Identification and Characterization of a Luteinizing Hormone Receptor (LHR) Homolog from the Chinese Mitten Crab Eriocheir sinensis.

Luteinizing hormone (LH), a pituitary gonadotropin, coupled with LH receptor (LHR) is essential for the regulation of the gonadal maturation in vertebrates. Although LH homolog has been detected by immunocytochemical analysis, and its possible role in ovarian maturation was revealed in decapod crustacean, so far there is no molecular evidence for the existence of LHR. In this study, we cloned a novel LHR homolog (named EsLHR) from the Chinese mitten crab Eriocheir sinensis. The complete sequence of the EsLHR cDNA was 2775bp, encoding a protein of 924 amino acids, sharing 71% amino acids identity with the ant Zootermopsis nevadensis LHR. EsLHR expression was found to be high in the ovary, while low in testis, gill, brain, and heart, and no expression in the thoracic ganglion, eye stalk, muscle, and hepatopancreas. Quantitative PCR revealed that the expression level of EsLHR mRNA was significantly higher in the ovaries in previtellogenic (Pvt), late vitellogenic (Lvt), and germinal vesicle breakdown (GVBD) stages than that in the vitellogenic (Mvt) and early vitellogenic (Evt) stages (P < 0.05), and, the highest and the lowest expression were in Lvt, and Evt, respectively. The strong signal was mainly localized in the ooplasm of Pvt oocyte as detected by in situ hybridization. The crab GnRH homolog can significantly induce the expression of EsLHR mRNA at 36 hours post injection in vivo (P < 0.01), suggesting that EsLHR may be involved in regulating ovarian development through GnRH signaling pathway in the mitten crab.

Synthesis of azo-linked covalent organic polymers for pipette tip solid-phase extraction of sedative residues from animal tissues samples.

Azo-linked covalent organic polymers (ACOPs) were synthesized by a simple azo reaction, with 2,2'-bis(trifluoromethyl)benzidine and 1,3,5-trihydroxybenzene as the monomers. The preparation process was mild, green, and environmental-friendly, avoiding the use of high temperature, metal catalysis, and harmful organic reagent. The obtained ACOPs were characterized by scanning electron microscopy, Fourier transform infrared spectroscopy, thermogravimetric analysis, powder X-ray diffraction, and Brunauer-Emmett-Teller. With the prepared ACOPs as adsorbent, a method of pipette tip solid-phase extraction-liquid chromatography-tandem mass spectrometry detection (PTSPE-LC-MS/MS) was proposed for the analysis of target sedatives in animal tissues. Furthermore, the parameters for the extraction of five sedatives, including the amount of adsorbent, pH value, ion strength, elution solvent and volume, were investigated. Under the optimized conditions, the linear dynamic range was found from 0.1 to 10.0 µg kg-1, and the limits of detection were ranged from 0.02 to 0.1 µg kg-1. The method was assessed by the analysis of target sedatives in animal tissues, and the recoveries for the spiked pork muscle and pork liver samples were 84-102% and 83-101%, respectively. The results show that the developed method of PTSPE-LC-MS/MS with ACOPs as adsorbent is efficient for the analysis of trace sedatives in animal tissues.

Detection of azithromycin residue in broiler feathers by liquid chromatography-tandem mass spectrometry.

In the study, a sensitive and reproducible method for the quantitative analysis of azithromycin in broiler feather samples by liquid chromatography-tandem mass spectrometry (LC-MS/MS) was developed. Feather samples were rinsed after being wrapped in medical gauze, then chopped and then added to 5% (v/v) ammonia in methanol solution for ultrasonic extraction. The extract was purified by the combination of commercial polymeric microparticles (Oasis MCX) and Fe3O4 nanoparticles. The LC separation was performed on an Agilent Eclipse plus C18 column. Multiple reaction monitoring was used for the selective detection of azithromycin. The good linearity curve of azithromycin in feather sample was in the range from 1.0 µg kg-1 to 100.0 µg kg-1 with 0.9935 of correlation coefficient. And the limit detection and limit of quantification was 0.5 µg kg-1 and 2.0 µg kg-1 in spiked feather samples. The recoveries of azithromycin were 85.2-94.7% with the relative standard deviation less than 10%. The established method is simple, rapid, sensitive and specific, and could meet the need of government and enterprises to monitor the illegal use of azithromycin in livestock and poultry breeding.

