Changes of the Endothelium and Extracellular Matrix in Thoracic Aortic Aneurysm Revealed by Scanning Electron Microscopic Investigations Are there Structural Parallels to Aortic Valve Degeneration?

Objective: Thoracic aneurysms occur most frequently with 60% in ascending aorta and arch, und in 40% the descending aorta, although the etiological factors are different for individual segments. While, depending on localization, factors such as atherosclerosis, bicuspid aortic valve, familial thoracic aortic aneurysm syndrome, Turner syndrome, Marfan syndrome, aortic arteritis in Takayasu’s arteritis, trauma and dissection lead to the formation of thoracic aortic aneurysms, the structural changes of the basement membrane and the exact causes of the mechanism of thoracic aortic aneurysm are not known. The aim of this study was to investigate the pathophysiological origin of thoracic aortic aneurysms from a morphological point of view. Patients and methods: For this study, between 2007 and 2017, 12 patients with thoracic aortic aneurysms with contraindications to interventional therapy underwent open surgery. All patients underwent multi- detector row CT with three-dimensional (3D) reformation in 3 mm slices and transesophageal echocardiography, Tissue samples were taken from descending aorta of two female (average age, 72 .32 years) and 10 male patients (63.12 years), and were fixed for 6 hours in a solution containing 2.5% glutaraldehyde and 0.2 mMol cacodylate. The samples were finally visualized using the digital scanning microscope. Results: In addition to partial desquamation of the endothelium, severe alterations of the collagen fibers and basal membrane and repair attempts in the form of endothelial islets and deep tears in the fibers, cancellation and destruction of the helical structure of collagen fibers are visible. Conclusion: In summary, in thoracic aortic aneurysms we found severe structural changes of collagen fibres, endothelium and basement membrane, which occur in a similar form in acquired aortic valve degeneration.


Introduction
Thoracic aortic aneurysms (TAAs) are potentially devastating, and due to their asymptomatic behavior, pose a serious health risk characterized by the lack of medical treatment options and high rates of surgical morbidity and mortality [1,2]. Independent of the inciting stimuli (biochemical/mechanical), TAA development proceeds by a multifactorial process influenced by both cellular and extracellular mechanisms, resulting in alterations of the structure and composition of the vascular extracellular matrix (ECM) [3,4].
The molecular mechanisms leading to ascending thoracic aortic aneurysms (ATAAs) remain unknown. Diseases affecting the ascending aorta, such as these thoracic aortic aneurysms and type I and II dissections, are primarily associated with medial necrosis, which is characterized by fragmentation and loss of elastic fibers, 276 loss of muscle cells and interstitial collections of collagenous fibers and basophilic ground substance [5,6]. The aim of this study was the evaluation of the changes of endocardium and extracellular matrix in thoracic aortic aneurysms.  [7]. Table 1 presents summarized data of the basic patient characteristics.

Scanning Electron Microscopy
In order to reveal the morphology of the tissue sample, scanning electron microscopy was performed in all tissue samples.
Specimens from the descending aortic were fixed for 6 hours in a solution containing 2.5% glutaraldehyde and 0.2 mMol cacodylate.
Afterwards, samples were dehydrated in a series of increasing concentrations of alcohol. After critical point drying, all samples were sputtered with gold-palladium. Samples were visualized using the digital scanning microscope (Zeiss DSM 960, Germany).

Discussion
Thoracic aortic aneurysms and dissections constitue a critical condition that often goes undiagnosed with fatal consequences.
Besides, various inherited forms discussed, genetic factors are discussed, genetic factors being important in the development of thoracic aneurysms [8,9]. Familial aortic aneurysm (FAA) has been described in such conditions as the Marfan and Ehlers-Danols syndrome type IV, which are due to defects in the fibrillin-1 and III procollagen genes respectively [10][11][12][13][14]. In this group of a novel locus at chromosome 11q23.3-q24 was detected, a critical step toward elucidating a gene defect responsible for aortic dilatation [15]. Hasham et al. Determined one locus, mapped to 5q13-14, which will enhance the chances to determine persons at risk for aortic aneurysm in non-familial aortic aneurysm diseases [5].
Thoracic aortic aneurysms and infrarenal aortic aneurysms exhibit distinct patterns of gene expression relative to the normal aorta from the same sites [14]. Keramat mapped in familial TAA, a single

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locus on chromosome 15q21 with the peak lod score of 6.3 at the fibrillin-1 gene [16]. The molecular changes of the extracellular matrix have been the subject of many studies. In this study, we have investigated the morphological structural changes of the endothelium and the extracellular matrix in aneurysmal changes in the descending aorta. The endothelial cells were desquamated, for the most part, the remaining once grouped into smaller islands.
These showed a transformation to villi-like structures on their surface, which showed morphological similarities with to cells of the duodenum. These changes are associated with different pathophysiological dysfunction of the endothelium. The Nogo-B is highly expressed in endothelial cells following downregulated induction by lysophosphatidylcholine, which contributed to atherossclerotic lesions [17]. In TAA, the Nogo-B mRNA and protein expression are downregulated, which correlate to the formation of aneurysm [10]. The ageassociated changes in endothelial nitric oxide synthase expression have not been definitively linked to pathophysiology of aortic aneurysm [18]. As a sign of endothelial inflammatory response the increased VCAM-1 expression has been detected in human aorta with atheromatous changes [19]. In descending thoracic aortic anuerysms as a sign of the transmural inflammation, IFN-gamma, IP-10 and mig chemikines are correlated with vascular remodeling and expansion of aneurysms [20]. Smooth muscle cells can exert a tiggr function in the development of inflammatory processes involved in transmural inflammation by increased secretion of GM-CSF [21][22][23]. The bulk of the extracellular matrix was destroyed, the triple helical structure of collagen fibers removed.
In addition, some pronounced hypertrophy was present in collagen fibers. The basal lamina is a crucial component of mechanical and functional blood vessels, constituting a sensor of extracellular microenvironment for endothelial cells and pericytes. In segments of thoracic aortic aneurysms the collagen I, laminin alpha 2 chain and fibonectin amount decreased [24,25]. Consistently with the common macroscopic observation that ascending aorta dilatations tend to expand asymmetrically, with prevalent involvement of vessel convexity and relative loss of the concavity, collagen type IV is more evident in the concavity and transcin in the convexity [26].
However, we could not detect any differences in the destruction of the extracellular matrix in the convex or concave aortic arcus.
The synthesis of other chains of collagen, discussed already in the degeneration of aortic and mitral valves is a sign of overloading of the collagen fibers as primary stimuli [27]. Toumpoulis et al.
demonstrate that increased collagen alpha 1 (XI) and V collagen mRNA and protein levels are linearly correlated with the size of the aneurysm and provide a potential mechanism for generation and progression of aneurysmal enlargement [28]. The hypothesis of an increased conversion of the basement membrane by a variant of the synthesized collagen fibers is underlined by the demonstration of increased expression of metalloproteinases ADAM10, ADAM17, matrix metalloproteinases MMP-2 and 9 [29][30][31][32]. A similar finding with increased MMP-8 in acute thoracic aortic dissection was recently reported [33]. Surprisingly, we detecded in the examined aortic, endothelial defects in the form of craters which we have described in a similar form in the degeneration of aortic valves.
The functional significance of this change is completely unknown.

Conflict of Interest
I hereby declare that there were no financial or other interests in the execution and evaluation of this work.