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Abstract Although avian influenza A viruses (avian IAVs) bind preferentially to terminal sialic acids (Sia) on glycans that possess Siaα2-3Gal, the actual glycan structures found in chicken respiratory tracts have not been reported. Herein, we analyzed N-glycan structures in chicken trachea and lung, the main target tissues of low pathogenic avian IAVs. 2-Aminopyridine (PA)-labeled N-glycans from chicken tissues were analyzed by combined methods using reversed-phase liquid chromatography (LC), electrospray ionization (ESI)-mass spectrometry (MS), MS/MS, and multistage MS (MSn), with or without modifications using exoglycosidases, sialic acid linkage-specific alkylamidation (SALSA), and/or permethylation. The results of SALSA indicated that PA-N-glycans in both chicken trachea and lung harbored slightly more α2,6-Sia than α2,3-Sia. Most α2,3-Sia on N-glycans in chicken trachea was a fucosylated form (sialyl Lewis X, sLex), whereas no sLex was detected in lung. By contrast, small amounts of N-glycans with 6-sulfo sialyl LacNAc were detected in lung but not in trachea. Considering previous reports that hemagglutinins (HAs) of avian IAVs originally isolated from chicken bind preferentially to α2,3-Sia with or without fucosylation and/or 6-sulfation but not to α2,6-Sia, our results imply that avian IAVs do not evolve to possess HAs that bind preferentially to α2,6-Sia, regardless of the abundance of α2,6-Sia.

Nonalcoholic steatohepatitis (NASH) is often associated with obesity, but some patients develop NASH without obesity. The physiological processes by which nonobese patients develop NASH and cirrhosis have not yet been determined. Here, we analyzed the effects of dietary methionine content on NASH induced in mice fed on a choline‐deficient, methionine‐lowered, l‐amino acid‐defined high‐fat diet (CDAHFD). CDAHFD with insufficient methionine induced insulin sensitivity and enhanced NASH pathology, but without obesity. In contrast, CDAHFD with sufficient methionine induced steatosis, and unlike CDAHFD with insufficient methionine, also induced obesity and insulin resistance. Gene profile analysis revealed that the disease severity in CDAHFD may partially be due to upregulation of the Rho family GTPases pathway and mitochondrial and nuclear receptor signal dysfunction. The signaling factors/pathways detected in this study may assist in future study of NASH regulation, especially its ‘nonobese’ subtype.

Abstract Agonistic profiles of AMPA receptor (AMPA-R) potentiators may be associated with seizure risk and bell-shaped dose-response effects. Here, we report the pharmacological characteristics of a novel AMPA-R potentiator, TAK-653, which exhibits minimal agonistic properties. TAK-653 bound to the ligand binding domain of recombinant AMPA-R in a glutamate-dependent manner. TAK-653 strictly potentiated a glutamate-induced Ca2+ influx in hGluA1i-expressing CHO cells through structural interference at Ser743 in GluA1. In primary neurons, TAK-653 augmented AMPA-induced Ca2+ influx and AMPA-elicited currents via physiological AMPA-R with little agonistic effects. Interestingly, TAK-653 enhanced electrically evoked AMPA-R-mediated EPSPs more potently than AMPA (agonist) or LY451646 (AMPA-R potentiator with a prominent agonistic effect) in brain slices. Moreover, TAK-653 improved cognition for both working memory and recognition memory, while LY451646 did so only for recognition memory, and AMPA did not improve either. These data suggest that the facilitation of phasic AMPA-R activation by physiologically-released glutamate is the key to enhancing synaptic and cognitive functions, and nonselective activation of resting AMPA-Rs may negatively affect this process. Importantly, TAK-653 had a wide safety margin against convulsion; TAK-653 showed a 419-fold (plasma Cmax) and 1017-fold (AUC plasma) margin in rats. These findings provide insight into a therapeutically important aspect of AMPA-R potentiation.

