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Sample GSM590832 Query DataSets for GSM590832
Status Public on Aug 25, 2017
Title 5532 - 06h
Sample type RNA
 
Source name DENV/5532-infected
Organism Homo sapiens
Characteristics infection: DENV/5532
time point: 06h
cell type: Dendrític Cells
Treatment protocol Ten mdDCs cultures were mock-infected or infected with DENV3/5532 at an MOI of 5 for two hours in the absence of FBS. The viral inoculum was removed and cells were washed and cultured in RPMI 1640 plus 10% of FBS and 100 IU/µg/mL of penicillin/streptomycin (Gibco-BRL, Grand Island, NY).
Growth protocol Monocyte-derived dendritic cells (mdDCs) were generated using peripheral blood from healthy volunteers after informed consent and approval of the FIOCRUZ Research Ethics Committee (number 514/09). Mononuclear cells were separated by density gradient using Lymphocyte Separation Medium (Lonza, Walkersville, USA) and CD14+ cells were purified by magnetic immunosorting (Miltenyi Biotec, Auburn, CA), according to the manufacturer’s recommendations. CD14+ cells were cultured for 6-7 days in RPMI 1640 medium containing 100 ng/mL interleukin-4 (IL-4) and 50 ng/mL granulocyte-monocyte colony stimulating factor (GM-CSF; PeproTech, Rocky Hill, NJ), 10% fetal bovine serum (FBS) and 100 IU/µg/mL of penicillin/streptomycin and 2 mM of L-Glutamine (Gibco-BRL, Grand Island, NY). The cultures were assessed by flow cytometry and only cultures with more than 80% of CD1a+ and less than 5% of CD14+ purity were used. C6/36 cells were grown in Leibovitz-15 medium supplemented with 5% FBS and 0.26% of tryptose and 25 µg/mL of gentamicin (all reagents from Gibco-BRL, Grand Island, NY).
Extracted molecule total RNA
Extraction protocol Cells were collected at 6, 12, 24 and 48 hours post infection (hpi) and used to extract total RNA using RNaesy Mini Kit according to the manufacturer’s recommendations (QIAGEN, Valencia, CA). The RNA of all ten mdDC cultures was pooled (75 µg of each) for microarray experiments and compared to mock infected cells using Human Gene 1.0 st v. 1 array GeneChip slides from Affymetrix (Santa Clara, CA). The RNA pool was processed as according to the manufacturer’s recommendations (Affymetrix, Santa Clara, CA) for the amplification and in vitro transcription, purification, tagging, hybridization and scanning of the slides (GeneChip® Scanner 3000).
Label biotin
Label protocol Single-stranded cDNA was generated from the amplified cRNA with the WT cDNA Synthesis Kit (Affymetrix) and then fragmented and labeled with the WT Terminal Labeling Kit (Affymetrix).
 
Hybridization protocol Samples were hybridized with GeneChip Human Gene 1.0 ST Arrays (Affymetrix) and scanned at the LNLS- Campinas Brazil facility.
Scan protocol Array scanning was performed according to the manufacturer's instruction (Affymetrix)
Description RMA expression value derived from Expression Console software; core-gene analysis
Data processing The generated files were analyzed for quality by the software Expression Console (Affymetrix, Santa Clara, CA). The same software was used to normalize the data by the RMA method. Log-ratio between DENV3/5532 infected-mdDCs and mock-infected-mdDCs intensity signal values were generated and a list of differential expressed genes was obtained by using a 2 our higher fold-change. Data clustering was performed by the Cluster 3.0 software with a Euclidian’s distance. Tool for Analysis of GO Enrichments (TANGO), in Expander software was used to obtain a GO with statistical enhancer values of 0.05, and, NCBIEntrez Gene database (www.ncbi.nih.gov) and the Gene Ontology database (www.geneontology.org), to define the most relevant cell signaling and metabolic pathways during mdDC infection with DENV3 5532.
 
Submission date Sep 07, 2010
Last update date Aug 25, 2017
Contact name Guilherme Ferreira Silveira
E-mail(s) gfsilveira@gmail.com
Organization name Instituto Carlos Chagas
Street address Prof Algacyr Munhoz Máder
City Curitiba
ZIP/Postal code 81350-010
Country Brazil
 
Platform ID GPL6244
Series (1)
GSE23986 Dengue virus type-3 isolated from a fatal case with visceral complications induces potent inflammatory responses associated with apoptosis in human monocyte-derived dendritic cells

Data table header descriptions
ID_REF
VALUE RMA data

Data table
ID_REF VALUE
7892501 7.1821
7892502 6.07935
7892503 5.46834
7892504 9.80554
7892505 4.23021
7892506 6.08264
7892507 5.80605
7892508 7.02788
7892509 11.3559
7892510 5.23048
7892511 3.90561
7892512 7.64247
7892513 5.57899
7892514 10.4384
7892515 8.87221
7892516 6.23745
7892517 7.15909
7892518 4.60267
7892519 6.309
7892520 9.6122

Total number of rows: 33297

Table truncated, full table size 516 Kbytes.




Supplementary file Size Download File type/resource
GSM590832_5532_6h_HuGene-1_0-st-v1_.CEL.gz 4.1 Mb (ftp)(http) CEL
GSM590832_5532_6h_HuGene-1_0-st-v1_.rma-gene-default.chp.gz 256.0 Kb (ftp)(http) CHP
Processed data included within Sample table
Processed data provided as supplementary file

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