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Abstract Introduction Complexities of robotic distal gastrectomy (RDG) give reason to assess physician's surgical skill. Varying levels in surgical skill affect patient outcomes. We aim to investigate how a novel artificial intelligence (AI) model can be used to evaluate surgical skill in RDG by recognizing surgical instruments. Methods Fifty‐five consecutive robotic surgical videos of RDG for gastric cancer were analyzed. We used Deeplab, a multi‐stage temporal convolutional network, and it trained on 1234 manually annotated images. The model was then tested on 149 annotated images for accuracy. Deep learning metrics such as Intersection over Union (IoU) and accuracy were assessed, and the comparison between experienced and non‐experienced surgeons based on usage of instruments during infrapyloric lymph node dissection was performed. Results We annotated 540 Cadiere forceps, 898 Fenestrated bipolars, 359 Suction tubes, 307 Maryland bipolars, 688 Harmonic scalpels, 400 Staplers, and 59 Large clips. The average IoU and accuracy were 0.82 ± 0.12 and 87.2 ± 11.9% respectively. Moreover, the percentage of each instrument's usage to overall infrapyloric lymphadenectomy duration predicted by AI were compared. The use of Stapler and Large clip were significantly shorter in the experienced group compared to the non‐experienced group. Conclusions This study is the first to report that surgical skill can be successfully and accurately determined by an AI model for RDG. Our AI gives us a way to recognize and automatically generate instance segmentation of the surgical instruments present in this procedure. Use of this technology allows unbiased, more accessible RDG surgical skill.

ABSTRACT Bone and soft tissue infections caused by biofilm-forming bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA), remain a significant clinical challenge. While the control of local infection is necessary, systemic treatment is also required, and biofilm eradication is a critical target for successful management. Topical antibiotic treatments, such as antibiotic-loaded bone cement (ALBC), have been used for some time, and continuous local antibiotic perfusion therapy, a less invasive method, has been developed by our group. However, the optimal antibiotics and concentrations for biofilms of clinical isolates are still not well understood. We examined the efficacy of high concentrations of gentamicin against MRSA biofilms and the role of gentamicin resistance genes in biofilm eradication. We collected 101 MRSA samples from a hospital in Japan and analyzed their gene properties, including methicillin and gentamicin resistance, and their minimum biofilm eradication concentration (MBEC) values. Our results showed that high concentrations of gentamicin are effective against MRSA biofilms and that even concentrations lower than the MBEC value could eliminate biofilms after prolonged exposure. We also identified three aminoglycoside/gentamicin resistance genes [aac(6′)-aph(2″), aph(3′)-III, and ant(4′)-IA] and found that the presence or absence of these genes may inform the selection of treatments. It was also found that possession of the aac(6′)-aph(2″) gene correlated with the minimum inhibitory concentration/MBEC values of gentamicin. Although this study provides insight into the efficacy of gentamicin against MRSA biofilms and the role of gentamicin resistance genes, careful selection of the optimal treatment strategy is needed for clinical application.IMPORTANCEOur analysis of 101 MRSA clinical isolates has provided valuable insights that could enhance treatment selection for biofilm infections in orthopedics. We found that high concentrations of gentamicin were effective against MRSA biofilms, and even prolonged exposure to concentrations lower than the minimum biofilm eradication concentration (MBEC) value could eliminate biofilms. The presence of the aac(6′)-aph(2″) gene, an aminoglycoside resistance gene, was found to correlate with the minimum inhibitory concentration (MIC) and MBEC values of gentamicin, providing a potential predictive tool for treatment susceptibility. These results suggest that extended high concentrations of local gentamicin treatment could effectively eliminate MRSA biofilms in orthopedic infections. Furthermore, testing for gentamicin MIC or the possession of the aac(6′)-aph(2″) gene could help select treatment, including topical gentamicin administration and surgical debridement.

