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Background: Transcription factors regulate gene expression and play important role in tumor genesis. Especially, the E2F transcription factor family controls the cell cycle and regulate many tumor suppressors. Missense variants in E2F family genes, which change the amino acid sequence, may alter the capacity for DNA binding or the protein structure, leading to a functional alteration. Material and Methods: We here searched for missense variants in E2F transcription family genes (E2F1~E2F8) and identified two (rs2075995 for E2F2 and rs3829295 for E2F7) with minor allele frequencies >0.01 in Chinese Han Beijing population from the 1000 genome project. We genotyped these two variants in 1,055 colorectal cancer (CRC) patients and 1,936 healthy controls using Taqman genotyping assays and assessed associations between SNPs and risk of CRC using logistic regression adjusted for gender and age. Results: We found rs3829295 at E2F7 to be significantly associated with risk of CRC. Compared with TT genotype carriers, CT and CT+CC genotype carriers had lower risks of CRC with ORs of 0.61 (95% CI: 0.44-0.85, P=0.003) and 0.61 (95% CI: 0.44-0.84, P=0.003), respectively. When stratified by gender and age, significant associations were observed in males (OR= 0.56, 95% CI: 0.38-0.83, P=0.004) for rs3829295, but not females (OR= 0.73, 95% CI: 0.43-1.22, P=0.232). Conclusion: Through a systematic assessment of variants in the E2F transcription factor family, we identified a lowfrequent missense variant in E2F7 significantly associated with CRC risk, indicating that E2F7 may play an important role in development of this tumor type., (Creative Commons Attribution License)

A highly sensitive and selective fluorescence method of quantitative detection for mercury in soil was developed using non-labeled molecular beacon (MB), single-stranded nucleic acid (ssDNA) and fluorescent dye Hoechst 33258. In this analytical method, the loop of MB was designed to be a sequence that was complementary to the ssDNA with multiple T-T mismatches, the stem of MB was completely designed as C-G base pairs, and both ends of the MB are not modified by any fluorophore and quencher. In the absence of Hg 2+ , the interaction between Hoechst 33258 and the MB was very weak, and the fluorescence signal of Hoechst 33258 was very low. In the presence of Hg 2+ , the MB and ssDNA with multiple T-T mismatches formed a double-stranded nucleic acid (dsDNA) via the T-Hg 2+ -T coordination structure which provided binding sites for Hoechst 33258. Then Hoechst 33258 binded to A-T base pairs of dsDNA, and the fluorescence intensity of Hoechst 33258 was significantly enhanced. Thus, a highly sensitive fluorescence quantitative detection method for Hg 2+ can be realized. In this strategy, the optimal determination conditions for Hg 2+ were a buffer solution pH of 8.2, an incubated temperature of 50°C, an incubated time of 5 min and NaCl of 60 mmol L -1 . Under the optimum conditions, the fluorescence intensity of Hoechst 33258 exhibited a good linear dependence on the concentration of Hg 2+ in the range of 5 × 10 -9 - 400 × 10 -9 mol L -1 . The fitted regression equation was ΔI = 2.1084C - 8.9587 with a correlation coefficient of 0.9943 (R 2 ), and the detection limit of this method was 3 × 10 -9 mol L -1 (3σ). The proposed method had a high selection; the common substances in soil such as Ca 2+ , Mg 2+ , Mn 2+ , Fe 3+ , Cu 2+ , Pb 2+ , Al 3+ , K + , Na + , Ni 2+ , Cd 2+ , Cr 3+ , SiO 3 2- , Cl - , PO 4 3- , NH 4 + and S 2- had no interference to the detection of mercury. The proposed method had a high accuracy, and it was applied to detect mercury of ten different types of soil; the recoveries were 97.65 - 103.22%. In addition, the proposed method had a low background emission, fast detection speed and low detection cost.

Programmed cell death (PCD) and immunity in plants are tightly controlled to promote antimicrobial defense while preventing autoimmunity. However, the mechanisms contributing to this immune homeostasis are poorly understood. Here, we isolated a rice mutant ebr1 (enhanced blight and blast resistance 1) that shows enhanced broad-spectrum bacterial and fungal disease resistance, but displays spontaneous PCD, autoimmunity, and stunted growth. EBR1 encodes an E3 ubiquitin ligase that interacts with OsBAG4, which belongs to the BAG (Bcl-2-associated athanogene) family that functions in cell death, growth arrest, and immune responses in mammals. EBR1 directly targets OsBAG4 for ubiquitination-mediated degradation. Elevated levels of OsBAG4 in rice are necessary and sufficient to trigger PCD and enhanced disease resistance to pathogenic infection, most likely by activating pathogen-associated molecular patterns-triggered immunity (PTI). Together, our study suggests that an E3-BAG module orchestrates innate immune homeostasis and coordinates the trade-off between defense and growth in plants., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Th17 and Th9 cells have been demonstrated to possess immune regulatory functions in malignant pleural effusion (MPE). However, whether IL-17 can affect differentiation and function of Th9 cells in MPE remains unknown. The objective of the present study was to explore the impact of IL-17 on the in vivo differentiation of Th9 cells in relation to Th2 cells in a murine model of MPE, and to explore whether IL-17 inhibits MPE formation via IL-9‒dependent mechanism. It was found that Th9 and Th2 cells were decreased in MPE from IL-17 -/- mice as compared with wild type mice. IL-17 deficiency inhibited Th9 and Th2 cell differentiation via suppressing transcription factors IRF4 and GATA-3, respectively. IL-17 deficiency enhanced MPE formation by promoting angiogenesis and proliferation of pleural tumors, and thus accelerated the death of mice bearing MPE. The in vivo administration of anti-IL-9 neutralizing mAb accelerated the death of WT mice; whereas administration of exogenous IL-9 improved the survival of IL-17 -/- mice. Our data provide the first definitive evidence that IL-17 promotes the differentiation of Th9 and Th2 cells in MPE. Our findings also demonstrate that IL-17 inhibits the formation of MPE and improves the survival of mice bearing MPE via an IL-9-dependent mechanism.

The YiQiFuMai powder injection (YQFM), a traditional Chinese medicine (TCM) prescription re-developed based on the well-known TCM formula Sheng-maisan, showed a wide range of pharmacological activities in cardiovascular diseases in clinics. However, its role in protection against myocardial ischemia/reperfusion (MI/R) injury has not been elucidated. The present study not only evaluated the cardioprotective effect of YQFM from MI/R injury but also investigated the potential molecular mechanisms both in vivo and in vitro. The myocardium infarct size, production of lactate dehydrogenase (LDH), creatine kinase (CK), cardiac function, TUNEL staining, and caspase-3 activity were measured. Cell viability was determined, and cell apoptosis was measured by Hoechst 33342 staining and flow cytometry. Mitochondrial membrane potential (ΔΨm) was measured, and ATP content was quantified by bioluminescent assay. Expression of apoptosis-related proteins, including Caspase-3, Bcl-2, Bax, AMPKα, and phospho-AMPKα, was analyzed by western blotting. AMPKα siRNA transfection was also applied to the mechanism elucidation. YQFM at a concentration of 1.06 g/kg significantly reduced myocardium infarct size and the production of LDH, CK in serum, improved the cardiac function, and also produced a significant decrease of apoptotic index. Further, combined treatment with compound C partly attenuated the anti-apoptotic effect of YQFM. In addition, pretreatment with YQFM ranging from 25 to 400 μg/mL markedly improved cell viability and decreased LDH release. Moreover, YQFM inhibited H9c2 apoptosis, blocked the expression of caspase-3, and modulated Bcl-2 and Bax proteins, leading to an increased mitochondrial membrane potential and cellular ATP content. Mechanistically, YQFM activated AMP-activated protein kinase (AMPK) signaling pathways whereas pretreatment with AMPK inhibitor Compound C and application of transfection with AMPKα siRNA attenuated the anti-apoptotic effect of YQFM. Our results indicated that YQFM could provide significant cardioprotection against MI/R injury, and potential mechanisms might suppress cardiomyocytes apoptosis, at least in part, through activating the AMPK signaling pathways.

