Evaluation of 3-week-old layer chicks intratracheally challenged with Salmonella isolates from serogroup c1 (O:6,7) and Salmonella Enteritidis.

With the exception to Salmonella enterica serotype Typhimurium and S. Enteritidis (serogroups B [O:4] and D [O:9], respectively), there have been very few studies conducted on the respiratory tract as route of infection in chickens with salmonellas from serogroup C1 (O:6,7). Therefore, the purpose of this present study was to determine the potential organ invasion by Salmonella enterica serotype Potsdam (SP), S. Mbandaka (SM), and S. Infantis (SI) from serogroup C1 (O:6,7) and compare their characteristics with those of S. Enteritidis (SE) on intratracheally (IT) challenged 3-week-old layer chicks. A total of 360 one-day-old White Leghorn layer chicks were acquired from a commercial hatchery and randomly assigned into four treatment groups (SP, SM, SI, and SE, respectively), consisting of three independent trials. Chicks were grown up to 21 days (3 weeks) and IT-challenged thereafter with 106 CFU of respective salmonella organisms per group (n = 30). Chicks (n = 5) were humanely sacrificed every 24 h for 6 days post-IT infection and organs such as lung, heart, liver, spleen, kidney and caecal content were cultured for salmonella. All treatment groups exhibited colonization of lungs and caecal contents at 1 d (P = 0.475) and 4 d (P = 0.696) post-IT infection, respectively. There was no isolation of SP, SM, and SI in heart, liver, spleen, and kidney. In contrast, SE was obtained from heart, liver, and spleen of IT-infected chicks. The findings of this study contribute to a better understanding of the importance of the respiratory route in salmonella infection in poultry.

Detection of synchronized burst firing in cultured human induced pluripotent stem cell-derived neurons using a 4-step method.

Human induced pluripotent stem cell-derived neurons are promising for use in toxicity evaluations in nonclinical studies. The multi-electrode array (MEA) assay is used in such evaluation systems because it can measure the electrophysiological function of a neural network noninvasively and with high throughput. Synchronized burst firing (SBF) is the main analytic parameter of pharmacological effects in MEA data, but an accurate method for detecting SBFs has not been established. In this study, we present a 4-step method that accurately detects a target SBF confirmed by the researcher's interpretation of a raster plot. This method calculates one set parameter per step, in the following order: the inter-spike interval (ISI), the number of spikes in an SBF, the inter-SBF interval, and the number of spikes in an SBF again. We found that the 4-step method is advantageous over the conventional method because it determines the preferable duration of an SBF, accurately distinguishes continuous SBFs, detects weak SBFs, and avoids false detection of SBFs. We found also that pharmacological evaluations involving SBF analysis may differ depending on whether the 4-step or conventional threshold method is used. This 4-step method may contribute to improving the accuracy of drug toxicity and efficacy evaluations using human induced pluripotent stem cell-derived neurons.

Comprehensive molecular exploration identified promoter DNA methylation of the CRBP1 gene as a determinant of radiation sensitivity in rectal cancer.

BACKGROUND: Neoadjuvant chemoradiotherapy (NCRT) for advanced rectal cancer (RC) is a well-evidenced therapy; however, some RC patients have no therapeutic response. Patient selection for NCRT so that non-responsive patients are excluded has been subjective. To date, no molecular markers indicating radiation sensitivity have been reported. METHODS: We irradiated six colorectal cancer (CRC) cell lines and identified HCT116 cells as radiation-sensitive and HCT15 and DLD-1 cells as radiation resistant. Using a microarray, we selected candidate radiation sensitivity marker genes by choosing genes whose expression was consistent with a radiation-resistant or sensitive cell phenotype. RESULTS: Among candidate genes, cellular retinol binding protein 1 (CRBP1) was of particular interest because it was not only induced in HCT116 cells by tentative 10 Gy radiation treatments, but also its expression was increased in HCT116-derived radiation-resistant cells vs parental cells. Forced expression of CRBP1 decreased the viability of both HCT15 and DLD-1 cells in response to radiation therapy. We also confirmed that CRBP1 was epigenetically silenced by hypermethylation of its promoter DNA, and that the quantitative methylation value of CRBP1 significantly correlated with histological response in RC patients with NCRT (P=0.031). CONCLUSIONS: Our study identified CRBP1 as a radiation-sensitive predictor in RC.

