Fig. 3: Detection of ferroptosis markers in CDAHFD.

Markers of hepatic cell death and ferroptosis were measured in groups of mice fed the standard diet (SD) or choline-deficient L-amino acid-defined high-fat diet (CDAHFD) with an emphasis on the 4-week timepoint (n = 6 per group). A Hepatic MDA measured in mice on SD or CDAHFD for 1, 2, 3, 4 and 6 weeks. B Representative images and quantification of the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) stain (red), with DAPI counterstain (blue), in mice fed SD or CDAHFD for 4 weeks. C, D Representative images of immunohistochemistry (IHC) for 4HNE and Gpx4 in mice on SD or CDAHFD for 4 weeks. To quantify the change in distribution of these epitopes, liver lobules were reconstructed based on the Voronoi principle. The proportion of area positive for 4HNE or Gpx4 is plotted with regards to the relative distance (0–1) within the liver lobule, between the portal triads (value 0) to the centrolobular veins (value 1 on x-axis). Slides for IHC were pooled from two independent experiments. E Hepatic mRNA expression of Gpx4 in mice on SD or CDAHFD on different time points, expressed as calibrated normalized relative quantities (CNRQ). F Western blot to visually inspect the protein level of Gpx4 in mice on SD or CDAHFD for 4 weeks. Results are presented as mean ± standard deviation. Mann–Whitney U test with correction for multiple hypothesis testing; *p < 0.05; **p < 0.01. Magnification 100×, scale bar represents 200 µm.