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| Status |
Public on Apr 03, 2023 |
| Title |
Progenitor Endothelial Cells, 0 hours, Replicate 3 |
| Sample type |
RNA |
| |
|
| Source name |
Progenitor Endothelial Cells, 0 hours, Replicate 3
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Progenitor Endothelial Cells
estradiol treatment: 00hrs
|
| Treatment protocol |
When P-SMC and P-EC reached 80% confluence, the cells washed thrice with phenol red free DMEM/F12 medium and subsequently fed medium (DMEM/F12 supplemented with 2.5% steroid free serum and 1x AA) containing either vehicle (DMSO 1ul/ml) or 17β-estradiol (10nM). Following treatment with estradiol for the prescribed times (0, 4 and 30 hours) the cells were washed with PBS and processed for total RNA isolation.
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| Growth protocol |
Progenitor SMCs were grown to confluence in M231 medium supplemented with smooth muscle growth supplement (SMGS; Cascade Biologics Inc.), 1x antibiotic/antimycotic (AA) mix and 20% FCS. Progenitor ECs were cultured in phenol red free endothelial basal medium (EBM-2; Lonza) supplemented with 1x AA, endothelial growth supplement (EGM-2 bullet kit; Lonza) and 20% FCS.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from vehicle-treated and estradiol-treated samples using QIA shredder and RNAeasy Mini kit (Qiagen) according to manufacturer’s instruction. The quality of the isolated RNA was determined with a NanoDrop ND 1000 (NanoDrop Technologies, Delaware, USA) and a Bioanalyzer 2100 (Agilent, Waldbronn, Germany). Only the samples with a 260 nm/280 nm ratio between 1.8–2.1 and a 28S/18S ratio within 1.5–2.0 were further processed.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA were prepared from 2 ug total RNA using Affymetrix GeneChip One-Cycle Target Labeling and Control Reagents according to the manufacturer’s protocol.
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| |
|
| Hybridization protocol |
10 μg of cRNA/sample were hybridized for 16 h at 45°C. Washing and staining with streptavidin-conjugated phycoerythrin were done using a GeneChip Fluidics Station 400 (Affymetrix).
|
| Scan protocol |
Standard Affymetrix procedures using a Hewlett-Packard scanner
|
| Description |
Endothelial Cells, 0 hours, Replicate 3
|
| Data processing |
Raw gene expression data generated by the GeneChip Operating Software (Affymetrix) were normalized for all probe sets on the array using the Robust Multichip Average (RMA) method.
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| |
|
| Submission date |
Apr 08, 2021 |
| Last update date |
Apr 03, 2023 |
| Contact name |
Lennart Opitz |
| Organization name |
FGCZ Zuerich
|
| Street address |
Winterthurerstr. 190
|
| City |
Zuerich |
| ZIP/Postal code |
8057 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL16329 |
| Series (1) |
| GSE171709 |
Differential Effects of Estradiol on Progenitor -Smooth Muscle Cell and -Endothelial Cell Growth and Gene Response. |
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