機械循環拉伸力對3D培養肺癌細胞之影響

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賴聖皓(Sheng-Hao Lai) 查詢紙本館藏

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系統生物與生物資訊研究所

論文名稱

機械循環拉伸力對3D培養肺癌細胞之影響
(The effect of cyclic mechanical stretch on the 3D culture model of lung cancer cells)

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摘要(中)

在人體中存在著許多種機械力而拉伸力即屬於其中一種，循環拉伸存在於如:心臟、肺臟等等…在過去已知循環拉伸會對細胞的增殖、分化及基因調控造成影響且在維持組織恆定上扮演重要的角色。本實驗分別對人類肺部纖維母細胞NHLF和肺腺癌細胞A549給予機械循環拉伸處理，並探討纖維化程度、癌細胞於三維環境中的侵襲以及和周遭免疫細胞的互動。結果顯示給予一定程度的循環拉伸會使肺部纖維母細胞表現較多的成肌纖維母細胞標誌物α-SMA。在癌細胞部份，循環拉伸對於三維肺腺癌細胞球在膠原蛋白水凝膠中的侵襲有抑制的現象，而使用經由形變量10%、頻率0.21 hz拉伸的A549條件培養基處理的M0未成熟巨噬細胞在24、48小時有較明顯的M1標誌物TLR2、IL-1β、TNF-α上升及M2標誌物CD206下降，這說明經拉伸的A549可能具有促使巨噬細胞朝M1表型發展之能力。

摘要(英)

There are many mechanical forces in the human body, and stretching is one of them. Cyclic stretching exists in the heart, lungs, etc. In the past, it was known that cyclic stretching would affect cell proliferation, differentiation, and gene regulation. And it plays an important role in tissue maintenance. In this study, human lung fibroblasts NHLF and lung adenocarcinoma cells A549 were treated with mechanical circulation stretching, and the degree of fibrosis, the invasion of cancer cells in a three-dimensional environment and the interaction with surrounding immune cells were explored. The results show that given a certain degree of cyclic stretching can make lung fibroblasts express more myofibroblast marker α-SMA. In the cancer cell part, cyclic stretching can inhibit the invasion of lung adenocarcinoma spheroid in the collagen hydrogel. After stretched A549 conditioned medium with a magnitude of 10% and a frequency of 0.21 hz treating, Naïve macrophages have obvious M1 markers TLR2, IL-1β, TNF-α increase and M2 marker CD206 decrease at 24 and 48 hours, which indicates that the stretched A549 may promote the macrophages to move toward M1 phenotype.

關鍵字(中)

★ 機械循環拉伸
★ 肺癌
★ 3D培養

關鍵字(英)

論文目次

目錄
中文摘要 i
Abstract ii
目錄 iii
圖目錄 v
表目錄 vi
一、緒論 1
1-1 肺癌 1
1-2 機械循環拉伸 2
1-3 腫瘤微環境 2
1-4 三維細胞培養 3
1-5 肺部纖維化 4
二、實驗材料與方法 5
2-1實驗方法 5
2-1-1 肺腺癌三維細胞球 5
2-1-2 水凝膠配製 6
2-1-3 裝置製作 7
2-1-4 三維細胞循環拉伸實驗 8
2-1-5 二維細胞循環拉伸實驗 8
2-1-6 巨噬細胞、單核球極化實驗 9
2-1-7 細胞固定及染色 9
2-1-8 螢光數據計算方法 10
2-1-9 RNA萃取 13
2-1-10 qPCR(即時定量聚合酶連鎖反應) 14
2-2實驗材料 18
2-2-1 細胞培養 18
2-2-2 循環拉伸系統 19
2-2-3 細胞固定及染色 20
2-2-4 qPCR、RNA萃取及反轉錄套件 21
2-2-5 DNA膠體電泳 21
三、實驗流程 22
四、實驗結果 23
4-1以機械循環拉伸刺激人類肺纖維母細胞(NHLF) 23
4-1-1 以形變量15%對人類肺纖維母細胞進行機械循環拉伸 23
4-1-2 以形變量20%對人類肺纖維母細胞進行機械循環拉伸 25
4-2以人類肺腺癌A549三維細胞球於水凝膠中給予機械循環拉伸刺激 27
4-2-1 膠原蛋白水凝膠濃度對肺腺癌細胞球之影響 27
4-2-2 肺腺癌細胞球於裝置不同位置與形變量之關係 28
4-2-3 肺腺癌細胞球於水凝膠中位置與侵襲面積之關係 29
4-2-4 肺腺癌細胞球經機械循環拉伸刺激之結果 30
4-2-4-1 肺腺癌細胞球於濃度1 mg/ml膠原蛋白水凝膠中循環拉伸結果 30
4-2-4-2 於濃度1mg/ml膠原蛋白水凝膠加入10 μg/ml纖連蛋白(Fibronectin)之循環拉伸結果 33
4-3以機械循環拉伸後之肺腺癌細胞條件培養基處理未成熟巨噬細胞
(Naïve macrophage, M0)及人類單核球細胞株(THP-1) 35
4-3-1 經由條件培養基處理之未成熟巨噬細胞及人類單核球RT-qPCR結果 37
4-3-2 以DNA膠體電泳確認RT-qPCR產物 40
五、討論與結論 41
5-1一定程度的機械循環拉伸能使人類纖維母細胞表達較多α-SMA 41
5-2機械循環拉伸能減少肺腺癌細胞球於水凝膠中的侵襲面積 41
5-3經機械循環拉伸之A549肺腺癌細胞可能使巨噬細胞朝M1方向發展 43
參考文獻 44

