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High-resolution remote sensing technology is an efficient and low-cost space-to-earth observation strategy, which can carry out simultaneous monitoring of large-scale areas. It has incomparable advantages over ground monitoring solutions. Traditional road extraction methods are mainly based on image processing techniques. These methods usually only use one or a few features of images, which is difficult to fully deal with the real situation of roads. This work proposes a two-steps network for the road extraction. First, we optimize a pix2pix model for image translation to obtain the required map style image. Images output by the optimized model is full of road features and can relief the occlusion issues. It can intuitively reflect information such as the position, shape and size of the road. After that, we propose a new FusionLinkNet model, which has a strong stability in the road information by fusing the DenseNet, ResNet and LinkNet. Experiments show that our accuracy and learning rate have been improved. The MIOU (Mean Intersection Over Union) value of the proposed model in road extraction is over 80% in both DeepGlobe and Massachusetts road dataset. The figures are available from https://github.com/jsit-luwei/training-dataset.

COVID-19 is a global pandemic. In response to this unprecedented crisis, Chinese government formulated a series of policies. This research is dedicated to exploring the dynamic evolution of China's policy mix in response to COVID-19 in different crisis response stages from a network perspective. First, a three-dimensional analysis framework of "policy subject-policy target-policy instrument" was developed. Then, based on the data sets collected by textual analysis, the dynamic evolution of policy subject, policy target, policy instrument in China's policy mix in response to COVID-19 was discussed by using the method of SNA. This study concluded that the core policy subject, policy instrument, and policy target of China's response to COVID-19 changed with time. National Health Commission (NHC), Ministry of Finance (MOF), Ministry of Transport (MOT) and Ministry of Human Resources and Social Security (MHRSS) have important influences in the network of policy subjects. Other subjects are more at the edge of the network, and there are few joint issuances among policy subjects. The study also found that the core policy target was adjusted over time, with phased dynamic characteristics. At the initial stage of China's response to COVID-19, "reduce infection and mortality" and "steadily carry out economic and social work" were the core policy targets. With the COVID-19 under control, "enterprise development and work resumption" becomes a new core policy target. In addition, this study also revealed the dynamic evolution and unbalanced use of China's policy instruments in response to COVID-19 in different stages. The combination of policy instruments is mainly composed of "mandatory administration instruments" and "economic incentive instruments", and supplemented by "health promotion instruments" and "voluntary plan instruments". These findings may enrich the literature on COVID-19 policy to help researchers understand the dynamics of policy from a network perspective. Moreover, these findings may provide several valuable implications for policymakers and other countries to formulate more effective policies for epidemic response.

The plastic hinge is a key factor in the ductile and plastic design of structures and an important basis for the seismic strengthening of structures. The formation and behavior of plastic hinges is critical for the seismic performance of an entire structure. The relocation of plastic hinges away from the beam end is an effective way of addressing brittle failure. In this paper, the cause of the shear buckling failure of prefabricated steel beams with corrugated webs and the strain variation at the flanges of steel beams are theoretically analyzed through structural tests. Based on the analysis results, a local strengthening method is proposed, and the effects of the beam sizes and the strengthening steel plate on the plastic hinges are obtained. In addition, a calculation method for determining the size of the strengthening steel plate that promotes the relocation of the plastic zone away from the beam end is given, a design method for plastic hinge relocation is proposed based on the test data, and its validity is verified.

ObjectivesHCV infection status awareness is crucial in the HCV care continuum for both HCV-seropositive (HCV-positive status awareness) and seronegative (HCV-negative status awareness) populations. However, trends in the unawareness of HCV infection status (UoHCV) remain unknown in HIV-positive patients. This study investigated UoHCV prevalence, the associated factors of UoHCV, and its association with HCV-related knowledge in HIV-positive patients.MethodsFor this cross-sectional, multicenter, questionnaire-based study, 844 HIV-infected participants were recruited from three hospitals in Taiwan from June 2018 to March 2020. Participants were grouped by HCV serostatus (HCV-seronegative [n = 734] and HCV-seropositive [n = 110]) and categorized by their HIV diagnosis date (before 2008, 2008-2013, and 2014-2020). Exploratory factor analysis was used to categorize the 15 items of HCV-related knowledge into three domains: route of HCV transmission, HCV course and complications, and HCV treatment.ResultsThe prevalence of UoHCV was 58.7%-62.6% and 15.1%-31.3% in the HCV-seronegative and HCV-seropositive groups, respectively, across 3 periods. More participants with UoHCV believed that HCV infection was only contracted by intravenous injection. In the HCV-seropositive group, participants with UoHCV were more likely to have HIV diagnosis before 2008 (vs. 2014-2020), be men who have sex with men (vs. people who inject drugs), and have hepatitis A virus seronegativity. In the HCV-seronegative group, participants with UoHCV were more likely to have a recent history of sexually transmitted diseases, but had a lower education level, had received less information on HCV infection from clinicians, and were less likely to have heard of HCV infection prior to the research. UoHCV was associated with lower scores for three domains of HCV-related knowledge in both groups.ConclusionsThe negative association of UoHCV with HCV-related knowledge suggests that strategies targeting patients according to their HCV serostatus should be implemented to reduce UoHCV and eradicate HCV infection among HIV-positive patients.

IntroductionThis retrospective study investigated the implications of changes in blood parameters and cellular immune function in patients with Coronavirus Disease 2019 (COVID-19).MethodsRecords were reviewed of 85 patients admitted with COVID-19 between February 4 and 16, 2020. The primary outcome was in-hospital death.ResultsFourteen patients died. The baseline leukocyte count, neutrophil count and hemoglobin was significantly higher in non-survivors compared with survivors, while the reverse was true of lymphocyte count, platelet, PaO2/FiO2, CD3+ count and CD4+ count. The percentage of neutrophil count > 6.3×109/L in death group was significantly higher than that in survival group, and multivariate logistic regression showed neutrophil count > 6.3×109/L was independently associated with mortality. However, there were not significant difference in IgG, IgM, IgA, C3, C4 and the percentage of IgE > 100 IU/ml between the death group and survival group. Areas under the receiver operating characteristic curves of the following at baseline could significantly predict mortality: leukocyte, neutrophil, lymphocyte, CD3+ and CD4+ counts.ConclusionsFor hospitalized patients with COVID-19, lymphocyte, CD3+ and CD4+ counts that marked decrease suggest a poor outcome. Admission neutrophil count > 6.3 ×109/L is independently associated with mortality. At admission, leukocyte, neutrophil, lymphocyte, CD3+ and CD4+ counts should receive added attention.

BackgroundChina adopted family doctor (FD) to help achieve "Healthy China 2030" through providing continuous, comprehensive, and life-cycle contract services. However, there is a disparity between actual and targeted FD use, as residents continue to visit specialists in large hospitals. The government implemented initiatives to improve residents' willingness to sign up with and visit their FDs. Factors that influence contract behavior are therefore significant for frontier policy research.MethodsTwo survey waves were conducted in Shanghai (2013 and 2016). The first wave included 2754 people and the second 1995 people. Exploratory factor analysis was used to synthesize "satisfaction" as a predictor of contract behavior. Pearson's chi-square, pooled and logistic regression models were used to estimate associations between influencing factors and contract behavior, and clarify variations in factors across the two waves.ResultsFour factors were extracted from 15 satisfaction items: "Treatment Environment," "Medical Technology," "Service Specification" and "Service Attitude". Consistent with descriptive analysis, longitudinal analysis showed sociodemographic characteristics (age, education, marital status, and hukou) were significant predictors of contract behavior. The odds ratio of non-communicable diseases (NCD) patients for contract behavior was 2.218 times that of residents without NCD. Contract behavior was positively correlated with awareness of FD services (OR = 21.674, 95%CI = 15.043-31.229), satisfaction with Service Attitude (OR = 1.210, 95%CI = 1.009-1.451), and visit compliance (OR = 1.959, 95%CI = 1.564-2.452). Over time, the odds ratios of the married, Shanghai hukou, NCD, and awareness of FD services declined from 0.456, 1.795, 2.492, 28.690 to 0.443, 1.678, 1.910 and 14.031 respectively, while those of age, and visit compliance increased from 1.027, 1.521 to 1.041 and 2.305 respectively. In 2016, an education-contract gradient had formed (the higher the education level, the higher probability of signing with a FD), whereas high school education had the highest odds ratio (OR = 1.163,95%CI = 0.740-1.827) in 2013. Service Attitude was the only significant satisfaction-related predictor (OR = 1.358, 95%CI = 1.001-1.842) in 2016, compared with "Treatment Environment" (OR = 1.224, 95%CI = 1.001-1.496) and "Service Specification" in 2013(OR = 1.270, 95%CI = 1.040-1.552).ConclusionsExcept for the socio-demographic variables, NCD, awareness of FD services, satisfaction and visit compliance were significant predictors of contract behavior with FDs. The effect of visit compliance had increased over time while NCD and awareness of FD services were losing impact over time. Significant satisfaction factors had also changed from "Treatment Environment" and "Service Specification" to "Service Attitude".

