Skip to main content
SpringerLink
Log in

Phenotype Transformation of Aortic Valve Interstitial Cells Due to Applied Shear Stresses Within a Microfluidic Chip

  • Published:
Annals of Biomedical Engineering Aims and scope Submit manuscript

Abstract

Despite valvular heart diseases constituting a significant medical problem, the acquisition of information describing their pathophysiology remains difficult. Due to valvular size, role and location within the body, there is a need for in vitro systems that can recapitulate disease onset and progression. This study combines the development of an in vitro model and its application in the mechanical stimulation of valvular cell transformation. Specifically, porcine aortic valvular interstitial cells (PAVIC) were cultured on polydimethylsiloxane microfluidic devices with or without exposure to shear stresses. Mechanobiological responses of valvular interstitial cells were evaluated at shear stresses ranging from 0 to 4.26 dyn/cm2. When flow rates were higher than 0.78 dyn/cm2, cells elongated and aligned with the flow direction. In addition, we found that shear stress enhanced the formation of focal adhesions and up-regulated PAVIC transformation, assessed by increased expression of α-smooth muscle actin and transforming growth factor β. This study reveals a link between the action of shear forces, cell phenotype transformation and focal adhesion formation. This constitutes the first step towards the development of co-cultures (interstitial-endothelial cells) on organ-on-a-chip devices, which will enable studies of the signaling pathways regulating force-induced valvular degeneration in microtissues and potential discovery of valvular degeneration therapies.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
Figure 8

Similar content being viewed by others

References

  1. Aikawa, E., P. Whittaker, M. Farber, K. Mendelson, R. F. Padera, M. Aikawa, and F. J. Schoen. Human semilunar cardiac valve remodeling by activated cells from fetus to adult: implications for postnatal adaptation, pathology, and tissue engineering. Circulation. 113:1344–1352, 2006.

    Article  PubMed  Google Scholar 

  2. Arjunon, S., S. Rathan, H. Jo, and A. P. Yoganathan. Aortic valve: mechanical environment and mechanobiology. Ann. Biomed. Eng. 41(7):1331–1346, 2013.

    Article  PubMed  PubMed Central  Google Scholar 

  3. Aupperle, H., I. März, J. Thielebein, and H. Schoon. Expression of transforming growth factor-beta1, -beta2 and -beta3 in normal and diseased canine mitral valves. J. Comp. Pathol. 139(2–3):97–107, 2008.

    Article  CAS  PubMed  Google Scholar 

  4. Balachandran, K., P. Sucosky, H. Jo, and A. P. Yoganathan. Elevated cyclic stretch alters matrix remodeling in aortic valve cusps: implications for degenerative aortic valve disease. Am J Physiol Heart Circ Physiol 296(3):H756–H764, 2009.

    Article  CAS  PubMed  Google Scholar 

  5. Balachandran, K., P. Sucosky, H. Jo, and A. P. Yoganathan. Elevated cyclic stretch induces aortic valve calcification in a bone morphogenic protein-dependent manner. Am. J. Pathol. 177(1):49–57, 2010.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Balachandran, K., P. Sucosky, and A. P. Yoganathan. Hemodynamics and mechanobiology of aortic valve inflammation and calcification. Int. J. Inflamm. 2011:263870, 2011.

    Article  Google Scholar 

  7. Balachandran, K., S. Hussain, C.-H. Yap, M. Padala, A. H. Chester, and A. P. Yoganathan. Elevated cyclic stretch and serotonin result in altered aortic valve remodeling via a mechanosensitive 5-HT(2A) receptor-dependent pathway. Cardiovasc. Pathol. Off. J. Soc. Cardiovasc. Pathol. 21:206–213, 2011.

    Article  Google Scholar 

  8. Balachandran, K., M. A. Bakay, J. M. Connolly, X. Zhang, A. P. Yoganathan, and R. J. Levy. Aortic valve cyclic stretch causes increased remodeling activity and enhanced serotonin receptor responsiveness. Ann. Thorac. Surg. 92(1):147–153, 2011.

    Article  PubMed  PubMed Central  Google Scholar 

  9. Bhat, V. D., Truskey, G. A., Reichert, W. M., and Carolina, N. Fibronectin and avidin—biotin as a heterogeneous ligand system for enhanced endothelial cell adhesion, 14–16, 1997.

