|
| Status |
Public on May 17, 2018 |
| Title |
ATMSC-MCEV-c |
| Sample type |
RNA |
| |
|
| Source name |
ATMSC-MCEV
|
| Organism |
Homo sapiens |
| Characteristics |
gender: female
cell type: adipose tissue-derived mesenchymal stem/stromal cells
treatment: MC-EVs
|
| Treatment protocol |
The culture medium was removed, the wells were washed in PBS and the contents were collected for analysis.
|
| Growth protocol |
AT-MSCs at passages 4-6 were seeded in 96-well plates at 2.5 * 103 cells/cm2. ls. The cells were cultured for 18 days at 37°C and 5% CO2 in a humidified atmosphere during which the medium was refreshed every 2-3 days. Each time, EVs were freshly added.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from the AT-MSCs using the miRCURY™ RNA Isolation Kit (Exiqon A/S) according to the manufacturer's instructions.
|
| Label |
biotin
|
| Label protocol |
The starting amount of total RNA was 100 ng. 15 ug of cRNA was used for single-stranded cDNA-synthesis (sscDNA). This single-stranded DNA sample is then treated with a combination of uracil DNA glycosylase (UDG) and apurinic/apyrimidinic endonuclease 1 (APE 1) that specifically recognizes the unnatural dUTP residues and breaks the DNA strand. DNA is labeled by erminal deoxynucleotidyl transferase (TdT) with the Affymetrix® proprietary DNA Labeling Reagent that is covalently linked to biotin. GeneChip® WT Plus Reagent Kit, Manual Target Preparation for GeneChip® Whole Transcript Expression Arrays, P/N 703174 Rev.5, Affymetrix.
|
| |
|
| Hybridization protocol |
A hybridization cocktail is prepared including the labeled, fragmented target and hybridization controls. The target is then hybridized to the GeneChip array in cartridge format during 16-hour incubation at 45 °C. Immediately following hybridization, the array is washed and stained with streptavidin phycoerythrin conjugate using an automated protocol on the GeneChip® Fluidics Station 450, followed by scanning on a GeneChip® Scanner. GeneChip® Expression Wash, Stain and Scan User Manual For Cartridge Arrays, P/N 702731 Rev. 4, Affymetrix
|
| Scan protocol |
GeneChips were scanned using the Affymetrix GeneChip Scanner 3000
|
| Description |
MC-EVs (c)
|
| Data processing |
The data were analyzed with R Statistical Software (R 3.4.0) using the Robust Multichip Average (RMA) algorithm of the oligo package as a normalization method. A modified t-statistics was calculated using the limma package. A false discovery rate (FDR) of < 0.05 was used to filter differentially expressed genes between treatments and control (EV-depleted MM).
|
| |
|
| Submission date |
Aug 09, 2017 |
| Last update date |
May 17, 2018 |
| Contact name |
Arjen WH Gebraad |
| E-mail(s) |
arjen.gebraad@helsinki.fi
|
| Organization name |
University of Helsinki
|
| Department |
Department of Oral and Maxillofacial Diseases
|
| Lab |
Translational Research on Oral and Maxillofacial Sciences Group
|
| Street address |
Haartmaninkatu 8, PO box 63
|
| City |
Helsinki |
| State/province |
NA |
| ZIP/Postal code |
00014 |
| Country |
Finland |
| |
|
| Platform ID |
GPL23126 |
| Series (1) |
| GSE102401 |
Effect of monocyte- and osteoclast-derived extracellular vesicles on adipose tissue-derived mesenchymal stem/stromal cells |
|
