使用結構照明顯微術觀察活體小鼠毛囊生長週期之變化

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黃建翔(Chien-Hsiang Huang) 查詢紙本館藏

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光電科學與工程學系

論文名稱

使用結構照明顯微術觀察活體小鼠毛囊生長週期之變化

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摘要(中)

在生醫研究上使用光學顯微系統觀察生物樣本的時候，需要能夠快速並且穩定的獲取即時資訊以及提供樣本的完整性，而結構照明顯微術 (Structured Illumination Microscopy, SIM) 除了能夠提供光學切片的功能和有著不錯的影像對比度之外，還可以穩定並且快速的取得即時影像，因此成為了一種能夠提供生醫研究上觀察樣本的工具。
以往使用光學顯微系統觀察毛囊的不同生長週期的型態時，需要將許多隻經過密蠟除毛後的小鼠在不同的毛囊生長週期時間上，將其背部一部份的皮膚切下並且犧牲掉小鼠，接下來再把皮膚毛囊幹細胞染上不同的螢光標記並做成樣本，藉此來觀察不同時間毛囊的生長週期狀態，但是這樣的實驗過程中沒有辦法觀察由同一隻小鼠毛囊生長周期變化的連續性，而且樣本染色的成功與否也會影響觀察的方向性和正確性，最後小鼠的犧牲也會造成小鼠資源浪費並且無法再次利用。
本論文以綠螢光蛋白(Green Fluorescent Protein, GFP)基因轉殖的小鼠做為樣本，其背部經過密蠟除毛使所有毛囊同步進入毛囊生長的休止期，利用以數位微型投影機 (Digital Light Projector, DLP) 做為系統光源的結構造明顯微系統來進行活體小鼠的觀察，實驗將持續一整個毛囊生長週期的循環時間觀察毛囊生長周期過程中不同的型態樣貌。

摘要(英)

In biomedical research, when using microscope system to observe biological sample, it is necessary to have fast and stable data acquisition with completeness of samples. Structured illumination microscopy not only can provide optical sectioning and nice image contrast, but also capture image fast and stable. These advantages make structured illumination microscopy become a popularity-used system for biomedical research.
When observing hair follicles with optical microscopy system in the past, different mice have to be waxed and sacrificed to get back skin samples at different stages of the hair growth cycle. The hair follicle stem cells of the samples should be strained with dye of fluorophores to enhance their contrast for observation. However, with these process, the observation can-not be focused on the same mouse in a complete growth cycle and the staining process plays a key role in the sample completeness and data accuracy.
In this paper, green fluorescence protein (GFP) transgenic mice were used as the specimens. The mice were waxed to synchronize all hair follicles into anagen state of the growth cycle. By using the structured illumination microscopy system based on a digital light projector to observe the mice hair follicle in vivo. The observation will be last for a complete hair follicle growth cycle to long-term monitior the hair follicle stem cell morphology at different stages of the growth cycle.

關鍵字(中)

★ 結構照明顯微術
★ 小鼠
★ 毛囊生長週期

關鍵字(英)

論文目次

中文摘要.............................................................................................................. II
Abstract.............................................................................................................. III
目錄.................................................................................................................... IV
圖目錄............................................................................................................... VII
表目錄.................................................................................................................. X
第一章緒論...................................................................................................... 1
1-1研究目的與動機.......................................................................................... 1
1-2毛囊構造及生長週期................................................................................. 3
1-2-1皮膚構造............................................................................................... 3
1-2-2 毛髮及毛囊結構.................................................................................. 4
1-2-3 毛囊幹細胞.......................................................................................... 5
1-2-4 毛囊生長週期...................................................................................... 7
1-3毛囊研究文獻回顧.................................................................................... 10
第二章基本理論............................................................................................... 12
2-1光學顯微術................................................................................................ 12
2-2具有光學切片能力的顯微術................................................................... 12
2-3結構照明顯微術....................................................................................... 16
2-3-1發展及基本觀念................................................................................. 16
2-3-2成像理論推導..................................................................................... 18
2-3-3延伸發展介紹..................................................................................... 20
第三章實驗方法.............................................................................................. 22
3-1實驗架構................................................................................................... 22
3-1-1 結構照明顯微術系統架構................................................................ 22
3-1-2 數位微型投影機................................................................................ 24
3-1-3 二維光偵測器.................................................................................... 26
3-1-4 影像擷取控制.................................................................................... 26
3-1-5分光鏡選擇......................................................................................... 27
3-1-6系統解析度分析................................................................................. 28
3-2 條紋型式.................................................................................................. 32
3-3樣本準備................................................................................................... 35
3-3-1切片樣本............................................................................................. 35
3-3-2活體樣本............................................................................................. 36
第四章實驗結果............................................................................................... 38
4-1切片樣本................................................................................................... 38
4-1-1以螢光標定CK14的切片樣本......................................................... 38
4-1-2以螢光標定CD34的切片樣本......................................................... 39
4-1-3以螢光標定CK14及CD34的Whole Mount 樣本...................... 41
4-2 GFP螢光小鼠毛囊生長之活體觀察...................................................... 43
4-2-1 第1到第8天................................................................................... 44
4-2-2 第11天到第17天............................................................................ 48
4-2-3 第20到第23天................................................................................ 53
4-3 毛囊整體比較.......................................................................................... 56
4-3-1 膚色.................................................................................................... 56
4-3-2 毛根及毛髮型態................................................................................ 57
4-3-3 毛根、髮幹以及毛囊孔洞大小........................................................ 60
第五章結論....................................................................................................... 63
參考文獻............................................................................................................. 65

