Abstract
BackgroundAortic diseases are diverse and involve a multiplicity of biological systems in the vascular wall. Aortic dissection, which is usually preceded by aortic aneurysm, is a leading cause of morbidity and mortality in modern societies. Although the endothelium is now known to play an important role in vascular diseases, its contribution to aneurysmal aortic lesions remains largely unknown. The aim of this study was to define a reliable methodology for the isolation of aortic intimal and adventitial endothelial cells in order to throw light on issues relevant to endothelial cell biology in aneurysmal diseases.Methodology/Principal FindingsWe set up protocols to isolate endothelial cells from both the intima and the adventitia of human aneurysmal aortic vessel segments. Throughout the procedure, analysis of cell morphology and endothelial markers allowed us to select an endothelial fraction which after two rounds of expansion yielded a population of >90% pure endothelial cells. These cells have the features and functionalities of freshly isolated cells and can be used for biochemical studies. The technique was successfully used for aortic vessel segments of 20 patients and 3 healthy donors.Conclusions/SignificanceThis simple and highly reproducible method allows the simultaneous preparation of reasonably pure primary cultures of intimal and adventitial human endothelial cells, thus providing a reliable source for investigating their biology and involvement in both thoracic aneurysms and other aortic diseases.
Highlights
Human thoracic aneurysm of the ascending aorta (TAA) is a chronic, asymptomatic and potentially lethal disease
The wall of the aorta consists of three well-defined layers [1]: i) the intima, which is the innermost layer composed of the endothelium whose cells rest and adhere onto a basement membrane; ii) the media which consists of ~70 layers of alternating elastic laminae [2] and embedded smooth muscle cells (SMCs), collagen fibers, proteoglycans (PG) and glycosaminoglycans (GAG); and iii) the tunica adventitia, the outermost and most complex layer, which is composed of a variety of cells and connective tissue where collagen fibers stabilize and anchor the aorta to adjacent tissues [1]
Recent studies point to the endothelium as being important player in the pathogenesis of TAAs, most of them were conducted with animal models
Summary
Human thoracic aneurysm of the ascending aorta (TAA) is a chronic, asymptomatic and potentially lethal disease. It is characterized by dilatation of the aortic wall, which can progress to vessel dissection and rupture. There is an urgent need to understand the mechanisms underlying the pathogenesis of the disease in order to develop alternative therapies for treating the causes of human TAAs. Clinically, arterial enlargement is attributed to unbearable wall tension superimposed on defective aortic wall integrity and impaired aortic repair mechanisms. The endothelium is known to play an important role in vascular diseases, its contribution to aneurysmal aortic lesions remains largely unknown. The aim of this study was to define a reliable methodology for the isolation of aortic intimal and adventitial endothelial cells in order to throw light on issues relevant to endothelial cell biology in aneurysmal diseases
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