Comparative study on the activities of part-time occupational physicians in Japan between 2008 and 2016: effects of the stress-check program.

The Ministry of Health, Labor, and Welfare of Japan recommends that an occupational physician (OP) play an important role in implementing the stress-check program since 2015. This study aimed to compare the activities and encountered difficulties of Japanese part-time OPs in 2008 and 2016, and to investigate the effects of the stress-check program. Questionnaires were sent via mail to 946 part-time OPs in Kyoto prefecture in 2016. Completed questionnaires were returned by 181 OPs who were private practitioners or physicians in hospitals, and served as OPs on a part-time basis. In 2016, OPs utilized long hours for activities related to general health examination and to stress-check. Hours for specific health examination, health and hygiene education, health promotion activity, development of a comfortable workplace, and guidance of workers on sick leave reduced from 2008 to 2016. A total of 62% OPs frequently encountered difficulties in the stress-check-related activities in 2016. Many OPs also reported difficulties in the mental health care and the prevention of health hazard due to overwork both in 2008 and 2016. Enforcement of the stress-check program in 2015 changed the activities of part-time OPs in Japan. OPs should be given opportunities to gain more information in this area.

[Dose effect of alcohol on sex differences in blood alcohol metabolism--cases where healthy subjects with ALDH2*1/1 genotype drunk beer with meal].

It is said that blood alcohol concentrations (BAG) are higher in female than in male due to the smaller distribution volume of alcohol in female, whereas the rate of alcohol metabolism is faster in female than in males due to a higher activity of liver alcohol dehydrogenase (ADH) in female. However, it is also known that alcohol metabolism varies depending on drinking conditions. In this study, we evaluated the dose effect of alcohol on sex differences in alcohol metabolism in daily drinking conditions, where young adults (16 males, 15 females) with ALDH2*1/1 genotype drunk beer at a dose of 0.32g or 1.0g ethanol/kg body weight with a test meal (460kcal). This study was conducted using a randomized cross-over design. In the considerable drinking condition (1.0g/kg), BAG was significantly higher in females than in males, whereas the rate of alcohol metabolism (beta) was higher in female than in male. In the moderate drinking condition (0.32g/kg), however, no sex differences in alcohol metabolism including BAG were seen. These results suggest that an increased first pass metabolism through liver ADH in female, which may be caused by the reduction of gastric emptying rate due to the meal intake, contribute to the vanishing of sex difference in BAC in the moderate drinking condition.

Characteristics, risk factors and mortality of stroke patients in Kyoto, Japan.

OBJECTIVES: The aim of the study was to evaluate the characteristics, risk factors and outcome of recent stroke patients in Kyoto, Japan. DESIGN: We analysed stroke patients in the registry with regard to their characteristics, risk factors and mortality. Cox proportional hazards regressions were used to calculate adjusted HRs for death. SETTINGS: The Kyoto prefecture of Japan has established a registry to enrol new stroke patients in cooperation with the Kyoto Medical Association and its affiliated hospitals PARTICIPANTS: The registry now has data on 14 268 patients enrolled from 1 January 1999 to 31 December 2009. Of these, 12 774(89.5%) underwent CT, 9232 (64.7%) MRI, 2504 (17.5%) angiography and 342 (2.4%) scintigraphy. Excluding 480 (3.3%) unclassified patients, 13 788 (96.6%) patients formed the basis of further analyses which were divided into three subtypes: cerebral infarction (CI), cerebral haemorrhage (CH) and subarachnoid haemorrhage (SAH). RESULTS: A total of 13 788 confirmed stroke patients in the study cohort comprised 9011 (86.3%) CI, 3549 (25.7%) CH and 1197 (8.7%) SAH cases. The mean age ±SD was 73.3±11.8, 69.1±13.6 and 62.7±13.5 in the CI, CH and SAH cases, respectively. Men were predominant in the CI and CH cases, whereas women were predominant in the SAH cases. The frequencies of risk factors were different among the subtypes. Mortality was worst in SAH, followed by CH, and least in CI. HRs for death adjusted for age, sex, histories of hypertension, arrhythmia, diabetes mellitus and hyperlipaemia and use of tobacco and/or alcohol showed a significant (p<0.001) difference among CI (as reference), CH (3.71; 3.11 to 4.43) and SAH (8.94; 7.21 to 11.11). CONCLUSIONS: The characteristics, risk factors and mortality were evaluated in a quantitative manner in a large Japanese study cohort to shed light on the present status of stroke medicine.

