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T follicular helper (Tfh) cells and T peripheral helper (Tph) cells produce interleukin (IL)-21 and are thought to contribute to follicular and extra-follicular B-cell activation, respectively, in autoimmune diseases. It is known that programmed cell death-1 (PD-1)-positive CXCR5+ Tfh-like cells are differentiated from human naive CD4+ T cells by IL-12 plus transforming growth factor (TGF)-β. However, it remains unclear what cytokines are required for Tph differentiation. In this study, we found that interferon (IFN)-α and IFN-β reduce the frequency of Tfh-like cells under the IL-12 plus TGF-β condition, whereas they promote generation of PD-1+CXCR5-CD4+ T cells and secretion of IL-21, IFN-γ and CXCL13. Intracellular cytokine staining and T-cell-B-cell co-culture studies indicated that IFN-α promotes generation of IL-21+IFN-γ +CXCR5-CD4+ T cells thereby enhancing B-cell helper function. By IFN-α treatment, the mRNA levels of IL21, IFNG, CXCL13, CD244, SLAMF7, GZMB and PRDM1 were significantly up-regulated but BCL6 mRNA expression was down-regulated, suggesting a Tph-related gene expression pattern. On the other hand, IL-2-neutralization increased mRNA levels of IL21, CXCL13 and CXCR5, retained BCL6, but showed no clear effect on IFNG or PRDM1. RNA sequencing analyses revealed that PD-1hiCXCR5-CD4+ T cells prepared from in vitro culture show a Tph-related gene expression pattern similar with that of PD-1hiCXCR5- Tph cells obtained from the blood of patients with systemic lupus erythematosus. From our findings, it is highly probable that type I IFNs play a key role in differentiation of Tph cells and trigger Tph cell expansion in autoimmune diseases., (© The Author(s) 2022. Published by Oxford University Press on behalf of The Japanese Society for Immunology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The aim of this study was to clarify the clinical significance of bone metabolism in the mandibular condyles in determining condylar resorptive changes. Twelve condyles of patients with idiopathic condylar resorption and degenerative joint disease were analysed using 99m Tc HMDP SPECT/CT at baseline and subsequent computed tomography during the follow-up period. Twenty-two healthy condyles were enrolled as controls. After generating three-dimensional SPECT/CT images, two independent observers scored the degree of condylar uptake and measured the morphological changes in the condylar height and condylar volume. In the group with positive condylar uptake, the follow-up computed tomography showed significant decreases in condylar height (-1.69 ± 0.93 mm) and condylar volume (-12.51 ± 10.30%) when compared to healthy controls (condylar height, 0.09 ± 0.54 mm; condylar volume, -0.29 ± 4.22%) (P < 0.001). Moreover, the degree of uptake correlated with the changes in condylar height (observer 1, P = 0.012; observer 2, P = 0.039) and condylar volume (observer 1, P = 0.005; observer 2, P = 0.037). These results suggest that condylar bone metabolism is closely related to the resorptive activity. Thus, SPECT/CT would be useful in the prognostic evaluation or determination of treatment strategies for idiopathic condylar resorption and degenerative joint disease., (Copyright © 2021 International Association of Oral and Maxillofacial Surgeons. Published by Elsevier Inc. All rights reserved.)

Background: This study aimed to determine the rate, timing, and predictors of diabetes and exocrine pancreatic insufficiency after pancreatectomy in order to inform preoperative patient counseling and risk management strategies., Methods: Using prescription claims as a surrogate for disease prevalence, IBM Watson Health MarketScan was queried for claims patterns pre- and post-pancreatectomy. Multivariable models explored associations between clinical characteristics and medication use within 2 years of surgery., Results: In total, 18.96% of 2,848 pancreaticoduodenectomy (PD) patients and 18.95% of 1,858 distal pancreatectomy (DP) patients had preoperative diabetic medication prescription claims. Fewer (6.6% and 3.88%, respectively) had pancreatic enzyme replacement therapy (PERT) claims. Diabetic medication claims increased to 28.69% after PD and 38.59% after DP [adjusted relative risk (aRR) = 1.36 (95% CI 1.27, 1.46)]. Other associated factors included age > 45, medical comorbidity, and obesity. The incidence of new diabetic medication claims among medication naïve patients was 13.78% for PD and 24.7% for DP (p < 0.001) with a median 4.7 and 4.9 months post-operatively. The prevalence of PERT claims was 55.97% after PD and 17.06% after DP [aRR = 0.32 (0.29, 0.36)]. The incidence of postoperative PERT claims 53.98% (PD) and 14.84% (DP) (p < 0.0001). The median time to new PERT claim was 3.0 (PD) and 3.2 (DP) months, respectively. Claims for both diabetic medications and PERT rose sharply after surgery and plateaued within 6 months., Conclusions: This study defines prevalence, timing, and predictors for post-pancreatectomy insufficiency to inform preoperative counseling, risk modification strategies, and interventions related to quality of life., (© 2022. The Society for Surgery of the Alimentary Tract.)

Stream-dwelling white-spotted charr, Salvelinus leucomaenis , populations tend to have unique color and spot patterns in different regions and may even display stream-specific patterns. An extreme edge of such diversity, found in individuals with atypical body color patterns (so-called nagaremon-type charr, a rare morphotype of Salvelinus leucomaenis [hereafter termed nagaremon-charr]), sympatrically occurring with normal-charr, has been reported from only six small isolated populations in Japan. Based on morphological and ecological perspectives, nagaremon-charr has been considered as an intraspecific color variant of white-spotted charr, although the genetic status of nagaremon-charr has not been determined. In this study, genetic diversity and population structure of the nagaremon-charr in a tributary of the Ane River (Lake Biwa system) were investigated through microsatellite and mtDNA analyses. Nagaremon-charr and sympatric normal-charr in the tributary shared the mtDNA haplotypes and were assigned to the same cluster in the STRUCTURE analysis and discriminant analysis of principal components (DAPC). These results suggested that nagaremon-charr in the Ane River is an intra-populational specific color variant of white-spotted charr. Above a waterfall, nagaremon-charr specimens exhibited extremely reduced genetic diversity, indicating that genetic drift may account for the fixation of the nagaremon-morphotype. Normal-charr below the waterfall clustered separately from hatchery-reared charr, indicative of native status of the former. Thus, both nagaremon-charr and normal-charr in the entire Ane River tributary should be conserved.

Oxygen and carbon stable isotope ratios ( 18 O/ 16 O, 13 C/ 12 C, and 17 O/ 16 O) of CO 2 have been crucial in helping us understand Earth and planetary systems. These ratios have also been used in medicine for the noninvasive diagnosis of diseases from exhaled breath and for quantifying biochemical or metabolic reactions and in determining the production area of agricultural products. The current method for measuring the stable isotope ratios of CO 2 is primarily gas-source isotope ratio mass spectroscopy (IRMS). Due to the recent demand for isotopic microanalysis of carbonates and organic compounds, the sample size required for isotopic measurements has been reduced to approximately 2 nmol CO 2 (equivalent to 0.2 μg CaCO 3 and 24 ng carbon) by using high-precision IRMS. We report a novel method using tunable mid-infrared laser direct absorption spectroscopy (TILDAS) for sensitive measurements of 18 O/ 16 O and 13 C/ 12 C in subnanomolar CO 2 . This method can accurately measure 18 O/ 16 O and 13 C/ 12 C in CO 2 with a repeatability of less than 0.03‰ ( n = 28) in a range of 0.3 nmol (equivalent to 0.03 μg CaCO 3 and 3.8 ng carbon) to 30 nmol. This is a sample size 1 order of magnitude smaller than currently available sensitive analytical techniques. In addition, the TILDAS system measures 17 O/ 16 O simultaneously with a repeatability of less than 0.06‰ ( n = 28). Our method is a major advance in supersensitive CO 2 stable isotopic analyses for various fields.