Translocator Protein (18kDa, TSPO) is a mitochondrial outer membrane transmembrane protein. Its expression is elevated during inflammation and injury. However, the function of TSPO in vivo is still controversial. Here, we constructed a TSPO global knockout (KO) mouse with a Cre-LoxP system that abolished TSPO protein expression in all tissues and showed normal phenotypes in the physiological condition. The birth rates of TSPO heterozygote (Het) x Het or KO x KO breeding were consistent with Mendel's Law, suggesting a normal viability of TSPO KO mice at birth. RNA-seq analysis showed no significant difference in the gene expression profile of lung tissues from TSPO KO mice compared with wild type mice, including the genes associated with bronchial alveoli immune homeostasis. The alveolar macrophage population was not affected by TSPO deletion in the physiological condition. Our findings contradict the results of Papadopoulos, but confirmed Selvaraj's findings. This study confirms TSPO deficiency does not affect viability and bronchial alveolar immune homeostasis., Competing Interests: Beijing Thorgene Medical Laboratory provided support in the form of salaries for author FL. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Four porous carbon samples denoted as LSC-1, LSC-2, LCS-3, and LSC-4 were prepared by carbonization of loofah sponge pretreated by ZnCl₂ activation, immersion in N , N -dimethylformamide (DMF), DMF-assisted solvothermal and melamine-assisted hydrothermal processes, and the specific surface areas were 1007, 799, 773, and 538 m²·g -1 with mainly micropores, respectively. Electrocapacitive properties of four porous carbon-based electrodes were investigated with cyclic voltammetry, galvanostatic charge-discharge, and electrochemical impedance spectroscopy in symmetric supercapacitors. All the cyclic voltammetries of four types of supercapacitors showed a rectangular shape, even under a high scan rate of 500 mV·s -1 . The capacitances of LSC-1, LSC-2, LSC-3, and LSC-4 were 107.4, 92.5, 60.3, and 82.3 F·g -1 at the current density of 0.1 A·g -1 , respectively, and LSC-1 displayed the excellent capacitance retention of about 81.3% with a current density up to 5 A·g -1 . All supercapacitors showed excellent electrochemical stability, and the LSC-1-based supercapacitor showed a cycle stability with 92.6% capacitance retention after 5000 cycles at 1 A·g -1 . The structure-property relationship of LSC samples is discussed and analyzed on the basis of the experimental data.

Chitosan-montmorillonite composites can provide hydrophobicity and amino groups to enhance the performances of montmorillonite in wastewater treatment. In this paper, low molar mass chitosan (LC) and medium molar mass chitosan (MC) were selected to intercalate a sodium montmorillonite (Mt). The adsorption isotherm of LC and MC on the Mt and the pH dependency were measured. Thermo X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR) were used to characterize the chitosan-Mt composites. Results showed that with hydrochloric acid as the solvent of chitosans, increasing pH from 3 to 5.5 enhanced the adsorption of both LC and MC on the Mt, and when the equilibrium concentration of chitosans approached 300mg/L at pH 5.5, the amount of adsorbed chitosan reached the highest level of 0.203mg/mg for LC and 0.190mg/mg for MC, respectively. When the Mt was saturated with the chitosan, the amino groups on the chitosan were bonded with the Mt through electrostatic interaction and there was monolayer of the chitosan in the interlayer space of Mt, which were confirmed by FTIR spectra and XRD patterns, respectively., (Copyright © 2016. Published by Elsevier B.V.)

The C-2/C-2U Thomson scattering system has been substantially upgraded during the latter phase of C-2/C-2U program. A Rayleigh channel has been added to each of the three polychromators of the C-2/C-2U Thomson scattering system. Onsite spectral calibration has been applied to avoid the issue of different channel responses at different spots on the photomultiplier tube surface. With the added Rayleigh channel, the absolute intensity response of the system is calibrated with Rayleigh scattering in argon gas from 0.1 to 4 Torr, where the Rayleigh scattering signal is comparable to the Thomson scattering signal at electron densities from 1 × 10 13 to 4 × 10 14 cm -3 . A new signal processing algorithm, using a maximum likelihood method and including detailed analysis of different noise contributions within the system, has been developed to obtain electron temperature and density profiles. The system setup, spectral and intensity calibration procedure and its outcome, data analysis, and the results of electron temperature/density profile measurements will be presented.

The coexistence and coupling between magnetization and electric polarization in multiferroic materials provide extra degrees of freedom for creating next-generation memory devices. A variety of concepts of multiferroic or magnetoelectric memories have been proposed and explored in the past decade. Here we propose a new principle to realize a multilevel nonvolatile memory based on the multiple states of the magnetoelectric coefficient (α) of multiferroics. Because the states of α depends on the relative orientation between magnetization and polarization, one can reach different levels of α by controlling the ratio of up and down ferroelectric domains with external electric fields. Our experiments in a device made of the PMN-PT/Terfenol-D multiferroic heterostructure confirm that the states of α can be well controlled between positive and negative by applying selective electric fields. Consequently, two-level, four-level, and eight-level nonvolatile memory devices are demonstrated at room temperature. This kind of multilevel magnetoelectric memory retains all the advantages of ferroelectric random access memory but overcomes the drawback of destructive reading of polarization. In contrast, the reading of α is nondestructive and highly efficient in a parallel way, with an independent reading coil shared by all the memory cells.

Aims/hypothesis: CUG-binding protein 1 (CUGBP1) is a multifunctional RNA-binding protein that regulates RNA processing at several stages including translation, deadenylation and alternative splicing, as well as RNA stability. Recent studies indicate that CUGBP1 may play a role in metabolic disorders. Our objective was to examine its role in endocrine pancreas function through gain- and loss-of-function experiments and to further decipher the underlying molecular mechanisms., Methods: A mouse model in which type 2 diabetes was induced by a high-fat diet (HFD; 60% energy from fat) and mice on a standard chow diet (10% energy from fat) were compared. Pancreas-specific CUGBP1 overexpression and knockdown mice were generated. Different lengths of the phosphodiesterase subtype 3B (PDE3B) 3' untranslated region (UTR) were cloned for luciferase reporter analysis. Purified CUGBP1 protein was used for gel shift experiments., Results: CUGBP1 is present in rodent islets and in beta cell lines; it is overexpressed in the islets of diabetic mice. Compared with control mice, the plasma insulin level after a glucose load was significantly lower and glucose clearance was greatly delayed in mice with pancreas-specific CUGBP1 overexpression; the opposite results were obtained upon pancreas-specific CUGBP1 knockdown. Glucose- and glucagon-like peptide1 (GLP-1)-stimulated insulin secretion was significantly attenuated in mouse islets upon CUGBP1 overexpression. This was associated with a strong decrease in intracellular cAMP levels, pointing to a potential role for cAMP PDEs. CUGBP1 overexpression had no effect on the mRNA levels of PDE1A, 1C, 2A, 3A, 4A, 4B, 4D, 7A and 8B subtypes, but resulted in increased PDE3B expression. CUGBP1 was found to directly bind to a specific ATTTGTT sequence residing in the 3' UTR of PDE3B and stabilised PDE3B mRNA. In the presence of the PDE3 inhibitor cilostamide, glucose- and GLP-1-stimulated insulin secretion was no longer reduced by CUGBP1 overexpression. Similar to CUGBP1, PDE3B was overexpressed in the islets of diabetic mice., Conclusions/interpretation: We conclude that CUGBP1 is a critical regulator of insulin secretion via activating PDE3B. Repressing this protein might provide a potential strategy for treating type 2 diabetes.

Neospora caninum is one of the important causes of abortion in dairy cattle worldwide. The dog is known as a definitive host of N. caninum and can transmit the parasite to cattle by shedding oocysts. The aim of the present study is to detect the presence of N. caninum in feces of dairy farm dogs and determine the genetic characteristics of N. caninum in Central China. A total of 78 fecal samples were collected from dogs in dairy farms from May to November 2014 and examined by microscopy and nested PCR based on Nc5 gene. Neospora-like oocysts were microscopically detected in two fecal samples, of which only one (Nc-LY1) was confirmed to be N. caninum by nested PCR. Seven out of 78 fecal samples (9.0 %) were N. caninum DNA positive, of which Neospora-like oocysts were simultaneously microscopically detected only in one sample (Nc-LY1). No statistical associations were found between the positive rates and age or sex of dogs (P > 0.05). The N. caninum-positive DNA samples were further analyzed by multilocus microsatellite (MS) genotyping for MS4, MS5, MS6A, MS7, MS8, MS10, MS12, and Cont-14. Only the fecal sample in which oocysts were detected was successfully genotyped at all genetic loci, and a new genotype was identified. To our knowledge, this study is the first report of genetic characterization of N. caninum isolates from naturally infected dogs based on multilocus microsatellites in China.

