Regulators specifying cell fate activate cell cycle regulator genes to determine cell numbers in ascidian larval tissues.

In animal development, most cell types stop dividing before terminal differentiation; thus, cell cycle control is tightly linked to cell differentiation programmes. In ascidian embryos, cell lineages do not vary among individuals, and rounds of the cell cycle are determined according to cell lineages. Notochord and muscle cells stop dividing after eight or nine rounds of cell division depending on their lineages. In the present study, we showed that a Cdk inhibitor, Cdkn1.b, is responsible for stopping cell cycle progression in these lineages. Cdkn1.b is also necessary for epidermal cells to stop dividing. In contrast, mesenchymal and endodermal cells continue to divide even after hatching, and Myc is responsible for maintaining cell cycle progression in these tissues. Expression of Cdkn1.b in notochord and muscle is controlled by transcription factors that specify the developmental fate of notochord and muscle. Likewise, expression of Myc in mesenchyme and endoderm is under control of transcription factors that specify the developmental fate of mesenchyme and endoderm. Thus, cell fate specification and cell cycle control are linked by these transcription factors.

Revealing Molecular-Scale Structural Changes in Polymer Nanocomposites during Thermo-Oxidative Degradation Using Evolved Gas Analysis with High-Resolution Time-of-Flight Mass Spectrometry Combined with Principal Component Analysis and Kendrick Mass Defect Analysis.

This study introduces a novel method that utilizes evolved gas analysis with time-of-flight mass spectrometry (EGA-TOFMS) coupled with principal component analysis (PCA) and Kendrick mass defect (KMD) analysis, called EGA-PCA-KMD, to analyze complex structural changes in polymer materials during thermo-oxidative degradation. While EGA-TOFMS captures exact mass data related to the degradation components in the temperature-dependent mass spectra of the evolved products, numerous high-resolution mass spectra with large amounts of ion signals and varying intensities provide challenges for interpretation. To address this, we employed mathematical decomposition through PCA to selectively extract information about the ion series specific to the products that evolved from the degradation components. Additionally, KMD analysis was applied to the attribution of the exact mass signals extracted from the PCA, which categorizes and visualizes depending on the molecular compositions in a two-dimensional plot. The complex structural changes of the triblock copolymer thermoplastic elastomer and its nanocomposites containing nanodiamonds during thermo-oxidative degradation were elucidated using EGA-PCA-KMD to demonstrate the effectiveness of this characterization technique for polymer degradation. Furthermore, it is revealed that the formation of rigid matrix-filler interfacial interaction via the π-π stacking and chemical bonds in the nanocomposites contributes to improvement in the stability toward thermo-oxidative degradation. Our results highlight the benefits of EGA-PCA-KMD and provide valuable insights into polymer degradation.

An Autopsy Case of Dextromethorphan Poisoning.

A woman in her 30s who was being treated for a mental illness with several psychotropic drugs was admitted to the hospital after being found in a state of unconsciousness and respiratory arrest at home. She was pronounced dead 12 hours after she was discovered. Her autopsy revealed symmetrical hemorrhagic necrosis in the putamen on both sides of her cerebrum. Although many drugs were detected in her blood, all of those other than dextromethorphan (DXM) were within or below the therapeutic range. Her blood DXM was 1.73 µg/ml at admission and 1.61 µg/ml at autopsy, which were within the toxic range or coma-to-death range. The cause of death was diagnosed as DXM poisoning. DXM can cause hallucinations and euphoria if taken in excess, but since it is available as an over-the-counter drug at general pharmacies, an increasing number of young people are overdosing on it, mistakenly believing it to be a safe drug with few side effects. We believe that further social measures against DXM are necessary in Japan, such as disseminating correct knowledge in society and regulating over-the-counter sales.

Overcoming cancer-associated fibroblast-induced immunosuppression by anti-interleukin-6 receptor antibody.