Purpose/objectives: Simulated educational models of teeth, bones, and gums have been used for a long time in procedural skills training in dentistry.  The advent of three-dimensional (3D) printing technologies and additive manufacturing has facilitated the production of more advanced 3D printed models that can be utilized for surgical and dental training together with other technologies (e.g., 3D scanners and image analysis software)., Methods: We conducted training on osteotomy in the maxilla and mandible using maxillofacial simulation software (MSS) and a 3D-printed model for 5th-year undergraduate dental students (13 students in 2017 and 11 students in 2018 with more than half of their results). We compared the group (13 students) who performed osteotomy after participating in the simulation lecture with those (11 students) who performed osteotomy after performing self-simulation (they were instructed to bring their personal computers and install the MSS) using tests and questionnaires (pre- and post-curriculum)., Results: The average test score was significantly higher in the group who performed osteotomy using the 3D-printed model after performing self-simulation; participants in this group had acquired a better understanding of the surgical procedure. Comparison using questionnaires showed a significant difference in the students' understanding of instruments and surgical techniques between both groups., Conclusion: The MSS and 3D printed models are widely used clinically. Incorporating these in the curriculum will help accelerate student development. In addition, prompt education on the purpose and usefulness of these tools will not only facilitate simulation software and 3D-printed model-based treatment plan in the clinic but also promote further research., (© 2022 The Authors. Journal of Dental Education published by Wiley Periodicals LLC on behalf of American Dental Education Association.)

Background: With improvements in the safety and stability of surgeries, the number of orthognathic surgeries is increasing. Most patients who undergo orthognathic surgeries are younger, and the number of orthognathic surgeries for patients with comorbidities is also increasing. We report a survey and clinical investigation of patients with comorbidities who underwent orthognathic surgeries at our department to improve the safety of orthognathic surgery., Results: The participants included 296 men and 712 women, with a mean age of 28 years (13-19 years, n=144; 20-29 years, n=483; 30-39 years, n=236; 40-49 years, n=102; 50-59 years, n=39; ≥60 years, n=4). In total, 347 patients underwent one-stage Le Fort type I osteotomy and sagittal split ramus osteotomy (SSRO), 243 underwent SSRO, 287 underwent plate removal, 126 underwent genioplasty and plate removal, and five underwent other surgeries. In total, 529 patients had comorbidities (52%), including allergic diseases (n=220, 33%), respiratory diseases (n=107, 16%), neurologic and psychiatric diseases (n=69, 10%), gynecologic diseases (n=28, 4%), hematologic diseases (n=27, 4%), cardiovascular diseases (n=24, 4%), digestive diseases (n=22, 3%), metabolic and endocrine diseases (n=18, 3%), spinal diseases (n=11, 2%), ophthalmologic diseases (n=11, 2%), renal and urological diseases (n=9, 1%), and other diseases (n=117, 18%). Among the patients with comorbidities, 11 with hemorrhagic diatheses (hemophilia and von Willebrand disease), arrhythmia (atrioventricular block), psychiatric disease (adjustment disorder), and metabolic disease (diabetes) required cautious perioperative management. The patient with hemophilia was managed with regular low-dose recombinant factor VIII replacement therapy, and the patient with type I diabetes mellitus was administered continuous insulin infusion and sliding-scale insulin therapy; both patients had an uneventful course., Conclusions: The study findings suggest that with the increase in orthognathic surgeries, oral and maxillofacial surgeons should adequately manage cases requiring cautious perioperative control and highlight the importance of preoperative screening. Despite the well-established safety and postoperative stability of orthognathic surgeries, oral surgeons should adopt appropriate additional preventive measures for patients with comorbidities., (© 2022. The Author(s).)

Background: Excessive bleeding is a major intraoperative risk associated with orthognathic surgery. This study aimed to investigate the factors involved in massive bleeding during orthognathic surgeries so that safe surgeries can be performed. Patients (n=213) diagnosed with jaw deformities and treated with bimaxillary orthognathic surgery (Le Fort I osteotomy and bilateral sagittal split ramus osteotomy) in the Department of Oral and Maxillofacial Surgery at the Suidobashi Hospital, Tokyo Dental College between January 2014 and December 2016 were included. Using the patients' medical and operative records, the number of cases according to sex, age at the time of surgery, body mass index (BMI), circulating blood volume, diagnosis of maxillary deformity, direction of maxillary movement, operative duration, incidence of bad split, injury of nasal mucosa, and blood type were analyzed., Results: The results revealed that BMI, circulating blood volume, nasal mucosal injury, and operative time were associated with the risk of intraoperative massive bleeding in orthognathic surgeries. Chi-square tests and binomial logistic regression analyses showed significant differences in BMI, circulating blood volume, direction of maxillary movement, operative duration, and injury to the nasal mucosa. Operative duration emerged as the most important risk factor. Furthermore, a >4-mm upward migration of the posterior nasal spine predicted the risk of massive bleeding in orthognathic surgery., Conclusions: The upward movement of the maxilla should be recognized during the preoperative planning stage as a risk factor for intraoperative bleeding, and avoiding damage to the nasal mucosa should be considered a requirement for surgeons to prevent massive bleeding during surgery., (© 2022. The Author(s).)

Aim: Sarcopenia - aging-related loss of muscle mass and muscle strength - is a key feature of the frailty model. In the present study, we aimed to elucidate the molecular biological changes associated with aging in the extensor digitorum longus muscle of senescence-accelerated mouse prone 8 mouse model by capillary electrophoresis-mass spectrometry., Methods: Three groups of senescence-accelerated mouse prone 8 mice were used, namely, 12-week-old (young; n = 5), 40-week-old (elderly; n = 5) and 55-week-old mice (late elderly; n = 5). The extensor digitorum longus muscle was collected. After preliminary analyses, metabolome analysis was carried out by capillary electrophoresis-mass spectrometry. Additionally, we examined whether the activity of enzymes in the metabolic pathway fluctuated with aging, by real-time polymerase chain reaction., Results: Among the 116 water-soluble metabolites associated with the central energy metabolism pathway, changes were observed in 19 metabolites between 12- and 40 -weeks-old, in 40 metabolites between 40- and 55-weeks-old, and in 57 metabolites between 12- and 55-weeks-old. The fluctuated metabolites that were common among the groups were Val, putrescine and His. The levels of putrescine, associated with cell proliferation, protein synthesis and nucleic acid synthesis, and β-Ala and His, a component of carnosine that is characterized by its anti-oxidant and anti-fatigue effects, decreased with age., Conclusions: We confirmed that there were two aging-related metabolic changes in the extensor digitorum longus muscle of senescence-accelerated mouse prone 8 mice. Based on the changes in metabolites, cell senescence and fatigue in the extensor digitorum longus muscle might increase in old mice compared with those in young mice, showing molecular biological changes with aging. Geriatr Gerontol Int 2022; 22: 160-167., (© 2021 The Authors. Geriatrics & Gerontology International published by John Wiley & Sons Australia, Ltd on behalf of Japan Geriatrics Society.)

Paeoniflorin and albiflorin, which are functional isomers, are the major constituents of an herbal medicine derived from Paeonia lactiflora . Those functional isomers and their galloylated derivatives, which are positional isomers, were studied by matrix-assisted laser desorption/ionization-tandem mass spectrometry (MALDI-MS/MS). The resulting mass spectra are discussed based on the fragmentation patterns of the sodium adducts. The product ion spectra of 4- O -galloylalbiflorin and 4'- O -galloylpaeoniflorin differed, even though they were positional isomers. The fragmentations of the ester parts were influenced by the neighboring hydroxyl groups. The ionization efficiency of the sodium adduct of albiflorin was higher than that for paeoniflorin. These results indicate that the carboxylic ester group has a higher affinity for sodium ions than the acetal group, which can be attributed to the carbonyl oxygen being negatively polarized, allowing it to function as a Lewis base., (Copyright © 2022 Tohru Yamagaki, Kohtaro Sugahara, Kohki Fujikawa, and Kazuto Washida.)

