Candida auris: A pathogen difficult to identify, treat, and eradicate and its characteristics in Japanese strains.

Candida auris is a multidrug-resistant and emergent pathogen that has caused healthcare-associated infection outbreaks. Recently, C. auris has spread worldwide; nevertheless, it was unexpectedly rare before 2009. Based on the molecular epidemiological analysis, C. auris may independently emerge at specific areas at first and recently may be transmitted to other continents. As C. auris cannot be detected using conventional methods, internally transcribed spacers, D1/D2 regions of the 26S rDNA sequencing, and/or matrix-assisted laser desorption ionization time-of-flight mass spectrometry method can be selected as comparatively accessible choices. Thus, detection of C. auris using the conventional method might be underestimated. In Japan, all C. auris strains were isolated from ear specimen and not from invasive mycoses. Japan strains were classified as an East Asian clade under a single clone. Although colonization, virulence, and infection pattern are almost the same as with other Candida species, its antifungal resistance is different. Fluconazole resistance is notably common, but resistance to all three classes of antifungals (azole, polyene, and echinocandin) rarely exists. Once C. auris is detected, screening, emphasis on hand hygiene adherence, use of single-patient room isolation, contact precaution, surveillance, and eradication from the environment and patients are appropriately required for infection control.

IMAGING URANIUM DISTRIBUTION ON RAT KIDNEY SECTIONS THROUGH DETECTION OF ALPHA TRACKS USING CR-39 PLASTIC NUCLEAR TRACK DETECTOR.

Uranium is renowned as a global contaminant, and attracts major concern with regards to the health risks involved because its nephrotoxicity. This paper discusses the development of a simple method to identify accumulated regions or localized sites of uranium within kidneys using the CR-39 plastic nuclear track detector. To demonstrate the proposed method, renal cryo-sections (5 µm-t) from Wistar male rats, subcutaneously administered with uranyl acetate (2 mg/kg), were prepared on day one after administration. Concerned sections were subsequently placed on CR-39, stored for 1.25 years, and then etched in a 7 M NaOH solution at 70°C for 3 h. α-tracks were then detected in the form of etch pits, corresponding to uranium, and also the tissue shape and structure were transferred as a roughness on the surface of CR-39. As observed, the proposed method served to facilitate simultaneous detection and identification of localized regions of uranium accumulation within kidneys.

Brain-derived neurotrophic factor regulates glucose metabolism by modulating energy balance in diabetic mice.

We previously reported that brain-derived neurotrophic factor (BDNF) regulates both food intake and blood glucose metabolism in rodent obese diabetic models such as C57BL/KsJ-lepr(db)/lepr(db) (db/db) mice. To elucidate the effect of BDNF on glucose metabolism, we designed a novel pellet pair-feeding apparatus to eliminate the effect of appetite alteration on glucose metabolism. The apparatus was used to synchronize food intake precisely between BDNF-treated and vehicle-treated db/db mice. It was shown using this pellet pair-feeding apparatus that BDNF administered daily (20 mg x kg(-1) x day(-1)) to db/db mice significantly lowered blood glucose compared with pellet pair-fed db/db mice. To evaluate the effect of BDNF on insulin action, we used streptozotocin-induced type 1 diabetic mice. In this case, BDNF did not lower blood glucose concentration but rather enhanced the hypoglycemic action of insulin. In hyperglycemic db/db mice, pancreatic insulin content was reduced and glucagon content was increased compared with normoglycemic db/m mice. BDNF administered to db/db mice significantly restored both pancreatic insulin and glucagon content. Histological observations of aldehyde-fuchsin staining and immunostaining with anti-insulin indicated that insulin-positive pancreatic beta-cells were extensively regranulated by BDNF administration. We also studied the effect of BDNF on KK mice, normoglycemic animals with impaired glucose tolerance. In these mice, BDNF administration improved insulin resistance in the oral glucose tolerance test. To elucidate how blood glucose was metabolized in BDNF-treated animals, we investigated the effect of BDNF on the energy metabolism of db/db mice. Body temperature and oxygen consumption of the pellet pair-fed vehicle-treated mice were remarkably lower than the ad libitum-fed vehicle-treated mice. Daily BDNF administration for 3 weeks completely ameliorated both of the reductions. Finally, to clarify its action mechanism, the effect of intracerebroventricular administration of BDNF on db/db mice was examined. Here, a small dose of BDNF was found to be effective in lowering blood glucose concentration. This indicates that BDNF regulates glucose metabolism by acting directly on the brain.

