Short Telomere Lesions with Dysplastic Metaplasia Histology May Represent Precancerous Lesions of Helicobacter pylori-Positive Gastric Mucosa.

Gastric cancers are strongly associated with Helicobacter pylori infection, with intestinal metaplasia characterizing the background mucosa in most cases. However, only a subset of intestinal metaplasia cases proceed to carcinogenesis, and the characteristics of high-risk intestinal metaplasia that link it with gastric cancer are still unclear. We examined telomere reduction in five gastrectomy specimens using fluorescence in situ hybridization, and identified areas with localized telomere loss (outside of cancerous lesions), which were designated as short telomere lesions (STLs). Histological analyses indicated that STLs were characteristic of intestinal metaplasia accompanied by nuclear enlargement but lacking structural atypia, which we termed dysplastic metaplasia (DM). A review of gastric biopsy specimens from 587 H. pylori-positive patients revealed 32 cases of DM, 13 of which were classified as high-grade based on the degree of nuclear enlargement. All high-grade DM cases exhibited a telomere volume reduced to less than 60% of that of lymphocytes, increased stemness, and telomerase reverse transcriptase (TERT) expression. Two patients (15%) exhibited low levels of p53 nuclear retention. After a 10-year follow-up, 7 (54%) of the high-grade DM cases had progressed to gastric cancer. These results suggest that DM is characterized by telomere shortening, TERT expression, and stem cell proliferation, and high-grade DM is a high-grade intestinal metaplasia that likely represents a precancerous lesion of gastric cancer. High-grade DM is expected to effectively prevent progression to gastric cancer in H. pylori-positive patients.

Intracellular interplay between cholecystokinin and leptin signalling for satiety control in rats.

Cholecystokinin (CCK) and leptin are satiety-controlling peptides, yet their interactive roles remain unclear. Here, we addressed this issue using in vitro and in vivo models. In rat C6 glioma cells, leptin pre-treatment enhanced Ca2+ mobilization by a CCK agonist (CCK-8s). This leptin action was reduced by Janus kinase inhibitor (AG490) or PI3-kinase inhibitor (LY294002). Meanwhile, leptin stimulation alone failed to mobilize Ca2+ even in cells overexpressing leptin receptors (C6-ObRb). Leptin increased nuclear immunoreactivity against phosphorylated STAT3 (pSTAT3) whereas CCK-8s reduced leptin-induced nuclear pSTAT3 accumulation in these cells. In the rat ventromedial hypothalamus (VMH), leptin-induced action potential firing was enhanced, whereas nuclear pSTAT3 was reduced by co-stimulation with CCK-8s. To further analyse in vivo signalling interplay, a CCK-1 antagonist (lorglumide) was intraperitoneally injected in rats following 1-h restricted feeding. Food access was increased 3-h after lorglumide injection. At this timepoint, nuclear pSTAT3 was increased whereas c-Fos was decreased in the VMH. Taken together, these results suggest that leptin and CCK receptors may both contribute to short-term satiety, and leptin could positively modulate CCK signalling. Notably, nuclear pSTAT3 levels in this experimental paradigm were negatively correlated with satiety levels, contrary to the generally described transcriptional regulation for long-term satiety via leptin receptors.

Regulatory perspectives on next-generation sequencing and complementary diagnostics in Japan.

INTRODUCTION: The 'one biomarker/one drug' scenario is unsustainable because cancer is a complex disorder that involves a number of molecular defects. In the past decade, major technological advances have lowered the overall cost and increased the efficiency of next-generation sequencing (NGS). AREAS COVERED: We review recent regulations on NGS and complementary diagnostics in Japan, mainly focusing on high-quality studies that utilized these new diagnostic modalities and were published within the last 5 years. We highlight significant changes in regulation, and explain the direction of efforts to translate the results of NGS and complementary diagnostics into clinical practice. EXPERT OPINION: NGS holds a number of advantages over conventional companion and complementary diagnostics that enable simultaneous analyzes of multiple cancer genes to detect actionable mutations. Parallel technological developments and regulatory changes have led to the rapid adoption of NGS into clinical practice. NGS-based genomic data have been leveraged to better understand the characteristics of a disease that affects its patient's response to a given therapy. As NGS-based tests become more widespread, however, Japanese authorities will face significant challenges particularly with respect to the complexity of genomic data, which will have to be managed if NGS is to benefit patients.