  10. Butcher, J. T., and R. M. Nerem. Valvular endothelial cells regulate the phenotype of interstitial cells in co-culture: effects of steady shear stress. Tissue Eng. 12:905–915, 2006.

    Article  CAS  PubMed  Google Scholar 

  11. Butcher, J. T., and R. M. Nerem. Valvular endothelial cells and the mechanoregulation of valvular pathology. Philos. Trans. R. Soc. Lond. B Biol. Sci. 362(1484):1445–1457, 2007.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Butcher, J. T., A. M. Penrod, A. J. Gaecía, and R. M. Nerem. Unique morphology and focal adhesion development of valvular endothelial cells in static and fluid flow environments. Arterioscler, Thromb. Vasc. Biology. 24(8):1429–1434, 2004.

    Article  CAS  Google Scholar 

  13. Butcher, J. T., C. A. Simmons, and J. N. Warnock. Review: mechanobiology of the aortic heart valve. J Heart Valve Dis 17:62–73, 2007.

    Google Scholar 

  14. Chen, H., J. Cornwell, H. Zhang, T. Lim, R. Resurreccion, T. Port, G. Rosengarten, and R. E. Nordon. Cardiac-like flow generator for long-term imaging of endothelial cell responses to circulatory pulsatile flow at microscale. Lab Chip 13(15):2999–3007, 2013.

    Article  CAS  PubMed  Google Scholar 

  15. Clark-Greuel, J. N., J. M. Connolly, E. Sorichillo, N. R. Narula, H. S. Rapoport, E. R. Mohler, J. H. Gorman, R. C. Gorman, and R. J. Levy. Transforming growth factor-β1 mechanisms in aortic valve calcification: increased alkaline phosphatase and related events. Ann. Thorac. Surg. 83(3):946–953, 2007.

    Article  PubMed  Google Scholar 

  16. Corcoran, B. M., A. Black, H. Anderson, J. D. McEwan, A. French, P. Smith, and C. Devine. Identification of surface morphologic changes in the mitral valve leaflets and chordae tendineae of dogs with myxomatous degeneration. Am. J. Vet. Res. 65(2):198–206, 2004.

    Article  PubMed  Google Scholar 

  17. Dubash, A. D., M. M. Menold, T. Samson, E. Boulter, R. García-Mata, R. Doughman, and K. Burridge. Chapter 1 focal adhesions: new angles on an old structure. Int. Rev. Cell Mol. Biol. 277(C):1–65, 2009.

    CAS  PubMed  Google Scholar 

  18. Farrar, E. J., and J. T. Butcher. Heterogeneous susceptibility of valve endothelial cells to mesenchymal transformation in response to TNFα. Ann. Biomed. Eng. 42(1):149–161, 2014.

    Article  PubMed  Google Scholar 

  19. Gould, R. A., K. Chin, T. P. Santisakultarm, A. Dropkin, J. M. Richards, C. B. Schaffer, and J. T. Butcher. Cyclic strain anisotropy regulates valvular interstitial cell phenotype and tissue remodeling in three-dimensional culture. Acta Biomater. 8(5):1710–1719, 2012.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Gupta, V., J. A. Werdenberg, J. S. Mendez, and K. J. Grande-Allen. Influence of strain on proteoglycan synthesis by valvular interstitial cells in three-dimensional culture. Acta Biomater. 4(1):88–96, 2008.

    Article  CAS  PubMed  Google Scholar 

  21. Hagler, M. A., T. M. Hadley, H. Zhang, K. Mehra, C. M. Roos, H. V. Schaff, R. M. Suri, and J. D. Miller. TGF-β signalling and reactive oxygen species drive fibrosis and matrix remodelling in myxomatous mitral valves. Cardiovasc. Res. 99(1):175–184, 2013.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Halldorsson, S., E. Lucumi, R. Gómez-Sjöberg, and R. M. T. Fleming. Advantages and challenges of microfluidic cell culture in polydimethylsiloxane devices. Biosens. Bioelectron. 63:218–231, 2015.

