腫瘤發展初期須藉由血管進行養分輸送，腫瘤細胞也會透過血管進行轉移，因此對於腫瘤早期發展異常新生血管為重要的診斷指標。雷射散斑對比成像(Laser speckle contrst imaging, LSCI)技術可以以非破壞性方式獲得生物組織血液動態微循環訊息，包含血管型態、流速、血氧飽和濃度等重要參數，因此在此研究中我們建立了一套雷射散斑對比顯微系統，利用雙波長雷射作為照明雷射並整合於商用顯微鏡可用於小動物活體量測，並透過所建立的演算法可以即時獲得上述血管微循環參數。另外，我們也將此套雷射散斑對比顯微系統應用於探討下列主題，包含外加壓力對於組織微循環的動態效應、腫瘤生長初期微循環的變化、即時監控與評估雷射於腫瘤治療可行性、連續波與脈衝式雷射治療特性比較等。由實驗結果已驗證組織微循環在外加壓力下所造成的動態變化可由此套系統觀測，腫瘤生長初期的微循環變化也可以透過成像結果可進一步量化流速變化、血管型態變化、血氧飽和濃度變化，經由雷射治療而觀察血管中血氧濃度變化，可以達到阻斷血液中的養分輸送以抑制腫瘤生長。相較於連續式雷射治療，脈衝式雷射治療可以達到漸進治療成效避免組織過度傷害。由此論文的結果可發現此套顯微成像系統可透過多項參數活體評估小動物疾病模型，也可以用於小動物治療模型，希望未來透過微小化工程預計將可用於人體即時量測。

The initial development of tumor needs blood vessels to transport nutrient. Tumor cells divert through blood vessels. Therefore, the newborn blood vessels are important index to diagnose in regard to the abnormal development of tumor. Laser speckle contrast imaging (LSCI) is the technology which can acquire the blood circle of organism organization through undamaged way. The information of blood circle includes blood vessels situation, flow speed, and blood oxygen saturated concentration. Thus, we form the LSCI system in this research. This system combines double wavelength laser to the microscope, which can be used on the tiny organism biopsy. Through the system and the calculation, we can get the blood circle of organism organization. In addition, we will use LSCI to the following topics, includes moment effect of organism organization with additional pressure, the organism organization variation in the initial development of tumor, the feasibility of immediately monitoring and evaluating tumor treatment, and the contrast of continuous wave and pulse laser treatments’ property. The result of this research confirms that organism organization variation by additional pressure can be observed by LSCI. The organism organization variation can further quantifies the variation of flow speed, blood vessels situation, and blood oxygen saturated concentration through imaging results. The observation of blood oxygen saturated concentration in the blood vessels by LSCI, which can obstruct the nutrient transportation in the blood vessels to keep down the development of tumors. In compare with the continuous laser treatment, pulse laser treatment can achieve the effect of gradual progress treatment to keep organization from being damaged. The result of this thesis indicates that LSCI can evaluate the disease model of tiny creatures through parameters. It also can be used on model treatment of tiny creatures. In the future, hope that infinitesimal project can be immediately used on human body treatment.

[1] A. S. Popel and P. C. J. A. R. F. M. Johnson, "Microcirculation and hemorheology," vol. 37, pp. 43-69, 2005.
[2] S. J. Lunt, C. Gray, C. C. R. Aldasoro, S. J. Matcher, and G. M. J. J. o. b. o. Tozer, "Application of intravital microscopy in studies of tumor microcirculation," vol. 15, no. 1, p. 011113, 2010.
[3] G. Ilangovan, A. Bratasz, H. Li, P. Schmalbrock, J. L. Zweier, and P. J. M. R. i. M. A. O. J. o. t. I. S. f. M. R. i. M. Kuppusamy, "In vivo measurement and imaging of tumor oxygenation using coembedded paramagnetic particulates," vol. 52, no. 3, pp. 650-657, 2004.
