氣喘(Asthma)是一種下呼道發炎反應疾病，目前對於氣喘的實際致病原仍然不清楚。醫學研究上難以直接進行人體內的氣喘機制實驗研究，在體外重建類似於人體組織功能與微環境能使研究更方便。
關於氣喘體外細胞研究中，大多數研究團隊著重於探討氣喘發病時支氣管平滑肌收縮情況下所產生的反應。在本研究中，我們利用微機電技術製作一個微流體晶片，在體外重建支氣管的構造與環境，致力於研究呼吸過程產生拉伸現象對於氣道內細胞所產生的影響，探討氣喘潛在的致病原因。
此研究設計一個簡易拉伸系統與微流體晶片搭配，使培養於微流體晶片內的細胞受到類似於人體呼吸時的拉伸環境，此拉伸系統不用複雜操作與龐大儀器，組成為3D列印元件搭配一個伺服馬達。
實驗結果顯示人類平滑肌在我們晶片灌流培養下四天細胞激素介白素-6的濃度是427.5pg/ml，高於孔盤靜置培養11.05 pg/ml，本研究設計的拉伸系統能夠使細胞在微流體晶片中受到10%伸長量拉伸，拉伸頻率為每分鐘10次和30次，類似正常呼吸和氣喘發作的呼吸頻率，拉伸會造成細胞重新排列，每分鐘10次拉伸作用三天後，細胞角度從未拉伸平均60.93度(標準差15.43)到三天後平均85.84度(標準差1.66)。氣喘用藥格隆溴銨(glycopyrronium bromide)濃度5nM作用在受每分鐘30次拉伸的平滑肌細胞，有效降低發炎物質介白素6分泌，從三天增加82.4% (未給藥)降至三天增加4.9%，介白素8 濃度從三天增加229.7%降至三天增加39.1%。我們成功在體外重建類似氣喘發作時的支氣管平滑肌發炎模型，並藉由臨床常見的氣喘治療藥物抑制發炎反應。

Asthma is a disease about lower respiratory tract inflammation. The actual pathogen of asthma is still unclear, because it is difficult to do the experiments inside human body. Thus, many state-of-art bioengineering technology and studies focus on rebuilding tissue structure and function of human in vitro.
These years, many top research groups focused their research dealing with asthma studies on the contraction responses of bronchial smooth muscle cells in vitro. In our study, we used microelectromechanical techniques to develop a microfluidic chip to rebuild the structure and micro environment of the bronchium in vitro. We studied the responses of bronchial smooth muscle cells under regularly stretching and investigated the pathogen of asthma.
In this master program study, the interleukin 6 concentrations of human bronchial smooth muscle cell (BSMC) were characterized as 427.5pg/ml in our microfluidic chip, 11.05pg/ml in 96 well culture plate for 4 days culture. We developed a stretching system for our microfluidic chip. The stretching system could give the cells 10% elongation 10 times per minute and 30 per minute. The frequencies mimicked the conditions of normal breathing and under asthma. The cell stretching changed the cells alignment. After giving 10% elongation 10 times per minute stretching for 3 days, the cell angel of BSMC varied from 60.93 degree (standard deviation 15.3) to 85.84 degree (standard deviation 1.66). We treated BSMC with 5nM glycopyrronium bromide, the drug for asthma, at 30 times stretching per minute. It decreases the secretion of interleukin 6 and interleukin 8. The interleukin 6 concentration increases 82.4% without drug treatment and 4.9% at drug treatment in 3 days. The interleukin 8 concentration increases 229.7% without drug treatment and 39.1% at drug treatment in 3 days. We successfully rebuilt a bronchial smooth muscle inflammation model in vitro and demonstrated the drug treatment.

[1] 吳家興, 林瑞雄, 謝貴雄, 邱文達, 陳麗美, and 邱淑媞, "台灣北區國中學生 氣喘盛行率調查," 中華公共衛生雜誌, vol. 17, pp. 214-225, 1998.
[2] Andrew Harver and H. Kotses, Asthma, health and society a public health perspective. New York: Springer, 2010.
[3] H. A. Werner, "Status asthmaticus in children - A review," Chest, vol. 119, pp. 1913-1929, Jun 2001.
[4] J. L. Wright, M. Cosio, and A. Churg, "Animal models of chronic obstructive pulmonary disease," American Journal of Physiology-Lung Cellular and Molecular Physiology, vol. 295, pp. L1-L15, Jul 2008.
[5] Wikipedia. lung.
https://en.wikipedia.org/wiki/Lung
[6] wikiwand. Respiratory tract.
http://www.wikiwand.com/en/Respiratory_tract
[7] Elward, G. Douglas, and K. S, Asthma. London: Manson Pub.
