小鼠糖尿病性角膜病变模型的建立及上皮病变的初步研究
发布时间：2018-12-14 18:44
【摘要】：小鼠糖尿病性角膜病变模型的建立及上皮病变的初步研究 目的：研究链脲佐菌素(Streptozocin, STZ)腹腔注射诱导的糖尿病小鼠模型中角膜病变的病理变化及其机制。 方法：建立STZ腹腔注射诱导C57BL/6小鼠的糖尿病模型,在成模2-16周分别行1%虎红染色检查角膜上皮完整性；角膜上皮刮除后,荧光素钠染色观察比较角膜上皮的愈合速度；病理检测角膜的组织形态和结构的变化；免疫组化检测刮除上皮和未刮除上皮DNA损伤标记8-OHdG的表达情况；透射电子显微镜检查角膜在超微结构水平的变化。 结果：STZ腹腔注射后,C57BL/6小鼠逐渐表现出糖尿病多饮、多食、多尿、体重下降等典型症状,其血糖稳定维持在较高水平(18mmol/L),体重较正常小鼠明显偏低。虎红染色发现,成模2周的小鼠角膜即已出现点状着色,且随着病程的延长,染色面积和程度逐渐加重,到成模16周几乎呈全角膜着色。在上皮愈合速度方面,刮除角膜上皮后,成模2周的小鼠角膜上皮在40小时即完全愈合(与正常小鼠相似),成模4周小鼠上皮愈合时间为120小时,成模8周、12周、16周上皮完全愈合的时间为144小时左右,其中成模16周的小鼠在96小时至120小时上皮缺损的范围再次扩大,后逐渐缩小愈合,呈现反复现象。病理检测显示,STZ诱导的糖尿病小鼠角膜上皮细胞排列疏松极性逐渐消失,细胞水肿,细胞层数较正常对照小鼠明显减少,基质层纤维增粗排列紊乱。8-OHdG免疫组化检测显示,STZ诱导的小鼠在未刮除角膜上皮和刮除角膜上皮的角膜上皮层和基质层可见其阳性染色,尤其在中央角膜上皮刮除后残存的角膜上皮中表达水平较高,且随着糖尿病病程延长,表达量逐渐升高。透射电镜检测发现,STZ诱导的糖尿病小鼠角膜上皮表面的微绒毛减少,上皮细胞间以及上皮与基底膜间的紧密连接数目明显减少。 结论：STZ腹腔注射诱导的糖尿病小鼠角膜,表现出角膜上皮损害,角膜上皮损伤后延迟愈合等症状,说明其可以作为研究糖尿病角膜病变的动物模型。此外,糖尿病小鼠角膜上皮在超微结构和DNA氧化应激损伤方面的异常可能是造成上皮损伤愈合延迟的部分原因。
[Abstract]:Establishment of Diabetic Corneal lesion Model and preliminary study on epithelial lesions in mice objective: to study the pathological changes and mechanism of corneal lesions in diabetic mice induced by intraperitoneal injection of streptozotocin (Streptozocin, STZ). Methods: the diabetic model of C57BL/6 mice induced by intraperitoneal injection of STZ was established and the corneal epithelial integrity was examined by 1% tiger red staining at 2-16 weeks. The healing rate of corneal epithelium was observed by fluorescein sodium staining, the changes of corneal tissue morphology and structure were detected by pathology, and the expression of 8-OHdG was detected by immunohistochemistry. The ultrastructural changes of cornea were examined by transmission electron microscope (TEM). Results: after intraperitoneal injection of STZ, C57BL/6 mice gradually showed typical symptoms of diabetes mellitus, polydipsia, polyuria and weight loss, and their blood glucose remained stable at a higher level (18mmol/L), and their body weight was significantly lower than that of normal mice. It was found by tiger red staining that the cornea of mice in the second week of model formation had been stained with dots, and with the prolongation of the course of disease, the staining area and degree became more and more serious, and almost the whole cornea was stained at the 16th week of model formation. In terms of the speed of epithelial healing, the corneal epithelium healed completely in 40 hours (similar to normal mice) in 2 weeks after scraping corneal epithelium, 120 hours in 4 weeks, 8 and 12 weeks in model. The time of complete epithelial healing at 16 weeks was about 144 hours, and the range of epithelial defect in mice with model 16 weeks was enlarged again from 96 hours to 120 hours, and then gradually reduced and healed, showing repeated phenomena. Pathological examination showed that the loose polarity of corneal epithelial cells in diabetic mice induced by STZ disappeared gradually, the cell edema and the number of cell layers decreased significantly compared with the normal control mice. 8-OHdG immunohistochemical examination showed that STZ induced mice had positive staining in corneal epithelium and stroma of unscraped and erased corneal epithelium. Especially in the corneal epithelium remaining after central corneal epithelial curettage, the expression level was higher, and with the prolongation of diabetes course, the expression level gradually increased. Transmission electron microscopy (TEM) showed that the number of tight junctions between epithelial cells and basement membrane decreased significantly in diabetic mice induced by STZ. Conclusion: the cornea of diabetic mice induced by intraperitoneal injection of STZ showed corneal epithelial damage and delayed healing after corneal epithelial injury, which indicated that it could be used as an animal model for the study of diabetic keratopathy. In addition, the abnormal ultrastructure of corneal epithelium and oxidative stress injury of DNA in diabetic mice may be part of the cause of delayed healing of epithelial injury.