Biological monitoring of occupational exposure to dichloromethane by means of urinalysis for un-metabolized dichloromethane.

The objective of the study is to establish exposure-excretion relationship between dichlorometane (DCM) in air (DCM-A) and in urine (DCM-U) in workplace to confirm a previous report. Male workers in a screen-printing plant participated in the study. Time-weighted average DCM-A was measured by diffusive sampling followed by gas-chromatography (GC), and DCM in end-of-shift urine samples was by head-space GC. The data were subjected to regression and other statistical analyses. In practice, 30 sets of DCM-A and DCM-U values were available. The geometric mean DCM-A was 8.4 ppm and that of DCM-U (as observed) was 41.1 µg/l. The correlation coefficients (0.70-0.85) were statistically significant across the correction for urine density. Thus, the analysis for un-metabolized DCM in end-of-shift urine samples is applicable for biological monitoring of occupational exposure to DCM, in support of and in agreement with the previous report. In conclusion, biological monitoring of occupational DCM exposure is possible by use of analysis for un-metabolized DCM in end-of-shift urine.

Biogenically induced bedded chert formation in the alkaline palaeo-lake of the Green River Formation.

Rhythmically bedded cherts are observed in both pelagic marine and lacustrine deposits, but the formation mechanism in the latter remains highly uncertain. Our study of alternating chert-dolomite beds in the Eocene Green River Formation, Utah, USA reveals dense accumulations of organic-matter spheres (30-50 µm diameter) of probable algal cyst origin in the chert layers, and centennial- to millennial-scale periodicities in chert layer deposition. A positive correlation between the degree of degradation of the organic spheres and Si distribution implies decomposition of algal organic matter lead to precipitation of lacustrine chert. As both alkalinity and dissolved silica were likely high in the palaeo-lake waters of the Green River Formation, we hypothesize that decomposition of algal organic matter lowered the pH of sediment pore waters and caused silica precipitation. We propose a formation model in which the initial abundance of algal organic matter in sediment varies with productivity at the lake surface, and the decomposition of this algal matter controls the extent of silica precipitation in sediment. The formation of rhythmically bedded chert-dolomite may be linked to centennial- to millennial-scale climatic/environmental factors that modulate algal productivity, which are possibly tied to solar activity cycles known to have similar periodicities.

Simplified procedures for estimation of biological occupational exposure limits.

OBJECTIVES: To simplify the procedures to estimate biological occupational exposure limits (BOELs) by use of the ratio of geometric mean (GM) concentration of un-metabolized organic solvent in urine (U-GM) over GM organic solvent concentration in air (A-GM) (the [U-GM/A-GM] ratio). METHODS: Occupational Exposure Limits (OELs) and BOELs were cited from publications from the Japan Society of Occupational Health (JSOH) and the American Conference of Governmental Industrial Hygienists (ACGIH). Data on [U-GM/A-GM] and the SLOPE of exposure-excretion regression line were collected from published articles (men and women were treated separately). Correlation analysis and paired t test were employed as the method to examine statistical significances. RESULTS: Significant linear correlation was established between the SLOPE and the [U-GM/A-GM]. Thus, it was considered to be possible to calculate the SLOPE value from the [U-GM/A-GM]. Previously established equation of BOEL = SLOPE × OEL allowed to estimate BOEL values in 22 cases of data sets. The comparison of the estimated BOELs with the existing BOELs (JSOH's BOELs and ACGIH's BEIs) in terms of the ratio of [(estimated BOEL)/(existing BOEL)] showed that the ratios for the 22 cases probably distributed log-normally with a GM of 0.85, and the maximum was 5. Therefore, the estimated BOEL may be generally applicable in occupational health when BOEL remains yet to be established. In the worst case, the estimated BOEL may be five times greater than it should be. The recommended procedures for application of estimated BOEL values were described. CONCLUSION: Simplified procedures for estimation of BOEL values are proposed.

