Effect of aspirin on cholesterol crystallization: A potential mechanism for plaque stabilization

Abstract

Background and aims: Cholesterol crystals (CCs) have been found to be critical in the evolution and progression of atherosclerotic plaque leading up to rupture. This includes triggering inflammation and mechanically traumatizing the plaque and surrounding tissues. Thus, inhibition of crystal formation and degrading the crystals could be an important therapeutic approach in the prevention of cardiovascular events. Because of its physico-chemical properties we examined the effect of aspirin (ASA) on cholesterol crystallization.

Methods: A first experiment tested three amounts of cholesterol (1, 2, 3 g) with a wide range of ASA (0-60 mg) on cholesterol crystallization and volume expansion. A second experiment tested the effect of CCs with and without ASA in perforation of fibrous membrane during crystallization. A third experiment evaluated the effect of ASA on melting CCs in human atherosclerotic plaques. Scanning electron microscopy (SEM) was used to evaluate crystal morphology.

Results: Aspirin significantly inhibited cholesterol crystallization and volume expansion in a dose related fashion and even at physiologic levels (0.3 mg/ml). Moreover, ASA prevented perforation of fibrous membranes. By SEM, crystals in human atherosclerotic plaques were found melted with ASA.

Conclusions: Cholesterol volume expansion during crystallization was significantly inhibited and CCs were dissolved in the presence of ASA. Fibrous membranes were not perforated with ASA because of both these effects.

Keywords: Aspirin; Cholesterol crystals; Plaque stabilization.

Fig. 1

Change in volume expansion (ΔVE) during crystallization. Graduated cylinders demonstrating crystal expansion with (a) pure cholesterol (3 g); (b) partial attenuation in ΔVE with cholesterol and aspirin at 10 mg; (c) total attenuation of ΔVE with cholesterol and aspirin at 50 mg.

Fig. 2

Absolute volume expansion with and without aspirin. Effect of aspirin on volume expansion during cholesterol crystallization (a = 3 g, b = 2 g, c = 1 g cholesterol).

Fig. 3

Change in volume expansion (ΔVE) at physiologic level of aspirin. Reduction of ΔVE at physiologic levels of aspirin at 0.3 mg compared to higher dose at 3.0 mg.

Fig. 4

Effect of aspirin on cholesterol crystal morphology (a) SEM demonstrating cholesterol crystals with sharp tipped edges; (b, c) SEM demonstrating blunted tipped cholesterol crystals when formed in the presence of aspirin.

Fig. 5

Composition of pericardial membrane. Histology of pericardial membrane stained with toluene blue and transmission electron microscopy of the same tissue demonstrating extensive collagen composition. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Fig. 6

Effect of aspirin on fibrous tissue injury. (a) Tube with cholesterol crystals expanding and disrupting pericardium during crystallization. (b, c) SEM of cholesterol crystals perforating the pericardial membrane during crystallization. (d) Tube with cholesterol crystals and aspirin (50 mg) with minimal encroachment on the pericardial membrane. (e, f) SEM of cholesterol crystals formed in the presence of aspirin demonstrating very few crystals perforating the membrane surface.

Fig. 7

Effect of aspirin on cholesterol crystal morphology. Scanning electron micrographs of matching segments of human carotid plaques (left: a, b, c) low power and higher power demonstrating presence of sharp edged crystals in plaque incubated in physiologic buffered saline. (right: d, e, f) Segment of plaque incubated physiologic buffered saline with aspirin demonstrates plaque with loss of sharp edges and fewer crystals perforating the intimal surface.