Oral administration of Cimicifuga racemosa extract attenuates psychological and physiological stress responses.

Dried rhizomes of Cimicifuga racemosa (CR), which are known as black cohosh, have been widely used as herbal dietary supplements to treat menopausal symptoms. The present study examined the effect of CR extracts on human psychological and physiological responses to acute stress induced by mental arithmetic tests, by measuring the subjective stress intensity, the brain-wave patterns according to electroencephalography, and the concentrations of salivary chromogranin-A and cortisol. The experiments were performed double-blind and their order was counterbalanced. Treatment with CR significantly attenuated the elevated subjective perception of stress and the increased salivary chromogranin-A levels compared with placebo treatment. CR extract also rapidly recovered the decrease in alpha waveband induced by performing the mental arithmetic task. We therefore propose that CR extracts might be suitable for the prevention and treatment of stress-related disorders.

Oral administration of Cimicifuga racemosa extract affects immobilization stress-induced changes in murine cerebral monoamine metabolism.

We investigated the effects of Cimicifuga racemosa (CR) plant extracts on the changes in levels of the cerebral monoamines norepinephrine (NE), dopamine (DA), and serotonin (5-HT), the respective metabolites 3-methoxy-4-hydroxyphenylglycol (MHPG), 3,4-dihydroxyphenylacetic acid (DOPAC), and 5-hydroxyindoleacetic acid (5-HIAA), and plasma corticosterone in mice subjected to acute immobilization stress. Single oral administration of the CR extract (1,000 mg/kg) significantly attenuated plasma corticosterone levels that had been increased as a result of enforced immobilization. Acute immobilization stress caused significant changes in the corresponding amine-to-metabolite ratios in the hypothalamus, hippocampus, and cortex; however, CR-extract treatment significantly attenuated the MHPG/NE change in the hypothalamus, and the 5-HIAA/5- HT changes in each region of the brain. Our results suggest that the CR extract interacts not only with the hypothalamic-pituitary-adrenal (HPA) axis but also with the sympathetic adrenomedullary (SAM) system under stress conditions. Thus the CR extract can alleviate acute stress responses by suppressing the changes of amine-to-metabolite ratio in brain.

Oral administration of Cimicifuga racemosa extract attenuates immobilization stress-induced reactions.

Dried rhizomes of Cimicifuga racemosa (CR), known as black cohosh, have been widely used as a herbal dietary supplement in the treatment of menopausal symptoms. Here we used experimental mouse stress models to investigate the role of anti-stress food factors, and found that a CR extract had stress-relieving effects. A single oral administration of CR extract (1,000 mg/kg) significantly attenuated plasma corticosterone and aspartate aminotransferase (AST) levels that had increased as a result of enforced immobilization. Bioassay-guided fractionation of the CR extract resulted in the isolation of 10 triterpenes, among which actein, 23-epi-26-deoxyactein, and cimiracemoside F (100 mg/kg, per os) were shown to contribute to the anti-stress effects. Furthermore, the CR extract significantly prevented the development of water immersion stress-induced gastric mucosal ulcers in rats. We propose that the CR extract might be suitable for the prevention and treatment of stress-related disorders.

Intraspecies discrimination of Lactobacillus paraplantarum by PCR.