Cancer-associated fibroblasts (CAFs) are a critical component of the tumor microenvironment and play a central role in tumor progression. Previously, we reported that CAFs might induce tumor immunosuppression via interleukin-6 (IL-6) and promote tumor progression by blocking local IL-6 in the tumor microenvironment with neutralizing antibody. Here, we explore whether an anti-IL-6 receptor antibody could be used as systemic therapy to treat cancer, and further investigate the mechanisms by which IL-6 induces tumor immunosuppression. In clinical samples, IL-6 expression was significantly correlated with α-smooth muscle actin expression, and high IL-6 cases showed tumor immunosuppression. Multivariate analysis showed that IL-6 expression was an independent prognostic factor. In vitro, IL-6 contributed to cell proliferation and differentiation into CAFs. Moreover, IL-6 increased hypoxia-inducible factor 1α (HIF1α) expression and induced tumor immunosuppression by enhancing glucose uptake by cancer cells and competing for glucose with immune cells. MR16-1, a rodent analog of anti-IL-6 receptor antibody, overcame CAF-induced immunosuppression and suppressed tumor progression in immunocompetent murine cancer models by regulating HIF1α activation in vivo. The anti-IL-6 receptor antibody could be systemically employed to overcome tumor immunosuppression and improve patient survival with various cancers. Furthermore, the tumor immunosuppression was suggested to be induced by IL-6 via HIF1α activation.

Multi-enzyme Machinery for Chitin Degradation in the Chitinolytic Bacterium Chitiniphilus shinanonensis SAY3T.

The chitinolytic bacterium, Chitiniphilus shinanonensis SAY3T was examined to characterize its chitin-degrading enzymes in view of its potential to convert biomass chitin into useful saccharides. A survey of the whole-genome sequence revealed 49 putative genes encoding polypeptides that are thought to be related to chitin degradation. Based on an analysis of the relative quantity of each transcript and an assay for chitin-degrading activity of recombinant proteins, a chitin degradation system driven by 19 chitinolytic enzymes was proposed. These include sixteen endo-type chitinases, two N-acetylglucosaminidases, and one lipopolysaccharide monooxygenase that catalyzes the oxidative cleavage of glycosidic bonds. Among the 16 chitinases, ChiL was characterized by its remarkable transglycosylation activity. Of the two N-acetylglucosaminidases (ChiI and ChiT), ChiI was the major enzyme, corresponding to > 98% of the total cellular activity. Surprisingly, a chiI-disrupted mutant was still able to grow on medium with powdered chitin or GlcNAc dimer. However, its growth rate was slightly lower compared to that of the wild-type SAY3. This multi-enzyme machinery composed of various types of chitinolytic enzymes may support SAY3 to efficiently utilize native chitin as a carbon and energy source and may play a role in developing an enzymatic process to decompose and utilize abundant chitin at the industrial scale.

Accuracy of the Nova StatStrip® glucometer in patients undergoing major abdominal surgery: an observational study.

PURPOSE: While the Nova StatStrip® Glucose Hospital Meter System (Nova Biomedical, Waltham, MA, USA) is approved for point-of-care testing (POCT) in critically ill patients, its use during major abdominal surgery has not been evaluated. The purpose of this study was to assess the accuracy of the Nova StatStrip glucometer in patients undergoing major hepatobiliary procedures using the Parkes error grid (ISO15197:2013) and criteria defined by the Clinical and Laboratory Standards Institute (CLSI) POCT12-A3 guideline. METHODS: This study was a post hoc exploratory study of patients participating in a prospective randomized controlled trial on the effects of hyperinsulinemic normoglycemia (HNC) on infectious outcomes after hepatobiliary surgery. Arterial blood samples were collected before surgery and one hour, two hours, and three hours after baseline. Blood glucose levels were analyzed by the Nova StatStrip glucometer and the GEM® PremierTM 5000 blood gas analyzer. Accuracy of the StatStrip glucometer was assessed using the Parkes error grid for type 1 diabetes mellitus (when 99% of samples were within zones A and B on the Parkes error grid and clinical accuracy was acceptable) and the CLSI POCT12-A3 criteria. RESULTS: Blood glucose levels were analyzed in 135 patients, 70 of whom received the HNC. In the Parkes error grid plotted, all samples at all time-points were within zones A and B. The Nova StatStrip glucometer also satisfied CLSI POCT12-A3 criteria at all time-points. CONCLUSION: The Nova StatStrip glucometer was accurate in patients undergoing major upper abdominal surgery, independent of the administration of high-dose insulin therapy. STUDY REGISTRATION: ClinicalTrials.gov (NCT01528189); registered 7 February 2012.