Thirteen anthocyanins were isolated from the flowers of two Aeschynanthus species, A. fulgens and A. pulcher, and six cultivars, 'Mahligai', 'Mona Lisa', 'SoeKa', 'Redona', 'Freshya' and 'Bravera', and identified as pelargonidin and cyanidin 3-O-sambubiosides and their malonates, succinates, p-coumarates and caffeates, and pelargonidin 3-O-glucoside by acid hydrolysis, HR-MS and NMR. Of their anthocyanins, pelargonidin 3-O-[xylosyl-(1 → 2)-(6''-malonylglucoside)] (2), pelargonidin 3-O-[xylosyl-(1 → 2)-(6''-succinylglucoside)] (3), pelargonidin 3-O-[xylosyl-(1 → 2)-(6''-E-p-coumaroylglucoside)] (4), pelargonidin 3-O-[xylosyl-(1 → 2)-(6''-Z-p-coumaroylglucoside)] (5), pelargonidin 3-O-[xylosyl-(1 → 2)-(6''-E-caffeoylglucoside)] (6) and cyanidin 3-O-[xylosyl-(1 → 2)-(6''-succinylglucoside)] (9) were reported in nature for the first time., (Crown Copyright © 2021. Published by Elsevier Ltd. All rights reserved.)

Little is known regarding the DNA methyltransferases (MTases) in hyperthermophilic archaea. In this study, we focus on an MTase from Aeropyrum pernix K1, a hyperthermophilic archaeon that is found in hydrothermal vents and whose optimum growth temperature is 90°C to 95°C. From genomic sequence analysis, A. pernix K1 has been predicted to have a restriction-modification system (R-M system). The restriction endonuclease from A. pernix K1 (known as ApeKI from New England BioLabs Inc. [catalog code R06435]) has been described previously, but the properties of the MTase from A. pernix K1 (M.ApeKI) have not yet been clarified. Thus, we demonstrated the properties of M.ApeKI. In this study, M.ApeKI was expressed in Escherichia coli strain JM109 and affinity purified using its His tag. The recognition sequence of M.ApeKI was determined by methylation activity and bisulfite sequencing (BS-seq). High-performance liquid chromatography (HPLC) was used to detect the position of the methyl group in methylated cytosine. As a result, it was clarified that M.ApeKI adds the methyl group at the C-5 position of the second cytosine in 5'-GCWGC-3'. Moreover, we also determined that the MTase optimum temperature was over 70°C and that it is strongly tolerant to high temperatures. M.ApeKI is the first highly thermostable DNA (cytosine-5)-methyltransferase to be evaluated by experimental evidence. IMPORTANCE In general, thermophilic bacteria with optimum growth temperatures over or equal to 60°C have been predicted to include only N 4 -methylcytosine or N 6 -methyladenine as methylated bases in their DNA, because 5-methylcytosine is susceptible to deamination by heat. However, from this study, A. pernix K1, with an optimum growth temperature at 95°C, was demonstrated to produce a DNA (cytosine-5)-methyltransferase. Thus, A. pernix K1 presumably has 5-methylcytosine in its DNA and may produce an original repair system for the expected C-to-T mutations. M.ApeKI was demonstrated to be tolerant to high temperatures; thus, we expect that M.ApeKI may be valuable for the development of a novel analysis system or epigenetic editing tool.

Nutritional status is well-known to influence patient recovery after resection of head and neck cancer (HNC). The influence of preoperative nutritional status on dysphagia was assessed in patients who underwent surgical resection of HNC along with the assessment of nutritional status during the acute and subacute phases. Eighty-six patients underwent surgical resection and dysphagia assessments (repetitive saliva-swallowing test, water-swallowing test, and functional oral intake scale) and had their tongue pressure assessed five times (before surgery, after 1-2 weeks, and 1, 2, and 3 months after surgery). The nutritional status was assessed according to the body mass index, total protein, and albumin. The prognostic nutritional index was calculated from preoperative data, and the subjects were classified into three groups: Low-risk, Attention and High-risk groups. After surgery, the nutritional status index values were low, and the High-risk group showed significantly lower values in comparison to the other two groups. The water-swallowing test and functional oral intake scale findings were worse than they had been preoperatively until 2 months after surgery, and a significant correlation was noted between the postoperative nutritional status and the presence of dysphagia. The results indicated that the preoperative nutritional status of HNC patients influenced their ability to ingest/swallow, which in turn influenced their nutritional status after HNC resection.

The aim of this study was to verify the reproducibility and accuracy of preoperative planning in maxilla repositioning surgery performed with the use of computer-aided design/manufacturing technologies and mixed reality surgical navigation, using new registration markers and the HoloLens headset. Eighteen patients with a mean age of 26.0 years were included. Postoperative evaluations were conducted by comparing the preoperative virtual operation three-dimensional image (Tv) with the 1-month postoperative computed tomography image (T 1 ). The three-dimensional surface analysis errors ranged from 79.9% to 97.1%, with an average error of 90.3%. In the point-based analysis, the errors at each point on the XYZ axes were calculated for Tv and T 1 in all cases. The median signed value deviation of all calculated points on the XYZ axes was -0.03mm (range -2.93mm to 3.93mm). The median absolute value deviation of all calculated points on the XYZ axes was 0.38mm (range 0mm to 3.93mm). There were no statistically significant differences between any of the points on any of the axes. These values indicate that the method used was able to reproduce the maxilla position with high accuracy., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: Pancreatogenic diabetes is common after pancreatectomy, and the impact on quality of life (QOL) is poorly understood. The objective of this study was to investigate QOL between diabetic and non-diabetic patients at least five years after pancreatectomy., Methods: Patients were recruited from a prospectively maintained institutional database. Participants were administered the Audit of Diabetes-Dependent Quality of Life (ADDQOL). Quality of life was compared between diabetics and non-diabetics using validated European Organization for Research and Treatment of Cancer questionnaires., Results: 80 individuals completed surveys. 55% were female, 80% non-Hispanic white, 44% underwent Whipple, 48% were cystic neoplasms and 39% were adenocarcinoma. Diabetic patients (42.5%) reported comparable EORTC QLQ-C30 and Pan26 scores to non-diabetic patients. Pre-operative diabetic patients reported more dyspnea (p = 0.02) and greater pain (p = 0.02) than new-onset diabetics. Diabetic patients reported an overall ADDQOL quality of life score 'very good' (IQR: excellent, good) though felt life would be much better without diabetes (IQR: very much better, little better). While operation type was not influential, patients diagnosed with cystic neoplasms were almost twice as likely as those with other pathologies to report that life would be much better without diabetes (p < 0.01)., Conclusion: At a median of 9.3 years from pancreatic surgery, ADDQoL scores of patients were similar to cohorts of non-pancreatogenic diabetics in the general population. Patients without cancer were more likely to report that diabetes affected their overall QOL, regardless of operation. This study provides nuanced understanding of long-term QOL to improve the informed consent process and post-operative long-term care., Competing Interests: Declaration of competing interest We have no conflicts of interest, sources of financial support, corporate involvement or patent holdings to report., (Copyright © 2021 IAP and EPC. Published by Elsevier B.V. All rights reserved.)