Lactobacillus paraplantarum is a species phenotypically close to Lactobacillus plantarum. Several PCR methods were evaluated to discriminate L. paraplantarum strains and among them, a PCR using an enterobacterial repetitive intergenic consensus (ERIC) sequence differentiated L. paraplantarum from other Lactobacillus species. In addition, a combination of ERIC and random amplified polymorphic DNA (RAPD) analysis distinguished among seven strains of L. paraplantarum tested. ERIC-PCR profiles showed several strain-specific DNA fragments in L. paraplantarum, among them, a 2.2-kb ERIC marker, termed LpF1, found to be specific to strain FBA1, which improved the skin integrity in an animal model. The LpF1 encodes three proteins similar to Lactobacillus fermentum AroA, TyrA, and AroK, which are involved in the shikimate pathway. A primer pair specific to FBA1 based on the internal sequence of LpF1 amplified a 950-bp FBA1-specific fragment LpF2. Southern blot analysis of Dra I-digested genomic DNA of L. paraplantarum strains using LpF2 as a probe showed that LpF2 is distinctive of strain FBA1 among 16 L. paraplantarum strains. Because both ERIC- and RAPD-PCR are fast and technically simple methods, they are useful for the rapid discrimination of L. paraplantarum strains and for the development of new strain-specific DNA markers for identifying industrially important strains.

Identification of the layered morphology of the esophageal wall by optical coherence tomography.

AIM: To assess each layer of the optical coherence tomography (OCT) image of the esophageal wall with reference to the histological structure. METHODS: Resected specimens of fresh pig esophagus was used as a model for the esophageal wall. We injected cyanoacrylate adhesive into the specimens to create a marker, and scanned them using a miniature OCT probe. The localization of these markers was assessed in the OCT images. Then we compared the OCT-imaged morphology with the corresponding histological section, guided by the cyanoacrylate adhesive markers. We prepared a second set of experiments using nylon sutures as markers. RESULTS: The OCT image of the esophageal specimen has a clear five-layered morphology. First, it consisted of a relatively less reflective layer; second, a more reflective layer; third, a less reflective layer; fourth, a more reflective layer; and fifth, a less reflective layer. Comparing the OCT images with marked histological sections showed that the first layer corresponded to stratified squamous epithelium; the second to lamina propria; the third to muscularis mucosa; fourth, submucosa; and fifth, muscularis propria with deeper structures of the esophageal wall. CONCLUSION: We demonstrated that the OCT image of the normal esophageal wall showed a five-layered morphology, which corresponds to histological esophageal wall components.

HorC, a hop-resistance related protein, presumably functions in homodimer form.

To determine whether two HorC molecules coordinately form a single unit, the functional properties of covalently linked dimers of HorC encoded by tandemly fused horC genes were studied. Lactobacillus brevis introduced with the fused horC genes and a single horC gene exhibited same degree of resistance to hop compounds and cetyltrimethylammonium bromide. This suggests that HorC functions as a homodimer.

Determination of a yellow rice toxin, luteoskyrin, in rice by using liquid chromatography-tandem mass spectrometry with electrospray ionization.

Penicillium islandicum produces luteoskyrin (LUT), a yellow rice toxin that has been found frequently in rice. However, conventional analytical methods for determining LUT are limited, are complicated, and exhibit low sensitivity. In this study, an analytical method more sensitive and simple based on high-performance liquid chromatography combined with electrospray ionization mass spectrometry was developed. The cleanup procedure of the method was one step, using a solid-phase extraction cartridge. An isocratic mobile-phase system, consisting of acetonitrile-water-acetic acid (50:49:1 [vol/vol/vol]) at a flow rate of 0.2 ml/min, was utilized to obtain the best resolution. Our method showed good linearity (r = 0.9993, 0.5 to 50 ng/g) and high repeatability (relative standard deviation = 8.9 and 5.1% at levels of 0.5 and 10 ng/g, respectively) in the fortification test. The detection and quantification limits for the method in multiple-reaction monitoring mode were 0.1 and 0.3 ng/g, respectively. The average recovery of LUT in spiked rice at 0.5 and 10 ng/g was 80.7 and 85.2%, respectively. The method developed in this study should be applicable to survey LUT in rice, with high sensitivity, selectivity, and rapidity.

