- Department of Neurosurgery, American Hospital, Istanbul, Turkey
- Department of Biochemistry, Marmara University, Faculty of Medicine, Istanbul, Turkey
- Department of Radiology, Sisli Etfal State Hospital, Istanbul, Turkey
- Department of Neurology, Boston University, School of Medicine, Boston, MA, USA
- Department of Neurosurgery, Pendik State Hospital, Istanbul, Turkey
- Department of Neurosurgery, Koc University School of Medicine, Istanbul, Turkey
Department of Neurosurgery, Koc University School of Medicine, Istanbul, Turkey
DOI:10.4103/2152-7806.77600Copyright: © 2011 Sasani M This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
How to cite this article: Sasani M, Yazgan B, Celebi I, Aytan N, Catalgol B, Oktenoglu T, Kaner T, Ozer NK, Ozer AF. Hypercholesterolemia increases vasospasm resulting from basilar artery subarachnoid hemorrhage in rabbits which is attenuated by Vitamin E. Surg Neurol Int 14-Mar-2011;2:29
How to cite this URL: Sasani M, Yazgan B, Celebi I, Aytan N, Catalgol B, Oktenoglu T, Kaner T, Ozer NK, Ozer AF. Hypercholesterolemia increases vasospasm resulting from basilar artery subarachnoid hemorrhage in rabbits which is attenuated by Vitamin E. Surg Neurol Int 14-Mar-2011;2:29. Available from: http://sni.wpengine.com/surgicalint_articles/hypercholesterolemia-increases-vasospasm-resulting-from-basilar-artery-subarachnoid-hemorrhage-in-rabbits-which-is-attenuated-by-vitamin-e/
Background:Aneurysm rupture results in subarachnoid hemorrhage (SAH) with subsequent vasospasm in the cerebral and cerebellar major arteries. In recent years, there has been increasing evidence that hypercholesterolemia plays a role in the pathology of SAH. It is known that hypercholesterolemia is one of the major risk factors for the development of atherosclerosis. Among the factors that have been found to retard the development of atherosclerosis is the intake of a sufficient amount of Vitamin E. An inverse association between serum Vitamin E and coronary heart disease mortality has been demonstrated in epidemiologic studies. Therefore, we tested, in an established model of enhanced cholesterol feed in rabbits, the effects of hypercholesterolemia on vasospasm after SAH by using computed tomography (CT) angiograms of the rabbit basilar artery; in addition, we tested the effects of Vitamin E on these conditions, which have not been studied up to now.
Methods:In this study rabbits were divided into 3 major groups: control, cholesterol fed, and cholesterol + Vitamin E fed. Hypercholesterolemia was induced by a 2% cholesterol-containing diet. Three rabbit groups were fed rabbit diet; one group was fed a diet that also contained 2% cholesterol and another group was fed a diet containing 2% cholesterol and they received i.m. injections of 50 mg/kg of Vitamin E. After 8 weeks, SAH was induced by the double-hemorrhage method and distilled water was injected into cisterna magna. Blood was taken to measure serum cholesterol and Vitamin E levels. Basilar artery samples were taken for microscopic examination. CT angiography and measurement of basilar artery diameter were performed at days 0 and 3 after SAH.
Results:Two percent cholesterol diet supplementation for 8 weeks resulted in a significant increase in serum cholesterol levels. Light microscopic analysis of basilar artery of hypercholesterolemic rabbits showed disturbances in the subendothelial and medial layers, degeneration of elastic fibers in the medial layer from endothelial cell desquamation, and a reduction of waves in the endothelial layer. However, the cholesterol + Vitamin E group did not exhibit these changes. The mean diameter of the basilar artery after SAH induction in the cholesterol-treated group was decreased 47% compared with the mean diameter of the control group. This value was less affected in cholesterol + Vitamin E-treated rabbits, which decreased 18% compared with the mean diameter of the control group.