[4] B. Dutta, R. Yan, S. K. Lim, J. P. Tam, S. K. J. M. Sze, and C. Proteomics, "Quantitative profiling of chromatome dynamics reveals a novel role for HP1BP3 in hypoxia-induced oncogenesis," vol. 13, no. 12, pp. 3236-3249, 2014.
[5] A. Bratasz et al., "In vivo imaging of changes in tumor oxygenation during growth and after treatment," vol. 57, no. 5, pp. 950-959, 2007.
[6] A. Gouliamos et al., "Magnetic resonance angiography compared to intra-arterial digital subtraction angiography in patients with subarachnoid haemorrhage," vol. 35, no. 1, pp. 46-49, 1992.
[7] E. Z. Kapsalaki, C. D. Rountas, and K. N. J. I. j. o. v. m. Fountas, "The role of 3 Tesla MRA in the detection of intracranial aneurysms," vol. 2012, 2012.
[8] P. H. Tomlins and R. J. J. o. P. D. A. P. Wang, "Theory, developments and applications of optical coherence tomography," vol. 38, no. 15, p. 2519, 2005.
[9] T. E. De Carlo, A. Romano, N. K. Waheed, J. S. J. I. j. o. r. Duker, and vitreous, "A review of optical coherence tomography angiography (OCTA)," vol. 1, no. 1, p. 5, 2015.
[10] U. Baran and R. K. J. N. Wang, "Review of optical coherence tomography based angiography in neuroscience," vol. 3, no. 1, p. 010902, 2016.
[11] J. Hsieh, "Computed tomography: principles, design, artifacts, and recent advances," 2009: SPIE Bellingham, WA.
[12] G. L. Raff, M. J. Gallagher, W. W. O’Neill, and J. A. J. J. o. t. A. C. o. C. Goldstein, "Diagnostic accuracy of noninvasive coronary angiography using 64-slice spiral computed tomography," vol. 46, no. 3, pp. 552-557, 2005.
[13] E. A. Hulten, S. Carbonaro, S. P. Petrillo, J. D. Mitchell, and T. C. J. J. o. t. A. C. o. C. Villines, "Prognostic value of cardiac computed tomography angiography: a systematic review and meta-analysis," vol. 57, no. 10, pp. 1237-1247, 2011.
[14] E. Frampas et al., "CT angiography for brain death diagnosis," vol. 30, no. 8, pp. 1566-1570, 2009.
[15] R. F. Spaide, J. M. Klancnik, and M. J. J. J. o. Cooney, "Retinal vascular layers imaged by fluorescein angiography and optical coherence tomography angiography," vol. 133, no. 1, pp. 45-50, 2015.
[16] K. R. Mendis et al., "Correlation of histologic and clinical images to determine the diagnostic value of fluorescein angiography for studying retinal capillary detail," vol. 51, no. 11, pp. 5864-5869, 2010.
[17] D. A. Salz et al., "Select features of diabetic retinopathy on swept-source optical coherence tomographic angiography compared with fluorescein angiography and normal eyes," vol. 134, no. 6, pp. 644-650, 2016.
[18] J. W. Goodman, "Statistical properties of laser speckle patterns," in Laser speckle and related phenomena: Springer, 1975, pp. 9-75.
[19] A. Fercher and J. D. J. O. c. Briers, "Flow visualization by means of single-exposure speckle photography," vol. 37, no. 5, pp. 326-330, 1981.
[20] J. E. Krook et al., "Effective surgical adjuvant therapy for high-risk rectal carcinoma," vol. 324, no. 11, pp. 709-715, 1991.
[21] G. E. J. C. Sheline, "Radiation therapy of brain tumors," vol. 39, no. S2, pp. 873-881, 1977.
[22] N. A. Vick, J. D. Khandekar, and D. D. J. A. o. N. Bigner, "Chemotherapy of brain tumors: The blood-brain barrier is not a factor," vol. 34, no. 9, pp. 523-526, 1977.