Rapid detection and identification of beer-spoilage lactic acid bacteria by microcolony method.

We evaluated a microcolony method for the detection and identification of beer-spoilage lactic acid bacteria (LAB). In this approach, bacterial cells were trapped on a polycarbonate membrane filter and cultured on ABD medium, a medium that allows highly specific detection of beer-spoilage LAB strains. After short-time incubation, viable cells forming microcolonies were stained with carboxyfluorescein diacetate (CFDA) and counted with muFinder Inspection System. In our study, we first investigated the growth behavior of various beer-spoilage LAB by traditional culture method, and Lactobacillus lindneri and several L. paracollinoides strains were selected as slow growers on ABD medium. Then the detection speeds were evaluated by microcolony method, using these slowly growing strains. As a result, all of the slowly growing beer-spoilage LAB strains were detected within 3 days of incubation. The specificity of this method was found to be exceptionally high and even discriminated intra-species differences in beer-spoilage ability of LAB strains upon detection. These results indicate that our microcolony approach allows rapid and specific detection of beer-spoilage LAB strains with inexpensive CFDA staining. For further confirmation of species status of detected strains, subsequent treatment with species-specific fluorescence in situ hybridization (FISH) probes was shown as effective for identifying the CFDA-detected microcolonies to the species level. In addition, no false-positive results arising from noise signals were recognized for CFDA staining and FISH methods. Taken together, the developed microcolony method was demonstrated as a rapid and highly specific countermeasure against beer-spoilage LAB, and compared favorably with the conventional culture methods.

Modified multiplex PCR methods for comprehensive detection of Pectinatus and beer-spoilage cocci.

Specific PCR primers were designed based on the 16S rRNA genes of recently proposed beer-spoilage species, Pectinatus haikarae, Megasphaera sueciensis, and M. paucivorans, and two sets of our previously reported multiplex PCR methods for Pectinatus spp. and beer-spoilage cocci were reconstructed. Each modified multiplex PCR method was found specifically to detect beer-spoilage species of Pectinatus and cocci, including new species.

Determination of ochratoxin A in wine by immunoaffinity cleanup and liquid chromatography tandem mass spectrometry.

A simple and accurate method has been developed for determining ochratoxin A (OTA), using an immunoaffinity column for cleanup and liquid chromatography-tandem mass spectrometry for identification and quantification. Wine samples were diluted with a solution containing polyethylene glycol 8000 and sodium hydrogen carbonate, filtered through a glass microfiber filter, and cleaned up on an immunoaffinity column. OTA was then eluted with methanol-acetic acid (98:2) and analyzed by liquid chromatography-tandem mass spectrometry. The average recoveries of OTA from red and white wines were 95 and 96.7% (spiked OTA level was 0.05 ng/ml) and repeatabilities (relative standard deviation) were 3.8 and 2.4%, respectively. The detection limit was 0.0003 ng/ml based on the signal-to-noise ratio in wine of 3:1. Analysis of 74 samples of domestic and imported wines showed OTA levels ranging from < 0.0003 to 0.82 ng/ml, with an incidence of contamination of 92.1% for red wines, and < 0.0003 to 0.51 ng/ml, with an incidence of contamination of 77.8% for white wines. These detection rates were higher than those rates of past reports of OTA contamination in wine, due to the high sensitivity of this method. However, all samples analyzed in this study complied with European Union regulations. It is concluded that this method is a useful tool for the quality assurance of wine.

Long-term outcomes of gastric mucosa-associated lymphoid tissue lymphomas after Helicobacter pylori eradication therapy.

