AQP1在大鼠放射性白内障晶状体上皮的变化及其机制的实验研究

发布时间：2018-07-03 15:14

本文选题：X射线 + 放射性白内障　；参考：《苏州大学》2010年硕士论文

【摘要】： 目的:利用动物实验,通过X射线照射S-D大鼠双眼建立放射性白内障动物模型,观察X射线剂量与白内障严重程度之间的关系;检测受不同剂量X射线照射的大鼠晶状体中水通道蛋白1(AQP1)的表达水平,探讨水通道蛋白1与放射性白内障形成之间的关系。方法:照射前选取8只大鼠作为正常对照组,实验照射组大鼠24只,平均分成3组每组8只,一次性分别照射X射线(双眼同时照射)5、15、25 Gy.实验对照组24只,分成3组每组8只,分别作为5、15、25 Gy的对照组行假照射(照射光线中不含X射线,其它条件分别与对应各照射组相同)。每只大鼠随机取一只眼睛(本实验取右眼)为研究对象。所有大鼠分别于照射后1、3、5、7、15、30、45、60、90 d,腹腔注射3.6%水合氯醛麻醉后,用双星明滴眼液充分散瞳,裂隙灯显微镜观察并记录晶状体混浊情况;运用免疫组织化学方法(SP法)对AQP1在大鼠晶状体上的表达进行定位检测,并采用Motic Med 6.0数码医学图像分析系统半定量检测AQP1的表达;运用逆转录聚合酶链反应(RT-PCR)及计算机图像分析技术检测AQP1在各组大鼠晶状体上的表达变化;应用SPSS11.5统计软件对结果进行统计分析,采用Independent Samples T-Test法和One-Way-ANOVA法分别对计量资料进行独立样本T检验和单因素方差分析,并使用LSD法进行组间比较。结果: 1.组织学:观察3个月,各实验对照组和5 Gy组大鼠的晶状体均未出现混浊。15 Gy照射组于照射后45 d晶状体后囊和后皮质开始出现细小、散在白点状混浊,继而融合成不均匀较密集点状、片状混浊,进一步发展后皮质出现较均匀致密混浊,前皮质、前囊和核部轻度混浊。到3个月时62.5%的晶状体达Ⅱ期混浊,37.5%达Ⅲ期混浊。25 Gy照射组亦出现类似变化,但是晶体发生混浊的时间更早、速度更快、混浊更明显。于照射后30d后囊即开始出现点状混浊,此后病变迅速发展,到45d时即有37.5%的晶体全部混浊。到3个月的时候除1例为后囊下明显混浊外,其余7例晶状体均完全混浊。 2.免疫组织化学和计算机图像分析结果显示:AQP1在大鼠晶状体上阳性表达的部位位于晶状体前囊膜的晶体上皮细胞(lens epithelial cells,LECs)的胞膜,胞膜上可见程度不等的棕黄色颗粒。各组均可见到其阳性表达,随照射量增大,各实验组LECs上的AQP1均较空白组表达减少,差异有显著性意义(P0.05)。各实验组AQP1的表达随照射剂量的增加而减少,组内差异有统计学意义(P0.05)。 3.RT-PCR和计算机图像分析结果显示:在mRNA水平AQP1在大鼠晶状体LECs胞膜表达,实验组和空白组的晶状体进行RT-PCR均可获得AQP1的阳性目的基因,不同照射量实验组晶状体的AQP1的表达均较空白组有不同程度的减少,差异有统计学意义(P0.05)。而且随着照射量的增加,各亚组之间AQP1的表达水平降低,变化呈剂量依赖性,差异有统计学意义(P0.05)。结论:大剂量X射线在短期内即可引起晶状体混浊;放射性白内障的形成速度和程度与射线剂量成正相关。在辐射线诱导的白内障模型,大鼠晶状体中AQP1的表达发生改变,辐射损伤可能通过减少AQP1在晶状体上皮细胞上的表达,影响晶状体的水代谢,以诱发白内障在放射性白内障中晶状体上皮细胞内AQP1表达减少,在白内障形成中起重要作用。
[Abstract]:Objective: to establish a radioactive cataract animal model in the eyes of S-D rats by X ray irradiation, and to observe the relationship between the dose of X ray and the severity of cataract, and to detect the expression level of aquaporin 1 (AQP1) in the lens of rats exposed to different doses of X ray, and to explore the formation of aquaporin 1 and the formation of radioactive cataract. The relationship between them.
Methods: before irradiation, 8 rats were selected as the normal control group, and 24 rats in the experimental group were divided into 3 groups, with an average of 8 rats in each group. 24 rats were irradiated with X ray (double binocular simultaneous irradiation) 5,15,25 Gy. experimental control group, and 8 rats in each group were divided into 3 groups of 8, respectively, which were respectively irradiated with no X rays in the light rays and other conditions. Each rat was treated with one eye (the right eye) randomly. All rats were treated with 1,3,5,7,15,30,45,60,90 d after irradiation. After intraperitoneal injection of 3.6% chloral anaesthesia, double star eyedrops were used to fully diffuse pupil, slit lamp microscopes were observed and recorded in the lens opacity; immunization was used. The expression of AQP1 in rat lens was detected by histochemical method (SP method), and the expression of AQP1 was detected by Motic Med 6 digital medical image analysis system. Reverse transcription polymerase chain reaction (RT-PCR) and computer image analysis were used to detect the expression of AQP1 in the lens of rats in each group; and SPSS11. was applied. 5 statistical software was used to analyze the results. The Independent Samples T-Test method and One-Way-ANOVA method were used to carry out independent sample T test and single factor analysis of variance, and the LSD method was used to compare the data between groups.