Abstract Glycans in tissues are structurally diverse and usually include a large number of isomers that cannot be easily distinguished by mass spectrometry (MS). To address this issue, we developed a combined method that can efficiently separate and identify glycan isomers. First, we separated 2-aminopyridine (PA)-derivatized N-glycans from chicken colon by reversed-phase liquid chromatography (LC) and directly analyzed them by electrospray ionization (ESI)-MS and MS/MS to obtain an overview of the structural features of tissue glycans. Next, we deduced the structures of isomers based on their elution positions, full MS, and MS/MS data, before or after digestions with several exoglycosidases. In this method, the elution position differed greatly depending on the core structure and branching pattern, allowing multiantennary N-glycan structures to be easily distinguished. To further determine linkages of branch sequences, we modified PA-N-glycans with sialic acid linkage-specific alkylamidation and/or permethylation, and analyzed the products by LC–MS and multistage MS. We determined the relative abundances of core structures, branching patterns, and branch sequences of N-glycans from chicken colon, and confirmed presence of characteristic branch sequences such as Lex, sialyl Lex, sulfated LacNAc, LacNAc repeat, and LacdiNAc. The results demonstrated that our method is useful for comparing N-glycomes among various tissue samples.

Abstract Background Nonalcoholic steatohepatitis (NASH) is a form of liver disease characterized by steatosis, necroinflammation, and fibrosis, resulting in cirrhosis and cancer. Efforts have focused on reducing the intake of trans fatty acids (TFAs) because of potential hazards to human health and the increased risk for NASH. However, the health benefits of reducing dietary TFAs have not been fully elucidated. Here, the effects of TFAs vs. a substitute on NASH induced in mice by feeding a choline-deficient, methionine-lowered, L-amino acid-defined, high-fat diet (CDAA-HF) were investigated. Methods Mice were fed CDAA-HF containing shortening with TFAs (CDAA-HF-T(+)), CDAA-HF containing shortening without TFAs (CDAA-HF-T(−)), or a control chow for 13 or 26 weeks. Results At week 13, NASH was induced in mice by feeding CDAA-HF-T(+) containing TFAs or CDAA-HF-T(−) containing no TFAs, but rather mostly saturated fatty acids (FAs), as evidenced by elevated serum transaminase activity and liver changes, including steatosis, inflammation, and fibrosis. CDAA-HF-T(−) induced a greater extent of hepatocellular apoptosis at week 13. At week 26, proliferative (preneoplastic and non-neoplastic) nodular lesions were more pronounced in mice fed CDAA-HF-T(−) than CDAA-HF-T(+). Conclusions Replacement of dietary TFAs with a substitute promoted the development of proliferation lesions in the liver of a mouse NASH model, at least under the present conditions. Attention should be paid regarding use of TFA substitutes in foods for human consumption, and a balance of FAs is likely more important than the particular types of FAs.

Background and aims: Endoscopic submucosal dissection (ESD) is the preferred technique for en bloc resection of superficial colorectal neoplasms. Resection of extensive lesions with ESD can be challenging, owing to loss of orientation in the submucosal space. In this case series, we describe the double-tunneling (DoT) butterfly method for ESD of extensive rectal neoplasms. Methods: The key feature of the DoT butterfly method is the creation of 2 tunnels that are transformed into bilateral flaps, leaving a submucosal septum between them. Results: Four rectal neoplasms measuring (maximum diameter) 7 cm, 8 cm, 9 cm, and 18 cm, respectively, were resected in 4 patients by use of the DoT butterfly method. The lesions included recurrent adenoma (n = 1) and dysplasia (n = 1) in longstanding ulcerative colitis. Curative R0 resection was confirmed in all 4 cases. Histologic examination showed tubular adenomas with low-grade dysplasia in 1 of 4 patients and focal high-grade dysplasia in 3 of 4 patients. One patient experienced postprocedural bleeding that required endoscopic reintervention. Conclusion: The DoT butterfly method appears to be useful for the resection of extensive rectal neoplasms. A prospective study is required to assess whether these results can be reproduced in a large cohort of patients.