The clinical significance of cysteine dioxygenase type 1 methylation in Barrett esophagus adenocarcinoma.

Methylation of cysteine dioxygenase type 1 (CDO1) gene, a tumor suppressor gene, has been studied in various cancers; however, there is no information regarding Barrett esophagus cancer. In this study, the clinical significance of CDO1 methylation in Barrett esophagus adenocarcinoma (BEA) was clarified. CDO1 gene promoter methylation was analyzed for DNA from the patient's specimens using quantitative methylation-specific polymerase chain reaction. Thirty-eight BEA patients who underwent resection were identified between 2000 and 2014. Hypermethylation of CDO1 gene was demonstrated to be frequently recognized even at early stage in BEA by quantitative methylation-specific polymerase chain reaction. In BEA, there is a robust prognostic difference between stage I and stage II/III/IV with regard to 5-year relapse-free survival (P = 0.0016) and 5-year overall survival (P = 0.0024), and the tumor size separated by 7 cm was also a prognostic factor. There was significant difference in CDO1 gene methylation according to the tumor size (P = 0.036). BEA patients with CDO1 gene methylation were shown marginally significantly poorer prognosis (P = 0.054) than otherwise patients. In conclusion, higher CDO1 gene methylation was seen in BEA at earlier stage than in squamous cell carcinoma, and it may account for aggressive phenotype of BEA.

Induction of long-term potentiation and depression phenomena in human induced pluripotent stem cell-derived cortical neurons.

Plasticity such as long-term potentiation (LTP) and long-term potentiation depression (LTD) in neuronal networks has been analyzed using in vitro and in vivo techniques in simple animals to understand learning, memory, and development in brain function. Human induced pluripotent stem cell (hiPSC)-derived neurons may be effectively used for understanding the plasticity mechanism in human neuronal networks, thereby elucidating disease mechanisms and drug discoveries. In this study, we attempted the induction of LTP and LTD phenomena in a cultured hiPSC-derived cerebral cortical neuronal network using multi-electrode array (MEA) systems. High-frequency stimulation (HFS) produced a potentiated and depressed transmission in a neuronal circuit for 1 h in the evoked responses by test stimulus. The cross-correlation of responses revealed that spike patterns with specific timing were generated during LTP induction and disappeared during LTD induction and that the hiPSC-derived cortical neuronal network has the potential to repeatedly express the spike pattern with a precise timing change within 0.5 ms. We also detected the phenomenon for late-phase LTP (L-LTP) like plasticity and the effects for synchronized burst firing (SBF) in spontaneous firings by HFS. In conclusion, we detected the LTP and LTD phenomena in a hiPSC-derived neuronal network as the change of spike pattern. The studies of plasticity using hiPSC-derived neurons and a MEA system may be beneficial for clarifying the functions of human neuronal circuits and for applying to drug screening.

Predictive index for the onset of medication overuse headache in migraine patients.

Migraine patients are particularly prone to develop medication overuse headache (MOH). However, the risk factors for the transformation of migraine to MOH are still not clear. We investigated gene polymorphisms, personality traits, and characteristics of headache and lifestyle in 47 migraine patients (aged 36.4 ± 10.3) and 22 MOH patients (aged 39.6 ± 9.9) who progressed from migraine and made a scoring system for a predictive index (PI) of the onset of MOH in patients with migraine. By multivariate logistic stepwise regression analysis, type of migraine, regular and sufficient dietary intake, and methylenetetrahydrofolate reductase (MTHFR) C677T (rs1801133) and dopamine D2 receptor (DRD2) C939T (rs6275) polymorphisms were selected as significant factors that contribute independently to the development from migraine to MOH (P < 0.05). The regression coefficients (ß) of these four selected factors were approximated and scored. The PI score in MOH patients (7.32 ± 1.60) was significantly higher than that in migraine patients (4.62 ± 1.83, P < 0.001). The proposed scoring system should in the future be the object of larger studies to confirm its validity.

Identification of a distinct lineage of Cacopsylla chinensis (Hemiptera: Psyllidae) in Japan on the basis of two mitochondrial DNA sequences.