Although it is curable, tuberculosis remains one of the most frequent causes of pleural effusions on a global scale, especially in developing countries. Tuberculous pleural effusion (TPE) is one of the most common forms of extrapulmonary tuberculosis. TPE usually presents as an acute illness with fever, cough and pleuritic chest pain. The pleural fluid is an exudate that usually has predominantly lymphocytes. The gold standard for the diagnosis of TPE remains the detection of Mycobacterium tuberculosis in pleural fluid, or pleural biopsy specimens, either by microscopy and/or culture, or the histological demonstration of caseating granulomas in the pleura along with acid fast bacilli, Although adenosine deaminase and interferon-γ in pleural fluid have been documented to be useful tests for the diagnosis of TPE. It can be accepted that in areas with high tuberculosis prevalence, the easiest way to establish the diagnosis of TPE in a patient with a lymphocytic pleural effusion is to generally demonstrate a adenosine deaminase level above 40 U/L. The recommended treatment for TPE is a regimen with isoniazid, rifampin, and pyrazinamide for two months followed by four months of two drugs, isoniazid and rifampin.

The stretching of smooth muscle tissue modulates contraction through augmentation of Ca(2+) transients, but the mechanism underlying stretch-induced Ca(2+) transients is still unknown. We found that mechanical stretching and maintenance of mouse urinary bladder smooth muscle strips and single myocytes at 30% and 18% beyond the initial length, respectively, resulted in Ca(2+) oscillations. Experiments indicated that mechanical stretching remarkably increased the production of nitric oxide (NO) as well as the amplitude and duration of muscle contraction. Stretch-induced Ca(2+) oscillations and contractility increases were completely abolished by the NO inhibitor L-NAME or eNOS (also known as NOS3) gene inactivation. Moreover, exposure of eNOS-knockout myocytes to exogenous NO donor induced Ca(2+) oscillations. The stretch-induced Ca(2+) oscillations were greatly inhibited by the selective inositol 1,4,5-trisphosphate receptor (IP3R) inhibitor xestospongin C and partially inhibited by ryanodine. Moreover, the stretch-induced Ca(2+) oscillations were also suppressed by the phosphoinositide 3-kinase (PI3K) inhibitor LY294002, but not by the soluble guanylyl cyclase (sGC) inhibitor ODQ. These results suggest that stretching myocyte and maintenance at a certain length results in Ca(2+) oscillations that are NO dependent , and sGC and cGMP independent, and results from the activation of PI3K in smooth muscle., (© 2016. Published by The Company of Biologists Ltd.)

Cancer cell migration is involved in tumour metastasis. However, the relationship between calcium signalling and cancer migration is not well elucidated. In this study, we used the human lung adenocarcinoma A549 cell line to examine the role of endoplasmic reticulum protein 44 (ERP44), which has been reported to regulate calcium release inside of the endoplasmic reticulum (ER), in cell migration. We found that the inositol 1,4,5-trisphosphate receptors (IP3Rs/ITPRs) inhibitor 2-APB significantly inhibited A549 cell migration by inhibiting cell polarization and pseudopodium protrusion, which suggests that Ca2+ is necessary for A549 cell migration. Similarly, the overexpression of ERP44 reduced intracellular Ca2+ release via IP3Rs, altered cell morphology and significantly inhibited the migration of A549 cells. These phenomena were primarily dependent on IP3R2 because wound healing in A549 cells with IP3R2 rather than IP3R1 or IP3R3 siRNA was markedly inhibited. Moreover, the overexpression of ERP44 did not affect the migration of the human neuroblastoma cell line SH-SY5Y, which mainly expresses IP3R1. Based on the above observations, we conclude that ERP44 regulates A549 cell migration mainly via an IP3R2-dependent pathway.

Background: Disrupted Ca2+ homeostasis contributes to the development of colonic dysmotility in ulcerative colitis (UC), but the underlying mechanisms are unknown. This study aimed to examine the alteration of colonic smooth muscle (SM) Ca2+ signaling and Ca2+ handling proteins in a rat model of dextran sulfate sodium (DSS)-induced UC., Methods: Male Sprague-Dawley rats were randomly divided into control (n = 18) and DSS (n = 17) groups. Acute colitis was induced by 5% DSS in the drinking water for 7 days. Contractility of colonic SM strips (controls, n = 8 and DSS, n = 7) was measured in an organ bath. Cytosolic resting Ca2+ levels (n = 3 in each group) and Ca2+ transients (n = 3 in each group) were measured in single colonic SM cells. Ca2+ handling protein expression was determined by Western blotting (n = 4 in each group). Differences between control and DSS groups were analyzed by a two-sample independent t-test., Results: Average tension and amplitude of spontaneous contractions of colonic muscle strips were significantly enhanced in DSS-treated rats compared with controls (1.25 ± 0.08 g vs. 0.96 ± 0.05 g, P= 0.007; and 2.67 ± 0.62 g vs. 0.52 ± 0.10 g, P= 0.013). Average tensions of carbachol-evoked contractions were much weaker in the DSS group (1.08 ± 0.10 g vs. 1.80 ± 0.19 g, P= 0.006). Spontaneous Ca2+ transients were observed in more SM cells from DSS-treated rats (15/30 cells) than from controls (5/36 cells). Peak caffeine-induced intracellular Ca2+ release was lower in SM cells of DSS-treated rats than controls (0.413 ± 0.046 vs. 0.548 ± 0.041, P= 0.033). Finally, several Ca2+ handling proteins in colonic SM were altered by DSS treatment, including sarcoplasmic reticulum calcium-transporting ATPase 2a downregulation and phospholamban and inositol 1,4,5-trisphosphate receptor 1 upregulation., Conclusions: Impaired intracellular Ca2+ signaling of colonic SM, caused by alteration of Ca2+ handing proteins, contribute to colonic dysmotility in DSS-induced UC.

Bitter taste receptors (TAS2Rs) are traditionally thought to be expressed exclusively on the taste buds of the tongue. However, accumulating evidence has indicated that this receptor family performs non-gustatory functions outside the mouth in addition to taste. Here, we examined the role of TAS2Rs in human and mouse detrusor smooth muscle (DSM). We showed that mRNA for various TAS2R subtypes was expressed in both human and mouse detrusor smooth muscle (DSM) at distinct levels. Chloroquine (CLQ), an agonist for TAS2Rs, concentration-dependently relaxed carbachol- and KCl-induced contractions of human DSM strips. Moreover, 100 µM of CLQ significantly inhibited spontaneous and electrical field stimulation (EFS)-induced contractions of human DSM strips. After a slight contraction, CLQ (1 mM) entirely relaxed carbachol-induced contraction of mouse DSM strips. Furthermore, denatonium and quinine concentration-dependently decreased carbachol-induced contractions of mouse DSM strips. Finally, we demonstrated that CLQ treatment significantly suppressed the overactive bladder (OAB) symptoms of mice with partial bladder outlet obstruction (PBOO). In conclusion, we for the first time provide evidence of the existence of TAS2Rs in the urinary DSM and demonstrate that TAS2Rs may represent a potential target for OAB. These findings open a new approach to develop drugs for OAB in the future., Competing Interests: CONFLICTS OF INTERESTS The authors declare no financial conflicts of interest.