Aphids are major pests of cotton crops in the Xinjiang Uygur Autonomous Region in China, and parasitoids are considered as important natural enemies in regulating aphid populations. However, information on aphid parasitoids in the Xinjiang cotton fields is limited, which hinders the study of aphid-parasitoid interactions and the application of conservation biological control against cotton aphids. In this study, a 3-year survey was conducted in a large geographical range that included three primary cotton planting areas in southern and northern Xinjiang. The population dynamics and the parasitism levels of an assemblage of aphids in the cotton fields were investigated along with the composition of the parasitoid community associated with these aphids. Aphid parasitization varied significantly within both years and seasons, with parasitism levels ranging from 0 to 26%, indicating that there is less effective biological control of parasitoids on aphids under field conditions. Among the primary parasitoids described, Binodoxys communis (Gahan) constituted 95.19% of the parasitoid species, followed by Praon barbatum Mackauer (3.15%), Trioxys asiaticus Telenga (1.01%) and Lysiphlebus fabarum Marshall (0.65%). Significant differences were found in the composition of the primary parasitoid species between the cotton seedling period (June) and the flowering period (July-August), and two more primary aphid parasitoids were found in the seedling period. Twelve hyperparasitoid species belonging to six genera were found in our study, of which Pachyneuron aphidis (Bouché), Syrphophagus species and Dendrocerus laticeps (Hedicke) were the dominant species. The composition of the hyperparasitoid community also differed significantly between the seedling and the flowering periods. The description of this parasitoid community-associated assemblage of aphids in cotton fields will facilitate the study of aphid-parasitoid interactions and promote the development of effective cotton aphid management strategies in Xinjiang.

OBJECTIVE:Periodontitis is a microbe-induced chronic inflammatory disease. Previous exposure of the host to bacteria or their virulence factors leads to refractory responses to further stimuli, which is called tolerance. Porphyromonas gingivalis (P. gingivalis) is one of the most important pathogenic microorganisms associated with periodontitis, and is a potent inducer of pro- and anti-inflammatory cytokines. The aim of this study was to explore the roles and possible mechanisms of tolerance induced by P. gingivalis. METHODS:THP-1-derived macrophages were pretreated with 1x108 colony-forming units/ml P. gingivalis ATCC 33277 or 21 clinical isolates from moderate to severe chronic periodontitis patients (24 h), washed (2 h) and treated with P. gingivalis ATCC 33277 or the same clinical isolates again (24 h). Levels of pro-inflammatory cytokines TNF-α and IL-1β and anti-inflammatory cytokine IL-10 in supernatants were detected by ELISA. Moreover, to identify the possible mechanisms for the changes in cytokine secretion, Toll-like receptor 2 (TLR2) and TLR4 protein expressions were explored in these cells by flow cytometry. RESULTS:After repeated challenge with P. gingivalis ATCC 33277 or clinical isolates, production of TNF-α and IL-1β in macrophages was decreased significantly compared with that following a single stimulation (p0.05). In addition, there was interstrain variability in the ability to induce IL-1β and IL-10 production after repeated P. gingivalis stimulation. However, no significant changes in TLR2 or TLR4 were detected in macrophages that were repeatedly treated with P. gingivalis ATCC 33277 or clinical isolates compared with those stimulated with P. gingivalis only once (p>0.05). CONCLUSIONS:Repeated P. gingivalis stimulation triggered tolerance, which might contribute to limiting periodontal inflammation. However, tolerance induced by P. gingivalis might develop independently of TLR2 and TLR4 and be related to molecules in signaling pathways downstream of TLR2 and TLR4.

BACKGROUND:The rising cost of healthcare is of great concern in China, as evidenced by the media features negative reports almost daily. However there are only a few studies from well-developed cities, like Beijing or Shanghai, and little is known about healthcare costs in rest of the country. In this study, we use hospitalization summary reports (HSRs) from admitted cardiovascular diseases patients in Guangxi hospitals during 2013-2016, and we investigate temporal trends of healthcare costs and associated factors. METHODS:By generalized additive model, we compute temporal trends of cost per stay (CPS), cost per day (CPD) and others. We then use generalized linear models to assess which factors associate with CPS and CPD. FINDINGS:Using a total of 760,000 HSRs, we find that CPS appears to be stabilized around $1040 until the middle of year 2015, before exhibiting a downward trend. Similarly, CPD exhibits similar stable pattern. Meanwhile, surgery-specific CPS showed an increase in year 2013-2014, and then stabilized. Drug costs account for over 1/3 of CPS, but they are gradually declining. Costs associated with physicians' and nurses' services represent less than 5% of CPS. We found that age, sex, marital status, occupation and payment methods are significantly associated with CPS or CPD. Interestingly, we found no association between patient ethnicity and these costs. However, we did find that minority patients use more secondary hospitals than Han patients. INTERPRETATIONS:Healthcare costs in Guangxi are stable, contrary to the rise portrayed by Chinese mass media. Several factors can be associated with healthcare costs, and these may be useful for developing evidence-based policies. In particular, there is a need to encourage more Han patients to seek care in primary and secondary hospitals.

Chinese university students often suffer from acute stress, which can affect their mental health. We measured and evaluated perceived stress in this population using the Simplified Chinese version of the 10-item Perceived Stress Scale (SCPSS-10). The SCPSS-10, Patient Health Questionnaire (PHQ), and Generalized Anxiety Disorder 7-item scale (GAD-7) were conducted in 1096 university students. Two weeks later, 129 participants were re-tested using the SCPSS-10. Exploratory factor analysis yielded two factors with Eigen values of 4.76 and 1.48, accounting for 62.41% of the variance. Confirmatory factor analysis demonstrated good fit of this two-factor model. The internal consistency reliability, as measured by Cronbach's α, was 0.85. The test-retest reliability coefficient was 0.7. The SCPSS-10 exhibited high correlation with the PHQ-9 and GAD-7, indicating an acceptable concurrent validity. The SCPSS-10 exhibited satisfactory psychometric properties in Chinese university students.

China has high burden on both of tuberculosis (TB) and tobacco use. This study aims to explore the potential link between smoking and TB infection using baseline survey data of a large-scale population-based prospective study in rural China.Between July 1 and Sept 30, 2013, based on the baseline survey of a population-based, prospective study in rural China, the relationship between smoking and TB infection, assessed by interferon-gamma release assays (IGRA), was investigated among the total study population and only among those smokers, respectively.A total of 21,008 eligible rural registered residents (≥ 5 years old) from 4 rural sites were included in the analysis. Ever-smokers were more likely to be QuantiFERON-TB Gold In-Tube (QFT) positive than never smokers with an adjusted odds ratio (OR) of 1.34 (95% confidence interval (CI): 1.21-1.49). Among ever smokers, a significant linear dose-response relation was observed between duration of smoking (by years) and QFT positivity (p < 0.001). Stratified analysis suggested that such an association was not influenced by gender and age. Evidence for interaction of smoking status with age was found.Our results provide further evidence to support smoking might increase host susceptibility to TB infection. Populations under high risk of infection, such as elderly smokers, should be prior to TB infection controlling under a premise of community level intervention.

Spermatogenesis is a complex process that involves many elements. However, until now, little is known at the molecular level about spermatogenesis in poultry. Here we investigated microRNAs and their target genes that may be involved in germ cell development and spermatogonial in chicken. We used next-generation sequencing to analyze miRNA expression profiles in three types of germline cells: primordial germ cells (PGCs), spermatogonial stem cells (SSCs), and spermatogonia (Sp) during early stage of spermatogenesis. After validated the candidate miRNAs and corresponding genes' expression in three types of cells, we found 15 miRNAs that were enriched 21 target genes that may be involved in spermatogenesis. Among the enriched miRNAs, miR-202-5p/3p were up-regulated in the Sp library and down-regulated in the PGCs library. Through RT-qPCR and Dual-Luciferase reporter assay, we confirmed that miR-202-5p bind to LIMK2 and involved in germ cell development. Collectively, we firstly discover the novel miRNAs, like miR-202-5p, and its related genes and pathways, expressed during the early spermatogonial stage in chicken, which will provide new clues for deciphering the molecular mechanism of the miRNAs regulating germline stem cell differentiation and spermatogenesis in chicken.

