DSS-induced colitis suggesting a potential novel application for this immunobiotic strain, (b) the modulation of the transcriptomic response of intestinal epithelial cells would play a key role in the beneficial effects of the TL2937 strain on colitis, and (c) the in vitro PIE cell immunoassay system could be of value for the screening and selection of new immunobiotic strains for their application in IBD.Fil: Sato, Nana. Tohoku University; JapónFil: García Castillo, Valeria. Tohoku University; Japón. Universidad de Concepción; ChileFil: Yuzawa, Mao. Tohoku University; JapónFil: Islam, Md Aminul. Tohoku University; Japón. Bangladesh Agricultural University; BangladeshFil: Albarracín, Leonardo Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Universidad Nacional de Tucumán. Facultad de Ciencias Exactas y Tecnología; Argentina. Tohoku University; JapónFil: Tomokiyo, Mikado. Tohoku University; JapónFil: Ikeda Ohtsubo, Wakako. Tohoku University; JapónFil: García Cancino, Apolinaria. Universidad de Concepción; ChileFil: Takahashi, Hideki. Tohoku University; JapónFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; JapónFil: Kitazawa, Haruki. Tohoku University; Japó

Attitude of young psychiatrists toward coercive measures in psychiatry: a case vignette study in Japan

<p>Abstract</p> <p>Background</p> <p>Every psychiatrist must pay careful attention to avoid violating human rights when initiating coercive treatments such as seclusion and restraint. However, these interventions are indispensable in clinical psychiatry, and they are often used as strategies to treat agitated patients. In this study, we investigated young psychiatrists' attitudes toward psychiatric coercive measures.</p> <p>Methods</p> <p>A total of 183 young psychiatrists participated as subjects in our study. A questionnaire with a case vignette describing a patient with acute psychosis was sent to the study subjects via the Internet or by mail. This questionnaire included scoring the necessity for hospitalization, and the likelihood of prescribing seclusion and/or restraint, on a 9-point Likert scale (with 9 indicating strong agreement).</p> <p>Results</p> <p>There was general agreement among the study subjects that the case should be admitted to a hospital (8.91 ± 0.3) and secluded (8.43 ± 1.0). The estimated length of hospitalization was 13.53 ± 6.4 weeks. Regarding the likelihood of prescribing restraint, results showed great diversity (5.14 ± 2.5 on 9-point scale); psychiatrists working at general hospitals scored significantly higher (6.25 ± 2.5) than those working at university hospitals (5.02 ± 2.3) or psychiatric hospitals (4.15 ± 2.6). A two-group comparison of the length of inpatient care revealed a significant difference between those psychiatrists who scored 1-3 (n = 55, 14.22 ± 7.4 wks) and those who scored 7-9 (n = 62, 12.22 ± 4.0) regarding the need to use restraint.</p> <p>Conclusion</p> <p>Our results may reflect the current dilemma in Japanese psychiatry wherein psychiatrists must initiate coercive measures to shorten hospitalization stays. This study prompted its subject psychiatrists to consider coercive psychiatric treatments.</p

Exopolysaccharides From Streptococcus thermophilus ST538 Modulate the Antiviral Innate Immune Response in Porcine Intestinal Epitheliocytes

