Effects Of Alcohol On Promoting Apoptosis Of Vascular Endothelial Cells In Rats

Objective: To study the effects of alcohol on promoting apoptosis of vascular endothelial cells (EC) and its mechanisms, and evaluate the relationship between excessive intake of alcohol and atherosclerosis (AS).Methods: The rats were divided into four groups according to the time of using gavage of alcohol: (1)Control group: Rats were administrated distilled water by gavage at a dose of 1ml three times daily; (2)Experimental group I: Rats were administrated alcohol (concentration: 40V/V) by gavage at a dose of 8g/(kg d) three times daily for four weeks; (3)Experimental group II: Rats were administr ated alcohol (concentration: 40V/V) by gavage at a dose of 8g/(kg d) three times daily for six weeks; (4)Experimental group III: Rats were administrated alcohol (concentration: 40V/V) by gavage at a dose of 8g/(kg d) three times daily for eight weeks.The serum was obtained from heart by puncture, and samples of the aortas were taken to prepare "en-face" specimens of aortic endothelium after fixing by perfusion with 4% Paraformaldehyde at the end of each expiration. The morphological characteristics of EC in each group were observed by Hematoxylin-Eosin (H-E) stain; the expression of intercellular adhesion molecule-1 (ICAM-1) and EC labeling-CD34 in each group were detected by immunohistochemistry; ratios of EC apoptosis in each group were detected by terminal deoxynucleotidy transferase-mediated dUTP-biotin nick end labeling(TUNEL) method. In addition, the morphological characteristics of EC in each group were observed by scanning electron microscopy (SEM), Maleic Dialdehyde (MDA) values and total antioxidation capability (T-AOC) in serum were detected in each group by Spectrophotometer. The results were analyzed by Statistic software-SPSS 11.0 system.Results: Abnormity aortic EC nucleus was not seen in the Control group, the nucleus was stained blue and the cytoplasm red by Hematoxylin-Eosin (HE) stain. However, all experimental groups showed that the affected cells were shrinks, the cytoplasm was intensely eosinophilic and the nucleus was small and dense or disappeared. The results of immunohistochemistry indicated that CD34 expressed in the cytoplasm in all control and experimental groups. ICAM-1 was negative in control group while all experimental groups were positive in the cytoplasm, and the expression showed significantly time-dependence. Apoptosis couldn't be observed in EC of control group by TUNEL method, and all experimental groups showed many apoptosis EC. The ratios of EC apoptosis were 5.56% (group I), 8.86% (group II) and 12.5% (group III), respectively, which showed time-dependence. There were significant differences (p<0.01) among group II, group III and group I. SEM showed that EC size of control group was uniform, the distribution was ordered; the cell membrane was integrity and lubricity, and normal microvillus and pseudopod could be observed, in the while, the distribution was ordered; the characteristics of apoptosis EC that disappear of structure of cell membrane was observed in all experimental groups such as microvillus and pseudopod; the cell membrane was undulate; in the meanwhile, the characteristics of injured EC were wrinkly and small, cellmembrane existed splits and apertures, and the microvillus were decreased, disappeared, stiff or broken and irregular distribution, and the cell gaps were wide.Detection of MDA values in serum indicated that the differences between experimental groups and control group were significant (p<0.01). The MDA values were 6.46±0.66nmol/ml (group I), 7.20 + 0.70nmol/ml (group II), 7.53 ±0.54nmol/ml (group III), respectively, which showed time-dependence, but the MDA values in control group was 4.68 0.15nmol/ml. Detection of T-AOC in serum indicated that the differences between experimental groups and control group were significant (p<0.0\). The T-AOC values were 15.28 ± 1.40U/ml(group I), 9.73 + l.OOU/ml (group II), 4.74±2.36U/ml (group III), respectively, which showed time-dependence, however the T-AOC values in control group was 20.44 + 2.56U/ml. Conclusions:1. Excessive intake of alcohol could induce imbalance of oxidation-antioxidation system.2. Imbalance of oxidation-antioxidation system increased oxygen free radical (OFR); the OFR reaction was pathologically exacerbated, in the meanwhile, OFR could induce endothelial cell apoptosis.3. EC apoptosis could induce AS.4. Excessive intake of alcohol could induce EC apoptosis, AS.