This paper presents an adaption of the fractional order terminal sliding mode control (AFTSMC) strategy for DC-DC Buck converter. The following strategy aims to design a novel nonlinear sliding surface function, with a double closed-loop structure of voltage and current. This strategy is a fusion of two characteristics: terminal sliding mode control (TSMC) and fractional order calculation (FOC). In addition, the influence of "the controller parameters" on the "performance of double closed-loop system" is investigated. It is observed that the value of terminal power has to be chosen to make a compromise between start-up and transient response of the converter. Therefore the AFTSMC strategy chooses the value of the terminal power adaptively, and this strategy can lead to the appropriate number of fractional order as well. Furthermore, through the fractional order analysis, the system can reach the sliding mode surface in a finite time. And the theoretical considerations are verified by numerical simulation. The performance of the AFTSMC and TSMC strategies is tested by computer simulations. And the comparison simulation results show that the AFTSMC exhibits a considerable improvement in terms of a faster output voltage response during load changes. Moreover, AFTSMC obtains a faster dynamical response, smaller steady-state error rate and lower overshoot.

Chronic stress induces altered energy metabolism and plays important roles in the etiology of depression, in which the glucocorticoid negative feedback is disrupted due to imbalanced glucocorticoid receptor (GR) functions. The mechanism underlying the dysregulation of GR by chronic stress remains elusive. In this study, we investigated the role of AMP-activated protein kinase (AMPK), the key enzyme regulating cellular energy metabolism, and related signaling pathways in chronic stress-induced GR dysregulation. In cultured rat cortical astrocytes, glucocorticoid treatment decreased the level, which was accompanied by the decreased expression of liver kinase B1 (LKB1) and reduced phosphorylation of AMPK. Glucocorticoid-induced effects were attenuated by glucocorticoid-inducible kinase 1 (SGK1) inhibitor GSK650394, which also inhibited glucocorticoid induced phosphorylation of Forkhead box O3a (FOXO3a). Furthermore, glucocorticoid-induced down-regulation of GR was mimicked by the inhibition of AMPK and abolished by the AMPK activators or the histone deacetylase 5 (HDAC5) inhibitors. In line with the role of AMPK in GR expression, AMPK activator metformin reversed glucocorticoid-induced reduction of AMPK phosphorylation and GR expression as well as behavioral alteration of rats. Taken together, these results suggest that chronic stress activates SGK1 and suppresses the expression of LKB1 via inhibitory phosphorylation of FOXO3a. Downregulated LKB1 contributes to reduced activation of AMPK, leading to the dephosphorylation of HDAC5 and the suppression of transcription of GR.

BACKGROUND:Several metals have been reported to be associated with childhood asthma. However, the results on relationships between metals and risk of childhood asthma are inconclusive, and the research on adult asthma in the Chinese general population is rare. OBJECTIVES:To investigate potential associations between levels of urinary metals and adult asthma. METHODS:A case-control study of 551 adult asthma cases and 551 gender- and age-matched controls was conducted in Wuhan, China. Demographic information was obtained, and lung function was assessed. The urinary concentrations of 22 metals were measured by inductively coupled plasma mass spectrometry. RESULTS:After adjusting for other metalsand other covariates, urinary cadmium, molybdenum, chromium, copper, uranium and selenium were positively associated with asthma, with odds ratios (95% CI) of 1.69 (1.00, 2.85), 3.76 (2.30, 6.16), 4.89 (3.04, 7.89), 6.06 (3.27, 11.21), 6.99 (4.37, 11.19) and 9.17 (4.16, 20.21), respectively. By contrast, urinary lead, barium, iron, zinc, nickel, manganese and rubidium were negatively associated with asthma, with odds ratios (95% CI) of 0.48 (0.29, 0.80), 0.44 (0.27, 0.71), 0.41 (0.26, 0.64), 0.40 (0.24, 0.66), 0.30 (0.22, 0.41), 0.23 (0.14, 0.39) and 0.07 (0.03, 0.15), respectively. When comparing urinary metals in different subgroups of cases with those in matched controls, the associations of above 13 metals with asthma prevalence were nearly the same. CONCLUSIONS:Our results suggested that asthma prevalence in the Chinese adults was positively associated with urinary chromium, chromium, selenium, molybdenum, cadmium, and uranium, and negatively associated with urinary manganese, iron, nickel, zinc, rubidium, barium and lead. Additional research with larger populations in different regions is required to support our findings.

OBJECTIVE:To determine factors influencing the utilization and accessibility to bacteriologic-based tuberculosis (TB) diagnosis among sputum smear positive (SS+) retreatment TB patients, and to develop strategies for improving the case detection rate of MDR-TB in rural China. STUDY DESIGN AND SETTING:A cross-sectional study of SS+ TB retreatment patients was conducted in eight counties from three provinces with different implementation period and strategy of MDR-TB program in China. Demographic and socioeconomic parameters were collected by self-reporting questionnaires. Sputum samples were collected and cultured by the laboratory of county-designated TB clinics and delivered to prefectural Centers for Disease Prevention and Control (CDC) labs for DST with 4 first-line anti-TB drugs. RESULTS:Among the 196 SS+ retreatment patients, 61.22% received culture tests during current treatment. Patients from more developed regions (OR = 24.0 and 3.6, 95% CI: 8.6-67.3 and 1.1-11.6), with better socio-economic status (OR = 3. 8, 95% CI: 1.3-10.7), who had multiple previous anti-TB treatments (OR = 5.0, 95% CI: 1.6-15.9), and who failed in the most recent anti-TB treatment (OR = 2.6, 95% CI: 1.0-6.4) were more likely to receive culture tests. The percentage of isolates resistant to any of first-line anti-TB drugs and MDR-TB were 50.0% (95% CI: 39.8%-60.2%) and 30.4% (95% CI: 21.0%-39.8%) respectively. CONCLUSIONS:Retreatment SS+ TB patients, high risk MDR-TB population, had poor utilization of access to bacteriologic-based TB diagnosis, which is far from optimal. The next step of anti-TB strategy should be focused on how to make bacteriological-based diagnosis cheaper, safer and more maneuverable, and how to assure the DST-guided treatment for these high-risk TB patients.

Necroptosis as a molecular program, rather than simply incidental cell death, was established by elucidating the roles of receptor interacting protein (RIP) kinases 1 and 3, along with their downstream partner, mixed lineage kinase-like domain protein (MLKL). Previous studies suggested that phosphoglycerate mutase family member 5 (PGAM5), a mitochondrial protein that associates with RIP1/RIP3/MLKL complex, promotes necroptosis. We have generated mice deficient in the pgam5 gene and surprisingly found PGAM5-deficiency exacerbated rather than reduced necroptosis in response to multiple in vitro and in vivo necroptotic stimuli, including ischemic reperfusion injury (I/R) in the heart and brain. Electron microscopy, biochemical, and confocal analysis revealed that PGAM5 is indispensable for the process of PINK1 dependent mitophagy which antagonizes necroptosis. The loss of PGAM5/PINK1 mediated mitophagy causes the accumulation of abnormal mitochondria, leading to the overproduction of reactive oxygen species (ROS) that worsen necroptosis. Our results revise the former proposal that PGAM5 acts downstream of RIP1/RIP3 to mediate necroptosis. Instead, PGAM5 protects cells from necroptosis by independently promoting mitophagy. PGAM5 promotion of mitophagy may represent a therapeutic target for stroke, myocardial infarction and other diseases caused by oxidative damage and necroptosis.