Objective: To clarify relevant proteins and clinical characteristics of a phenotype of IgG4-related disease (IgG4-RD) with lymphadenopathy., Methods: We enrolled patients newly diagnosed with IgG4-RD in our department between January 2000 and June 2018 and performed proteomic analysis to measure serum concentrations of 1305 proteins. We extracted proteins overexpressed in patients with IgG4-RD with lymphadenopathy by comparing between those with lymphadenopathy, those without lymphadenopathy and healthy controls. We further reviewed all the patients with IgG4-RD in our institution and investigated the characteristics and prognosis of the patients with IgG4-RD with lymphadenopathy., Results: Eighty-five patients with IgG4-RD were enrolled, of which, 55% had lymphadenopathy. Proteomic analysis in 31 patients with IgG4-RD and 6 healthy controls revealed that eotaxin-3 was a potential serum biomarker in the patients with lymphadenopathy versus those without lymphadenopathy and healthy controls. A cohort of 85 patients with IgG4-RD demonstrated that patients with lymphadenopathy showed a significantly higher serum IgG4, IgG4:IgG ratio, IgG4-RD responder index and eosinophilia (P < 0.001 for all), irrelevant of the extent to which organ involvement developed. Patients with lymphadenopathy treated with glucocorticoid alone relapsed with significantly higher rates than those without lymphadenopathy (P = 0.03)., Conclusion: Lymphadenopathy in IgG4-RD represents a phenotype associated with high disease activities, eosinophilia and relapsing disease. Eotaxin-3 is a novel biomarker related to IgG4-RD with lymphadenopathy., (© The Author(s) 2020. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Recently, a novel CS/DS 4-O-endosulfatase was identified from a marine bacterium and its catalytic mechanism was investigated further (Wang, W., et. al (2015) J. Biol. Chem.290, 7823-7832; Wang, S., et. al (2019) Front. Microbiol.10, 1309). In the study herein, we provide new insight about the structural characteristics of the substrate which determine the activity of this enzyme. The substrate specificities of the 4-O-endosulfatase were probed by using libraries of structure-defined CS/DS oligosaccharides issued from synthetic and enzymatic sources. We found that this 4-O-endosulfatase effectively remove the 4-O-sulfate of disaccharide sequences GlcUAβ1-3GalNAc(4S) or GlcUAβ1-3GalNAc(4S,6S) in all tested hexasaccharides. The sulfated GalNac residue is resistant to the enzyme when adjacent uronic residues are sulfated as shown by the lack of enzymatic desulfation of GlcUAβ1-3GalNAc(4S) connected to a disaccharide GlcUA(2S)β1-3GalNAc(6S) in an octasaccharide. The 3-O-sulfation of GlcUA was also shown to hinder the action of this enzyme. The 4-O-endosulfatase exhibited an oriented action from the reducing to the non-reducing whatever the saturation or not of the non-reducing end. Finally, the activity of the 4-O-endosulfatase decreases with the increase in substrate size. With the deeper understanding of this novel 4-O-endosulfatase, such chondroitin sulfate (CS)/dermatan sulfate (DS) sulfatase is a useful tool for exploring the structure-function relationship of CS/DS., (© 2021 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

To understand the unique green-blue color of Puya alpestris (Bromeliaceae) flowers, we investigated their constituent anthocyanin and related compounds. An anthocyanin, two undescribed flavonols, and two flavones were isolated and identified as delphinidin 3,3',5'-tri-O-β-glucopyranoside, myricetin 3-O-[α-rhamnopyranosyl-(1 → 6)-β-glucopyranoside]-3',5'-di-O-β-glucopyranoside, myricetin 3,3',5'-tri-O-β-glucopyranoside, luteolin 4'-O-glucoside, and apigenin 4'-O-glucoside. Furthermore, the presence of chlorophyll has also been revealed. P. alpestris petals show a gradient color appearance: Green-blue at the tip and blue at the base. This color difference between the tip and base was used to analyze the pigment components underlying the green-blue color expression. It was found that the petal tip contains the anthocyanin, flavonols, flavones, and chlorophyll in high quantities. Furthermore, the pH of petal juice was 6.2 and 5.6 at the tip and base, respectively. In vitro reconstruction revealed the blue color expression occurred via an intermolecular copigmentation between the anthocyanin and flavones, as well as yellow color expression, which was due to an increase in the absorption at 400-450 nm of the flavonols under the higher pH conditions. Furthermore, we found that the petal extract obtaining using 50% acetone containing chlorophyll showed the same absorption spectrum as that observed for the raw petal. These results indicate that the green-blue color of P. alpestris flowers is developed via an intermolecular co-pigmentation of the anthocyanin (delphinidin 3,3',5'-tri-O-β-glucopyranoside) with flavones, such as luteolin 4'-O-glucoside, the yellow color expression of flavonols, such as myricetin 3,3',5'-tri-O-glucoside under relatively high pH conditions in the cell sap, and the presence of chlorophyll., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Major bleeding associated with sagittal split ramus osteotomy (SSRO) involves vessels such as the inferior alveolar, facial, and maxillary arteries and veins, and the retromandibular vein (RMV). The present study aimed to clarify and classify the three-dimensional variations in RMV position and course direction in relation to the mandible. Specimens comprised a total of 15 scientific cadavers, and the relationship between RMV and the mandible lateral and posterior views was observed. We identified 3 patterns on the lateral view, the mean distance between the RMV and the posterior border of the ramus was 3.9 mm at the height of the lingula. A total of five course patterns were identified on the posterior view. In no course pattern, the RMV inferior to the lingula was lateral to its position superior to the lingual. The present findings suggest that it may be possible to predict correlations with intraoperative bleeding risk. Further study is planned using contrast computed tomography in patients with jaw deformity for skeletal classification.

Evolution of tumor-immune microenviroments (TIMEs) occurs during tumor growth and dissemination. Understanding inter-site tumor-immune heterogeneity is essential to harness the immune system for cancer therapy. While the development of immunotherapy against lung cancer with driver mutations and neuroendocrine tumors is ongoing, little is known about the TIME of large cell neuroendocrine carcinoma (LCNEC) or anaplastic lymphoma kinase ( ALK ) rearrangement-positive lung cancer. We present a case study of a 32-year-old female patient with ALK -rearrangement-positive LCNEC, who had multiple distant metastases including mediastinal lymph-node, bilateral breasts, multiple bones, liver and brain. Multiple biopsy samples obtained from primary lung and three metastatic tumors were analyzed by fluorescent multiplex immunohistochemistry. Tissue localizations of tumor-infiltrating lymphocytes in the tumor nest and surrounding stroma were evaluated. T cell and B cell infiltrations were decreased with distance from primary lung lesion. Although each tumor displayed a unique TIME, all tumors exhibited concomitant regression after treatment with an ALK-inhibitor. This study provides the first evidence of the coexistence of distinct TIME within a single individual with ALK -rearrangement-positive LCNEC. The present study contributes to our understanding of heterogeneous TIMEs between primary and metastatic lesions and provides new insights into the complex interplay between host-immunity and cancer cells in primary and metastatic lesions.

In recent years, non-invasive measurement of tissue stiffness (hardness) using ultrasound elastography has attracted considerable attention. It has been used to evaluate muscle stiffness in the fields of rehabilitation, sports, and orthopedics. However, ultrasonic diagnostic devices with elastography systems are expensive and clinical use of such devices has been limited. In this study, we proposed a novel estimation method for vibration-based shear wave elastography measurement of human skeletal muscle, then determined its reproducibility and reliability. The coefficient of variation and correlation coefficient were used to determine reproducibility and reliability of the method by measuring the shear wave velocities in konjac phantom gels and agar phantom gels, as well as skeletal muscle. The intra-day, day-to-day, and inter-operator reliabilities were good when measuring the shear wave velocities in phantom gels. The intra-day and day-to-day reliabilities were good when measuring the shear wave velocities in skeletal muscle. The findings confirmed adequate reproducibility and reliability of the novel estimation method for vibration-based shear wave elastography. Therefore, the proposed measurement method may be a useful tool for evaluation of muscle stiffness.

