人羊膜匀浆提取液对大鼠碱烧伤后角膜超微结构的影响

发布时间：2018-06-01 21:23

本文选题：羊膜匀浆提取液 + 共聚焦显微镜　；参考：《南京大学》2013年硕士论文

【摘要】：目的：观察人羊膜匀浆提取液对大鼠角膜碱烧伤后角膜超微结构的影响,探讨羊膜匀浆提取液应用于角膜碱烧伤后的临床价值及作用机理。方法：健康无眼疾的SD大鼠40只(40眼)随机分为实验组和对照组,每组20只(20只眼)。相同条件下,所有大鼠使用lmol/L的NaOH溶液建立右眼角膜碱烧伤模型。损伤后实验组立即行人羊膜匀浆提取液点眼,每日四次,对照组用PBS液点眼,每日四次。碱烧伤后每日在裂隙灯显微镜下观察大鼠眼表的总体情况,例如眼结膜充血,角膜透明性,以及新生血管生长情况。进行眼前节照相,通过图像分析系统记录两组大鼠角膜上皮愈合,以及角膜新生血管生长情况。隔日行共聚焦显微镜动态观察大鼠角膜超微结构的变化,分析角膜中炎症细胞、成纤维细胞以及内皮细胞的数量及形态改变。于烧伤后3d、7d、14d、28d每组随机处死3只试验大鼠,取大鼠眼球进行HE染色,行组织学染色观察角膜各层超微结构的变化。结果：碱烧伤后第1d至第7d,实验组角膜上皮愈合明显快于对照组(P0.01)。实验组角膜新生血管出现平均时间为(4.8±0.6)d,晚于对照组(3.1±0.5)d,(P0.01),对照组最大新生血管面积为(48.72±2.12)mm2,实验组为(35.12±1.77)mm2,各个时间点两组之间角膜新生血管面积的差异均有统计学意义(各p0.01)。碱烧伤后9d,对照组炎症细胞数量最多为(3164±306)个/mm2,实验组为(2830±254)个/mm2,两者之间差异有统计学意义(P0.01)；碱烧伤后11d,对照组成纤维细胞数量为(349±105)个/mm2,实验组为(248±84)个/mm2,两者之间差异有统计学意义(P0.01)；碱烧伤后28d,对照组内皮细胞计数为(1837±59)个/mm2,对照组内皮细胞计数为(2063±54)个/mm2,两者之间差异有统计学意义(P0.01)。HE染色提示实验组炎症细胞、成纤维细胞、角膜新生血管数量较少,基质层纤维结构排列较为规则。结论：新鲜人羊膜匀浆提取液可以有效促进大鼠碱烧伤后角膜上皮细胞的增殖,抑制炎症细胞浸润,减少角膜新生血管的形成,抑制成纤维细胞的激活和增殖,减少角膜瘢痕的形成,有利于碱烧伤后大鼠角膜的透明愈合。
[Abstract]:Objective: To observe the effect of human amniotic membrane homogenate extract on the ultrastructure of cornea after alkali burn of rat cornea, and to explore the clinical value and mechanism of application of amniotic membrane homogenate extract to corneal alkali burn. Methods: Forty SD rats without eye disease were randomly divided into experimental group and control group with 20 eyes in each group. Under the same conditions, all the rats used NaOH solution of lmol/L to establish the right eye corneal alkali burn model. After injury, the experimental group was treated with amniotic membrane homogenate extract, four times a day, while the control group was treated with PBS solution four times a day. After alkali burn, we observed the overall condition of eye surface under slit lamp microscope, such as conjunctival hyperemia, corneal transparency, and neovascularization. The corneal epithelium healing and corneal neovascularization were recorded by image analysis system. The changes of corneal ultrastructure were observed by confocal microscope on the following day, and the number and morphology of inflammatory cells, fibroblasts and endothelial cells in the cornea were analyzed. Three rats in each group were randomly killed on the 3rd day, 7th day, 14d and 28d after burn. The eyeballs of the rats were stained with HE, and the ultrastructure of each layer of cornea was observed by histological staining. Results: From the first day to the 7th day after alkali burn, the corneal epithelium healed faster in the experimental group than that in the control group (P 0.01). The mean time of corneal neovascularization in the experimental group was 4.8 卤0.6 days, which was later than that in the control group (3.1 卤0.5), the maximum neovascularization area in the control group was 48.72 卤2.12mm2, and in the experimental group was 35.12 卤1.77m2.There was significant difference in the corneal neovascularization area between the two groups at each time point (p0.01). At 9 days after alkali burn, the number of inflammatory cells in the control group was 3164 卤306 / m2, and in the experimental group was 2830 卤254 / m2, the difference was statistically significant (P 0.01); at 11 days after alkali burn, the number of fibroblasts in the control group was 349 卤105 / mm2, and in the experimental group was 248 卤84 / mm2, between the two groups. The number of endothelial cells in the control group was 1837 卤59 / m2 and that in the control group was 2063 卤54 / mm2 on the 28th day after alkali burn. The difference between the two groups was statistically significant (P 0.01) and HE staining indicated that the inflammatory cells were found in the experimental group. The number of corneal neovascularization in fibroblasts was less and the fibrous structure of stroma was regular. Conclusion: Fresh human amniotic membrane homogenate extract can effectively promote the proliferation of corneal epithelial cells, inhibit the infiltration of inflammatory cells, reduce the formation of corneal neovascularization, inhibit the activation and proliferation of fibroblasts, and reduce the formation of corneal scar. It is beneficial to the transparent healing of cornea after alkali burn in rats.
【学位授予单位】：南京大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R779.13

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