RéSUMé: OBJECTIF: Bien que le système hospitalier de lecture de la glycémie StatStrip® de Nova (Nova Biomedical, Waltham, MA, É.-U.) soit approuvé pour une utilisation au chevet (ou POCT, pour 'Point of Care Testing') chez la patientèle en état critique, son utilisation n'a pas été évaluée en chirurgie abdominale majeure. L'objectif de cette étude était d'évaluer la précision du glucomètre StatStrip de Nova chez la patientèle bénéficiant d'interventions hépatobiliaires majeures à l'aide de la grille d'erreur de Parkes (ISO15197:2013) et des critères définis par la directive POCT12-A3 du Clinical and Laboratory Standards Institute (CLSI). MéTHODE: Il s'agissait d'une étude exploratoire post-hoc auprès de patient·es participant à une étude randomisée contrôlée prospective sur les effets de la normoglycémie hyperinsulinémique (HNC) sur les issues infectieuses après une chirurgie hépatobiliaire. Des échantillons de sang artériel ont été prélevés avant la chirurgie et une heure, deux heures et trois heures après l'échantillon initial. Les taux de glycémie ont été analysés avec le glucomètre StatStrip de Nova et l'analyseur de gaz sanguin GEM® PremierTM 5000. La précision du glucomètre StatStrip a été évaluée à l'aide de la grille d'erreur de Parkes pour le diabète sucré de type 1 (lorsque 99 % des échantillons se trouvaient dans les zones A et B de la grille d'erreur de Parkes et que la précision clinique était acceptable) et des critères POCT12-A3 du CLSI. RéSULTATS: La glycémie a été analysée chez 135 personnes, dont 70 ont reçu une normoglycémie hyperinsulinémique. Dans la grille d'erreur de Parkes tracée, tous les échantillons à tous les points temporels se trouvaient dans les zones A et B. Le glucomètre StatStrip de Nova a également satisfait aux critères POCT12-A3 du CLSI à tous les points temporels. CONCLUSION: Le glucomètre StatStrip de Nova était précis chez la patientèle bénéficiant d'une chirurgie abdominale supérieure majeure, indépendamment de l'administration d'insulinothérapie à forte dose. ENREGISTREMENT DE L'éTUDE: ClinicalTrials.gov (NCT01528189); enregistrée le 7 février 2012.

A clinical investigation of recurrence and lost follow-up after renal cell carcinoma surgery: a single-center, long-term, large cohort, retrospective study.

BACKGROUND: Late recurrence of renal cell carcinoma (RCC) is observed in some postoperative patients. In addition, some of these patients are lost to long-term postoperative follow-up. We reviewed the treatment results and prognosis of postoperative patients with RCC at Chiba University Hospital, with the aim of clarifying the proportion and background of patients lost to follow-up. METHODS: This retrospective study included 1176 RCC patients who underwent radical or/and partial nephrectomy. Overall survival (OS), cancer-specific survival (CSS), recurrence-free survival (RFS), and lost follow-up free survival (LFFS) were evaluated and the risk factors for LFFS identified. RESULTS: The median RFS for stage II and II cases was 188.3 and 104.0 months, respectively. Even in stage I, recurrence was observed in about 20% of patients 20 years after surgery. The Kaplan-Meier curve for LFFS showed a linear descent over time, with 50% of patients lost to follow-up within 25 years. Older age (≥ 62 years), histological type (clear cell RCC), and no recurrence were significant risk factors for lost follow-up. CONCLUSIONS: Long-term follow-up is necessary after RCC surgery because late recurrence cases are not uncommon. We believe that lifelong follow-up with imaging studies is recommended for postoperative RCC patients. Early detection of recurrence in postoperative patients is a very important issue, and it may be worthwhile for improving the prognosis of postoperative patients to focus on patients lost to follow-up who may have been overlooked.

Nanog is a promising chemoresistant stemness marker and therapeutic target by iron chelators for esophageal cancer.