Parkinson's disease (PD) is the second most common age-related neurodegenerative disease. MicroRNA-7 (miR-7) displays neuroprotective properties against PD. However, the biological roles of miR-7 and its underlying molecular mechanisms in PD remain unclear. We demonstrated herein that 1-methyl-4-phenylpyridinium ion (MPP(+)) confers toxic effects on dopaminergic neuron in a dose-dependent manner in a cellular PD model, although this phenomenon is attenuated by miR-7 treatment. Introduction of miR-7 inhibits MPP(+)-induced neuronal apoptosis as reflected by the reduced terminal transferase-mediated dUTP nick end labeling-positive rate, mitochondrial permeability potential, caspase 3 activity, and nucleosomal enrichment factor. Bax and sirtuin 2 (Sirt2) are the direct targets of miR-7. Moreover, the effects of miR-7 were counteracted by Bax and Sirt2 overexpression, respectively. The altered molecular expressions downstream of Bax and Sirt2 are also involved in miR-7 regulation of the MPP(+)-triggered neuronal apoptosis. These findings have implications on the potential application of miR-7 in PD treatment.

Background: It is often challenging to distinguish tuberculous pleural effusion (TPE) from malignant pleural effusion (MPE); thoracoscopy is among the techniques with the highest diagnostic ability in this regard. However, such invasive examinations cannot be performed on the elderly, or on those in poor physical condition. The aim of this study was to explore the differential diagnostic value of carbohydrate antigen 125 (CA125), carbohydrate antigen 199 (CA199), carcinoembryonic antigen (CEA), neuron-specific enolase (NSE), and squamous cell carcinoma (SCC) associated antigen in patients with TPE and MPE., Methods: Using electrochemiluminescence, we measured the concentration of tumor markers (TMs) in the pleural effusion and serum of patients with TPE (n = 35) and MPE (n = 95). We used receiver operating characteristic (ROC) curve analysis to evaluate the TMs and differentiate between TPE and MPE., Results: The cut-off values for each TM in serum were: CA125, 151.55 U/ml; CA199, 9.88 U/ml; CEA, 3.50 ng/ml; NSE, 13.27 ng/ml; and SCC, 0.85 ng/ml. Those in pleural fluid were: CA125, 644.30 U/ml; CA199, 12.08 U/ml; CEA, 3.35 ng/ml; NSE, 9.71 ng/ml; and SCC, 1.35 ng/ml. The cut-off values for the ratio of pleural fluid concentration to serum concentration (P/S ratio) of each TM were: CA125, 5.93; CA199, 0.80; CEA, 1.47; NSE, 0.76; and SCC, 0.90. The P/S ratio showed the highest specificity in the case of CEA (97.14%). ROC curve analysis revealed that, for all TMs, the area under the curve in pleural fluid (0.95) was significantly different from that in serum (0.85; P < 0.001)., Conclusions: TMs in TPE differ significantly from those in MPE, especially when detected in pleural fluid. The combined detection of TMs can improve diagnostic sensitivity.

The numbers of IL-27(+) CD4(+) and IL-27(+) CD8(+) T cells have been found to be increased in tuberculous pleural effusion (TPE) as compared with malignant pleural effusion (MPE). The objective of the present study was to investigate whether pleural IL-27(+) CD4(+) and IL-27(+) CD8(+) T cells can distinguish patients with TPE from those with MPE. Paired specimen of pleural fluid and peripheral blood were collected from 35 patients with TPE and 46 MPE. The numbers of IL-27(+) CD4(+) and IL-27(+) CD8(+) T cells were simultaneously determined by flow cytometry. Receiver operating characteristic curve analysis was used to evaluate the capacity of IL-27(+) CD4(+) and IL-27(+) CD8(+) T cells to differentiate TPE from MPE. The sensitivity, specificity, positive likelihood ratio (PLR), negative likelihood ratio (NLR), positive predictive value (PPV), and negative predictive value (NPV) of IL-27(+) CD4(+) T cells were 94.3%, 93.5%, 14.46, 0.06, 91.7%, and 95.6%, respectively. The sensitivity, specificity, PLR, NLR, PPV and NPV of IL-27(+) CD8(+) T cells were 80.0%, 93.5%, 12.27, 0.21, 90.3% and 86.0%, respectively. The number of IL-27(+) CD4(+) in pleural fluid is a helpful diagnostic biomarker for the diagnosis of TPE, which performs better than that of IL-27(+) CD8(+) T cells.

The diagnosis of extrapulmonary tuberculosis (EPTB) is difficult. In recent years, T-cell interferon-γ release assays (IGRAs) are widely used in diagnosing tuberculosis. The aim of this meta-analysis is to evaluate the diagnostic accuracy of body fluid IGRAs in diagnosing EPTB. The PubMed, EMBASE, Web of Science, and Cochrane bibliographies were searched for English language articles. 22 studies met the inclusion criteria. The pooled sensitivity and specificity of body fluid IGRAs for diagnosing EPTB were 0.87 [95% confidence interval (CI): 0.83-0.92] and 0.85 (95% CI: 0.79-0.90), respectively. For the fluid T-SPOT.TB, the pooled sensitivity and specificity were 0.92 (95% CI: 0.88-0.95) and 0.85 (95% CI: 0.78-0.91), respectively. The diagnostic odds ratio (DOR) of the fluid T-SPOT.TB was 46.99 (95% CI: 13.69-161.28) for tuberculosis pleurisy, 26.46 (95% CI: 11.38-61.56) for tuberculosis peritonitis, and 97.86 (95% CI: 25.31-378.45) for tuberculosis meningitis. The application of T-SPOT. TB in the diagnosis of EPTB performed better in the body fluid than in the blood. The diagnostic values of the fluid T-SPOT.TB varied for different fluid categories. However, the utility of T-SPOT.TB was limited due to its suboptimal accuracy and higher cost compared with conventional tests.

Toll-like receptor 4 (TLR4) is involved in multiple malignancies; however, the role of TLR4 in the pathogenesis of malignant pleural effusion (MPE) remains unknown. The objectives of this study were to explore the impact of TLR4 signaling on the development of MPE in a murine model and to define the underline mechanisms by which TLR works. Development of MPE as well as proliferation and angiogenesis of pleural tumor were determined in TLR4(-/-) and wild type mice. Differentiation of Th1 and Th17 cells as well as their signal transductions was explored. The effects of TLR4 signaling on survival of mice bearing MPE were also investigated. Compared with wild type mice, Th1 cells were augmented, and Th17 cells were suppressed in MPE from TLR4(-/-) mice. The in vitro experiments showed that TLR4 deficiency promoted Th1 cell differentiation via enhancing STAT1 pathway and inhibited Th17 cell differentiation via suppressing STAT3 pathway. TLR4 deficiency promoted MPE formation and, thus, accelerated the death of mice bearing MPE, whereas intraperitoneal injection of anti-IFN-γ mAb or recombinant mouse IL-17 protein into TLR4(-/-) mice was associated with improved survival. Our data provides the first definitive evidence of a role for TLR4 signaling in protective immunity in the development of MPE. Our findings also demonstrate that TLR4 deficiency promotes MPE formation and accelerates mouse death by enhancing Th1 and suppressing Th17 response., (© 2015 International Federation for Cell Biology.)

Toxoplasma gondii is an obligate intracellular parasite that has a remarkable ability to infect almost all warm-blooded animals, including humans. This study was aimed to determine the genetic characteristics of T. gondii isolates from domestic animals in Henan Province, central China. A total of 363 DNA samples, including 208 from hilar lymph nodes of pigs, 36 from blood samples of cats, 12 from tissues of aborted bovine fetuses and 107 from blood samples of dams with history of abortion in Henan Province, were examined for the presence of T. gondii by nested PCR based on B1 gene. The positive DNA samples were further genotyped by PCR-RFLP at 11 markers, including SAG1, (3'+ 5') SAG2, alt.SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico. DNA samples from 9 pigs, 5 cats, and 4 dairy cows were T. gondii B1 gene positive. Nine samples were successfully genotyped at all genetic loci, of which 5 samples from pigs, and 2 from cats were identified as ToxoDB genotype #9, and 2 samples from cows belonged to ToxoDB genotype #225. To our knowledge, the present study is the second report of genetic typing of T. gondii isolates from cattle in China, and the first report of T. gondii ToxoDB#225 from cattle.