We recently reported increased levels of urinary free-glycans in some cancer patients. Here, we focused on cancer related alterations in the levels of high molecular weight free-glycans. The rationale for this study was that branching, elongation, fucosylation and sialylation, which lead to increases in the molecular weight of glycans, are known to be up-regulated in cancer. Urine samples from patients with gastric cancer, pancreatic cancer, cholangiocarcinoma and colorectal cancer and normal controls were analyzed. The extracted free-glycans were fluorescently labeled with 2-aminopyridine and analyzed by multi-step liquid chromatography. Comparison of the glycan profiles revealed increased levels of glycans in some cancer patients. Structural analysis of the glycans was carried out by performing chromatography and mass spectrometry together with enzymatic or chemical treatments. To compare glycan levels between samples with high sensitivity and selectivity, simultaneous measurements by reversed-phase liquid chromatography-selected ion monitoring of mass spectrometry were also performed. As a result, three lactose-core glycans and 78 free-N-glycans (one phosphorylated oligomannose-type, four sialylated hybrid-type and 73 bi-, tri- and tetra-antennary complex-type structures) were identified. Among them, glycans with α1,3-fucosylation ((+/- sialyl) Lewis X), triply α2,6-sialylated tri-antennary structures and/or a (Man3)GlcNAc1-core displayed elevated levels in cancer patients. However, simple α2,3-sialylation and α1,6-core-fucosylation did not appear to contribute to the observed increase in the level of glycans. Interestingly, one tri-antennary free-N-glycan that showed remarkable elevation in some cancer patients contained a unique Glcβ1-4GlcNAc-core instead of the common GlcNAc2-core at the reducing end. This study provides further insights into free-glycans as potential tumor markers and their processing pathways in cancer.

This paper investigates the effect of a hearer's attitude toward a speaker's multimodal behaviors. Twenty-one university students, nine males and twelve females, participated in this experiment as speakers. Two females, aged 22 years old, performed in the role of hearers. The experiment was carried out under two conditions regarding the hearer's attitude: polite and impolite conditions. The polite hearer nodded, produced back-channel response, and gazed at the speaker, while the impolite hearer did these actions to a lesser extent. Under each condition, the speakers talked about their own experiences to the two different hearers through a within-participants design. The speech, gaze, and body motion of speakers were recorded with a video camera. In addition, the communication skills of speakers, including expressivity, sensitivity, regulation, assertiveness, and responsiveness skills, were measured after the experiment. The speakers' silent pauses, filled pauses, gaze aversion, and representational gestures were compared between the hearers' two politeness conditions. The relationship between the multimodal behaviors and the speakers' communication skills was also analyzed. The results suggest that the speakers' silent pauses, filled pauses, and gaze aversion decreased in the polite hearer condition, while representational gestures increased. From the communication-skills test, the speakers with higher expressivity, sensitivity, and regulation skills but lower assertiveness and responsiveness skills show longer silent pauses under the impolite hearer condition. Moreover, we discuss how the implications of our findings can enhance the relationship between social robots and people.

Abstract Background Aortic stenosis (AS) is common in patients on dialysis as well as in the general population. AS leads to difficulty with dialysis therapy because of unstable conditions such as intradialytic hypotension due to low cardiac output. However, the precise morbidity rates and risk factors of AS in patients on dialysis are unknown. Moreover, there are no large-scale observational studies regarding the association between AS in patients on dialysis and mortality. Therefore, we will investigate whether morbidity of AS in patients on dialysis is associated with mortality. Methods This is a multicenter prospective cohort analysis in the Tokai region of Japan. The 75 participating centers in this study will enroll approximately 2400 patients during 12 months, with or without AS. We started enrollment in July 2017 and will follow patents until June 2023. Transthoracic echocardiography will be performed to evaluate aortic valve. Parameters used for evaluation of aortic valve are mean pressure gradient between left ventricle and ascending aorta, aortic valve area, and maximum aortic jet velocity. We will diagnose AS using the criteria based on the 2014 American Heart Association/ American College of Cardiology Guideline. We will also perform transthoracic echocardiography at 12, 24, 36, 48, and 60 months. Survival prognosis and CV events will be determined at the end of June 2019, 2020, 2021, 2022, and 2023. Development of AS will be also evaluated as new onset or annual change in AS parameters. We will classify patients based on the presence or absence of AS and the stages of AS and will compare outcomes. Study outcomes will include the following: 1) all-cause mortality rates; 2) incidence of cardiovascular (CV) events; 3) CV-related mortality rates; 4) infection-related mortality rates; 5) new onset or development of AS. Discussion We will consider the following hypotheses in this study, among others: The prevalence of AS is higher in dialysis patients; new onset and development of AS are associated with factors that are specific for dialysis, such as hyperphosphatemia, hyperparathyroidism, and medication; and outcomes in AS patients are poorer than in patients without AS at baseline. Trial registration UMIN000026756, Registered March 29 2017.