[23] I. Bozic et al., "Evolutionary dynamics of cancer in response to targeted combination therapy," vol. 2, p. e00747, 2013.
[24] H.-J. Schwarzmaier, F. Eickmeyer, V. U. Fiedler, and F. J. M. L. A. Ulrich, "Basic principles of laser induced interstitial thermotherapy in brain tumors," vol. 17, no. 2, pp. 147-158, 2002.
[25] D. E. Dolmans, D. Fukumura, and R. K. J. N. r. c. Jain, "Photodynamic therapy for cancer," vol. 3, no. 5, p. 380, 2003.
[26] D. J. I. J. o. R. Gabor and Development, "Laser speckle and its elimination," vol. 14, no. 5, pp. 509-514, 1970.
[27] J. I. Trisnadi, "Method and apparatus for reducing laser speckle," ed: Google Patents, 2001.
[28] J. D. J. O. A. Briers, "Laser speckle contrast imaging for measuring blood flow," vol. 37, 2007.
[29] D. A. Boas and A. K. J. J. o. b. o. Dunn, "Laser speckle contrast imaging in biomedical optics," vol. 15, no. 1, p. 011109, 2010.
[30] J. Senarathna, A. Rege, N. Li, and N. V. J. I. r. i. b. e. Thakor, "Laser speckle contrast imaging: theory, instrumentation and applications," vol. 6, pp. 99-110, 2013.
[31] J. D. Briers and S. J. J. o. b. o. Webster, "Laser speckle contrast analysis (LASCA): a nonscanning, full-field technique for monitoring capillary blood flow," vol. 1, no. 2, pp. 174-180, 1996.
[32] Z. Li et al., "Calibration of diffuse correlation spectroscopy blood flow index with venous-occlusion diffuse optical spectroscopy in skeletal muscle," vol. 20, no. 12, p. 125005, 2015.
[33] D. Wen, Y. Li, X. Zhu, M. Chen, J. Lu, and P. J. O. l. Li, "Selective photoactivation of neural activity combined with laser speckle imaging of cerebral blood flow," vol. 43, no. 15, pp. 3798-3801, 2018.
[34] M. Draijer, E. Hondebrink, T. van Leeuwen, and W. J. L. i. m. s. Steenbergen, "Review of laser speckle contrast techniques for visualizing tissue perfusion," vol. 24, no. 4, p. 639, 2009.
[35] H. M. Adera, K. James, J. J. Castronuovo Jr, M. Byrne, R. Deshmukh, and J. J. J. o. v. s. Lohr, "Prediction of amputation wound healing with skin perfusion pressure," vol. 21, no. 5, pp. 823-829, 1995.
[36] K. Wardell, I. M. Braverman, D. G. Silverman, and G. E. J. M. r. Nilsson, "Spatial heterogeneity in normal skin perfusion recorded with laser Doppler imaging and flowmetry," vol. 48, no. 1, pp. 26-38, 1994.
[37] M. J. Daly and R. E. J. J. o. n. m. o. p. Henry, Society of Nuclear Medicine, "Quantitative measurement of skin perfusion with xenon-133," vol. 21, no. 2, pp. 156-160, 1980.
[38] H. Schatz, T. Burton, L. Yannuzzi, and M. J. S. L. M. Rabb, "Interpretation of fundus fluorescein angiography," pp. 550-631, 1978.
[39] R. H. Webb, G. W. Hughes, and F. C. J. A. o. Delori, "Confocal scanning laser ophthalmoscope," vol. 26, no. 8, pp. 1492-1499, 1987.
[40] W. Drexler, J. G. J. P. i. r. Fujimoto, and e. research, "State-of-the-art retinal optical coherence tomography," vol. 27, no. 1, pp. 45-88, 2008.
[41] H. Isono, S. Kishi, Y. Kimura, N. Hagiwara, N. Konishi, and H. J. A. o. O. Fujii, "Observation of choroidal circulation using index of erythrocytic velocity," vol. 121, no. 2, pp. 225-231, 2003.
