通心络对家兔动脉粥样硬化模型VCAM-1及PAI-1表达的影响

发布时间：2018-01-31 01:19

本文关键词： 动脉粥样硬化血管细胞黏附因子1 纤溶酶原激活剂抑制剂1 通心络　出处：《吉林大学》2011年硕士论文　论文类型：学位论文

【摘要】：背景与目的： AS已成为威胁人类健康,引起死亡的主要疾病。可导致全身多个靶器官损害,严重影响人民的生活质量。为从根本解决AS为人民带来的困扰,大量基础实验及临床观察致力于对AS发病机制及治疗方法的研究。本研究通过高脂饮食建立家兔AS模型,了解PAI-1和VCAM-1在AS家兔血清和病变局部的表达,观察通心络对PAI-1和VCAM-1的影响及其抗AS的作用机制。资料与方法：一、建立家兔AS模型,采用酶联免疫夹心法、免疫组化方法检测VCAM-1和PAI-1在正常饮食组和高脂饮食组家兔血清中及动脉血管壁的表达。 (一)动物模型的建立 1、实验动物和分组雄性大耳白兔24只。随机分为常规饮食组、高脂饮食组、通心络组,每组8只,饲养14周。 2、病理标本的制作 14周末处死动物,常规病理切片,H-E染色。 3、高脂血症的确立正常饮食组和高脂饮食组实验前(0周)血清中TC、TG、LDL无显著性差异(P0.01)；正常饮食组14周TC、TG、LDL与0周比较无显著性差异(P0.01)；高脂饮食组14周TC、TG、LDL明显增高,与0周比较有显著性差异(P0.01)。 (二)酶联免疫夹心法检测VCAM-1和PAI-1在正常饮食组和高脂饮食组血清中的表达 1、VCAM-1在正常饮食组和高脂饮食组血清中的表达正常饮食组的血清中VCAM-1水平在0周、14周无显著差别(P0.01)；高脂饮食组血清中VCAM-1水平14周较0周显著增高(P0.01)。 2. PAI-1在正常饮食组和高脂饮食组血清中的表达正常饮食组血清中PAI-1水平在0周、14周无显著差别(P0.01);高脂饮食组血清中PAI-1的水平14周较0周显著增高(P0.01)。 (三)免疫组化检测VCAM-1和PAI-1在正常饮食组和高脂饮食组主动脉壁的表达 1、VCAM-1在正常饮食组和高脂饮食组主动脉壁的阳性表达正常饮食组主动脉内膜和中膜仅见散在棕黄色颗粒状阳性染色物质,阳性染色百分比为2.92±0.31%。高脂饮食组增厚的动脉内膜及中膜内可见大量的棕黄色颗粒状阳性染色物质,阳性染色百分比为18.38±2.55%,明显高于正常饮食组(P0.01)。 2、PAI-1在正常饮食组和高脂饮食组主动脉壁的阳性表达正常饮食组主动脉内膜和中膜仅见散在棕黄色颗粒状阳性染色物质,阳性染色百分比为3.24±0.21%。高脂饮食组增厚的动脉内膜及中膜内可见大量的棕黄色颗粒状阳性染色物质,尤以内膜明显,阳性染色百分比为15.46±2.23%,明显高于正常饮食组(P0.01)。二、通心络的调脂作用实验前三组动物血清TC、TG、LDL无差异(P0.01),14周时正常饮食组TC、TG、LDL与0周比较无显著性差异(P0.01)；14周时高脂饮食组和通心络组TC、TG、LDL明显增高,明显高于正常饮食组(P0.01)；高脂饮食组和通心络组比较,14周时通心络组血清TC、TG、LDL较高脂饮食组明显下降(P0.01)。三、通心络对VCAM-1和PAI-1表达的影响 (一)通心络对血清VCAM-1和PAI-1表达的影响 1、通心络对血清VCAM-1表达的影响三组动物血清中VCAM-1水平在0周无显著差别(P0.01)；饲养14周时高脂饮食组和通心络组血清中VCAM-1水平较0周显著增高(P0.01),并高于正常饮食组(P0.01)；14周时通心络组血清中VCAM-1较高脂饮食组明显下降(P0.01)。 2、通心络对血清PAI-1表达的影响三组动物血清中PAI-1水平在0周无显著差别(P0.01)；饲养14周时高脂饮食组和通心络组血清中PAI-1水平较0周增高(P0.01),并高于正常饮食组(P0.01)；14周时通心络组血清中PAI-1较高脂饮食组明显下降(P0.01)。 (二)通心络对主动脉壁VCAM-1和PAI-1阳性染色百分比的影响 1、通心络对主动脉壁VCAM-1阳性染色百分比的影响高脂饮食组和通心络组增厚的动脉内膜及中膜内可见较多的棕黄色颗粒状阳性染色物质,表达量分别为18.38±2.55%和13.18±1.25%。通心络组的阳性表达百分比较高脂饮食组明显降低(P0.01)。 2、通心络对主动脉壁PAI-1阳性染色百分比的影响高脂饮食组和通心络组增厚的动脉内膜及中膜内可见较多的棕黄色颗粒状阳性染色物质,表达量分别为15.46±2.23%和9.45±1.09%。通心络组的阳性表达百分比较高脂饮食组明显降低(P0.01)。以上实验结果以平均数±标准差(X±S)表示,自身及组间对照采用t检验,P0.05有显著性差异。结果： 1.高脂饮食组和通心络组14周后血清TC、TG、LDL较实验前明显升高(P0.01),并且高于对照组(P0.01),通心络组血清TC、TG、LDL较高脂饮食组明显下降(P0.01)。 2.高脂饮食组和通心络组14周后血清PAI-1和VCAM-1的水平较0周显著增高(P0.01),并且高于对照组(P0.01),通心络组PAI-1和VCAM-1的水平明显低于高脂饮食组(P0.01)。 3.免疫组化分析显示：高脂饮食组增厚的动脉内膜及中膜内可见大量的棕黄色颗粒状阳性染色物质,部分融合成片,尤以内膜明显,通心络组阳性染色物质明显低于高脂饮食组,三组组间比较有显著性差异(P0.01)。结论： 1、AS家兔血清中VCAM-1和PAI-1的表达增高；家兔主动脉粥样硬化病变局部VCAM-1和PAI-1阳性表达百分比增高。VCAM-1和PAI-1的过度表达可能是AS发生机制之一。 2、通心络能够减少AS家兔VCAM-1和PAI-1在血清中及主动脉粥样硬化病变局部阳性表达百分比。抑制VCAM-1和PAI-1的表达可能是通心络抗AS机制之一。 3、通心络不但具有调脂的作用,并且具有减轻和抑制AS的作用。 4、检测血清中VCAM-1和PAI-1,对于早期诊断AS性疾病以及监测AS的发生、发展及转归有着重要的意义。
[Abstract]:Background and purpose:
AS has become a threat to human health, mainly caused by the disease can lead to death. More than one target organ damage, seriously affecting people's quality of life. In order to fundamentally solve the AS problems for people, a large number of basic experiments and clinical observation of AS is devoted to the study of pathogenesis and therapy. The study of rabbit AS model the high fat diet, understand PAI-1 and VCAM-1 expression in serum and AS local lesions in rabbits, observe the mechanism effect of Tongxinluo on PAI-1 and VCAM-1 and anti AS.
Information and methods:
First, the rabbit AS model was established. The expression of VCAM-1 and PAI-1 in serum and arterial wall of normal diet group and high-fat diet group were detected by enzyme-linked immunosorbent assay and immunohistochemistry.
(I) establishment of animal model
1, experimental animals and groups
24 male large white rabbits were randomly divided into regular diet group, high fat diet group, Tongxinluo group and 8 rats in each group for 14 weeks.
