Carotid Atherosclerosis: Why Some Patients are Symptomatic and Others Not?
Occlusive vascular disease (OVD) is the leading cause of morbidity and mortality in the United States. OVD of the carotid arteries results in stroke or transient ischemic attacks. A significant number of strokes are caused by carotid artery atherosclerosis. Atherosclerosis, commonly referred to as "hardening of the arteries," is a disease where there is build up of plaque in the blood vessel wall. Advanced plaque typically consists of lipid, such as cholesterol, a necrotic core and calcification, surrounded by cells and connective tissue matrix. Over the course of many years, build up of plaque material can compromise the carotid artery lumen Many studies have shown that the risk for stroke increases as the degree of luminal narrowing increases in the carotid artery. Despite the availability of quantitative diagnostic tools such as duplex ultrasound and carotid endarterectomy as the current treatment for occlusive carotid artery disease, incidence of stroke is increasing. In coronary vasculature it has been shown that plaques from symptomatic patients are friable and thus more likely to undergo plaque rupture to precipitate symptoms. If this happens in the carotid arteries, this can lead to occlusion of cerebral arteries to cause transient ischemic attack (TIA) or stroke. Therefore, the goal of this study was to identify morphological features of carotid plaques and determine changes in the function of vascular smooth muscle cells in the plaque at the cellular level. I proposed the hypothesis; Plaques from symptomatic patients have thinner fibrous caps overiying a iarge necrotic core, contain higher numbers of macrophages and exhibit higher teveis of apoptosis in vascuiar smooth muscie ceiis (VSMCs) than piaques from asymptomatic patients In order to test this hypothesis, carotid endarterectomy tissue specimens following surgery were obtained from symptomatic and asymptomatic patients with carotid stenosis. Symptomatic patients exhibited hemispheric transient ischemic attack, stroke, and amaurosis fugax within the last two weeks prior to surgery, and asymptomatic patients were those who had angiographic stenosis of 70 - 98%. Histologic sections of the plaque were examined using computer-assisted morphometric analysis. The cross-sectional area of plaque occupied by necrotic/lipid core, the thickness of the fibrous cap as well as the area of the fibrous cap were determined. The presence of macrophages/monocytes collections within the fibrous cap, in the shoulders of the plaque, and at the base of the plaque were measured to determine plaque inflammation, /n-s/fu TUNEL was performed to determine percent apoptotic cells in the fibrous cap and base of plaques. Ten plaques within this cohort were used to isolate VSMCs for /n-v/fro studies, and the purity of the cells was confirmed by positive staining to smooth muscle a-actin and caldesmon. TUNEL and Annexin V binding assays were performed to identify apoptotic death in cultured VSMCs isolated from the plaques followed by cell proliferation assay and cel) cycle analysis. Total protein expression and the activity of caspases were performed to identify intrinsic and/or extrinsic pathways involved in the apoptosiis of VSMCs. The mean percent angiographic stenosis in the asymptomatic and symptomatic plaques was 84 + 11% and 83 + 13%, respectively. The area of the necrotic core in asymptomatic plaques was 0.254 ± 0.013 mrn^ and in symptomatic plaques was 0.321 ± 0.012 mrm (p = 0.0006). The mean thickness of the fibrous cap from asymptomatic plaques was 0.129 ± 0.010 mm, while that from symptomatic plaques was 0.035 ± 0.003 mm (p = 0.0001). The area of the fibrous cap was not significantly different (p > 0.05) between symptomatic (0.027 ± 0.002 mnrf)and asymptomatic plaques (0.034 ± 0.003 mrrf). The area of the macrophage colonization, as defined by positive immunoreactivity to CD68, in fibrous caps of symptomatic plaques (0.023 ± 0.001 mm^) was significantly greater (p <0.001) as compared to that in asymptomatic plaques (0.016 + 0.002 mm^). The apoptotic nuclei in the fibrous cap of the plaques from asymptomatic plaques were 9.92 ± 0.55% and in the symptomatic plaques were 23.88 ± 0.80% (p = 0.0001). The TUNEL and Annexin V labeling showed symptomatic plaque VSMCs had five times greater apoptosis than asymptomatic plaque VSMCs. This was supported by increased expression of cell survival protein, bcl-xL, and cyclin D1 in the fibrous cap of the plaques from asymptomatic patients that that in symptomatic patients. The VSMCs from asymptomatic plaques showed higher proliferation with and without serum stimulation than symptomatic plaque VSMCs. Activity of caspase 9 and caspase 3 was significantly increased in symptomatic plaque VSMCs as compared to asymptomatic plaque VSMCs. The cel) cycle analysis revealed that VSMCs from symptomatic plaques were arrested in the S-phase while the VSMCs from asymptomatic plaques proceeded through the cell cycle. These findings indicate that there is a significant diference in the thickness of fibrous cap, area of the necrotic core, inflammation and apoptosis around the fibrous cap between plaques from symptomatic and asymptomatic patients with carotid stenosis. In symptomatic plaques, decreased proliferation and increased apoptosis of VSMCs, may result in plaque rupture leading to the development of symptoms, are associated with clinical ischemic events. Further investigation of intra-plaque events, which condition VSMCs to undergo apoptosis, is warranted to identify vulnerable plaques. Additionally, imaging techniques that precisely resolve the position of the necrotic core and evidence of inflammatory reactions within carotid plaques should help identify high-risk stenoses before disruption and symptomatic carotid disease.
Networked Digital Library of Theses and Dissertations