Esophageal cancer is a disease showing poor prognosis. Although combination chemotherapy using cisplatin (CDDP) and 5-fluorouracil is standard for unresectable esophageal cancer, the response rate is 35%. Cancer stem cells (CSCs) and inflammation are reportedly responsible for the poor prognosis of esophageal cancer. However, comprehensive analyses have not been conducted and proposals for progress remain lacking. Iron is known to be a key factor in the stemness of CSCs. Our study focused on the therapeutic potential of iron control using iron chelators for CSCs in esophageal cancer. Among 134 immunohistochemically analyzed cases, Nanog expression was high in 98 cases and low in 36 cases. High Nanog expression correlated with low overall and disease-free survivals. The iron chelators deferasirox (DFX) and SP10 suppressed the proliferation and expression of stemness markers in TE8 and OE33 cells. DFX and SP10 did not induce compensatory interleukin (IL)-6 secretion, although CDDP did result in high induction. Moreover, BBI608 and SSZ, as other CSC-targeting drugs, could not suppress the expression of stemness markers. Overall, Nanog expression appears related to poor prognosis in esophageal cancer patients, and inhibition of stemness and compensatory IL-6 secretion by iron chelators may offer a novel therapeutic strategy for esophageal cancer.

Albumin and fibrinogen kinetics in sepsis: a prospective observational study.

BACKGROUND: The measurement of circulating substrate concentrations does not provide information about substrate kinetics. It, therefore, remains unclear if a decrease in plasma concentration of albumin, as seen during critical illness, is a consequence of suppressed production in the liver or increased peripheral clearance. In this study, using stable isotope tracer infusions, we measured albumin and fibrinogen kinetics in septic patients and in a control group of non-septic subjects. METHODS: With the approval from the institutional Research Ethics Board and after obtaining written informed consent from patients or their substitute decision maker, mechanically ventilated patients with sepsis and patients scheduled for elective coronary artery bypass grafting were enrolled. Patients in the non-sepsis group were studied on the day before surgery. The stable isotope L-[ring-2H5]phenylalanine was used to measure absolute synthesis rates (ASR) of albumin and fibrinogen. A priming dose of L-[ring-2H5]phenylalanine (4 µmol/kg) was given followed by a six-hour infusion at a rate of 0.15 µmol/kg/min. At baseline and hourly thereafter, blood was drawn to measure isotope enrichments by gas chromatography/mass spectrometry. Very low density lipoprotein apolipoprotein-B 100 isotopic enrichment was used to represent the isotopic enrichment of the phenylalanine precursor pool from which the liver synthesizes proteins. Plasma albumin and fibrinogen concentrations were also measured. RESULTS: Mean plasma albumin in septic patients was decreased when compared to non-septic patients, while synthesis rates were comparable. Mean plasma fibrinogen and ASR in septic patients was increased when compared to non-septic patients. In non-septic patients, no statistically significant correlation between plasma albumin and ASR was observed but plasma fibrinogen significantly correlated with ASR. In septic patients, plasma albumin and fibrinogen significantly correlated with ASR. CONCLUSIONS: While septic patients showed lower plasma albumin levels than non-septic patients, albumin synthesis was similar in the two groups suggesting that hypoalbuminemia during sepsis was not caused by suppressed hepatic production but a result of enhanced clearance from the circulation. Hyperfibrinogenemia in septic patients was a consequence of increased fibrinogen production. TRIAL REGISTRATION: ClinicalTrials.gov: NCT02865408 (registered on August 12, 2016) and ClinicalTrials.gov: NCT02549443 (registered on September 15, 2015).

Intense browsing by sika deer (Cervus nippon) drives the genetic differentiation of hairy nettle (Urtica thunbergiana) populations.

Many studies have inferred the way in which natural selection, genetic drift and gene flow shape the population genetic structures, but very few have quantified the population differentiation under spatially and temporally varying levels of selection pressure, population fluctuation and gene flow. In Nara Park (6.6 km2), central Japan, where several hundred sika deer (Cervus nippon) have been protected for more than 1,200 years, heavily- or moderately-haired nettle (Urtica thunbergiana) populations have evolved probably in response to intense deer browsing. Here, we analysed the genetic structure of two Nara Park populations and five surrounding populations using amplified fragment length polymorphism markers. A total of 546 marker loci were genotyped from 210 individuals. A Bayesian method estimated 5.5% of these loci to be outliers, which are putatively under natural selection. Neighbour-joining, principal coordinates and Bayesian clustering analyses using all-loci, non-outlier loci and outlier loci datasets showed that the Nara Park populations formed a cluster distinct from the surroundings. These results indicate the genome-wide differentiation of the Nara Park populations from the surroundings. Moreover, these imply the following: (1) gene flow is limited between these populations and thus genetic drift is a major factor causing the differentiation; and (2) natural selection imposed by intense deer browsing has contributed to some extent to the differentiation. In conclusion, sika deer seems to have counteracted genetic drift to drive the genetic differentiation of hairy nettles in Nara Park. This study suggests that a single herbivore species could lead to genetic differentiation among plant populations.

