|
| Status |
Public on Jul 11, 2020 |
| Title |
TGF-b treatment |
| Sample type |
RNA |
| |
|
| Source name |
Human myofibroblast
|
| Organism |
Homo sapiens |
| Characteristics |
gender and age: Male, 46-year-old
tissue: Lung
cell type: myofibroblast
diagnosis: Idiopathic pulmonary fibrosis (IPF)
|
| Treatment protocol |
Expanded myofibroblasts at passage 5 were used for the following experiments. After 24 h of serum deprivation in DMEM containing 0.2% fat-free bovine serum albumin, myofibroblasts were treated with/without JQ1 (1 µM) and/or TGF- (10 ng/ml) for a further 48 h.
|
| Growth protocol |
Lung tissue from a 46-y old male patient who underwent lung transplantation by reason of severe IPF (usual interstitial pneumonia pattern) was obtained after surgery. The patient provided informed consent, and the study was approved by the Ethics Committee of Chiba University, Graduate School of Medicine and Hospital, Japan. Lung tissue was minced into small pieces, and then were incubated in DMEM supplemented with 1 mg/ml collagenase type I, 0.5 mg/ml dispase, 2 U/ml DNase, 0.1 mg/ml streptomycin, and 100 U/ml penicillin at 37°C for 15 min with gentle shaking. After washing twice with DMEM, the resulting pieces were transferred to a 90 mm-culture dish and were dipped with culture medium (10% FBS in DMEM supplemented with streptomycin and penicillin) and cultured at 37°C and 5% CO2. Outgrowth of cells was monitored daily with changing the culture medium every 4 d. When the dish reached confluence (about 14 days in vitro), outgrown cells were harvested as cells at passage 0. Expanded cells at passage 3 were subjected to the immunofluorescent study to verify population of α-SMAhighED-A-FN+S100A4- cells as myofibroblasts. More than 90% of the cells were myofibroblasts.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA of the cells was extracted using ISOGEN plus (Nippon Gene, Toyama, Japan) according to manufacturer instructions.
|
| Label |
biotin
|
| Label protocol |
Biotinylated RNAs were prepared according to the standard FlashTagTM Biotin HSR RNA Labeling Kit protocol from 1000 ng of total RNA.
|
| |
|
| Hybridization protocol |
After fragmentation, total RNAs were hybridized for 18 hr at 48°C in a rotating (60 rpm) hybridization oven using GeneChipTM Hybridization Oven 645. GeneChips were washed and stained in the GeneChipTMFluidics Station 450.
|
| Scan protocol |
GeneChips were scanned using GeneChipTM Scanner 3000 7G (Thermo Fisher Scientific, Inc.).
|
| Description |
miRNA expression data for TGF-b treatment
|
| Data processing |
The expression data was normalized using the Robust Multi-array Average (RMA)
|
| |
|
| Submission date |
Nov 15, 2019 |
| Last update date |
Jul 12, 2020 |
| Contact name |
Yoshitoshi Kasuya |
| Organization name |
Chiba Univ.
|
| Department |
Grad. Sch. Med
|
| Lab |
Biomedical Sci.
|
| Street address |
1-8-1 Inohana Chuo-ku
|
| City |
Chiba |
| ZIP/Postal code |
260-8670 |
| Country |
Japan |
| |
|
| Platform ID |
GPL21572 |
| Series (2) |
| GSE140475 |
Identification of microRNA expression patterns in human myofibroblasts with/without the JQ-1 treatment and/or TGF-b |
| GSE140477 |
Comprehensive transcriptome analysis of human myofibroblasts with/without the JQ-1 treatment |
|