Background: Le Fort I osteotomy is a highly effective treatment for skeletal jaw deformities and is commonly performed. High Le Fort I osteotomy is a modified surgical procedure performed for improving the depression of the cheeks by setting the osteotomy higher than the conventional Le Fort I osteotomy. Developments in three-dimensional (3D) technology have popularized the use of 3D printers in various institutions, especially in orthognathic surgeries. In this study, we report a safe and inexpensive method of performing a high Le Fort I osteotomy using a novel 3D device and piezosurgery, which prevent tooth root injury without disturbing the operation field for patients with a short midface and long tooth roots., Results: A 17-year-old woman presented with facial asymmetry, mandibular protrusion, a short midface, and long tooth roots. We planned high Le Fort I osteotomy and bilateral sagittal split ramus osteotomy. Prevention of damage to the roots of the teeth and the infraorbital nerve and accurate determination of the posterior osteotomy line were crucial for clinical success. Le Fort I osteotomy using 3D devices has been reported previously but were particularly large in size for this case. Additionally, setting the fixing screw of the device was difficult, because of the risk of damage to the roots of the teeth. Therefore, a different surgical technique, other than the conventional Le Fort I osteotomy and 3D device, was required. The left and right parts of the 3D device were fabricated separately, to prevent any interference in the surgical field. Further, the 3D device was designed to accurately cover the bone surface from the piriform aperture to the infra-zygomatic crest with two fixation points (the anterior nasal spine and the piriform aperture), which ensured stabilization of the 3D device. The device is thin and does not interfere with the surgical field. Safe and accurate surgical performance is possible using this device and piezosurgery. The roots of the teeth and the infraorbital nerve were unharmed during the surgery., Conclusions: This device is considerably smaller than conventional devices and is a simple, low-cost, and efficient method for performing accurate high Le Fort I osteotomy., Competing Interests: Competing interestsThe authors declare that they have no competing interests., (© The Author(s) 2020.)

Disorders of the oral mucosa are considered easy to diagnose since they can be visualized and examined directly. A change in the color of the oral mucosa reflects histopathological changes and is an important diagnostic parameter. However, the subjective perception of color varies. To determine the extent of resection for oral mucosa conditions, it is necessary to digitize the color and perform objective assessments. In recent years, fluorescence visualization devices and analysis software that measure tissue luminance G have been employed for the identification of oral mucosa diseases. Fluorescence visualization is presumably based on the decrease in epithelial flavin adenine dinucleotide content and luminance G values due to the destruction of collagen cross-links [fluorescence visualization loss (FVL)]. However, cases with differences between luminance values and histopathological presentation exist. Therefore, additional factors may affect fluorescence visualization. The present study used a portable, non-contact oral mucosa fluorescence visualization device for luminance measurements in seven patients with oral squamous cell carcinoma. Furthermore, Picro-Sirius Red and immunohistochemical staining were performed for CK13, CK17, Ki67, p53 and E-cadherin in the FVL(+) (lesion) and FVL(-) (resection stump) areas to elucidate the principle of fluorescence visualization. Fluorescence was significantly lower in the FVL(+) than in the FVL(-) areas, and the mean luminance G value was 56. The Picro-Sirius Red stain revealed collagen destruction in the FVL(+) areas but no collagen disruption in the FVL(-) areas. CK13 was negative in the FVL(+) and positive in the FVL(-) areas, whereas the opposite pattern was observed for CK17. In the FVL(+) area, p53 staining was positive. E-cadherin expression was enhanced in the FVL(-) areas and reduced in the FVL(+) areas. Furthermore, the luminance G value tended to be lower in cases with weaker E-cadherin staining. The aforementioned results suggest that decreased E-cadherin expression may be a factor that regulates fluorescence visualization., (Copyright: © Sugahara et al.)

Tryptophan has a unique role as a nutritional signaling molecule that regulates protein synthesis in mouse and rat liver. However, the mechanism underlying the stimulating actions of tryptophan on hepatic protein synthesis remains unclear. Proteomic and metabolomic analyses were performed to identify candidate proteins and metabolites likely to play a role in the stimulation of protein synthesis by tryptophan. Overnight-fasted rats were orally administered L-tryptophan and then sacrificed 1 or 3 h after administration. Four differentially expressed protein spots were detected in rat liver at 3 h after tryptophan administration, of which one was identified as an ornithine aminotransferase (OAT) precursor. OAT is the main catabolic enzyme for ornithine, and its expression was significantly decreased by tryptophan administration. The concentration of ornithine was increased in the liver at 3 h after tryptophan administration. Ornithine is a precursor for polyamine biosynthesis. Significantly increased concentrations of polyamines were found in the liver at 3 h after administration of tryptophan. Additionally, enhanced hepatic protein synthesis was demonstrated by oral administration of putrescine. We speculate that the increase in ornithine level through suppression of OAT expression by tryptophan administration may lead to accelerated polyamine synthesis, thereby promoting protein synthesis in the liver.

Accumulating evidences suggested that the overactivation of epidermal growth factor receptor (EGFR) was involved in the development of adult respiratory distress syndrome and pulmonary fibrosis. Elucidation of the mechanisms that regulate EGFR residence on the plasma membrane during inflammatory lung conditions is important for identifying potential therapies. We have demonstrated that flagellin phosphorylated EGFR at Ser1047 and induced transient EGFR internalization. In this study, we examined the molecular pathway and effect of interleukin 1 beta (IL-1β) on EGFR in alveolar epithelial cells. Treatment of A549 cells with IL-1β induced the activation of p38 mitogen-activated protein kinase (MAP kinase) and MAP kinase-activated protein kinase-2 (MAPKAPK-2), as well as EGFR phosphorylation at serine 1047. Both MAPKAPK-2 activation and EGFR phosphorylation were inhibited by SB203580, a p38 MAP kinase inhibitor. In addition, MK2a inhibitor (a MAPKAPK-2 inhibitor) suppressed EGFR phosphorylation. Assessment of the biotinylation of cell surface proteins indicated that IL-1β induced EGFR internalization. Furthermore, long-term treatment of A549 cells with IL-1β caused morphological changes and loss of cell-cell contact. Moreover, IL-1β augmented the effect of transforming growth factor beta 1 on the epithelial-mesenchymal transition. These results suggested that IL-1β regulates EGFR functions and induces morphological changes of alveolar epithelial cells., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.)

Background: Pseudodiastrophic dysplasia (PDD) is a severe skeletal dysplasia associated with prenatal manifestation and early lethality. Clinically, PDD is classified as a 'dysplasia with multiple joint dislocations'; however, the molecular aetiology of the disorder is currently unknown., Methods: Whole exome sequencing (WES) was performed on three patients from two unrelated families, clinically diagnosed with PDD, in order to identify the underlying genetic cause. The functional effects of the identified variants were characterised using primary cells and human cell-based overexpression assays., Results: WES resulted in the identification of biallelic variants in the established skeletal dysplasia genes, B3GAT3 (family 1) and CANT1 (family 2). Mutations in these genes have previously been reported to cause 'multiple joint dislocations, short stature, and craniofacial dysmorphism with or without congenital heart defects' ('JDSCD'; B3GAT3) and Desbuquois dysplasia 1 (CANT1), disorders in the same nosological group as PDD. Follow-up of the B3GAT3 variants demonstrated significantly reduced B3GAT3/GlcAT-I expression. Downstream in vitro functional analysis revealed abolished biosynthesis of glycosaminoglycan side chains on proteoglycans. Functional evaluation of the CANT1 variant showed impaired nucleotidase activity, which results in inhibition of glycosaminoglycan synthesis through accumulation of uridine diphosphate., Conclusion: For the families described in this study, the PDD phenotype was caused by mutations in the known skeletal dysplasia genes B3GAT3 and CANT1 , demonstrating the advantage of genomic analyses in delineating the molecular diagnosis of skeletal dysplasias. This finding expands the phenotypic spectrum of B3GAT3-related and CANT1-related skeletal dysplasias to include PDD and highlights the significant phenotypic overlap of conditions within the proteoglycan biosynthesis pathway., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2020. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Objective: The temporal fascia has been widely used in tympanoplasty. In addition, the preserved fascia has been also used in the ear surgery. In this study, we planned the experiments to determine whether physical and chemical properties of the fascia preserved at a low temperature., Methods: Preserved temporal fasciae from 21 patients were used in this study. The thickness of the temporal fascia was measured under a 3D laser microscope. The tensile strength was evaluated using a tensile tester. In addition, the chemical property evaluated was the biologic antioxidative potential of samples., Results: The results showed that the strength of the fascia was not affected by the retention period. The thick fascia tended to show the less tensile strength. The intensity was highest in middle-aged donors when compared to young and older donor. The antioxidative potentials did not affect the preservation., Conclusion: The results suggested that the preserved temporal fascia could be safely used for tympanoplasty., Competing Interests: Declaration of Competing Interest No potential conflict of interest was reported by the authors., (Copyright © 2019 Oto-Rhino-Laryngological Society of Japan Inc. Published by Elsevier B.V. All rights reserved.)

