PDMS is not a hospitable surface for cell adhesion
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•PDMS is not a hospitable surface for cell adhesion •Lack of adhesion shows MCF7 cells retain characteristics of noncancerous cells •PDMS is not a hospitable surface for cell adhesion •Adhesion of cells shows that MB231 cells have unique adaptive abilities not found in noncancerous cells Actin Fibers Focal Adhesions * * Actin Fibers Focal Adhesions 0 2 4 6 8 10 12 14 16 Fluorescence (average intensity/cell) Day 1 Day 7 Cells Structure Analysis MB231 Cell Line Hypothesis Effect of Substrate Rigidity On Breast Cancer Cells Elizabeth L. Smith, RET Fellow 2011 West Aurora High School RET Mentor: Dr. Michael Cho, PhD NSF- RET Program Motivation Abstract Conclusion Material and Methods Results NSF Grant CBET-EEC-0743068 Prof. A. Linninger, RET Program Director Dr. Michael Cho, Research Advisor Brandon Lutz, Research Mentor University of Illinois- Chicago Acknowledgements References 1. Breastcancer.org (April 19 th , 2011). U.S. Breast CancerStatistics. Breastcancer.org. Retrieved July 26, 2011, from http://www.breastcancer.org/symptoms/understand_bc/statistics.jsp 2. Curtis, A., & Wilkinson, C. (January 01, 1997). Topographical control of cells. Biomaterials, 18, 24, 1573. 3. Guo, W. H., Frey, M. T., Burnham, N. A., & Wang, Y. L. (January 01, 2006). Substrate rigidity regulates the formation and maintenance of tissues. Biophysical Journal,90, 6, 2213-20. 4. Rapier, R., Huq, J., Vishnubhotla, R., Bulic, M., Perrault, C. M., Metlushko, V., Cho, M., ... Glover, S. C. (January 01, 2010). The extracellular matrix microtopography drives critical changes in cellular motility and Rho A activity in colon cancer cells. Cancer Cell International, 10. 5. Tzvetkova-Chevolleau, T., Stphanou, A., Fuard, D., Ohayon, J., Breast cancer cells will change structure in response to varying substrate stiffness Average Number of Cells per View MB231 Cell Line Rigid Substrate 10:1 PDMS Soft Substrate 30:1 PDMS Statistical Significance * Two Substrate s Polymer : Cross- linking Ratio Rigidity Rigid PDMS 10:1 2.2 MPa Soft PDMS 30:1 0.3 MPa Two Cell Lines Tissue Mobility MB231 Breast Invasive MCF7 Breast Less invasive Analysi s 2 samples - Each Cell line on Each Substrate Collected - Day 1, Day 3, Day 7 Florescent Microscopy Dye Target Indicates Color Actin Cytoskelton Red Vinculin Focal Adhesions Green DNA Nucleus Blue Metamorph Software -Images psuedocolored and overlaid ImageJ Software - Cell count determined using nuclei - RGB intensities quantified for each image then normalized by cell count Microsoft Excel - Statistical Analysis & Graph Making Hypothesis 1 in 8 women in the U.S. will be diagnosed with breast cancer in their lifetime Breast cancer is the second deadliest cancer in women 1 Cancer cells interact with the environment around them in a bidirectional manner. While metastatic cells modify the environment and navigate through it, the environment exerts significant influence over the cell’s shape, structure, and behavior. 2,3,4,5 This study was designed to examine how two breast cancer lines, MB231 and MCF7, respond to PDMS (polydimethylsiloxane) substrates that differ in the rigidity by an order of magnitude. Less invasive MCF7 cells adhere poorly to PDMS independent of the rigidity, suggesting lack of focal adhesion. Highly invasive MB231 cells showed greater proliferation on the soft substrate, as well as significantly greater actin fibers and focal adhesions. Invasive MB231 cells alter their structure based on substrate rigidity. Understanding the rigidity-dependent cancer cell behavior may lead to development of better cancer diagnoses, therapies, and potentially cures. MB231 cell growing on 10:1 PDMS at 40x magnification 100 um Invasive MB231 Cells on Rigid Substrate - Lower cell density - Greater actin fibers - Greater focal adhesions Invasive MB231 Cells on Soft Substrate - Higher cell density - Lesser actin fibers - Lesser focal adhesions Day 1 Day 7 0 2 4 6 8 10 12 14 16 Average Number of Cells per View (40x Magnification) * Future studies may include further analysis of: - Cell mobility - Surface proteins - Size & shape of focal adhesions - Cell rigidity - Intercellular signaling and actin MB231 Invasive Breast Cancer MCF7 Less invasive Breast Cancer
description
Effect of Substrate Rigidity On Breast Cancer Cells Elizabeth L. Smith, RET Fellow 2011 West Aurora High School RET Mentor: Dr. Michael Cho, PhD NSF- RET Program. Abstract. Motivation. - PowerPoint PPT Presentation