ABSTRACT “The infectious organism lurking in human airways (IOLA)” is a candidate pathogenic bacterium detected in bronchoalveolar lavage fluid specimens derived from adult patients with chronic lower respiratory tract infections. Genomic analyses of IOLA have revealed that it possesses the smallest and most AT-rich genome among human-derived bacteria. However, its biological properties remain unclear because no culture method has been established for IOLA. Here, we conducted a large-scale IOLA surveillance study of nasal discharge specimens from children in Japan and investigated the correlation between IOLA detection frequency and patient characteristics. We detected IOLA in 5.4% (103 of 1,920) of pediatric nasal discharge samples. No significant differences were observed in the frequency of detection based on the patient’s background. However, with respect to age, the frequency of detection tended to be significantly high at 2–3 and 6 years old. Phylogenetic analysis revealed five phylotypes in the IOLA 16S rRNA gene sequences, and the sequences detected in adult patients with respiratory infections in a previous study belonged to one of the five phylotypes. The involvement of IOLA in the symptoms is not clear, but IOLA is detected at a relatively high frequency in pediatric nasal discharge. Many subjects with detected IOLA were not always IOLA positive, and IOLA was detected transiently. Our findings suggest that IOLA is horizontally transmitted through groups in nursery and elementary schools, and there are differences in biological characteristics among the IOLA phylotypes.IMPORTANCE“The infectious organism lurking in human airways (IOLA)” is a candidate pathogenic bacterium strongly suspected to be infectious to the respiratory tracts of humans and animals. However, a culture method for IOLA has not been established yet, and its properties remain unclear. In this study, IOLA was detected at a relatively high frequency in the nasal discharge of children, and five phylotypes of IOLA were identified. One of these phylotypes was found in the bronchoalveolar lavage fluid from adult patients, suggesting lineage-specific differences in the pathogenicity of IOLA. Moreover, it was suggested that IOLA is horizontally transmitted when children gather in groups such as nursery and elementary schools. These findings strongly indicate that IOLAs have been clinically undetected so far but are spreading among children, with one lineage being involved in respiratory diseases in adults. Examining the presence of IOLA in clinical specimens may help to understand the etiology of respiratory diseases with unknown causes.

Patients with pancreatic neuroendocrine tumors (pNETs) have limited access to effective targeted agents and invariably succumb to progressive disease. MUC1-C is a druggable oncogenic protein linked to driving pan-cancers. There is no known involvement of MUC1-C in pNET progression. The present work was performed to determine if MUC1-C represents a potential target for advancing pNET treatment. We demonstrate that the MUC1 gene is upregulated in primary pNETs that progress with metastatic disease. In pNET cells, MUC1-C drives E2F- and MYC-signaling pathways necessary for survival. Targeting MUC1-C genetically and pharmacologically also inhibits self-renewal capacity and tumorigenicity. Studies of primary pNET tissues further demonstrate that MUC1-C expression is associated with (i) an advanced NET grade and pathological stage, (ii) metastatic disease, and (iii) decreased disease-free survival. These findings demonstrate that MUC1-C is necessary for pNET progression and is a novel target for treating these rare cancers with anti-MUC1-C agents under clinical development.

Abstract The oral microbiota associated with mucosal diseases, including oral squamous cell carcinoma and oral potentially malignant disorders, have been extensively analyzed at the phylum and genus levels. However, the details of the oral microbiota remain unclear at the species and operational taxonomic unit (OTU) levels. We aimed to determine differences in the microbiota of oral rinse, lesion and normal site swab samples of patients with mucosal abnormalities on the tongues. Oral samples were obtained from 10 patients with oral mucosal abnormalities. Alpha and beta diversity at the OTU and genus levels of the microbiota samples were analyzed using OTUs clustered with 99.6% similarity based on 16S rRNA gene sequences obtained using the Sanger method. At the OTU level, the microbiota of the lesions were the least diverse but were different from those of the normal site and oral rinse samples. The OTUs corresponding to Streptococcus infantis and Haemophilus parainfluenzae were suggested to contribute to the differences between the microbiota of the lesions and normal sites. At the genus level, no significant differences between these microbiota were observed. In conclusion, strict OTU-level microbiota analysis might be able to discriminate lesions from normal sites of patients with mucosal abnormalities.

