Relationship between mucin expression and preoperative bile juice cytology in biliary tract carcinoma

The present study evaluated correlations between preoperative bile juice cytology and mucin expression of surgical specimens in biliary tract carcinoma. Twenty-five patients with biliary tract carcinoma surgically treated at our hospital, whose bile juice cytology had been evaluated before operation, were allocated to this study. Biliary cytology was classified into three categories based on the Papanicolaou classification. Immunohistochemical staining of tissues was performed using MUC1 and MUC2 monoclonal antibodies. Lesions showing MUC1 expression of ＋＋ or higher and MUC2 expression of - were classified as Group A, and the remaining lesions as Group B. According to the epithelial site, preoperative cytology was highly correlated in Group A, while it was negative in Group B (p<0.05). In the advanced site of carcinomas, preoperative cytology tended to highly be positive in Group A, while it tended to be negative in Group B (p<0.05). These results suggest that the bile juice cytology results are affected by characteristics of mucin expression in the tissue. Based on the possibility that mucin expression correlates with the prognosis of each carcinoma, a positive cytological result suggests a poor prognosis for the carcinoma, which may be informative for predicting the post-operative courses and choosing treatments

Assay for Glycosaminoglycans by Tandem Mass Spectrometry and its Applications

Glycosaminoglycans (GAGs) are distributed in the whole body and play a variety of important physiological roles associated with inflammation, growth, coagulation, fibrinolysis, lipolysis, and cell-matrix biology. Accumulation of undegraded GAGs in lysosomes gives rise to a distinct clinical syndrome, mucopolysaccharidoses. Measurement of each specific GAG in a variety of specimens is urgently required to understand GAG interaction with other molecules, physiological status of patients, and prognosis and pathogenesis of the disease. We established a highly sensitive and accurate tandem mass spectrometry (LC-MS/MS) method for measurements of disaccharides derived from four specific GAGs [dermatan sulfate (DS), heparan sulfate (HS), keratan sulfate (KS), and chondroitin sulfate (CS)]. Disaccharides were produced by specific enzyme digestion of each GAG, and quantified by negative ion mode of multiple reaction monitoring. Subclasses of HS and GAGs with identical molecular weights can be separated using a Hypercarbcolumn (2.0 mm×50 mm, 5 μm) with an aectonitrile gradient in ammonium acetate (pH 11.0). We also developed a GAG assay by RapidFire with tandem mass spectrometry (RF-MS/MS). The RF system consists of an integrated solid phase extraction robot that binds and de-salts samples from assay plates and directly injects them into a MS/MS detector, reducing sample processing time to ten seconds. RF-MS/MS consequently yields much faster throughput than conventional LC-MS/MS-based methods. However, the RF system does not have a chromatographic step, and therefore, cannot distinguish GAGs that have identical molecular weights. Both methods can be applied to analysis of dried blood spots, blood, and urine specimens. In this article, we compare the assay methods for GAGs and describe their potential applications

Establishment of glycosaminoglycan assays for mucopolysaccharidoses

Mucopolysaccharidoses (MPS) are a group of lysosomal storage disorders caused by deficiency of the lysosomal enzymes essential for catabolism of glycosaminoglycans (GAGs). Accumulation of undegraded GAGs results in dysfunction of multiple organs, resulting in distinct clinical manifestations. A range of methods have been developed to measure specific GAGs in various human samples to investigate diagnosis, prognosis, pathogenesis, GAG interaction with other molecules, and monitoring therapeutic efficacy. We established ELISA, liquid chromatography tandem mass spectrometry (LC-MS/MS), and an automated high-throughput mass spectrometry (HT-MS/MS) system (RapidFire) to identify epitopes (ELISA) or disaccharides (MS/MS) derived from different GAGs (dermatan sulfate, heparan sulfate, keratan sulfate, and/or chondroitin sulfate). These methods have a high sensitivity and specificity in GAG analysis, applicable to the analysis of blood, urine, tissues, and cells. ELISA is feasible, sensitive, and reproducible with the standard equipment. HT-MS/MS yields higher throughput than conventional LC-MS/MS-based methods while the HT-MS/MS system does not have a chromatographic step and cannot distinguish GAGs with identical molecular weights, leading to a limitation of measurements for some specific GAGs. Here we review the advantages and disadvantages of these methods for measuring GAG levels in biological specimens. We also describe an unexpected secondary elevation of keratan sulfate in patients with MPS that is an indirect consequence of disruption of catabolism of other GAGs