Transcript of PDMS is not a hospitable surface for cell adhesion
•PDMS is not a hospitable surface for cell adhesion
•Lack of adhesion shows MCF7 cells retain characteristics of noncancerous cells
• PDMS is not a hospitable surface for cell adhesion
• Adhesion of cells shows that MB231 cells have unique adaptive abilities not found in noncancerous cells
Actin Fibers Focal Adhesions
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Actin Fibers Focal Adhesions0
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Cells Structure AnalysisMB231 Cell Line
Hypothesis
Effect of Substrate RigidityOn Breast Cancer Cells
Elizabeth L. Smith, RET Fellow 2011West Aurora High School
RET Mentor: Dr. Michael Cho, PhD NSF- RET Program
MotivationAbstract
Conclusion
Material and Methods
Results
NSF Grant CBET-EEC-0743068 Prof. A. Linninger, RET Program Director Dr. Michael Cho, Research Advisor Brandon Lutz, Research Mentor University of Illinois- Chicago
Acknowledgements
References1. Breastcancer.org (April 19th, 2011). U.S. Breast CancerStatistics. Breastcancer.org. Retrieved
July 26, 2011, from http://www.breastcancer.org/symptoms/understand_bc/statistics.jsp2. Curtis, A., & Wilkinson, C. (January 01, 1997). Topographical control of
cells. Biomaterials, 18, 24, 1573.3. Guo, W. H., Frey, M. T., Burnham, N. A., & Wang, Y. L. (January 01, 2006). Substrate rigidity
regulates the formation and maintenance of tissues. Biophysical Journal,90, 6, 2213-20.4. Rapier, R., Huq, J., Vishnubhotla, R., Bulic, M., Perrault, C. M., Metlushko, V., Cho, M., ...
Glover, S. C. (January 01, 2010). The extracellular matrix microtopography drives critical changes in cellular motility and Rho A activity in colon cancer cells. Cancer Cell International, 10.
5. Tzvetkova-Chevolleau, T., Stephanou, A., Fuard, D., Ohayon, J., Schiavone, P., & Tracqui, P. (January 01, 2008). The motility of normal and cancer cells in response to the combined influence of the substrate rigidity and anisotropic microstructure. Biomaterials,29, 10, 1541-51.
Breast cancer cells will change structure in response to varying substrate stiffness
Average Number of Cells per ViewMB231 Cell Line
Rigid Substrate10:1 PDMS
Soft Substrate30:1 PDMS
StatisticalSignificance*
Two Substrates
Polymer : Cross-linkingRatio Rigidity
Rigid PDMS 10:1 2.2 MPa
Soft PDMS 30:1 0.3 MPa
Two Cell Lines
Tissue Mobility
MB231 Breast Invasive
MCF7 Breast Less invasive
Analysis 2 samples - Each Cell line on Each SubstrateCollected - Day 1, Day 3, Day 7
Florescent Microscopy
Dye Target Indicates Color Actin Cytoskelton Red
Vinculin Focal Adhesions Green
DNA Nucleus Blue
MetamorphSoftware -Images psuedocolored and overlaid
ImageJSoftware
-Cell count determined using nuclei-RGB intensities quantified for each image then normalized by cell count
Microsoft Excel
-Statistical Analysis & Graph Making
Hypothesis
1 in 8 women in the U.S. will be diagnosed with breast cancer in their lifetime
Breast cancer is the second deadliest cancer in women1
Cancer cells interact with the environment around them in a bidirectional manner. While metastatic cells modify the environment and navigate through it, the environment exerts significant influence over the cell’s shape, structure, and behavior.2,3,4,5
This study was designed to examine how two breast cancer lines, MB231 and MCF7, respond to PDMS (polydimethylsiloxane) substrates that differ in the rigidity by an order of magnitude.
Less invasive MCF7 cells adhere poorly to PDMS independent of the rigidity, suggesting lack of focal adhesion. Highly invasive MB231 cells showed greater proliferation on the soft substrate, as well as significantly greater actin fibers and focal adhesions. Invasive MB231 cells alter their structure based on substrate rigidity.
Understanding the rigidity-dependent cancer cell behavior may lead to development of better cancer diagnoses, therapies, and potentially cures.
MB231 cell growing on 10:1 PDMS at 40x magnification
100 um
Invasive MB231 Cells on Rigid Substrate- Lower cell density- Greater actin fibers- Greater focal adhesions
Invasive MB231 Cells on Soft Substrate- Higher cell density- Lesser actin fibers- Lesser focal adhesions
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Future studies may include further analysis of: - Cell mobility- Surface proteins - Size & shape of focal adhesions- Cell rigidity - Intercellular signaling and actin fibers
MB231Invasive Breast Cancer
MCF7 Less invasive Breast Cancer