Psyllids are a menacing pests of pears (Pyrus spp.) grown in temperate and subtropical regions of the world, including Taiwan and China. Pear psyllids belong to the large genus Cacopsylla (Psyllidae: Psyllinae). Among the 28 psyllid species that infest pear trees, Cacopsylla chinensis (Yang and Li, 1981) is considered the most harmful. Two psyllid outbreaks involving exotic species affected Japanese pear (Pyrus pyrifolia variety culta) orchards in the Saga Prefecture in July and November 2011. The psyllids were morphologically identical to the summer and winter forms of C. chinensis. In this study, we performed DNA sequence analysis of the mitochondrial cytochrome oxidase I-leucine tRNA-cytochrome oxidase II (COI-tRNA(Leu)-COII) and 16S rDNA regions to elucidate the phylogenetic relationships among 82 summer-form psyllids, five winter-form psyllids from several orchards in Japan, and those reported from Taiwan and China. The sequences of the COI-tRNA(LEU)-COII and 16S rDNA regions were identical among all 87 psyllids from Japan, regardless of summer/winter forms or orchards in Saga, Japan. Comparison of nucleotide sequences and phylogenetic analysis differentiated Japanese psyllids from the Taiwanese and Chinese C. chinensis isolates, with approximately 8 and 7% nucleotide difference in the COI-tRNA(LEU)-COII and 16S rDNA regions, respectively. The results suggest that C. chinensis possess a high level of genetic variability and that the psyllids responsible for the outbreak in Saga, Japan belong to a distinct lineage of C. chinensis.

Ku-band multichannel frequency comb Doppler reflectometer on the GAMMA 10/potential control and divertor simulating experiment (PDX) tandem mirror.

A Ku-band (12-18 GHz) multichannel Doppler reflectometer (DR) has been developed in the GAMMA 10/potential control and divertor simulating experiment (PDX) tandem mirror device to improve the applicability of DR measurement for simultaneous monitoring of velocity of electron density turbulence at different locations. Our previous single-channel DR circuit has been replaced by the multichannel microwave system using a nonlinear transmission line based comb generator with heterodyne technique. The multichannel DR system has been installed in the central cell of GAMMA 10/PDX. Initial results of application to GAMMA 10/PDX plasma are presented, showing Doppler frequency shifts during an additional ion cyclotron resonance frequency heating and gas-puffing experiment.

Prognostic significance of peritoneal tumour cells identified at surgery for colorectal cancer.

BACKGROUND: The prognostic significance of intraperitoneal tumour cells (IPCs) in colorectal cancer is not clear. This study aimed to determine whether detection of IPCs could be used a prognostic marker for selecting patients at high risk of recurrence. METHODS: The study included 226 patients with colorectal cancer who underwent elective resection. Clinical variables, including the presence of IPCs, were analysed for their prognostic significance. RESULTS: Thirty-three patients (14.6 per cent) were positive for IPCs. Univariable analysis indicated that the presence of IPCs was a significant prognostic factor in patients with stage III colorectal cancer; the 5-year disease-specific survival rate was 14 per cent in IPC-positive patients versus 79 per cent in those without IPCs (P < 0.001). Multivariable analysis showed that IPC positivity was the most robust prognostic factor in stage III disease (hazard ratio 2.2; P = 0.003), whereas nodal category (N1 or N2) showed no significant association with prognosis. In addition, IPCs were associated with haematogenous recurrence (P = 0.004) rather than peritoneal or local recurrence (P = 0.077) in patients with stage III disease. CONCLUSION: The presence of IPCs is a significant prognostic factor in patients with stage III colorectal cancer.

Structural and electronic properties of ZnO polycrystals doped with Co.

Zn(1-x)Co(x)O samples were prepared by a standard solid-state reaction method. Zn(1-x)Co(x)O crystals in the wurtzite structure were obtained with a Co composition of up to 22.1%. The a- and c-axis lengths increased and decreased, respectively, with an increase in Co composition. Raman spectra showed systematic broadening of the E(2) (high) phonon mode associated with the increase in Co composition, and electronic transitions of Co in the oxygen tetrahedron were observed in optical absorption measurement. These results indicated systematic substitution of Co into the Zn sites. Furthermore, an additional broad absorption band at 2.4-3.3 eV corresponding to the charge transfer (CT) process [Formula: see text] was also observed. The Raman spectra showed strong enhancement of the LO phonon due to a resonant Raman process induced with the coupling of the LO phonon and a photo-excited carriers mediated CT gap. These results suggest the possibility of carrier-induced ferromagnetism based on double exchange interaction in Zn(1-x)Co(x)O by visible light irradiation.

