本文選題：激光　切入點(diǎn)：堿　出處：《中國人民解放軍軍事醫(yī)學(xué)科學(xué)院》2011年博士論文

【摘要】：本課題旨在研究角膜燒傷的特點(diǎn)及發(fā)展規(guī)律,探索其損傷的機(jī)制,重點(diǎn)研究激光角膜損傷的分子機(jī)理及促進(jìn)修復(fù)措施,同時(shí)還應(yīng)用傅里葉變換紅外光譜技術(shù)(FT-IR),建立了FT-IR檢測角膜蛋白分子結(jié)構(gòu)改變的新方法。 1.目的 眼部堿燒傷是臨床常見而處理棘手的眼外傷,國內(nèi)外相關(guān)研究報(bào)道較多。激光角膜燒傷的研究報(bào)道甚少。隨著激光技術(shù)的迅速發(fā)展,激光武器已經(jīng)成為用于戰(zhàn)爭且技術(shù)最成熟的一種新概念武器,各國都在進(jìn)行相關(guān)研究,但由于世界各軍事強(qiáng)國都將激光武器的開發(fā)研制列入絕密的國家高新技術(shù)武器研究計(jì)劃,因此激光生物效應(yīng)的研究報(bào)道寥寥無幾。隨著激光武器研發(fā)的深入和實(shí)戰(zhàn)應(yīng)用,激光生物學(xué)效應(yīng)、損傷機(jī)制和治療措施的研究已成為各國軍事醫(yī)學(xué)的重要而緊迫的研究領(lǐng)域。 眼是激光輻射最敏感的靶器官,也是激光武器重要致傷部位之一。本課題用堿和激光建立眼燒傷模型,重點(diǎn)研究激光對眼角膜的損傷效應(yīng)、損傷機(jī)制以及成纖維細(xì)胞生長因子-10(Fibroblast growth factor-10,FGF-10)對角膜燒傷的治療效應(yīng)。激光眼角膜損傷效應(yīng)與機(jī)制研究以及FGF-10治療效應(yīng)研究,將為臨床角膜燒傷提供新的治療方案,為我國激光武器的研制和防護(hù)提供生物學(xué)依據(jù),對國家安全和保障人民身體健康均具有重大意義。 2.方法 采用日本大耳白兔分別構(gòu)建堿和激光角膜燒傷模型,實(shí)驗(yàn)分燒傷對照組、bFGF治療對照組、FGF-10 12.5μg/ml組、FGF-10 25μg/ml組和FGF-10 50μg/ml組。通過裂隙燈顯微鏡與病理組織學(xué)觀察、燒傷斑圖像分析、角膜光密度值測定和MTT法檢測損傷后角膜的恢復(fù)情況,比較治療后7、14天各組的差異。建立FT-IR技術(shù)用于角膜研究的方法。分析正常兔角膜的化學(xué)組成。在此基礎(chǔ)上進(jìn)一步應(yīng)用FT-IR技術(shù)研究激光角膜損傷的分子機(jī)制,采用解卷積、曲線擬合等方法對FT-IR圖譜酰胺I帶中蛋白質(zhì)二級結(jié)構(gòu)各吸收峰進(jìn)行定量分析,比較對照組與照射組特征峰的位移與積分值差異,得出蛋白質(zhì)結(jié)構(gòu)的變化特征。 3.結(jié)果 3.1成功構(gòu)建穩(wěn)定的角膜堿燒傷和激光燒傷模型,在角膜中央表面造成白色圓形燒灼斑。 3.2物理和化學(xué)因素雖然性質(zhì)不同,但引起的角膜損傷效應(yīng)相似,主要表現(xiàn)為角膜上皮細(xì)胞和內(nèi)皮細(xì)胞的壞死脫落、基質(zhì)中以中性粒細(xì)胞為主的炎細(xì)胞浸潤及成纖維細(xì)胞增生等。 3.3 FGF-10有利于眼燒傷修復(fù),表現(xiàn)為燒傷斑面積縮小,角膜透光度恢復(fù),且藥物濃度為25μg/ml時(shí),即可達(dá)到最大修復(fù)率,增加藥物濃度并不能相應(yīng)增加藥效。 3.4 FGF-10對角膜上皮細(xì)胞沒有表現(xiàn)促增殖作用。 3.5對FGF-10與陽性對照藥bFGF的療效進(jìn)行比較,結(jié)果表明,二者療效相當(dāng),但在改善角膜損傷后組織結(jié)構(gòu)的恢復(fù)和抑制角膜新生血管(CNV)形成方面,濃度為25μg/ml的FGF-10治療作用更顯著。 3.6 FGF-10可能通過緩和促進(jìn)角膜上皮細(xì)胞的再生和遷移、減輕角膜燒傷后的炎癥反應(yīng)及抑制成纖維細(xì)胞過度增生等機(jī)制,促進(jìn)堿和激光燒傷后受損角膜的修復(fù),具有一定的治療效應(yīng)。 3.7針對兔眼角膜上皮層和基質(zhì)層的FT-IR研究發(fā)現(xiàn),代表蛋白質(zhì)酰胺Ⅰ帶的紅外吸收最大,表明FT-IR是研究角膜損傷分子機(jī)制的有效手段。 3.8角膜受激光輻射后,發(fā)生蛋白質(zhì)二級結(jié)構(gòu)特征峰的位移,α-螺旋減少與β-折疊結(jié)構(gòu)增加,表明蛋白質(zhì)結(jié)構(gòu)穩(wěn)定性下降,趨向無序化。 4結(jié)論 4.1物理和化學(xué)因素引起的角膜損傷效應(yīng)相似,主要表現(xiàn)為急性炎癥反應(yīng)、角膜上皮細(xì)胞和內(nèi)皮細(xì)胞壞死脫落、以中性粒細(xì)胞為主的炎細(xì)胞浸潤及成纖維細(xì)胞增生等。 4.2 FGF-10可通過緩和促進(jìn)角膜上皮細(xì)胞的再生和遷移、減輕角膜燒傷后的炎癥反應(yīng)及抑制成纖維細(xì)胞過度增生等機(jī)制,促進(jìn)堿和激光燒傷后受損角膜的修復(fù),具有一定的治療效應(yīng)。 4.3在改善受損后組織結(jié)構(gòu)的恢復(fù)和抑制CNV方面,濃度為25μg/ml的FGF-10治療效應(yīng)優(yōu)于臨床常用藥bFGF。 4.4 FT-IR是研究角膜分子結(jié)構(gòu)組成的有效方法。 4.5激光輻射使蛋白質(zhì)二級結(jié)構(gòu)的特征峰發(fā)生位移,構(gòu)象發(fā)生改變,進(jìn)而引起蛋白質(zhì)生物功能的紊亂或喪失。熱效應(yīng)可能是激光角膜燒傷的主要機(jī)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.

【學(xué)位授予單位】：中國人民解放軍軍事醫(yī)學(xué)科學(xué)院
【學(xué)位級別】：博士
【學(xué)位授予年份】：2011
【分類號】：R772.21

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