Mouse models have become the most common model for defining mechanisms of atherosclerotic disease. Many genetic manipulations have enabled the development of atherosclerosis in mice due to either endogenous or diet-induced hypercholesterolemia. This availability of lesion-susceptible mice has facilitated many studies using pharmacological and genetic approaches. Unfortunately, this expansive literature on mouse atherosclerosis has generated many contradictions on the role of specific pathways. A contributor to these inconsistencies may be the multiple modes in which atherosclerosis is evaluated. Also, for each specific technique, there are no consistent standards applied to the measurements. This chapter will discuss the imaging, biochemical, and compositional modes of evaluating atherosclerosis with suggestions for standard execution of these techniques.
"Animal models are useful both for the definition of dietary regimes that induce and for the assessment of treatments that prevent or revert vascular lesions. For a thorough evaluation of such effects it is useful to make reference to a large panel of biological markers (Daugherty et al. 2009). Proteomic investigations on whole serum (murine data reviewed in Gianazza et al. 2002, 2012a, b) provide quantitative data on several highly expressed proteins. "
[Show abstract][Hide abstract] ABSTRACT: To investigate the influence of diet on serum protein pattern, mice were fed for 8 weeks either control chow or a high-fat diet (containing 21 % w/w milk fat and 0.2 % w/w cholesterol); sera were collected and analyzed by 2-DE. The main positive acute-phase reactant proteins, haptoglobin and hemopexin, were significantly up-regulated in animals receiving the high-fat diet. Data on all other proteins also pointed to an inflammatory condition in these animals. The largest change in concentration was observed for carboxylesterase N, a circulating enzyme seldom connected with lipid metabolism in earlier reports. These observations agree with the notion of a link between diet-induced hyperlipidemia and the inflammatory component of its cardiovascular sequels in humans, but the effects in the experimental animals are massive and obviously affect most of the major serum proteins.
[Show abstract][Hide abstract] ABSTRACT: It is well established that activation of the unfolded protein response (UPR) occurs during all stages of plaque development and progression. Mouse models of atherosclerosis have allowed the study of the UPR in the complex environment of lesions which are composed of different cell types, abundant extracellular matrix, as well as necrotic and apoptotic areas. For proper evaluation of the UPR, the processing, sectioning, and staining of lesions have to be performed in a consistent and reproducible manner. In this chapter, we describe in detail how to embed aortic roots in paraffin, cut serial sections through the aortic root, measure lesion size, and stain paraffin sections by immunohistochemistry, immunofluorescence, and the terminal dUTP nick-end-labeling (TUNEL) technique, with special attention to the proteins involved in the UPR.
Methods in enzymology 01/2011; 489:23-46. DOI:10.1016/B978-0-12-385116-1.00002-9 · 2.09 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The architectonics and cell composition of the human large arteries are not sufficiently understood. The present study is the first to undertake an analysis of the distribution and quantities of HLA-DR-expressing cells in grossly undiseased human intima using immunohistochemical and immunofluorescent analysis, complemented by the advantages of confocal microscopy. The study revealed a widespread distribution of HLA-DR-expressing cells throughout the intimal space where the cells were integrated into continuous networks via long cell processes. Numbers of HLA-DR+ cells were found to be significantly larger in the middle third of the intima than in the superficial and deep intimal portions. We speculate that a widespread distribution of HLA-DR-expressing cells in the intima of normal human aorta might play a role in the surveillance and maintenance of vascular homeostasis.
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