A chitosanase was purified from jelly fig latex by ammonium sulfate fractionation (50-80% saturation) and three successive column chromatography steps. The purified enzyme was almost homogeneous, as determined by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and gel activity staining. The molecular mass of the enzyme was 20.5 kDa. The isoelectric point (pI) was

Periodontitis is a chronic inflammatory disease induced by bacteria. Exposure of the host to periodontal pathogens and their virulence factors induces a state of hyporesponsiveness to subsequent stimulations, which is termed endotoxin tolerance. The role and mechanism of lipopolysaccharide (LPS)-tolerized monocytes in inflammatory responses in neutrophils are currently unclear. Here, conditioned supernatants were collected from THP-1 cells treated with or without repeated 1 μg/ml Porphyromonas gingivalis (P.gingivalis) LPS. The chemotactic response of freshly isolated neutrophils recruited by supernatants was determined by a transwell migration assay, which demonstrated a reduced migration of neutrophils stimulated with supernatants from tolerized THP-1 cells in comparison to non-tolerized THP-1 cells. In addition, there was a marked increase in reactive oxygen species (ROS) generation and a significant decrease in Caspase 3 activities in neutrophils treated with supernatants from THP-1 cells that were treated repeatedly with P.gingivalis LPS in comparison to single treatment. A cytokine antibody array was then used to assess cytokine expression patterns in THP-1 cells. In tolerized THP-1 cells, 43 cytokine (43/170) expression levels were decreased, including chemokine ligand 23 (CCL23) and IFN-γ, while 11 cytokine (11/170) expression levels were increased, such as death receptor 6 (DR6). Furthermore, there was decreased production of IFN-γ and epithelial neutrophil activating peptide-78 (ENA-78) in THP-1 cells after stimulation with repeated P. gingivalis LPS in comparison to single challenge, which was confirmed by ELISA. Therefore, P.gingivalis LPS- tolerized THP-1 cells were able to depress neutrophil chemotaxis and apoptosis, and contribute to respiratory burst, which might be related to the changes in cytokine expression patterns in THP-1 cells.

The Cancer Genetic Markers of Susceptibility genome-wide association study (GWAS) originally identified a single nucleotide polymorphism (SNP) rs11249433 at 1p11.2 associated with breast cancer risk. To fine-map this locus, we genotyped 92 SNPs in a 900kb region (120,505,799-121,481,132) flanking rs11249433 in 45,276 breast cancer cases and 48,998 controls of European, Asian and African ancestry from 50 studies in the Breast Cancer Association Consortium. Genotyping was done using iCOGS, a custom-built array. Due to the complicated nature of the region on chr1p11.2: 120,300,000-120,505,798, that lies near the centromere and contains seven duplicated genomic segments, we restricted analyses to 429 SNPs excluding the duplicated regions (42 genotyped and 387 imputed). Per-allelic associations with breast cancer risk were estimated using logistic regression models adjusting for study and ancestry-specific principal components. The strongest association observed was with the original identified index SNP rs11249433 (minor allele frequency (MAF) 0.402; per-allele odds ratio (OR) = 1.10, 95% confidence interval (CI) 1.08-1.13, P = 1.49 x 10-21). The association for rs11249433 was limited to ER-positive breast cancers (test for heterogeneity P≤8.41 x 10-5). Additional analyses by other tumor characteristics showed stronger associations with moderately/well differentiated tumors and tumors of lobular histology. Although no significant eQTL associations were observed, in silico analyses showed that rs11249433 was located in a region that is likely a weak enhancer/promoter. Fine-mapping analysis of the 1p11.2 breast cancer susceptibility locus confirms this region to be limited to risk to cancers that are ER-positive.

BackgroundAcidosis impairs cognitions and behaviors presumably by acidification-induced changes in neuronal metabolism. Cortical GABAergic neurons are vulnerable to pathological factors and their injury leads to brain dysfunction. How acidosis induces GABAergic neuron injury remains elusive. As the glia cells and neurons interact each other, we intend to examine the role of the astrocytes in acidosis-induced GABAergic neuron injury.ResultsExperiments were done at GABAergic cells and astrocytes in mouse cortical slices. To identify astrocytic involvement in acidosis-induced impairment, we induced the acidification in single GABAergic neuron by infusing proton intracellularly or in both neurons and astrocytes by using proton extracellularly. Compared the effects of intracellular acidification and extracellular acidification on GABAergic neurons, we found that their active intrinsic properties and synaptic outputs appeared more severely impaired in extracellular acidosis than intracellular acidosis. Meanwhile, extracellular acidosis deteriorated glutamate transporter currents on the astrocytes and upregulated excitatory synaptic transmission on the GABAergic neurons. Moreover, the antagonists of glutamate NMDA-/AMPA-receptors partially reverse extracellular acidosis-induced injury in the GABAergic neurons.ConclusionOur studies suggest that acidosis leads to the dysfunction of cortical GABAergic neurons by astrocyte-mediated excitotoxicity, in addition to their metabolic changes as indicated previously.

BACKGROUND:Elevated heavy metals and fasting plasma glucose (FPG) levels were both associated with increased risk of cardiovascular diseases. However, studies on the associations of heavy metals and essential elements with altered FPG and diabetes risk were limited or conflicting. The objective of this study was to evaluate the potential associations of heavy metals and essential trace elements with FPG and diabetes risk among general Chinese population. METHODS:We conducted a cross-sectional study to investigate the associations of urinary concentrations of 23 metals with FPG, impaired fasting glucose (IFG) and diabetes among 2242 community-based Chinese adults in Wuhan. We used the false discovery rate (FDR) method to correct for multiple hypothesis tests. RESULTS:After adjusting for potential confounders, urinary aluminum, titanium, cobalt, nickel, copper, zinc, selenium, rubidium, strontium, molybdenum, cadmium, antimony, barium, tungsten and lead were associated with altered FPG, IFG or diabetes risk (all P< 0.05); arsenic was only dose-dependently related to diabetes (P< 0.05). After additional adjustment for multiple testing, titanium, copper, zinc, selenium, rubidium, tungsten and lead were still significantly associated with one or more outcomes (all FDR-adjusted P< 0.05). CONCLUSIONS:Our results suggest that multiple metals in urine are associated with FPG, IFG or diabetes risk. Because the cross-sectional design precludes inferences about causality, further prospective studies are warranted to validate our findings.

The best way to increase patient survival rate is to identify patients who are likely to progress to muscle-invasive or metastatic disease upfront and treat them more aggressively. The human cell lines HCV29 (normal bladder epithelia), KK47 (low grade nonmuscle invasive bladder cancer, NMIBC), and YTS1 (metastatic bladder cancer) have been widely used in studies of molecular mechanisms and cell signaling during bladder cancer (BC) progression. However, little attention has been paid to global quantitative proteome analysis of these three cell lines. We labeled HCV29, KK47, and YTS1 cells by the SILAC method using three stable isotopes each of arginine and lysine. Labeled proteins were analyzed by 2D ultrahigh-resolution liquid chromatography LTQ Orbitrap mass spectrometry. Among 3721 unique identified and annotated proteins in KK47 and YTS1 cells, 36 were significantly upregulated and 74 were significantly downregulated with >95% confidence. Differential expression of these proteins was confirmed by western blotting, quantitative RT-PCR, and cell staining with specific antibodies. Gene ontology (GO) term and pathway analysis indicated that the differentially regulated proteins were involved in DNA replication and molecular transport, cell growth and proliferation, cellular movement, immune cell trafficking, and cell death and survival. These proteins and the advanced proteome techniques described here will be useful for further elucidation of molecular mechanisms in BC and other types of cancer.

Baicalein, a widely used Chinese herbal medicine, has multiple pharmacological activities. However, the precise mechanisms of the anti-proliferation and anti-metastatic effects of baicalein on gallbladder cancer (GBC) remain poorly understood. Therefore, the aim of this study was to assess the anti-proliferation and anti-metastatic effects of baicalein and the related mechanism(s) on GBC. In the present study, we found that treatment with baicalein induced a significant inhibitory effect on proliferation and promoted apoptosis in GBC-SD and SGC996 cells, two widely used gallbladder cancer cell lines. Additionally, treatment with baicalein inhibited the metastasis of GBC cells. Moreover, we demonstrated for the first time that baicalein inhibited GBC cell growth and metastasis via down-regulation of the expression level of Zinc finger protein X-linked (ZFX). In conclusion, our studies suggest that baicalein may be a potential phytochemical flavonoid for therapeutics of GBC and ZFX may serve as a molecular marker or predictive target for GBC.