【学位授予单位】：青岛大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R772.2
本文编号：2379132
[Abstract]:Establishment of Diabetic Corneal lesion Model and preliminary study on epithelial lesions in mice objective: to study the pathological changes and mechanism of corneal lesions in diabetic mice induced by intraperitoneal injection of streptozotocin (Streptozocin, STZ). Methods: the diabetic model of C57BL/6 mice induced by intraperitoneal injection of STZ was established and the corneal epithelial integrity was examined by 1% tiger red staining at 2-16 weeks. The healing rate of corneal epithelium was observed by fluorescein sodium staining, the changes of corneal tissue morphology and structure were detected by pathology, and the expression of 8-OHdG was detected by immunohistochemistry. The ultrastructural changes of cornea were examined by transmission electron microscope (TEM). Results: after intraperitoneal injection of STZ, C57BL/6 mice gradually showed typical symptoms of diabetes mellitus, polydipsia, polyuria and weight loss, and their blood glucose remained stable at a higher level (18mmol/L), and their body weight was significantly lower than that of normal mice. It was found by tiger red staining that the cornea of mice in the second week of model formation had been stained with dots, and with the prolongation of the course of disease, the staining area and degree became more and more serious, and almost the whole cornea was stained at the 16th week of model formation. In terms of the speed of epithelial healing, the corneal epithelium healed completely in 40 hours (similar to normal mice) in 2 weeks after scraping corneal epithelium, 120 hours in 4 weeks, 8 and 12 weeks in model. The time of complete epithelial healing at 16 weeks was about 144 hours, and the range of epithelial defect in mice with model 16 weeks was enlarged again from 96 hours to 120 hours, and then gradually reduced and healed, showing repeated phenomena. Pathological examination showed that the loose polarity of corneal epithelial cells in diabetic mice induced by STZ disappeared gradually, the cell edema and the number of cell layers decreased significantly compared with the normal control mice. 8-OHdG immunohistochemical examination showed that STZ induced mice had positive staining in corneal epithelium and stroma of unscraped and erased corneal epithelium. Especially in the corneal epithelium remaining after central corneal epithelial curettage, the expression level was higher, and with the prolongation of diabetes course, the expression level gradually increased. Transmission electron microscopy (TEM) showed that the number of tight junctions between epithelial cells and basement membrane decreased significantly in diabetic mice induced by STZ. Conclusion: the cornea of diabetic mice induced by intraperitoneal injection of STZ showed corneal epithelial damage and delayed healing after corneal epithelial injury, which indicated that it could be used as an animal model for the study of diabetic keratopathy. In addition, the abnormal ultrastructure of corneal epithelium and oxidative stress injury of DNA in diabetic mice may be part of the cause of delayed healing of epithelial injury.
【学位授予单位】：青岛大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R772.2
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相关期刊论文 前1条
1 姜琨,秦樾,童坦君;erbB-2表达抑制与表皮生长因子刺激对信号转导与转录激活分子及细胞周期蛋白D1的影响[J];北京医科大学学报;1998年03期
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