高同型半胱氨酸血症大鼠视网膜ICAM-1表达的研究

发布时间：2018-05-15 06:04

本文选题：高同型半胱氨酸血症 + 同型半胱氨酸　；参考：《中国人民解放军军医进修学院》2010年硕士论文

【摘要】： 目的： 1、建立高同型半胱氨酸血症大鼠模型。 2、采用免疫组化染色和Western blot免疫印迹法研究高同型半胱氨酸血症大鼠视网膜表达ICAM-1的部位和强度,印证高同型半胱氨酸血症对视网膜的毒性作用机制。方法： 1、采用在大鼠每升饮水中添加1.5g L-甲硫氨酸,使大鼠摄入过量甲硫氨酸,饲养大鼠6周,制造高同型半胱氨酸血症模型。 2、采用HE染色法观察高同型半胱氨酸大鼠视网膜的结构。 3、应用抗ICAM-1抗体免疫组化方法,检测大鼠视网膜上ICAM-1的表达量及其表达部位。 4、应用Western blot免疫印迹检测大鼠视网膜ICAM-1的表达量,印证明免疫组化的检测结果。结果： 1、采用在大鼠每升饮水中添加1.5g L-甲硫氨酸,饲养大鼠6周后,采大鼠尾静脉血检测血浆同型半胱氨酸,对照组血浆Hcy平均浓度为7.0146±2.00μmol／L,模型组为42.9081±12.02μmol／L,对照组中所有大鼠个体血浆Hcy浓度均小于15μmol／L,模型组中所有大鼠个体血浆Hcy浓度均大于15μmol／L,没有大鼠从实验中剔除。两组大鼠血浆Hcy浓度经t检验,P=0.000,P＜0.05,故两组大鼠血同型半胱氨酸浓度差别有统计学意义。显示造模成功。 2、在HE切片上,对照组和模型组大鼠视网膜组织结构未见明显变化。两组大鼠视网膜均未见出血、炎性细胞浸润、血管闭塞等改变。 3、免疫组化检查发现对照组OD值为0.284±0.0383；模型组为0.486±0.0329。两组之间有明显统计学差异。 4、对照组平均ICAM-1／β-actin比值为0.35±0.04,模型组为0.49±0.03,两组大鼠视网膜组织Western blot实验显色条带OD值经t检验,P=0.000,P＜0.05,故两组OD值差别有统计学意义。结论 1、0.15%浓度的甲硫氨酸饮用水能顺利诱导出中度升高的高同型半胱氨酸血症大鼠,显示动物模型制造成功； 2、用免疫组化和Western bolt免疫印迹法在大鼠视网膜上检测到了ICAM-1; 3、高同型半胱氨酸血症能促进大鼠视网膜ICAM-1表达量上升； 4、在参加实验的所有动物中,未发现眼底有明显病变,提示高同型半胱氨酸血症致病作用的发生可能还需要更长的时间,或同型半胱氨酸需要联合其他致病因素才能起作用。
[Abstract]:Objective: 1. The model of hyperhomocysteinemia was established in rats. 2. The localization and intensity of ICAM-1 expression in the retina of rats with hyperhomocysteinemia were studied by immunohistochemical staining and Western blot immunoblotting to confirm the toxic mechanism of hyperhomocysteinemia on retina. Methods: 1. By adding 1.5 g L-methionine to every liter of drinking water, the rats were fed with excessive methionine for 6 weeks, and the model of hyperhomocysteinemia was established. 2. The structure of high homocysteine rat retina was observed by HE staining. 3. The expression and location of ICAM-1 in rat retina were detected by immunohistochemical method with anti-ICAM-1 antibody. 4. Western blot Western blot was used to detect the expression of ICAM-1 in rat retina. Results: 1. The rats were fed with 1.5 g L-methionine per liter of drinking water for 6 weeks. The plasma homocysteine was detected in the caudal vein blood of the rats. The average plasma Hcy concentration was 7.0146 卤2.00 渭 mol / L in the control group, 42.9081 卤12.02 渭 mol / L in the model group, and less than 15 渭 mol / L in all the rats in the control group. The plasma Hcy concentration of all the rats in the model group was greater than 15 渭 mol / L, and no rats were excluded from the experiment. The plasma Hcy concentration of the two groups was tested by t test (P < 0.05), so the difference of plasma homocysteine concentration between the two groups was statistically significant. The model is shown to be successful. 2. In HE section, there was no obvious change in retinal tissue structure in control group and model group. No hemorrhage, inflammatory cell infiltration and vascular occlusion were found in the retina of both groups. 3The OD value of the control group was 0.284 卤0.0383, and that of the model group was 0.486 卤0.0329. There was a significant difference between the two groups. 4. The average ratio of ICAM-1/ 尾 -actin was 0.35 卤0.04 in the control group and 0.49 卤0.03 in the model group. The OD value of the Western blot in the two groups was significantly higher than that in the control group (P < 0.05). Conclusion (1) 0.15% methionine drinking water could induce moderately elevated hyperhomocysteinemia rats, indicating that the animal model was made successfully. 2ICAM-1 was detected in rat retina by immunohistochemistry and Western bolt immunoblotting. 3. Hyperhomocysteinemia could promote the increase of ICAM-1 expression in rat retina. 4. In all the animals involved in the experiment, there were no obvious lesions in the fundus, suggesting that the pathogenicity of hyperhomocysteinemia may take longer, or that homocysteine needs to be combined with other pathogenic factors to play its role.
【学位授予单位】：中国人民解放军军医进修学院
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R774.1

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