| COMMENTARIES |
Role of Cathepsin K in Structural Changes of the Brachiocephalic Artery during Progression of Atherosclerosis in ApoE-Deficient Mice
Andriy O. Samokhin, Andre Wong, Paul Saftig*, Dieter Brömme, Department of Oral Biological and Medical Sciences, Faculty of Dentistry, University of British Columbia, Vancouver, Canada, *University of Kiel, Kiel, Germany
Introduction
Since the discovery of the potent collagenolytic and elastolytic activities of cathepsin K several research groups have studied its role in atherosclerotic lesion development [1-3]. Extracellular matrix remodeling in the brachiocephalic artery of ApoE-deficient mouse on a high fat diet has been widely used as a model relevant for human atherosclerotic plaque development [4]. Different compositions of the high fat diet are used to accelerate the development of atherosclerosis. The use of cholate in those diets is usually discouraged as it might induce hepatic fibrosis with collagen accumulation and the activation of proinflammatory genes [5-7]. However, atherosclerosis is also recognized as a chronic inflammatory disease and it was shown that the inclusion of cholate to an atherogenic diet increases the destruction of elastic fibers in the tunica media [8]. The link between dietary cholate, collagen/elastin metabolism and increased arterial inflammation prompted us to consider a high fat diet with 1% cholesterol and 0.5% sodium cholate as a useful model to study the role of cathepsin K in atheroma development in brachiocephalic arteries of apoE-/- mice with potentially similar features to human atherosclerotic plaques.
Experimental Design
The effects of an 8- and 16-week cholate-containing high fat diet (HFD) on the structural changes of brachiocephalic arteries in ApoE-/- mice was compared between cathepsin K-expressing and cathepsin K-deficient mice.
Results
After 8 weeks of high fat diet, the sizes of atherosclerotic plaques in brachiocephalic arteries did not differ significantly between cathepsin K-deficient and cathepsin K-expressing mice but the amount of collagen was more than 5 times higher in the cathepsin K-deficient litter group. Here, collagen was distributed more evenly throughout the plaques, whereas in cathepsin K-expressing mice collagen was accumulated in the fibrous cap region that allowed an easy recognition of so-called buried fibrous caps. Buried caps are considered as evidence of previous plaque ruptures [9]. Though the number of buried fibrous caps was significantly smaller in cathepsin K-deficient mice, they were thicker as revealed by their increased collagen content. These results testify that the collagenolytic activity of cathepsin K may directly control plaque stability. Cathepsin K-deficient mice on 16 weeks of HFD had also smaller plaque sizes. Simultaneously, the collagen content in cathepsin K-expressing mice increased 4 times from 8 to 16 weeks on HFD and became almost undistinguishable from those in cathepsin K-deficient litters.
This finding raised the question about other collagenolytically independent activities of cathepsin K that could contribute to the increased collagen content. Smooth muscle cells are the main source of de novo collagen synthesis in plaques [10]. During the atherosclerotic progression smooth muscle cells migrate from the media into the intima where they undergo a transformation from a contractile to a synthetic type and are associated with the formation of a plaque-forming extracellular matrix, mainly consisting of collagen and elastin fibers. After 16 weeks of HFD immunostaining for smooth muscle cells revealed a significant difference between the two groups of mice: medial smooth muscle cell content decreased almost two-fold in cathepsin K-expressing mice when compared to the cathepsin K-deficient litter group. DNA fragmentation assays revealed that mice lacking cathepsin K have a significantly lower percentage of cells undergoing apoptosis and apoptotic signals were exclusively found in plaque areas that testifies in favor of smooth muscle cell migration rather then their death in the medial area. To migrate from the media into the intima, smooth muscle cells have to pass through basement membranes and the elastic laminae sheets. As cathepsin K exhibits a potent elastolytic activity it can accelerate smooth muscle cell migration. Light and electron microscopy analysis revealed that cathepsin K-expressing mice have a significantly higher number of elastic fiber breaks that also correlated with higher levels of serum desmosine, a surrogate marker of elastin degradation. These results and the lower number of medial macrophages at 8 and 16 weeks of HFD in cathepsin K-deficient mice suggest that cathepsin K facilitates both the migration of smooth muscle cells from the tunica media and macrophages into medial areas.
Conclusion
Results of this study show that the elastolytic and collagenolytic activities of cathepsin K play a significant role in the remodeling of the brachiocephalic artery. We suggest that the inclusion of cholate in a high fat diet permits the observation of striking differences in extracellular matrix synthesis/degradation between apoE-/- and apoE-/-catK-/- mice as mirrored in smaller plaque sizes and reduced smooth muscle cell loss in the tunica media of cathepsin K-deficient mice. Plaques in these mice contained less buried fibrous caps which may indicate a higher stability against rupture. Thus, a selective inhibition of cathepsin K might be a beneficial treatment strategy for atherosclerosis and may stabilize plaques.
References