    Article  CAS  PubMed  Google Scholar 

  23. Harasaki, H., H. Hanano, J. Tanaka, K. Tokunaga, and M. Torisu. Surface structure of the human cardiac valve. A comparative study of normal and diseased valves. J. Cardiovasc. Surg. (Torino) 19(3):281–290, 1978.

    CAS  Google Scholar 

  24. Hinton, R. B., and K. E. Yutzey. Heart valve structure and function in development and disease. Ann. Rev. Physiol. 73:29–46, 2011.

    Article  CAS  Google Scholar 

  25. Jalali, S., M. A. del Pozo, K. Chen, H. Miao, Y. Li, M. A. Schwartz, J. Y. Shyy, and S. Chien. Integrin-mediated mechanotransduction requires its dynamic interaction with specific extracellular matrix (ECM) ligands. Proc. Natl. Acad. Sci. USA 98(3):1042–1046, 2001.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Jean, C., P. Gravelle, J.-J. Fournie, and G. Laurent. Influence of stress on extracellular matrix and integrin biology. Oncogene. 30(24):2697–2706, 2011.

    Article  CAS  PubMed  Google Scholar 

  27. Jian, B., N. Narula, Q. Li, E. R. M. Iii, and R. J. Levy. Progression of aortic valve stenosis: TGF- B1 is present in calcified aortic valve cusps and promotes aortic valve interstitial cell calcification via apoptosis. Ann. Thorac. Surg. 75:457–466, 2003.

    Article  PubMed  Google Scholar 

  28. Lacerda, C. M. R., and Orton, E. C. Evidence of a role for tensile loading in the pathogenesis of mitral valve degeneration. Clin. Exp. Cardiol. S3, 2012.

  29. Lacerda, C. M. R., H. B. MacLea, J. D. Kisiday, and E. C. Orton. Static and cyclic tensile strain induce myxomatous effector proteins and serotonin in canine mitral valves. J. Vet. Cardiol. 14(1):223–230, 2012.

    Article  PubMed  Google Scholar 

  30. Lacerda, C. M. R., J. D. Kisiday, B. Johnson, and E. C. Orton. Local serotonin mediates cyclic strain-induced phenotype transformation, matrix degradation, and glycosaminoglycan synthesis in cultured sheep mitral valves. Am. J. Physiol. Heart Circ. Physiol 302(10):H1983–H1990, 2012.

    Article  CAS  PubMed  Google Scholar 

  31. Lane, W. O., Jantzen, A. E., Carlon, T. A., Jamiolkowski, R. M., Grenet, J. E., Ley, M. M., Haseltine, J. M., Galinat, L. J., Lin, F.-H., Allen, J. D., Truskey, G. A., and Achneck, H. E. Parallel-plate flow chamber and continuous flow circuit to evaluate endothelial progenitor cells under laminar flow shear stress. J. Vis. Exp. (59):1–12, 1997.

  32. Lee, J., M. E. Razu, X. Wang, C. Lacerda, and J. J. Kim. Biomimetic cardiac microsystems for pathophysiological studies and drug screens. J. Lab Autom. 20(2):96–106, 2015.

    Article  CAS  PubMed  Google Scholar 

  33. Lindsey, S. E., J. T. Butcher, and H. C. Yalcin. Mechanical regulation of cardiac development. Front. Physiol. 5:1–15, 2014.

    Article  Google Scholar 

  34. Liu, A. C., V. R. Joag, and A. I. Gotlieb. The emerging role of valve interstitial cell phenotypes in regulating heart valve pathobiology. Am. J. Pathol. 171(5):1407–1418, 2007.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Lu, H., L. Y. Koo, W. M. Wang, D. A. Lauffenburger, L. G. Griffith, and K. F. Jensen. Microfluidic shear devices for quantitative analysis of cell adhesion. Anal. Chem. 76(18):5257–5264, 2004.

    Article  CAS  PubMed  Google Scholar 

  36. Mozaffarian, D., E. J. Benjamin, A. S. Go, D. K. Arnett, M. J. Blaha, M. Cushman, S. R. Das, S. de Ferranti, J.-P. Després, H. J. Fullerton, V. J. Howard, M. D. Huffman, C. R. Isasi, M. C. Jiménez, S. E. Judd, et al. Heart disease and stroke statistics—2016 update. Circulation 133:e38–e360, 2015.