[8] Guidelines for the Diagnosis and Management of Asthma, 2007.
[9] L. United States-National Institute of Health: National Heart, Blood Institute What Is Asthma? http://www.nhlbi.nih.gov/health/health-topics/topics/asthma/
[10] K. G. Birukov, J. R. Jacobson, A. A. Flores, S. Q. Ye, A. A. Birukova, A. D. Verin, et al., "Magnitude-dependent regulation of pulmonary endothelial cell barrier function by cyclic stretch," American Journal of Physiology-Lung Cellular and Molecular Physiology, vol. 285, pp. L785-L797, Oct 2003.
[11] D. Huh, B. D. Matthews, A. Mammoto, M. Montoya-Zavala, H. Y. Hsin, and D. E. Ingber, "Reconstituting Organ-Level Lung Functions on a Chip," Science, vol. 328, pp. 1662-1668, Jun 2010.
[12] S. J. Gunst and M. F. Wu, "Selected contribution: Plasticity of airway smooth muscle stiffness and extensibility: role of length-adaptive mechanisms," Journal of Applied Physiology, vol. 90, pp. 741-749, Feb 2001.
[13] T. G. Klingele, "Terminal bronchiole diameter changes with volume in isolated, air-filled lobes of cat lung," Journal of Applied Physiology, vol. 30, pp. 224-+, 1971.
[14] J. M. B. Hughes, F. G. Hoppin, and J. Mead, "Effect of lung inflation on bronchial length and diameter in excised lungs," Journal of Applied Physiology, vol. 32, pp. 25-&, 1972.
[15] H. L. Hahn, A. Watson, A. G. Wilson, and N. B. Pride, "Influence of bronchomotor tone on airway dimensions and resistance in excised dog lungs," Journal of Applied Physiology, vol. 49, pp. 270-278, 1980.
[16] N. Lechtzin. Control of Breathing.
http://www.merckmanuals.com/home/lung-and-airway-disorders/biology-of-the-lungs-and-airways/control-of-breathing
[17] K. E. Barrett, S. Boitano, S. M. Barman, and H. L. Brooks, Ganong's Review of Medical Physiology, 24 ed. United States The McGraw-Hill Companies, 1976.
[18] D. Huh, D. C. Leslie, B. D. Matthews, J. P. Fraser, S. Jurek, G. A. Hamilton, et al., "A human disease model of drug toxicity-induced pulmonary edema in a lung-on-a-chip microdevice," Science Translational Medicine, vol. 4, Nov 2012.
[19] K. H. Benam, R. Villenave, C. Lucchesi, A. Varone, C. Hubeau, H. H. Lee, et al., "Small airway-on-a-chip enables analysis of human lung inflammation and drug responses in vitro," Nature Methods, vol. 13, pp. 151-+, Feb 2016.
[20] K. H. Benam, R. Novak, J. Nawroth, M. Hirano-Kobayashi, T. C. Ferrante, Y. Choe, et al., "Matched-comparative modeling of normal and diseased human airway responses using a microengineered breathing lung chip," Cell Systems, vol. 3, pp. 456-+, Nov 2016.
[21] T. H. Punde, W. H. Wu, P. C. Lien, Y. L. Chang, P. H. Kuo, M. D. T. Chang, et al., "A biologically inspired lung-on-a-chip device for the study of protein-induced lung inflammation," Integrative Biology, vol. 7, pp. 162-169, 2015.
[22] A. T. Nials and S. Uddin, "Mouse models of allergic asthma: acute and chronic allergen challenge," Disease Models & Mechanisms, vol. 1, pp. 213-220, Nov-Dec 2008.
[23] M. Liu, S. J. M. Skinner, J. Xu, R. N. N. Han, A. K. Tanswell, and M. Post, "Stimulation of fetal-rat lung-cell proliferation invitro by mechanical stretch," American Journal of Physiology, vol. 263, pp. L376-L383, Sep 1992.
[24] W. Cheng, B. S. Li, J. Kajstura, P. Li, M. S. Wolin, E. H. Sonnenblick, et al., "Stretch-induced programmed myocyte cell-death," Journal of Clinical Investigation, vol. 96, pp. 2247-2259, Nov 1995.
[25] X. Trepat, L. H. Deng, S. S. An, D. Navajas, D. J. Tschumperlin, W. T. Gerthoffer, et al., "Universal physical responses to stretch in the living cell," Nature, vol. 447, pp. 592-+, May 2007.
[26] V. Vogel and M. Sheetz, "Local force and geometry sensing regulate cell functions," Nature Reviews Molecular Cell Biology, vol. 7, pp. 265-275, Apr 2006.
