Figure 1.
Parameters of hepatic inflammation.
(A, B) Liver sections were stained for infiltrating macrophages and neutrophils (Mac-1) and neutrophils (NIMP). From each liver, 6 random pictures were taken at 200x magnification to cover the whole slide. Positive cells for the specific staining were then counted being indicative for inflammation (C) Result of scoring for inflammation by an experienced pathologist using the HE staining in all groups. (D) Liver sections were stained for T-cells (CD3) and positive cells counted. (E, F) Representative pictures of Mac-1 staining and NIMP staining in the four experimental groups (200x magnification). *P<0.05, **P<0.01, and ***P<0.001, respectively.
Figure 2.
(A–E) Gene expression analysis of the macrophage marker Cd68, monocyte chemoattractant protein 1 (Mcp-1), interleukin 1β (IL-1β), tumor necrosis factor α (Tnf-α) and intercellular adhesion molecule 1 (Icam). Relative expression was normalized to endogenous control gene Cyclophilin A. Data were set relative to group on chow diet. n = 10 per group. *P<0.05, **P<0.01, and ***P<0.001, respectively.
Figure 3.
(A) Scoring for the size and foamy appearance of Kupffer cells using CD68 staining. (B) Scoring for the size and foamy appearance of Kupffer cells using HE staining. A score ranging from 0–3 was given by an experienced pathologist. (C) Representative pictures of the foamy Kupffer cell appearance with CD68 staining (200x magnification). *P<0.05, **P<0.01, and ***P<0.001, respectively.
Figure 4.
(A, B) Plasma and liver cholesterol measurements. (C, D) Plasma and liver triacylglycerol levels. (E) Scoring of liver slides for the accumulation of fat (steatosis) using HE staining. A score ranging from 0–3 (3 = highest steatosis) was given by an experienced pathologist. *P<0.05, **P<0.01, and ***P<0.001, respectively.
Figure 5.
Serum lipid and lipoprotein profiles.
(A, B) Using FPLC, serum lipid and lipoprotein profiles were analyzed in all experimental groups. On the chromatogram, the X-axis represents the fractions present in the mixture as a peak, thereby identifying the different components of the mixture. On the Y-axis, the amount of the different fractions can then be read (nMol/l).
Figure 6.
Hepatic non-cholesterol sterol concentrations.
Hepatic concentrations of (A) campesterol, (B) sitosterol, (C) campestanol and (D) sitostanol were measured. To analyze endogenous cholesterol synthesis, hepatic (E) lathosterol and (F) desmosterol were measured. All values are shown as absolute concentrations (ng/mg tissue). n = 10 per group. *P<0.05, ***P<0.001.
Figure 7.
Effect of plant stanols on macrophages in vitro.
Changes in Tnf-α concentrations in supernatant and LXR target gene expression of bone marrow derived macrophages after incubation with sitostanol (0.6 and 1.2 µm) or desmosterol (0.25, 0.5 and 1.0 µm) and 4 h LPS stimulation. (A) Tnf-α concentrations, (B) LXRα mRNA, (C) Abca1 mRNA, and (D) Abcg1 mRNA expression after sitostanol exposure, (E) Tnf-α mRNA, (F) LXRα mRNA, (G) Abca1 mRNA, and (H) Abcg1 mRNA expression after desmosterol exposure. Data were set relative to cells incubated with cyclodextrin (carrier control). *P<0.05; **P<0.01; ***P<0.001.
Figure 8.
Plasma cholesterol precursor levels in severely obese patients.
Serum levels of (A) desmosterol, (B) lathosterol and (C) cholestanol were measured in control (n = 20), NAFLD (n = 8) and NASH (n = 25) patients. All values are shown as absolute levels (mg/dl serum). *P<0.05.
Table 1.
Population characteristics.
Table 2.
Histological scoring of liver biopsies from NASH subjects.
Figure 9.
Schematic representation of direct effects of sitostanol vs desmosterol.
Table 3.
Composition of the experimental and chow diets.