Fig. 3: Targeted knock-in of the 17β-HSD13 S33A mutation drives NAFLD pathogenesis.

a Schematic representation of the gene targeting strategy of 17β-HSD13 S33A. Homologous recombination was used to replace exon 1 with the corresponding mutant sequence, resulting in a substitution of alanine for serine 33 in 17β-HSD13. b Representative liver gross view, H&E staining, Oil red O staining, and immunohistochemistry (IHC) pictures in the Hsd17b13^33+/+(WT) and Hsd17b13^33A/A (S33A) fed a chow at the age of 5 months; Scale bar, 100μm. WT: n = 5; S33A: n = 5 biologically independent animals. c Transmission electron microscopy (TEM) analysis of lipid droplets (LDs) in the livers of WT and S33A mice. Nu nucleus; mi mitochondria; RER Rough endoplasmic reticulum. Scale bar = 2 μm (upper panels); 1 μm (lower panels); WT: n = 3; S33A: n = 3 biologically independent animals. Serum triglyceride (TG) and cholesterol (TC) levels (d), liver TG and TC contents (e), serum ALT and AST levels (f), serum insulin (g) and serum non-esterified fatty acid (NEFA) (h) were analyzed in the S33A mutant mice compared with their WT littermates at the age of 5 months. WT: n = 9; S33A: n = 10 biologically independent animals. Glucose tolerance test (GTT) (i), insulin tolerance test (ITT) (j), and pyruvate tolerance test (k) were performed at the age of 14–15 weeks. AUC, the area under the curve. WT: n = 5; S33A: n = 5 biologically independent animals. l Lipidomics analysis was performed in the livers from WT and the S33A mutant mice. The expression levels of various lipids species were displayed by heat map.WT: n = 6; S33A: n = 6 biologically independent animals. Data represent mean ± SEM; Significance was calculated by two-tailed student’s t test (d–h). Two-way ANOVA with Bonferroni post hoc analysis was performed for i–k.