| Summary |
We performed transcriptomic profiling of mouse whole liver cells perfused in situ with collagenase and then mechanically dissociated into single cell suspensions, to garner about 40% hepatocytes and 60% non-parenchymal cells. Cells were isolated from male and female C57BL/6J mice that received either PBS vehicle control, or 400 mg/kg acetaminophen (APAP) and perfused 48 hours after injury.
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| Samples |
| GSM ID |
Sample info |
Characteristics |
Description |
| GSM6262596 |
Male APAP |
tissue: liver
Sex: male
strain: C57BL/6J
: 12 weeks
isolation method: Mice were anaesthetized with ketamine/xylazine and then perfused by through the inferior vena cava with 30 mL 1X Liver Perfusion Medium (Gibco by Life Technologies), while the portal vein was cut to allow the perfusate to flow out. In total, 15 mL of Earle’s Balanced Salt Solution (EBSS) containing 10mM Hepes (Ca++ and Mg++, pH 7.4) was then perfused, and finally followed with 30 mL of Liver Digest Medium (Gibco by Life Technologies) to allow complete dissociation of liver cells in situ. All solutions were pre-warmed to 40 °C and delivered at a rate of 4 ml/min. Livers were then extracted and mechanically dissociated in 10 mL of Liver Digest Medium. Cells were filtered through 100 μm cell strainer and filtrate was centrifuged at 50 × g for 2 min at 4 °C to obtain hepatocytes in the pellet while the supernatant was collected as the fraction of NPCs. The hepatocyte pellet was resuspended in wash media (Hepatocyte Wash Medium by Gibco + 0.1 mg/mL DNAse 1 + 10% FBS) and saved. The cell fraction caught in the 100 μm strainer was further digested in NPC Digest Medium (2.5 mg/mL Collagenase IV + 0.1 mg/mL DNAse 1), filtered through a 40 μm strainer, then centrifuged at 300 × g for 5 min at 4 °C. The supernatant was discarded, pellet was resuspended in wash media, and solution was added to the other collection of NPCs. This NPC fraction was used for the single-cell RNA sequencing analysis. |
EE_MA |
| GSM6262597 |
Female APAP |
tissue: liver
Sex: female
strain: C57BL/6J
: 12 weeks
isolation method: Mice were anaesthetized with ketamine/xylazine and then perfused by through the inferior vena cava with 30 mL 1X Liver Perfusion Medium (Gibco by Life Technologies), while the portal vein was cut to allow the perfusate to flow out. In total, 15 mL of Earle’s Balanced Salt Solution (EBSS) containing 10mM Hepes (Ca++ and Mg++, pH 7.4) was then perfused, and finally followed with 30 mL of Liver Digest Medium (Gibco by Life Technologies) to allow complete dissociation of liver cells in situ. All solutions were pre-warmed to 40 °C and delivered at a rate of 4 ml/min. Livers were then extracted and mechanically dissociated in 10 mL of Liver Digest Medium. Cells were filtered through 100 μm cell strainer and filtrate was centrifuged at 50 × g for 2 min at 4 °C to obtain hepatocytes in the pellet while the supernatant was collected as the fraction of NPCs. The hepatocyte pellet was resuspended in wash media (Hepatocyte Wash Medium by Gibco + 0.1 mg/mL DNAse 1 + 10% FBS) and saved. The cell fraction caught in the 100 μm strainer was further digested in NPC Digest Medium (2.5 mg/mL Collagenase IV + 0.1 mg/mL DNAse 1), filtered through a 40 μm strainer, then centrifuged at 300 × g for 5 min at 4 °C. The supernatant was discarded, pellet was resuspended in wash media, and solution was added to the other collection of NPCs. This NPC fraction was used for the single-cell RNA sequencing analysis. |
EE_FA |
| GSM6262598 |
Male Control |
tissue: liver
Sex: male
strain: C57BL/6J
: 12 weeks
isolation method: Mice were anaesthetized with ketamine/xylazine and then perfused by through the inferior vena cava with 30 mL 1X Liver Perfusion Medium (Gibco by Life Technologies), while the portal vein was cut to allow the perfusate to flow out. In total, 15 mL of Earle’s Balanced Salt Solution (EBSS) containing 10mM Hepes (Ca++ and Mg++, pH 7.4) was then perfused, and finally followed with 30 mL of Liver Digest Medium (Gibco by Life Technologies) to allow complete dissociation of liver cells in situ. All solutions were pre-warmed to 40 °C and delivered at a rate of 4 ml/min. Livers were then extracted and mechanically dissociated in 10 mL of Liver Digest Medium. Cells were filtered through 100 μm cell strainer and filtrate was centrifuged at 50 × g for 2 min at 4 °C to obtain hepatocytes in the pellet while the supernatant was collected as the fraction of NPCs. The hepatocyte pellet was resuspended in wash media (Hepatocyte Wash Medium by Gibco + 0.1 mg/mL DNAse 1 + 10% FBS) and saved. The cell fraction caught in the 100 μm strainer was further digested in NPC Digest Medium (2.5 mg/mL Collagenase IV + 0.1 mg/mL DNAse 1), filtered through a 40 μm strainer, then centrifuged at 300 × g for 5 min at 4 °C. The supernatant was discarded, pellet was resuspended in wash media, and solution was added to the other collection of NPCs. This NPC fraction was used for the single-cell RNA sequencing analysis. |
EE_MC |
| GSM6262599 |
Female Control |
tissue: liver
Sex: female
strain: C57BL/6J
: 12 weeks
isolation method: Mice were anaesthetized with ketamine/xylazine and then perfused by through the inferior vena cava with 30 mL 1X Liver Perfusion Medium (Gibco by Life Technologies), while the portal vein was cut to allow the perfusate to flow out. In total, 15 mL of Earle’s Balanced Salt Solution (EBSS) containing 10mM Hepes (Ca++ and Mg++, pH 7.4) was then perfused, and finally followed with 30 mL of Liver Digest Medium (Gibco by Life Technologies) to allow complete dissociation of liver cells in situ. All solutions were pre-warmed to 40 °C and delivered at a rate of 4 ml/min. Livers were then extracted and mechanically dissociated in 10 mL of Liver Digest Medium. Cells were filtered through 100 μm cell strainer and filtrate was centrifuged at 50 × g for 2 min at 4 °C to obtain hepatocytes in the pellet while the supernatant was collected as the fraction of NPCs. The hepatocyte pellet was resuspended in wash media (Hepatocyte Wash Medium by Gibco + 0.1 mg/mL DNAse 1 + 10% FBS) and saved. The cell fraction caught in the 100 μm strainer was further digested in NPC Digest Medium (2.5 mg/mL Collagenase IV + 0.1 mg/mL DNAse 1), filtered through a 40 μm strainer, then centrifuged at 300 × g for 5 min at 4 °C. The supernatant was discarded, pellet was resuspended in wash media, and solution was added to the other collection of NPCs. This NPC fraction was used for the single-cell RNA sequencing analysis. |
EE_FC |
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