Solubilization of excess sludge in activated sludge process using the solar photo-Fenton reaction.

The solubilization of excess sludge by the solar photo-Fenton reaction has been investigated for the reduction of excess sludge in the activated sludge process. The solubilization kinetics depended on the dosages of the Fenton reagents, Fe and H(2)O(2). Increases of initial Fe and H(2)O(2) concentrations in their ranges studied in this work continuously enhanced the sludge solubilization. Cell lysis by the photo-Fenton reaction caused the increase in dissolved chemical oxygen demand (COD) in the first step of sludge solubilization. The further oxidative decomposition of the discharged organic compounds by the photo-Fenton reaction led to the decrease in the dissolved COD as the second step of sludge solubilization. The increase of dissolved COD in the first step of sludge solubilization and the consumption of H(2)O(2) could be described by the pseudo-zero order kinetics based on the accumulated light energy. About 40% reduction of mixed-liquor suspended solids (MLSS) by the solar photo-Fenton reaction was found. It was found that solar light used as a light energy source instead of costly and hazardous artificial UV light was very effective. The dissolved COD for solar photo-Fenton reaction increased faster and by 1.5 times as compared with that by artificial UV light.

Clinical application of the Fricke-glucomannan gel dosimeter for high-dose-rate (192)Ir brachytherapy.

This study investigates the efficacy of a new Fricke dosimeter formulation consisting of a standard Fricke gel dosimeter gelled with glucomannan (FrGDG). FrGDG was irradiated using a (192)Ir gamma-ray source with a remote afterloading system based on computed tomography images. (60)Co irradiation was performed for measuring the absorption of FrGDG and water. The distribution maps of T2 values from the irradiated containers were obtained by MR imaging and converted to the absorbed dose to visualize the dose distribution. We found that FrGDG was produced easily and quickly at room temperature. R2 (1/T2) values were reproducible and linearly correlated with the absorbed doses in the range from 0 to 30 Gy for irradiation with (192)Ir (the correlation coefficient was 0.99). The mean deviation between the doses obtained from the MR images of the FrGDG and those calculated by the treatment planning system for doses of 37.5, 40, 50, 62.5 and 75 Gy was 4.9%, 4.8%, 3.5%, 2.3% and 2.4%, respectively. In conclusion, MR imaging of FrGDG can visualize the dose distribution successfully, and thus serves as a useful quality assurance tool for complicated three-dimensional radiotherapy treatments.

Characterization of Ornithobacterium rhinotracheale from commercial layer chickens in eastern Japan.

From a total of 72 commercial layer and pullet farms that were monitored in the eastern Japan area, 4 farms had mild to severe respiratory disease accompanied by decreased feed intake and drop in egg production. Microbiological analysis showed that 3 of the 4 farms, particularly from Fukushima, Tochigi, and Ibaraki prefectures, were positive for Ornithobacterum rhinotracheale (ORT). Out of 65 birds examined, ORT was isolated in 21 birds (32.31%). All isolates were Gram-negative pleomorphic rods with a colony size of 0.05 mm, translucent with grayish coloration, and with butyric smell after 48 h of incubation in 10% chicken blood agar at 37°C under microaerophilic conditions. All isolates reacted positively in the p-nitrophenyl-ß-d-galactopyranoside test within 3 h and were positive in cytochrome oxidase tests with an API 20NE identification system biocode of 0-0-2-0-0-0-4. An agar gel precipitation test showed that all isolates were serotype-A. All strains were positive in PCR by yielding a 784 bp amplicon of the 16S rRNA gene. All strains were resistant to amikacin, colistin, gentamicin, kanamycin, neomycin, polymyxin b, streptomycin, and sulfamethoxazole trimethoprim and susceptible to amoxicillin clavulanic acid, ampicillin, doxycycline, spectinomycin, and tetracycline. This study is the first characterization of ORT from commercial layer chickens in eastern Japan.