Abstract Background C–C chemokine receptor type 7 (CCR7) participates in chemotactic and metastatic responses in various cancers, including in esophageal squamous cell carcinoma (ESCC). The microRNA (miRNA) let-7a suppresses migration and invasion of various types of cancer cells by downregulating CCR7 expression. Methods The expression levels of CCR7 and let-7a were measured in the cell lines, tumor, and peritumoral tissues of ESCC patients. KYSE cell lines were transfected with synthetic let-7a miRNA and a let-7a miRNA inhibitor, and their CCR7 expression levels as well as invasive ability were evaluated. A highly invasive cell line was established via an invasion assay, and CCR7 expression level along with let-7a level was subsequently evaluated. Cancer cells overexpressing CCR7 were injected subcutaneously into mice, and the animals were monitored for tumor growth along with lymph node metastasis. Results A negative correlation between CCR7 and let-7a expression was observed in the ESCC cell lines as well as in tissue samples from patients. Synthetic let-7a decreased CCR7 expression level, while the let-7a inhibitor increased it. In vitro, the established highly invasive cancer cells with high and low levels of CCR7 and let-7a expression, respectively, exhibited a greater invasive ability than the wild-type cell line. The cells were associated with tumor growth and lymph node metastasis in mice. Patients in the high-CCR7/low-let-7a group had the worst prognosis, with a five-year recurrence free survival (5-RFS) rate of 37.5%, followed by the high-CCR7/high-let-7a (5-RFS: 60.0%) and low-CCR7 (5-RFS: 85.7%; p = 0.038) groups. Conclusions The expression of CCR7 was downregulated by let-7a miRNA in esophageal cancer cells. The decrease in let-7a expression level led to the increased expression level of CCR7 in ESCC cells, consequently increasing their invasive ability and malignancy and resulting in a worse prognosis for ESCC patients. Trial registration. Retrospectively registered.

Abstract Next-generation sequencing (NGS) technologies have been applied in bacterial flora analysis. However, there is no standardized protocol, and the optimal clustering threshold for estimating bacterial species in respiratory infection specimens is unknown. This study was conducted to investigate the optimal threshold for clustering 16S ribosomal RNA gene sequences into operational taxonomic units (OTUs) by comparing the results of NGS technology with those of the Sanger method, which has a higher accuracy of sequence per single read than NGS technology. This study included 45 patients with pneumonia with aspiration risks and 35 patients with lung abscess. Compared to Sanger method, the concordance rates of NGS technology (clustered at 100%, 99%, and 97% homology) with the predominant phylotype were 78.8%, 71.3%, and 65.0%, respectively. With respect to the specimens dominated by the Streptococcus mitis group, containing several important causative agents of pneumonia, Bray Curtis dissimilarity revealed that the OTUs obtained at 100% clustering threshold (versus those obtained at 99% and 97% thresholds; medians of 0.35, 0.69, and 0.71, respectively) were more similar to those obtained by the Sanger method, with statistical significance (p

Sarcoidosis is a systemic inflammatory disease characterized by noncaseating epithelioid cell granulomas. However, certain infections can exhibit similar histological findings. We present a case of a 69-year-old man who was initially diagnosed with sarcoidosis and later was confirmed, through 16S rRNA sequencing, to have disseminated Mycobacterium genavense infection. Acid-fast bacteria were detected in the bone marrow biopsy using Ziehl–Neelsen staining, but routine clinical tests did not provide a definitive diagnosis. The patient tested negative for HIV, anti-interferon-gamma antibodies, and genetic immunodeficiency disorders. He was treated with multiple drugs, including aminoglycosides and macrolides, but showed no improvement in fever and pancytopenia. However, these clinical signs responded favorably to steroid therapy. We reviewed 17 Japanese cases of M. genavense infection. All cases were in males; 7/17 (41%) were HIV-negative; and 12/17 (71%) had a decreased CD4 count. Genetic analysis confirmed M. genavense isolation, and macrolides were used universally. Mycobacterium genavense infection is challenging to identify and mimics other systemic inflammatory diseases such as sarcoidosis. There are no standard treatment protocols. Our case report and Japanese case review contribute to understanding this rare disease.

