| BackgroundAtherosclerosis is the pathological basis of cardiovascular and cerebrovascular diseases. Especially to coronary heart disease, atherosclerosis is the key. Atherosclerosis can decrease the function of vasomotion, narrow and block the vascular lumens, form thrombsis, effect the function of the heart and all the circulation system, thereby cause to occur kinds of acute cardiovascular incident. Therefore, the investigation on the mechanism and the treatment of atherosclerosis is attracting more and more attention. For a long time, AS is considered as a disease of aging and degeneration. However, the researchers recently find that AS is actually a kind of chronic inflammation ,which mediating by many cytokine and participating with vascular endothelial cells, smooth muscle cells , white blood cells and others, in the intimae of large and medial arteries which functions and structures are damaged by various factors. Grape seed proanthocyanidin extract (GSPE) is a sort of polyphenols extracted from grape seeds and it has potent antioxidant activity. Tong xinluo is a traditional Chinese medicine compound preparation, including many natural medicinal materials such as Ginseng, Leech, Scorpion, Ground Beetle, Red Peony Root, Borneol etc. It can promote blood circulation to remove meridian obstruction and benefit vital energy. In this research, we fed the rabbits high cholesterol diet, and observed the different effect of anti-atherosclerosis by GSPE, by Tong xinluo and by GSPE plus Tong xinluo in order to approach theanti-atherosclerosis mechanism of GSPE and Tong xinluo, to investigate the anti-atherosclerosis effect of combination, and to offer the clinical therapy experiment data. ObjectiveTo investigate the antiatherogenic effect and the possible mechanism of GSPE, Tong xinluo and GSPE plus Tong xinluo through the animal model of experimental atherosclerosis:(1) To observe the effects of GSPE, Tong xinluo and GSPE plus Tong xinluo on the serum lipid profile in atherosclerosis rabbits;(2) To measure the effects of GSPE, Tong xinluo and GSPE plus Tong xinluo on the serum lipid peroxidation in atherosclerosis rabbits;(3) To measure the effects of GSPE, Tong xinluo and GSPE plus Tong xinluo on the serum nitric oxide in atherosclerosis rabbits;(4) To observe the effects of GSPE, Tong xinluo and GSPE plus Tong xinluo on the serum interleukin 6 in atherosclerosis rabbits;(5) To measure the effects of GSPE, Tong xinluo and GSPE plus Tong xinluo on the serum vascular cell adhension molecule in atherosclerosis rabbits;(6) To confirm the effects of GSPE, Tong xinluo and GSPE plus Tong xinluo on the anti-atherosclerosis by observing the rabbits' aorta, heart and brain tissues through optical microscope, electron microscope separately and Immunohistochemicalchemistry.(7) To investigate whether there is synergism on atherosclerosis prevention between GSPE and Tong xinluo.Methods(1) Animals and diets: 24 male New Zealand rabbits were randomized into four groups: cholesterol group (control), GSPE group (a), Tong xinluo group (b), GSPE plus Tong xinluo group (ab), respectively 6 rabbits in each group. They were individually housed in metal cages in an air-conditioned room. Throughout the experimental period, they were given restricted amounts of food (100-120g/head per day) of each diet,without limit of drinking. The control group was fed with standard diet containing 1%cholesterol. The group was fed with standard diet containing 1%cholesterol plus 1%GSPE. The group b was fed with standard diet containing 1%cholesterol plus Tong xinluo (1g/kg.d). The group ab was fed with standard diet containing 1%cholesterol plus 1%GSPE plus Tong xinluo (1g/kg.d).(2) Sample taking: Blood samples were drawn from ear middle arteries of rabbits just before the experiment at the forth, the eighth and the twelfth weekend of the experiment. All the rabbits were fasted for at least eight hours before the blood was drawn. All the rabbits were sacrificed at the twelfth weekend and aorta, heart,and brain of the rabbits were excised for the morphological observation and immunohistochemical examination..(3) Measurements: Serum TC, LDL-C, TG, and HDL-C were measured enzymatically by automatic facilities. Serum MDA was measured as thiobarbituric-acid-reactive substances (TBARS) by fluorometric assay. Serum NO was measured by nitrate reductase technique. Serum IL-6 and VCAM-1 were determined using commercially available enzyme linked immunosorbent assay kits. Aorta, heart, and brain samples were observed through optical microsope and electron microcope separately. Immunohistochemical stain of CD68, a-actin and VCAM-1 were used to determine the changes of aortal intima.(4) Stastical analysis: Results were expressed as means±SD. One-way-anova of SPSS 10.0 soft ware was used to evaluate the differences between groups.Results(1) Compared with cholesterol group , the serum lipid profile did not change appreciably in the GSPE group; while the serum MDA level was declined evidently (8th to 12th weekend, P<0.05) , the serum NO level was increased definitely(4th to 12th weekend, P<0.001), the serum VCAM-1 and the serum IL-6 level were decreased remarkably (VCAM-1,4th to 12thweekend, P<0.001; IL-6, 4th weekend, P<0.05,8th to 12th weekend, P<0.001) in the GSPE group. And GSPE relieved the extent and severity of atherosclerosis, and strengthened the stability of the atherosclerotic plaque.(2) Compared with cholesterol group , the serum lipid profile did not change appreciably in the Tong xinluo group; while the serum MDA level was declined evidently (12th weekend, P<0.05) , the serum NO level was increased definitely(4th to 12th weekend, P<0.001), the serum VCAM-1 and the serum IL-6 level were decreased remarkably (VCAM-1,4th weekend, P<0.05,8th to 12th,P<0.001; IL-6, 8th to 12th weekend, P<0.001) in the Tong xinluo group. And Tong xinluo relieved the extent and severity of atherosclerosis, and strengthened the stability of the atherosclerotic plaque.(3) Compared with cholesterol group , the serum lipid profile did not change appreciably in the GSPE plus Tong xinluo group; while the serum MDA level was declined evidently (4th weekend, P<0.05,8th to 12th weekend, P<0.001) , the serum NO level was increased definitely(4th to 12th weekend, P<0.001), the serum VCAM-1 and the serum IL-6 level were decreased remarkably (VCAM-1,4th to 12th,P<0.001; IL-6, 4th weekend, P<0.01,8th to 12th weekend, P<0.001) in the GSPE plus Tong xinluo group. And GSPE combining with Tong xinluo relieved the extent and severity of atherosclerosis, and strengthened the stability of the atherosclerotic plaque.(4) From our experimental results, we found that there were synergistic effects in decreasing the serum MDA level, VCAM-1 level and descending the serum IL-6 level between GSPE and Tong xinluo.ConclusionsBoth GSPE and Tong xinluo have the efficacies of anti-atherosclerosis, and the mechanism seems to be concerned with the activities that they can inhibit the oxidation of lipoproteins, protect the functions of blood vessel endothelium, and prevent inflammation andblood white cells adhension at the endothelium. Fourthermore, there is synergism on anti-atherosclerosis between GSPE and Tong xinluo. The combination of these two drugs has more effects than each one.
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