Small GTPase RhoG is a key regulator for neurite outgrowth in PC12 cells.

The Rho family of small GTPases has been implicated in cytoskeletal reorganization and subsequent morphological changes in various cell types. Among them, Rac and Cdc42 have been shown to be involved in neurite outgrowth in neuronal cells. In this study, we examined the role of RhoG, another member of Rho family GTPases, in nerve growth factor (NGF)-induced neurite outgrowth in PC12 cells. Expression of wild-type RhoG in PC12 cells induced neurite outgrowth in the absence of NGF, and the morphology of wild-type RhoG-expressing cells was similar to that of NGF-differentiated cells. Constitutively active RhoG-transfected cells extended short neurites but developed large lamellipodial or filopodial structures at the tips of neurites. RhoG-induced neurite outgrowth was inhibited by coexpression with dominant-negative Rac1 or Cdc42. In addition, expression of constitutively active RhoG elevated endogenous Rac1 and Cdc42 activities. We also found that the NGF-induced neurite outgrowth was enhanced by expression of wild-type RhoG whereas expression of dominant-negative RhoG suppressed the neurite outgrowth. Furthermore, constitutively active Ras-induced neurite outgrowth was also suppressed by dominant-negative RhoG. Taken together, these results suggest that RhoG is a key regulator in NGF-induced neurite outgrowth, acting downstream of Ras and upstream of Rac1 and Cdc42 in PC12 cells.

Overexpression of a Gene Encoding a Catabolite Repression Element in Alternaria citri Causes Severe Symptoms of Black Rot in Citrus Fruit.

ABSTRACT A gene (AcCreA) encoding a catabolite repression element (CreA) with (two zinc fingers of the Cys(2)His(2) type was isolated from the postharvest fungal pathogen Alternaria citri. The AcCreA overexpression mutant AcOEC2 of A. citri showed normal growth on pectin medium and on segments of peel or the juice sac area from citrus fruit. Production of endopolygalacturonase, an essential virulence factor of this pathogen, was similar in AcOEC2 and the wild type in pectin-containing media. However, addition of glucose to the medium showed that carbon catabolite repression of endopolygalacturonase gene (Acpg1) expression, as well as endopolygalacturonase production, was lost in AcOEC2. The wild-type strain of A. citri causes rot mainly in the central axis of citrus fruit without development of rotting in the juice sac area; however, AcOEC2 caused severe black rot symptoms in both the central axis and juice sac areas. These results indicate that AcCreA-mediated catabolite repression controls the virulence or infection of this pathogen, and that the wild-type A. citri does not cause symptoms in the juice sac area due to carbon catabolite repression by sugars in the juice of the juice sac area.

Structural properties of nanometre-sized ZnO crystals doped with Co.

Nanometre-sized ZnO crystals doped with Co were synthesized by a co-precipitation method combined with a thermal treatment. By changing the reaction temperature, we can control the crystallite size from roughly 10 nm particles to 20 nm × 200 nm nm rods grown along the hexagonal c-direction. X-ray diffraction and Raman scattering showed growth of high-quality wurtzite ZnO crystals incorporating Co systematically in the ZnO host lattice in the tested range of [Co]<3.0 mol%. Electronic transitions of Co in the oxygen tetrahedron were also observed in optical absorption, giving supporting evidence for systematic substitution of Co into the Zn site.

Transcriptional targeting of adenovirus vectors with the squamous cell carcinoma-specific antigen-2 promoter for selective apoptosis induction in lung cancer.

