SCDb: an integrated database of stomach cancer.

BACKGROUND: Stomach cancer (SC) is a type of cancer, which is derived from the stomach mucous membrane. As there are non-specific symptoms or no noticeable symptoms observed at the early stage, newly diagnosed SC cases usually reach an advanced stage and are thus difficult to cure. Therefore, in this study, we aimed to develop an integrated database of SC. METHODS: SC-related genes were identified through literature mining and by analyzing the publicly available microarray datasets. Using the RNA-seq, miRNA-seq and clinical data downloaded from The Cancer Genome Atlas (TCGA), the Kaplan-Meier (KM) survival curves for all the SC-related genes were generated and analyzed. The miRNAs (miRanda, miRTarget2, PicTar, PITA and TargetScan databases), SC-related miRNAs (HMDD and miR2Disease databases), single nucleotide polymorphisms (SNPs, dbSNP database), and SC-related SNPs (ClinVar database) were also retrieved from the indicated databases. Moreover, gene_disease (OMIM and GAD databases), copy number variation (CNV, DGV database), methylation (PubMeth database), drug (WebGestalt database), and transcription factor (TF, TRANSFAC database) analyses were performed for the differentially expressed genes (DEGs). RESULTS: In total, 9990 SC-related genes (including 8347 up-regulated genes and 1643 down-regulated genes) were identified, among which, 65 genes were further confirmed as SC-related genes by performing enrichment analysis. Besides this, 457 miRNAs, 20 SC-related miRNAs, 1570 SNPs, 108 SC-related SNPs, 419 TFs, 44,605 CNVs, 3404 drug-associated genes, 63 genes with methylation, and KM survival curves of 20,264 genes were obtained. By integrating these datasets, an integrated database of stomach cancer, designated as SCDb, (available at http://www.stomachcancerdb.org/) was established. CONCLUSIONS: As a comprehensive resource for human SC, SCDb database will be very useful for performing SC-related research in future, and will thus promote the understanding of the pathogenesis of SC.

Cloning, expression and cellular localization of the Doublesex gene in the water flea, Daphnia carinata, during different developmental stages.

In this study, one of Doublesex genes from the common freshwater cladoceran Daphnia carinata, designated DapcaDsx1, was cloned using primers based on homologous sequences and rapid amplification of cDNA ends (RACE). qPCR was employed to quantify differences in DapcaDsx1 expression between the different sexual phases, with expression levels being higher in sexual females. The role of DapcaDsx1 in the reproductive transformation was further investigated in parthenogenetic-phase females and sexual-phase females using whole-mount in situ hybridization. This cellular localization study showed specific expression of DapcaDsx1 in the thoracic segments, second antenna and part of the ventral carapace. Higher expression levels were exhibited in sexual females compared to parthenogenetic females. This suggests that the DapcaDsx1 gene plays significant roles in switching modes of reproduction and during sexual differentiation.

Cloning and expression profiling of a cuticular protein gene in Daphnia carinata.

The cladoceran Daphnia carinata undergoes an unusual transition from asexual to sexual reproduction in response to environmental stimuli. Previously, a D. carinata cuticular protein (CP) was identified in an EST library. In this study, the full-length CP cDNA was cloned and sequenced (GenBank accession number: KF551931), and the expression levels in different reproductive states were assessed. Parthenogenetic and sexual female D. carinata were isolated, and CP expression was investigated using semiquantitative reverse transcription polymerase chain reaction (RT-PCR). CP was expressed during both reproductive stages, but expression was higher in sexual females. Cellular localization was also investigated using digoxin-labeled RNA probes in RNA whole-mount in situ hybridization assays, and CP was mainly expressed in the first pair of thoracic appendages, the surface of the head, shell spines, and other parts of the epidermis in parthenogenetic organisms. In contrast, CP expression was restricted to the thoracic appendages in sexual females.

Cloning, expression and cellular localization of Daphnia pulex senescence-associated protein, DpSAP.

Daphnia (water fleas) are small crustaceans that undergo an unusual switch from asexual to sexual reproduction that is dependent on environmental conditions. In this study, a senescence-associated protein (SAP) from the common freshwater species Daphnia pulex was cloned using primers based on homologous sequences and rapid amplification of cDNA ends (RACE). Real-time PCR was employed to quantify the expression of D. pulex SAP (DpSAP) in individual organisms. The role of DpSAP in the reproductive transformation was further investigated in both parthenogenetic and sexual females by using digoxin-labeled SAP RNA probes and RNA whole-mount in situ hybridization. DpSAP was more highly expressed in sexual females, indicating a role in growth and reproduction. Cellular localization studies using RNA whole-mount in situ hybridization showed specific expression in the second tentacle joints. These expression patterns suggest an important role for DpSAP in the reproductive transformation of D. pulex.

Characterization of Cdc2 kinase in the red claw crayfish (Cherax quadricarinatus): evidence for its role in regulating oogenesis.

