臺灣博碩士論文加值系統

在角膜的研究上，建構高解析三維影像，能為診斷角膜疾病、辨識不同角膜組織細胞形態提供更完整的資訊，像是由準確分層的角膜縱平面來計算各層厚度，以提供角膜水腫診斷參數。此外，角膜為高密度神經支配組織，角膜神經形態的改變與多種神經疾病有關，像是糖尿病在早期階段會出現外圍神經數量的減少，因此定量角膜神經形態參數為早期診斷提供重大的幫助。
在本研究中，以等向性之微米級空間解析度的可見光Ce3+:YAG自發輻射與近紅外光Ti:sapphire放大自發輻射兩套光源之全域式光學同調斷層掃描術來展示二維平面及三維立體空間中角膜各層及體外培養細胞的結構，且比較角膜各層組織在不同中心波長下的光學特性。此外，本系統具備大面積掃描能力，藉由拼接相鄰三維影像來重建大範圍的可視區域。
利用高空間解析度的優勢提出以立體影像來解決基質神經斜向生長在定量上的爭議，並且將其應用在可見光系統活體大鼠基質神經定量上，而神經形態參數定量結果為: 寬度 、厚度 且可得知基質神經為具有0.87橢圓率似皮帶狀(belt-like)結構。此外藉由可見光與近紅外光系統檢體小鼠角膜實驗，驗證兩系統在定量亞基底神經形態參數的可行性，而神經形態參數定量結果分別為: 神經密度 、直徑 、扭曲度 ；神經密度 、直徑 、扭曲度 。
在探討角膜上皮細胞的癒合、眼輪部上皮細胞與神經三者之間的關係，藉由OCT非侵入掃描、快速成像的特性，提供神經母細胞瘤細胞株(Neuro-2A cell line)與眼輪部上皮細胞共同培養實驗的高空間解析度的立體影像，且辨識多面相的細胞團塊以及和眼輪部上皮細胞結構明顯不同的神經細胞，也在Neuro-2A細胞與眼輪部上皮細胞培養於羊膜基底實驗，利用高解析度的縱平面分別辨識出Neuro-2A細胞向外延伸的神經突觸及眼輪部上皮細胞的三層細胞結構。

Construct clear delineation of corneal three-dimensional, en-face and cross sectional imaging provide more complete information to accurate diagnosis of corneal pathology and morphological identification of different corneal layers. Changes in corneal neuromorphic parameters are associated with ocular diseases. Thus, more precise technologies for analyzing innervation architecture of the cornea will be required.
Here we develop Ce3+:YAG and Ti:sapphire light source full-field optical coherence tomography (FF-OCT) systems with isotropic micron spatial resolution in en-face and cross sectional views. Our systems also provide wide area scanning to reconstruct large field of view by stitching tomograms side by side. We think of a new three-dimensional concept to deal with wide range quantitative results of oblique stromal nerve. Otherwise, we applied this method on in vivo rat stromal image and got quantitative results in corneal nerve width and thickness of 17.9(2.5) and 2.2(0.2), respectively. With high spatial resolution we could quantify more reliable neuromorphic parameters of sub-basal nerve plexus. Ex vivo mouse quantitative results of corneal nerve density, diameter and tortuosity were 28.85(1.2), 1.29(0.06), 0.029(0.002) and 9.73(2.44), 1.52(0.02), 0.014(0.005), respectively, from Ce3+:YAG and Ti:sapphire system.
OCT as a novel approach to assess cellular activity nondestructively. We observed different types cell colonies and nerve cells from Neuro-2A cell and limbal epithelial cell co-culture system. Moreover, we identify neurite of Neuro-2A cell and three layer structures in limbal epithelium on amniotic substrate.

致謝 I
摘要 II
Abstract IV
圖目錄 VII
表目錄 XII
第一章 緒論 1
第二章 Mirau-based 全域式光學同調斷層掃描術及角膜結構介紹 3
2.1 OCT基本原理 3
2.2 Mirau-based 全域式OCT系統 11
2.2.1 系統簡介 11
2.2.2 影像處理 19
2.2.3 系統橫向及縱向解析度 22
2.2.4 系統干涉效率與訊雜比實驗及模擬 25
2.3 角膜結構介紹 32
第三章 Sprague Dawley活體大鼠角膜量測與分析 35
3.1 In vivo正常大鼠角膜量測與分析 35
3.1.1 In vivo大鼠量測方法 35
3.1.2 可見光與近紅外光源掃描大鼠角膜影像結果之分析 36
3.2 大鼠角膜神經影像定量分析 46
第四章 FVB檢體小鼠角膜量測與分析 50
4.1 Ex vivo正常小鼠角膜影像量測與分析 50
4.1.1 Ex vivo小鼠樣本製備與量測方法 50
4.1.2 可見光與近紅外光源掃描小鼠角膜影像結果之分析 51
4.2 小鼠角膜神經影像定量分析 61
第五章 In vitro細胞三維高解析度影像量測 73
5.1 Neuro-2A與眼輪部上皮細胞共同培養之量測 73
5.1.1 樣本製備及量測方法 73
5.1.2 Neuro-2A與眼輪部上皮細胞共同培養影像量測結果 74
5.2 Neuro-2A細胞與羊膜基底之量測 81
5.2.1 樣本製備及量測方法 81
5.2.2 Neuro-2A細胞與羊膜基底影像量測結果 81
5.3 眼輪部上皮細胞與羊膜基底之量測 85
5.3.1 樣本製備及量測方法 85
5.3.2 眼輪部上皮細胞與羊膜基底影像量測結果 85
第六章 結論與未來展望 88
6.1 結論 88
6.2 未來展望 92
參考文獻 93
附錄 97

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