Densification and Phase Transformation in Multi-Layered Graded Si3N4-TiN Components Produced by Field-Assisted Sintering.

The structural and/or functional design of multiphase ceramics, along with their processing, are timely research topics in the area of field-assisted sintering techniques, such as spark plasma sintering, especially for systems containing both electrically insulating and conductive phases. In the present study, spark plasma sintering of Si3N4-TiN composites was investigated by changing the TiN particle size and electrical current waveform. Their combined effects on both the densification behavior and α-to-ß phase conversion of the Si3N4 matrix was studied and compared by means of a thermodynamic approach and dilatometric measurements. Through the control of TiN phase characteristics and heating mode, double-layered Si3N4-based components were also prepared using a one-step spark plasma sintering process, which was compared with conventional hot-pressing. It was shown that the size of the conductive TiN phase has a significant influence on the particle rearrangement, with the formation of a liquid phase, and the solution-diffusion-precipitation process, through the field-induced local heating and electrowetting mechanisms. Moreover, the contribution of current pulsing to the densification and α-to-ß conversion of the layered Si3N4-based components was mostly dependent upon the particle size distribution and content of the TiN phase, indicating that the electric-field effect is dependent upon current path.

[Seasonal and Spatial Variations of Microcystins and Their Relationships with Physiochemical and Biological Factors in Poyang Lake].

Poyang Lake is the largest freshwater lake in China and an important drinking water source for Jiangxi Province. Since the year 2000, toxic cyanobacteria have been observed frequently in Poyang Lake. In this study, water samples were collected in the lake quarterly (April 2012, August 2012, October 2012, and January 2013) to examine the spatial and seasonal variations in the concentrations of microcystins (MCs; MC-RR, -YR, and -LR) and their relationships with physiochemical and biological factors. MCs were determined by ultra-high-performance liquid chromatography-electrospray ionization tandem triple quadrupole/mass spectrometry (UPLC-MS/MS). MC-RR (accounting for 75.01% and 71.34% of intracellular MC (IMC) and extracellular MC (EMC) concentrations, respectively) was the most dominant variant in Poyang Lake, followed by MC-LR (accounting for 21.95% and 24.97% of IMC and EMC concentrations, respectively), while MC-YR was detected in low concentrations (accounting for 3.01% and 3.69% of IMC and EMC concentrations, respectively). Total MC concentrations (IMC + EMC, TMC) ranged from 0.49 to 3517.85 ng·L-1, with an average of 337.43 ng·L-1 and only 2.53% (2 out of 79 water samples) of the water samples contained MCs concentrations exceeding the drinking water guideline level of 1 ng·L-1 for MC-LR proposed by World Health Organization (WHO). IMC concentrations showed significant relationships with Microcystis biomass (r=0.47, P<0.01), Oscillatoria biomass (r=0.68, P<0.01), Cyanobacteria biomass (r=0.56, P<0.01), and Chl-a (r=0.28, P<0.01), but no significant correlation was found between intracellular MC concentration and Anabena biomass (P>0.05), suggesting that Microcystis and Oscillatoria might be the main MCs-producing cyanobacteria in Poyang Lake. In addition, IMC concentrations were positively correlated with water temperature (r=0.51, P<0.01), transparence (r=0.69, P<0.01), Fe (r=0.43, P<0.01), and Zn contents (r=0.43, P<0.01), and negatively correlated with TN (r=-0.44, P<0.01), TP (r=-0.29, P<0.01), NH4+-N (r=-0.33, P<0.05), NO2--N (r=-0.28, P<0.05), Ca (r=-0.34, P<0.01), and Mg(r=-0.35, P<0.05), while no significant correlations were observed between IMC concentrations and pH, PO43--P, NO3--N, electrical conductivity, permanganate index, and Cu content (P>0.05). These results indicated that light intensity (represented by transparence), nitrogen, phosphorus, and water temperature might be the regulating factors of MCs production in Poyang Lake and trace elements (Fe, Zn, Ca, and Mg) can influence the MC production to a certain extent. IMCs and EMCs exhibited similar seasonal variations in Poyang Lake. The highest values of IMCs (531.87 ng·L-1) and EMCs (232.44 ng·L-1) were observed in summer. The concentrations of IMCs and EMCs in autumn were 31.97 ng·L-1 and 6.49 ng·L-1, respectively. Low concentrations were observed in spring (0.55 ng·L-1 and 0.88 ng·L-1 of IMCs and EMCs, respectively) and winter (0.69 ng·L-1 and 4.14 ng·L-1 of IMCs and EMCs, respectively). The highest IMCs and EMCs values of Poyang Lake in summer were 2298.08 ng·L-1 and 1219.77 ng·L-1, respectively, and the lowest values were 92.53 ng·L-1 and 38.80 ng·L-1, respectively. Overall, the concentrations of IMCs in eastern bays, the vicinity of Songmen Mountain, Banghu Lake, and its outlet were higher than those in other regions. However, the spatial distributions of EMCs in Poyang Lake were different from those of IMCs. EMCs concentrations in the vicinity of Songmen Mountain, Banghu Lake, and its outlet were higher than those in other regions.