Intranasal administration of 40 and 80 units of insulin does not cause hypoglycemia during cardiac surgery: a randomized controlled trial.

PURPOSE: Intranasal insulin administration may improve cognitive function in patients with dementia and may prevent cognitive problems after surgery. Although the metabolic effects of intranasal insulin in non-surgical patients have been studied, its influence on glucose concentration during surgery is unknown. METHODS: We conducted a randomized, double-blind, placebo-contolled trial in patients scheduled for elective cardiac surgery. Patients with type 2 diabetes mellitus (T2DM) and non-T2DM patients were randomly allocated to one of three groups (normal saline, 40 international units [IU] of intranasal insulin, and 80 IU intranasal insulin). Insulin was given after the induction of general anesthesia. Glucose and plasma insulin concentrations were measured in ten-minute intervals during the first hour and every 30 min thereafter. The primary outcome was the change in glucose concentration 30 min after intranasal insulin administration. RESULTS: A total of 115 patients were studied, 43 of whom had T2DM. In non-T2DM patients, 40 IU intranasal insulin did not affect glucose concentration, while 80 IU intranasal insulin led to a statistically significant but not clinically important decrease in blood glucose levels (mean difference, 0.4 mMol·L-1; 95% confidence interval, 0.1 to 0.7). In T2DM patients, neither 40 IU nor 80 IU of insulin affected glucose concentration. No hypoglycemia (< 4.0 mMol·L-1) was observed after intranasal insulin administration in any patients. In non-T2DM patients, changes in plasma insulin were similar in the three groups. In T2DM patients, there was an increase in plasma insulin concentrations ten minutes after administration of 80 IU of intranasal insulin compared with saline. CONCLUSIONS: In patients with and without T2DM undergoing elective cardiac surgery, intranasal insulin administration at doses as high as 80 IU did not cause clinically important hypoglycemia. TRIAL REGISTRATION: www.ClinicalTrials.gov (NCT02729064); registered 5 April 2016.

RéSUMé: OBJECTIF: L'administration intranasale d'insuline pourrait améliorer la fonction cognitive des patients souffrant de démence et pourrait prévenir les problèmes cognitifs après une chirurgie. Bien que les effets métaboliques de l'insuline intranasale chez les patients non chirurgicaux aient été étudiés, son influence sur la glycémie pendant une chirurgie est inconnue. MéTHODE: Nous avons réalisé une étude randomisée, à double insu, contrôlée par placebo auprès de patients devant subir une chirurgie cardiaque non urgente. Des patients atteints de diabète de type 2 et des patients non diabétiques ont été randomisés dans l'un de trois groupes (solution physiologique salée, 40 unités internationales [UI] d'insuline intranasale et 80 UI d'insuline intranasale). La solution intranasale a été administrée après l'induction de l'anesthésie générale. Les concentrations de glucose et d'insuline plasmatique ont été mesurées à des intervalles de dix minutes pendant la première heure et toutes les 30 minutes par la suite. Le critère d'évaluation principal était le changement de glycémie 30 min après l'administration intranasale d'insuline. RéSULTATS: Un total de 115 patients ont été étudiés, dont 43 souffraient de diabète de type 2. Chez les patients non diabétiques, 40 UI d'insuline intranasale n'ont pas affecté la glycémie, alors que 80 UI d'insuline intranasale ont entraîné une réduction statistiquement significative mais non cliniquement importante de la glycémie (différence moyenne, 0,4 mMol·L−1; intervalle de confiance de 95 %, 0,1 à 0,7). Chez les patients diabétiques, ni 40 UI ni 80 UI d'insuline n'ont affecté la glycémie. Aucune hypoglycémie (< 4,0 mMol·L−1) n'a été observée après administration intranasale d'insuline chez les patients diabétiques ou non diabétiques. Chez les patients non diabétiques, les changements de l'insuline plasmatique étaient semblables dans les trois groupes. Chez les patients diabétiques, une augmentation des concentrations d'insuline plasmatique a été observée dix minutes après l'administration de 80 UI d'insuline intranasale comparée à la solution saline. CONCLUSION: Chez les patients diabétiques et non diabétiques subissant une chirurgie cardiaque non urgente, l'administration intranasale d'insuline à des doses allant jusqu'à 80 UI n'a pas causé d'hypoglycémie cliniquement importante. ENREGISTREMENT DE L'éTUDE: www.ClinicalTrials.gov (NCT02729064); enregistrée le 5 avril 2016.