Genetic testing is widely used as a rapid diagnostic method to identify microorganisms and detect antibiotic resistance genes. The nucleic acid to be analyzed is located inside the cell wall, the cell membrane or nuclear envelope. Therefore, it is essential to disassemble them in nucleic acid extraction operation. It is also necessary to remove or inactivate interfering substances by exposing cytoplasmic components accompanying cell disruption. Nucleic acid extraction is an indispensable task, but depending on the selected method, it may have a significant effect on the genetic test results. However, the DNA extraction method that is actually selected tends to emphasize work efficiency, and the appropriate evaluation of the extraction operation is neglected. In this study, we focused on the purity of the extracted DNA, and examined six existing extraction methods and original extraction methods using Gram-negative bacilli as a simple model. As a result, there was a large difference in DNA purity depending on the extraction method. When used in a qualitative gene amplification test, there was a difference in the shading of the bands. However, the detection of resistance genes all gave similar results. Furthermore, as a result of using the original extraction method, the extraction method using sodium decylbenzenesulfonate (SDBS) was the most excellent extraction method from the viewpoint of recovered DNA and operability.

Rationale: The plant hormone auxin, indole-3-acetic acid, regulates many aspects of plant growth and development. Auxin quantification should offer broad insights into its mechanistic action in plants. However, limited auxin content in plant tissues hampers the establishment of quantification methods without the highest graded instruments or deeply specialized experimental techniques., Methods: In this study, we detailed optimized conditions for high-performance liquid chromatography coupled with triple-quadrupole mass spectrometry (LC/MS). We compared LC/MS conditions, such as columns, mobile phases, parameters of acquisition methods (selective or multiple ion monitoring), dwell times (DTs), and channel numbers, using differentially mixed authentic auxin and its related compounds. We further investigated pretreatment methods through the optimization of auxin recovery and irrelative compound removal from plant tissues prior to the LC/MS analysis., Results: Our LC/MS analysis demonstrated the particular importance of the column, DTs, and channel numbers on detection sensitivity. Our comparative analysis developed optimal pretreatment methods, including the pulverization of plants, concentration of extract through centrifugal evaporation, and removal of irrelative metabolites using liquid-liquid extraction and a spin filter. We injected plant samples into our LC/MS system, quantified auxin and eight related compounds in a single measurement, and determined the auxin increase in an auxin over-producing mutant., Conclusions: Our practical optimization of LC/MS conditions and pretreatment methods provides detailed experimental processes toward the sensitive quantification of auxin from 10 mg of plant tissue. These processes have not always been clearly documented; therefore, our protocol could broadly contribute to technical advances in plant growth and development research., (© 2019 John Wiley & Sons, Ltd.)

Congenital disorders of glycosylation (CDGs) comprise a large number of inherited metabolic defects that affect the biosynthesis and attachment of glycans. CDGs manifest as a broad spectrum of disease, most often including neurodevelopmental and skeletal abnormalities and skin laxity. Two patients with biallelic CSGALNACT1 variants and a mild skeletal dysplasia have been described previously. We investigated two unrelated patients presenting with short stature with advanced bone age, facial dysmorphism, and mild language delay, in whom trio-exome sequencing identified novel biallelic CSGALNACT1 variants: compound heterozygosity for c.1294G>T (p.Asp432Tyr) and the deletion of exon 4 that includes the start codon in one patient, and homozygosity for c.791A>G (p.Asn264Ser) in the other patient. CSGALNACT1 encodes CSGalNAcT-1, a key enzyme in the biosynthesis of sulfated glycosaminoglycans chondroitin and dermatan sulfate. Biochemical studies demonstrated significantly reduced CSGalNAcT-1 activity of the novel missense variants, as reported previously for the p.Pro384Arg variant. Altered levels of chondroitin, dermatan, and heparan sulfate moieties were observed in patients' fibroblasts compared to controls. Our data indicate that biallelic loss-of-function mutations in CSGALNACT1 disturb glycosaminoglycan synthesis and cause a mild skeletal dysplasia with advanced bone age, CSGALNACT1-CDG., (© 2019 The Authors. Human Mutation published by Wiley Periodicals, Inc.)

Amiselimod (MT-1303) is a novel and selective sphingosine 1-phosphate receptor-1 (S1P 1 ) modulator with a more favorable cardiac safety profile than other S1P 1 receptor modulators. In this study, we evaluated the effects of MT-1303 on the progression of lupus nephritis in two well-known murine systemic lupus erythematosus (SLE) models, MRL/ lpr and NZBWF1 mice, compared with those of FK506. Daily oral doses of 0.1 and 0.3 mg/kg MT-1303 not only inhibited the development of lupus nephritis when administered before onset in MRL/ lpr and NZBWF1 mice but also improved symptoms of lupus nephritis when administered after onset in MRL/ lpr mice. Its efficacy in these models was more potent or comparable to that of FK506 (1 and 3 mg/kg). In histological analysis, treatment with MT-1303 inhibited infiltration of T cells into the kidneys, mesangial expansion, and glomerular sclerosis. MT-1303 treatment resulted in a marked reduction in T cells and B cells in the peripheral blood and significantly inhibited increases in the number of plasma cells in the spleen and T cells in the kidneys. In addition, administration of MT-1303 suppressed elevations in serum anti-dsDNA antibody levels in MRL/ lpr mice, but not in NZBWF1 mice. Our findings show that MT-1303 exhibits marked therapeutic effects on lupus nephritis in two SLE models, likely by reducing the infiltration of autoreactive T cells into the kidneys. These results suggest that MT-1303 has the potential to be used as a therapeutic agent for patients suffering from SLE, including lupus nephritis., Competing Interests: The authors declare that there is no conflict of interest regarding the publication of this paper., (Copyright © 2019 Kunio Sugahara et al.)