We determined the role of endogenous hydrogen sulfide (H2S) in cerebral vasodilation/hyperemia and early BBB disruption following ischemic stroke. A cranial window was prepared over the left frontal, parietal and temporal cortex in mice. Transient focal cerebral Ischemia was induced by directly ligating the middle cerebral artery (MCA) for two hours. Regional vascular response and cerebral blood flow (CBF) during ischemia and reperfusion were measured in real time. Early BBB disruption was assessed by Evans Blue (EB) and sodium fluorescein (Na-F) extravasation at 3 hours of reperfusion. Topical treatment with DL-propargylglycine (PAG, an inhibitor for cystathionine γ-lyase (CSE)) and aspartate (ASP, inhibitor for cysteine aminotransferase/3-mercaptopyruvate sulfurtransferase (CAT/3-MST)), but not O-(Carboxymethyl)hydroxylamine hemihydrochloride (CHH, an inhibitor for cystathionine β-synthase (CBS)), abolished postischemic cerebral vasodilation/hyperemia and prevented EB and Na-F extravasation. CSE knockout (CSE-/-) reduced postischemic cerebral vasodilation/hyperemia but only inhibited Na-F extravasation. An upregulated CBS was found in cerebral cortex of CSE-/- mice. Topical treatment with CHH didn't further alter postischemic cerebral vasodilation/hyperemia, but prevented EB extravasation in CSE-/- mice. In addition, L-cysteine-induced hydrogen sulfide (H2S) production similarly increased in ischemic side cerebral cortex of control and CSE-/- mice. Our findings suggest that endogenous production of H2S by CSE and CAT/3-MST during reperfusion may be involved in postischemic cerebral vasodilation/hyperemia and play an important role in early BBB disruption following transient focal cerebral ischemia.

Many studies have addressed the diagnostic performance of echocardiography to evaluate acute cardiac allograft rejection compared with endomyocardial biopsy. But the existence of heterogeneity limited its clinical application. Thus, we conducted a comprehensive, systematic literature review and meta-analysis for the purpose.Studies prior to September 1, 2014 identified by Medline/PubMed, EMBASE and Cochrance were examined by two independent reviews. We conducted meta-analysis by using Meta-DiSc 1.4 software. An assessment tool of QUADAS-2 was applied to evaluate the risk of bias and applicability of the studies.Thirty studies met the inclusion criteria of meta-analysis. The four parameters of pressure half time, isovolumic relaxation time, index of myocardial performance and late diastolic mitral annular motion velocity were included in the meta-analysis, with a pooled diagnostic odds ratio of 10.43, 6.89, 15.95 and 5.68 respectively, and the area under the summary receiver operating characteristic curves value of 0.829, 0.599, 0.871 and 0.685 respectively.The meta-analysis and systematic review demonstrate that no single parameter of echocardiography showed a reliable diagnostic performance for acute cardiac allograft rejection. A result of echocardiography for ACAR should be comprehensively considered by physicians in the context of clinical presentations and imaging feature.

To disclose the associated risk factors for latent tuberculosis infection (LTBI) and the current situation of LTBI in the eastern China.A cross-sectional study was undertaken to evaluate the LTBI rate and risk factors.A total of 5305 subjects were finally included, with the IGRA positive rate of 19.98% (1060/5305). The LTBI rates were increasing with age (ORs were in significance from 6.60 to 20.92). Male gender significantly increased the risk of LTBI by 0.52 fold (OR = 1.52). Both smoking and drinking significantly increased the risk of LTBI (OR = 1.83 and OR = 1.67, respectively). Meanwhile, overweight and close contact with tuberculosis were risk factors for LTBI (OR = 1.36 and OR = 2.38, respectively). However, higher level of education and BCG vaccination lowered the risk of LTBI (OR = 0.16 and OR = 0.39, respectively). The multivariate logistic regression showed that age, male gender, smoking, overweight and close contacting with tuberculosis were risk factors for LTBI, but BCG vaccination was a protective factor for LTBI.BCG vaccination exerted protective effect on tuberculosis. However, LTBI rate in the Chinese rural area was critical and subjects above 30 years, male, smoking, overweight and close contact with tuberculosis wound be the targets for LTBI screening and source of tuberculosis.

Fibroblast growth factor 21 (FGF21) exerts wide-range effects on carbohydrate and lipid metabolism. However, its perturbation in type 2 diabetes mellitus (T2DM) remains elusive. Besides, previous human studies in T2DM simply investigated fasting or stimulated levels of FGF21. The current study sought to evaluate the temporal changes of circulating FGF21 in subjects with and without T2DM.Ten patients with T2DM and 16 normal controls (NC) were recruited. Participants were categorized as obese (BMI≥25 kg/m2) or lean (BMI 0.05).These findings suggest that the pattern of circulating FGF21 does not differ significantly between T2DM and NC,although T2DM patients showed a trend toward higher fasting FGF21 than healthy subjects. The pattern of circulating FFAs is significantly associated with that of FGF21.

The mosquito-borne Chikungunya virus (CHIKV) is a profound global threat due to its high rate of contagion and the lack of vaccine or effective treatment. Suramin is a symmetric polyanionic naphthylurea that is widely used in the clinical treatment of parasite infections. Numerous studies have reported the broad antiviral activities of suramin; however, inhibition effects against CHIKV have not yet been demonstrated. The aim of this study was thus to investigate the antiviral effect of suramin on CHIKV infection and to elucidate the molecular mechanism underlying inhibition using plaque reduction assay, RT-qPCR, western blot analysis, and plaque assay. Microneutralization assay was used to determine the EC50 of suramin in the CHIKV-S27 strain as well as in three other clinical strains (0611aTw, 0810bTw and 0706aTw). Time-of-addition was used to reveal the anti-CHIKV mechanism of suramin. We also evaluated anti-CHIKV activity with regard to viral entry, virus release, and cell-to-cell transmission. Cytopathic effect, viral RNA, viral protein, and the virus yield of CHIKV infection were shown to diminish in the presence of suramin in a dose-dependent manner. Suramin was also shown the inhibitory activities of the three clinical isolates. Suramin inhibited the early progression of CHIKV infection, due perhaps to interference with virus fusion and binding, which subsequently prevented viral entry. Results of a molecular docking simulation indicate that suramin may embed within the cavity of the E1/E2 heterodimer to interfere with their function. Suramin was also shown to reduce viral release and cell-to-cell transmission of CHIKV. In conclusion, Suramin shows considerable potential as a novel anti-CHIKV agent targeting viral entry, extracellular transmission, and cell-to-cell transmission.

To evaluate whether the genetic susceptibility of T2D was associated with overall survival (OS) and disease-free survival (DFS) outcomes for breast cancer (BC).Included in the study were 6346 BC patients who participated in three population-based epidemiological studies of BC and were genotyped with either GWAS or Exome-chip. We constructed a genetic risk score (GRS) for diabetes using risk variants identified from the GWAS catalog (http://genome.gov/gwastudies) that were associated with T2D risk at a minimum significance level of P ≤ 5.0E-8 among Asian population and evaluated its associations with BC outcomes with Cox proportional hazards models.During a median follow-up of 8.08 years (range, 0.01-16.95 years), 1208 deaths were documented in 6346 BC patients. Overall, the diabetes GRS was not associated with OS and DFS. Analyses stratified by estrogen receptor status (ER) showed that the diabetes GRS was inversely associated with OS among women with ER- but not in women with ER+ breast cancer; the multivariable adjusted HR was 1.38 (95% CI: 1.05-1.82) when comparing the highest to the lowest GRS quartiles. The association of diabetes GRS with OS varied by diabetes status (P for interaction

In this paper, we consider the Minimum Reaction Insertion (MRI) problem for finding the minimum number of additional reactions from a reference metabolic network to a host metabolic network so that a target compound becomes producible in the revised host metabolic network in a Boolean model. Although a similar problem for larger networks is solvable in a flux balance analysis (FBA)-based model, the solution of the FBA-based model tends to include more reactions than that of the Boolean model. However, solving MRI using the Boolean model is computationally more expensive than using the FBA-based model since the Boolean model needs more integer variables. Therefore, in this study, to solve MRI for larger networks in the Boolean model, we have developed an efficient Integer Programming formalization method in which the number of integer variables is reduced by the notion of feedback vertex set and minimal valid assignment. As a result of computer experiments conducted using the data of metabolic networks of E. coli and reference networks downloaded from the Kyoto Encyclopedia of Genes and Genomes (KEGG) database, we have found that the developed method can appropriately solve MRI in the Boolean model and is applicable to large scale-networks for which an exhaustive search does not work. We have also compared the developed method with the existing connectivity-based methods and FBA-based methods, and show the difference between the solutions of our method and the existing methods. A theoretical analysis of MRI is also conducted, and the NP-completeness of MRI is proved in the Boolean model. Our developed software is available at "http://sunflower.kuicr.kyoto-u.ac.jp/~rogi/minRect/minRect.html."

