Figure 4. HFD-feeding leads to hepatic steatosis after 24 weeks but inflammation becomes apparent after 40 weeks. (A) Representative pictures from H&E-stained liver sections of LFD and HFD mice after 24 (left), 40 (middle) and 52 (right) weeks of diet with inflammatory foci (arrows; insets). (B) Quantification of hepatic triglycerides. (C) Determination of steatosis grade (0=0-5%; 1=5-33%; 2=33-66%; 3=66-100% coverage), and (D) percentage of microvesicular over macrovesicular steatosis. (E) Determination of NAFLD Activity Score (NAS; sum of steatosis + lobular inflammation + ballooning), score >5 represents steatohepatitis (SH) and pathological scores <3 non-steatohepatitis (NSH). (F) Quantification of inflammatory foci per 5 random fields under 200x magnification and (G) determination of ballooning score. (H-L) mRNA expression levels of tumor necrosis factor (Tnf), monocyte chemotactic protein-1 (Mcp1), macrophage marker (F4/80), interleukin-1β (Il1β), and interleukin-10 (Il-10) in the AT. All mRNA expression data were normalized to the LFD24 group and expressed as mean ±SEM (n=7-8). Significance level set at p<0.05. *=significant from LFD at same time point, †=significant from same diet 24w, ‡=significant from same diet 40w.