Amiselimod (MT-1303) is a novel sphingosine 1-phosphate receptor-1 (S1P1 receptor) modulator with a more favorable cardiac safety profile than other S1P1 receptor modulators. MT-1303 phosphate (MT-1303-P), an active metabolite of MT-1303, exhibits S1P1 receptor agonism at a lower EC50 value than other S1P1 receptor modulators currently being developed. We aimed to evaluate the efficacy of MT-1303 and its mode of action in chronic colitis using an inflammatory bowel disease (IBD) model. Oral administration of MT-1303 (0.3 mg/kg) once daily for 3 days to mice almost completely abolished S1P1 receptor expression on CD4+ T cells from mesenteric lymph nodes, which corresponded to a marked decrease in CD4+ T cell count in peripheral blood, indicating that MT-1303-P acts as a functional antagonist of the S1P1 receptor. The potential benefit of MT-1303 for IBD was assessed using immunodeficient SCID mice with chronic colitis induced by adoptive transfer of CD4+CD45RBhigh T cells from BALB/c mice. An oral dose of 0.1 and 0.3 mg/kg MT-1303 administered daily one week after the cell transfer inhibited the development of chronic colitis with an efficacy comparable to that of an anti-mTNF-α mAb (250 μg/mouse). In addition, MT-1303 administration significantly reduced the number of infiltrating Th1 and Th17 cells into the lamina propria of the colon in colitis mice. Our results suggest that MT-1303 acts as a functional antagonist of the S1P1 receptor on lymphocytes, regulates lymphocyte trafficking, and inhibits infiltration of colitogenic Th1 and Th17 cells into the colon to inhibit the development of chronic colitis., Competing Interests: Competing interests: KS, YM, HK, MM, SM, HU, KO, and KS are employees of Mitsubishi Tanabe Pharma Corporation. This does not alter the authors’ adherence to PLOS ONE policies on sharing data and materials. MT-1303 is the product in development, and Mitsubishi Tanabe Pharma Corporation has the ownership of the patents of MT-1303.

Objective: Tsumura Suzuki Obese Diabetes (TSOD) mice exhibit early age-associated hearing loss. Histopathological analysis of these mice shows narrowing of capillaries in the stria vascularis and chronic reduction of blood flow in the cochlea. In this study, we investigated the effect of oral administration of a herbal medicine or calorie restriction on hearing in TSOD mice., Methods: TSOD mice were divided into 4 groups: CR (calorie restriction), BF and DS (treated with the herbal medicines, Bofutsushosan and Daisaikoto, respectively), and the control group. Body weight, blood glucose levels, and auditory brainstem responses (ABRs) were measured. The cochleae were excised and evaluated histopathologically., Results: Blood glucose levels were suppressed in the CR, BF, and DS groups. In addition, the elevation of ABR thresholds was inhibited in the CR, BF, and DS groups. Cochlear blood vessels remained wide in the three treatment groups compared with the control group. These results suggested that the administration of these herbal medicines improved glucose tolerance and yielded results similar to those on calorie restriction., Conclusion: Oral administration of 2 herbal medicines can prevent hearing function disorder in a model mouse of diabetes. The results may clarify the possibility of clinical application., (Copyright © 2019 Elsevier B.V. All rights reserved.)

TECTA is well known as a causative gene for autosomal dominant mid-frequency hearing loss observed in various populations. In this study, we performed next-generation sequencing analysis of a large Japanese hearing loss cohort, including eight hundred and twelve (812) subjects from unrelated autosomal dominant hearing loss families, to estimate the prevalence and phenotype-genotype correlations in patients with TECTA mutations. The prevalence of TECTA mutations in Japanese autosomal dominant sensorineural hearing loss families was found to be 3.2%. With regard to the type of hearing loss, the patients with mutations in the nidogen-like domain or ZA domain of TECTA showed varied audiograms. However, most of the patients with mutations in the ZP domain showed mid-frequency hearing loss. The rate of hearing deterioration in TECTA -associated hearing loss patients and in the normal hearing Japanese control population were the same and regression lines for each group were parallel. We carried out haplotype analysis for four families which had one recurring missense variant, c.5597C>T (p.Thr1866Met). Our results revealed four different haplotypes, suggesting that this mutation occurred independently in each family. In conclusion, TECTA variants represent the second largest cause of autosomal dominant sensorineural hearing loss in Japan. The hearing loss progression observed in the patients with TECTA mutations might reflect presbycusis. The c.5597C>T mutation occurred in a mutational hot spot and is observed in many ethnic populations.

More than 400 syndromes associated with hearing loss and other symptoms have been described, corresponding to 30% of cases of hereditary hearing loss. In this study we aimed to clarify the mutation spectrum of syndromic hearing loss patients in Japan by using next-generation sequencing analysis with a multiple syndromic targeted resequencing panel (36 target genes). We analyzed single nucleotide variants, small insertions, deletions and copy number variations in the target genes. We enrolled 140 patients with any of 14 syndromes (BOR syndrome, Waardenburg syndrome, osteogenesis imperfecta, spondyloepiphyseal dysplasia congenita, Stickler syndrome, CHARGE syndrome, Jervell and Lange-Nielsen syndrome, Pendred syndrome, Klippel-Feil syndrome, Alport syndrome, Norrie disease, Treacher-Collins syndrome, Perrault syndrome and auditory neuropathy with optic atrophy) and identified the causative variants in 56% of the patients. This analysis could identify the causative variants in syndromic hearing loss patients in a short time with a high diagnostic rate. In addition, it was useful for the analysis of the cases who only partially fulfilled the diagnostic criteria.

Chondroitin sulfate E (CS-E) is a glycosaminoglycan containing type-E disaccharide units (sulfated at C-4 and C-6 of N-acetylgalactosamine). CS-E is covalently linked to a core protein to form chondroitin sulfate proteoglycans (PGs) that are secreted or associated with the plasma membrane of several types of cells. CS-E-containing PGs selectively interact with growth factors and chemokines and control various cellular and/or tissue processes. Angiogenesis is a process that is highly regulated in physiological conditions but deregulated in pathologies, leading to excess or deficient blood vessel formation. Angiogenesis regulation is orchestrated by numerous growth factors, such as vascular endothelial growth factor A, fibroblast growth factors and pleiotrophin, whose functions can be affected by CS-containing PGs. In the present review, we focus on the emerging area of CS-mediated angiogenesis and particularly on the critical assessment of data related to a potential role of CS-E in controlling endothelial cell functions, focusing on angiogenesis regulation and vascular homeostasis in health and disease., (© 2019 Federation of European Biochemical Societies.)

Stereotactic body radiation therapy (SBRT) is usually verified with a dynamic phantom or solid phantom, but there is a demand for phantoms that can accurately simulate tumor dynamics within an individual that would allow customized validation in every patient. We developed a new 4D dynamic target phantom (multi-cell 4D phantom) that allows simulation of tumor movement in patients. The basic quality and dynamic reproducibility of this new phantom was verified in this investigation. The newly developed multi-cell 4D phantom comprises four main components: soft tissue, bones, lungs, and tumor (target). The phantom structure was based on computed tomography (CT) data of a male. In this study, we investigated the basic performance of a multi-cell 4D phantom. All the CT numbers of the phantom were very close to those of human data. The geometric maximum amplitudes were 4.57 mm in the lateral direction, 4.59 mm in the ventrodorsal direction, and 3.68 mm in the cranio-caudal direction. Geometric errors were 0.84, 0.58, and 0.40 mm, respectively. Movements of the abdominal surface were stable for 60 s. Repeated measurements show no actual differences in target movements between multiple measurements and indicated high reproducibility (r  >  0.97). End-to-end tests using Gafchromic film revealed a gamma pass rate of 98% or above (2 mm/3%). Although our phantom performed limited reproducibility in the movement of the patient tumor at present, a satisfactory level of precision was confirmed in general. This is a very promising device for use in the verification of radiation therapy for moving targets.