The presence of HPV in breast tissue and the potential causal association between human papillomavirus (HPV) and breast cancer (BC) remains controversial. The aim of the present study was to compare the HPV prevalence in BC tissues, adjacent normal breast tissues and breast benign disease tissues and to investigate the possible association between HPV and breast tumor development in Chinese women. Paraffin-embedded specimens from 187 pairs of BCs including tumor and normal breast tissue adjacent to tumors and 92 breast benign lesions between June 2009 and July 2014 were investigated by nested polymerase chain reaction (PCR) and type-specific PCR, respectively. With strictly quality control, HPV positive infection was detected in three BC tissues. No HPV positive infection was detected in all normal breast tissue adjacent to tumors and benign breast tissues. Through our detailed analysis, rare HPV infection in this study suggests that HPV might not be associated with BC progression.

Background: The numbers of IL-27-producing CD4(+) T cells and the concentration of soluble IL-27 have been found to be increased in tuberculous pleural effusion (TPE). The objective of the present study was to explore the mechanism by which IL-27(+)CD4(+) T cells are recruited into the pleural space, and to explore the impact of IL-27 on pleural mesothelial cells (PMCs)., Methods: The expression profiles of chemokine receptor (CCR) were determined by flow cytometry. The chemoattractant activity of chemokines CCL20 and CCL22 for IL-27(+)CD4(+) T cells in vitro was observed. Effects of IL-27 on wound healing, proliferation and apoptosis of PMCs were also investigated., Results: IL-27(+)CD4(+) T cells in TPE expressed high level of CCR6, medium level of CCR4, and low levels of CCR2, CCR3, CCR5, CCR7, CCR10, and CXCR3. Recruitment of IL-27(+)CD4(+) T cells into TPE could be induced by pleural CCL20 and CCL22. By activating STAT3 signaling, IL-27 significantly improved wound healing and promoted proliferation of PMCs, and completely prevented apoptosis of PMCs induced by IFN-γ., Conclusions: After being recruited into pleural space by CCL20 or/and CCL22, these pleural IL-27-producing CD4(+) T cells may play important roles in tuberculosis immunity by affecting PMC functions.

Tumour necrosis factor-α (TNF-α) is critical in the regulation of inflammation and tumour progression. TNF-α-308G > A is associated with constitutively elevated TNF-α expression. The purpose of this study was to assess the association between TNF-α-308G > A and breast cancer (BC) risk by subtype and the connection between genotypes and clinical features of BC. A total of 768 patients and 565 controls were enrolled in this study, and genotypes were detected using the TaqMan assay. No effect on susceptibility for any BC subtype was found for the TNF-α-308 polymorphism in our study or in the pooled meta-analysis. This polymorphism was shown to be associated with age at menarche in all BC and in progesterone receptor-negative BC. Interestingly, triple negative breast cancer (TNBC) patients with TNF-α-308A had an increased risk of distant tumour metastasis (OR = 3.80, 95% CI: 1.31-11.02, P = 0.009). Multi-regression analysis showed that TNF-α-308A was also a risk factor for distant tumour metastasis after adjustment for tumour size and lymph node metastasis status (OR = 6.26, 95% CI: 1.88-20.87, P = 0.003). These findings indicate that TNF-α might play a distinct role in the progression of TNBC, especially in distant tumour metastasis of TNBC.

Background: Advanced glycation end products (AGEs) are a group of stable covalent compounds generated by proteins, lipids, other macromolecules and sugar through a series of non-enzymatic reactions. As reported, AGEs can cause widespread pathophysiological responses through activation of AGE receptors (RAGEs) on the cell surface, and play an important role in the pathogenesis of osteoarthritis (OA). We hypothesized that the antioxidant and anti-glycan agent gallic acid (GA) can work against the effects of AGEs and can be used as a potential drug for the cure of OA., Methods: The present study first explored the negative functions of AGEs via AGE-treated chondrocytes isolated form rabbits. Then, we observed the protective role of GA in AGE-treated chondrocytes by measuring the reactive oxygen species (ROS), superoxide dismutase (SOD), collagen II, aggrecan, nitric oxide synthase (iNOs) and cyclooxygenase-2 (COX-2) in vitro. Finally, the changes in a cartilage lesion in a rabbit model of knee osteoarthritis was observed., Results: Exposure of chondrocytes to AGEs resulted in a reduction of ROS, SOD, collagen II and aggrecan, and an increase in iNOs and COX-2, which means exposure promoted OA lesions in a clinical setting. When AGE-treated chondrocytes were pretreated with GA, there were no significant changes in these key components compared to the normal chondrocytes. In vivo study showed cartilage degradation was reduced by GA as compared to the vehicle group., Conclusion: The results of this study confirmed the chondroprotective role of GA and provide a potential drug for the relief of OA.

Ophiopogonin-D is one of steroidal saponins isolated from the root of the Chinese medicinal plant Ophiopogon japonicas. It has been claimed to possess anti-inflammatory and anti-oxidant properties. The present study was the first to examine the anti-tumor metastasis properties of ophiopogonin-D. An MTT assay showed that ophiopogonin-D inhibited the proliferation of MDA-MB-435 melanoma cells, and decreased invasion was demonstrated using a Transwell invasion assay. Furthermore, adhesion of MDA-MB-435 cells to human umbilical vascular endothelial cells and to fibronectin was inhibited by ophiopogonin-D. Gelatin zymography and western blot analysis showed that ophiopogonin-D inhibited the expression and secretion of matrix metalloproteinase-9 (MMP-9), but not that of MMP-2. Inhibition of phosphorylation of p38 by ophiopogonin-D indicated its inhibition of the mitogen-activated protein kinase pathway. Overall, the results suggested that ophiopogonin-D may be considered as a candidate drug for treating or preventing tumor metastasis.

Non-muscle myosin heavy chain IIA (NMMHC IIA) has been shown to be involved in thrombus formation and inflammatory microparticle release in endothelial cells. However, the role of NMMHC IIA in regulating the expression of tissue factor (TF) and deep venous thrombosis remains to be elucidated. In the present study, endothelial cells were stimulated with tumour necrosis factor-α (TNF-α) to induce TF expression. Pretreatment with the NMMHC II inhibitor blebbistatin suppressed the mRNA and protein expressions as well as the procoagulant activity of TF in a dose-dependent manner. Blebbistatin enhanced Akt and GSK3β phosphorylation and inhibited NF-κB p65 nuclear translocation and IκBα degradation. These observations were similar to the effect of CHIR99021, a GSK3β inhibitor. TF downregulation by blebbistatin was antagonised by the PI3K inhibitor, wortmannin. Furthermore, siRNA knockdown of NMMHC IIA, but not IIB or IIC, inhibited TF expression, activated Akt/GSK3β and suppressed NF-κB signalling pathways, whereas the overexpression of NMMHC IIA increased TF expression. The binding of NMMHC IIA and TNF receptor 2 mediated signal internalisation in TNF-α-stimulated endothelial cells. Importantly, blebbistatin decreased endothelium NMMHC IIA and TF expression, deactivated GSK3β by inducing its phosphorylation, suppressed p65 nuclear translocation, and inhibited thrombus formation in a mouse deep venous thrombosis model.Our findings provide solid evidence that inhibition of NMMHC II, most likely NMMHC IIA, impedes TF expression and venous thrombosis via Akt/GSK3β-NF-κB signalling pathways in the endothelium both in vitro and in vivo. NMMHC IIA might be a potential novel target for the treatment of thrombotic disorders.

This study aimed to explore the effect of DT-13 (25(R,S)-ruscogenin- 1-O- [β-d-glucopyranosyl- (1→2)][β-d-xylopyranosyl-(1→3)]-β -d- fucopyranoside) on tumor necrosis factor (TNF)-α-induced vascular inflammation and the potential molecular mechanisms. In vitro, DT-13 suppressed TNF-α-induced adhesion and migration of human umbilical vein endothelial cells (HUVECs) by inhibiting the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). DT-13 markedly suppressed NF-кB p65 phosphorylation, and when NF-кB p65 was over-expressed, the inhibitory effect of DT-13 on adhesion molecular decreased. DT-13 also suppressed TNF-α induced luciferase activities of ICAM-1 and VCAM-1 promoter containing NF-κB binding sites. Furthermore, DT-13 markedly suppressed p38 phosphorylation and Src degradation induced by TNF-α, whereas had no significant effect on ERK and JNK activation. In vivo, DT-13 at 4 mg/kg prevented vascular inflammation and the expression of adhesion molecules induced by TNF-α in mice. These findings suggest that DT-13 abrogates vascular inflammation by down-regulating adhesion molecules associated with modulating the NF-кB, p38MAPK, Src signaling pathways, and NF-κB binding site is at least one of the targets of DT-13. This study provides novel information regarding the mechanism by which DT-13 exerts its effects on vascular inflammation, which is important for the onset and progression of various diseases.

