本文選題：板層角膜移植　切入點：前板層角膜供體　出處：《南昌大學》2012年碩士論文　論文類型：學位論文

【摘要】：目的：建立大鼠同種異體無縫線前、后板板層角膜移植(LKP)動物模型。觀察角膜上皮細的生長及組織學特點。 方法：選取Wistar大鼠27只54眼為供體制備前板（前彈力層+部分基質(zhì)層）和后板（后彈力層+部分基質(zhì)層）角膜植片；雌性SD大鼠108只為受體分為2組，每組54只，實驗組行后板LKP，對照組行前板LKP，兩組大鼠均右眼施行LKP手術(shù)，左眼為正常對照。用豬源纖維蛋白粘合劑（Porcine Fibrin Sealant Kit）替代角膜縫線行LKP，術(shù)畢行瞼緣縫合術(shù)[1]。兩組術(shù)后第l天拆除眼瞼縫合線，前3天滴用0.5%左氧氟沙星滴眼液（參天制藥可樂必妥）和妥布霉素地塞米松滴眼液（Alcon公司典必殊）4次/日，以后滴眼藥水2次/日，1月后停藥。兩組術(shù)后3、7、l4、28、42及56天裂隙燈下觀察術(shù)眼并行排斥反應指數(shù)(RI)評分，熒光素鈉染色觀察角膜缺損并用photoshop軟件計算角膜上皮生長面積。各觀察時間點處死實驗組和對照組大鼠各9只，取術(shù)眼角膜制備HE染色標本、透射電鏡標本觀察角膜上皮各層細胞形態(tài)、層數(shù)同時行角膜K12免疫熒光檢測角蛋白表達水平。 結(jié)果：選入的108只SD大鼠在實驗過程中因麻醉過度死亡占14只，術(shù)后植片出現(xiàn)感染占7只，均被清除后得到及時補充，108只SD大鼠全部進入最終結(jié)果分析。裂隙燈下觀察：實驗組和對照組角膜植片混濁水腫過程相同， RI圖表曲線幾乎重疊，其峰值及整體曲線走向大體一致，整體RI指數(shù)呈下降趨勢。熒光素鈉染色：兩組著色范圍縮小時間相同。photoshop畫圖軟件輔助分析角膜上皮生長面積觀察到兩組術(shù)后各時間點角膜上皮爬行面積差異甚小。對RI和Fls評分數(shù)據(jù)及上皮生長面積，采取兩樣本t檢驗，以a=0.05，p0.05為數(shù)據(jù)有統(tǒng)計學意義，計算得出各時間點p0.05，差異均無統(tǒng)計學意義。HE染色切片光鏡觀察：兩組術(shù)后均從早期上皮生長不典型到后期上皮結(jié)構(gòu)均長出，，形態(tài)接近正常上皮細胞。透射電鏡觀察：早期兩組角膜表皮細胞微絨毛均稀疏，至后期五層上皮細胞結(jié)構(gòu)均長出，微絨毛生長致密。同時觀察到角膜上皮基底膜與后彈力層連接有間隙，而與前彈力層連接緊密。K12免疫熒光檢測：兩組術(shù)后免疫熒光檢測可見上皮細胞從早期的弱熒光到后期的強熒光。 結(jié)論：1、首次建立大鼠同種異體無縫線前、后板LKP動物模型。 2、通過各項指標檢測，觀察到兩組在不同時間點上皮爬行的速率、面積及生長形態(tài)大致相同，證實后板同樣能作為角膜上皮細胞生長的載體，為后板進一步應用提供了實驗和理論依據(jù)。 3、通過透射電鏡觀察前、后板LKP術(shù)后角膜上皮移行中超微結(jié)構(gòu)的差異，為后續(xù)科研工作提供了重要參考。 4、豬源纖維蛋白粘合劑成功應用于大鼠板層角膜移植，達到無縫線手術(shù)目的，為其進一步應用于無縫線角膜移植手術(shù)提供了實驗基礎(chǔ)。
[Abstract]:Aim: to establish a rat model of LKP in anterior and posterior lamellar keratoplasty (LKP) and to observe the fine growth and histological characteristics of corneal epithelium. Methods: Twenty-seven Wistar rats (54 eyes) were selected as donors to prepare corneal grafts of anterior plate (partial stromal layer of anterior elastic layer) and posterior plate (partial stromal layer of posterior elastic layer), and 108 female SD rats were divided into 2 groups, 54 rats in each group. The posterior plate of the experimental group was treated with LKP and the control group with the anterior plate of LKP.The two groups of rats underwent LKP operation on the right eye. LKP was performed with porcine Fibrin Sealant Sealant instead of corneal suture, and eyelid margin suture was performed in both groups. The eyelid suture was removed on the 1st day after operation in both groups. 0.5% levofloxacin eye drops and tobramycin dexamethasone eye drops were used 4 times a day. Eye drops were given twice a day later and stopped after January. The patients in the two groups were observed under slit lamp for the score of rejection index (RI) on the 3rd and 56th day after operation. Cornea defect was observed by fluorescein sodium staining and photoshop software was used to calculate the corneal epithelial growth area. Nine rats of the experimental group and the control group were killed at each observation time point. The morphology of corneal epithelial cells was observed by transmission electron microscope (TEM), and keratin expression was detected by keratin expression with keratinocyte K12 immunofluorescence assay at the same time. Results: during the experiment, 14 SD rats died of excessive anaesthesia, and 7 rats were infected with grafts after operation. All 108 SD rats were removed and all entered the final result analysis. Observation under slit lamp: the process of turbid edema of corneal grafts in the experimental group and control group was the same, and the RI curve almost overlapped. The peak value and the trend of the whole curve are roughly the same. The overall RI index showed a downward trend. Fluorescein sodium staining: the two groups had the same color range reduction time. Photoshop software assisted the analysis of corneal epithelial growth area observed that there was little difference in corneal epithelial crawling area between the two groups at different time points after operation. RI and Fls score data and epithelial growth area, Using two samples t test, the data of aq.05 and p0.05 were statistically significant. The results showed that the difference was not statistically significant at each time point. The HE staining sections were observed under light microscope: after operation, both groups grew from atypical early epithelial growth to late epithelial structure, and the results showed that there was no significant difference between the two groups in the early stage of epithelium growth and in the late stage of epithelium structure. The morphology was close to normal epithelial cells. Transmission electron microscope observation: the microvilli of corneal epidermis cells in the early stage were sparse, and the structure of the five layers of epithelial cells grew up in the late stage. At the same time, there was a gap between the basal membrane of corneal epithelium and the posterior elastic layer. In the two groups, the epithelial cells were detected by immunofluorescence from early weak fluorescence to late strong fluorescence. ConclusionThe LKP model of seamless anterior and posterior plate of allograft in rats was established for the first time. 2. The rate, area and morphology of epithelial crawling at different time points in the two groups were approximately the same, which proved that the posterior plate could also be used as the carrier of corneal epithelial cell growth. It provides the experimental and theoretical basis for the further application of the back plate. 3. The difference of ultrastructure in corneal epithelial transition after LKP was observed by transmission electron microscope, which provided an important reference for further scientific research. 4. Porcine fibrin adhesives were successfully used in lamellar keratoplasty of rats, which provided experimental basis for further application of seamless keratoplasty.
【學位授予單位】：南昌大學
【學位級別】：碩士
【學位授予年份】：2012
【分類號】：R779.65

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