Basaloid-squamous carcinoma of the esophagus. A clinicopathologic, DNA ploidy, and immunohistochemical study of seven cases.

Basaloid-squamous carcinoma (BSC) of the esophagus is a rare but interesting neoplasm that occurs primarily in the upper aerodigestive tract. In this study, we reviewed 371 cases of esophageal malignancies and detected seven cases (1.9%) of BSC. The clinicopathologic features, light and electron microscopic findings, and immunohistochemical localization of various differentiation-related antigens, including cytokeratin (CK) subtypes, p53, and epidermal growth factor receptor (EGFR), were examined. DNA ploidy was also determined in an effort to characterize the biologic features of these tumors. The tumors were classified as stage I (n = 1), IIB (n = 3), III (n = 2) or IV (n = 1). Six patients had lymph node metastasis, in four the metastatic carcinoma exhibited basaloid components. Histologically, all the tumors displayed a biphasic pattern of basaloid and squamous components. The former predominated in three cases, the latter in four cases. All the tumors contained solid growth of basaloid cells with microcystic patterns and stromal hyalinosis as well as palisading of cells. Ultrastructurally, markedly replicated basement membrane was observed. Immunohistochemistry revealed staining with only CK 14 and CK 19 antibodies in the periphery of the basaloid tumor nests. These antibodies were also positive in the basal layer of normal esophagus. Diffuse immunoreactivity for EGFR was demonstrated in all the tumors. Five tumors displayed p53 nuclear immunoreactivity. All of the basaloid components demonstrated aneuploidy by DNA image cytometry. We conclude that BSC is a distinct type of esophageal carcinoma that should be differentiated from the usual types of esophageal carcinoma and may be associated with aggressive biologic behavior.

Autocrine stimulation of motility in SBC-5 human lung carcinoma cells by a two-kringle variant of HGF.

Hepatocyte growth factor (HGF), a ligand for the c-met protooncogene product, is a pleiotropic cytokine which elicits mitogenic, motogenic and morphogenic activities. Among various human lung carcinoma cells, we found that SBC-5 small cell lung carcinoma cells simultaneously expressed the c-Met/HGF receptor and a smaller variant-type of HGF composed of N-terminal two-kringle domains, without expressing authentic heterodimeric HGF. The addition of anti-HGF antibodies to cultures of SBC-5 cells specifically inhibited spreading and motility of the cells without affecting growth, and the conditioned medium of SBC-5 cells also induced scattering of other lineage lung carcinoma. Thus, simultaneous expression of the unique smaller variant HGF and its receptor, c-Met, in SBC-5 cells suggests the involvement of a smaller variant HGF in the development or progression of the lung carcinoma cells, through an autocrine mechanism.

Osteocompatibility of platinum-plated titanium assessed in vitro.

Suppression of dissolution is important to increase the biocompatibility of titanium implants. Therefore, the possibility of application of platinum-coated titanium as a biomaterial was explored in in vitro experiments using the MC3T3-E1 osteogenic cell line. The data obtained from long-term cultures indicated that pure platinum or titanium thickly coated with platinum inhibited calcification significantly, suggesting that the platinum ion fails to improve the osteocompatibility of titanium implants.

Intermittent administration of brain-derived neurotrophic factor ameliorates glucose metabolism in obese diabetic mice.

We have previously shown that brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family, interacts with the endocrine system in obese diabetic mice, and systemic peripheral administration of BDNF regulates glucose metabolism in this model. Results from the present study show that the hypoglycemic effect induced by 2 weeks' daily administration of BDNF (20 mg/kg/d) to db/db mice lasts for several weeks after treatment cessation, irrespective of food reduction. On the other hand, the antidiabetic agent, metformin had no lasting effect. This duration of the BDNF hypoglycemic action prompted us to examine the efficacy of BDNF intermittent administration on glucose metabolism. BDNF administered once or twice per week (70 mg/kg/wk) to db/db mice for 3 weeks significantly reduced blood glucose concentrations and hemoglobin A(1c), (HbA(1c)) as compared with ad libitum-fed phosphate-buffered saline (PBS)-treated and pair-fed PBS-treated groups. This suggests that BDNF not only temporarily reduced blood glucose concentrations but also ameliorated systemic glucose balance in this obese diabetic mouse model during the experimental period. Our results indicate that BDNF could be a novel hypoglycemic agent with an exceptional ability to normalize glucose metabolism even with treatment as infrequently as once per week.

