Role of Substrate Stiffness on Migratory Properties and Epithelial to Mesenchymal Transition in Human Lung Cancer Cells
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| Language: | English |
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The Ohio State University / OhioLINK
2012
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| Online Access: | http://rave.ohiolink.edu/etdc/view?acc_num=osu1356943256 |
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ndltd-OhioLink-oai-etd.ohiolink.edu-osu1356943256 |
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oai_dc |
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NDLTD |
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English |
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NDLTD |
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Biomedical Engineering Cell migration cell motility substrate stiffness PDMS Epithelial to Mesenchymal Transition metastasis 2D migration assay TGF-¿¿ A549 |
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Biomedical Engineering Cell migration cell motility substrate stiffness PDMS Epithelial to Mesenchymal Transition metastasis 2D migration assay TGF-¿¿ A549 Subisak, Angel Dharshini Role of Substrate Stiffness on Migratory Properties and Epithelial to Mesenchymal Transition in Human Lung Cancer Cells |
| author |
Subisak, Angel Dharshini |
| author_facet |
Subisak, Angel Dharshini |
| author_sort |
Subisak, Angel Dharshini |
| title |
Role of Substrate Stiffness on Migratory Properties and Epithelial to Mesenchymal Transition in Human Lung Cancer Cells |
| title_short |
Role of Substrate Stiffness on Migratory Properties and Epithelial to Mesenchymal Transition in Human Lung Cancer Cells |
| title_full |
Role of Substrate Stiffness on Migratory Properties and Epithelial to Mesenchymal Transition in Human Lung Cancer Cells |
| title_fullStr |
Role of Substrate Stiffness on Migratory Properties and Epithelial to Mesenchymal Transition in Human Lung Cancer Cells |
| title_full_unstemmed |
Role of Substrate Stiffness on Migratory Properties and Epithelial to Mesenchymal Transition in Human Lung Cancer Cells |
| title_sort |
role of substrate stiffness on migratory properties and epithelial to mesenchymal transition in human lung cancer cells |
| publisher |
The Ohio State University / OhioLINK |
| publishDate |
2012 |
| url |
http://rave.ohiolink.edu/etdc/view?acc_num=osu1356943256 |
| work_keys_str_mv |
AT subisakangeldharshini roleofsubstratestiffnessonmigratorypropertiesandepithelialtomesenchymaltransitioninhumanlungcancercells |
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1719418722836283392 |
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ndltd-OhioLink-oai-etd.ohiolink.edu-osu13569432562021-08-03T05:20:09Z Role of Substrate Stiffness on Migratory Properties and Epithelial to Mesenchymal Transition in Human Lung Cancer Cells Subisak, Angel Dharshini Biomedical Engineering Cell migration cell motility substrate stiffness PDMS Epithelial to Mesenchymal Transition metastasis 2D migration assay TGF-¿¿ A549 <p>Cancer affects a large percentage of the population in the United States, and innumerable research efforts are dedicated to studying the attributes of cancer to develop effective therapies. The mortality rate increases significant when the primary tumor begins to metastasize to different parts of the body. For example, women with localized breast cancer have a 93% survival rate while women with metastatic disease have a 15% survival rate. During metastasis, cancer cells acquire a highly mobile phenotype allowing them to invade into the surrounding tissues. One mechanism by which cancer cells acquire this highly invasive and mobile phenotype is known as Epithelial to Mesenchymal Transition (EMT). EMT is a fundamental biological process that plays an important role in organogenesis and wound healing but is also thought to play an important role in cancer progression where adherent epithelial cells acquire a highly invasive mesenchymal phenotype. Although EMT is believed to be a key process in cancer cell metastasis, the role of substrate stiffness on EMT and motility patterns is not completely understood. In addition, EMT can be induced in some cell types transforming growth factor-¿¿(TGF-¿¿)but it is not known how changes in substrate stiffness influence TGF-¿¿ induced EMT and the associated changes in cancer cell migration.</p><p>The goal of this thesis was to examine whether substrate stiffness affects migration pattern and behavior of cancer cells in vitro. We used a two dimensional invasion assay on substrates with varying stiffness in the physiologically relevant range, and assessed migration parameters including migration velocity, directionality and distance. We also examined how substrate stiffness alters cell migration patterns in the presence of TGF-¿¿. Finally, the migration results were compared with changes in cell morphology and EMT protein marker expression to investigate how substrate stiffness influences correlations between morphology/protein markers and migration patterns.</p><p>We used Polydimethylsiloxane (PDMS) coated with collagen type 1 which had a stiffness that varied from 6 to 4613 kPa and seeded A549 cells on these gels. Cells were treated with control medium (DMEM with 10% FBS) or 5ng of TGF-¿¿/ml of DMEM. Monolayers with a well-defined “edge” were created using ibidi¿¿ silicon inserts. After formation of a monolayer, the insert was removed and placed in live-cell imaging system to obtain migration data for 24 hours. 50 cells from 3 experiments were manually tracked and velocity (V), accumulated distance (daccum), Euclidean distance (deuclid) and directionality (deuclid / daccum) were measured. Western blotting was done to evaluate expression of E-cadherin, N-cadherin and vimentin. Immunofluorescence imaging was done for cytoskeletal actin.</p><p>Our study shows that cells migrating on stiff PDMS substrates (120-430 kPa) are more aggressive than cells on soft PDMS substrates (20-40 kPa) with increased velocity, directionality, and distance. However, cells migrating on very stiff substrates (> 4600 kPa) exhibit slower migration. This consistent change in migration due to matrix rigidity was not correlated with changes in cell morphology and biochemical marker expression. We also studied migration patterns after inducing classical EMT in the cells by treating them with TGF-¿¿. Upon treatment with TGF-¿¿, there is a pronounced change in migration, cell morphology, and protein markers. There is an increase in velocity and accumulated distance accompanied with a decrease in directionality and Euclidean distance which indicates that substrate stiffness can alter TGF-¿¿ induced changes in migration patterns. Although there is a decrease in E-cadherin and increase in N-cadherin expression, the changes in biochemical markers due to TGF-¿¿ do not correlated with the observed changes in migration as a function of substrate stiffness.</p><p>We conclude that changes in substrate stiffness alone have a significant impact on cell migration patterns and motility phenotype before and after inducing EMT by TGF-¿¿ treatment. However, changes in EMT biochemical markers do not correlate with changes in migration behaviors as a function of substrate stiffness. This novel finding suggests mechanical properties in the tumor microenvironment can alter cellular processes like migration independent of biochemical changes. Continued research to elucidate the relationship between substrate stiffness, EMT, metastasis, and migration in vitro may provide methods to control rate of pathological cell migration, thereby limiting formation of metastases.</p> 2012 English text The Ohio State University / OhioLINK http://rave.ohiolink.edu/etdc/view?acc_num=osu1356943256 http://rave.ohiolink.edu/etdc/view?acc_num=osu1356943256 unrestricted This thesis or dissertation is protected by copyright: all rights reserved. It may not be copied or redistributed beyond the terms of applicable copyright laws. |