The participation of large-conductance Ca2+ activated K+ channels (BKs) in chloroquine (chloro)-induced relaxation of precontracted airway smooth muscle (ASM) is currently undefined. In this study we found that iberiotoxin (IbTx, a selective inhibitor of BKs) and chloro both completely blocked spontaneous transient outward currents (STOCs) in single mouse tracheal smooth muscle cells, which suggests that chloro might block BKs. We further found that chloro inhibited Ca2+ sparks and caffeine-induced global Ca2+ increases. Moreover, chloro can directly block single BK currents completely from the intracellular side and partially from the extracellular side. All these data indicate that the chloro-induced inhibition of STOCs is due to the blockade of chloro on both BKs and ryanodine receptors (RyRs). We also found that low concentrations of chloro resulted in additional contractions in tracheal rings that were precontracted by acetylcholine (ACH). Increases in chloro concentration reversed the contractile actions to relaxations. In the presence of IbTx or paxilline (pax), BK blockers, chloro-induced contractions were inhibited, although the high concentrations of chloro-induced relaxations were not affected. Taken together, our results indicate that chloro blocks BKs and RyRs, resulting in abolishment of STOCs and occurrence of contraction, the latter will counteract the relaxations induced by high concentrations of chloro.

The potential causal association between human papillomavirus (HPV) and lung cancer (LC) remains controversial. We performed this meta-analysis to evaluate whether HPV infection in lung tissue is associated with LC compared with non-cancer controls. We also quantified this association in different LC subtypes. MEDLINE, EMBASE and Web of Science were searched through March 2014, using the search terms "lung cancer", "human papillomavirus", "HPV" and their combinations. Association was tested using odds ratio (OR) with 95% confidence intervals (95% CI). Heterogeneity was assessed using Q and I(2) statistic. Finally, nine studies, for a total of 1094 LCs and 484 non-cancer controls, were identified as eligible publications. The pooled results showed that HPV infection was associated with LC (OR=5.67, 95% CI: 3.09-10.40, P<0.001). Similar results were also observed in HPV16 and/or HPV18 (HPV16/18) infection analyses (OR=6.02, 95% CI: 3.22-11.28, P<0.001). HPV16/18 was significantly associated with lung squamous cell carcinoma (SCC) (OR=9.78, 95% CI: 6.28-15.22, P<0.001), while the pooled OR was 3.69 in lung adenocarcinoma (95% CI: 0.99-13.71, P=0.052). Our results suggest that lung tissue with HPV infection has a strong association with LC, and especially, HPV16/18 infection significantly increases SCC risk, which indicates a potential pathogenesis link between HPV and LC., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The prevalence and genotype of Toxoplasma gondii infection in dogs in Henan Province, Central China was investigated. A total of 125 blood samples were collected from pet dogs during April to June 2013, and all samples were examined by indirect hemagglutination antibody test (IHA) and nested PCR. The overall T. gondii prevalence in pet dogs was 24.0% (30/125), with 20.8% (26/125) in IHA and 10.4% (13/125) in PCR, respectively. No statistical associations were found between animal gender and age and the prevalence of T. gondii infection. Thirteen positive DNA samples were genotyped using 11 PCR-RFLP markers, including SAG1, (3'+5') SAG2, alt.SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico. Of these, only 2 samples were genotyped with complete data for all loci, and a novel genotype (type III at SAG3 and GRA6 loci, and type I at other loci) was identified. This is the first report of genetic characterization of T. gondii infection in dogs in China.

Calcium (Ca(2+)) oscillations play a central role in varieties of cellular processes including fertilization and immune response, but controversy over the regulation mechanisms still exists. It has been known that nitric oxide (NO) dependently regulates Ca(2+) signaling in most physiopathological processes. Previous study indicated that eNOS translocation during some pathological process influences intracellular Ca(2+) homeostasis. In this study, we investigated the role and mechanism of NO on Ca(2+) release by overexpressing eNOS in cytoplasm (Cyto-eNOS) and endoplasmic reticulum (ER-eNOS) of HeLa cells. We found that the properties of Ca(2+) release were altered by the overexpression of eNOS. The amplitude and frequency of extracellular ATP (eATP)-induced Ca(2+) oscillation were enhanced in both Cyto-eNOS and ER-eNOS cells, respectively. Especially, the enhancement of the amplitude and frequency of the Ca(2+) oscillation was much more significant in the ER-eNOS cells than that of Cyto-eNOS cells. Further study indicated that this effect was abrogated by NO inhibitor, L-NAME, suggesting it was not an artificial result induced by ER stress. Furthermore, an up-regulated phosphorylation of phospholamban (PLB) was observed and the sarco-endoplasmic reticulum Ca(2+)-ATPase (SERCA) function was activated followed by the significant increase in the ER Ca(2+) load. Taken together, we revealed a novel regulatory mechanism of Ca(2+) oscillation., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: Nonsyndromic oral clefts are complex in cause and have multiple genetic and environmental risk factors. This retrospective, questionnaire-based, case-control study investigated the relationship between oral clefts and parental mental and physical health and social support., Methods: Three hundred forty-seven parents of children with nonsyndromic oral clefts and 420 controls were included. Maternal and paternal health during the first trimester was assessed using interviews and questionnaires modeled from the Cornell Medical Index and the Social Support Rating Scale. Case-control analyses were performed using t tests, chi-square tests, and logistic regression., Results: Parental age, household income, and subsisting on farming were significantly different for cases and controls. The Cornell Medical Index for cases was significantly worse compared with controls for physical and psychological health. Logistic regression showed that nine factors were significantly associated with oral clefts: paternal respiratory health (OR, 1.56; p = 0.03), maternal gastrointestinal health (OR, 1.71; p < 0.01), maternal musculoskeletal health (OR, 1.50; p < 0.01), paternal nervous system health (OR, 2.82; p < 0.01), maternal frequency of illness (OR, 2.21; p = 0.01), maternal diseases (OR, 2.44; p < 0.01), maternal health habits (OR, 1.73; p < 0.01), paternal feelings of inadequacy (OR, 2.28; p = 0.03), and maternal anger (OR, 2.28; p < 0.01) in the first trimester. Weaker social support from the community was associated with oral clefts (p < 0.01)., Conclusion: Maternal and paternal health and social support may affect a family's risk of having a child with a cleft., Clinical Question/level of Evidence: Risk, III.

Background and Purpose: This study examined the role of the main vascular cAMP-hydrolysing phosphodiesterases (cAMP-PDE) in the regulation of basal vascular tone and relaxation of rat aorta mediated by β-adrenoceptors, following heart failure (HF)., Experimental Approach: Twenty-two weeks after proximal aortic stenosis, to induce HF, or SHAM surgery in rats, we evaluated the expression, activity and function of cAMP-PDE in the descending thoracic aorta., Key Results: HF rat aortas exhibited signs of endothelial dysfunction, with alterations of the NO pathway, and alteration of PDE3 and PDE4 subtype expression, without changing total aortic cAMP-hydrolytic activity and PDE1, PDE3 and PDE4 activities. Vascular reactivity experiments using PDE inhibitors showed that PDE3 and PDE4 controlled the level of PGF2α -stimulated contraction in SHAM aorta. PDE3 function was partially inhibited by endothelial NO, whereas PDE4 function required a functional endothelium and was under the negative control of PDE3. In HF, PDE3 function was preserved, but its regulation by endothelial NO was altered. PDE4 function was abolished and restored by PDE3 inhibition. In PGF2α -precontracted arteries, β-adrenoceptor stimulation-induced relaxation in SHAM aorta, which was abolished in the absence of functional endothelium, as well as in HF aortas, but restored after PDE3 inhibition in all unresponsive arteries., Conclusions and Implications: Our study underlines the key role of the endothelium in controlling the contribution of smooth muscle PDE to contractile function. In HF, endothelial dysfunction had a major effect on PDE3 function and PDE3 inhibition restored a functional relaxation to β-adrenoceptor stimulation., (© 2014 The British Pharmacological Society.)