Premature centromere division in patients with multiple endocrine neoplasia type 1.

Frequency of mitoses with premature centromere division (PCD) was examined in lymphocytes from subjects with multiple endocrine neoplasia type 1 (MEN 1). An increase in PCD after exposure to an alkylating agent was observed in subjects with MEN 1 who carry a heterozygous MEN1 gene mutation but not in normal controls or in affected subjects without the MEN1 gene mutation. These findings support the inclusion of MEN 1 as a chromosome instability syndrome and recognition of PCD as a manifestation of chromosome instability. Furthermore, these results suggest that the MEN1 gene product may function to maintain the integrity of DNA.

Antioxidant effects of fructosyl arginine, a Maillard reaction product in aged garlic extract.

The amino-carbonyl (Maillard) reaction of amino acids with sugars is a nonenzymatic browning reaction that takes place during the processing, cooking, and storage of foods. Maillard reaction products (MRPs) have been shown to possess interesting chemical and biological properties including antimutagenic and antioxidant activity. In this study, we determined the antioxidant effects of fructosyl arginine (Fru-Arg), a MRP in aged garlic extract. Low density lipoprotein (LDL) was incubated with Cu(2+) at 37 degrees C and 5% CO(2) for 24 hours, which resulted in an increase of thiobarbituric acid reactive substances (TBARS) indicating lipid peroxidation. Coincubation of Cu(2+) with Fru-Arg and LDL resulted in a significant inhibition of TBARS formation. Pulmonary artery endothelial cells (PAEC) were exposed to 0.1 mg/mL oxidized LDL (Ox-LDL) at 37 degrees C and 5% CO(2) for 24 hours. Lactate dehydrogenase (LDH) release, as an index of cell membrane damage, and TBARS were measured. Ox-LDL caused an increase of LDH release and TBARS formation. Pretreatment of PAEC with Fru-Arg inhibited these changes. Murine macrophages were incubated with Ox-LDL, and the release of peroxides was measured using a fluorometric assay. Ox-LDL caused an increased release of peroxides. Coincubation of macrophages with Fru-Arg and Ox-LDL inhibited the release of peroxides dose-dependently. In a cell free system, Fru-Arg was shown to scavenge hydrogen peroxide. These data suggest that Fru-Arg is a potent antioxidant, and thus may be useful for the prevention of atherosclerosis and other disorders associated with oxidative stress.

Absence of argininosuccinate lyase protein in the liver of two patients with argininosuccinic aciduria.

The enzyme defects in two cases of argininosuccinic aciduria were examined at the molecular level by enzymatic and immunological methods. No argininosuccinate lyase activity was detected in the liver or erythrocytes of either patient nor in the kidney or brain of one of the patients even in the presence of high concentrations of the substrate. The titration curve of antiserum to human argininosuccinate lyase with the liver extract from a control subject was not affected by the addition of the liver extracts from one of the patients. Double immunodiffusion analysis revealed a single precipitin line between the purified antiserum and the liver extract from a control, but no precipitin lines between the antisera and the liver extracts from the two patients. These results indicate a complete or almost complete defect of an immunologically cross-reactive material in the liver of the patients.

Increased platelet activating factor in the tracheal aspirates from neonates with patent ductus arteriosus.

We investigated platelet-activating factor (PAF) in the tracheal aspirate from 3 intubated low birth weight infants with symptomatic patent ductus arteriosus (PDA). PAF increased with the onset of symptomatic PDA and decreased to the control range soon after the ductal closure. The concentration of PAF in 26 samples taken during symptomatic PDA (median 16 pg/micrograms lipid phosphorus, range 1.4-1,200 pg/micrograms lipid phosphorus) was significantly higher than that of 31 samples from the same three patients during the periods without symptomatic PDA (median 1.9 pg/micrograms lipid phosphorus, range 0-12 pg/micrograms lipid phosphorus; P < 0.001). All 3 infants later developed chronic lung disease. These results suggest that large shunting PDA provokes PAF release to the air way of the neonate and that PAF might play a role in chronic lung disease developing after symptomatic PDA.

Tannins occurring in the toxic Brazilian plant Thiloa glaucocarpa.

