Exogenous interferon-gamma enhances atherosclerosis in apolipoprotein E-/- mice

Abstract

A role for interferon-gamma (IFN-gamma) has been implied in the atherogenic process. To determine whether exogenously administered IFN-gamma exerts an effect on the development of atherosclerosis, we intraperitoneally administered either recombinant IFN-gamma (100 U/g body weight) or phosphate buffered saline daily for 30 days to atherosclerosis-susceptible apolipoprotein E-/- mice (16-week-old male mice, n = 11 per group) fed a normal diet. Atherosclerotic lesion size was quantified in the ascending aorta. The number of T lymphocytes and major histocompatibility complex (MHC) class II-positive cells within lesions were also quantified in this region. IFN-gamma administration reduced serum cholesterol concentrations by 15% (P = 0.02). For both groups, the majority of cholesterol was present in very low density lipoproteins, which were modestly reduced in mice receiving IFN-gamma. Despite the decrease in serum cholesterol concentrations, IFN-gamma injections significantly increased lesion size twofold compared to controls (119,980 +/- 18, 536 vs. 59,396 +/- 20,017 micrometer(2); P = 0.038). IFN-gamma also significantly increased the mean number of T lymphocytes (19 +/- 4 vs. 7 +/- 1 cells; P = 0.03) and MHC class II-positive cells (10 +/- 3 vs. 3 +/- 1 cells; P = 0.04) within lesions. These data lend further support to a pro-atherogenic role of IFN-gamma.

Figure 1.

Serum (50 μl) from male apoE−/− mice receiving daily injection of either PBS alone (open symbols) or IFN-γ (solid symbols) was resolved by size exclusion chromatography using a Superose 6 column. Total cholesterol concentrations were then determined from each fraction and expressed as mean absorbency at 490 nm. Values are represented as mean ± SEM of 11 animals from each group. Fraction numbers correspond to VLDL; intermediate, low, and high density lipoproteins (IDL, LDL, and HDL, respectively) are indicated.

Figure 2.

Characteristics of lesions in the ascending aorta receiving daily injection of either PBS alone (open symbols) or IFN-γ (solid symbols) that include atherosclerotic lesion size (A), lesion T lymphocyte cell number (B), and lesion MHC class II-positive cell number (C). For all graphs, each circle represents the value for an individual mouse, square symbols positioned beside a group of circles represents the group mean, and bars represent the SEM. Segments of heart tissue spanning the aortic sinus and ascending aorta were embedded in OCT, sectioned, and stained with Oil Red O (A), a monoclonal anti-mouse Thy1.2 antibody (7 μg/ml) (B), or a monoclonal anti-mouse MHC class II antibody (1:5 dilution; C). Each point in A represents the mean lesion size of 4 sections per mouse, collected every 80 μm starting at the region where the aortic sinus becomes the ascending aorta. In B and C, each point represents the mean cell number associated within lesions for serial sections to those in A. Values are derived from 11 mice per group, with the exception of the IFN-γ-injected mice in B, where 8 mice per group were used. A: *P = 0.038. B: *P = 0.03. C: *P = 0.04.

Figure 3.

Representative sections from a region where the aortic sinus becomes the ascending aorta of PBS- (A, C, E, and G) and IFN-γ- (B, D, F and H) injected mice. A segment of heart tissue spanning the aortic sinus and ascending aorta was embedded in OCT, sectioned and stained with Oil Red O for neutral lipids (A and B); a rabbit antisera to mouse macrophages (1:3000 dilution; C and D); a monoclonal anti-mouse Thy1.2 antibody (7 μg/ml; E and F); or Gomori trichrome (G and H). C, E, G, and D, F, H are from regions encompassed in the boxes depicted in A and B, respectively. Original magnifications, ×40 (A and B) and ×200 (C−H).