An excessive release of glutamate is considered to be a molecular mechanism associated with several neurological diseases that causes neuronal damage. Therefore, searching for compounds that reduce glutamate neurotoxicity is necessary. In this study, the possibility that the natural flavone acacetin derived from the traditional Chinese medicine Clerodendrum inerme (L.) Gaertn is a neuroprotective agent was investigated. The effect of acacetin on endogenous glutamate release in rat hippocampal nerve terminals (synaptosomes) was also investigated. The results indicated that acacetin inhibited depolarization-evoked glutamate release and cytosolic free Ca(2+) concentration ([Ca(2+)]C) in the hippocampal nerve terminals. However, acacetin did not alter synaptosomal membrane potential. Furthermore, the inhibitory effect of acacetin on evoked glutamate release was prevented by the Cav2.2 (N-type) and Cav2.1 (P/Q-type) channel blocker known as ω-conotoxin MVIIC. In a kainic acid (KA) rat model, an animal model used for excitotoxic neurodegeneration experiments, acacetin (10 or 50 mg/kg) was administrated intraperitoneally to the rats 30 min before the KA (15 mg/kg) intraperitoneal injection, and subsequently induced the attenuation of KA-induced neuronal cell death and microglia activation in the CA3 region of the hippocampus. The present study demonstrates that the natural compound, acacetin, inhibits glutamate release from hippocampal synaptosomes by attenuating voltage-dependent Ca(2+) entry and effectively prevents KA-induced in vivo excitotoxicity. Collectively, these data suggest that acacetin has the therapeutic potential for treating neurological diseases associated with excitotoxicity.

Few studies have investigated vitamin D status in association with modifiable lifestyle factors and clinical characteristics among breast cancer patients, with no studies among Chinese women, who may be at higher risk of vitamin D deficiency. We aimed to evaluate circulating 25-hydroxyvitamin D (25(OH)D) levels in association with clinical and lifestyle factors among 1,940 Chinese breast cancer patients.Participants included breast cancer cases aged 22-77 from a population-based case-control study conducted in Shanghai, China during 1996-1998 (n = 1,044) and 2002-2005 (n = 896). Circulating 25(OH)D levels were measured in plasma samples (95% collected ≤6 months post-diagnosis). Prevalence ORs and 95% CIs were derived from multinomial logistic regression models, adjusting for age, season, and other factors.About 23% and 48% of women were vitamin D deficient (

We have previously reported in Xishuangbanna (Banna) Dai Autonomous Prefecture, a well-developed tourist destination in the southwest border of China, that HIV-1 transmitted dominantly through heterosexual contact with less divergent genotypes and few drug resistant mutations. Due to the rapid increase of newly diagnosed HIV-1 cases per year in Banna in recent years, it's important to evaluate the evolution of HIV-1 molecular epidemiology for the better understanding of ongoing HIV-1 outbreak in this region.By sequencing of HIV-1 pol genes and phylogenetic analysis, we conducted a molecular epidemiologic study in 352 HIV-1-seropositive highly active antiretroviral treatment (HAART)-naïve individuals newly diagnosed at the Banna Center for Disease Control and Prevention between 2009 and 2011. Of 283 samples (84.1% taken from heterosexually acquired adults, 10.6% from needle-sharing drug users, 2.8% from men who have sex with men, 0.4% from children born from HIV-1-infected mothers, and 2.1% remained unknown) with successful sequencing for pol gene, we identified 108 (38.2%) HIV-1 subtype CRF08_BC, 101 (35.7%) CRF01_AE, 49 (17.3%) CRF07_BC, 5 (1.8%) C/CRF57_BC, 3 (1.1%) B', 1 (0.4%) B/CRF51_01B, and 16 (5.7%) unique recombinants forms. Among these infected individuals, 104 (36.7%) cases showed drug resistant or resistance-relevant mutations, and 4 of them conferring high-level resistance to 3TC/FTC, EFV/NVP or NFV. Phylogenetic analysis revealed 21 clusters (2-7 sequences) with only 21.2% (60/283) sequences involved.In contrast to our previous findings, CRF08_BC, replaced CRF01_AE, became the dominant genotype of HIV-1 in Banna prefecture. The viral strains with drug resistance mutations were detected frequently in newly diagnosed HIV-1-infected individuals in this region.

The red swamp crawfish, Procambarus clarkii, has become one of the most economically important cultured species in China. Currently, little is known about the gonadal development of this species. Isolation and characterization of genes are an initial step towards understanding gonadal development of P. clarkii.Using the 454 pyrosequencing technology, we obtained a total of 1,134,993 high quality sequence reads from the crawfish testis and ovary libraries. We aimed to identify different genes with a potential role in gonad development. The assembly formed into 22,652 isotigs, distributed by GO analysis across 55 categories in the three ontologies, 'molecular function', 'cellular component', and 'biological processes'. Comparative transcript analysis showed that 1,720 isotigs in the ovary were up-regulated and 2138 isotigs were down-regulated. Several gonad development related genes, such as vitellogenin, cyclin B, cyclin-dependent kinases 2, Dmc1 and ubiquitin were identified. Quantitative real-time PCR verified the expression profiles of 14 differentially expressed genes, and confirmed the reliability of the 454 pyrosequencing.Our findings provide an archive for future research on gonadal development at a molecular level in P. clarkii and other crustacean. This data will be helpful to develop new ideas for artificial regulation of the reproductive process in crawfish aquaculture.

The WNK1 (WNK lysine deficient protein kinase 1) protein is a serine/threonine protein kinase with emerging roles in cancer. WNK1 causes hypertension and hyperkalemia when overexpressed and cardiovascular defects when ablated in mice. In this study, the role of Wnk1 in angiogenesis was explored using the zebrafish model. There are two zebrafish wnk1 isoforms, wnk1a and wnk1b, and both contain all the functional domains found in the human WNK1 protein. Both isoforms are expressed in the embryo at the initiation of angiogenesis and in the posterior cardinal vein (PCV), similar to fms-related tyrosine kinase 4 (flt4). Using morpholino antisense oligonucleotides against wnk1a and wnk1b, we observed that wnk1 morphants have defects in angiogenesis in the head and trunk, similar to flk1/vegfr2 morphants. Furthermore, both wnk1a and wnk1b mRNA can partially rescue the defects in vascular formation caused by flk1/vegfr2 knockdown. Mutation of the kinase domain or the Akt/PI3K phosphorylation site within wnk1 destroys this rescue capability. The rescue experiments provide evidence that wnk1 is a downstream target for Vegfr2 (vascular endothelial growth factor receptor-2) and Akt/PI3K signaling and thereby affects angiogenesis in zebrafish embryos. Furthermore, we found that knockdown of vascular endothelial growth factor receptor-2 (flk1/vegfr2) or vascular endothelial growth factor receptor-3 (flt4/vegfr3) results in a decrease in wnk1a expression, as assessed by in situ hybridization and q-RT-PCR analysis. Thus, the Vegf/Vegfr signaling pathway controls angiogenesis in zebrafish via Akt kinase-mediated phosphorylation and activation of Wnk1 as well as transcriptional regulation of wnk1 expression.

Cu2+, Zn2+, Fe2+ and I- are often supplemented to the diet of suckling and early weaning piglets, but little information is available regarding the effects of different Cu2+, Zn2+, Fe2+ and I- mixtures on bacteria growth, diversity and fermentation characteristics of fermented liquid diet for piglets. Pyrosequencing was performed to investigate the effect of Cu2+, Zn2+, Fe2+ and I- mixtures on the diversity, growth and fermentation characteristics of bacteria in the liquid diet fermented with Bacillus subtilis and Enterococcus faecalis under air-tight condition. Results showed that the mixtures of Cu2+, Zn2+, Fe2+ and I- at different concentrations promoted Bacillus growth, increased bacterial diversity and lactic acid production and lowered pH to about 5. The importance of Cu2+, Zn2+, Fe2+ and I- is different for Bacillus growth with the order Zn2+> Fe2+>Cu2+> I- in a 21-d fermentation and Cu2+>I->Fe2+>Zn2+ in a 42-d fermentation. Cu2+, Zn2+, Fe2+ and I- is recommended at a level of 150, 60, 150 and 0.6 mg/kg respectively for the production of fermented liquid diet with Bacillus subtilis. The findings improve our understanding of the influence of trace elements on liquid diet fermentation with probiotics and support the proper use of trace elements in the production of fermented liquid diet for piglets.