Squamous cell carcinoma antigens SCCA1 and SCCA2 are highly homologous serine proteinase inhibitors which have been widely utilized as serological markers for squamous cell cancers, but it has recently been demonstrated that only SCCA2 is truly specific for certain forms of lung cancer. Using a construct containing the 5'-flanking region of the SCCA2 gene between -460 and +0 bp and the luciferase reporter gene, SCCA2 promoter activity was detected in SCCA2-producing SCC cell lines (LK-2, LC-1), but not in SCCA2-nonproducing lung adenocarcinoma cell lines (A549, ABC-1, and RERF-LC-MS) or normal cells (WI-38, SAEC, and NHEK-Adult). Infection with a recombinant adenovirus vector, Ad-SCCA2-DsRed, resulted in cell-specific expression of the SCCA2 promoter-driven DsRed marker gene only in LK-2 and LC-1 cells. The same strategy was used for SCCA2-driven expression of a proapoptotic gene, (KLAKLAK)2, which can cause mitochondrial disruption by triggering mitochondrial permeabilization and swelling, resulting in the release of cytochrome c and induction of apoptosis. Infection with Ad-SCCA2-KLAKLAK2 specifically reduced the growth of the two human lung SCC cell lines compared to the SCCA2 nonproducing cell lines both in vitro and in vivo, suggesting that the SCCA2 promoter had a tumor-specific effect. These results suggest that transduction of SCCA2 promoter-controlled suicide genes by adenoviral vectors can confer transcriptionally targeted cytotoxicity in SCCA2-producing lung SCC cells, and represents a novel strategy for gene transfer specifically targeted to SCC in the lung.

Effect of perioperative steroid therapy on the postoperative course of patients with oesophageal cancer.

BACKGROUND: Perioperative steroid therapy is often used in oesophageal cancer surgery and we evaluate the effect of this therapy on the secretory leukocyte protease inhibitor levels in the lungs (a major antiprotease in the conducting airways) and postoperative course in oesophageal cancer patients. METHODS: Twenty-one patients operated on for oesophageal cancer in 2003-2004 were treated with perioperative steroid therapy (250 mg of methylprednisolone intravenously 1 h before the operation). Fifteen consecutive patients operated on in 2002 served as a control group. Secretory leukocyte protease inhibitor in bronchoalveolar lavage fluid and the postoperative course in the two groups were compared. RESULTS: The mortality rate was 0% and there was no significant difference in the morbidity rate between the two groups. Days of intubation and systemic inflammatory response syndrome were significantly shorter for the steroid group. The bronchoalveolar lavage fluid secretory leukocyte protease inhibitor level was significantly higher in the steroid group than in the control group on postoperative days 2 and 3. The secretory leukocyte protease inhibitor level on postoperative day 3 was remarkably lower for the patients intubated for > or = 5 days and for those with pulmonary complications. CONCLUSION: Perioperative steroid therapy increased the bronchoalveolar lavage fluid secretory leukocyte protease inhibitor level and reduced the days of intubation and systemic inflammatory response syndrome in patients with oesophagectomy.

A Virulence-Reducing Mutation in the Postharvest Citrus Pathogen Alternaria citri.

ABSTRACT Alternaria citri causes Alternaria black rot, a postharvest fruit disease, on a broad range of citrus cultivars. We previously described that an endopolygalacturonase minus mutant of A. citri caused significantly less black rot in citrus fruit. To search for other essential factors causing symptoms in addition to endopolygalacturonase, a random mutation analysis of pathogenicity was performed using restriction enzyme-mediated integration. Three isolates among 1,694 transformants of A. citri had a loss in pathogenicity in a citrus peel assay, and one of these three mutants was a histidine auxotroph. Gene AcIGPD that encodes imidazole glycerol phosphate dehydratase, the sixth enzyme in the histidine biosynthetic pathway, was cloned, and the mutant containing the disrupted target gene, AcIGPD, caused less black rot.

Comparative study of barnidipine and felodipine in Chinese patients with essential hypertension.

This study evaluated the efficacy and safety of barnidipine for the treatment of mild-to-moderate essential hypertension in Chinese patients. A total of 131 patients were randomized to receive either barnidipine (10 -15 mg) or felodipine (5 - 10 mg) once daily for 4 weeks. Both drugs reduced blood pressure significantly, with > or = 87% of patients obtaining a marked or moderate effect. The mean +/- SD reductions in systolic and diastolic blood pressure were 19.2 +/- 13.6 and 14.4 +/- 7.0 mmHg, respectively, for barnidipine treatment, and 20.3 +/- 11.3 and 14.7 +/- 7.7 mmHg, respectively, for felodipine treatment. There were no significant differences between the two drugs in terms of anti-hypertensive effect, heart rate, laboratory test results or incidence of adverse events. More patients taking felodipine experienced palpitations, but this difference was not statistically significant. Barnidipine is as efficacious and safe as felodipine in the treatment of essential hypertension in Chinese patients.