2, the preparation of pathological specimen
The animals were killed at the end of the 14 week, routine pathological section, H-E staining.
3, the establishment of hyperlipidemia
There was no significant difference in serum TC, TG and LDL between the normal diet group and the high-fat diet group before the experiment (0 weeks) (P0.01). There was no significant difference in TC, TG, LDL between the normal diet group and the 0 week group (P0.01). The 14 week TC, TG and LDL increased significantly in the high-fat diet group, which was significantly different from that in the 0 week (TC).
(two) the expression of VCAM-1 and PAI-1 in the serum of the normal diet group and the high fat diet group by enzyme linked immunosorbent assay
1, the expression of VCAM-1 in the serum of the normal diet group and the high fat diet group
The level of VCAM-1 in the serum of the normal diet group was 0 weeks, and there was no significant difference in the 14 weeks (P0.01), and the level of VCAM-1 in the serum of high fat diet group was significantly higher than that in the 0 week (P0.01).
2. PAI-1 expression in the serum of the normal diet group and the high fat diet group
There was no significant difference in the level of PAI-1 in the serum of the normal diet group for 0 weeks and 14 weeks (P0.01), and the level of PAI-1 in the serum of the high fat diet group was significantly higher than that in the 0 week (P0.01).
(three) immunohistochemical detection of the expression of VCAM-1 and PAI-1 in the aorta wall of the normal diet group and the high fat diet group
1, the positive expression of VCAM-1 in the aorta wall of the normal diet group and the high fat diet group
Normal diet group aortic intima and media were scattered in the yellow brown granular staining material, positive staining percentage was 2.92 + 0.31%. arterial intimal thickening of the high fat diet group and in the film can be seen in the large brown granular staining material, positive staining percentage was 18.38 + 2.55%, significantly higher than the normal diet group (P0.01).
2, the positive expression of PAI-1 in the aorta wall of the normal diet group and the high fat diet group
Normal diet group aortic intima and media were scattered in the yellow brown granular staining material, positive staining percentage was 3.24 + 0.21%. arterial intimal thickening of the high fat diet group and in the film can be seen in the large brown granular staining material, especially in endometrial. Positive staining percentage was 15.46 + 2.23%, significantly higher than the normal diet group (P0.01).
Two, the lipid-regulating effect of Tongxinluo
Before the experiment three group animal serum TC, TG, LDL no difference (P0.01), TC, normal diet group at 14 weeks compared with 0 weeks TG, LDL no significant difference (P0.01); 14 weeks of high fat diet group and Tongxinluo group TC, TG, LDL increased significantly, significantly higher than the normal diet group (P0.01); high fat diet group and Tongxinluo group, 14 weeks Shitong Tongxinluo group serum TC, TG, LDL in high fat diet group decreased significantly (P0.01).
Three, the effect of Tongxinluo on the expression of VCAM-1 and PAI-1
(1) the effect of Tongxinluo on the expression of serum VCAM-1 and PAI-1
1, the effect of Tongxinluo on the expression of serum VCAM-1
The VCAM-1 levels of the three groups in animal serum had no significant difference in the 0 week (P0.01); 14 weeks of feeding, high fat diet group and Tongxinluo group serum VCAM-1 levels were significantly increased in 0 weeks (P0.01), and higher than the normal diet group (P0.01); VCAM-1 high fat diet for 14 weeks Shitong Tongxinluo group decreased (P0.01).
