成纤维细胞生长因子-10对激光角膜烧伤的实验治疗及分子机制研究

发布时间：2018-04-04 23:07

本文选题：激光　切入点：碱　出处：《中国人民解放军军事医学科学院》2011年博士论文

【摘要】：本课题旨在研究角膜烧伤的特点及发展规律,探索其损伤的机制,重点研究激光角膜损伤的分子机理及促进修复措施,同时还应用傅里叶变换红外光谱技术(FT-IR),建立了FT-IR检测角膜蛋白分子结构改变的新方法。 1.目的眼部碱烧伤是临床常见而处理棘手的眼外伤,国内外相关研究报道较多。激光角膜烧伤的研究报道甚少。随着激光技术的迅速发展,激光武器已经成为用于战争且技术最成熟的一种新概念武器,各国都在进行相关研究,但由于世界各军事强国都将激光武器的开发研制列入绝密的国家高新技术武器研究计划,因此激光生物效应的研究报道寥寥无几。随着激光武器研发的深入和实战应用,激光生物学效应、损伤机制和治疗措施的研究已成为各国军事医学的重要而紧迫的研究领域。眼是激光辐射最敏感的靶器官,也是激光武器重要致伤部位之一。本课题用碱和激光建立眼烧伤模型,重点研究激光对眼角膜的损伤效应、损伤机制以及成纤维细胞生长因子-10(Fibroblast growth factor-10,FGF-10)对角膜烧伤的治疗效应。激光眼角膜损伤效应与机制研究以及FGF-10治疗效应研究,将为临床角膜烧伤提供新的治疗方案,为我国激光武器的研制和防护提供生物学依据,对国家安全和保障人民身体健康均具有重大意义。 2.方法采用日本大耳白兔分别构建碱和激光角膜烧伤模型,实验分烧伤对照组、bFGF治疗对照组、FGF-10 12.5μg/ml组、FGF-10 25μg/ml组和FGF-10 50μg/ml组。通过裂隙灯显微镜与病理组织学观察、烧伤斑图像分析、角膜光密度值测定和MTT法检测损伤后角膜的恢复情况,比较治疗后7、14天各组的差异。建立FT-IR技术用于角膜研究的方法。分析正常兔角膜的化学组成。在此基础上进一步应用FT-IR技术研究激光角膜损伤的分子机制,采用解卷积、曲线拟合等方法对FT-IR图谱酰胺I带中蛋白质二级结构各吸收峰进行定量分析,比较对照组与照射组特征峰的位移与积分值差异,得出蛋白质结构的变化特征。 3.结果 3.1成功构建稳定的角膜碱烧伤和激光烧伤模型,在角膜中央表面造成白色圆形烧灼斑。 3.2物理和化学因素虽然性质不同,但引起的角膜损伤效应相似,主要表现为角膜上皮细胞和内皮细胞的坏死脱落、基质中以中性粒细胞为主的炎细胞浸润及成纤维细胞增生等。 3.3 FGF-10有利于眼烧伤修复,表现为烧伤斑面积缩小,角膜透光度恢复,且药物浓度为25μg/ml时,即可达到最大修复率,增加药物浓度并不能相应增加药效。 3.4 FGF-10对角膜上皮细胞没有表现促增殖作用。 3.5对FGF-10与阳性对照药bFGF的疗效进行比较,结果表明,二者疗效相当,但在改善角膜损伤后组织结构的恢复和抑制角膜新生血管(CNV)形成方面,浓度为25μg/ml的FGF-10治疗作用更显著。 3.6 FGF-10可能通过缓和促进角膜上皮细胞的再生和迁移、减轻角膜烧伤后的炎症反应及抑制成纤维细胞过度增生等机制,促进碱和激光烧伤后受损角膜的修复,具有一定的治疗效应。 3.7针对兔眼角膜上皮层和基质层的FT-IR研究发现,代表蛋白质酰胺Ⅰ带的红外吸收最大,表明FT-IR是研究角膜损伤分子机制的有效手段。 3.8角膜受激光辐射后,发生蛋白质二级结构特征峰的位移,α-螺旋减少与β-折叠结构增加,表明蛋白质结构稳定性下降,趋向无序化。 4结论 4.1物理和化学因素引起的角膜损伤效应相似,主要表现为急性炎症反应、角膜上皮细胞和内皮细胞坏死脱落、以中性粒细胞为主的炎细胞浸润及成纤维细胞增生等。 4.2 FGF-10可通过缓和促进角膜上皮细胞的再生和迁移、减轻角膜烧伤后的炎症反应及抑制成纤维细胞过度增生等机制,促进碱和激光烧伤后受损角膜的修复,具有一定的治疗效应。 4.3在改善受损后组织结构的恢复和抑制CNV方面,浓度为25μg/ml的FGF-10治疗效应优于临床常用药bFGF。 4.4 FT-IR是研究角膜分子结构组成的有效方法。 4.5激光辐射使蛋白质二级结构的特征峰发生位移,构象发生改变,进而引起蛋白质生物功能的紊乱或丧失。热效应可能是激光角膜烧伤的主要机制。
[Abstract]:This paper aims at the characteristics and the development of the study on corneal burn injury, and explore its mechanism, focus on the molecular mechanism of laser corneal injury and promote the repair measures, but also the application of Fourier transform infrared spectroscopy (FT-IR), a new method was established to detect FT-IR protein structure changes of corneal points.
1. purposes
Ocular burn is a common clinical and treatment of intractable ocular trauma, many related research reports at home and abroad. The Research Report of laser corneal burn. With the rapid development of laser technology, laser weapon has become a new concept for weapons of war and the most mature technology, each country in the related research, but because of military power the world will be the development of laser weapons included in the top secret national high-tech weapons research program, so scanty research reports and laser biological effect. With the deepening of R & D laser weapons and combat application, the effect of laser biology, study the mechanism of injury and treatment measures has become an urgent and important research field of military medicine.
The eye is the most sensitive target organ of laser radiation, but also one of the important parts of laser weapon injury. This paper set up a model with alkali burns of the eye and the laser damage effect focuses on laser on cornea, injury mechanism and fibroblast growth factor -10 (Fibroblast growth factor-10, FGF-10) effects on the treatment of corneal burn. Research and the damage mechanism of effect of laser corneal research and FGF-10 treatment effect, will provide new treatment for clinical corneal burns, providing biological basis for the development and protection of China's laser weapons, and safeguarding the people's health is of great significance to national security.
2. method