Abstract Aim We have previously reported the existence of lymph nodes surrounding the thoracic duct ( TDLN) and transthoracic esophagectomy (TTE) with thoracic duct (TD) resection increased the number of lymph nodes (LNs) retrieved. The current study aims to evaluate the prognostic impact of TDLN metastasis in esophageal cancer patients subdivided by its location and comparing the patients’ survival with those with extra‐regional LN metastasis. Methods Patients who underwent TTE with TD resection for esophageal squamous cell carcinoma (ESCC) were reviewed. Patients were classified into those with or without TDLN metastasis, and clinicopathological factors were compared between groups. TDLN was further divided into TDLN‐Ut/Mt/Lt based on the location in the mediastinum. The relapse‐free survival (RFS) and overall survival (OS) were compared between groups. Results Of 232 patients, TDLN metastasis was observed in 17 (7%). RFS and OS were significantly worse in the TDLN metastasis group. TDLN metastasis was shown to be an independent prognostic factor for RFS and OS in the multivariate analysis. The negative prognostic impact of TDLN metastasis was evident in TDLN‐Mt/Lt. The RFS and OS of patients with TDLN metastasis were almost identical to those with positive LN metastasis in extra‐regional LNs. Conclusion TDLN metastasis was proven to be a strong prognostic indicator. Although the TDLN has been included in the classification of regional LN in the current staging systems, it could be independently classified from the current regional LNs. Given that neoadjuvant therapy has been a standard, we might need to introduce adjuvant therapy when TDLN metastasis is observed.

Kazumasa Fukuda et al. complete a new genome sequence for an uncultured bacterium detected in human respiratory tract named IOLA. The IOLA genome is found to be among the smallest and most AT-rich of known human-associated bacterial genomes and surveillance of clinical samples indicates that IOLA is in fact a human respiratory tract-associated bacterium.

Mesenchymal stromal/stem cells (MSCs) have shown potential in the treatment of degenerative diseases, including ischemia/reperfusion injury (IRI), which occurs during organ transplantation and represents the main cause of post-transplant graft dysfunction. However, MSCs have heterogeneous characteristics, and studies of MSCs therapy have shown a variety of outcomes. To establish a new effective MSCs therapy, we developed an adipose-derived mesenchymal stromal/stem cell line (ASCL) and compared its therapeutic effects on primary adipose-derived MSCs (ASCs) using a hepatocyte co-culture model of hypoxia/reoxygenation in vitro and a rat model of hepatic IRI in vivo . The results showed that both ASCL and ASCs protect against hypoxia by improving hepatocyte viability, inhibiting reactive oxygen species release, and upregulating transforming growth factor-β in vitro. In vivo , ASCL or ASCs were infused into the spleen 24 h before the induction of rat hepatic IRI. The results showed that ASCL significantly improved the survival outcomes compared with the control (normal saline infusion) with the significantly decreased serum levels of liver enzymes and less damage to liver tissues compared with ASCs. Both ASCL and ASCs suppressed NOD-like receptor family pyrin domain-containing 3 inflammasome activation and subsequently reduced the release of activated IL-1β and IL-18, which is considered an important mechanism underlying ASCL and ASCs infusion in hepatic IRI. In addition, ASCL can promote the release of interleukin-1 receptor antagonist, which was previously reported as a key factor in hampering the inflammatory cascade during hepatic IRI. Our results suggest ASCL as a new candidate for hepatic IRI treatment due to its relatively homogeneous characteristics.