參考文獻

參考文獻
1. Ferlay, J., et al., Estimates of worldwide burden of cancer in 2008: GLOBOCAN 2008. International journal of cancer, 2010. 127(12): p. 2893-2917.
2. Taiwan’s Cancer Registry Annual Report (2018) Bureau of Health, E. Promotion Department of Health, Taiwan 2020, Editor.
3. Reade, C.A., A.K. Ganti, and therapy, EGFR targeted therapy in non-small cell lung cancer: potential role of cetuximab. Biologics: targets, 2009. 3: p. 215.
4. Musani, A.I., Pulmonary Disease, An Issue of Medical Clinics of North America, E-Book. Vol. 103. 2019: Elsevier Health Sciences.
5. de Groot, P. and R.F. Munden, Lung cancer epidemiology, risk factors, and prevention. Radiologic Clinics, 2012. 50(5): p. 863-876.
6. Pope Iii, C.A., et al., Lung cancer, cardiopulmonary mortality, and long-term exposure to fine particulate air pollution. Jama, 2002. 287(9): p. 1132-1141.
7. Herbst, R.S., J.V. Heymach, and S.M. Lippman, Molecular origins of cancer. N Engl J Med, 2008. 359(13): p. 1367-80.
8. Matakidou, A., T. Eisen, and R. Houlston, Systematic review of the relationship between family history and lung cancer risk. British journal of cancer, 2005. 93(7): p. 825-833.
9. Wang, J.H.-C. and B.P. Thampatty, An introductory review of cell mechanobiology. Biomechanics modeling in mechanobiology, 2006. 5(1): p. 1-16.
10. Yang, G., R.C. Crawford, and J.H. Wang, Proliferation and collagen production of human patellar tendon fibroblasts in response to cyclic uniaxial stretching in serum-free conditions. Journal of biomechanics, 2004. 37(10): p. 1543-1550.
11. Li, N., et al., MicroRNA‐129‐1‐3p regulates cyclic stretch–induced endothelial progenitor cell differentiation by targeting Runx2. Journal of cellular biochemistry, 2019. 120(4): p. 5256-5267.
12. Inoh, H., et al., Uni‐axial cyclic stretch induces the activation of transcription factor nuclear factor κB in human fibroblast cells. The FASEB Journal, 2002. 16(3): p. 405-407.
13. Spill, F., et al., Impact of the physical microenvironment on tumor progression and metastasis. Current opinion in biotechnology, 2016. 40: p. 41-48.
14. Del Prete, A., et al., Leukocyte trafficking in tumor microenvironment. Current opinion in pharmacology, 2017. 35: p. 40-47.
15. Hanahan, D. and R.A. Weinberg, Hallmarks of cancer: the next generation. cell, 2011. 144(5): p. 646-674.
16. Mantovani, A., et al., Cancer-related inflammation. nature, 2008. 454(7203): p. 436-444.
17. Jarosz-Biej, M., et al., Tumor microenvironment as a “game changer” in cancer radiotherapy. International journal of molecular sciences, 2019. 20(13): p. 3212.
18. Mantovani, A., et al., Tumour-associated macrophages as treatment targets in oncology. Nature reviews Clinical oncology, 2017. 14(7): p. 399.
19. Lochter, A., et al., Matrix metalloproteinase stromelysin-1 triggers a cascade of molecular alterations that leads to stable epithelial-to-mesenchymal conversion and a premalignant phenotype in mammary epithelial cells. The Journal of cell biology, 1997. 139(7): p. 1861-1872.
20. Levental, K.R., et al., Matrix crosslinking forces tumor progression by enhancing integrin signaling. Cell, 2009. 139(5): p. 891-906.
21. Bruzzese, F., et al., Local and systemic protumorigenic effects of cancer-associated fibroblast-derived GDF15. Cancer research, 2014. 74(13): p. 3408-3417.
22. Walker, C., E. Mojares, and A. del Río Hernández, Role of extracellular matrix in development and cancer progression. International journal of molecular sciences, 2018. 19(10): p. 3028.
23. Baal, N., et al., In vitro spheroid model of placental vasculogenesis: does it work? Laboratory investigation, 2009. 89(2): p. 152-163.
24. Ma, H.-l., et al., Multicellular tumor spheroids as an in vivo–like tumor model for three-dimensional imaging of chemotherapeutic and nano material cellular penetration. Molecular imaging, 2012. 11(6): p. 7290.2012. 00012.