Antipsychotic drugs scavenge radiation-induced hydroxyl radicals and intracellular ROS formation, and protect apoptosis in human lymphoma U937 cells.

Antioxidant activity has been reported for some atypical antipsychotic drugs; however, the detailed mechanism is not well known. Here, we investigated the effects of atypical antipsychotic drugs on •OH radical formation, intracellular reactive oxygen species (ROS), and apoptosis induced by ionising radiation. The reaction rate constants with •OH radicals were determined for five antipsychotic drugs as follows, in descending order: olanzapine, aripiprazole, clozapine, haloperidol, and risperidone. Experiments with aminophenyl fluorescein, a fluorescent dye, showed that olanzapine and clozapine could scavenge intracellular ROS. However, experiments with hydroxyphenyl fluorescein showed that only olanzapine inhibited ROS generation. X-irradiation-induced apoptosis in human lymphoma U937 cells was inhibited by clozapine at relatively low concentrations and by olanzapine at higher concentrations. Clozapine inhibited caspase-8 and caspase-3 activation and prevented loss of mitochondrial membrane potential. In contrast, olanzapine inhibited X-irradiation-induced p-JNK activation. Although the atypical antipsychotic drugs used here have relatively high reaction rate constants with •OH radicals in aqueous solutions, inhibition of intracellular ROS was not due to •OH radical scavenging. In addition, suppression of X-irradiation-induced apoptosis was not directly linked with intracellular ROS scavenging. When apoptosis signalling pathways were studied, clozapine-mediated inhibition of apoptosis was dependent on caspase-3 and caspase-8. In contrast, olanzapine inhibited apoptosis via down regulation of X-irradiation-induced p-JNK. These results suggested that both olanzapine and clozapine have antioxidative and antiapoptotic activities via distinct pathways, and provide useful information for better understanding of drug characteristics.

Biological monitoring of occupational ethylbenzene exposure by means of urinalysis for un-metabolized ethylbenzene.

This study aimed to examine quantitative relation between ethylbenzene (EB) in air (EB-A) and un-metabolized EB in urine (EB-U) for biological monitoring of occupational EB exposure by urinalysis for EB. In total, 49 men in furniture production factories participated in the study. Time-weighted average EB-A was monitored by diffusive sampling. Urinalysis for EB was conducted by head-space gas-chromatography with end-of-shift samples. Data were subjected to regression analysis for statistical evaluation. A geometric mean (GM) and the maximum (Max) EB-A levels were 2.1 and 45.5 ppm, respectively. A GM and the Max for EB-U (observed values) were 4.6 and 38.7 µg/l. A significant linear correlation was observed. The regression equation was Y=3.1+0.73X where X is EB-A (ppm) and Y is EB-U (µg/l) (r=0.91, p<0.01). The significant correlation between EB-A and EB-U coupled with a small intercept suggests that biological monitoring of occupational EB exposure is possible by analysis for un-metabolized EB in end-of-shift urine samples. Further validation studies (including those on applicability to women) are envisaged. The feasibility should be examined for biological monitoring and the applicability of the equation among the workers exposed to EB at low levels.

Estimation of dietary intake of cadmium from cadmium in blood or urine in East Asia.

