Research Papers

The Role Of Extracellular Matrix Elasticity and Composition In Regulating the Nucleus Pulposus Cell Phenotype in the Intervertebral Disc: A Narrative Review

[+] Author and Article Information
Priscilla Y. Hwang

Department of Biomedical Engineering,
Duke University,
136 Hudson Hall, Box 90281,
Durham, NC 27708
e-mail: ph17@duke.edu

Jun Chen

Department of Orthopaedic Surgery,
Duke University,
375 Medical Science Research Building, Box 3093,
Durham, NC 27710
e-mail: junchen@duke.edu

Liufang Jing

Department of Biomedical Engineering,
Duke University,
136 Hudson Hall, Box 90281,
Durham, NC 27708
e-mail: lfjing@duke.edu

Brenton D. Hoffman

Department of Biomedical Engineering,
Duke University,
136 Hudson Hall, Box 90281,
Durham, NC 27708
e-mail: brenton.hoffman@duke.edu

Lori A. Setton

Department of Biomedical Engineering and Orthopaedic Surgery,
Duke University,
136 Hudson Hall, Box 90281,
Durham, NC 27708
e-mail: setton@duke.edu

1Corresponding author.

Contributed by the Bioengineering Division of ASME for publication in the JOURNAL OF BIOMECHANICAL ENGINEERING. Manuscript received October 3, 2013; final manuscript received December 13, 2013; accepted manuscript posted December 26, 2013; published online February 5, 2014. Editor: Beth Winkelstein.

J Biomech Eng 136(2), 021010 (Feb 05, 2014) (9 pages) Paper No: BIO-13-1460; doi: 10.1115/1.4026360 History: Received October 03, 2013; Revised December 13, 2013; Accepted December 26, 2013

Intervertebral disc (IVD) disorders are a major contributor to disability and societal health care costs. Nucleus pulposus (NP) cells of the IVD exhibit changes in both phenotype and morphology with aging-related IVD degeneration that may impact the onset and progression of IVD pathology. Studies have demonstrated that immature NP cell interactions with their extracellular matrix (ECM) may be key regulators of cellular phenotype, metabolism and morphology. The objective of this article is to review our recent experience with studies of NP cell-ECM interactions that reveal how ECM cues can be manipulated to promote an immature NP cell phenotype and morphology. Findings demonstrate the importance of a soft (<700 Pa), laminin-containing ECM in regulating healthy, immature NP cells. Knowledge of NP cell-ECM interactions can be used for development of tissue engineering or cell delivery strategies to treat IVD-related disorders.