It was reported that exopolysaccharides (EPSs) from lactobacilli are able to differentially modulate mucosal antiviral immunity. Although research has described the ability of EPSs derived from Streptococcus thermophilus to modulate the mucosal immune system, their impact on antiviral immunity was less explored. In this work, we investigated the capacity of the EPS-producing S. thermophilus ST538 to modulate the innate antiviral immune response triggered by the activation of the Toll-like receptor 3 (TLR3) in porcine intestinal epitheliocytes (PIE cells). Moreover, in order to study the immunomodulatory potential of S. thermophilus ST538 EPS, we successfully developed two mutant strains through the knockout of the epsB or epsC genes. High-performance liquid chromatography and scanning electron microscopy studies demonstrated that the wild type (WT) strain produced as high as 595 μg/ml of EPS in the skim milk medium, while none of the mutant strains (S. thermophilus ΔepsB and ΔepsC) were able to produce EPS. Studies in PIE cells demonstrated that the EPS of S. thermophilus ST538 is able to significantly improve the expression of interferon β (IFN-β), interleukin 6 (IL-6), and C-X-C motif chemokine 10 (CXCL10) in response to TLR3 stimulation. The role of EPS in the modulation of antiviral immune response in PIE cells was confirmed by comparative studies of cell free culture supernatants and fermented skim milks obtained from S. thermophilus ΔepsB and ΔepsC. These results suggest that S. thermophilus ST538 could be used as an immunobiotic strain for the development of new immunologically functional foods, which might contribute to improve resistance against viral infections.Fil: Mizuno, Akira. Tohoku University; JapónFil: Tomotsune, Kae. Tohoku University; JapónFil: Islam, Md Aminul. Tohoku University; JapónFil: Funabashi, Ryutaro. Tohoku University; JapónFil: Albarracín, Leonardo Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; Japón. Universidad Nacional de Tucumán. Facultad de Ciencias Exactas y Tecnología; ArgentinaFil: Ikeda Ohtsubo, Wakako. Tohoku University; JapónFil: Aso, Hisashi. Tohoku University; JapónFil: Takahashi, Hideki. Tohoku University; JapónFil: Kimura, Katsunori. Meiji Co.; JapónFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; JapónFil: Sasaki, Yasuko. Meiji University; JapónFil: Kitazawa, Haruki. Tohoku University; Japó

Evaluation of fat accumulation and adipokine production during the long-term adipogenic differentiation of porcine intramuscular preadipocytes and study of the influence of immunobiotics

The degree of fat accumulation and adipokine production are two major indicators of obesity that are correlated with increased adipose tissue mass and chronic inflammatory responses. Adipocytes have been considered effector cells for the inflammatory responses due to their capacity to express Toll-like receptors (TLRs). In this study, we evaluated the degree of fat accumulation and adipokine production in porcine intramuscular preadipocyte (PIP) cells maintained for in vitro differentiation over a long period without or with stimulation of either TNF-α or TLR2-, TLR3-, or TLR4-ligands. The cytosolic fat accumulation was measured by liquid chromatography and the expression of adipokines (CCL2, IL-6, IL-8 and IL-10) were quantified by RT-qPCR and ELISA at several time points (0 to 20 days) of PIP cells differentiation. Long-term adipogenic differentiation (LTAD) induced a progressive fat accumulation in the adipocytes over time. Activation of TLR3 and TLR4 resulted in an increased rate of fat accumulation into the adipocytes over the LTAD. The production of CCL2, IL-8 and IL-6 were significantly increased in unstimulated adipocytes during the LTAD, while IL-10 expression remained stable over the studied period. An increasing trend of adiponectin and leptin production was also observed during the LTAD. On the other hand, the stimulation of adipocytes with TLRs agonists or TNF-α resulted in an increasing trend of CCL2, IL-6 and IL-8 production while IL-10 remained stable in all four treatments during the LTAD. We also examined the influences of several immunoregulatory probiotic strains (immunobiotics) on the modulation of the fat accumulation and adipokine production using supernatants of immunobiotic-treated intestinal immune cells and the LTAD of PIP cells. Immunobiotics have shown a strain-specific ability to modulate the fat accumulation and adipokine production, and differentiation of adipocytes. Here, we expanded the utility and potential application of our in vitro PIP cells model by evaluating an LTAD period (20 days) in order to elucidate further insights of chronic inflammatory pathobiology of adipocytes associated with obesity as well as to explore the prospects of immunomodulatory intervention for obesity such as immunobiotics.Fil: Tada, Asuka. Tohoku University; JapónFil: Kober, Akm Humayun. Chittagong Veterinary and Animal Sciences University; Bangladesh. Tohoku University; JapónFil: Islam, Md Aminul. Bangladesh Agricultural University; Bangladesh. Tohoku University; JapónFil: Igata, Manami. Tohoku University; JapónFil: Takagi, Michihiro. Tohoku University; JapónFil: Suzuki, Masahiko. Tohoku University; JapónFil: Aso, Hisashi. Tohoku University; JapónFil: Ikeda Ohtsubo, Wakako. Tohoku University; JapónFil: Yoda, Kazutoyo. Takanashi Milk Products Co., Ltd.; JapónFil: Miyazawa, Kenji. Takanashi Milk Products Co., Ltd.; JapónFil: He, Fang. Takanashi Milk Products Co., Ltd.; JapónFil: Takahashi, Hideki. Tohoku University; JapónFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; JapónFil: Kitazawa, Haruki. Tohoku University; Japó

