miR-184在碱烧伤角膜新生血管中的表达和功能研究

发布时间：2018-01-03 11:40

本文关键词：miR-184在碱烧伤角膜新生血管中的表达和功能研究　出处：《重庆医科大学》2013年硕士论文　论文类型：学位论文

【摘要】：目的：探讨miR-184在碱烧伤诱导角膜新生血管形成过程中的表达变化和作用机制，及其对体外血管新生的影响。方法：碱烧伤诱导大鼠角膜新生血管形成，基质胶上建立人脐静脉血管内皮细胞体外三维培养体系，应用实时荧光定量PCR方法检测miR-184的体内外表达。生物信息学方法预测并分析miR-184靶基因。体外培养人脐静脉血管内皮细胞，脂质体法转染miR-184mimic进入细胞中，应用MTT、Transwell检测方法分别观察转染后血管内皮细胞增殖和迁移能力的变化，，同时观察miR-184对基质胶上细胞体外管腔形成能力的影响。结果：初步建立起血管新生的体内外研究模型，miR-184在碱烧伤大鼠角膜新生血管角膜中的表达（0.145±0.013）较正常角膜(1.015±0.189)显著下降(P＜0.01),而在人脐静脉血管内皮细胞中相对表达量极少（0.007±0.004）(P＜0.05)。预测的的靶基因与VEGF血管生成信号通路有关。miR-184mimic有效转染入人脐静脉血管内皮细胞中，转染后48h细胞光密度值（0.50±0.039）和转染后24h细胞迁移过膜数（55.4±5.857）均较对照组明显减少（P＜0.01），形成管腔样结构分支点数（13.33±2.082）较对照组明显减少（P＜0.05）。结论：新生血管进程中伴随着miR-184表达量降低，miR-184能抑制细胞的生长和无血清诱导的迁移能力，减弱内皮细胞基质胶上体外管腔形成能力，可能通过VEGF血管生成信号通路参与调控血管新生和维持角膜的透明无血管状态。
[Abstract]:Objective: to investigate the expression and mechanism of miR-184 in corneal neovascularization induced by alkali burn. Methods: the corneal neovascularization was induced by alkali burn and the three-dimensional culture system of human umbilical vein endothelial cells (HUVECs) was established on the matrix glue. The expression of miR-184 in vitro and in vivo was detected by real-time fluorescence quantitative PCR. The target gene of miR-184 was predicted and analyzed by bioinformatics. Human umbilical vein endothelial cells were cultured in vitro. After transfection of miR-184mimic into cells by liposome method, the changes of proliferation and migration ability of vascular endothelial cells were observed by MTTV-Transwell assay. At the same time, we observed the effect of miR-184 on the in vitro lumen formation of stromal cells. Results: an in vitro and in vivo model of angiogenesis was established. The expression of miR-184 in corneal neovascularization cornea after alkali burn was significantly lower than that in normal cornea (P < 0.01). However, the relative expression in human umbilical vein endothelial cells was very small (0.007 卤0.004) (P < 0.05). The predicted target gene was related to the VEGF angiogenesis signaling pathway. MiR-1844 mimic was effectively transfected into human umbilical vein endothelial cells. The optical density of cells at 48 h after transfection was 0.50 卤0.039) and the number of over membrane migration was 55.4 卤5.857) at 24 h after transfection, which was significantly lower than that in the control group (P < 0.01). The number of branches forming luminal structure was 13.33 卤2.082), which was significantly lower than that in control group (P < 0.05). Conclusion: the expression of miR-184 is decreased in the process of neovascularization. MiR-184 inhibited cell growth and serum-free migration, and weakened the ability of endothelial cells to form lumen on stroma in vitro. VEGF angiogenesis signaling pathway may be involved in the regulation of angiogenesis and the maintenance of a transparent and non-vascularized cornea.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R772.2

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