Reduced progression of atherosclerosis in apolipoprotein E-deficient mice treated with lacidipine is associated with a decreased susceptibility of low-density lipoprotein to oxidation

Abstract

A study has been carried out in the apolipoprotein (apo) E-deficient mouse to investigate the activity of lacidipine (a calcium antagonist with antioxidant properties) in inhibiting the development of atherosclerotic lesions; of particular interest were changes in the susceptibility of low-density lipoproteins (LDL) to oxidation. Mice receiving a Western-type diet to accelerate the development of atherosclerosis were treated orally with vehicle or lacidipine at 3 or 10 mg/kg/day for 8 weeks. Lacidipine treatment (at 3 or 10 mg/kg) had no effect on the plasma lipid profile. However, a significant (P < 0.01) dose-related reduction of 43 and 50% of the aortic lesion area in respect to vehicle-treated mice was observed. Moreover, the resistance of mouse plasma LDL to undergo lipid peroxidation was significantly (P < 0.01) increased in apo E-deficient mice treated with lacidipine. The native LDL-like particle, derived from apo E-deficient mice treated with lacidipine, contained significantly lower concentrations of malonyldialdehyde than the vehicle-treated control group (P < 0.01). After exposure to human umbilical vein endothelial cells, LDL-like particle vitamin E levels (expressed as area under the curve; AUC), were significantly higher (P < 0.01) in both the 3 and 10 mg/kg lacidipine-treated groups, in comparison with the vehicle-treated control animals. We conclude that lacidipine reduced the extent of the atherosclerotic area in hypercholesterolaemic apo E-deficient mice, and that this reduction may be associated with the capacity of the drug to decrease the susceptibility of LDL to oxidation.

Figure 1

Atherosclerotic lesions in sections of the aortic root of the heart in a control apo E-deficient mouse (a), and in an apo E-deficient mouse receiving 3 mg/kg lacidipine (b) (haematoxylin and eosin-stained; original magnification, ×40). In the control mouse (a) extensive, advanced lesions with a well-defined fibrous cap and a necrotic core with nodular deposits of calcification are present. The volume of the lesions in apo E-deficient mouse treated with lacidipine at 3 mg/kg (b) appears markedly reduced, and the lesion is less complex.

Figure 2

Effect of lacidipine at 0 (control), 3 and 10 mg/kg on atherosclerotic lesions measured in the aortic root of apo E-deficient mice after 8-week treatment. Results are expressed as mean ± SEM of the lesion area (×10³µm²) measured in 17 cross-sections through the aortic root of each mouse, over a distance of 240 µm n = 20 in each group; **P < 0.01, in lacidipine-treated animals compared with the controls.

Figure 3

The susceptibility of the mouse low-density lipoprotein (LDL) to undergo lipid peroxidation following incubation with CuSO₄. LDL-like particles were derived from plasma of apo E-deficient mice after 8-week treatment with vehicle (control) or lacidipine at 3 or 10 mg/kg. Results are expressed as mean ± SEM of the lag phase (min); n = 6 pooled samples in each dose-level group; *P < 0.05 in comparison with the 3 mg/kg lacidipine group; **P < 0.01 in comparison with the control group.

Figure 4

Effect of lacidipine on the hydroperoxide content of native low-density lipoprotein (LDL). Malondialdheyde (MDA) content in LDL-like particles derived from apoE-deficient mice after 8-week treatment with vehicle (control) or lacidipine at 3 or 10 mg/kg. The results are expressed as mean ± SEM; n = 6 pooled samples in each dose-level group; **P < 0.01 in comparison with the control group.

Figure 5

Relationship between the hydroperoxide content of native low-density lipoprotein (LDL) [measured as malonyldialdehyde (MDA) in LDL-like particles] and the resistance to oxidation of LDL-like particles (expressed as lag phase in minutes) from the plasma of individual apoE-deficient mice treated with vehicle (control; solid squares) or lacidipine at 3 mg/kg (open circles) or 10 mg/kg (solid triangles). n = 6 pooled samples in each dose-level group.

Figure 6

HUVEC-mediated oxidation of low-density lipoprotein (LDL)-like particles, expressed as the area under the curve (AUC, in nmol/ml/h; mean ± SEM) of malonyldialdehyde (MDA) production during a 24-h incubation period. LDL-like particles were derived from the plasma of apo E-deficient mice after 8-week treatment with vehicle, or lacidipine at 3 and 10 mg/kg. A dose-related reduction in AUC values was observed at both dose levels of lacidipine. **P < 0.01 in comparison with the cell-free medium (CFM) and the vehicle control; n = 6 for vehicle control; n = 11 for lacidipine at 3 mg/kg; n = 6 for lacidipine at 10 mg/kg and n = 8 for CFM.

Figure 7

Vitamin E content, expressed as the area under the curve (AUC, in nmol/ml/h; mean ± SEM) during a 24-h incubation period with human umbilical vein endothelial cells (HUVECs). Low-density lipoprotein (LDL)-like particles were derived from the plasma of apo E-deficient mice after 8-week treatment with vehicle or lacidipine at 3 or 10 mg/kg. Dose-related increases in the AUC values were observed at both dose levels of lacidipine in comparison with the vehicle-treated control group; **P < 0.01; n = 6 for vehicle control; n = 11 for lacidipine at 3 mg/kg and n = 6 for lacidipine at 10 mg/kg.