Conclusions:Hypercholesterolemia-related changes in the basilar artery aggravate vasospasm after SAH. Adding Vitamin E to cholesterol-treated rabbits decreased the degree of vasospasm following SAH in the rabbit basilar artery SAH model. We suggest that Vitamin E supplements and a low cholesterol diet may potentially diminish SAH complicated by vasospasm in high-risk patients.
Keywords: Aneurysm, atherosclerosis, hypercholesterolemia, subarachnoid hemorrhage, vasospasm, Vitamin E
Aneurysm rupture results in subarachnoid hemorrhage (SAH) with subsequent vasospasm in the cerebral and cerebellar major arteries. Vasospasm occurring prior to, during, and after surgery is one of the most important factors affecting the functional prognosis of patients.[
Most patients with SAH are hypercholesterolemic and atherosclerotic; therefore, it is important to determine the effects of cerebral vessel atherosclerosis resulting from hypercholesteremia on vasospasm after SAH. It is widely accepted that hypercholesterolemia, which is found with high low-density lipoprotein (LDL), plays a pivotal role in the progression of atherosclerosis.[
The aim of the current study was to investigate the effects of hypercholesterolemia on vasospasm, resulting from basilar artery SAH in rabbit model and to compare the effects of Vitamin E on vasospasm in hypercholesterolemia-related changes in the vessels after SAH using CT angiograms of the rabbit basilar artery.
Sixteen male albino rabbits (1–2 months old) were randomized to 3 groups. All rabbits were fed 100 g rabbit diet per day. Cholesterol was added to the diet as diethyl ether solution. The control diet was treated with the same amount of pure solvent. All diets were dried of the solvent before use. The concentrations of cholesterol and Vitamin E were based on previous reports.[
Determination of serum cholesterol and Vitamin E levels
Serum cholesterol levels were determined using an automated (Hitachi Modular P800) enzymatic technique (Roche, Boehringer Ingelheim). Vitamin E (α-tocopherol) levels were determined by extracting serum; concentrations of α-tocopherol were obtained by reverse phase high-pressure liquid chromatography and by using a C-18 Bondopak column and a UV detector at 294 nm.[
CT angiography and measurement of basilar artery diameter
A high-speed resolution (64 slice) CT was used to identify basilar artery vasospasm (The Phillips Brilliance CT 64-slice scanner, Phillips Company, Netherlands).
Axial sections of the arterial-phase CT angiogram of the cerebral arteries (in particular the basilar artery) were obtained using an injection of 623–769 mg/mL/kg Iopramid in the central ear vein. Parameters of the CT acquisition were 120 kV, 150 mAs, 64 × 0.625 detector collimation; pitch of 1.15, 0.5 s gantry rotation time; 512 × 512 matrix; and 20 cm field of view. Contrast enhancement was provided by the intravenous (central ear vein) administration at the rate of 3mL/s. The luminal diameter of the basilar artery was measured on the Phillips EBW workstation CT scanner using magnification and the appropriate window setting. Axial, coronal, and three-dimensional cranial CT angiograms were obtained after reconstruction of the CT slices [
The luminal diameters of the basilar artery were measured by a single observer. The CT angiograms were converted on a computer. Using public domain imaging software, the diameters of the basilar artery were measured on 3 randomly selected points along each artery above the bifurcation of the vertebral arteries. We used the average of the 3 diameters to calculate the final diameter. The luminal relative diameter was used to estimate arterial narrowing. To produce comparable data, the points were the same for all animals.
Preparation of basilar artery for histopathologic analyses
Basilar arteries from all groups were fixed for histopathology in 10% buffered paraformaldehyde for a minimum of 24 h. Tissue samples were prepared in an autotechnicon and embedded in paraffin. The specimens were sectioned (5 μm) with a microtome and deparaffinated 3 times with xylene in a 60°C incubator. The tissue samples were dehydrated with alcohol, washed with water, and stained with Elastica van Gieson. The morphometric characteristics of the tissue samples were evaluated under a light microscope (100× magnification).