Ventilator-associated pneumonia (VAP) is a common complication and cause of death in neonates on mechanical ventilation. However, it is difficult to define the causes of VAP. To understand the causes of VAP, we undertook a prospective study based on the diversity of the microflora in VAP. The experimental group consisted of newborns who suffered from respiratory distress syndrome (RDS) and VAP, while the control group suffered from RDS without VAP. Sputa were collected within 1, 3, and 5 days of ventilation and were divided into six groups. DNA was extracted from the samples, and the 16S rDNA was PCR amplified, separated using denaturing gradient gel electrophoresis (DGGE), cloned and sequenced. The resulting sequences were compared using BLAST. The DGGE pictures were measured, and the richness, Shannon-Wiener index, and cluster maps were analyzed. No differences were found regarding the constituent ratio of any genus between the Non-VAP and VAP group within 1 day after intubation. After 1 to 3 days, the constituent ratios of Klebsiella sp., Acinetobacter sp., and Streptococcus sp. in the VAP group were higher than those in the Non-VAP group, and the ratios of Serratia sp. and Achromobacter sp. were lower. After 3 to 5 days, the ratios of Klebsiella sp., Acinetobacter sp., Serratia sp., and Achromobacter sp. were lower than those in the Non-VAP group. The richness and Shannon-Wiener index of the Non-VAP group were higher than those of the VAP group from 1 to 3 days after intubation, while no differences were found within 1 day and from 3 to 5 days. We conclude that during the first three days of intubation, the microflora diversity in the lower respiratory tract was reduced due to VAP, and the greater constituent ratios of Klebsiella sp., Acinetobacter sp., and Streptococcus sp. in the sputum may be indicators of VAP.

Hepatocarcinogenesis commonly involves the gradual progression from hepatitis to fibrosis and cirrhosis, and ultimately to hepatocellular carcinoma (HCC). Endothelin 1 (Edn1) has been identified as a gene that is significantly up-regulated in HBx-induced HCC in mice. In this study, we further investigated the role of edn1 in hepatocarcinogenesis using a transgenic zebrafish model and a cell culture system. Liver-specific edn1 expression caused steatosis, fibrosis, glycogen accumulation, bile duct dilation, hyperplasia, and HCC in zebrafish. Overexpression of EDN1 in 293T cells enhanced cell proliferation and cell migration in in vitro and xenotransplantation assays and was accompanied with up-regulation of several cell cycle/proliferation- and migration-specific genes. Furthermore, expression of the unfolded protein response (UPR) pathway-related mediators, such as spliced XBP1, ATF6, IRE1, and PERK, was also up-regulated at both the RNA and protein levels. In the presence of an EDN1 inhibitor or an AKT inhibitor, these increases were diminished and the EDN1-induced migration ability also was disappeared, suggesting that the EDN1 effects act through activation of the AKT pathway to enhance the UPR and subsequently activate the expression of downstream genes. Additionally, p-AKT is enhanced in the edn1 transgenic fish compared to the GFP-mCherry control. The micro RNA miR-1 was found to inhibit the expression of EDN1. We also observed an inverse correlation between EDN1 and miR-1 expression in HCC patients. In conclusion, our data suggest that EDN1 plays an important role in HCC progression by activating the PI3K/AKT pathway and is regulated by miR-1.

AcrB is a multidrug transporter in the inner membrane of Escherichia coli. It is an obligate homotrimer and forms a tripartite efflux complex with AcrA and TolC. AcrB is the engine of the efflux machinery and determines substrate specificity. Active efflux depends on several functional features including proton translocation across the inner membrane through a proton relay pathway in the transmembrane domain of AcrB; substrate binding and migration through the substrate translocation pathway; the interaction of AcrB with AcrA and TolC; and the formation of AcrB homotrimer. Here we investigated two aspects of the inter-correlation between these functional features, the dependence of AcrA-AcrB interaction on AcrB trimerization, and the reliance of substrate binding and penetration on protein-protein interaction. Interaction between AcrA and AcrB was investigated through chemical crosslinking, and a previously established in vivo fluorescent labeling method was used to probe substrate binding. Our data suggested that dissociation of the AcrB trimer drastically decreased its interaction with AcrA. In addition, while substrate binding with AcrB seemed to be irrelevant to the presence or absence of AcrA and TolC, the capability of trimerization and conduction of proton influx did affect substrate binding at selected sites along the substrate translocation pathway in AcrB.

Berberine, an isoquinoline plant alkaloid, protects neurons against neurotoxicity. An excessive release of glutamate is considered to be one of the molecular mechanisms of neuronal damage in several neurological diseases. In this study, we investigated whether berberine could affect endogenous glutamate release in nerve terminals of rat cerebral cortex (synaptosomes) and explored the possible mechanism. Berberine inhibited the release of glutamate evoked by the K(+) channel blocker 4-aminopyridine (4-AP), and this phenomenon was prevented by the chelating extracellular Ca(2+) ions and the vesicular transporter inhibitor bafilomycin A1, but was insensitive to the glutamate transporter inhibitor DL-threo-beta-benzyl-oxyaspartate. Inhibition of glutamate release by berberine was not due to it decreasing synaptosomal excitability, because berberine did not alter 4-AP-mediated depolarization. The inhibitory effect of berberine on glutamate release was associated with a reduction in the depolarization-induced increase in cytosolic free Ca(2+) concentration. Involvement of the Cav2.1 (P/Q-type) channels in the berberine action was confirmed by blockade of the berberine-mediated inhibition of glutamate release by the Cav2.1 (P/Q-type) channel blocker ω-agatoxin IVA. In addition, the inhibitory effect of berberine on evoked glutamate release was prevented by the mitogen-activated/extracellular signal-regulated kinase kinase (MEK) inhibitors. Berberine decreased the 4-AP-induced phosphorylation of extracellular signal-regulated kinase 1 and 2 (ERK1/2) and synapsin I, the main presynaptic target of ERK; this decrease was also blocked by the MEK inhibition. Moreover, the inhibitory effect of berberine on evoked glutamate release was prevented in nerve terminals from mice lacking synapsin I. Together, these results indicated that berberine inhibits glutamate release from rats cortical synaptosomes, through the suppression of presynaptic Cav2.1 channels and ERK/synapsin I signaling cascade. This finding may provide further understanding of the mode of berberine action in the brain and highlights the therapeutic potential of this compound in the treatment of a wide range of neurological disorders.

Hepatocarcinogenesis is a multistep process that starts from fatty liver and transitions to fibrosis and, finally, into cancer. Many etiological factors, including hepatitis B virus X antigen (HBx) and p53 mutations, have been implicated in hepatocarcinogenesis. However, potential synergistic effects between these two factors and the underlying mechanisms by which they promote hepatocarcinogenesis are still unclear. In this report, we show that the synergistic action of HBx and p53 mutation triggers progressive hepatocellular carcinoma (HCC) formation via src activation in zebrafish. Liver-specific expression of HBx in wild-type zebrafish caused steatosis, fibrosis and glycogen accumulation. However, the induction of tumorigenesis by HBx was only observed in p53 mutant fish and occurred in association with the up-regulation and activation of the src tyrosine kinase pathway. Furthermore, the overexpression of src in p53 mutant zebrafish also caused hyperplasia, HCC, and sarcomatoid HCC, which were accompanied by increased levels of the signaling proteins p-erk, p-akt, myc, jnk1 and vegf. Increased expression levels of lipogenic factors and the genes involved in lipid metabolism and glycogen storage were detected during the early stages of hepatocarcinogenesis in the HBx and src transgenic zebrafish. The up-regulation of genes involved in cell cycle regulation, tumor progression and other molecular hallmarks of human liver cancer were found at later stages in both HBx and src transgenic, p53 mutant zebrafish. Together, our study demonstrates that HBx and src overexpression induced hepatocarcinogenesis in p53 mutant zebrafish. This phenomenon mimics human HCC formation and provides potential in vivo platforms for drug screening for therapies for human liver cancer.