參考文獻

[1]E. A. Olsen, "Female pattern hair loss," Journal of the American Academy of Dermatology, vol. 45, pp. S70-S80, 2001.
[2]F. C. Rossetti, L. V. Depieri, and M. V. L. B. Bentley, Confocal laser scanning microscopy as a tool for the investigation of skin drug delivery systems and diagnosis of skin disorders: INTECH Open Access Publisher, 2013.
[3]R. DasGupta and E. Fuchs, "Multiple roles for activated LEF/TCF transcription complexes during hair follicle development and differentiation," Development, vol. 126, pp. 4557-4568, 1999.
[4]B. Lehner, B. Sandner, J. Marschallinger, C. Lehner, T. Furtner, S. Couillard-Despres, et al., "The dark side of BrdU in neural stem cell biology: detrimental effects on cell cycle, differentiation and survival," Cell and tissue research, vol. 345, pp. 313-328, 2011.
[5]T. Schepeler, M. E. Page, and K. B. Jensen, "Heterogeneity and plasticity of epidermal stem cells," Development, vol. 141, pp. 2559-2567, 2014.
[6]K. Stenn and R. Paus, "Controls of hair follicle cycling," Physiological reviews, vol. 81, pp. 449-494, 2001.
[7]P. Rompolas, E. R. Deschene, G. Zito, D. G. Gonzalez, I. Saotome, A. M. Haberman, et al., "Live imaging of stem cell and progeny behaviour in physiological hair-follicle regeneration," Nature, vol. 487, pp. 496-499, 2012.
[8]A. Uchugonova, R. M. Hoffman, M. Weinigel, and K. Koenig, "Watching stem cells in the skin of living mice noninvasively," Cell Cycle, vol. 10, pp. 2017-2020, 2011.
[9]M. Ito, K. Kizawa, K. Hamada, and G. Cotsarelis, "Hair follicle stem cells in the lower bulge form the secondary germ, a biochemically distinct but functionally equivalent progenitor cell population, at the termination of catagen," Differentiation, vol. 72, pp. 548-557, 2004.
[10]V. Greco, T. Chen, M. Rendl, M. Schober, H. A. Pasolli, N. Stokes, et al., "A two-step mechanism for stem cell activation during hair regeneration," Cell stem cell, vol. 4, pp. 155-169, 2009.
[11]Y. V. Zhang, J. Cheong, N. Ciapurin, D. J. McDermitt, and T. Tumbar, "Distinct self-renewal and differentiation phases in the niche of infrequently dividing hair follicle stem cells," Cell stem cell, vol. 5, pp. 267-278, 2009.
[12]E. Legue and J.-F. Nicolas, "Hair follicle renewal: organization of stem cells in the matrix and the role of stereotyped lineages and behaviors," Development, vol. 132, pp. 4143-4154, 2005.
[13]R. Kopan, J. Lee, M.-H. Lin, A. J. Syder, J. Kesterson, N. Crutchfield, et al., "Genetic mosaic analysis indicates that the bulb region of coat hair follicles contains a resident population of several active multipotent epithelial lineage progenitors," Developmental biology, vol. 242, pp. 44-57, 2002.
[14]H. Kulessa, G. Turk, and B. L. Hogan, "Inhibition of Bmp signaling affects growth and differentiation in the anagen hair follicle," The EMBO journal, vol. 19, pp. 6664-6674, 2000.
[15]J. Kamimura, D. Lee, H. P. Baden, J. Brissette, and G. P. Dotto, "Primary mouse keratinocyte cultures contain hair follicle progenitor cells with multiple differentiation potential," Journal of Investigative Dermatology, vol. 109, pp. 534-540, 1997.
[16]S. Ghazizadeh and L. B. Taichman, "Multiple classes of stem cells in cutaneous epithelium: a lineage analysis of adult mouse skin," The EMBO journal, vol. 20, pp. 1215-1222, 2001.