Purpose: The aim of this study was to examine the effects of exposure to air pollution and cigarette smoke on respiratory function, respiratory symptoms, and the prevalence of COPD in individuals aged ≥50 years., Patients and Methods: We used spirometry and medical questionnaires to screen 433 individuals from Omuta City, Japan, an area with high levels of air pollution., Results: Non smokers had a high estimated COPD prevalence rate of 16%. Among smokers, the estimated prevalence of COPD was 29% in seniors (50- to 74-years group) and 37% in the elderly (>75 years group). We also found a correlation between levels of suspended particulate matter and COPD., Conclusion: Both smoking and chronic exposure to air pollution (>5 years) decreased respiratory function, exacerbated respiratory symptoms, and increased the prevalence of COPD. We strongly recommend periodic screening for the elderly patients to facilitate early detection of respiratory disease., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

Cryopyrin-associated periodic syndrome (CAPS) is one of the autoinflammatory disorders caused by mutations in NLRP3 gene. The over-production of interleukin (IL)-1β induced by NLRP3 gene mutations plays an important role in the pathophysiology of CAPS. We diagnosed 3 patients with CAPS, who were lineal family members having a novel mutation of NLRP3 gene. The objective of this report is to compare the characteristics of symptoms and differences in the therapeutic responses of them, who had the same mutation. In addition, we aimed to examine the usefulness of cytokine measurement for diagnosis or determination of treatment effect of CAPS. A 5-year-old Japanese boy (proband) came to our hospital because of short stature, reached the diagnosis of Muckle-Wells syndrome (MWS) due to a mutation in NLRP3 gene, which had not been reported so far (p.G328E, c.G983A). His mother and grandmother harbored the same mutation of NLRP3. We measured serum concentrations of cytokines in the proband assessed by flow-cytometric bead array. All of them had episodic skin eruptions with conjunctivitis, hearing loss, and arthralgia, but not periodic fever, cold-triggered episodes, and chronic aseptic meningitis. Only the proband had short stature. Canakinumab therapy led to a prompt relief of symptoms and normalized laboratory data in all patients. Audiograms demonstrated an improved hearing level in the proband, but not two others despite of the same mutation. All cytokines did not show any characteristic findings. Sensorineural hearing loss and itchless rash but not serum cytokine profile deserved attention to the diagnosis and treatment start of CAPS. The early intervention of IL-1β blockade may reduce the chance of complete deafness in patients with CAPS.

The color expression of anthocyanin pigments in blue flowers is precisely controlled by their chemical and physical properties such as pH and the presence of metal ions or colorless copigments. Despite the large number of known blue flowers, their coloration mechanisms have not been examined in sufficient detail. In this work, the blue coloration of Viola cornuta petals was expressed via the copigmentation of various flavonol 3- O-glycosides. By using a combination of imaging mass spectrometry with liquid chromatography mass spectrometry, the structures and contents of flavonols colocalized with violanin in the discrete blue-colored regions of the petal were identified. The obtained data allowed the in vitro reconstruction of the color expression that was consistent with the visible spectrum of the viola petal. The results of visible spectral analysis indicated that neither the increase in the solution pH inside the vacuole cells nor the presence of metal ions affected the color development process. Ultimately, it was experimentally confirmed that the excess amounts of flavonol 3- O-glycosides complexed with violanin, which prevented violanin molecules from forming a levorotatory helical self-assembly during the blue color expression via copigmentation.

Reliable detection and typing of carbapenemase is important in the treatment of infectious diseases. In this study we newly designed LAMP primer based on the latest information, and established a detection method for Carbapenemase Big five gene. For DNA extraction from strains, alkaline boiling method and commercial kit were used. The reaction temperatures of the LAMP method was VIM: 65°C, NDM: 63°C, KPC: 65°C, OXA-48-like: 65°C, IMP: 61°C. And simultaneous LAMP method was at 63°C, for 60 min. It was possible to detect up to 10 3 copies/ml. The reactivity of LAMP using 36 strains verified by Multiplex-PCR was VIM (4/4: number of LAMP method positive strains/number of strains evaluated), NDM (2/2), KPC (4/4), OXA-48-like (4/4), IMP (17/17). The type of carbapenemase determined by the LAMP method were all consistent with multiplex PCR. All strains were detected within 30 min. In VIM, both VIM-1-like and VIM-2-like were able to detect. In this study, although the number and variation of the strains evaluated was limited, LAMP method was clinically useful as a simple and rapid carbapenemase detection method.

Background: Due to advances in medicine, patients with pulmonary diseases have become candidates for surgery under general anesthesia. They often consult pulmonologists to assess their tolerability for surgery. The purpose of this study was to evaluate the significant characteristics responsible for postoperative pulmonary complications (PPCs) and the preclusion of the planned surgery., Methods: The clinical data of 462 consecutive patients who consulted at the Department of Respiratory Medicine before surgery under general anesthesia were used in this study. The relationship between the patient׳s characteristics and their outcomes were analyzed. The patients who were scheduled for lung resection were excluded., Results: Of the 386 patients who underwent planned surgery, 353 had no PPCs (Group A) and 33 developed PPCs (Group B). Planned surgery under general anesthesia was precluded in 31 patients due to respiratory problems (Group C). The significant predictors for PPCs consisted of a higher age, male gender, asthma, gastrointestinal surgery, cardiovascular surgery and a lower percentage of the predicted forced expiratory volume in 1 second (% predicted FEV 1 ). The significant factors associated with the preclusion of planned surgery included interstitial pneumonia (IP), dermatologic surgery and a lower % predicted FEV 1 . The predicted probability of PPCs in Group C was significantly higher than that in Group A and lower than that in Group B (all p-values < 0.05)., Conclusion: The common clinical finding for predicting PPCs and encouraging the preclusion of the planned surgery under general anesthesia was a lower % predicted FEV 1 ., (Copyright © 2018 The Japanese Respiratory Society. Published by Elsevier B.V. All rights reserved.)

The brilliant blue color of the Nemophila menziesii flower is derived from metalloanthocyanin, which consists of anthocyanin {petunidin 3-O-[6-O-(trans-p-coumaroyl)-β-glucoside]-5-O-[6-O-(malonyl)-β-glucoside]}, flavone [apigenin 7-O-β-glucoside-4'-O-(6-O-malonyl)-O-β-glucoside] and metal ions (Mg2+, Fe3+). Although the two glucosyl moieties at the apigenin 7-O and 4'-O positions are essential for metalloanthocyanin formation, the mechanism of glucosylation has not yet been clarified. In this study, we used crude protein extract prepared from N. menziesii petals to determine that apigenin is sequentially glucosylated by the catalysis of UDP-glucose:flavone 4'-O-glucosyltrasferase (F4'GT) and UDP-glucose:flavone 4'-O-glucoside 7-O-glucosyltransferase (F4'G7GT). We identified 150 contigs exhibiting homology with a UDP-glucose-dependent GT in the N. menziesii petal transcriptome and isolated 24 putative full-length GT cDNAs which were then subjected to functional analysis. Two GT cDNAs, NmF4'GT and NmF4'G7GT, which are highly expressed during the early stages of petal development and rarely in leaves, were shown to encode F4'GT and F4'G7GT activities, respectively. Biochemical characterization of the recombinant enzymes revealed that NmF4'GT specifically catalyzed 4'-glucosylation of flavonoids and that NmF4'G7GT specifically catalyzed 7-glucosylation of flavone 4'-O-glucosides and flavones. Apigenin 7,4'-O-diglucoside was efficiently synthesized from apigenin in the presence of recombinant NmF4'GT and NmF4'G7GT. Transgenic tobacco BY-2 cells expressing NmF4'GT and NmF4'G7GT converted apigenin into apigenin 7,4'-O-diglucoside, confirming their activities in vivo. Based on these results, we conclude that these two GTs act co-ordinately to catalyze apigenin 7,4'-O-diglucoside biosynthesis in N. menziesii.

We have been working on developing a low-cost tabletop NMR system. We reported that a field homogeneity as high as 50 ppm was achieved with a simple NMR magnet by employing two facing ferrite magnets with iron disks in between (Chonlathep et al., 2017). In this paper, we report two improvements added to our previous system: (1) an FPGA based signal processing unit to improve the S/N ratio and (2) a simple shimming mechanism to improve the field homogeneity. We obtained as high as 1 ppm field homogeneity in the best case. The signals from hydrogen nuclear spins in a methyl and carboxy group in acetic acid were resolved in NMR spectra., (Copyright © 2018 Elsevier Inc. All rights reserved.)