Risk assessments for broncho-pleural fistula and respiratory failure after lung cancer surgery by National Clinical Database Japan

BackgroundBroncho-pleural fistula (BPF) and respiratory failure (RF) are life-threatening complications after lung cancer surgery and can result in long-term hospitalization and decreased quality of life. Risk assessments for BPF and RF in addition to mortality and major morbidities are indispensable in surgical decision-making and perioperative care.MethodsThe characteristics and operative data of 80,095 patients who had undergone lung cancer surgery were derived from the 2014 and 2015 National Clinical Database (NCD) of Japan datasets. After excluding 1501 patients, risk models were developed from these data and validated by another dataset for 42,352 patients derived from the 2016 NCD dataset. Receiver operating characteristic curves were generated for postoperative BPF and RF development. The concordance-index was used to assess the discriminatory ability and validity of the model.ResultsBPF and RF occurred in 259 (0.3%) and 420 patients (0.5%), respectively, in the model development dataset and in 129 (0.3%) and 198 patients (0.5%), respectively, in the model validation dataset. Characteristic variables including types of surgery and comorbidities were identified as risk factors for BPF and RF, respectively. The concordance indexes of assessments for BPF and RF were 0.847 (p < 0.001) and 0.848 (p < 0.001), respectively, for the development dataset and 0.850 (p < 0.001) and 0.844 (p < 0.001), respectively, for the validation dataset.ConclusionsThese models are satisfactory for predicting BPF and RF after lung cancer surgery in Japan and could guide preoperative assessment and optimal measures for preventing BPF and RF

Time-resolved serial femtosecond crystallography reveals early structural changes in channelrhodopsin

X線自由電子レーザーを用いて、光照射によるチャネルロドプシンの構造変化の過程を捉えることに成功. 京都大学プレスリリース. 2021-03-26.Channelrhodopsins (ChRs) are microbial light-gated ion channels utilized in optogenetics to control neural activity with light . Light absorption causes retinal chromophore isomerization and subsequent protein conformational changes visualized as optically distinguished intermediates, coupled with channel opening and closing. However, the detailed molecular events underlying channel gating remain unknown. We performed time-resolved serial femtosecond crystallographic analyses of ChR by using an X-ray free electron laser, which revealed conformational changes following photoactivation. The isomerized retinal adopts a twisted conformation and shifts toward the putative internal proton donor residues, consequently inducing an outward shift of TM3, as well as a local deformation in TM7. These early conformational changes in the pore-forming helices should be the triggers that lead to opening of the ion conducting pore

The whole blood transcriptional regulation landscape in 465 COVID-19 infected samples from Japan COVID-19 Task Force

「コロナ制圧タスクフォース」COVID-19患者由来の血液細胞における遺伝子発現の網羅的解析 --重症度に応じた遺伝子発現の変化には、ヒトゲノム配列の個人差が影響する--. 京都大学プレスリリース. 2022-08-23.Coronavirus disease 2019 (COVID-19) is a recently-emerged infectious disease that has caused millions of deaths, where comprehensive understanding of disease mechanisms is still unestablished. In particular, studies of gene expression dynamics and regulation landscape in COVID-19 infected individuals are limited. Here, we report on a thorough analysis of whole blood RNA-seq data from 465 genotyped samples from the Japan COVID-19 Task Force, including 359 severe and 106 non-severe COVID-19 cases. We discover 1169 putative causal expression quantitative trait loci (eQTLs) including 34 possible colocalizations with biobank fine-mapping results of hematopoietic traits in a Japanese population, 1549 putative causal splice QTLs (sQTLs; e.g. two independent sQTLs at TOR1AIP1), as well as biologically interpretable trans-eQTL examples (e.g., REST and STING1), all fine-mapped at single variant resolution. We perform differential gene expression analysis to elucidate 198 genes with increased expression in severe COVID-19 cases and enriched for innate immune-related functions. Finally, we evaluate the limited but non-zero effect of COVID-19 phenotype on eQTL discovery, and highlight the presence of COVID-19 severity-interaction eQTLs (ieQTLs; e.g., CLEC4C and MYBL2). Our study provides a comprehensive catalog of whole blood regulatory variants in Japanese, as well as a reference for transcriptional landscapes in response to COVID-19 infection