Four tannins were isolated from dried leaves of Thiloa glaucocarpa, which is one of the plants causing poisoning of cattle in Brazil. These four tannins were determined from spectrometric and chemical evidence to be the ellagic C-glucosyl tannins vescalagin, castalagin, stachyurin and casuarinin. The main components, vescalagin and castalagin, are also known as constituents of oak (Quercus spp.), the toxicity of which for cattle is supposed to be due to tannins. Suggestions concerning the resemblance of the symptoms of poisoning by Thiloa glaucocarpa and by oak are supported by the presence of the constituents in both plants.

Allixin, a phytoalexin produced by garlic, and its analogues as novel exogenous substances with neurotrophic activity.

Effects of allixin, a phytoalexin of garlic, and its analogues were studied on the survival and morphology of primary cultured neurons from fetal rat brain. Addition of allixin (1-100 ng/ml) to medium significantly promoted the survival of neurons derived from various regions of brain and increased the number of branching points per axon in hippocampal neurons. Allixin, however, was cytotoxic at higher concentrations (>1 microg/ml). Among the analogues of allixin, 2,6-dimethyl-3-hydroxy-4H-pyran-4-one (DHP) possessed potent neurotrophic activity at concentrations over 10 ng/ml without any obvious cytotoxicity up to 10 microg/ml. DHP also retained the activity to promote axonal branching. These results indicate that DHP is a novel exogenous low molecular weight neurotrophic substance without apparent cytotoxicity. This compound may be a useful prototype leading chemical for developing therapeutic and/or prophylactic drugs for neurodegenerative disorders.

Tissue subtraction DNA in situ hybridization (TSDISH): in situ detection of DNA abnormalities in formalin-fixed paraffin-embedded tissue sections by subtraction hybridization with whole genomic DNA.

We have developed a novel in situ histochemical method of screening for genetic alterations of human malignancies by subtraction hybridization of genomic DNA without employing specific probes to give a colorimetric reaction. We identified a t(13;14) chromosome abnormality in the chromosome spread of a patient with multiple myeloma. In situ hybridization of a whole cell preparation of MCF-7 cell demonstrated reaction products as intranuclear dots-in all MCF-7 cells. We subsequently examined the cells of different foci of esophageal squamous cell carcinoma(10) and esophageal biopsy specimens (9) by this method. Hybridization with genomic DNA from the patients demonstrated no reaction products in the stromal cells of the esophagus. However, hybridization with reference DNA from a healthy individual demonstrated intranuclear reaction products in the stromal cells, possibly due to individual genomic differences. There were more intranuclear reaction products in the carcinoma cells than in the stromal cells when hybridized with reference DNA. When hybridized with the reference DNA above, the cells of the non-pathologic epithelium of 8 of 10 malignant esophagi demonstrated significantly more reaction product than the stromal cells of the some specimens. This was not detected in the cells of normal epithelium obtained from non-cancerous esophagi suggesting the accumulation of genetic alterations of the non-malignant epithelium of the cancerous esophagus. This method is thought to detect DNA alterations, including those which have not been previously identified, using genomic DNA for hybridization, and the results can be correlated with the morphological findings. Application of this in situ method, together with other molecular genetic techniques may contribute to the analysis of various genetic alterations of human malignancies using archival material.

DNA ploidy, P53 expression, and cellular proliferation in normal epithelium and squamous dysplasia of non-cancerous and cancerous human oesophagi.

Ki67 expression, S-phase fraction, p53 immunoreactivity and DNA content were examined in morphologically normal mucosa and squamous dysplasia of both cancerous and non-cancerous human oesophagi in order to understand possible early events in the development of esophageal squamous cell carcinoma. 103 different foci from cancerous esophagi including 17 non-pathological epithelium, 10 mild, 17 moderate and 15 severe dysplasia, 14 intraepithelial carcinomas and 30 invasive squamous cell carcinomas were examined. Also studied were 57 biopsy specimens from cancer-free individuals, including 12 normal epithelia, 15 oesophagitis, and 16 mild, 11 moderate and 3 severe dysplasia. Areas of squamous dysplasia from both cancer-free and cancerous oesophagi were morphologically indistinguishable and both demonstrated increased cellular proliferation compared to normal or non-pathological epithelia. However, squamous dysplasia in cancerous oesophagi demonstrated significantly larger ki67 labelling indices and smaller S-phase fractions than dysplasia in cancer-free patients. Squamous dysplasia in cancerous and non-cancerous oesophagi demonstrated an non-diploid DNA histogram in 67.9% and 43.3% respectively. However, dysplasia from cancer-free individuals demonstrated a non-diploid pattern with one or more peaks (Type I non-diploid histogram) and that from oesophageal cancer patients predominantly exhibited non-diploid histograms without any distinctive peaks (Type II non-diploid histogram). Significant differences in the frequency of p53 positive foci were observed between dysplasia of cancer-free (23.3%) and cancerous (56.8%) oesophagi. IN cancerous oesophagi, dysplasia associated with Type II non-diploid histograms had a significantly larger number of p53-positive foci than those with diploid histograms or Type I non-diploid histograms. These results indicated that the biological features of squamous dysplasia were different between cancerous and non-cancerous human oesophagi despite indistinguishable morphological features. In addition, the combination of p53 immuno-histochemistry and DNA ploidy analysis may contribute to identify possible high-risk squamous dysplasia of the oesophagus.

