叶酸和辛伐他汀对大鼠动脉粥样硬化平滑肌细胞增殖的影响

Effects of folic acid and simvastatin treatment on proliferation of smooth muscle cell in atherosclerosis of rat

目的研究叶酸和辛伐他汀对高同型半胱氨酸血症(HHcy)引起的大鼠动脉粥样硬化(AS)主动脉平滑肌细胞增殖治疗效果的影响。方法健康8周龄SD雄性大鼠36只,随机分为对照组、高蛋氨酸组、叶酸组和辛伐他汀组,对照组6只,其余各组均为10只。对照组给予普通饲料喂养;高蛋氨酸组在普通饲料喂养的基础上加质量分数2%的蛋氨酸;叶酸组以高蛋氨酸组饲料喂养4周后,叶酸按每天3 mg.kg-1剂量灌胃,同时继续喂食高蛋氨酸组饲料;辛伐他汀组以高蛋氨酸组饲料喂养4周后,辛伐他汀按每天5 mg.kg-1剂量灌胃,同时继续喂食高蛋氨酸组饲料。12周后,采用高效液相色谱法测定血浆同型半胱氨酸(Hcy)浓度,另取其胸主动脉用于苏木素-伊红(HE)染色和油红O染色。采用免疫组织化学染色法检测主动脉平滑肌细胞中增殖细胞核抗原(PCNA)蛋白的表达情况。结果高蛋氨酸组血浆Hcy含量明显高于对照组、辛伐他汀组和叶酸组(P<0.01);叶酸组血浆Hcy值明显低于辛伐他汀组(P<0.05)。HE染色结果显示对照组血管内皮细胞连续规整,内弹力膜完整,中膜平滑肌细胞排列整齐;高蛋氨酸组局部可见内皮细胞坏死脱落,中膜平滑肌细胞胞核固缩且排列紊乱,内膜下可见泡沫细胞;辛伐他汀组内膜和平滑肌细胞变化介于对照组和高蛋氨酸组之间,动脉壁细胞明显较叶酸组细胞排列紊乱;叶酸组和对照组动脉壁比较无差别。油红O染色结果显示对照组、叶酸组和辛伐他汀组动脉壁经油红O染色后无差别,不着色;高蛋氨酸组油红O染色局部阳性,主要位于内膜下。高蛋氨酸组PCNA蛋白阳性表达强度高于对照组、辛伐他汀组和叶酸组(P<0.01);叶酸组PCNA蛋白阳性表达强度高于辛伐他汀组(P<0.01)。结论高蛋氨酸饮食可以引起大鼠HHcy形成,进而引起动脉平滑肌细胞的增殖,促使AS发生。辛伐他汀及叶酸均可以降低Hcy浓度,减少Hcy对动脉壁的作用,抑制动脉平滑肌细胞的增殖,防止AS的发生,但在抑制平滑肌细胞增殖的效果方面辛伐他汀要优于叶酸。

Objective To investigate effects of folic acid and simvastatin treatment on proliferation of smooth muscle cell in atherosclerosis（AS） induced by hyperhomocysteinemia（HHcy） in rat.Methods Thirty-six male SD rats,eight weeks of age and healthy,were randomly divided into control group（n=6）,high methionine group（n=10）,folic acid group（n=10） and simvastatin group（n=10）.Normal food was fed in control group;Normal food and 2%（mass fraction） methionine were fed in high methionine group;After the feed in high methionine group were fed for four weeks,folic acid was lavaged in 3 mg·kg-1 dose per day,at the same time,the feed in high methionine group were fed continue in folic acid group;After the feed in high methionine group were fed for four weeks,simvastatin was lavaged in 5 mg·kg-1 dose per day,at the same time,the feed in high methionine group were fed continue in folic acid group.After twelve weeks,the plasma homocysteinemia（Hcy） concentration was detected with high performance liquid chromatography.And hematoxylin-eosin（HE） staining and oil red O staining were used in the thoracic aorta.The expressions of proliferating cell nuclear antigen（PCNA） protein in aortic smooth muscle cells were detected with immunohistochemistry.Results The plasma Hcy concentration in high methionine group were significantly higher than that in control group,simvastatin group and folic acid group（P0.01）;The plasma Hcy concentration in folic acid group was significantly lower than that in simvastatin group（P0.05）.HE staining showed vascular endothelial cells regular,internal elastic membrane intact,medial smooth muscle cells in order in control group.The local endothelial cell necrosis,the smooth muscle cell nucleus pyknosis and disorganized,foam cells can be seen in under intima in high methionine group.Changes of intima and smooth muscle cells were between the control group and high methionine groups in simvastatin group,aterial wall cells were more disordered than in folic acid group significantly.There was no significant difference in the arterial wall between folic acid group and control group.There was no significant difference in oil red O staining between folic acid group and simvastatin group,no stained.Oil red O staining showed the local location was positive,under the intima mainly in high methionine group.The positive expression of PCNA protein in high methionine group was significantly higher than control group,simvastatin group and folic acid group（P0.01）.The positive expression of PCNA protein in folic acid group was higher than that in simvastatin group（P0.01）.Conclusion High methionine diet can cause HHcy formation in rats,father more occurred aortic smooth cells proliferation and AS.Simvastatin and folic acid can reduce the concentration of Hcy,reduce the role of Hcy on the artery wall,inhibit aortic smooth cells proliferation,prevent the occurrence of AS.But simvastatin treatment is superior to folic acid in the effect of inhibit smooth muscle cell proliferation.