Epigenetic modifications in prostate cancer.

Prostate cancer is a major cause of cancer-related deaths among men worldwide. In addition to genomic alterations, epigenetic alterations accumulated in prostate cancer have been elucidated. While aberrant deoxyribonucleic acid hypermethylation in promoter CpG islands inactivates crucial genes associated with deoxyribonucleic acid repair, cell cycle, apoptosis or cell adhesion, aberrant deoxyribonucleic acid hypomethylation can lead to oncogene activation. Acetylation of histone is also deregulated in prostate cancer, which could cause aberrant super-enhancer formation and activation of genes associated with cancer development. Deregulations of histone methylation, such as an increase of trimethylation at position 27 of histone H3 by enhancer of zeste homolog2 overexpression, or other modifications, such as phosphorylation and ubiquitination, are also involved in prostate cancer development, and inhibitors targeting these epigenomic aberrations might be novel therapeutic strategies. In this review, we provide an overview of epigenetic alterations in the development and progression of prostate cancer, focusing on deoxyribonucleic acid methylation and histone modifications.

Adrenalectomy in Japanese patients with subclinical Cushing syndrome: 1-mg dexamethasone suppression test to predict the surgical benefit.

OBJECTIVES: To investigate whether the result of the 1-mg dexamethasone suppression test can predict the improvement of comorbidities after adrenalectomy in patients with subclinical Cushing syndrome. METHODS: This retrospective study included 117 subclinical Cushing syndrome patients who underwent adrenalectomy. The numbers of prescribed drugs for metabolic comorbidities and the clinical variables at diagnosis were compared with those at the follow up. Patients were classified into subgroups according to the result of the 1-mg dexamethasone suppression test. RESULTS: Significant improvements in blood pressure, serum cholesterol and body mass index were observed. Furthermore, a significant improvement in glycated hemoglobin was observed in patients with diabetes mellitus. These improvements led to a discontinuation or reduction of prescribed drugs after surgery. In addition, the greatest reduction of prescribed drugs was observed in patients whose serum cortisol levels were between 1.8 and 3.0 µg/dL after the 1-mg dexamethasone suppression test. CONCLUSIONS: The result of the 1-mg dexamethasone suppression test can be a useful factor predicting the improvement of comorbidities after adrenalectomy. Current data might give us a new insight into the decision-making for the treatment of subclinical Cushing syndrome.

The Role of Brain Methamidophos in Acephate Poisoning in Mice.

We gave mice a 540 mg/kg dose of LD50 acephate, followed by an assessment of acephate, methamidophos (MP), and choline esterase (ChE) activity for up to 4 hours (hr) in order to investigate the time course of acephate intoxication. At 1 hr, the blood acephate and MP levels were 428 ± 90 µg/ml (mean ± SEM) and 4.2 ± 0.4 µg/ ml, respectively. The liver acephate levels were similar to those in the blood, but the liver MP levels were approximately 3.5 times that of the blood at 1 hr. The brain MP level tended to be higher than the blood MP at 1 hr. These levels decreased gradually over 4 hr, but the brain acephate and MP levels surpassed the blood levels significantly at 4 hr, and after 2 hr, respectively. Serum, liver, cerebrum, cerebellum, and brainstem cholinesterase activity (ChE) were inhibited at 1 hr, and remained inhibited in all but the cerebellum until the end of the experiment. The obtained data were applied to previously reported autopsy cases of acephate intake. Experimental data suggest that brain MP is involved in acute acephate-induced poisoning, even after a reduction in blood acephate. In autopsy cases with suspected acephate poisoning, the MP level in the brain should be considered in addition to the ChE activity to diagnose the cause of death.