The objective of the present study was to figure out whether human IL-27-producing CD4(+) T cells represent a distinct T cell subset in tuberculosis pleural effusion (TPE). Distribution, phenotypic features of IL-27-producing CD4(+) T cells in TPE were determined. The required transcription factors and signal transductions for IL-27-producing CD4(+) T cell differentiation were explored. The immune regulation of IL-27 on pleural mesothelial cells was observed. We have determined the presence of a subset of human Th cells that infiltrated into tuberculous pleural effusion, which was characterized by the secretion of IL-27, and somehow IFN-γ, but not of IL-4, IL-9, IL-17, or IL-22. These IL-27-producing CD4(+) T cells were effector memory cells and exhibited a transcription profile clearly separated from those of Th2, Th17, Th9, and Th22 cells. The in vitro experiments showed that IL-1β, IL-2 and IL-12, or their various combinations could promote IL-27(+)CD4(+) T cell differentiation from naive CD4(+) T cells by means of phosphorylation of STAT3, STAT4, or/and STAT5. Transcription factors c-Fos and T-bet were required for IL-27(+)CD4(+) T cell differentiation. By activating STAT3 signaling, IL-27 not only restored a clear epithelial phenotype of pleural mesothelial cells, but also further reversed IFN-γ-induced epithelial-mesenchymal transition of pleural mesothelial cells. These data suggested that human IL-27(+)CD4(+) T cells might represent a distinct human T cell subset with unique expression profiles of transcription factors and proinflammatory cytokines, and these IL-27(+)CD4(+) T cells may play important roles in tuberculosis immunity by affecting pleural mesothelial cells.

A highly sensitive fluorescence method of quantitative detection for specific DNA sequence is developed based on molecular beacon (MB) and nucleic acid dye SYBR Green I by synchronous fluorescence analysis. It is demonstrated by an oligonucleotide sequence of wild-type HBV (target DNA) as a model system. In this strategy, the fluorophore of MB is designed to be 6-carboxyfluorescein group (FAM), and the maximum excitation wavelength and maximum emission wavelength are both very close to that of SYBR Green I. In the presence of targets DNA, the MBs hybridize with the targets DNA and form double-strand DNA (dsDNA), the fluorophore FAM is separated from the quencher BHQ-1, thus the fluorophore emit fluorescence. At the same time, SYBR Green I binds to dsDNA, the fluorescence intensity of SYBR Green I is significantly enhanced. When targets DNA are detected by synchronous fluorescence analysis, the fluorescence peaks of FAM and SYBR Green I overlap completely, so the fluorescence signal of system will be significantly enhanced. Thus, highly sensitive fluorescence quantitative detection for DNA can be realized. Under the optimum conditions, the total fluorescence intensity of FAM and SYBR Green I exhibits good linear dependence on concentration of targets DNA in the range from 2×10(-11) to 2.5×10(-9)M. The detection limit of target DNA is estimated to be 9×10(-12)M (3σ). Compared with previously reported methods of detection DNA with MB, the proposed method can significantly enhance the detection sensitivity., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The purpose of this study was to identify functional genetic variants in the promoter of tumor necrosis factor superfamily member 15 (TNFSF15) and evaluate their effects on the risk of developing gastric adenocarcinoma. Forty DNA samples from healthy volunteers were sequenced to identify single nucleotide polymorphisms (SNPs) in the TNFSF15 promoter. Two TNFSF15 SNPs (-358 T > C and -638 A > G) were identified by direct sequencing. Next, genotypes and haplotypes of 470 gastric adenocarcinoma patients and 470 cancer-free controls were analyzed. Odds ratios (ORs) and 95% confidence intervals (CIs) were estimated by logistic regression. Serologic tests for Helicobacter pylori infection were measured by enzyme-linked immuno-sorbent assay (ELISA). Subjects carrying the TNFSF15 -358 CC genotype were at an elevated risk for developing gastric adenocarcinoma, compared with those with the -358 TT genotype (OR 1.42, 95% CI, 1.10 to 2.03). H. pylori infection was a risk factor for developing gastric adenocarcinoma (OR 2.31, 95% CI, 1.76 to 3.04). In the H. pylori infected group, subjects with TNFSF15 -358 CC genotype were at higher risks for gastric adenocarcinoma compared with those carrying -358 TT genotype (OR: 2.01, 95%CI: 1.65 to 4.25), indicating that H. pylori infection further influenced gastric adenocarcinoma susceptibility. The -358 T>C polymorphism eliminates a nuclear factor Y (NF-Y) binding site and the -358 C containing haplotypes showed significantly decreased luciferase expression compared with -358 T containing haplotypes. Collectively these findings indicate that functional genetic variants in TNFSF15 may play a role in increasing susceptibility to gastric adenocarcinoma.

A high performance liquid chromatography method was established for simultaneously determining four bioactive components, salicin, liquiritin, paeonolum, and imperatorin in Fengshiding capsule, a widely used traditional Chinese medicine for treating rheumatic disease. The chromatographic separation was performed on a Shimadzu Shim-pack Stable Bond C18 column using gradient elution with methanol and water. The analytical method was validated through precision, repeatability and stability, and the relative standard deviation values were less than 3%, respectively. The recoveries of the four investigated compounds ranged from 95.80 to 101.21% with relative standard deviation values less than 3.2%. Then this proposed method was successfully applied to determine six batches of Fengshiding commercial products of capsule dosage form from two pharmaceutical factories. This study might provide a basis for quality control for this traditional Chinese medicine preparation.

We conducted a joint (pooled) analysis of three genome-wide association studies (GWAS) of esophageal squamous cell carcinoma (ESCC) in individuals of Chinese ancestry (5,337 ESCC cases and 5,787 controls) with 9,654 ESCC cases and 10,058 controls for follow-up. In a logistic regression model adjusted for age, sex, study and two eigenvectors, two new loci achieved genome-wide significance, marked by rs7447927 at 5q31.2 (per-allele odds ratio (OR) = 0.85, 95% confidence interval (CI) = 0.82-0.88; P = 7.72 × 10(-20)) and rs1642764 at 17p13.1 (per-allele OR = 0.88, 95% CI = 0.85-0.91; P = 3.10 × 10(-13)). rs7447927 is a synonymous SNP in TMEM173, and rs1642764 is an intronic SNP in ATP1B2, near TP53. Furthermore, a locus in the HLA class II region at 6p21.32 (rs35597309) achieved genome-wide significance in the two populations at highest risk for ESSC (OR = 1.33, 95% CI = 1.22-1.46; P = 1.99 × 10(-10)). Our joint analysis identifies new ESCC susceptibility loci overall as well as a new locus unique to the population in the Taihang Mountain region at high risk of ESCC.

Bitter tastants can induce relaxation in precontracted airway smooth muscle by activating big-conductance potassium channels (BKs) or by inactivating voltage-dependent L-type Ca2+ channels (VDLCCs). In this study, a new pathway for bitter tastant-induced relaxation was defined and investigated. We found nifedipine-insensitive and bitter tastant chloroquine-sensitive relaxation in epithelium-denuded mouse tracheal rings (TRs) precontracted with acetylcholine (ACH). In the presence of nifedipine (10 µM), ACH induced cytosolic Ca2+ elevation and cell shortening in single airway smooth muscle cells (ASMCs), and these changes were inhibited by chloroquine. In TRs, ACH triggered a transient contraction under Ca2+-free conditions, and, following a restoration of Ca2+, a strong contraction occurred, which was inhibited by chloroquine. Moreover, the ACH-activated whole-cell and single channel currents of non-selective cation channels (NSCCs) were blocked by chloroquine. Pyrazole 3 (Pyr3), an inhibitor of transient receptor potential C3 (TRPC3) channels, partially inhibited ACH-induced contraction, intracellular Ca2+ elevation, and NSCC currents. These results demonstrate that NSCCs play a role in bitter tastant-induced relaxation in precontracted airway smooth muscle.