    Article  PubMed  Google Scholar 

  37. Ohno, M., J. P. Cooke, V. J. Dzau, and G. H. Gibbons. Fluid shear stress induces endothelial transforming growth factor beta-1 transcription and production. Modulation by potassium channel blockade. J. Clin. Investig. 95(3):1363–1369, 1995.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Olson, E. N., and A. Nordheim. Linking actin dynamics and gene transcription to drive cellular motile functions. Nature reviews. Mol. Cell Biol. 11(5):353–365, 2010.

    CAS  Google Scholar 

  39. Paranya, G., S. Vineberg, E. Dvorin, S. Kaushal, S. J. Roth, E. Aikawa, F. J. Schoen, and J. Bischoff. Aortic valve endothelial cells undergo transforming growth factor-beta-mediated and non-transforming growth factor-beta-mediated transdifferentiation in vitro. Am. J. Pathol. 159:1335–1343, 2001.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Park, J. Y., S. J. Yoo, L. Patel, S. H. Lee, and S. H. Lee. Cell morphological response to low shear stress in a two-dimensional culture microsystem with magnitudes comparable to interstitial shear stress. Biorheology. 47(3–4):165–178, 2010.

    CAS  PubMed  Google Scholar 

  41. Platt, M. O., Y. Xing, H. Jo, and A. P. Yoganathan. Cyclic pressure and shear stress regulate matrix metalloproteinases and cathepsin activity in porcine aortic valves. J. Heart Valve Dis. 15:622–629, 2006.

    PubMed  Google Scholar 

  42. Provenzano, P. P., and P. J. Keely. Mechanical signaling through the cytoskeleton regulates cell proliferation by coordinated focal adhesion and Rho GTPase signaling. J. Cell Sci. 124(Pt 8):1195–1205, 2011.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Rosenkranz, S. TGF-beta1 and angiotensin networking in cardiac remodeling. Cardiovasc. Res. 63(3):423–432, 2004.

    Article  CAS  PubMed  Google Scholar 

  44. Schultz, J. E. J., S. A. Witt, B. J. Glascock, M. L. Nieman, P. J. Reiser, S. L. Nix, T. R. Kimballand, and T. Doetschman. TGF-β1 mediates the hypertrophic cardiomyocyte growth induced by angiotensin II. J. Clin. Investig. 109(6):787–796, 2002.

    Article  CAS  PubMed Central  Google Scholar 

  45. Schwarz, U. S., Erdmann, T., and Bischofs, I. B. Focal adhesions as mechanosensors: the two-spring model. BioSystems. 83(2–3 SPEC. ISS.):225–232, 2006.

  46. Sucosky, P., Balachandran, K., Elhammali, A., Jo, H., and Yoganathan, A. P. Altered shear stress stimulates upregulation of endothelial VCAM-1 and ICAM-1 in a BMP-4– and TGF-β1–dependent pathway. Arterioscler. Thromb. Vasc. Biol., 2008.

  47. Sun, L., N. M. Rajamannan, P. Sucosky, H. Jo, and A. Yoganathan. Defining the role of fluid shear stress in the expression of early signaling markers for calcific aortic valve disease. Aikawa E, ed. PLoS ONE. 8(12):e84433, 2013.

    Article  PubMed  PubMed Central  Google Scholar 

  48. Thayer, P., K. Balachandran, S. Rathan, C. H. Yap, S. Arjunon, H. Jo, and A. P. Yoganathan. The effects of combined cyclic stretch and pressure on the aortic valve interstitial cell phenotype. Ann. Biomed. Eng. 39(6):1654–1667, 2011.

    Article  PubMed  PubMed Central  Google Scholar 

  49. Tzima, E., M. A. del Pozo, S. J. Shattil, S. Chien, and M. A. Schwartz. Activation of integrins in endothelial cells by fluid shear stress mediates Rho-dependent cytoskeletal alignment. EMBO J. 20(17):4639–4647, 2001.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Velve-Casquillas, G., M. Le Berre, M. Piel, and P. T. Tran. Microfluidic tools for cell biological research. Nano Today. 5(1):28–47, 2010.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Walker, G. A., K. S. Masters, D. N. Shah, K. S. Anseth, and L. A. Leinwand. Valvular myofibroblast activation by transforming growth factor-beta: implications for pathological extracellular matrix remodeling in heart valve disease. Circ. Res. 95(3):253–260, 2004.

