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Single-cell RNA sequencing of liver CD45+ immune cells in mouse acute-on-chronic liver failure

ABSTRACT: Single-cell RNA sequencing was performed on liver CD45+ immune cells isolated from mice subjected to acute-on-chronic liver failure (ACLF). Three experimental groups were included: CCl4-treated control mice, ACLF mice treated with vehicle, and ACLF mice treated with the EP300 inhibitor CCS1477. Male C57BL/6J mice were intraperitoneally injected with carbon tetrachloride (CCl4, 0.2 ml/kg, twice weekly) for 8 weeks. ACLF was induced by administration of a double dose of CCl4 (0.4 ml/kg), followed by intraperitoneal injection of Klebsiella pneumoniae. CCS1477 was administered intraperitoneally 2 hours before bacterial challenge. Liver CD45+ immune cells were isolated 36 hours after K. pneumoniae injection and subjected to BD Rhapsody whole-transcriptome single-cell RNA sequencing.

ORGANISM(S): Mus musculus

PROVIDER: GSE338157 | GEO | 2026/07/14

REPOSITORIES: GEO

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Project description:Since xenobiotics enter the organism via the liver, hepatocytes must cope with numerous perturbations. In particular, exposure to hepatotoxic agents resulting in liver injury triggers a strong alteration in gene expression. The contribution of these transcriptional regulatory networks to the propagation of injury (i.e. by modulation of metabolic pathways, inflammation, proliferation) are not fully understood. Therefore, we conducted a time-resolved gene array study on mouse liver after exposure to a single intraperitoneal injection of the hepatotoxic compound carbon tetrachloride (CCl4). This model induces a transient injury which reaches its maximum between 2 to 3 days after injection, followed by a precise regenerative response. The experiment included early time points (i.e. 2 and 8h) and late time points (1 to 16 days) to cover the initial events induced by CCl4 metabolization and cell stress, and the regenerative phase between 2-6 days after intoxicaiton. C57BL/6N male mice (8-12 weeks old) were obtained from Charles Rivers (Sulzfeld, Germany). The local Ethics committee approved the experiments. 4-5 mice were used for each time point. Mice received a single intraperitoneal injection of CCl4 (1.6 g/kg body weight) dissolved in 0.5 ml olive oil. The liver tissue was collected after 2h, 8h, and 1, 2, 4, 6, 8 and 16 days following CCl4 administration. As control, mice received 0.5 ml of olive oil intraperitonealy for 1 or 8 days. Healthy liver was collected from untreated mice. At the indicated time points, mice were anesthetized and blood was collected from the heart with a 25 gauge needle, using EDTA as anticoagulant, for analysis of the liver transaminases GOT and GPT in serum. Afterwards, the liver was resected, washed in ice cold PBS and sectioned for further analysis. Liver tissue sections of about 0.5 cm2 were snap frozen in liquid nitrogen and stored at -80ﾰC for subsequent isolation of proteins or RNA. RNA was isolated from mouse liver tissue using the Phenol/Chloroform method (Trizolﾮ, Qiagen, Hilden, Germany) according to the manufacturerﾒs instructions. RNA concentration and integrity were determined spectrophotometrically in a Nanodropﾮ2000 (ThermoScientific, Waltham MA, USA) and in a Bioanalyzerﾮ (Agilent, Waldbronn, Germany), respectively.

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Single-cell RNA sequencing of liver CD45+ immune cells in mouse acute-on-chronic liver failure

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