Bm04862 is a novel hemocyte-specific gene, which has been identified and cloned through the microarray data in silkworm, Bombyx mori. Initially, we successfully obtained the full-length cDNA sequence of Bm04862 via the rapid amplification of cDNA ends (RACE). The sequence consists of an open reading frame (ORF) with 819 bp, which could encode 273 amino acid residues. The bioinformatics analysis predicted that Bm04862 was a transmembrane protein. Furthermore, qRT-PCR analysis revealed that the expression of Bm04862 was specifically detected in hemocyte and reached a peak at L4M and PP2 stage. Bm04862 was overexpressed in Sf9 insect cells, and the cellular localization indicated that Bm04862 was specifically located in cytoplasm and nuclei membrane. Interestingly, the expression of Bm04862 increas'd dramatically after challenged with Escherichia coli for 24 hours, which predicted its potential role in the innate immune system. Overall, this study could provide the fundamental knowledge for further research.
Animals ; Bombyx ; genetics ; Cloning, Molecular ; Computational Biology ; DNA, Complementary ; Hemocytes ; metabolism ; Insect Proteins ; genetics ; metabolism ; Membrane Proteins ; genetics ; metabolism ; Open Reading Frames

Objective To investigate the effects of down-regulation of polo-like kinase-1 (PLK1) gene by RNA interference (RNAi) on the invasion of a human malignant melanoma cell line A375 and their possible mechanisms.Methods Cultured A375 cells were classified into several groups:blank control group receiving no treatment,liposome group transfected with lipofectamine only,and three siRNA groups transfected with three concentrations of a small interference RNA (siRNA) targeting PLK1 respectively.After additional culture,real time quantitative PCR and Western blot analysis were performed to quantify the expressions of PLK1 mRNA and protein in A375 cells respectively,Transwell invasion assay to evaluate the invasive capacity of A375 cells,agarose gel electrophoresis and terminal deoxynucleotidyl-transferase-mediated dUTP-biotin nick end labelling (TUNEL) to detect anoikis in A375 cells.The colony-forming capacity was also evaluated for A375 cells.Statistical analysis was carried out by one-factor analysis of variance.Results There was a significant decrease in PLK1 mRNA and protein expressions as well as in colony-forming units in the siRNA groups compared with the blank control group (all P ＜ 0.05).The invasive capacity of A375 cells was significantly inhibited in the siRNA groups with the number of migrating cells in Transwell assay being 39 ± 5,19 ± 5 and 9 ± 3 in A375 cells transfected with 3.125,6.250 and 12.500 nmol/L siRNAs respectively,compared to 56 ± 5 in the blank control group (all P ＜ 0.05).A characteristic DNA ladder was observed on agarose gel electrophoresis in the siRNA (6.250 nmol/L) group.Compared with the blank control group and liposome group,the three siRNA groups showed increased apoptotic index (3.86％ ± 0.35％ (3.125 nmol/L siRNA),7.35％ ± 0.36％ (6.250 nmol/L siRNA) and 17.56％ ± 0.38％ (12.500 nmol/L siRNA) vs.1.15％ ± 0.25％ (blank control group) and 1.18％ ± 0.22％ (liposome group),all P ＜ 0.05).Conclusions PLK1 siRNA can inhibit the invasion of malignant melanoma cells,likely by inducing anoikis in these cells.

Objective To study the effects of Forkhead box protein M1 (FoxM1) down regulation by small interfering RNA(siRNA) on chemosensitivity and mechanism of human pancreatic cancer cell and its mechanism.Methods Three FoxM1 siRNAs were designed and constructed.All cancer cells were divided into different groups,after transfected with FoxM1 siRNA for different time,the cultured cells were harvested to carry on the next tests.Expression of FoxM1 were determined by red-time PCR and Western blot,and prolifearion and chemosensitivity were evaluated by MTT assay,and the phosphorylation of Akt protein was examined by Western blot.Results FoxM1 siRNA could down-regulate the FoxM1 expression in a dose-and time-dependent manner.The MTF results showed that the inhibit rates was 17.78％,17.56％,35.39％,52.81％,70.98％ indifferentgroups [Con-A + Gemcitabine,Con-B + Gemcitabine,siRNA (3.125nM) + Gemcitabine,siRNA (6.25nM) + Gemcitabine and siRNA(12.5nM) + Gemcitabine,respectively.The phosphorylation of Akt protein was inhibited in a dose-dependent manner.Conclusions FoxM1 siRNA could sensitize human pancreaticr cancer cells chemotherapy sensitivity,it is the one of the important mechanisms through down-regulate Akt phosphorylated levels,but the molecular mechanism need to be explored further.

OBJECTIVETo examine the effect of acetylcholine (ACh) on the electric activities of pain-excitation neurons (PEN) and pain-inhibitation neurons (PIN) in the hippocampal CA1 area of normal rats or morphinistic rats, and to explore the role of ACh in regulation of pain perception in CA1 area under normal condition and morphine addiction.