Mucosa-associated lymphoid tissue (MALT) lymphomas are localized primarily in the gastrointestinal tract and are characterized by an indolent nature and favorable outcome with specific therapy. Gastric MALT lymphomas are closely linked to Helicobacter pylori (H. pylori) infection, for which eradication therapy is recognized as an effective primary treatment for the disease. However, there is little information about long-term outcomes after the therapy. In the present study, we elucidated the long-term outcomes of 74 patients (70 H. pylori-positive and 4 negative cases) followed up by endoscopy at least 12 months after exclusive eradication therapy alone. The median follow-up period was 46 months. When the remission status was estimated at the time point of 12 months post-eradication, the numbers of patients with complete remission (CR), histologically residual disease with macroscopic normalization (hRD), partial remission with more than 50% tumor reduction (PR) or no response (NR) were 56, 12, 2 and 4, respectively. During follow-ups of over 12 months post-eradication, 11 of the 12 hRD cases were belatedly induced to CR but one CR case histologically relapsed into hRD. One of the 2 PR cases eventually turned into hRD 20 months later. Therefore, 66 CR, 3 hRD, 1 PR, and 4 NR cases (including 3 H. pylori-negative) were identified at the last follow-up of the present study. All 74 patients were followed up without any second-line therapies, but none exhibited disease progression. Thus, the long-term outcome of localized gastric MALT lymphoma after H. pylori eradication therapy was favorable. A watch and wait strategy may be a reasonable approach for hRD since the majority might be in the process of turning into delayed CR.

Effects of morphological changes in beer-spoilage lactic acid bacteria on membrane filtration in breweries.

Membrane filter performance was investigated using beer-spoilage lactic acid bacteria (LAB). As a result, beer-adapted LAB strains showed considerably increased penetration rate through filters, as compared with non-adapted strains. Further statistical analyses demonstrated the significant shifts in cell size distribution towards shorter rods, when Lactobacillus brevis and L. lindneri strains were precultured in beer. These results indicate that diminished cell size is responsible for the deteriorated filter performance and, therefore, beer-adapted lactic acid bacteria are regarded as a serious threat to the production of unpasteurized beers. In addition, the selection of test strains and preculture conditions are suggested to be important for the rigorous and standardized evaluation of membrane filter performance in the brewing industry.

Preservation of gastric acid secretion may be important for the development of gastroesophageal junction adenocarcinoma in Japanese people, irrespective of the H. pylori infection status.

BACKGROUND: We have previously reported that Helicobacter pylori infection prevents reflux esophagitis (RE) and Barrett's esophagus (BE) by decreasing gastric acid secretion. Gastroesophageal (GE) junction adenocarcinoma, including Barrett's adenocarcinoma, has been thought to be a complication of gastroesophageal reflux disease (GERD). However, the relationship between H. pylori infection, gastric acid secretion, and GE junction adenocarcinoma has not yet been investigated in Japan. The aim of this study was to evaluate this relationship in the Japanese population. METHODS: A total of 168 Japanese patients (RE alone: 80, short-segment BE (SSBE): 16, long-segment BE (LSBE): 20, GE junction adenocarcinoma: 12, distal early gastric cancer (EGC): 40; male/female = 106/62; mean age 61.5 yr) and 80 Japanese control subjects who had no localized lesions in the upper gastrointestinal tract (male/female = 43/37, mean age 58.1 yr) were enrolled for this study. The prevalence of H. pylori infection was determined by biopsy, the rapid urease test, and measurement of the serum H. pylori IgG antibody. Gastric acid secretion was assessed by the endoscopic gastrin test (EGT). RE was diagnosed according to the Los Angeles classification. RESULTS: The prevalence of H. pylori infection in the patients with RE alone (30%) was significantly lower than that in control subjects (71.2%). There was also a tendency for the prevalence of H. pylori infection to be lower in patients with BE (SSBE, 18.7%; LSBE, 0%) when compared to that in patients with RE alone. On the other hand, while the prevalence of H. pylori infection in patients with GE junction adenocarcinoma (58.3%) was significantly lower than that in patients with EGC (87.5%), it tended to be higher than that in patients with RE alone or BE. The mean EGT value in patients with RE alone (3.74 mEq/10 min) was significantly higher than that in control subjects (1.83). The mean EGT value in patients with BE (SSBE, 4.74; LSBE, 4.76) tended to be even higher than that in patients with RE alone. The mean EGT value in patients with GE junction adenocarcinoma (3.94) was significantly higher than that in control subjects and patients with EGC (0.67), but it was comparable to that independent of the H. pylori infection status in patients with RE alone or BE. CONCLUSION: Preservation of gastric acid secretion may be important for the development of GE junction adenocarcinoma in Japanese people, irrespective of the H. pylori infection status.