[17]E. Legue, I. Sequeira, and J.-F. Nicolas, "Hair follicle renewal: authentic morphogenesis that depends on a complex progression of stem cell lineages," Development, vol. 137, pp. 569-577, 2010.
[18]Y. Liu, S. Lyle, Z. Yang, and G. Cotsarelis, "Keratin 15 promoter targets putative epithelial stem cells in the hair follicle bulge," Journal of Investigative Dermatology, vol. 121, pp. 963-968, 2003.
[19]C. S. Trempus, R. J. Morris, C. D. Bortner, G. Cotsarelis, R. S. Faircloth, J. M. Reece, et al., "Enrichment for living murine keratinocytes from the hair follicle bulge with the cell surface marker CD34," Journal of Investigative Dermatology, vol. 120, pp. 501-511, 2003.
[20]K. K. Youssef, A. Van Keymeulen, G. Lapouge, B. Beck, C. Michaux, Y. Achouri, et al., "Identification of the cell lineage at the origin of basal cell carcinoma," Nature cell biology, vol. 12, pp. 299-305, 2010.
[21]V. Jaks, N. Barker, M. Kasper, J. H. Van Es, H. J. Snippert, H. Clevers, et al., "Lgr5 marks cycling, yet long-lived, hair follicle stem cells," Nature genetics, vol. 40, pp. 1291-1299, 2008.
[22]J. A. Nowak, L. Polak, H. A. Pasolli, and E. Fuchs, "Hair follicle stem cells are specified and function in early skin morphogenesis," Cell stem cell, vol. 3, pp. 33-43, 2008.
[23]Y.-C. Hsu, H. A. Pasolli, and E. Fuchs, "Dynamics between stem cells, niche, and progeny in the hair follicle," Cell, vol. 144, pp. 92-105, 2011.
[24]I. Brownell, E. Guevara, C. B. Bai, C. A. Loomis, and A. L. Joyner, "Nerve-derived sonic hedgehog defines a niche for hair follicle stem cells capable of becoming epidermal stem cells," Cell stem cell, vol. 8, pp. 552-565, 2011.
[25]M. E. Page, P. Lombard, F. Ng, B. Gottgens, and K. B. Jensen, "The epidermis comprises autonomous compartments maintained by distinct stem cell populations," Cell Stem Cell, vol. 13, pp. 471-482, 2013.
[26]S. Claudinot, M. Nicolas, H. Oshima, A. Rochat, and Y. Barrandon, "Long-term renewal of hair follicles from clonogenic multipotent stem cells," Proceedings of the National Academy of Sciences of the United States of America, vol. 102, pp. 14677-14682, 2005.
[27]M. Ito, Y. Liu, Z. Yang, J. Nguyen, F. Liang, R. J. Morris, et al., "Stem cells in the hair follicle bulge contribute to wound repair but not to homeostasis of the epidermis," Nature medicine, vol. 11, pp. 1351-1354, 2005.
[28]V. Levy, C. Lindon, Y. Zheng, B. D. Harfe, and B. A. Morgan, "Epidermal stem cells arise from the hair follicle after wounding," The FASEB Journal, vol. 21, pp. 1358-1366, 2007.
[29]F. Liu, A. Uchugonova, H. Kimura, C. Zhang, M. Zhao, L. Zhang, et al., "The bulge area is the major hair follicle source of nestin-expressing pluripotent stem cells which can repair the spinal cord compared to the dermal papilla," Cell Cycle, vol. 10, pp. 830-839, 2011.
[30]M. Minsky, "Microscopy apparatus," ed: Google Patents, 1961.
[31]W. Denk, J. H. Strickler, and W. W. Webb, "Two-photon laser scanning fluorescence microscopy," Science, vol. 248, pp. 73-76, 1990.
[32]P. J. Keller, A. D. Schmidt, J. Wittbrodt, and E. H. Stelzer, "Reconstruction of zebrafish early embryonic development by scanned light sheet microscopy," science, vol. 322, pp. 1065-1069, 2008.
[33]P. T. So, C. Y. Dong, B. R. Masters, and K. M. Berland, "Two-photon excitation fluorescence microscopy," Annual review of biomedical engineering, vol. 2, pp. 399-429, 2000.