Objectives: The aim of this study is to determine the optimal duration of antimicrobial therapy for lower respiratory tract infection (LRTI) in neuromuscular disorder (NMD) patients. Methods: This prospective study included 13 episodes from 9 NMD patients hospitalized for bacterial LRTI. Sputum samples were collected from these patients during the three consecutive days after their admission. Bacterial cell counts and the proportion of the most predominant bacterium identified by a clone library analysis of the bacterial 16S rRNA gene sequence in the samples obtained before antimicrobial therapy were serially investigated. Results: All episodes were initially treated with ampicillin/sulbactam. In 12 episodes with a therapeutic effect, the bacterial cell counts in the samples obtained on the third day of therapy were significantly lower than those before therapy (P = 0.0013). In most of these episodes, the most predominant bacterium in the sample obtained before therapy was undetectable by the third day of therapy. In the one patient without a therapeutic effect, neither the bacterial cell counts nor the proportion of the most predominant bacterium in the sample obtained before therapy decrease after therapy. Conclusion: Short-term antimicrobial therapy is sufficiently effective for LRTI in NMD patients if the initial therapy is effective.

Abstract Background Gastric cancer that mimics a submucosal tumor (SMT) is infrequently encountered in routine clinical settings, and histopathological analysis is often negative for malignant cells. In such cases, excisional biopsy of the entire tumor may be necessary to make a definitive pathological diagnosis, and laparoscopic and endoscopic cooperative surgery (LECS) is a viable method of excisional biopsy. Case presentation An 80-year-old male patient diagnosed with stomach wall irregularities at routine medical check-up was referred to our facility, and consequent endoscopic examination detected a 20-mm protruded lesion in the greater curvature at the middle third of the stomach. Endoscopic ultrasound (EUS) showed a thick, low echoic lesion with an irregular margin in the second layer of the gastric wall. Further, a nodular part of the lesion had infiltrated into the submucosa, with an appearance similar to that of linitis plastica of the stomach. The lesion was highly suspected to be a gastric carcinoma with submucosal invasion. However, mucosal-incision-assisted biopsy revealed no malignant cells. Computed tomography (CT) identified no metastatic lymph nodes. Therefore, an excisional biopsy using LECS was performed, and to avoid peritoneal dissemination, we used a modified version, namely, combination of laparoscopic and endoscopic approaches to neoplasia with non-exposure technique (CLEAN-NET). The procedure ended without any complications, and post-operative course was uneventful. As histopathology returned a diagnosis of adenocarcinoma pT4a, we performed radical gastrectomy and D2 lymphadenectomy. Post-operative course was unremarkable and the patient underwent follow-up examinations without adjuvant chemotherapy because of old age. Conclusions Local resection using LECS for gastric tumors with a high suspicion of malignancy is useful and feasible. LECS could be used in similar cases.

Intraperitoneal (i.p.) tumor dissemination and the consequent malignant ascites remain unpredictable and incurable in patients with gastrointestinal (GI) cancer, and practical advances in diagnosis and treatment are urgently needed in the clinical settings. Here, we explored tumor biological and immunological mechanisms underlying the i.p. tumor progression for establishing more effective treatments.We established mouse tumor ascites models that murine and human colorectal cancer cells were both i.p. and subcutaneously (s.c.) implanted in mice, and analyzed peritoneal exudate cells (PECs) obtained from the mice. We then evaluated anti-tumor efficacy of agents targeting the identified molecular mechanisms using the ascites models. Furthermore, we validated the clinical relevancy of the findings using peritoneal lavage fluids obtained from gastric cancer patients.I.p. tumor cells were giant with large nuclei, and highly express AURKA, but less phosphorylated TP53, as compared to s.c. tumor cells, suggesting polyploidy-like cells. The i.p. tumors impaired phagocytic activity and the consequent T-cell stimulatory activity of CD11b+Gr1+PD1+ myeloid cells by GDF15 that is regulated by AURKA, leading to treatment resistance. Blocking AURKA with MLN8237 or siRNAs, however, abrogated the adverse events, and induced potent anti-tumor immunity in the ascites models. This treatment synergized with anti-PD1 therapy. The CD11b+PD1+ TAMs are also markedly expanded in the PECs of gastric cancer patients.These suggest AURKA is a determinant of treatment resistance of the i.p. tumors. Targeting the AURKA-GDF15 axis could be a promising strategy for improving clinical outcome in the treatment of GI cancer.