[27] D. E. Ingber, "Tensegrity II. How structural networks influence cellular information processing networks," Journal of Cell Science, vol. 116, pp. 1397-1408, Apr 2003.
[28] S. Boyden, "Chemotactic effect of mixtures of antibody and antigen on polymorphonuclear leucocytes," Journal of Experimental Medicine, vol. 115, pp. 453-&, 1962.
[29] D. Zicha, G. A. Dunn, and A. F. Brown, "A new direct-viewing chemotaxis chamber," Journal of Cell Science, vol. 99, pp. 769-775, Aug 1991.
[30] S. H. Zigmond and J. G. Hirsch, "Leukocyte locomotion and chemotaxis - new methods for evaluation and demonstration of a cell-derived chemotactic factor," Journal of Experimental Medicine, vol. 137, pp. 387-410, 1973.
[31] H. Kamble, M. J. Barton, M. Jun, S. Park, and N. T. Nguyen, "Cell stretching devices as research tools: engineering and biological considerations," Lab on a Chip, vol. 16, pp. 3193-3203, 2016.
[32] Y. L. Huang and N. T. Nguyen, "A polymeric cell stretching device for real-time imaging with optical microscopy," Biomedical Microdevices, vol. 15, pp. 1043-1054, Dec 2013.
[33] D. Tremblay, S. Chagnon-Lessard, M. Mirzaei, A. E. Pelling, and M. Godin, "A microscale anisotropic biaxial cell stretching device for applications in mechanobiology," Biotechnology Letters, vol. 36, pp. 657-665, Mar 2014.
[34] A. O. Stucki, J. D. Stucki, S. R. R. Hall, M. Felder, Y. Mermoud, R. A. Schmid, et al., "A lung-on-a-chip array with an integrated bio-inspired respiration mechanism," Lab on a Chip, vol. 15, pp. 1302-1310, 2015.
[35] C. Moraes, M. Likhitpanichkul, C. J. Lam, B. M. Beca, Y. Sun, and C. A. Simmons, "Microdevice array-based identification of distinct mechanobiological response profiles in layer-specific valve interstitial cells," Integrative Biology, vol. 5, pp. 673-680, 2013.
[36] K. Naruse, T. Yamada, and M. Sokabe, "Involvement of SA channels in orienting response of cultured endothelial cells to cyclic stretch," American Journal of Physiology-Heart and Circulatory Physiology, vol. 274, pp. H1532-H1538, May 1998.
[37] K. Naruse, T. Yamada, X. R. Sai, M. Hamaguchi, and M. Sokabe, "Pp125(FAK) is required for stretch dependent morphological response of endothelial cells," Oncogene, vol. 17, pp. 455-463, Jul 1998.
[38] Y. Shao, X. Y. Tan, R. Novitski, M. Muqaddam, P. List, L. Williamson, et al., "Uniaxial cell stretching device for live-cell imaging of mechanosensitive cellular functions," Review of Scientific Instruments, vol. 84, Nov 2013.
[39] S. Akbari and H. R. Shea, "Microfabrication and characterization of an array of dielectric elastomer actuators generating uniaxial strain to stretch individual cells," Journal of Micromechanics and Microengineering, vol. 22, Apr 2012.
[40] A. Poulin, C. S. Demir, S. Rosset, T. V. Petrova, and H. Shea, "Dielectric elastomer actuator for mechanical loading of 2D cell cultures," Lab on a Chip, vol. 16, pp. 3788-3794, 2016.
[41] D. Cei, J. Costa, G. Gori, G. Frediani, C. Domenici, F. Carpi, et al., "A bioreactor with an electro-responsive elastomeric membrane for mimicking intestinal peristalsis," Bioinspiration & Biomimetics, vol. 12, Feb 2017.
[42] Y. Kamotani, T. Bersano-Begey, N. Kato, Y. C. Tung, D. Huh, J. W. Song, et al., "Individually programmable cell stretching microwell arrays actuated by a Braille display," Biomaterials, vol. 29, pp. 2646-2655, Jun 2008.
[43] D. Huh, H. J. Kim, J. P. Fraser, D. E. Shea, M. Khan, A. Bahinski, et al., "Microfabrication of human organs-on-chips," Nature Protocols, vol. 8, pp. 2135-2157, Nov 2013.
[44] O. Kilic, D. Pamies, E. Lavell, P. Schiapparelli, Y. Feng, T. Hartung, et al., "Brain-on-a-chip model enables analysis of human neuronal differentiation and chemotaxis," Lab on a Chip, vol. 16, pp. 4152-4162, 2016.
[45] "National Heart, Lung and Blood Institute Guidelines for the Diagnosis and Management of Asthma, Expert Panel Report 2 Bethesda: National Institutes of Health publication number 97–4051," 1997.