2, the effect of Tongxinluo on the expression of serum PAI-1
The PAI-1 levels of the three groups in animal serum had no significant difference in the 0 week (P0.01); 14 weeks of feeding, high fat diet group and Tongxinluo group in the serum PAI-1 level was 0 Zhou Zenggao (P0.01), and higher than the normal diet group (P0.01); PAI-1 high fat diet for 14 weeks Shitong Tongxinluo group was significantly decreased (P0.01).
(two) the effect of Tongxinluo on the percentage of VCAM-1 and PAI-1 positive staining in the aortic wall
1, the effect of Tongxinluo on the percentage of VCAM-1 positive staining in the aortic wall
In the high fat diet group and Tongxinluo group, there were more brown and granular positive staining substances in the intima and middle membrane. The expression levels were 18.38 + 2.55% and 13.18 + 1.25%. respectively. The percentage of positive expression in Tongxinluo group was significantly lower than that in the high-fat diet group (P0.01).
2, the effect of Tongxinluo on the percentage of PAI-1 positive staining in the aortic wall
In the high fat diet group and Tongxinluo group, there were more brown and granular positive staining substances in the intima and middle membrane. The expression levels were 15.46 + 2.23% and 9.45 + 1.09%. respectively. The percentage of positive expression in Tongxinluo group was significantly lower than that in the high-fat diet group (P0.01).
The above experimental results were expressed with mean standard deviation (X + S). T test was used in self and group control, and there were significant differences in P0.05.
1., after 14 weeks of high fat diet group and Tongxinluo group, serum TC, TG and LDL increased significantly (P0.01), which was higher than that of the control group (P0.01). The serum TC, TG and LDL of the Tongxinluo group were significantly lower than those of the high-fat diet group (P0.01).
2., after 14 weeks of high fat diet group and Tongxinluo group, serum PAI-1 and VCAM-1 levels increased significantly compared with 0 weeks (P0.01), and higher than those in control group (P0.01). The levels of PAI-1 and VCAM-1 in Tongxinluo group were significantly lower than those in high-fat diet group (P0.01).
3. immunohistochemical analysis showed that intimal thickening in cholesterol diet group and in the film can be seen in the large brown granular staining material, partial integration into the film, especially in endometrial. Tongxinluo group positive substance was significantly lower than that in high fat diet group, there were significant differences between the three groups (P0.01).
Conclusion:
1, the expression of VCAM-1 and PAI-1 increased in serum of AS rabbits. The percentage of positive expression of VCAM-1 and PAI-1 in aorta atherosclerotic lesion of rabbits increased, and the over expression of.VCAM-1 and PAI-1 might be one of the mechanisms of AS.
2, Tongxinluo can reduce the percentage of positive expression of VCAM-1 and PAI-1 in serum and atherosclerotic lesion in AS rabbits. Inhibition of VCAM-1 and PAI-1 expression may be one of the mechanisms of Tongxinluo AS.
3, Tongxinluo not only has the effect of lipid-regulating, but also has the effect of reducing and inhibiting AS.
4, the detection of VCAM-1 and PAI-1 in serum is of great significance for the early diagnosis of AS disease and the monitoring of the occurrence, development and prognosis of AS.

【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R-332

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