Figure 5

Effect of SPARC deficiency in mice fed with a non-alcoholic steatohepatitis-inducing diet. (A) Hematoxylin/eosin (H/E) and oil red staining of liver sections from SPARC^+/+ mice and SPARC^−/− mice after feeding with WD or RD diet for 20 weeks. H/E staining demonstrated more inflammation and necrosis in WD-fed SPARC^+/+ mice compare with WD-fed SPARC^−/− mice. Arrows indicate inflammatory cell infiltration. WD diet increased fat deposition visualized by oil red staining in SPARC^+/+ and SPARC^−/−mice. WD-fed SPARC^−/− exhibited enlarged lipid droplets compared to SPARC^+/+ with WD. Image magnification: 200×. (B) NAFLD activity score (NAS) showing more steatosis, inflammation and ballooning in SPARC^+/+ mice in comparison with SPARC^−/− mice. n = 6–8 mice per group. Data are presented as median and range. *Compared WD vs RD diet in both SPARC^+/+ and SPARC^−/− mice; ^σcompared WD-fed SPARC^+/+ versus WD-fed SPARC^−/− mice. *p < 0.05, ***p < 0.001; ^σp < 0.05; ^σσp < 0.01, ^σσσp < 0.001; Kruskal-Wallis test with Dunn’s post test C) Serum AST levels. Data are mean ± sem. *p < 0.05, Kruskal-Wallis test with Dunn’s post test. n = 6–8 mice per group. D) Analysis of hepatic F4/80 positive cell infiltration by immunofluorescence in liver sections of WD diet-fed SPARC^+/+ and SPARC^−/− mice. Magnification: 200× (E) F4/80 positive cells quantification. Ten random fields were analyzed for each group (x200 magnification), n = 4 mice per group. ***p < 0.001, Mann-Whitney T test. (F) Hepatic IL6 mRNA expression assessed by qrt-PCR ***p < 0.001, Mann-Whitney T test. nd, non-detectable amplification (G–I) Hepatic mRNA expression of tumor necrosis factor-α (TNF-α), IP-10, and FAS/CD95 in response to WD diet in both SPARC^−/− mice and SPARC^+/+ mice compared with animals fed with RD. (J) Hematoxylin/eosin (H/E) staining of adipose tissue sections from SPARC^+/+ and SPARC^−/− mice after feeding with WD or RD diet for 20 weeks. Magnification: x200. (K) Inflammatory cell quantification. Ten random fields were analyzed for each group (x200 magnification). (L) Adipose IL6 mRNA expression assessed by qPCR. *Compared WD vs RD diet in both SPARC^+/+ and SPARC^−/− mice; ^σcompared WD-fed SPARC^+/+ versus WD-fed SPARC^−/− mice. * or ^σp < 0.05, ** or ^σσp < 0.01, Kruskall-Wallis test, with Dunn’s post test.