Abstract Background The emergence of multi-drug resistant pathogens is an urgent health-related problem, and the appropriate use of antibiotics is imperative. It is often difficult to identify the causative bacteria in patients with aspiration pneumonia because tracheal aspirate contains contaminants of oral bacteria. We investigated the dynamics of microbiota in mechanically ventilated patients with aspiration pneumonia to develop a treatment strategy. Methods Twenty-two intubated patients with aspiration pneumonia were recruited. Saliva and tracheal aspirate of the subjects were collected at three time points: (A) within 2 h after intubation, (B) just before administration of antibiotics, and (C) 48-72 h after administration of antibiotics. The microbiota in each specimen was analyzed by using the 16S rRNA gene clone library sequencing method. Bacterial floras of the samples were analyzed by principal component analysis. Results Principal component analysis based on the composition of genus revealed that although the changes of microbiota in the saliva from (A) to (B) were not clear, the composition of anaerobes in the tracheal aspirate (B) was lower than (A). In fact, the reduction of anaerobes, not in the saliva but in the tracheal aspirate from (A) to (B), was confirmed by incident rate ratios estimated by a multilevel Poisson regression model (p

We performed gene amplification methods for the detections of bacteria and viruses using sputum samples to clarify the microbiological characteristics of lower respiratory tract infection in patients with neuromuscular disorders. The tendencies of higher proportion of respiratory virus detection and lower diversity of bacteria in sputum were observed. Keywords: Neuromuscular disorders, Lower respiratory tract infection, Respiratory viruses, 16S rRNA gene, Microbiome in respiratory tract

BackgroundLeptospirosis has been described as a biphasic disease consisting of hematogenous dissemination to major organs in the acute phase and asymptomatic renal colonization in the chronic phase. Several observational studies have suggested an association between leptospirosis and chronic kidney disease (CKD). We investigated the dynamics of leptospires and histopathological changes in the kidney to understand the relationship between them, and also investigated the extent of renal dysfunction in the acute and chronic phases of leptospirosis using a hamster model.FindingsHamsters (n = 68) were subcutaneously infected with 1 × 104 cells of the Leptospira interrogans serovar Manilae strain UP-MMC-SM. A total of 53 infected hamsters developed fatal acute leptospirosis, and the remaining 15 hamsters recovered from the acute phase, 13 of which showed Leptospira colonization in the kidneys in the chronic phase. Five asymptomatic hamsters also had renal colonization in the chronic phase. Immunofluorescence staining showed that leptospires were locally distributed in the renal interstitium in the early acute phase and then spread continuously into the surrounding interstitium. The kidneys of the surviving hamsters in the chronic phase showed patchy lesions of atrophic tubules, a finding of chronic tubulointerstitial nephritis, which were substantially consistent with the distribution of leptospires in the renal interstitium. The degree of atrophic tubules in kidney sections correlated statistically with the serum creatinine level in the chronic phase (rs = 0.78, p = 0.01).ConclusionSubcutaneous infection with pathogenic leptospires could cause acute death or chronic leptospirosis in hamsters after surviving the acute phase. We suggest that the renal distribution of leptospires during the acute phase probably affected the extent of tubular atrophy, leading to CKD.