Autopsy case of fatal hypoglycemia following ingestion of a therapeutic dose of tramadol.

An 86-year-old female was found unconscious the day after taking a prescribed tablet containing a combination of tramadol and acetaminophen. At admission to the hospital, marked hypoglycemia (blood glucose: 4 mg/dL) was confirmed, but serum insulin and C-peptide were within the normal range, which suggested that neither endogenous hyperinsulinemia nor exogenous insulin administration was responsible for the hypoglycemia. Despite resuscitation efforts, the woman subsequently died. At autopsy, there was renal disorder, but any pathological abnormalities that could have caused hypoglycemia were not observed. Blood tramadol and acetaminophen were in the therapeutic range. We speculate that the cause of fatal hypoglycemia was tramadol intake at the therapeutic dose. Older age and renal insufficiency are factors that could have potentially caused the fatal hypoglycemia in this case despite tramadol having been taken at a therapeutic dose. This is the first case report of fatal hypoglycemia following ingestion of a therapeutic dose of tramadol.

Rapid Fingerprinting of High-Molecular-Weight Polymers by Laser Desorption-Ionization Using Through-Hole Alumina Membrane High-Resolution Mass Spectrometry.

Residual acid found in the desorption ionization using through-holes alumina membranes (DIUTHAME) induces a reproducible protonation/in-source dissociation of polymers made of ester, amide, or siloxane moieties during their surface-assisted laser desorption ionization (SALDI) mass analysis. Deposited on the DIUTHAME chips in solution (solvent-based) or in pure form by melting the polymer powder in situ (solvent-free), high-molecular-weight nylons, silicone, or functionalized celluloses among other polymers are instantly fingerprinted by laser DIUTHAME high-resolution mass spectrometry (MS) with specific patterns resembling their direct analysis in real-time (DART) single-stage or tandem mass spectra. Depending on the polymer, two main types of fingerprints are observed with either the protonated monomer or product ions revealing the nature of the repeating unit or its functionalization. This technique allows a rapid molecular analysis of industrial homopolymers regardless of their molecular weight and complementary to DART with simple or no sample preparation and also promisingly applicable for copolymers.

MicroRNA biomarkers for chemical hazard screening identified by RNA deep sequencing analysis in mouse embryonic stem cells.

We investigated the responses of microRNAs (miRNAs) using mouse embryonic stem cells (mESCs) exposed to nine chemicals (bis(2-ethylhexyl)phthalate, p-cresol, p-dichlorobenzene, phenol, pyrocatecol, chloroform, tri-n-butyl phosphate, trichloroethylene, and benzene), which are listed as "Class I Designated Chemical Substances" from the Japan Pollutant Release and Transfer Register. Using deep sequencing analysis (RNA-seq), several miRNAs were identified that show a substantial response to general chemical toxicity (i.e., to these nine chemicals considered as a group) and several miRNA biomarkers that show a substantial and specific response to benzene. The functions of the identified miRNAs were investigated in accordance with Gene Ontology terms of their predicted target genes, indicating regulation of cellular processes. We compared the results with those for the long non-coding RNAs (ncRNAs) and mRNAs reported in our previous studies in addition to previously identified miRNAs that are either up- or down-regulated in response to the benzene as stimuli. We also observed that the changes in expression of miRNAs were smaller than those for long ncRNAs and mRNAs. Taken together the current and previous results revealed that toxic chemical stimuli regulate the expression of miRNAs. We believe that the use of miRNAs, including the thus identified miRNAs, as biomarkers contribute to predicting the potential toxicity of particular chemicals or identifying human individuals that have been exposed to chemical hazards.

Molecular characterization of polyethylene oxide based oligomers by surface-assisted laser desorption/ionization mass spectrometry using a through-hole alumina membrane as active substrate.