Result:
1. histology: after 3 months of observation, the lens of the experimental control group and the 5 Gy group had no cloudy.15 Gy irradiation group, and the posterior capsule and the posterior cortex began to appear in the posterior capsule and the posterior cortex after the irradiation. In the cortex, the anterior capsule and the nucleus, 62.5% of the lens reached phase II turbidity by 3 months, and the 37.5% period of phase III cloudy.25 Gy irradiation group also appeared similar changes, but the time of turbidity in the crystal was earlier, the speed was faster, and the turbidity was more obvious. After 30d, the PACA began to appear turbid, and then the pathological changes developed rapidly, when 45d was 37.. 5% of the lenses were all cloudy. At 3 months, 7 of the 7 patients were completely opacification except for 1 cases of posterior capsular opacification.
2. the results of immuno histochemistry and computer image analysis showed that the positive expression of AQP1 in the lens of the rat was located in the membrane of the lens epithelial cells (lens epithelial cells, LECs) in the anterior capsule of the lens, and the brown yellow granules of different degree were found on the membrane. The positive expression was seen in each group, with the increase of the irradiation amount, the experimental group was LE. The expression of AQP1 on Cs was less than that in the blank group, and the difference was significant (P0.05). The expression of AQP1 in the experimental group decreased with the increase of irradiation dose, and the difference in the group was statistically significant (P0.05).
The results of 3.RT-PCR and computer image analysis showed that the expression of LECs cell membrane in the rat lens at the level of AQP1 at mRNA level, the positive target gene of AQP1 in the experimental group and the blank group could be obtained by RT-PCR, and the expression of AQP1 in the lens of the experimental group of different irradiated groups were all lower than that in the blank group, and the difference was statistically significant (P0.05). Moreover, with the increase of irradiation dose, the expression level of AQP1 in all subgroups decreased, and the change was dose-dependent, and the difference was statistically significant (P0.05).
Conclusion: large dose X rays can cause lens opacities in a short period of time. The velocity and degree of radiation cataract formation is positively correlated with the dose of ray. In the cataract model induced by radiation, the expression of AQP1 in the lens of rat is changed. The radiation damage may affect the crystalline form by reducing the expression of AQP1 on the lens epithelial cells and affecting the crystalline form. The water metabolism of the body can induce the decrease of AQP1 expression in MICROTEK epithelial cells, and play an important role in the formation of cataract.
【学位授予单位】：苏州大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R776.1

【参考文献】