All data are expressed as mean ± standard deviation (SD). Data were analyzed by one-way ANOVA test using the “Graph Pad Prism 5” statistical program, and the individual comparisons of groups were obtained using Bonferroni's multiple comparison test. P values of less than 0.05 were selected as the levels of significance.
Analyses of serum cholesterol and Vitamin E
Blood analysis of the animals was performed after 8 weeks of treatment, just before SAH induction. Cholesterol and Vitamin E serum concentrations of the 3 animal groups are shown in
Histological analyses of basilar artery
Rabbit basilar arteries from the 3 groups were examined by light microscopy. Rabbits in control group, which were fed a pure diet, exhibited normal arterial wall layers [
CT angiographic evaluation of basilar artery vasospasm
CT angiography was used to measure the basilar artery diameter before and after SAH in all rabbits.
The average diameter of the atherosclerotic basilar artery after vasospasm induction in the control, cholesterol, and cholesterol + Vitamin E groups (was 1.20 ± 0.01, 0.66 ± 0.02, and 0.95 ± 0.03, respectively. These values were compared by ANOVA test and found to be significantly different (P < 0.05). The mean diameter of the basilar artery in the cholesterol-treated groups was decreased approximately 47% compared to the mean diameter of the control group. This value was less affected in the cholesterol + Vitamin E-treated rabbits, which was decreased 18% compared with the mean diameter of the control group. The mean diameter of the basilar artery after SAH induction decreased in the following order: control group > cholesterol + Vitamin E group > cholesterol group [
The development of atherosclerosis is a multifactorial process in which elevated plasma cholesterol levels play a major role; however, most patients with SAH have atherosclerotic vessels. Numerous studies have shown the involvement of the oxidative processes in the pathogenesis of atherosclerosis and the protective effect of Vitamin E against atherosclerosis.[
Rabbit basilar arteries from the 3 groups were examined by light microscopy. Rabbits in cholesterol group developed atherosclerosis similar lesions in the basilar artery, and endothelial cell integrity was impaired relative to the control group. They showed disturbances in the subendothelial and medial layers, degeneration of elastic fibers in the medial layer from endothelial cell desquamation, and reduced waves in the endothelial layer. However, the cholesterol + Vitamin E group did not exhibit these changes.
In fact, the atherosclerotic plaque could not be demonstrated in histological examination. Elastica van Gieson stain was used to demonstrate histopathologic analyses. However, histological analysis of the basilar artery after acute SAH using Elantica Van Gieson staining might be more effective for showing degeneration of the tunica muscularis. Nevertheless, initial changes in the subendothelial degeneration of elastic fibers and a reduction of waves in the endothelial layer were observed.
Our results are the first to suggest the potential of Vitamin E as a treatment for vasospasm in hypercholesterolemia-related changes in the vessels after SAH. The present study used CT angiography to measure changes in basilar artery diameter after SAH, while similar studies have used conventional angiography and/or solely histological examination to determine experimental basilar artery vasospasm after SAH.[
We could study only acute stage vasospasm; studies of chronic vasospasm would require a more prolonged study. Cerebral vasospasm after subarachnoid hemorrhage occurs on different time scales among species. Angiographic vasospasm appears 2 days after injecting autologous blood in the rabbit and after about 4-14 days in humans.[
Early or late period vasospasm following SAH is known. Although various spasmogens and their origins have been studied, the pathogenesis of vasospasm following SAH remains unclear.[
In conclusion, hypercholesterolemia increases vasospasm resulting in basilar artery subarachnoid hemorrhage; however, we found that supplementation with Vitamin E decreased the degree of vasospasm in hypercholesterolemic animals following SAH in the rabbit basilar artery SAH model. For the first time, we suggest that Vitamin E supplements and a low cholesterol diet may potentially diminish SAH complicated by vasospasm in hypercholesterolemia patients who are at a high risk.
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