Transcriptome Modifications in the Porcine Intramuscular Adipocytes during Differentiation and Exogenous Stimulation with TNF-α and Serotonin

Adipocytes are dynamic cells that have critical functions to maintain body energy homeostasis. Adipocyte physiology is affected by the adipogenic differentiation, cell program, as well as by the exogenous stimulation of biochemical factors, such as serotonin and TNF-α. In this work, we investigated the global transcriptome modifications when porcine intramuscular preadipocyte (PIP) was differentiated into porcine mature adipocyte (pMA). Moreover, we studied transcriptome changes in pMA after stimulation with serotonin or TNF-α by using a microarray approach. Transcriptome analysis revealed that the expression of 270, 261, and 249 genes were modified after differentiation, or after serotonin and TNF-α stimulation, respectively. Expression changes in APP, HNF4A, ESR1, EGR1, SRC, HNF1A, FN1, ALB, STAT3, CBL, CEBPB, AR, FOS, CFTR, PAN2, PTPN6, VDR, PPARG, STAT5A and NCOA3 genes which are enriched in the ‘PPAR signaling’ and ‘insulin resistance’ pathways were found in adipocytes during the differentiation process. Dose-dependent serotonin stimulation resulted in a decreased fat accumulation in pMAs. Serotonin-induced differentially expressed genes in pMAs were found to be involved in the significant enrichment of ′GPCR ligand-binding′, ‘cell chemotaxis’, ‘blood coagulation and complement’, ‘metabolism of lipid and lipoproteins’, ‘regulation of lipid metabolism by PPARA’, and ‘lipid digestion, mobilization and transport’ pathways. TNF-α stimulation also resulted in transcriptome modifications linked with proinflammatory responses in the pMA of intramuscular origin. Our results provide a landscape of transcriptome modifications and their linked-biological pathways in response to adipogenesis, and exogenous stimulation of serotonin- and TNF-α to the pMA of intramuscular origin.Fil: Tada, Asuka. Tohoku University; JapónFil: Islam, Md Aminul. Tohoku University; Japón. Bangladesh Agricultural University; BangladeshFil: Kober, Akm Humayun. Tohoku University; JapónFil: Fukuyama, Kohtaro. Tohoku University; JapónFil: Takagi, Michihiro. Tohoku University; JapónFil: Igata, Manami. Tohoku University; JapónFil: Albarracín, Leonardo Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; JapónFil: Ikeda Ohtsubo, Wakako. Tohoku University; JapónFil: Miyazawa, Kenji. Takanashi Milk Products Co.; JapónFil: Yoda, Kazutoyo. Takanashi Milk Products Co.; JapónFil: He, Fang. Takanashi Milk Products Co.; JapónFil: Takahashi, Hideki. Tohoku University; JapónFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; JapónFil: Aso, Hisashi. Tohoku University; JapónFil: Kitazawa, Haruki. Tohoku University; Japó

Selection of immunobiotic ligilactobacillus salivarius strains from the intestinal tract of wakame-fed pigs: Functional and genomic studies