Tumor necrosis factor-α (TNF-α) is an immunoregulatory cytokine involved in B- and T-cell function, and also plays an important role in inflammation and cancer. TNF-α-308G>A has been associated with constitutively elevated TNF-α expression. Several studies have reported the association between the TNF-α-308G>A polymorphism and non-Hodgkin lymphomas (NHL) risk, however, results are still inconsistent. To solve these conflicts, we conducted the first meta-analysis to assess the effect of TNF-α-308G>A polymorphism on the risk of NHL and various subtypes (additive model) including 10,619 cases and 12,977 controls in Caucasian and Asian populations. Our meta-analysis indicated that TNF-α-308G>A polymorphism is not associated with NHL risk when pooling all studies together (OR=1.06, 95% CI: 0.92-1.23, p=0.413). In stratified analyses, we found TNF-α-308A allele was significantly associated with higher risk of NHL, B-cell lymphomas (BCL), T-cell lymphomas (TCL) and diffuse large B-cell lymphomas (DLBCL) in Caucasians (OR=1.22, 95% CI: 1.06-1.40, p=0.007; OR=1.18, 95% CI: 1.03-1.34, p=0.014; OR=1.20, 95% CI: 1.01-1.42, p=0.040; OR=1.21, 95% CI: 1.11-1.32, p<0.001, respectively). Interestingly, it was associated with decreased risk of NHL, BCL and DLBCL in Asians (OR=0.75, 95% CI: 0.66-0.86, p<0.001; OR=0.70, 95% CI: 0.52-0.94, p=0.018; OR=0.70, 95% CI: 0.57-0.86, p=0.001). These findings also suggest TNF-α might play a distinct role in pathogenesis of NHL in different populations.

Objective: To analyze the association of micoRNA-related genes DROSHA single nucleotide polymorphisms (SNP) rs10719 and rs6877842, DICER1 rs3742330and GEMIN4 rs3744741 with prognosis of T-cell lymphoma., Methods: Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method was used to determine the genotypes of the above 4SNPs and their associations with complete remission (CR) rate and overall survival (OS) in 163 patients with TCL., Results: Patients carrying the rs6877842 CG genotype had a significantly higher CR rate compared with those carrying the CC genotype (OR=0.07, 95% CI 0.01-0.72, P=0.026); the same for patients carrying the DICER1 rs3742330 GG genotype compared with those carrying the GA genotype (OR=0.15, 95% CI 0.02-0.97, P=0.047) or the AA genotype (OR=0.11, 95% CI 0.02-0.71, P=0.020). In addition, patients with the DICER1 rs3742330 GG genotype had a significantly improved OS compared with those carrying the GA (HR=9.02, 95% CI 1.22-66.92, P=0.031) or AA genotype (HR=8.77, 95% CI 1.19-64.67, P=0.033). The other two SNPs of rs10719 and rs3744741 had no significant association with CR or OS., Conclusion: DROSHA rs6877842 and DICER1 rs3742330 were independent factors for TCL CR, and DICER1 rs3742330 was also an independent prognostic factor for TCL OS.

Activation of the cyclic AMP (cAMP) pathway reduces bladder contractility. However, the role of phosphodiesterase (PDE) families in regulating this function is poorly understood. Here, we compared the contractile function of the cAMP hydrolyzing PDEs in neonatal rat bladder smooth myocytes. RT-PCR and Western blotting analysis revealed that several isoforms of PDE1-4 were expressed in neonatal rat bladder. While 8-methoxymethyl-3-isobutyl-1-methylxanthine (a PDE1 inhibitor) and BAY-60-7550 (a PDE2 inhibitor) had no effect on the carbachol-enhanced phasic contractions of bladder strips, cilostamide (Cil, a PDE3 inhibitor) and Ro-20-1724 (Ro, a PDE4 inhibitor) significantly reduced these contractions. This inhibitory effect of Ro was blunted by the PKA inhibitor H-89, while the inhibitory effect of Cil was strongly attenuated by the PKG inhibitor KT 5823. Application of Ro in single bladder smooth myocytes resulted in an increase in Ca(2+) spark frequency but a decrease both in Ca(2+) transients and in sarcoplasmic reticulum (SR) Ca(2+) content. In contrast, Cil had no effect on these events. Furthermore, Ro-induced inhibition of the phasic contractions was significantly blocked by ryanodine and iberiotoxin. Taken together, PDE3 and PDE4 are the main PDE isoforms in maintaining the phasic contractions of bladder smooth myocytes, with PDE4 being functionally more active than PDE3. However, their roles are mediated through different mechanisms., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Objective: Although the values of soluble mesothelin-related peptides (SMRPs), including mesothelin and megakaryocyte potentiating factor, in serum and/or pleural fluid for diagnosing malignant pleural mesothelioma (MPM) have been extensively studied, the exact diagnostic accuracy of these SMRPs remains controversial. The purpose of the present meta-analysis is to update the overall diagnostic accuracy of SMRPs in serum and, furthermore, to establish diagnostic accuracy of SMRPs in pleural fluid for MPM., Design: Systematic review and meta-analysis., Methods: A total of 30 articles of diagnostic studies were included in the current meta-analysis. Sensitivity, specificity and other measures of accuracy of SMRPs in serum and pleural fluid for the diagnosis of MPM were pooled using random effects models. Summary receiver operating characteristic curves were used to summarise overall test performance., Results: The summary estimates of sensitivity, specificity, positive likelihood ratio, negative likelihood ratio and diagnostic OR were 0.61, 0.87, 5.71, 0.43 and 14.43, respectively, for serum and 0.79, 0.85, 4.78, 0.30 and 19.50, respectively, for pleural fluid. It was also found that megakaryocyte potentiating factor in serum had a superior diagnostic accuracy compared with mesothelin for MPM., Conclusions: SMRPs in both serum and pleural fluid are helpful markers for diagnosing MPM with similar diagnostic accuracy. The negative results of SMRP determinations are not sufficient to exclude non-MPM, and the positive test results indicate that further invasive diagnostic steps might be necessary for the diagnosis of MPM.

Objective: Tumour biomarkers are used as indicators for cancer screening and as predictors for therapeutic responses and prognoses in cancer patients. We aimed to identify genetic loci that influence concentrations of cancer antigen 19-9 (CA19-9), carcinoembryonic antigen (CEA) and α fetoprotein (AFP), and investigated the associations between the significant single nucleotide polymorphisms (SNPs) with risks of oesophageal squamous cell (OSCC), pancreatic and hepatocellular cancers., Design: We carried out a genome wide association study on plasma CA19-9, CEA and AFP concentrations in 3451 healthy Han Chinese and validated the results in 10 326 individuals. Significant SNPs were further investigated in three case control studies (2031 OSCC cases and 2044 controls; 981 pancreatic cancer cases and 1991 controls; and 348 hepatocellular cancer cases and 359 controls)., Results: The analyses showed association peaks on three genetic loci for CA19-9 (FUT6-FUT3 at 19p13.3, FUT2-CA11 at 19q13.3 and B3GNT3 at 19p13.1; p=1.16×10(-13)-3.30×10(-290)); four for CEA (ABO at 9q34.2, FUT6 at 19p13.3, FUT2 at 19q13.3 and FAM3B at 21q22.3; p=3.33×10(-22)-5.81×10(-209)); and two for AFP (AFP at 4q11-q13 and HISPPD2A at 15q15.3; p=3.27×10(-18) and 1.28×10(-14)). These explained 17.14% of the variations in CA19-9, 8.95% in CEA and 0.57% in AFP concentrations. Significant ABO variants were also associated with risk of OSCC and pancreatic cancers, and AFP variants with risk of hepatocellular cancer (p<0.05)., Conclusions: This study identified several loci associated with CA19-9, CEA and AFP concentrations. The ABO variants were associated with risk of OSCC and pancreatic cancers and AFP variants with risk of hepatocellular cancer.