The transport and accumulation of anticancer nanodrugs in tumor tissues are affected by many factors including particle properties, vascular density and leakiness, and interstitial diffusivity. It is important to understand the effects of these factors on the detailed drug distribution in the entire tumor for an effective treatment. In this study, we developed a small-scale mathematical model to systematically study the spatiotemporal responses and accumulative exposures of macromolecular carriers in localized tumor tissues. We chose various dextrans as model carriers and studied the effects of vascular density, permeability, diffusivity, and half-life of dextrans on their spatiotemporal concentration responses and accumulative exposure distribution to tumor cells. The relevant biological parameters were obtained from experimental results previously reported by the Dreher group. The area under concentration-time response curve (AUC) quantified the extent of tissue exposure to a drug and therefore was considered more reliable in assessing the extent of the overall drug exposure than individual concentrations. The results showed that 1) a small macromolecule can penetrate deep into the tumor interstitium and produce a uniform but low spatial distribution of AUC; 2) large macromolecules produce high AUC in the perivascular region, but low AUC in the distal region away from vessels; 3) medium-sized macromolecules produce a relatively uniform and high AUC in the tumor interstitium between two vessels; 4) enhancement of permeability can elevate the level of AUC, but have little effect on its uniformity while enhancement of diffusivity is able to raise the level of AUC and improve its uniformity; 5) a longer half-life can produce a deeper penetration and a higher level of AUC distribution. The numerical results indicate that a long half-life carrier in plasma and a high interstitial diffusivity are the key factors to produce a high and relatively uniform spatial AUC distribution in the interstitium.

Age at natural menopause (ANM) is a complex trait with high heritability and is associated with several major hormonal-related diseases. Recently, several genome-wide association studies (GWAS), conducted exclusively among women of European ancestry, have discovered dozens of genetic loci influencing ANM. No study has been conducted to evaluate whether these findings can be generalized to Chinese women.We evaluated the index single nucleotide polymorphisms (SNPs) in 19 GWAS-identified genetic susceptibility loci for ANM among 3,533 Chinese women who had natural menopause. We also investigated 3 additional SNPs which were in LD with the index SNP in European-ancestry but not in Asian-ancestry populations. Two genetic risk scores (GRS) were calculated to summarize SNPs across multiple loci one for all SNPs tested (GRSall), and one for SNPs which showed association in our study (GRSsel). All 22 SNPs showed the same association direction as previously reported. Eight SNPs were nominally statistically significant with P≤0.05: rs4246511 (RHBDL2), rs12461110 (NLRP11), rs2307449 (POLG), rs12611091 (BRSK1), rs1172822 (BRSK1), rs365132 (UIMC1), rs2720044 (ASH2L), and rs7246479 (TMEM150B). Especially, SNPs rs4246511, rs365132, rs1172822, and rs7246479 remained significant even after Bonferroni correction. Significant associations were observed for GRS. Women in the highest quartile began menopause 0.7 years (P = 3.24×10(-9)) and 0.9 years (P = 4.61×10(-11)) later than those in the lowest quartile for GRSsel and GRSall, respectively.Among the 22 investigated SNPs, eight showed associations with ANM (P

BACKGROUND: There are urgent needs for rapid and accurate drug susceptibility testing of M. tuberculosis. GenoType MTBDRsl is a new molecular kit designed for rapid identification of the resistance to the second-line antituberculosis drugs with a single strip. In recent years, it has been evaluated in many settings, but with varied results. The aim of this meta-analysis was to synthesize the latest data on the diagnostic accuracy of GenoType MTBDRsl in detecting drug resistance to fluoroquinolones, amikacin, capreomycin, kanamycin and ethambutol, in comparison with the phenotypic drug susceptibility test. METHODS: This systematic review followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guideline. The search terms of "MTBDRsl" and "tuberculosis" were used on PubMed, EMBASE, and Web of Science. QUADAS-2 was used to assess the quality of included studies. Data were analyzed by Meta-Disc 1.4. We calculated the sensitivity, specificity, positive likelihood ratio (PLR), negative likelihood ratio (NLR), diagnostic odds ratio (DOR) and corresponding 95% confidence interval (CI) for each study. From these calculations, forest plots and summary receiver operating characteristic (SROC) curves were produced. RESULTS: Patient selection bias as well as flow and timing bias were observed in most studies. The summarized sensitivity (95% CI) was 0.874(0.845-0.899), 0.826(0.777-0.869), 0.820(0.772-0.862), 0.444(0.396-0.492), and 0.679(0.652-0.706) for fluoroquinolones, amikacin, capreomycin, kanamycin, and ethambutol, respectively. The specificity (95% CI) was 0.971(0.961-0.980), 0.995(0.987-0.998), 0.973(0.963-0.981), 0.993(0.985-0.997), and 0.799(0.773-0.823), respectively. The AUC (standard error) were 0.9754(0.0203), 0.9300(0.0598), 0.9885(0.0038), 0.9689(0.0359), and 0.6846(0.0550), respectively. CONCLUSION: Genotype MTBDRsl showed good accuracy for detecting drug resistance to fluoroquinolones, amikacin and capreomycin, but it may not be an appropriate choice for kanamycin and ethambutol. The lack of data did not allow for proper evaluation of the test on clinical specimens. Further systematic assessment of diagnostic performance should be carried out on direct clinical samples.

We compared exemplar strains from two hypervirulent clonal complexes, strain NMB-CDC from ST-8/11 cc and strain MC58 from ST-32/269 cc, in host cell attachment and invasion. Strain NMB-CDC attached to and invaded host cells at a significantly greater frequency than strain MC58. Type IV pili retained the primary role for initial attachment to host cells for both isolates regardless of pilin class and glycosylation pattern. In strain MC58, the serogroup B capsule was the major inhibitory determinant affecting both bacterial attachment to and invasion of host cells. Removal of terminal sialylation of lipooligosaccharide (LOS) in the presence of capsule did not influence rates of attachment or invasion for strain MC58. However, removal of either serogroup B capsule or LOS sialylation in strain NMB-CDC increased bacterial attachment to host cells to the same extent. Although the level of inhibition of attachment by capsule was different between these strains, the regulation of the capsule synthesis locus by the two-component response regulator MisR, and the level of surface capsule determined by flow cytometry were not significantly different. However, the diplococci of strain NMB-CDC were shown to have a 1.89-fold greater surface area than strain MC58 by flow cytometry. It was proposed that the increase in surface area without changing the amount of anchored glycolipid capsule in the outer membrane would result in a sparser capsule and increase surface hydrophobicity. Strain NMB-CDC was shown to be more hydrophobic than strain MC58 using hydrophobicity interaction chromatography and microbial adhesion-to-solvents assays. In conclusion, improved levels of adherence of strain NMB-CDC to cell lines was associated with increased bacterial cell surface and surface hydrophobicity. This study shows that there is diversity in bacterial cell surface area and surface hydrophobicity within N. meningitidis which influence steps in meningococcal pathogenesis.

Menopausal symptoms have been suggested to be an indicator of better prognosis among patients treated for breast cancer, because women who experience these symptoms usually have a lower level of estrogen. We tested this hypothesis in a population-based, prospective cohort study involving 4,842 women with stage 0 to III primary breast cancer who were enrolled in the Shanghai Breast Cancer Survival Study between March 2002 and April 2006, were aged 20 to 75 years, and were recruited 6 months post-diagnosis. They were followed-up by in-person surveys and record linkages with the vital statistics registry. Cox regression analysis was used to evaluate the association of menopausal symptoms at baseline with breast cancer recurrence. Approximately 56% of patients experienced at least one menopausal symptom, including hot flashes, night sweats, and/or vaginal dryness at baseline. During a median follow-up period of 5.3 years, 720 women had a recurrence. Experiencing hot flashes or having ≥2 menopausal symptoms was associated with lower risk of recurrence among premenopausal women (hazard ratio [HR]=0.77, 95% confidence interval [CI]: 0.62-0.96 for hot flashes; 0.73, 0.56-0.96 for ≥2 menopausal symptoms). Lower recurrence risk in relation to hot flashes was also observed among women who were not overweight/obese (HR=0.78, 95% CI: 0.64-0.99), those with relatively low waist-to-hip ratio (WHR) (HR=0.77, 95% CI: 0.61-0.97), and those who used tamoxifen (HR=0.75, 95% CI: 0.58-0.98). Consistently experiencing multiple menopausal symptoms was associated with lower recurrence risk among women with low WHR or who used tamoxifen. This large, population-based cohort study of women with breast cancer confirms that experiencing menopausal symptoms is an indicator of favorable breast cancer prognosis.