BACKGROUND AND AIM: Cadmium (Cd) is an ubiquitous environmental toxic pollutant. As daily foods are an almost exclusive source of exposure for general populations, it is of public health importance to know the level of dietary intake of cadmium (Cd-D). The purpose of this study is to examine whether Cd in blood (Cd-B) or urine (Cd-U) correlates with Cd-D in East Asia, and in case it is, whether it is possible to estimate Cd-D from Cd-B or Cd-U. It should be added that the measurement of Cd-D is quite hand-consuming in practice. MATERIALS AND METHODS: Literature was retrieved for publication on Cd-B and Cd-U in combination with Cd-D. Twenty three data sets thus obtained for East Asia were subjected to regression analysis to investigate the possibility to estimate Cd-D from Cd-B or Cd-U. RESULTS: In Japan and Korea, large correlation coefficients (p > 0.7) were observed between Cd-B and Cd-D, as well as between Cd-U and Cd-D. In China, the coefficient was >0.7 between Cd-B and Cd-D. Furthermore, correlation was significant for Cd-B and Cd-D, as well as Cd-U and Cd-D, when 19 sets for Japan, Korea and China were combined for analysis. DISCUSSION: Major reasons for successful analysis may be predominant use of women-based data. Women have been less smoking than men in East Asia, and possible confounding effects of smoking on Cd exposure might be minimized. CONCLUSION: Based on significant correlations, Cd-D can be estimated from Cd-B or Cd-U in East Asia.

Further examination of log Pow-based procedures to estimate biological occupational exposure limits.

OBJECTIVES: To test the reliability of the procedures (described in a previous article) for estimation of biological occupational exposure limits (BOELs). METHODS: Data on four organic solvents (styrene, ethyl benzene, isopropyl alcohol and tetrachloroethylene) were obtained from recent publications and added to previously cited data for 10 organic solvents. Regression analysis was used for statistical evaluation. RESULTS AND DISCUSSION: The previously reported results obtained using 10 solvents were reproduced by the analysis with 14 solvents. Repeated randomized division of the 14 sets into two subgroups of equal size followed by statistical comparisons did not show a significant difference between two regression lines. This reproducibility suggests that the procedures used to estimate BOELs may be applicable across many solvents, and this may be of particular benefit for protecting the health of workers who work with skin-penetrating solvents.

Multidirectional analyses of hepatic chronotoxicity induced by cadmium in mice.

The expression levels or activities of biological defense factors can fluctuate daily following biological rhythms. We have focused on the relationship between injection timing and the degree of toxicity of cadmium (Cd) to promote the concept of "chronotoxicology," which introduces chronobiology to the field of toxicology. Our previous studies have clearly indicated that Cd may be subject to chronotoxicity. In this report, to confirm the character of the Cd-induced chronotoxicity, we performed multidirectional examinations. Male C57BL/6J mice that received a single intraperitoneal injection of CdCl2 at ZT6 showed drastic hepatic injury estimated by histopathological analyses, i.e., nuclear condensations, fatty degenerations, and hemorrhages, but showed no injury when injected at ZT18. This difference was supported by several biochemical analyses that were indicators of hepatic injury (levels of alanine aminotransferase, aspartate aminotransferase, and malondialdehyde). The chronotoxicity of Cd was also observed in multiple strains (ICR and Balb/c), in a different injection route (subcutaneous), and in multiple injections (5 injections). Based on these results, we propose that chronotoxicology may provide important information not only for toxicology but also for occupational health, i.e., the importance of injection timing for toxicity evaluation tests, the reproducibility of animal experiments, and the improvement in the quality of risk assessments for night shift workers who may be exposed to toxic substances at various times of the day.

Inhibitory Effect of Bovine Lactoferrin on Catechol-O-Methyltransferase.