We report a patient with highly advanced gastric carcinoma who was treated successfully with chemoradiotherapy (CRT) comprising S-1 and cisplatin. The patient was a 71-year-old male who was diagnosed with advanced gastric carcinoma by esophagogastroduodenoscopy (EGD) by medical examination. EGD demonstrated type 3 advanced gastric carcinoma in the posterior wall of the upper gastric body. An abdominal computed tomography (CT) scan showed that the gastric wall was thickened due to gastric primary tumor, and large lymph nodes (LNs) including the lesser curvature LN, anterosuperior LN along the common hepatic artery and some para-aortic LNs were detected. The patient was diagnosed with stage IV advanced gastric carcinoma according to the Japanese classification of gastric carcinoma (cT4a, cN3, cM1 [para-aortic LN], cStage IV). Preoperative CRT was carried out in an attempt to downstage the disease. Remarkable reduction of the primary tumor and metastatic LNs was observed after initial CRT, and radiological examination determined that a partial response had been achieved. Adverse effects included grade 2 anorexia and grade 3 ALP elevation (919 U/ml). No grade 4 or more severe adverse event was observed. After CRT, although we recommended curative surgery, the patient refused surgical treatment and opted for conservative treatment. Thus, we continued S-1 oral administration for 1 year. Five months after beginning CRT, upper endoscopy showed that the tumor had maintained regression and scar formation, in which no cancer cells were detected by endoscopic biopsy. The patient is doing well and has maintained a clinical complete response for more than 42 months without curative surgery. CRT could be considered as an option for treatment of patients with locally advanced gastric carcinoma diagnosed as unresectable, or for those who refuse surgical treatment.

Tumor draining lymph nodes (TDLNs) are located in the routes of lymphatic drainage from a primary tumor and have the highest risk of metastasis in various types of solid tumors. TDLNs are also considered as a tissue to activate the antitumor immunity, where antigen-specific effector T cells are generated. However, T cell receptor (TCR) repertoires in TDLNs have not been well characterized. We collected 23 colorectal cancer tumors with 203 lymph nodes with/without metastatic cancer cells (67 were metastasis-positive and the remaining 136 were metastasis-negative) and performed TCR sequencing. Metastasis-positive TDLNs showed a significantly lower TCR diversity and shared TCR clonotypes more frequently with primary tumor tissues compared to metastasis-negative TDLNs. Principal component analysis indicated that TDLNs with metastasis showed similar TCR repertoires. These findings suggest that cancer-reactive T cell clones could be enriched in the metastasis-positive TDLNs.

We previously observed an inverse correlation between stroke and urinary 3-hydroxypropyl mercapturic acid (3-HPMA), an acrolein-glutathione metabolite, through its measurement by liquid chromatography with tandem mass spectrometry (LC-MS/MS). However, the cost of equipment for LC-MS/MS and its maintenance fee is very expensive and a cost-efficient method is required. In this study, we have developed a sensitive enzyme-linked immunosorbent assay (ELISA) system to measure 3-HPMA using a chicken antibody recognizing 3-HPMA-conjugated chicken albumin as antigen. Linearity to measure 3-HPMA was obtained from 0 to 10 μM, indicating that this ELISA system is useful for measurement of urine 3-HPMA. It was confirmed that 3-HPMA in urine of stroke patients decreased significantly compared with that of control subjects using the ELISA system. Using the ELISA kit, it became possible to evaluate the risk of brain stroke by not only plasma but also by urine. These results confirm that shortage of glutathione to detoxify acrolein is one of the major causes of stroke incidence. Our method contributes to maintenance of quality of life (QOL) of the elderly.

Corynebacterium ulcerans infection is emerging in humans. We conducted phylogenetic analyses of C. ulcerans and C. diptheriae, which revealed diverse diphtheria toxin in C. ulcerans. Diphtheria toxin diversification could decrease effectiveness of diphtheria toxoid vaccine and diphtheria antitoxin for preventing and treating illnesses caused by this bacterium.