The landscape of immune cells in the mouse HCC model with F1 and F3 subtype

In this study, we identified and validated a molecular classification of hepatocellular carcinoma (HCC) patients based on 42 fatty acid degradation (FAD) genes. The F1 subtype was characterized by the lowest expression of FAD genes, whereas F3 subtype had the highest expression levels, and F2 had the intermediate expression levels of FAD genes. We characterized the immune microenvironment in HCC patients from different FAD subtypes. To further explore the immune landscape of HCC, we generated the Nras-driven HCC model (belonging to the F1 subtype) and the Akt1-driven HCC model (belonging to the F3 subtype) by hydrodynamic tail vein injection (HTVi) of oncogenes together with the sleeping beauty transposase. The tumor tissues were resected from the liver 14 weeks after the hydrodynamic delivery of the plasmids, isolated for single cells, and prepared for scRNA-seq. Mouse liver tumors were induced by hydrodynamic tail vein injection (HTVi) of oncogenes together with the sleeping beauty transposase. To generate the Nras-driven HCC model, two expression constructs for human N-Ras (19 μg/mouse) and human c-Myc (1 μg/mouse), together with a plasmid expressing Sleeping Beauty transposase (2 μg/mouse) were co-transfected. To generate the AKT1.MYC model, two expression constructs for Akt1 (10 μg/mouse) and human c-Myc (10 μg/mouse), together with a plasmid expressing Sleeping Beauty transposase (5 μg/mouse) were co-injected. All plasmid DNAs were diluted in phosphate-buffered saline (PBS) and injected at a final volume of 10% of the mouse’s body weight through the tail vein in less than 8 seconds. The tumor tissues were resected from the liver 14 weeks after the hydrodynamic delivery of the plasmids.