The Japanese big ear rabbit models were constructed of alkali and laser corneal burns were divided into burn control group, bFGF treatment group, FGF-10 12.5 g/ml group, FGF-10 group and 25 g/ml FGF-10 50 g/ml group. Through the slit lamp microscope and histological observation and analysis of burn image corneal optical density measurement and MTT method detection of corneal injury recovery, the difference of 7,14 day groups after treatment was compared. The establishment of FT-IR method for corneal research. Chemical analysis of normal rabbit cornea. The molecular mechanism on the basis of further study of the application of FT-IR technique in laser corneal injury, using deconvolution, curve fitting method of FT-IR spectra in the amide I band two protein secondary structures were quantitative analysis of the absorption peak and integral displacement between the control group and the irradiation group characteristic peak values, the variation of protein structure.
3. results
3.1 a stable corneal alkali burn and laser burn model was successfully constructed, and a white circular cauterization was created on the central surface of the cornea.
3.2, although physical and chemical factors are different in nature, the corneal injury effect is similar, mainly manifested as necrosis and abscission of corneal epithelial cells and endothelial cells, infiltration of inflammatory cells and fibroblast proliferation in the matrix.
3.3 FGF-10 is conducive to the repair of eye burns. It shows that the area of burn spots is reduced, the transmittance of cornea is restored, and the maximum repair rate can be achieved when the drug concentration is 25 g/ml. Increasing the concentration of drugs can not increase the efficacy of eye burns.
3.4 FGF-10 did not promote the proliferation of corneal epithelial cells.
3.5, we compared the efficacy of FGF-10 and bFGF. The results showed that the two treatments had the same effect. But when improving the tissue structure and inhibiting the formation of corneal neovascularization (CNV) after corneal injury, the therapeutic effect of FGF-10 with a concentration of 25 g/ml was more significant.
3.6 FGF-10 may play a certain therapeutic role in promoting corneal epithelial cell regeneration and migration, alleviating inflammatory reaction after corneal burn and inhibiting fibroblast proliferation, and promoting the repair of damaged cornea after alkali and laser burn.
3.7 FT-IR studies on rabbit corneal epithelium and stroma showed that the largest infrared absorption of protein amide I band showed that FT-IR is an effective way to study the molecular mechanism of corneal injury.
3.8, after the cornea is irradiated by laser, the displacement of protein two level structural characteristic peak, the decrease of alpha helix and the increase of the beta fold structure indicate that the stability of protein structure decreases and tends to disorder.
4 Conclusion
4.1, corneal injury caused by physical and chemical factors is similar, mainly manifested as acute inflammatory reaction, corneal epithelial cells and endothelial cells necrosis and abscission, neutrophil infiltration and fibroblast proliferation.
4.2 FGF-10 can help promote corneal epithelial cell regeneration and migration, relieve inflammatory reaction after corneal burn and inhibit fibroblast proliferation, and promote the repair of damaged cornea after alkali and laser burn. It has certain therapeutic effect.
4.3 the therapeutic effect of FGF-10 with a concentration of 25 mu g/ml is better than that of common clinical drug bFGF. in improving the recovery of tissue structure and inhibiting CNV after damage.
4.4 FT-IR is an effective method to study the molecular structure of the cornea.
4.5, laser radiation causes the displacement of the characteristic peak of protein two level structure, and the conformational change leads to disorder or loss of protein biological function. Thermal effect may be the main mechanism of laser corneal burn.

【学位授予单位】：中国人民解放军军事医学科学院
【学位级别】：博士
【学位授予年份】：2011
【分类号】：R772.21

【参考文献】