Cardiovascular diseases, in which EC dysfunction is known to be a major cause, are the number one cause of death in the world. It is known that blood flow (hemodynamics) is one of the key factors which governs the EC dysfunction. Although much work has investigated the effects of flow on ECs in vitro, accurate quantification of the flow over the ECs and quantitative analysis of cell response in some key aspects remains elusive. The current research aims to apply quantitative methods to examine the cell response under various flow conditions through determining the morphometric parameters as well as cytoskeletal remodeling to link the descriptive and quantitative cell response due to flow with EC dysfunction. The following chapter will discuss in detail the in vitro
hemodynamic facility, the associated experimental set-up and protocols that were developed as the initial phase of the research project.
1.9 References
[1] World Health Organization (WHO), " Cardiovascular Disease," retrieved from: http://www.who.int/mediacentre/factsheets/fs317/en/. [Accessed 3 February 2015]. [2] G. Hansson and A. Hermansonn, "The Immune System in Atherosclerosis," Nat.
Immunol., vol. 13, no. 2, pp. 204-212, 2011.
[3] B. Alberts, A. Johnson, J. Lewis, M. Raff, K. Roberts and P. Walter, Molecular Biology of the Cell, New York: Garland Sience, 2008.
[4] G. A. Truskey, F. Yuan and D. F. Katz, Transport Phenomena in Biological Systems, New York: Pearson, 2009.
[5] S. Tojkander, G. Gateva and P. Lappalainen, "Actin Stress Fibers: Assembly, Dynamics and Biological Roles," J. Cell Sci., vol. 125, pp. 1855-1864, 2012. [6] B. M. Gumbiner, "Cell Adhesion: The Molecular Basis of Tissue Architecture and
Morphogenesis," Cell, vol. 84, pp. 345-357, 1996.
[7] D. E. Jackson, "The Unfolding Tale of PECAM-1," FEBS Lett., vol. 540, pp. 7-14, 2003.
[8] S. M. Albelda, W. A. Muller, C. A. Buck and P. J. Newman, "Molecular and Cellular Properties of PECAM-1(endoCAM/CD31): A Novel Vascular Cell-Cell Adhesion Molecule," J. Cell Biol., vol. 114, no. 5, pp. 1059-1068, 1991.
[9] S. Chien, "Mechanotransduction and Endothelial Cell Homeostasis; the Wisdom of the Cell," Am J Physiol Heart Circ Physiol, vol. 292, pp. 1209-1224, 2007.
[10] J. A. Berliner, M. Navab, A. M. Fogelman, J. S. Frank, L. L. Demer, P. Edwards, A. D. Watson and A. J. Lusis, "Atherosclerosis: Basic Mechanisms, Oxidation,
[11] F. M. White, Fluid mechanics, New York: McGraw Hill, 2009. [12] M. A. Gimbrone, "Vascular Endothelium, Hemodynamic Forces and
Atherogenesis," Am. J. Pathol., vol. 155, no. 1, pp. 537-539, 1999.
[13] D. E. Ingber, "Mechanobiology and Diseases of Mechanotransduction," Ann. Med., vol. 35, pp. 564-577, 2003.
[14] R. M. Nerem, "Hemodynamics and Vascular Endothelium," J. Biomech. Eng. , vol. 115, pp. 510-514, 1993.
[15] H. S. Baldwin, H. M. Shen, H. C. Yan, H. M. DeLiss, A. Chunh, C. Mickanin, T. Trask, N. E. Kirschbuaum, P. J. Newman and S. M. Albelda, "Platelet Endothelial Cell Adhesion Molecule-1 (PECAM-1/CD31): Alternatively Spliced, Functionally distinct Isoforms Expressed During Mammalian Cardiovascular Development," Development, vol. 120, pp. 2539-2553, 1994.
[16] L. Cao, A. Wu and G. A. Truskey, "Biomechanical Effects of Flow and Conculture on Human Aortic and Cord Blood Derived Endothelial Cells," J Biomech., vol. 44, pp. 2150-2157, 2011.
[17] S. Chien, "Effects of Disturbed Flow on Endothelial Cell," Anns. Biomed. Eng., vol. 36, no. 4, pp. 554-562, 2007.
[18] J.-J. Chiu and S. Chien, "Effects of Disturbed Flow on Vascular Endothelium; Pathophysiological Basis and Clinical Perspectives," Physiol Rev, vol. 91, pp. 327- 387, 2011.
[19] R. Ross, "The Pathogenesis of Atherosclerosis: A Prospective for 1990s," Nature, vol. 363, pp. 801-809, 1993.
[20] Q. Guo, "The effect of Biomechanical and Biochemical Factors on endothelial Cells: Relevance to Atherosclerosis," PhD theis, University of Western Ontarion, London, Canada, p. 137, 2011.
Pathol., vol. 86, no. 3, p. 675–684, 1977.
[22] S. R. Bussolari, F. Dewey and M. A. Gimbrone, "Apparatus for Subjecting Living Cells to Fluid Shear Stress," Rev. Sci. Instrum., vol. 53, no. 12, pp. 1851-1854, 1982.