[34]J. Huisken and D. Y. Stainier, "Selective plane illumination microscopy techniques in developmental biology," Development, vol. 136, pp. 1963-1975, 2009.
[35]W. Lukosz, "Optical systems with resolving powers exceeding the classical limit," JOSA, vol. 56, pp. 1463-1471, 1966.
[36]W. Lukosz, "Optical systems with resolving powers exceeding the classical limit. II," JOSA, vol. 57, pp. 932-941, 1967.
[37]M. Saxena, G. Eluru, and S. S. Gorthi, "Structured illumination microscopy," Advances in Optics and Photonics, vol. 7, pp. 241-275, 2015.
[38]M. Neil, R. Ju?kaitis, and T. Wilson, "Method of obtaining optical sectioning by using structured light in a conventional microscope," Optics letters, vol. 22, pp. 1905-1907, 1997.
[39]L. M. Hirvonen, K. Wicker, O. Mandula, and R. Heintzmann, "Structured illumination microscopy of a living cell," European Biophysics Journal, vol. 38, pp. 807-812, 2009.
[40]R. Fiolka, M. Beck, and A. Stemmer, "Structured illumination in total internal reflection fluorescence microscopy using a spatial light modulator," Optics letters, vol. 33, pp. 1629-1631, 2008.
[41]Q. Song, K. Isobe, F. Kannari, H. Kawano, A. Kumagai, A. Miyawaki, et al., "Multiphoton 3D structured illumination microscopy for enhanced axial resolution in deep imaging," in Conference on Lasers and Electro-Optics/Pacific Rim, 2015, p. 27H2_2.
[42]P. W. Winter, P. Chandris, R. S. Fischer, Y. Wu, C. M. Waterman, and H. Shroff, "Incoherent structured illumination improves optical sectioning and contrast in multiphoton super-resolution microscopy," Optics express, vol. 23, pp. 5327-5334, 2015.
[43]R. Fiolka, L. Shao, E. H. Rego, M. W. Davidson, and M. G. Gustafsson, "Time-lapse two-color 3D imaging of live cells with doubled resolution using structured illumination," Proceedings of the National Academy of Sciences, vol. 109, pp. 5311-5315, 2012.
[44]N. Bozinovic, C. Ventalon, T. Ford, and J. Mertz, "Fluorescence endomicroscopy with structured illumination," Optics express, vol. 16, pp. 8016-8025, 2008.
[45]D. Xu, T. Jiang, A. Li, B. Hu, Z. Feng, H. Gong, et al., "Fast optical sectioning obtained by structured illumination microscopy using a digital mirror device," Journal of biomedical optics, vol. 18, pp. 060503-060503, 2013.
[46]D. Dan, M. Lei, B. Yao, W. Wang, M. Winterhalder, A. Zumbusch, et al., "DMD-based LED-illumination Super-resolution and optical sectioning microscopy," Scientific reports, vol. 3, 2013.
[47](2012). DLPR LightCrafter? Evaluation Module (EVM) User′s Guide.
[48]J. A. Rodrigo and T. Alieva, "Fast control of temporal and spatial coherence properties of microscope illumination using DLP projector," in SPIE BiOS, 2015, pp. 93360F-93360F-6.
[49]N. Chakrova, B. Rieger, and S. Stallinga, "Development of a DMD-based fluorescence microscope," in SPIE BiOS, 2015, pp. 933008-933008-11.
[50]"measured LightCrafter LED spectra," 2014.
[51]ORCA-Flash4.0 V2 Digital CMOS camera C11440-22CU.
[52]Semrock 515 nm blocking edge BrightLineR long-pass filter.
[53]Semrock 532 nm EdgeBasic? best-value long-pass edge filter.
[54]D. Karadagli? and T. Wilson, "Image formation in structured illumination wide-field fluorescence microscopy," Micron, vol. 39, pp. 808-818, 2008.
[55]Thermal Fisher Scientific Fluorescence Spectraviewer.
[56]徐鈺, "DLP-Based Hyperspectral Imaging via Optical Sectioning Microscopy," 2016.

指導教授

陳思妤(Szu-Yu Chen)

審核日期

2017-1-24

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