BACKGROUND:Transcatheter aortic valve implantation (TAVI) is an alternative therapy for surgically high-risk patients with severe aortic stenosis (AS). Although TAVI improves survival of patients with severe AS, the mechanism of this effect remains to be clarified. We investigated the effects of TAVI on left ventricular (LV) function and identified the predictive parameters for cardiac events after TAVI. METHODS AND RESULTS:We studied 128 patients with severe symptomatic AS who underwent TAVI. Echocardiographic assessments were performed before and after TAVI. In addition to the conventional echocardiographic parameters such as LV ejection fraction (LVEF) and LV mass index (LVMI), the LV global longitudinal strain (GLS) and early diastolic peak strain rate (SR_E) using two-dimensional speckle tracking echocardiography were also evaluated. All patients were assessed for clinical events including major adverse cardiac events and stroke according to Valve Academic Research Consortium-2 criteria. GLS, early diastolic peak velocity (e'), aortic regurgitation (AR) severity, and SR_E were significantly improved after TAVI. Thirteen patients had an event during the observational period of 591 days (median). Patients with events had higher LVMI, more severe AR, and worse GLS compared to those without events. Furthermore, receiver-operating curve analysis revealed that GLS was the strongest predictor for clinical events (p = 0.009; area under the curve, 0.73). CONCLUSION:Preoperative LV geometric deformation and dysfunction, as a consequence of the cumulative burden of pressure overload, improved after TAVI and could predict cardiac events after TAVI.

Non-alcoholic fatty liver disease (NAFLD), the hepatic manifestation of metabolic syndrome, can progress to steatohepatitis (NASH) and advanced liver damage, such as that from liver cirrhosis and cancer. Recent studies have shown the benefits of consuming n-3 polyunsaturated fatty acids (PUFAs) for the treatment of NAFLD. In the present study, we investigated and compared the effects of the major n-3 PUFAs-eicosapentaenoic acid (EPA, C20:5) and docosahexaenoic acid (DHA, C22:6)-in preventing atherogenic high-fat (AHF) diet-induced NAFLD. Mice were fed the AHF diet supplemented with or without EPA or DHA for four weeks. Both EPA and DHA reduced the pathological features of AHF diet-induced NASH pathologies such as hepatic lobular inflammation and elevated serum transaminase activity. Intriguingly, EPA had a greater hepatic triacylglycerol (TG)-reducing effect than DHA. In contrast, DHA had a greater suppressive effect than EPA on AHF diet-induced hepatic inflammation and ROS generation, but no difference in fibrosis. Both EPA and DHA could be effective for treatment of NAFLD and NASH. Meanwhile, the two major n-3 polyunsaturated fatty acids might differ in a relative contribution to pathological intermediate steps towards liver fibrosis.

Ovarian cancer remains the most lethal gynecologic cancer and new targeted molecular therapies against this miserable disease continue to be challenging. In this study, we analyzed the expressional patterns of Interleukin-6 (IL-6) and its receptor (IL-6R) expression in ovarian cancer tissues, evaluated the impact of these expressions on clinical outcomes of patients, and found that a high-level of IL-6R expression but not IL-6 expression in cancer cells is an independent prognostic factor. In in vitro analyses using ovarian cell lines, while six (RMUG-S, RMG-1, OVISE, A2780, SKOV3ip1 and OVCAR-3) of seven overexpressed IL-6R compared with a primary normal ovarian surface epithelium, only two (RMG-1, OVISE) of seven cell lines overexpressed IL-6, suggesting that IL-6/IL-6R signaling exerts in a paracrine manner in certain types of ovarian cancer cells. Ovarian cancer ascites were collected from patients, and we found that primary CD11b+CD14+ cells, which were predominantly M2-polarized macrophages, are the major source of IL-6 production in an ovarian cancer microenvironment. When CD11b+CD14+ cells were co-cultured with cancer cells, both the invasion and the proliferation of cancer cells were robustly promoted and these promotions were almost completely inhibited by pretreatment with anti-IL-6R antibody (tocilizumab). The data presented herein suggest a rationale for anti-IL-6/IL-6R therapy to suppress the peritoneal spread of ovarian cancer, and represent evidence of the therapeutic potential of anti-IL-6R therapy for ovarian cancer treatment.