Lactoferrin (LF) is a well-known multifunctional protein. In this study, we report the inhibitory potency of bovine LF (bLF) on catechol-O-methyltransferase (COMT), which catalyzes methylation of catechol substrates. We found that bLF binds to and inhibits COMT using its N-terminal region. An N-terminal peptide fragment obtained from bLF by trypsin digestion showed a higher inhibitory activity than intact bLF. A synthetic fragment of the bLF N-terminal residues 6-50, with two pairs of disulfide bonds, also showed higher inhibitory activity than intact bLF. Enzyme kinetic studies proved that bLF did not compete with S-adenosylmethionine (the methyl donor substrate) as well as methyl acceptor substrates such as dihydroxybenzoic acid, (-)-epicatechin, norepinephrine, or l-3,4-dihydroxyphenylalanine. The inhibitory potency of bLF decreased against a COMT preparation pretreated with dithiothreitol, suggesting that the oxidation status of COMT is relevant to interaction with bLF. We further confirmed that COMT activity in the cell extracts form Caco-2 and HepG2 cells was inhibited by bLF and by the synthesized fragment. Enzyme kinetic study indicated that bLF functions as a non-competitive inhibitor by binding to an allosteric surface of COMT.

Approval status and evidence for WHO essential medicines for children in the United States, United Kingdom, and Japan: a cross-sectional study.

BACKGROUND: The WHO Model List of Essential Medicines for Children (EMLc) covers medicines for globally high-burden diseases. Regulatory approval in high-income countries ensures evidence and dosage form but usually focuses on diseases common in those countries and not in low- and middle-income countries. METHODS: This cross-sectional study assessed supporting evidence for the 346 medicines in the 5th WHO EMLc and their approval data from the United States, United Kingdom, and Japan. RESULTS: Of the 346 EMLc medicines, 307 were approved in one or more of the three countries, 278 of which had supporting evidence of efficacy. The percentage of medicines approved in one or more of the three countries was lowest for antiparasitics (60%) whereas 100% for medicines for cancers and musculoskeletal and respiratory conditions were approved. Five of the 30 EMLc antineoplastics had no supporting paediatric evidence. Of the 39 EMLc medicines unapproved in all three countries, 26 were indicated for neglected infectious diseases (NIDs). Ten of the 26 had supporting paediatric evidence. Seventeen of the 39 unapproved medicines had no paediatric dosage form available, and all 17 were indicated for NIDs. CONCLUSIONS: Most EMLc medicines for diseases common in the three countries had supporting evidence, which was closely associated with approval, whereas a substantial number of medicines for NIDs were unapproved in the three countries, regardless of whether they had supporting evidence. Because of the limited contribution to the EMLc from high income countries, appropriate incentive mechanisms for pharmaceutical companies are required to make paediatric development for NIDs feasible and effective.

Bimodal modulation of store-operated Ca2+ channels by clozapine in astrocytes.

Clozapine (Clz) and olanzapine (Olz) are second generation (atypical) antipsychotics, used widely for treating schizophrenia and bipolar disorder. These drugs share multiple sites of actions, however their mechanisms remain incompletely understood. Here, we analyzed the effects of these drugs on primary cultures of rat cortical astrocytes and C6 glioma cells using fura-2-based Ca2+ imaging. C6 cells, but not cortical astrocytes, express the serotonin 2A receptor subtype, which couples to phospholipase C. Clz (1µM) significantly blocked serotonin-induced Ca2+ transients in C6 cells, consistent with known antagonistic actions of Clz. Interestingly, at higher concentrations (>10µM), Clz but not Olz increased intracellular Ca2+ concentrations in both cortical astrocytes and C6 cells. This Clz-induced Ca2+ increase was concentration-dependent and completely blocked by removal of extracellular Ca2+ using ethylene glycol tetraacetic acid (EGTA). Furthermore, 2-aminoethyl diphenylborinate or SKF-96365, blockers for store-operated Ca2+ channels, significantly inhibited the Clz-induced Ca2+ increase. Therefore, we analyzed the effects of Clz and Olz during Ca2+ re-entry through store-operated Ca2+ channels, which was maximized following depletion of internal Ca2+ stores by thapsigargin and EGTA. The results demonstrated that Clz decreased Ca2+ re-entry through store-operated Ca2+ channels in cortical astrocytes and C6 cells whereas Olz failed to modulate the Ca2+ re-entry. These results suggest Clz-specific bimodal actions via store-operated Ca2+ channels in astrocytic cells. Since intracellular Ca2+ homeostasis in astrocytes is an important determinant for neighboring synaptic signal transmission, our results may explain Clz-specific adverse effects or differential actions between Clz and Olz reported in the treatment of psychiatric disorders.