Two surgical procedures for esophagogastric variceal bleeding in patients with portal hypertension.

AIM: To determine the clinical value of a splenorenal shunt plus pericardial devascularization (PCVD) in portal hypertension (PHT) patients with variceal bleeding. METHODS: From January 2008 to November 2012, 290 patients with cirrhotic portal hypertension were treated surgically in our department for the prevention of gastroesophageal variceal bleeding: 207 patients received a routine PCVD procedure (PCVD group), and 83 patients received a PCVD plus a splenorenal shunt procedure (combined group). Changes in hemodynamic parameters, rebleeding, encephalopathy, portal vein thrombosis, and mortality were analyzed. RESULTS: The free portal pressure decreased to 21.43 ± 4.35 mmHg in the combined group compared with 24.61 ± 5.42 mmHg in the PCVD group (P < 0.05). The changes in hemodynamic parameters were more significant in the combined group (P < 0.05). The long-term rebleeding rate was 7.22% in the combined group, which was lower than that in the PCVD group (14.93%), (P < 0.05). CONCLUSION: Devascularization plus splenorenal shunt is an effective and safe strategy to control esophagogastric variceal bleeding in PHT. It should be recommended as a first-line treatment for preventing bleeding in PHT patients when surgical interventions are considered.

Increased immunohistochemical expression of YKL-40 in the spleen of patients with portal hypertension.

YKL-40 has been identified as a growth factor in connective tissue cells and also a migration factor in vascular smooth muscle cells. To a large extent, the increase of serum YKL-40 is attributed to liver fibrosis and asthma. However, the relationship of the expression and clinical/prognostic significance of YKL-40 to the splenomegaly of patients with portal hypertension is unclear. In the present study, the expression of YKL-40 was studied by immunohistochemistry in 48 splenomegaly tissue samples from patients with portal hypertension and in 14 normal spleen specimens. All specimens were quickly stored at -80°C after resection. Primary antibodies YKL-40 (1:150 dilution, rabbit polyclonal IgG) and MMP-9 (1:200 dilution, rabbit monoclonal IgG) and antirabbit immunoglobulins (HRP K4010) were used in this study. The relationship of clinicopathologic features with YKL-40 is presented. The expression of YKL-40 indicated by increased immunochemical reactivity was significantly up-regulated in splenomegaly tissues compared to normal spleen tissues. Overexpression of YKL-40 was found in 68.8% of splenomegaly tissues and was significantly associated with Child-Pugh classification (P = 0.000), free portal pressure (correlation coefficient = 0.499, P < 0.01) and spleen fibrosis (correlation coefficient = 0.857, P < 0.01). Further study showed a significant correlation between YKL-40 and MMP-9 (correlation coefficient = -0.839, P < 0.01), indicating that YKL-40 might be an accelerator of spleen tissue remodeling by inhibiting the expression of MMP-9. In conclusion, YKL-40 is an important factor involved in the remodeling of spleen tissue of portal hypertension patients and can be used as a therapeutic target for splenomegaly.

Polymorphisms of tumor-related genes IL-10, PSCA, MTRR and NOC3L are associated with the risk of gastric cancer in the Chinese Han population.