The Galβ1-4Gal epitope is rarely found in mammals, and the natural antibody against Galβ1-4Gal is rich in human. In contrast, we have previously demonstrated the presence of Galβ1-4Gal in pigeon and ostrich, and the absence of this epitope in chicken. Here, to further investigate the expression of this glycan among birds, egg white glycoproteins and egg yolk IgG from nine species of birds, namely, chicken, duck, emu, guineafowl, ostrich, peafowl, pigeon, quail, and turkey, were analyzed by western blot using an anti-(Galβ1-4Gal) antibody. The results indicated that some egg white glycoproteins from emu, ostrich, and quail, and heavy chains of IgG from all of the birds, except chicken and quail, were stained with the antibody. The presence of Galβ1-4Gal on N-glycans of IgGs from guineafowl, peafowl, and turkey were confirmed by mass spectrometry (MS), MS/MS, and MS(n) analyses. In quail, the presence of Galβ1-4Gal was confirmed by detecting the activities of UDP-galactose: β-galactoside β1,4-galactosyltransferase (β4GalT(Gal)) in various tissues, and by detecting Galβ1-4Gal by western blotting. In contrast, bamboo partridge, which is a close relative of chicken, did not show any detectable activities of β4GalT(Gal) or Galβ1-4Gal on glycoproteins. Because quail, peafowl, turkey, chicken, and bamboo partridge belong to the same family, i.e., Phasianidae, expression of Galβ1-4Gal was most likely differentiated within this family. Considering that Galβ1-4Gal is also expressed in ostrich, emu, and pigeon, which are phylogenetically distant relatives within modern birds, Galβ1-4Gal expression appears to be widely distributed among birds, but might have been abolished in the ancestors of chicken and bamboo partridge.

Cannabinoid (CB) receptor agonists are expected to alleviate ischemic brain damage by modulating neurotransmission and neuroinflammatory responses via CB(1) and CB(2) receptors, respectively. In a previous study, TAK-937, a novel potent and selective CB(1) and CB(2) receptor agonist, was shown to exert significant cerebroprotective effects accompanied by hypothermia after transient middle cerebral artery occlusion (MCAO) in rats. Sustained hypothermia itself induces significant neuroprotective effects. In the present studies, we examined the relative contribution of hypothermia and CB(1) receptor activation to the cerebroprotective effects of TAK-937.Using a multichannel brain temperature controlling system we developed, the brain temperature of freely moving rats was telemetrically monitored and maintained between 37 and 38°C during intravenous infusion of TAK-937 (100 µg/kg/h) or vehicle for 24 h after 2 h MCAO. AM251, a selective CB(1) receptor antagonist, was administered intraperitoneally at 30 mg/kg 30 min before starting intravenous infusion of TAK-937 (100 µg/kg/h) for 24 h. Rats were sacrificed and their brains were isolated 26 h after MCAO in both experiments. When the hypothermic effect of TAK-937 was completely reversed by a brain temperature controlling system, the infarct-reducing effect of TAK-937 was attenuated in part, but remained significant. On the other hand, concomitant AM251 treatment with TAK-937 completely abolished the hypothermic and infarct-reducing effects of TAK-937.We conclude that the cerebroprotective effects of TAK-937 were at least in part mediated by induction of hypothermia, and mainly mediated by CB(1) receptor activation.

