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| Status |
Public on Apr 30, 2024 |
| Title |
human_Deionized_water_24_0_na_F06 |
| Sample type |
SRA |
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| Source name |
primary hepatocytes
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| Organism |
Homo sapiens |
| Characteristics |
strain: 10 donors pooled
cell type: primary hepatocytes
chemical: Deionized_water
concentration: solvent control
concentration units: na
exposure duration_(hours): 24
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| Treatment protocol |
Using 48-well plates, CD-1, B6129SF2/J and PPARa-KO mouse hepatocytes were seeded at a density of 0.6 x 10^6 cells/mL, and rat and human hepatocytes were seeded at a density of 1.3 x 10^6 cells/mL. After a 24 h adaptation period, hepatocytes from each species/strain were treated for 12, 24 or 72 h with supplemented MCM media containing solvent control in the presence or absence of HFPO-DA (0.1, 5, 50 or 500 μM) or one of the following positive controls: GW7647 (0.01, 0.1, 1 or 10 μM), rosiglitazone (0.01, 0.1, 1 or 10 μM), acetaminophen (0.3, 1, 3 or 10 mM) or d-galactosamine (0.3, 1, 3 or 10 mM). Deionized water (1%) served as the solvent control for HFPO-DA and dimethylsulfoxide (DMSO, 0.1%; cell culture grade; Sigma Aldrich) served as the solvent control for the remaining test chemicals. Treatment solutions were replaced every 24 h. For each species/strain, treatment groups were performed in triplicate wells for 12 and 72 h treatment durations, and quadruplicate wells for the 24 h treatment duration.
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| Growth protocol |
Hepatocytes were isolated from the livers of 8 – 12 week-old male CD-1 mice (CD-1 IGS, strain code: 022) purchased from Charles River (Raleigh, NC) and 11 week-old male B6129SF2/J mice (stock #101045) and 11 week-old male PPARa-null mice (B6;129S4-Pparatm1Gonz/J, stock #008154) purchased from The Jackson Laboratory (Bar Harbor, ME). Rat hepatocytes were isolated from the livers of 8 – 12 week-old male Sprague Dawley (SD, strain code: 001) rats purchased from Charles River. Human hepatocytes were isolated from a pool of 10 donor livers (5 male and 5 female). Hepatocytes from all species/strains were isolated using a two-step enzymatic digestion of liver tissue as described in Mudra and Parkinson (2001). Primary hepatocytes were plated in a collagen-sandwich configuration on 48-well plates. Hepatocytes were maintained in Modified Eagle's medium, Dr. Chee’s modification (MCM; Gibco, Grand Island, NY) supplemented with 11.5% NaHCO3 (Sigma Aldrich), 0.88% L-arginine (Sigma Aldrich), 3% L-glutamine (Sigma Aldrich), 0.05% thymidine (Sigma Aldrich), 1% (v/v) ITS+ (BD Biosciences, Bedford, MA), 1% (v/v) penicillin/streptomycin (5,000 U/mL penicillin, 5,000 μg/mL streptomycin (Sigma Aldrich), and 0.001% 10 mM dexamethasone (Sigma Aldrich). Cell cultures were incubated in a humidified culture chamber (37 ± 2 °C at 95% relative humidity, 95/5% air/CO2).
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| Extracted molecule |
total RNA |
| Extraction protocol |
Following treatment with HFPO-DA or positive controls (i.e., GW7647, rosiglitazone, acetaminophen or d-galactosamine) for 12, 24 or 72 h, hepatocytes were lysed using TempO-Seq® Enhanced Lysis Buffer and processed according to the TempO-Seq® protocol by BioSypder Technologies.
TempO-Seq® protocol by BioSypder Technologies
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
Sequencing reads are demultiplexed using the standard sequencing instrument software for each sample using the barcodes to give a FASTQ file for each
no adapter trimming for TempO-Seq fastqs, default settings for clusters passing filters
bcl2fastq v2.17.1.14 or v2.18.0.12 for demultiplexing
fastqs were aligned to a pseudo transcriptome using Bowtie allowing up to 2 mismatches
Assembly: probe-based targeted sequencing
Supplementary files format and content: comma-separated text files include raw count values for each gene_probe
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| Submission date |
Nov 20, 2023 |
| Last update date |
Apr 30, 2024 |
| Contact name |
Melissa Marie Heintz |
| E-mail(s) |
mheintz@toxstrategies.com
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| Phone |
6106986908
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| Organization name |
ToxStrategies
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| Street address |
31 College Place, Suite B118
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| City |
Asheville |
| State/province |
NC |
| ZIP/Postal code |
28801 |
| Country |
USA |
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| Platform ID |
GPL16791 |
| Series (1) |
| GSE248251 |
Comparison of transcriptomic profiles between HFPO-DA and positive controls for PPARa, PPARg or cytotoxicity in mouse, rat, and pooled human hepatocytes |
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| Relations |
| BioSample |
SAMN38333832 |
| SRA |
SRX22584623 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
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