METHODSThe trains of electric impulses applied to sciatic nerve were set as noxious stimulation. The discharges of PEN and PIN in the CA1 area were recorded extracellularly by glass microelectrode. We observed the influence of intracerebroventricular (i.c.v.) injection of ACh and atropine on the noxious stimulation-evoked activities of PEN and PIN in the CA1 area.

RESULTSNoxious stimulation enhanced the electric activity of PEN and depressed that of PIN in the CA1 area of both normal and addiction rats. In normal rats, ACh decrease the pain-evoked discharge frequency of PEN, while increased the frequency of PIN. These effects reached the peak value at 4 min after injection of ACh. In morphinistic rats, ACh also inhibited the PEN electric activity and potentialized the PIN electric activity, but the maximum effect appeared at 6 min after administration. The ACh-induced responses were significantly blocked by muscarinic receptor antagonist atropine.

CONCLUSIONCholinergic neurons and muscarinic receptors in the hippocampal CA1 area are involved in the processing of nociceptive information and they may play an analgesia role in pain modulation. Morphine addiction attenuated the sensitivity of pain-related neurons to the noxious information.

Acetylcholine ; administration & dosage ; metabolism ; Adaptation, Physiological ; drug effects ; physiology ; Animals ; Electric Stimulation ; Evoked Potentials ; physiology ; Female ; Hippocampus ; cytology ; metabolism ; Injections, Intraventricular ; Male ; Morphine ; pharmacology ; Morphine Dependence ; metabolism ; Narcotics ; pharmacology ; Neuronal Plasticity ; physiology ; Neurons ; drug effects ; physiology ; Pain ; metabolism ; Pain Threshold ; physiology ; Rats ; Rats, Wistar ; Receptors, Cholinergic ; drug effects ; metabolism ; Sciatic Nerve ; physiopathology ; Signal Transduction ; physiology

Objective To investigate the effects of sevoflurane pretreatment on renal ischemia-reperfusion (I/R)-induced apoptosis in kidney in rats. Methods Thirty pathogen-free male SD rats weighing 220-260 g were randomized into 3 groups (n=10 each):group control (group C);group I/R and group sevoflurane(group S). Renal I/R was induced by clamping the left renal pedicle for 45 min in I/R and S groups. In group S inhalation of 2.2% sevoflurane in O2 was started at 30 min before operation and maintained throughout the experiment.Venous blood samples were taken at 3 h of reperfusion for determination of serum BUN and Cr concentrations. The animals were then sacrificed and the left kidneys were removed for microscopic examination, detection of apoptosis(by TUNEL)and determination of heme oxygenase-1(HO-1) mRNA and protein expression (by RT-PCR and Western blot).Results Renal I/R significantly increased serum BUN and Cr concentrations, apoptotic index(percentage of apoptotic cells) and the severity of necrosis of renal proximal convoluted tubules (0=normal,4=necrosis of whole segment of proximal convoluted tubules).Sevoflurane inhalation attenuated the I/R-induced changes mentioned above.HO-1 mRNA and protein expression was up-regulated by I/R and HO-1 mRNA expression was further up-regulated by sevoflurane inhalation.Conclusion Sevoflurane pretreatment can protect kidney against I/R injury by attenuating cell apoptosis.Up-regulation of HO-1 mRNA expression may be involved in the mechanism.

Miscanthus sinensis Anderss is a perennial C4-grass. It is a promising bioenergy plant, which has been proposed as general feedstock for biomass and lignocellulosic biofuel production. In this study, the flower and leaf buds transcriptomes of Miscanthus sinensis Anderss were sequenced by the platform of Illumina HiSeq 2000. In total 98 326 Unigenes were generated by de novo assembly with an average length of 822 bp and N50 of 1 023 bp. Based on the NR, NT, Swiss-Prot, KEGG, GO and COG databases (Evalue < le-5), 74 134 (75.40%) Unigenes were annotated. A total of 45 507 Unigenes were mapped into different GO terms. In KEGG pathways identification, 36 710 sequences were assigned to 128 KEGG pathways. Sorghum bicolor (37 731, 60.86%), Zea mays (16 258, 26.22%), and Oryza sativa (3 065, 4.94%) showed high similarity to Miscanthus sinensis Anderss. And 24 photosynthesis-related enzyme genes were identified. The result provides a foundation for further characterizing the functional genes in Miscanthus sinensis Anderss.
Biofuels ; Gene Expression Profiling ; Genes, Plant ; Poaceae ; genetics ; metabolism ; RNA, Plant ; genetics ; Sequence Analysis, RNA ; Transcriptome