Suppression by incadronate of invasion and growth of A-375 human melanoma in mandible in nude mice.

Frequent invasion of oral cancers into the neighboring jaw bones is a major cause of increased mortality. Moreover, majority of patients with these oral cancers suffer from functional inconveniences and esthetic disadvantages during clinical course and after surgical treatments. Understanding of the mechanisms underlying oral cancer invasion into jaw bones is, therefore, critical to the better management of oral cancer patients. To study the pathophysiology of mandibular invasion of oral cancers, we established an animal model by inoculating the A-375 human melanoma cells onto mandible through masseter muscle in nude mice. Histological study showed that A-375 tumor cells aggressively destroyed the mandibular cortical bones and invaded into the bone marrow cavities. Histochemical examination revealed that there were numerous tartrate-resistant acid phosphatase-positive osteoclasts on the residual bone surfaces facing A-375 tumors. To examine the role of osteoclasts in mandibular bone invasion of A-375 tumors, we next studied the effects of the bisphosphonate incadronate (INC), a potent inhibitor of osteoclastic bone resorption, using this model. Daily subcutaneous injections of INC (1 mg/kg) markedly decreased A-375-increased osteoclasts, increased numbers of apoptotic osteoclasts and inhibited mandibular cortical bone destruction. Furthermore, INC significantly suppressed the growth of A-375 tumors, increased numbers of apoptotic A-375 cells and decreased the tumor-associated blood vessel density. These results suggest that invasion and growth of oral cancers in mandibular bone is dependent on osteoclastic bone resorption and that INC can inhibit the aggressive behavior of oral cancer through inhibiting osteoclastic bone resorption.

[Reflux esophagitis after H. pylori eradication].

We have reported that prevalence of H. pylori infection and grade of atrophic gastritis were significantly lower, while acid secretion was significantly greater in the patients with reflux esophagitis than those without it. We have also reported that increased gastric acid secretion after H. pylori eradication may explain a reason for developing reflux esophagitis. Therefore, H. pylori is considered to be a protective factor for the development of reflux esophagitis via the induction of gastric hyposecretion. Conversely, according to recent large population-based studies, no evidence was obtained that supports the increased frequency of heartburn symptoms or reflux oesophagitis after the eradication of H. pylori. One reason for the discrepancy may be the difference in the evaluation of gastro-esophageal reflux, another reason may be the difference in acid secretion before the clearance of H. pylori in individual subjects, because the effects of H. pylori on acid secretion vary from patient to patient.

[Influence of urease activity of the oral cavity and oropharynx on 13C-urea breath test].

We examined whether 13C-urea breath test with powdered 13C-Urea is affected with urease activity in the oral cavity and oropharynx in H. pylori negative cases. We also examined whether the degree of gargling affects the results of 13C-UBT. The results demonstrated that the urease activity not only in the oral cavity, but in the oropharynx may influence the UBT value. Therefore, it is possible that the UBT value may be affected with relic urease activity in the oropharynx if only the oral cavity is gargled. In conclusion, Gargling of the oropharynx in addition to the oral cavity is important to obtain true negative diagnosis with 13C-UBT in patients without Helicobacter pylori infection.