BACKGROUND: Gastric cancer is the fourth most common cancer in the world. Environmental and genetic factors both play critical roles in the etiology of gastric cancer. Hundreds of SNPs have been identified to have association with the risk of gastric cancer in many races. In this study, 25 SNPs in genes for IL-10, IL-1B, MTRR, TNF-а, PSCA, PLCE1 and NOC3L were analyzed to further evaluate their associations with gastric cancer susceptibility in the Chinese Han population. METHODS: Two hundred and seventy nine gastric cancer patients and 296 healthy controls were recruited in this study. SNP genotyping was conducted using Sequenom MassARRAY RS1000. Data management and statistical analyses were conducted by Sequenom Typer 4.0 Software and Pearson's χ(2) test. RESULTS: One protective allele and three risk alleles for gastric cancer patients were found in this study. The allele "G" of rs1801394 in MTRR showed an association with a decreased risk of gastric cancer: odds ratio (OR) = 0.74, 95% confidence interval (95% CI) = 0.57-0.97, P = 0.030 in the additive model; OR = 0.495, 95% CI = 0.26-0.95, P = 0.034 in the recessive model. The other three SNPs, the allele "C" of rs1800871 in IL10 (OR = 1.33, 95% CI = 1.04-1.90; P = 0.026 in the additive model; OR = 1.46, 95% CI = 1.04-2.06; P = 0.030 in the recessive model), the allele "A" of rs2976391 in PSCA (OR = 1.30, 95% CI = 1.01-1.66; P = 0.041 in the additive model and OR = 1.48, 95% CI = 1.04-2.11, P = 0.028 in the recessive model), and the allele "G" of rs17109928 in NOC3L gene (OR = 1.34, 95% CI = 1.01-1.78; P = 0.042 by additive model analysis; OR = 1.47, 95% CI = 1.04-2.07, P = 0.028 by dominant model analysis), showed an association with an increased risk of gastric cancer. CONCLUSIONS: These results indicate the importance of four gastric cancer susceptibility polymorphisms of IL-10, NOC3L, PSCA and MTRR in the Chinese Han population, which could be used in the determination of gastric cancer risk in clinical practice.

[Use of dinoflagellates as a metal toxicity assessment tool in aquatic system].

Although dinoflagellates have been used to assess biological toxicity of contaminants, this method still lacks of corresponding toxicity assessment standard. This study appraised the toxicity of selected heavy metals to dinoflagellates based on the dinoflagellates bioluminescence with QwikLite developed by the United States Navy. The results show that single heavy metal biological toxicity is in the order: Hg2+ > Cu2+ > Cd2+ > As5+ > Pb2+ > Cr6+; Two, three and four heavy metal mixture experiments show synergism primarily, antagonism is in minority. pH has not remarkable effect on dinoflagellates, they can be applied directly in natural water, but pH influence Hg2+ and Cu2+ toxicity greatly, eliminating the influence of pH is essential when doing these two kind of ions measurements. The nutrients has little influence on dinoflagellates, change in COD has obvious effect on the response relationships between dinoflagellates and Hg2+ or CU2+. Metal toxicity assessment using dinoflagellates shows great sensitivity, narrow response scope and high stability. Dinoflagellates are good species for heavy metal biological toxicity test in aquatic system.

Increased immunohistochemical expression of YKL-40 in the spleen of patients with portal hypertension

YKL-40 has been identified as a growth factor in connective tissue cells and also a migration factor in vascular smooth muscle cells. To a large extent, the increase of serum YKL-40 is attributed to liver fibrosis and asthma. However, the relationship of the expression and clinical/prognostic significance of YKL-40 to the splenomegaly of patients with portal hypertension is unclear. In the present study, the expression of YKL-40 was studied by immunohistochemistry in 48 splenomegaly tissue samples from patients with portal hypertension and in 14 normal spleen specimens. All specimens were quickly stored at -80°C after resection. Primary antibodies YKL-40 (1:150 dilution, rabbit polyclonal IgG) and MMP-9 (1:200 dilution, rabbit monoclonal IgG) and antirabbit immunoglobulins (HRP K4010) were used in this study. The relationship of clinicopathologic features with YKL-40 is presented. The expression of YKL-40 indicated by increased immunochemical reactivity was significantly up-regulated in splenomegaly tissues compared to normal spleen tissues. Overexpression of YKL-40 was found in 68.8 percent of splenomegaly tissues and was significantly associated with Child-Pugh classification (P = 0.000), free portal pressure (correlation coefficient = 0.499, P < 0.01) and spleen fibrosis (correlation coefficient = 0.857, P < 0.01). Further study showed a significant correlation between YKL-40 and MMP-9 (correlation coefficient = -0.839, P < 0.01), indicating that YKL-40 might be an accelerator of spleen tissue remodeling by inhibiting the expression of MMP-9. In conclusion, YKL-40 is an important factor involved in the remodeling of spleen tissue of portal hypertension patients and can be used as a therapeutic target for splenomegaly.