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研究生(外文):Chen-Yu Lin
論文名稱(外文):Characterization of Oral Precancerous Lesions Based on Higher Harmonic Generation Microscopy
指導教授(外文):Chi-Kuang Sun
口試委員(外文):Yih-Leong ChangKung-Bin SungDar-Bin Shieh
外文關鍵詞:Harmonic generation microscopyNon-invasiveOral precancerous lesion
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Oral cancer is the fourth most common cause of male cancer death in Taiwan and ranks first among the cancers that affect the young male population. About 5,400 new oral cancer cases are reported in Taiwan each year. However, the five-year survival rates of oral cancer patients are still low. The only way to address this problem is early diagnosis. In the oral cancer diagnosis, surgical biopsy followed by pathological exam is the standard operating procedure. But, surgical biopsy is always accompanied by pain, bleeding, trauma, and spreading of cancer cells. Thus, developing a non-invasive diagnosis tool is relatively important.
In this research, we used a noninvasive harmonic generation microscope to observe the ex vivo oral mucosa of 23 precancerous specimens, including leukoplakia and erythroplakia. In all of these specimens, there were 4 specimens excluded in our analysis due to the fact that we could not obtain the whole epithelium imaging of these samples.
According to pathological diagnosis results, we separated 19 samples to carcinoma in situ, severe dysplasia, moderate dysplasia, mild dysplasia, and hyperplasia. We compared the five groups with their H&E stained slides. Moreover, based on the histopathological diagnosis criteria, we further analyzed the cellular density at one-third and two-third of epithelium to quantify the difference between the oral dysplasia lesions. In this analysis, we grouped the carcinoma in situ and severe dysplasia as severe dysplasia. The cellular density of severe dysplasia lesions had a significant increase comparing with other lesions at one third of oral epithelium (P value <0.05). In the two third of epithelium, the cellular density of moderate dysplasia also had statistically difference comparing with mild dysplasia and hyperplasia (P value<0.05).
This result showed that we could distinguish the severe dysplasia and moderate dysplasia from all of precancerous lesions. We then analyzed the changes of cellular density versus depth. Additionally, we also calculated the changes in cellular density versus depth. The results of this analysis indicated that the cellular density of severe dysplasia had a marked increase at first 60μm depth. For these samples, we could not define the one-third and two-third of the epithelium; however, we could still analyze the trend of cellular density versus depth to separate severe/carcinoma in situ lesions from other lesions.
These results indicated that HGM could be a broad potential application to precancerous diagnosis, intraoperative margin, and prognosis.

致謝 I
摘要 IV
Chapter 1 Introduction 1
1.1 Motivation 1
1.2 Optical imaging in oral cavity over view 2
1.3 Structure of oral epithelium 5
1.3.1 Keratinized epithelium 6
1.3.2 Non-keratinized epithelium 7
1.4 Oral precancerous lesion 7
1.4.1 Leukoplakia 8
1.4.2 Erythroplakia 8
1.5 Thesis aim and organization 9
Chapter 2 Basic principles and properties of harmonic generation microscopy 12
2.1 Conventional optical microscopy 12
2.2 Confocal microscopy 16
2.3 Nonlinear Optical Microscopy 18
2.3.1 Second harmonic generation (SHG) 20
2.3.2 Third harmonic generation (THG) 22
Chapter 3 Experimental Setup 27
3.1 Choosing laser source 27
3.2 HGM system 30
3.3 Sample preparation 32
Chapter 4 HGM Study on oral precancerous lesion 33
4.1 Samples 33
4.2 Ex vivo HGM imaging of dysplasia lesion 34
4.3 Ex vivo HGM imaging of early stage OSCC 47
4.4 Cellular density analysis 50
4.4.1 Analysis method 50
4.4.2 Results of cellular density analysis 52
Chapter 5 Summary 57
Reference 58
Appendix A 62
Appendix B 63
Appendix C 64

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