Enhanced atherosclerosis and kidney dysfunction in eNOS(-/-)Apoe(-/-) mice are ameliorated by enalapril treatment

Abstract

Hypertension and atherosclerosis are each important causes of morbidity and mortality in the developed world. We have investigated the interaction between these conditions by breeding mice that are atherosclerotic due to lack of apolipoprotein (apo) E with mice that are hypertensive due to lack of endothelial nitric oxide synthase (eNOS). The doubly deficient mice (nnee) have higher blood pressure (BP) and increased atherosclerotic lesion size but no change in plasma lipoprotein profiles compared with normotensive but atherosclerotic (NNee) mice. The nnee mice also develop kidney damage, evidenced by increased plasma creatinine, decreased kidney weight/body weight ratio, and glomerular lipid deposition and calcification. Enalapril treatment abolishes the deleterious effects of eNOS deficiency on BP, atherosclerosis, and kidney dysfunction in nnee mice. In striking contrast, a genetic lack of inducible NOS, which does not affect BP, has no effect on the development of atherosclerotic lesions in Apoe(-/-) mice. We also observed a positive relationship between BP and size of atherosclerotic lesions These results suggest that the atherogenic effects of eNOS deficiency can be partially explained by an increase in BP and reemphasize the importance of controlling hypertension in preventing atherosclerosis.

Figure 1

(a) BP of NNee and nnee mice. An asterisk indicates a significant increase in the BP of nnee animals compared with NNee controls (P < 0.0001). Number inside bar is sample size. (b) BP of NNee and nnee mice treated with enalapril (0.12 g/L drinking water). Two asterisks indicate a significant decrease in BP of nnee animals treated with enalapril vs. untreated animals (P < 0.001) (but not in BP of enalapril-treated vs. untreated NNee animals); (P = 0.75). Error bars represent SE.

Figure 2

(a) Left panel: Atherosclerotic lesion size expressed on a log scale in 4-month-old female NNee and nnee animals. There is an increase in the lesion size of female nnee mice compared with NNee mice. *P < 0.002. Each dot represents the mean lesion size of 4 sections from each animal. Logarithmic mean value of lesion size (in μm²) from each group is shown to the right of the dots. Data are shown as the range of logarithmic mean ± SE. Right panel: Atherosclerotic lesion size in nnee female mice is reduced by treatment with enalapril (0.12 g/L drinking water) for 2 months. **P = 0.05 compared with untreated nnee female mice. (b) Arrangement is the same as in a, except male mice are shown. *P < 0.0001 compared with NNee mice. **P < 0.02 compared with untreated nnee mice.

Figure 3

Representative histological sections from 4-month-old nnee animals. (a) Proximal aorta section showing atherosclerotic lesions. Sudan IVB with hematoxylin counterstain; ×40. Arrows indicate microaneurysms. (b) A plaque in a small vessel in myocardium. Sudan IVB with hematoxylin counterstain; ×200 (initial magnification). Arrow indicates lumen. (c) Aneurysms in an abdominal aortic section. Sudan IVB with hematoxylin counterstain; ×40 (original magnification). Arrows indicate sites of dissection. (d) Foam cells in glomerulus. Hematoxylin and eosin; ×165. Arrow indicates small area of calcification. (e) Four glomeruli demonstrating the transition from heavy lipid deposition (which appears orange) to dystrophic calcification (which appears dark blue). Sudan IVB with hematoxylin counterstain; ×82.5. (f) Glomerulus of enalapril-treated nnee mouse. Lipid deposition is light (arrow) and is confined to extraglomerular mesangial cells. Sudan IVB and hematoxylin; ×165. This pattern of lipid staining is similar to that seen in NNee mice.

Figure 4

Atherosclerotic lesion size on a log scale expressed as a function of BP in female (left) and male (right) 4-month-old mice. Genotype and drug treatments are as indicated. There is a significant relationship between BP and lesion size in both female (P < 0.01) and male (P < 0.02) mice, independent of how BP was altered (by genotype, treatment, or a combination of the two). n = 36 for females and n = 52 for males. Enal, enalapril.

Figure 5

Analysis of plasma cholesterol by fast performance liquid chromatography. There were no differences in distribution of lipoproteins from NNee mice (closed circles) and nnee mice (open diamonds).

Figure 6

Effect of iNOS deficiency on the size of atherosclerotic plaques. There is no significant difference in the atherosclerotic lesion size between iNOS^+/+Apoe^–/– (IIee) mice and iNOS^–/–Apoe^–/– (iiee) mice on normal chow. Error bars represent SE.

Figure 7

Ratios of kidney weight to body weight in 4-month-old NNee mice and nnee mice treated (+) and untreated (–) with enalapril in drinking water (0.12 g/L) for 2 months. The nnee mice show a significant decrease in the kidney weight/body weight ratio compared with NNee control mice (*P < 0.01). This decrease is abolished by enalapril treatment (**P < 0.002, nnee untreated compared with nnee treated). Error bars represent SE.