This paper experimentally shows the importance of perceptual continuity of the expressive strength in vocal timbre for natural change in vocal expression. In order to synthesize various and continuous expressive strengths with vocal timbre, we investigated gradually changing expressions by applying the STRAIGHT speech morphing algorithm to singing voices. Here, a singing voice without expression is used as the base of morphing, and singing voices with three different expressions are used as the target. Through statistical analyses of perceptual evaluations, we confirmed that the proposed morphing algorithm provides perceptual continuity of vocal timbre. Our results showed the following: (i) gradual strengths in absolute evaluations, and (ii) a perceptually linear strength provided by the calculation of corrected intervals of the morph ratio by the inverse (reciprocal) function of an equation that approximates the perceptual strength. Finally, we concluded that applying continuity was highly effective for achieving perceptual naturalness, judging from the results showing that (iii) our gradual transformation method can perform well for perceived naturalness.

Background: Endoscopic submucosal dissection (ESD) is a reliable method that can replace surgery in the treatment of early colorectal cancer under certain conditions. Aim: The aim of the study was to analyze factors influencing the ESD procedure in early colorectal cancer, with the intention of improving its effectiveness. Patients and Methods: We conducted a multicenter, retrospective cohort study on 214 patients who underwent ESD procedures for early colorectal cancer from January 2016 to October 2023. Results:En bloc resection was achieved in 197 (92.1%) of ESD procedures, R0 resection in 149 (69.6%), and curative resection in 54 (40.9%). The submucosal invasion was classified as level 1 (SM1) in 96 cases (45.3%), level 2 (SM2) in 61 cases (28.8%), and level 3 (SM3) in 36 cases (17%). R0 resection was achieved more often in the rectum—92 (81.4%), compared to the right—24 (64.9%) and left colon—33 (61.1%), p = 0.009. In rectal tumors, R0 resection was achieved in 51 (98.1%) SM1 invasion, 27 (73%) SM2 invasion, and 13 (65%) SM3 invasion (p < 0.001). Lateral and vertical resection margins were positive in 12 (7.7%) and 52 (25.2%) cases, respectively. Vertical resection margins were statistically more often positive in lesions located in the right colon—11 cases (28.9%) and left colon—21 cases (38.9%), than in rectum—20 cases (17.5%); p = 0.010. Complications were found in 32 (15%) cases of ESD procedure—perforation in 12 cases (5.6%) and delayed bleeding in four cases (1.9%). Procedures performed in the right colon were associated with a significantly higher risk of any complications (30%) and perforations (15%) than those performed in the rectum (10.3% and 2.6%) or the left colon (13.8% and 5.2%; p = 0.016; p = 0.015), respectively. Conclusions: ESD for early colon cancer is a viable strategy due to its effectiveness and low complication rate. The ESD technique performed in the rectum yields the best results; however, in the right colon, it still requires careful attention. [ABSTRACT FROM AUTHOR]

Titin is a giant protein that is specifically expressed in striated muscle and essential for the maintenance of sarcomere structure and function. Recently, the N-terminal fragment of the Titin (N-titin) has been reported to show high levels in human urine in patients with muscular diseases and is expected to serve as a diagnostic biomarker for these diseases. In this study, we examined the utility of N-titin as a biomarker to detect muscle atrophy in mice. Male BALB/c mice (6 weeks of age, n=5 per group) were given 10 mg/L dexamethasone (DEX) dissolved in drinking water for 4 weeks. The gastrocnemius muscle (GAS) weight was significantly decreased and mRNA levels of muscle atrophy-related genes (Atrogin-1 and MuRF-1) were increased in the GAS after 4 weeks of DEX treatment. Although there were no degenerative/necrotic changes in the histopathological examination, the muscle fiber cross-sectional area significantly decreased in the GAS. On the other hand, there were no DEX treatment-related changes in the muscle weights and the muscle fiber cross-sectional area in the soleus muscle. These results suggest that 4-week of DEX treatment preferentially caused atrophy of fast-dominant muscle. Under the condition of this study, urinary N-titin/CRN ratio markedly increased from Week 2 of the DEX treatment. From the above results, the urinary N-titin/CRN ratio could be a biomarker for monitoring skeletal muscle atrophy in mice. [ABSTRACT FROM AUTHOR]