RATIONALE: Molecular characterization of industrial oligomeric products is performed using surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS), termed desorption ionization using a through-hole alumina membrane (DIUTHAME). This paper describes the unique feature of a DIUTHAME chip applying active SALDI, which generates specific types of fragments of polyglycol samples. METHODS: Polyethylene oxide (PEO) and PEO-based materials were subjected to SALDI-MS. The influence of the presence or absence of a cationization salt on the mass spectrum was investigated. The resulting mass spectra composed of fragment ions were compared with those obtained by collision-induced dissociation (CID)-MS/MS. The specific fragment ions generated using the DIUTHAME chip were further subjected to high-energy CID-MS/MS. RESULTS: The addition of a cationization salt resulted in SALDI mass spectra with fewer fragment peaks. The mass spectra obtained without adding the cationization salt were composed of many more fragment ions caused by in-source decay. The fragmentation pattern was similar to that seen with low-energy CID. The resulting fragment ions were formed by selective cleavage at the C-O bond. High-energy CID-MS/MS can be performed for the specific fragment ions generated by in-source decay fragmentation. CONCLUSIONS: Molecular characterization of PEO-based oligomers by SALDI-MS using the DIUTHAME chip was successfully demonstrated. The selective fragmentation and high-energy CID-MS/MS of the in-source decay fragments made it possible to provide more detailed structural information. This unique feature of DIUTHAME gives it potential for use in new molecular characterization techniques.

A mass spectrometry imaging method for visualizing synthetic polymers by using average molecular weight and dispersity as indices.

RATIONALE: Matrix-assisted laser desorption/ionization mass spectrometric imaging (MSI) is considered to be a powerful tool for visualizing the spatial distribution of synthetic polymers. However, a conventional method extracting an image of a specific m/z value is not suitable for polymers, which have a mass distribution. It is necessary to develop the visualization method to show the spatial distribution of entire polymer series. METHODS: The mass peaks included in polymer series were specified from the average mass spectrum of the entire MSI measurement region by using Kendrick mass defect analysis. The images of those mass peaks were extracted and the number average molecular weight (Mn ), the weight average molecular weight (Mw ) and dispersity (D) were calculated for each pixel. Finally, the spatial distribution of the polymer series was summarized to images using Mn , Mw and D as indices. RESULTS: The effects of the methods were investigated by (i) polymers with different mass distributions and (ii) polymers with different repeat units and end-groups. In both cases, the spatial distribution of specific polymer series including several dozens to hundreds of mass peaks was summarized into three images related to Mn , Mw and D, which are familiar indices in polymer analysis. The results are able to provide an overview of the spatial variation of each polymer more intuitively. CONCLUSIONS: The visualization of Mn , Mw and D will help provide an overview of the spatial distribution of polymer series combined with ion intensity distribution made by conventional methods. It can be also applied to other mass spectrometric imaging methods such as desorption electrospray ionization (DESI) or time-of-flight secondary ion mass spectrometry (TOF-SIMS).

An arsenal of tools based on Kendrick mass defects to process congested electrospray ionization high-resolution mass spectra of polymers with multiple charging.

RATIONALE: Electrospray ionization (ESI) favors the multiple charging of high molecular weight polymer samples and allows their high-resolution mass analysis in the low-mass range. It also induces the detection of numerous ion series at different charge states with different adducts complicating the interpretation of the mass spectrum which should be facilitated by an appropriate data processing. METHODS: An arsenal of tools based on the Kendrick mass defect (KMD) is proposed to process congested ESI high-resolution mass spectra of poly(propylene oxide) (PPO) samples. The combination of regular, charge-dependent, and resolution-enhanced KMD plots in addition to a "remainders" plot and a new three-dimensional plot offers unrivaled capabilities of filtering for any minor series among thousands of points. The sequential data processing is conducted using Kendo, a spreadsheet developed in-house for an advanced KMD analysis. RESULTS: The charge-state distribution is easily evaluated by counting the parallel lines in a regular KMD plot. A charge-dependent resolution-enhanced KMD plot instantly reveals the variation of adducted ions at a given charge state, helping the user to choose the best analytical conditions. Ion series at different charge states from PPO oligomers carrying different end-groups are also efficiently extracted using several combinations of KMD and remainders plots and assigned using a new simulator tool. CONCLUSIONS: The innovative combination of existing and new KMD-related plots, selection tools, and simulator all combined in a single spreadsheet dramatically facilitates the processing and interpretation of complex ESI mass spectral data. The presented tools may be extended to any other class of homo-, co- and terpolymers.