In this article, Ligilactobacillus salivarius FFIG strains, isolated from the intestinal tract of wakame-fed pigs, are characterized according to their potential probiotic properties. Strains were evaluated by studying their interaction with porcine intestinal epithelial (PIE) cells in terms of their ability to regulate toll-like receptor (TLR)-3- or TLR4-mediated innate immune responses, as well as by assessing their adhesion capabilities to porcine epithelial cells and mucins. These functional studies were complemented with comparative genomic evaluations using the complete genome sequences of porcine L. salivarius strains selected from subgroups that demonstrated different ‘immune’ and ‘adhesion’ phenotypes. We found that their immunomodulatory and adhesion capabilities are a strain-dependent characteristic. Our analysis indicated that the differential immunomodulatory and adhesive activities of FFIG strains would be dependent on the combination of several surface structures acting simultaneously, which include peptidoglycan, exopolysaccharides, lipoteichoic acid, and adhesins. Of note, our results indicate that there is no correlation between the immunomodulatory capacity of the strains with their adhesion ability to mucins and epithelial cells. Therefore, in the selection of strains destined to colonize the intestinal mucosa and modulate the immunity of the host, both properties must be adequately evaluated. Interestingly, we showed that L. salivarius FFIG58 functionally modulated the innate immune responses triggered by TLR3 and TLR4 activation in PIE cells and efficiently adhered to these cells. Moreover, the FFIG58 strain was capable of reducing rotavirus replication in PIE cells. Therefore, L. salivarius FFIG58 is a good candidate for further in vivo studying the protective effect of lactobacilli against intestinal infections in the porcine host. We also reported and analyzed, for the first time, the complete genome of several L. salivarius strains that were isolated from the intestine of pigs after the selective pressure of feeding the animals with wakame. Further genomic analysis could be of value to reveal the metabolic characteristics and potential of the FFIG strains in general and of the FFIG58 strain, in particular, relating to wakame by-products assimilation.Fil: Zhou, Binghui. Tohoku University; JapónFil: Albarracín, Leonardo Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Universidad Nacional de Tucumán. Facultad de Ciencias Exactas y Tecnología. Departamento de Ciencias de la Computación; Argentina. Tohoku University; JapónFil: Indo, Yuhki. Tohoku University; JapónFil: Arce, Lorena Paola. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto Superior de Investigaciones Biológicas. Universidad Nacional de Tucumán. Instituto Superior de Investigaciones Biológicas; Argentina. Tohoku University; JapónFil: Masumizu, Yuki. Tohoku University; JapónFil: Tomokiyo, Mikado. Tohoku University; JapónFil: Islam, Md Aminul. Bangladesh Agricultural University; Bangladesh. Tohoku University; JapónFil: Garcia Castillo, Valeria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; JapónFil: Ikeda Ohtsubo, Wakako. Tohoku University; JapónFil: Nochi, Tomonori. Tohoku University; JapónFil: Morita, Hidetoshi. Okayama University; JapónFil: Takahashi, Hideki. Tohoku University; JapónFil: Kurata, Shoichiro. Tohoku University; JapónFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; JapónFil: Kitazawa, Haruki. Tohoku University; Japó

Suicidal ideation and burnout among psychiatric trainees in Japan

AIM: Burnout is a psychological condition that may occur in all workers after being exposed to excessive work-related stresses. We investigated suicidal ideation and burnout among Japanese psychiatric trainees as a part of the Burnout Syndrome Study (BoSS) International.  METHODS: In the Japanese branch, 91 trainees fully completed suicide ideation and behaviour questionnaire (SIBQ) and Maslach Burnout Inventory-General Survey (MBI-GS).  RESULTS: Passive suicidal ideation was reported by 38.5% of Japanese trainees and 22.0% of them had experienced active suicidal ideation. The burnout rate among Japanese subjects was 40.0%. These results were worse compared to the all 1980 trainees who fully completed the main outcome measure in BoSS International, 25.9%, 20.4% and 36.7%, respectively.  CONCLUSIONS: Our results suggest a higher risk of suicide among Japanese residents. Japan has a higher suicide rate than other countries. Early detection of, and appropriate intervention for, suicidal ideation is important in preventing suicide in psychiatry residents