Cdc2 kinase is a catalytic subunit of the maturation-promoting factor (MPF), a central factor for inducing the meiotic maturation of oocytes. MPF has been studied in a wide variety of animal species; however, its expression in crustaceans is poorly characterized. In this study, a complete cDNA sequence of Cdc2 kinase was cloned from the red claw crayfish, Cherax quadricarinatus, and its spatiotemporal expression profiles were analyzed. The Cdc2 cDNA (1,769 bp) encodes for a 299 amino acid protein with a calculated molecular weight of 34.7 kDa. Quantitative real-time PCR demonstrated that Cdc2 mRNA was expressed mainly in the ovary tissue and the expression decreased as the ovaries developed. Immunohistochemistry analysis revealed that the Cdc2 protein relocated from the cytoplasm to the nucleus during oogenesis. These findings suggest that Cdc2 kinase may play an important role in the gametogenesis and gonad development in C. quadricarinatus.

Cancer stem cell labeling using poly(L-lysine)-modified iron oxide nanoparticles.

Cell labeling using magnetic nanoparticles is an increasingly used approach in noninvasive behavior tracking, in vitro separation of cancer stem cells (CSCs), and CSC-based research in cancer therapy. However, the impact of magnetic labeling on the biological properties of targeted CSCs, such as self-renewal, proliferation, multi-differentiation, cell cycle, and apoptosis, remains elusive. The present study sought to explore the potential effects on biological behavior when CSCs are labeled with superparamagnetic iron oxide (SPIO) nanoparticles in vitro. The glioblastoma CSCs derived from U251 glioblastoma multiforme were labeled with poly(L-lysine) (PLL)-modified γ-Fe(2)O(3) nanoparticles. The iron uptake of glioblastoma CSCs was confirmed through prussian blue staining, and was further quantified using atomic absorption spectrometry. The cellular viability of the SPIO-labeled glioblastoma CSCs was assessed using a fluorescein diacetate and propidium iodide double-staining protocol. The expressed specific markers and multi-differentiation of SPIO-labeled glioblastoma CSCs were comparatively assessed by immunocytochemistry and semi-quantitative RT-PCR. The effects of magnetic labeling on cell cycle and apoptosis rate of glioblastoma CSCs and their differentiated progenies were assayed using a flow cytometer. The results demonstrated that the cell viability and proliferation capacity of glioblastoma CSCs and their differentiated progenies were not affected by SPIO labeling compared with their unlabeled counterparts. Moreover, the magnetically labeled CSCs displayed an intact multi-differentiation potential, and could be sub-cultured to form new tumor spheres, which indicates the CSCs capacity for self-renewal. In addition, cell cycle distribution, apoptosis rate of the magnetically labeled glioblastoma CSCs, and their differentiated progenies were not impaired. Therefore, the SPIO-labeled CSCs could be a feasible approach in conducting further functional analysis of targeted CSCs.

Capillary-composited microfluidic device for heat shock transformation of Escherichia coli.

This work describes chemical heat shock transformation of foreign plasmid DNA into bacterial host Escherichia coli cells using a capillary-composited microfluidic device. Transformation processes of the loading, mixing, heat shock and recovery of the transformation mixture were carried out automatically in a linear fashion. In addition, by utilizing the capillary with a hollow cylindrical chamber as heating source, simple, low cost local heat shock with accurate heat shock time to transformation mixture was obtained on the microdevice. Results demonstrated that plasmid DNA could be effectively transformed into E. coli, and the transformation efficiency and frequency were as the same level or better than conventional tube-based method. This work complements other microfluidic technologies for potential gene cloning and functional genomics studies.

Superparamagnetic microsphere-assisted fluoroimmunoassay for rapid assessment of acute myocardial infarction.

Rapid assessment of acute myocardial infarction (AMI) was successfully demonstrated using an improved superparamagnetic polymer microsphere-assisted sandwich fluoroimmunoassay to detect two early cardiac markers-myoglobin and human heart-type fatty acid binding protein (H-FABP). This assay used a preparation of superparamagnetic poly(styrene-divinylbenzene-acrylamide) microspheres, glutaraldehyde-coupled capture antibodies (monoclonal anti-myoglobin 7C3 and anti-H-FABP 10E1) grafted onto the polymer microspheres, and a sequential sandwich fluoroimmunoassay using detection antibodies (FITC-labeled anti-myoglobin 4E2 and FITC-labeled anti-H-FABP 9F3). Characterization of the polymer microspheres by TEM, SEM and Fourier transform infrared spectroscopy (FT-IR) showed that the microspheres were uniformly round with an average diameter of 1.12 microm, and had a Fe(3)O(4)-polymer core-shell structure (shell thickness was about 84 nm) with 0.22 mmol/g amino groups on their surfaces. The magnetic behavior of the Fe(3)O(4)-polymer microspheres was superparamagnetic (M(s)=13 emu/g, H(c)=13.1 Oe). Fluorescence images of the post-immunoassay microspheres recorded using a confocal laser-scanning microscope showed that the average fluorescence intensity was correlated with the concentration of cardiac markers, in agreement with the results obtained by an F-4500 FL spectrophotometer; this indicated that the fluoroimmunoassay could be used to semi-quantitatively detect both myoglobin and H-FABP. The detection limit was 25 ng/mL for myoglobin and 1 ng/mL for H-FABP.