25. Kimlin, L.C., G. Casagrande, and V.M. Virador, In vitro three‐dimensional (3D) models in cancer research: an update. Molecular carcinogenesis, 2013. 52(3): p. 167-182.
26. Collins, A., et al., Patient-derived explants, xenografts and organoids: 3-dimensional patient-relevant pre-clinical models in endometrial cancer. Gynecologic oncology, 2020. 156(1): p. 251-259.
27. Wan, L., C. Neumann, and P. LeDuc, Tumor-on-a-chip for integrating a 3D tumor microenvironment: chemical and mechanical factors. Lab on a Chip, 2020. 20(5): p. 873-888.
28. Nath, S. and G.R. Devi, Three-dimensional culture systems in cancer research: Focus on tumor spheroid model. Pharmacology therapeutics
2016. 163: p. 94-108.
29. Henderson, N.C., F. Rieder, and T.A. Wynn, Fibrosis: from mechanisms to medicines. Nature, 2020. 587(7835): p. 555-566.
30. Sundarakrishnan, A., et al., Engineered cell and tissue models of pulmonary fibrosis. Advanced drug delivery reviews, 2018. 129: p. 78-94.
31. Rittié, L., Type I collagen purification from rat tail tendons, in Fibrosis. 2017, Springer. p. 287-308.
32. Kisling, A., R.M. Lust, and L.C. Katwa, What is the role of peptide fragments of collagen I and IV in health and disease? Life sciences, 2019. 228: p. 30-34.
33. Doyle, A.D., et al., Local 3D matrix microenvironment regulates cell migration through spatiotemporal dynamics of contractility-dependent adhesions. Nature communications, 2015. 6(1): p. 1-15.
34. Sia, S.K. and G.M. Whitesides, Microfluidic devices fabricated in poly (dimethylsiloxane) for biological studies. Electrophoresis, 2003. 24(21): p. 3563-3576.
35. Luo, Z., et al., Effect of NR5A2 inhibition on pancreatic cancer stem cell (CSC) properties and epithelial‐mesenchymal transition (EMT) markers. Molecular carcinogenesis, 2017. 56(5): p. 1438-1448.
36. Tedesco, S., et al., Convenience versus biological significance: are PMA-differentiated THP-1 cells a reliable substitute for blood-derived macrophages when studying in vitro polarization? Frontiers in pharmacology, 2018. 9: p. 71.
37. Morón-Calvente, V., et al., Inhibitor of apoptosis proteins, NAIP, cIAP1 and cIAP2 expression during macrophage differentiation and M1/M2 polarization. PloS one, 2018. 13(3): p. e0193643.
38. Roan, E. and C.M. Waters, What do we know about mechanical strain in lung alveoli? American Journal of Physiology-Lung Cellular
Molecular Physiology, 2011. 301(5): p. L625-L635.
39. Chang, C.-H., H.-H. Lee, and C.-H. Lee, Substrate properties modulate cell membrane roughness by way of actin filaments. Scientific reports, 2017. 7(1): p. 1-11.
40. Gordon, S., The macrophage: past, present and future. European journal of immunology, 2007. 37(S1): p. S9-S17.
41. Gordon, S., Alternative activation of macrophages. Nature reviews immunology, 2003. 3(1): p. 23-35.
42. Mosser, D.M. and J.P. Edwards, Exploring the full spectrum of macrophage activation. Nature reviews immunology, 2008. 8(12): p. 958-969.
43. Xu, F., et al., Astragaloside IV inhibits lung cancer progression and metastasis by modulating macrophage polarization through AMPK signaling. Journal of Experimental Clinical Cancer Research, 2018. 37(1): p. 1-16.
44. Peyser, R., et al., Defining the activated fibroblast population in lung fibrosis using single-cell sequencing. American journal of respiratory cell molecular biology, 2019. 61(1): p. 74-85.
45. Friedl, P., et al., Classifying collective cancer cell invasion. Nature cell biology, 2012. 14(8): p. 777-783.
46. Zhang, B., et al., Cyclic mechanical stretching promotes migration but inhibits invasion of rat bone marrow stromal cells. Stem cell research, 2015. 14(2): p. 155-164.
47. Solinas, G., et al., Tumor-conditioned macrophages secrete migration-stimulating factor: a new marker for M2-polarization, influencing tumor cell motility. The Journal of Immunology, 2010. 185(1): p. 642-652.

指導教授

許藝瓊

審核日期

2021-10-4

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