Objective To determine the prognostic value of standardized uptake value(SUV)of fluorodeoxyglucose(FDG)by positron emission tomography and computed tomography(PET-CT)in nonsmall cell lung cancer(NSCLC).Methods Forty-eight patients(39 male,9 female)with stage ⅢNSCLC were reviewed.All patients had at least two repeated FDG PET-CT scans either before and after therapy and the maximum standardized uptake value(SUVmax)of the primary lung lesion was calculated. Resuits Of the 45 eligible patients,after a median follow-up of 22.5 months(rang,13 to 35 months),24 patients had local and regional recurrenee or metastasis and 21 remain disease-free.The mean SUVmax of patients who had local recurrence or metastasis before and after treatment was 12.30±3.17 and 5.35±2.29,respectively.The mean SUVmax of patients who had no loeal recurrence or metastasis before and after treatment was 8.46±3.00 and 2.82±0.63,respectively.Significant differences(tbefore=4.15,Pbefore＜0.01;Pafter=4.88,Pafter＜0.01)in SUVmax were observed either before and after treatment.However,the percent change of SUVmax between pretreatment and post-treatment were not significiantly different(t=1.99,P＞0.05).Using the SUVbefore of 9.0 yielded 92% sensitivity,62% specificity,73% positive predictive value and 87%negaffve Dredictive value in predicting regional recurrence or metastasis. While using the SUVafter of 4.3 yielded 71% sensitivity,100% specificity,100% positive predictive valne and 72% negative Dredictive value. Conclusions PET-CT may have the potientials to predict response to therapy and the SUVmax is a significant predictor for recurrent or metastasis in patients of NSCLC.

Objective To optimize a W/O microemulsion formulation of troxerutin and evaluate its physical properties such as morphology, droplet size, stability and content of troxerutin. Methods The W/O microemulsion was optimized using a pseudoternary phase diagram and the area of the microemulsion region was used to screen and determine microemulsion components.HPLC assay was used for determination of the loading content. Results The optimal formulation contained lecithin, ethanol, isopropyl myristate and water (23.30:11.67:52.45:12.59).The microemulsion was physicochemically stable with round shape and uniform size, and the mean droplet size was about 50. 20 nm. Conclusion Microemulsion was developed successfully.It will expect to be the new preparation for troxerutin.

Objective To investigate the effects of miR-10a expression on migration and invasion of human pancreatic cancer cells AsPC-1.Methods Small interfering RNA targeting at miR-10a (miR-10a-siRNA) was constructed,then it was transfected into pancreatic cancer AsPC-1 cells,and nonsense siRNA (Nc-siRNA) group and blank control group was established.Real time PCR assay was used to detect the expression of miR-10a in the 3 groups,and wound healing assay and Transwell assay were used to determine the migration and invasion abilities of cancer cells.The amount of matrix metalloproteinase-13 (MMP-13) in supernatant of cancer cell culture of each group was examined by ELISA assay.Results The miR-10a levels in control group,NC-siRNA group and miR-10a-siRNA group were 1.05 ±0.08,1.03 ±0.06,0.02 ±0.01 ; and the number of transmembrane cell were (150 ± 2.6),(145 ± 2.2),(62 ± 1.8),the levels of MMP-13 in the supernatant were (108.5 ± 2.8),(107.8 ± 2.5),(35.8 ± 1.5) pg/ml.The values were significantly lower in miR-10a-siRNA group than those in control group and NC-siRNA group (P ＜ 0.01).The distance of cultured clone in miR-10a treated cancer cells (736± 18 μm) was significantly longer than those in the controls (385 ±5 μm) and NC-siRNA group (395± 13 μm,P＜0.01).Conclusions Down-regulation of miR-10a by siRNA may inhibit migration and invasion of pancreatic cancer AsPC-1 cells,and the downregulated expression of MMP-13 may be one of the important mechanisms.

Objective To determine PD-L2 expression in human cervical squamous cell carcinomas and analyze its association with the clinical and pathologic characteristics of these cases ;observe the role of recombinant PD-L2 protein on apoptosis of active peripheral blood T lymphocytes of cervical carcinoma patients .Methods PD-L2 expression in cervical squamous cell carcinomas was determined by immunohistochemistry staining ,and the association between PD-L2 expression with the clinical and pathologic characteristics of these cases was analyzed .In vitro ,the peripheral blood T lymphocytes of cervical carcinoma patients were divided into blank group ,PD-L2 group and PD-L2+anti-PD-1 group respectively .After these active T cells were cultured 72 h ,their apop-totic rates were detected by flow cytometry .Results PD-L2 expressed in 53 .3% (32/60)cervical squamous cell carcinomas ,and it′s expression associated with lymph node metastasis of these cases (P< 0 .05) .In vitro ,PD-L2 promoted apoptosis of CD4+ T and CD8+ T lymphocytes ,the apoptotic rates were 17 .0% and 22 .4% respectively ,which were higher than 9 .0% and 16 .2% of blank group;however ,the apoptotic rates dropped to 11 .1% and 17 .5% in PD-L2+anti-PD-1 group .Conclusion PD-L2 aberrantly ex-presses in cervical squamous cell carcinomas and is associated with their lymph node metastasis .PD-L2 promotes apoptosis of T lymphocytes and depresses the anti-tumor immunity of cervical microenvironment ,and promotes lymph node metastasis of these cancers .PD-L2/PD-1 pathway may be a potential immunotherapy target of cervical squamous cell carcinomas .