共聚焦显微镜下糖尿病兔的角膜病变研究

发布时间：2018-05-21 01:13

本文选题：糖尿病 + 角膜　；参考：《暨南大学》2012年硕士论文

【摘要】：目的：1.采用共聚焦显微镜对糖尿病兔角膜组织的三维立体结构进行活体观察，了解糖尿病角膜组织病变的病理形成过程。2.糖尿病兔角膜组织切片HE染色，并与活体共聚焦显微镜的观察结果结合起来，宏观分析糖尿病角膜病变的发病机制。3.通过免疫组织化学方法检测半胱氨酸天冬氨酸蛋白酶3(caspase-3)在糖尿病兔角膜组织的表达水平，，并观察角膜细胞与caspase-3表达变化的关系，蛋白水平分析糖尿病角膜病变发病的分子机制。方法：1.四氧嘧啶静脉注射新西兰兔（18只），并给予高糖高脂饲料喂养来复制糖尿病模型；对照组新西兰兔（3只）给予同等剂量的生理盐水注射，且普通饲料喂养。2.分别在给药前和疾病模型建立后4w,8w,12w采用角膜共聚焦显微镜进行活体观察。3.分别在疾病模型建立后4w,8w各处死3只糖尿病兔，12w处死12只糖尿病兔和对照组3只正常新西兰兔，角膜组织切片HE染色，显微镜下观察。4.免疫组化观察角膜组织切片Caspase-3的表达，并与共聚焦活体观察和角膜组织切片HE染色的结果结合，综合分析和研究糖尿病角膜病变的发病机制。结果：1.糖尿病兔血糖增高显著，血糖最低17.3mmol/L,最高31.2mmol/L，平均22.97±4.03mmol/L。2.随着糖尿病模型建立时间的延长，糖尿病兔角膜逐渐出现不同层次的上皮脱落且脱落的面积逐渐增大。前、后基质细胞数目减少,基质水肿，基质细胞核出现形态改变。内皮层多形性细胞增加，内皮细胞数目减少。上皮下神经纤维出现弯曲，变形等形态学改变。基质神经纤维分枝减少，神经出现断裂。3. Caspase-3在糖尿病兔角膜组织的上皮和基质层都有表达，且随着糖尿病时间延长，Caspase-3表达增强。结论：糖尿病血糖增高导致糖尿病角膜病变的发生。随糖尿病疾病时间的延长，角膜神经纤维和角膜上皮细胞核均发生变性，且上皮细胞凋亡、脱落；角膜基质细胞排列疏松，水肿，细胞核变形，凋亡。我们考虑糖尿病角膜上皮细胞和基质细胞的这些病变与角膜神经纤维变性有关。在没有角膜神经分布的角膜内皮层，角膜内皮细胞最先被观察到出现细胞形态改变，暗示这可能与角膜内皮与房水直接接触相关。
[Abstract]:Purpose 1. A confocal microscope was used to observe the three-dimensional structure of the cornea of diabetic rabbits in vivo, and to understand the pathological process of diabetic corneal lesions. The pathological mechanism of diabetic keratopathy was analyzed macroscopically by HE staining in corneal sections of diabetic rabbits combined with the observation of confocal microscope in vivo. The expression of cysteine aspartate proteinase 3 (caspase-3) in the cornea of diabetic rabbits was detected by immunohistochemical method. The relationship between corneal cells and caspase-3 expression was observed. The protein level was used to analyze the molecular mechanism of diabetic keratopathy. Method 1: 1. Alloxan was injected intravenously into 18 New Zealand rabbits and fed with high sugar and high fat diet to make diabetic model; in the control group, 3 New Zealand rabbits were given the same dose of normal saline, and the normal diet was fed. 2. The corneal confocal microscope was used to observe in vivo before administration and at 8 weeks and 12 weeks after the establishment of the disease model. 12 diabetic rabbits and 3 normal New Zealand rabbits were killed at 4 weeks and 8 weeks after the establishment of the model. The corneal tissue sections were stained with HE and observed under microscope. The expression of Caspase-3 in corneal sections was observed by immunohistochemistry and combined with the results of confocal in vivo and HE staining in corneal sections to analyze and study the pathogenesis of diabetic keratopathy. The result is 1: 1. Diabetic rabbits had a significant increase in blood glucose, with the lowest 17.3 mmol / L and the highest 31.2 mmol / L, with an average of 22.97 卤4.03 mmol / L. With the prolongation of the diabetic model, the corneal epithelium of diabetic rabbits gradually appeared different levels of epithelial exfoliation and the area of exfoliation gradually increased. The number of anterior and posterior stromal cells decreased, matrix edema and morphologic changes of stromal nucleus appeared. The number of endothelial cells decreased with the increase of pleomorphic cells in the endodermis. Morphologic changes such as curvature and deformation of the nerve fibers in the subepithelial region. The branching of stromal nerve fibers was decreased, and the nerve was broken. 3. Caspase-3 was expressed in corneal epithelium and stroma of diabetic rabbits, and the expression of Caspase-3 was increased with the prolongation of diabetic time. Conclusion: diabetic keratopathy is caused by hyperglycemia. With the extension of diabetic disease, corneal nerve fibers and corneal epithelial nuclei were denatured, and epithelial cells were apoptotic, and corneal stromal cells were arranged loosely, edema, deformed nucleus and apoptosis. We consider that these lesions in diabetic corneal epithelial and stromal cells are associated with corneal nerve fiber degeneration. In the corneal endothelium without corneal nerve distribution, the morphology of corneal endothelial cells was first observed, suggesting that this may be related to the direct contact between corneal endothelium and aqueous humor.
【学位授予单位】：暨南大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R772.2;R587.2

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