Time trend of cadmium intake in Korea.

OBJECTIVES: The aim of this study was to elucidate past and current levels of cadmium (Cd) intake among the general populations in Korea. METHODS: For this purpose, publications reporting dietary intake of cadmium (Cd-D), cadmium concentration in blood (Cd-B) and that in urine (Cd-U) in Korea were retrieved through literature survey for a period from 1975 to 2015. RESULTS: In practice, 9, 21 and 14 articles were available on Cd-D, Cd-B and Cd-Ucr (Cd-U as corrected for creatinine concentration), respectively. Linear regression analyses of the reported values as a function of years (i.e., the year when each survey was conducted) showed steady decreases in all of the three exposure markers of Cd-D, Cd-B and Cd-U(cr). Factors possibly contributing for the reduction were discussed including the government-set guideline of 0.2 mg/kg for rice and changes in food habits among general populations. CONCLUSIONS: There have been steady decreases in Cd-D, Cd-B and Cd-U(cr). The current estimates for Cd-D, Cd-B and Cd-Ucr were 6.0-7.4 µg/day, 0.73-0.83 µg/L and 0.60-0.95 µg/g cr, respectively.

Drug-diagnostic co-development: challenges and issues.

Diagnostic tests have become increasingly important for optimizing drug use. Ideally, a companion diagnostic test is developed concurrently with a corresponding therapeutic product (co-development). However, the diagnostic test may also be developed to optimize treatment with previously approved therapeutic agents (follow-up-development). In co-development, the effectiveness of an agent in marker-defined patients is confirmed by an enrichment trial design. In follow-up-development, a biomarker is validated by prospective and/or retrospective analyses of unselected design trials. A prospectively designed trial is the gold-standard approach to biomarker validation, but retrospective validation can be used to efficiently determine effective treatments for marker-defined patients. Accumulation and systematization of examples of drug-diagnostic development will aid in the effective development of companion diagnostics.

Estimation of daily cadmium intake from cadmium in blood or cadmium in urine.

OBJECTIVES: It would be quite convenient if daily dietary cadmium intake (Cd-D) can be estimated either from Cd in blood (Cd-B) or from Cd in urine (Cd-U). The aim of the study was to examine if Cd-D can be estimated from Cd-B or Cd-U. METHODS: The data available in a previous publication were employed for regression analyses between Cd-D and Cd-B, and between Cd-D and Cd-U. 30 sites in various prefectures throughout Japan were surveyed and 20 adult women/site on average provided food duplicate, peripheral blood, and second morning urine samples. Geometric means were taken as representative values and employed in regression analyses. RESULTS: Cd-D, Cd-B, and Cd-U(cr) [i.e., Cd-U after correction for creatinine (cr) concentration] distributed in ranges of 12.5-70.5 µg/day, 0.46-3.98 µg/l, and 1.16-11.02 µg/g cr, respectively. A close correlation was observed between Cd-D and Cd-B, and also between Cd-D and Cd-U(cr) with r = 0.76 and r = 0.79 (p < 0.001 for both), respectively. Both regression lines passed close to the origins. Application of 1.23 µg Cd/l blood and 1.26 µg Cd/g cr in urine (average levels for adult Japanese women) to the regression equations gave 16.5 and 11.5 µg Cd/day. CONCLUSIONS: The analyses suggested that it may be possible to estimate Cd-D from Cd-B or Cd-U. Cd-B-based estimation should be more respected. As variations in the estimation parameters and estimated values are inherent to field surveys, care should be taken in the application of the study results. Application on a group basis (and not on an individual basis) should be considered.