[23] T. Asakura and T. Karino, "Flow Patterns and Spatial Distribution of
Atherosclerotic Lesions in Human Coronary Arteries," Circ. Res., vol. 66, pp. 1045- 1066, 1990.
[24] C. M. Gibson, L. Diaz, K. Kandarpa, F. M. Sacks, R. C. Pasternak, T. Sandor, C. Feldman and P. H. Stone, "Relation of Vessel Wall Shear Stress to Atherosclerosis Progression in Human Coronary Arteries," J. Atheroscler. Thromb., vol. 13, pp. 310-315, 1993.
[25] R. M. Nerem, M. J. Levesque and J. F. Cornhill, "Vascular Endothelial Morphology as an Indicator of the Pattern of Blood Flow," J. Biomech. Eng., vol. 103, pp. 172- 176, 1981.
[26] S. Chien, "Molecular and Mechanical Bases of Focal Lipid Accumulation in Arterial Wall," Prog. Biophys. Mol. Biol., vol. 83, pp. 131-151, 2003.
[27] M. L. Albuquerque, C. M. Waters, U. Savla, H. W. Schnaper and A. S. Flozak, "Shear Stress Enhances Human Endothelial Cell Wound Closure in Vitro," Am. J. Physiol., vol. 279, pp. H293-H302, 2000.
[28] Y. Tardy, T. Resnick, T. Nagel, M. A. Gimbrone and C. Dewey, "Shear Stress Gradients Remodel Endothelial Monolayers in Vitro via a Cell Proliferation- Migration-Loss Cycle," Arterioscl. Thromb. Vasc. Biol., vol. 17, pp. 3102-3106, 1997.
[29] P. A. VanderLaan, C. A. Reardon and G. S. Getz, "Site Specificity of Atherosclerosis: Site-Selective Responses to Atherosclerotic Modulators," Arterioscler. Thromb. Vasc. Biol., vol. 24, pp. 12-22, 2004.
Flow Patterns on Endothelial Cell Migration into a Zone of Mechanical
Denudation," Biochem. Biophys. Res. Commun., vol. 285, no. 3, pp. 751-9, 2001. [31] K. Cunningham and A. I. Gotlieb, "The Role of Shear Stress in the Pathogenesis of
Atherosclerosis," Lab. Invest., vol. 85, pp. 9-23, 2005.
[32] N. Depaola, P. F. Davies, W. F. Pritchard, L. Florez, N. Harbeck and D. C. Polacek, "Spatial and Temporal Regulation of Gap Junction Connexin43 in Vascular
Endothelial Cells Exposed to Controlled Disturbed Flows in Vitro," Proc. Natl. Acad. Sci. USA, vol. 56, pp. 3154-3159, 1999.
[33] N. Sabrena, D. B. Cowan, A. I. Gotlieb and B. L. Langille, "Transient and Steady- State Effects of Shear Stress on Endothelial Cell Adherens Junctions," Circ. Res., vol. 85, no. 6, pp. 504-14, 1999.
[34] T. Nagel, N. Resnick, W. J. Atkinson, C. F. Dewey and M. A. Gimbrone, "Shear Stress Selectively Upregulates Intercellular Adhesion Molecule-1 Expression in Cultured Human Vascular Endothelial Cells," J. Clin. Invest., vol. 94, no. 2, p. 885– 891, 1994.
[35] A. J. Wheeler and A. R. Ganji, Introduction to Engineering Experimentation, Upper Saddle River: Prentice Hall, 2010.
[36] TSI Incorporated, Operation Manual: Phase Doppler Particle Analyzer (PDPA)/ Laser Doppler Velocimeter (LDV), Shoreview: MN, 2006.
[37] I. Flemin, B. Fisslthaler, M. Dixit and R. Busse, "Role of PECAM-1 in the Shear- Stress-Induced Activation of Akt and the Endothelail Nitric Oxide Synthase (eNOS) in Endothelail Cells," J. Cell Sci., vol. 118, pp. 4103-4111, 2005.
[38] D. E. Conway and M. A. Schwartz, "Mechanotransduction of Shear Stress Occurs through Changes in VE-Cadherin and PECAM-1 Tension: Implications for Cell Migration," Cell Adh. Migr., p. DOI:10.4161/19336918.2014.968498, 2014. [39] D. E. Conway, M. T. Breckennridge, E. Hinde, E. Gratton and C. S. Chen, "Fluid
Cadherin and PECAM-1," Curr. Biol., vol. 23, no. 11, pp. 1024-1030, 2013. [40] J. Eyckmans, T. Boudou, X. Yu, C. S. Chen, “A Hitchhiker’s Guide to
Chapter 2
2
An in Vitro Hemodynamic Facility to Study the Effects of
Quantified Shear Stresses on Endothelial Cells
The following chapter describes the development and validation of an in vitro hemodynamic facility for studying the effect of shear stress on endothelial cells. First, a detailed literature review will be presented, followed by the experimental details and results and finally, a discussion of the results obtained.