    Article  CAS  PubMed  Google Scholar 

  52. Walshe, T. E., N. G. Dela Paz, and P. A. D’Amore. The role of shear-induced transforming growth factor-β signaling in the endothelium. Arterioscler. Thromb. Vasc. Biol. 33(11):2608–2617, 2013.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Waltenberger, J., L. Lundin, K. Oberg, E. Wilander, K. Miyazono, C. H. Heldin, and K. Funa. Involvement of transforming growth factor-beta in the formation of fibrotic lesions in carcinoid heart disease. Am. J. Pathol. 142(1):71–78, 1993.

    CAS  PubMed  PubMed Central  Google Scholar 

  54. Waxman, A. S., B. G. Kornreich, R. A. Gould, N. S. Moïse, and J. T. Butcher. Interactions between TGFβ1 and cyclic strain in modulation of myofibroblastic differentiation of canine mitral valve interstitial cells in 3D culture. J. Vet. Cardiol. 14(1):211–221, 2012.

    Article  PubMed  PubMed Central  Google Scholar 

  55. Waxman, A. S., B. G. Kornreich, R. A. Gould, N. S. Moïse, and J. T. Butcher. Interactions between TGFβ1 and cyclic strain in modulation of myofibroblastic differentiation of canine mitral valve interstitial cells in 3D culture. J. Vet. Cardiol. 14(1):211–221, 2012.

    Article  PubMed  PubMed Central  Google Scholar 

  56. Weston, M. W., and A. P. Yoganathan. Biosynthetic activity in heart valve leaflets in response to in vitro flow environments. Ann. Biomed. Eng. 29(9):752–763, 2001.

    Article  CAS  PubMed  Google Scholar 

  57. Wheatley, D. J., J. Fisher, I. J. Reece, T. Spyt, and P. Breeze. Primary tissue failure in pericardial heart valves. J. Thorac. Cardiovasc. Surg. 94(3):367–374, 1987.

    CAS  PubMed  Google Scholar 

  58. Yang, B., C. Radel, D. Hughes, S. Kelemen, and V. Rizzo. P190 RhoGTPase-activating protein links the β1 integrin/caveolin-1 mechanosignaling complex to RhoA and actin remodeling. Arterioscler. Thromb. Vasc. Biol. 31(2):376–383, 2011.

    Article  CAS  PubMed  Google Scholar 

  59. Young, E. W. K., A. R. Wheeler, and C. A. Simmons. Matrix-dependent adhesion of vascular and valvular endothelial cells in microfluidic channels. Lab Chip 7:1759–1766, 2007.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

The authors would like to acknowledge Texas Tech University for new investigator start-up funds for Drs. Kim and Lacerda.

Author information

Authors and Affiliations

  1. Department of Chemical Engineering, Texas Tech University, Lubbock, TX, 79409, USA

    Xinmei Wang, Mir Ali & Carla M. R. Lacerda

  2. Department of Mechanical Engineering, Texas Tech University, Lubbock, TX, 79409, USA

    Joohyung Lee & Jungkyu Kim

  3. Department of Chemical Engineering, Texas Tech University, 6th and Canton Avenue, Box 43121, Lubbock, TX, 79409-3121, USA

    Carla M. R. Lacerda

Authors
  1. Xinmei Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Joohyung Lee
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Mir Ali
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jungkyu Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Carla M. R. Lacerda
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Carla M. R. Lacerda.

Additional information

Associate Editor Umberto Morbiducci oversaw the review of this article.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Wang, X., Lee, J., Ali, M. et al. Phenotype Transformation of Aortic Valve Interstitial Cells Due to Applied Shear Stresses Within a Microfluidic Chip. Ann Biomed Eng 45, 2269–2280 (2017). https://doi.org/10.1007/s10439-017-1871-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10439-017-1871-z

Keywords

Search

Navigation