[42] M. Nagahara, Y. Tamaki, M. Araie, and H. J. J. j. o. o. Fujii, "Real-time blood velocity measurements in human retinal vein using the laser speckle phenomenon," vol. 43, no. 3, pp. 186-195, 1999.
[43] K. Khaksari and S. J. J. J. o. B. o. Kirkpatrick, "Combined effects of scattering and absorption on laser speckle contrast imaging," vol. 21, no. 7, p. 076002, 2016.
[44] E. Zarahn, G. K. Aguirre, and M. J. N. D'Esposito, "Empirical analyses of BOLD fMRI statistics," vol. 5, no. 3, pp. 179-197, 1997.
[45] J. Mayhew, D. Johnston, J. Berwick, M. Jones, P. Coffey, and Y. J. N. Zheng, "Spectroscopic analysis of neural activity in brain: increased oxygen consumption following activation of barrel cortex," vol. 12, no. 6, pp. 664-675, 2000.
[46] A. K. Dunn et al., "Simultaneous imaging of total cerebral hemoglobin concentration, oxygenation, and blood flow during functional activation," vol. 28, no. 1, pp. 28-30, 2003.
[47] T. Durduran et al., "Spatiotemporal quantification of cerebral blood flow during functional activation in rat somatosensory cortex using laser-speckle flowmetry," vol. 24, no. 5, pp. 518-525, 2004.
[48] B. Weber, C. Burger, M. Wyss, G. Von Schulthess, F. Scheffold, and A. J. E. J. o. N. Buck, "Optical imaging of the spatiotemporal dynamics of cerebral blood flow and oxidative metabolism in the rat barrel cortex," vol. 20, no. 10, pp. 2664-2670, 2004.
[49] S. Sakadžić et al., "Simultaneous imaging of cerebral partial pressure of oxygen and blood flow during functional activation and cortical spreading depression," vol. 48, no. 10, pp. D169-D177, 2009.
[50] A. A. J. J. o. n. Leao, "Spreading depression of activity in the cerebral cortex," vol. 7, no. 6, pp. 359-390, 1944.
[51] M. J. B. Lauritzen, "Pathophysiology of the migraine aura: the spreading depression theory," vol. 117, no. 1, pp. 199-210, 1994.
[52] G. G. J. P. r. Somjen, "Mechanisms of spreading depression and hypoxic spreading depression-like depolarization," vol. 81, no. 3, pp. 1065-1096, 2001.
[53] A. K. Dunn, H. Bolay, M. A. Moskowitz, D. A. J. J. o. C. B. F. Boas, and Metabolism, "Dynamic imaging of cerebral blood flow using laser speckle," vol. 21, no. 3, pp. 195-201, 2001.
[54] H. Bolay, U. Reuter, A. K. Dunn, Z. Huang, D. A. Boas, and M. A. J. N. m. Moskowitz, "Intrinsic brain activity triggers trigeminal meningeal afferents in a migraine model," vol. 8, no. 2, p. 136, 2002.
[55] H. K. Shin et al., "Vasoconstrictive neurovascular coupling during focal ischemic depolarizations," vol. 26, no. 8, pp. 1018-1030, 2006.
[56] S. Zhang and T. H. J. P. b. Murphy, "Imaging the impact of cortical microcirculation on synaptic structure and sensory-evoked hemodynamic responses in vivo," vol. 5, no. 5, p. e119, 2007.
[57] D. N. Atochin et al., "Mouse model of microembolic stroke and reperfusion," vol. 35, no. 9, pp. 2177-2182, 2004.
[58] 李承育，"開發雷射散斑對比成像顯微平台於活體小動物研究"，碩士論文，長庚大學電機電機工程研究所，2018。
[59] J. E. J. T. A. j. o. e. m. Sinex, "Pulse oximetry: principles and limitations," vol. 17, no. 1, pp. 59-66, 1999.