Enhanced sensitivity to alkylating agent in lymphocytes from patients with multiple endocrine neoplasia type 1.

Chromosome instability is known to be associated with certain autosomal recessive cancer-prone disorders such as Fanconi's anemia. Multiple endocrine neoplasia type 1 (MEN 1) is an autosomal dominant disorder characterized by development of tumors in two or more endocrine organs, and chromosome instability in patients with MEN 1 has been described. The clinical features of MEN 1 are, however, distinct from other DNA instability syndromes except predisposition to tumors. Therefore, we reevaluated chromosome instability in patients with familial MEN 1. An increase in the frequency of chromosome aberrations was observed in MEN 1 patients but not in control subjects when peripheral mononuclear cells were exposed to an alkylating agent, diepoxybutane (DEB). DEB reduced survival of mononuclear cells in a dose-dependent manner in both MEN 1 patients and control subjects, but this effect was more prominent in MEN I patients. There was no apparent correlation between certain MEN1 gene mutations and sensitivity to DEB. From these results, we conclude that hypersensitivity to alkylating agents exists in patients with MEN 1. Molecular mechanisms of this phenomenon and relationship to tumorigenesis in endocrine organs should be elucidated.

Severe infantile congenital myopathy with nemaline and cytoplasmic bodies: a case report.

A Japanese boy had marked generalized hypotonia and weakness and progressive respiratory failure since birth. Left biceps brachii muscle biopsy at 47 days of age showed marked variation in muscle fiber size, and nemaline and/or cytoplasmic bodies in approximately 10% of the muscle fibers. To our knowledge, the presence of nemaline and cytoplasmic bodies in the same muscle has not been previously reported. The severity of his respiratory failure and muscle weakness were thought to be related to muscle immaturity since there were many undifferentiated type 2C fibers.

Polymorphism of 17 STRs by multiplex analysis in Japanese population.

Genotype and distribution of allele frequencies at 17 STRs were studied in 526 unrelated Japanese individuals using the PowerPlex 16 system and the AmpFlSTR Identifiler.

Novel detection of plankton from lung tissue by enzymatic digestion method.

Studies are reported on the enzymatic digestion method for detection of plankton from lung tissue by using proteinase K with sodium dodecyl sulphate. This method is simple, safe and effective for detection of not only phytoplankton including diatoms but zooplankton which are destroyed by the acid digestion method. The present method is, therefore, much more advantageous for diagnosis of drowning than the disorganization method using strong acids.

Perinatal lung function measurements and prediction of respiratory problems in infancy.

The aim of this study was to determine which lung function test employed in the perinatal period gave the results most significantly associated with respiratory problems in infancy. The ratio of the proportion of time to reach peak tidal expiratory flow to total expiratory time (tPTEF:tE), thoracic gas volume (TGV) and airway resistance (R(aw)) (from which specific conductance (SG(aw)) was calculated) measurements were examined from 85 infants born at or near term. The infants were followed until at least one year of age and described as symptomatic if they wheezed for at least 24 hours. Twenty-three infants were symptomatic in the first year. The symptomatic group, compared to the asymptomatic, had a higher median FRC (p < 0.01) and R(aw) (p < 0.001); their median SG(aw) was lower (p < 0.001). It was possible to obtain tPTEF:tE results from only 61 infants; the median tPTEF:tE did not differ significantly between symptomatic and asymptomatic infants. Logistic regression analysis demonstrated a high R(aw) and FRC, but not a low tPTEF:tE, independently related to positive symptom status. A high R(aw) (>26 cm H2O (1 s(-1))(-1)) was the most sensitive (83%) predictor of subsequent respiratory problems, but all the tests examined had low positive predictive values.