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Bogduk, N., 2005, Clinical Anatomy of the Lumbar Spine and Sacrum, Elsevier, New York.
Urban, J. P., and Roberts, S., 2003, “Degeneration of the Intervertebral Disc,” Arthritis Res. Ther., 5(3), pp. 120–30. [CrossRef] [PubMed]
Baer A. E., Laursen, T. A., Guilak, F., and Setton, L. A., 2003, “The Micromechanical Environment of Intervertebral Disc Cells Determined by a Finite Deformation, Anisotropic, and Biphasic Finite Element Model,” ASME J. Biomech. Eng., 125(1), pp. 1–11. [CrossRef]
Hsieh, A. H., Wagner, D. R., Cheng, L. Y., and Lotz, J. C., 2005, “Dependence of Mechanical Behavior of the Murine Tail Disc on Regional Material Properties: A Parametric Finite Element Study,” ASME J. Biomech. Eng., 127(7), pp. 1158–1167. [CrossRef]
Cao, L., Guilak, F., and Setton, L. A., 2009, “Pericellular Matrix Mechanics in the Anulus Fibrosus Predicted by a Three-Dimensional Finite Element Model and In Situ Morphology,” Cell. Mol. Bioeng., 2(3), pp. 306–319. [CrossRef] [PubMed]
Jackson, A. R., Huang, C. Y., Brown, M. D., and Gu, W. Y., 2011, “3D Finite Element Analysis of Nutrient Distributions and Cell Viability in the Intervertebral Disc: Effects of Deformation and Degeneration,” ASME J. Biomech. Eng., 133(9), p. 091006. [CrossRef]
Cao, L., Guilak, F., and Setton, L. A., 2011, “Three-Dimensional Finite Element Modeling of Pericellular Matrix and Cell Mechanics in the Nucleus Pulposus of the Intervertebral Disk Based on In Situ Morphology,” Biomech. Model Mechanobiol., 10(1), pp. 1–10. [CrossRef] [PubMed]
Korecki, C. L., MacLean, J. J., and Iatridis, J. C., 2008, “Dynamic Compression Effects on Intervertebral Disc Mechanics and Biology,” Spine, 33(13), pp. 1403–1409. [CrossRef] [PubMed]
Boos, N., Weissbach, S., Rohrbach, H., Weiler, C., Spratt, K.F., and Nerlich, A.G., 2002, “Classification of Age-Related Changes in Lumbar Intervertebral Discs: 2002 Volvo Award in Basic Science,” Spine, 27(23), pp. 2631–2644. [CrossRef] [PubMed]
Liebscher, T., Haefeli, M., Wuertz, K., Nerlich, A.G., and Boos, N., 2011, “Age-Related Variation in Cell Density of Human Lumbar Intervertebral Disc,” Spine, 36(2), pp. 153–159. [CrossRef] [PubMed]
Roberts, S., Evans, H., Trivedi, J., and Menage, J., 2006, “Histology and Pathology of the Human Intervertebral Disc,” J. Bone Joint Surg. Am., 88(2), pp. 10–14. [CrossRef]
Johnson, W. E., Eisenstein, S. M., and Roberts, S., 2001, “Cell Cluster Formation in Degenerate Lumbar Intervertebral Discs is Associated With Increased Disc Cell Proliferation,” Connect Tissue Res., 42(3), pp. 197–207. [CrossRef] [PubMed]
Johnson, W. E., and Roberts, S., 2003, “Human Intervertebral Disc Cell Morphology and Cytoskeletal Composition: A Preliminary Study of Regional Variations in Health and Disease,” J. Anat., 203(6), pp. 605–612. [CrossRef] [PubMed]
Hastreiter, D., Ozuna, R. M., and Spector, M., 2001, “Regional Variations in Certain Cellular Characteristics in Human Lumbar Intervertebral Discs, Including the Presence of Alpha-Smooth Muscle Actin,” J. Orthop. Res., 19, pp. 597–604. [CrossRef] [PubMed]
Choi, K. S., Cohn, M. J., and Harfe, B. D., 2008, “Identification of Nucleus Pulposus Precursor Cells and Notochordal Remnants in the Mouse: Implications for Disk Degeneration and Chordoma Formation,” Dev. Dyn., 2008; 237(12), pp. 3953–3958. [CrossRef] [PubMed]
Roughley, P. J., 2004, “Biology of Intervertebral Disc Aging and Degeneration: Involvement of the Extracellular Matrix,” Spine, 29(23), pp. 2691–2699. [CrossRef] [PubMed]
Urban, J. P., 2002, “The Role of the Physicochemical Environment in Determining Disc Cell Behaviour,” Biochem. Soc. Trans., 30(6), pp. 858–864. [CrossRef] [PubMed]
Johnstone, B., and Bayliss, M. T., 1995, “The Large Proteoglycans of the Human Intervertebral Disc. Changes in Their Biosynthesis and Structure With Age, Topography, and Pathology,” Spine, 20(6), pp. 674–684. [CrossRef] [PubMed]
Nachemson, A., 1992, “Lumbar Mechanics as Revealed by Lumbar Intradiscal Pressure Measurements, The Lumbar Spine and Back Pain, 4th ed., M. I. V.Jayson, Churchill Livingstone, New York, pp. 157–171.
McNally, D. S., and Adams, M. A., 1992, “Internal Intervertebral Disc Mechanics as Revealed by Stress Profilometry,” Spine, 17, pp. 66–73. [CrossRef] [PubMed]
Hurri, H., and Karppinen, J., “Discogenic Pain,” Pain, 112(3), pp. 225–228. [CrossRef] [PubMed]
Peacock, A., 1951, Observations on the Prenatal Development of the Intervertebral Disc in Man,” J. Anat., 85(3), pp. 260–274. [PubMed]
Walmsley, R., 1953, The Development and Growth of the Intervertebral Disc, Edinburgh Med. J., 60(8), pp. 341–364.
Ellis, K., Bagwell, J., and Bagnat, M., 2013, “Notochord Vacuoles are Lysosome-Related Organelles That Function in Axis and Spine Morphogenesis,” J. Cell. Biol., 200(5), pp. 667–679. [CrossRef] [PubMed]
Gilchrist, C. L., Chen, J., Richardson, W. J., Loeser, R. F., and Setton, L. A., 2007, “Functional Integrin Subunits Regulating Cell-Matrix Interactions in the Intervertebral Disc,” J. Orthop. Res., 25(6), pp. 829–840. [CrossRef] [PubMed]
Hunter, C. J., Matyas, J. R., and Duncan, N. A., 2003, “The Three-Dimensional Architecture of the Notochordal Nucleus Pulposus: Novel Observations on Cell Structures in the Canine Intervertebral Disc,” J. Anat., 202(3), pp. 279–291. [CrossRef] [PubMed]
Trout, J. J., Buckwalter, J. A., Moore, K. C., and Landas, S. K., 1982, “Ultrastructure of the Human Intervertebral Disc I Changes in Notochordal Cells With Age,” Tissue Cell, 14(2), pp. 359–369. [CrossRef] [PubMed]
Minogue, B. M., Richardson, S.M., Zeef, L. A., Freemont, A. J., and Hoyland, J. A., 2010, “Characterization of the Human Nucleus Pulposus Cell Phenotype and Evaluation of Novel Marker Gene Expression to Define Adult Stem Cell Differentiation,” Arthritis Rheum., 62(12), pp. 3695–3705. [CrossRef] [PubMed]
Gotz, W., Kasper, M., Fischer, G., and Herken, R., 1995, “Intermediate Filament Typing of the Human Embryonic and Fetal Notochord,” Cell Tissue Res., 280(2), pp. 455–462. [CrossRef] [PubMed]
Buxboim, A., Ivanovska, I. L., and Discher, D. E., 2010, “Matrix Elasticity, Cytoskeletal Forces and Physics of the Nucleus: How Deeply do Cells ‘Feel’ Outside and In?” J. Cell. Sci., 123(3), pp. 297–308. [CrossRef] [PubMed]
Hayes, A. J., Benjamin, M., and Ralphs, J. R., 1999, “Role of Actin Stress Fibres in the Development of the Intervertebral Disc: Cytoskeletal Control of Extracellular Matrix Assembly,” Dev. Dyn., 215(3), pp. 179–189. [CrossRef] [PubMed]
Lehtonen, E., Stefanovic, V., and Saraga-Babic, M., 1995, “Changes in the Expression of Intermediate Filaments and Desmoplakins During Development of Human Notochord,” Differentiation, 59(1), pp. 43–49. [CrossRef] [PubMed]
Rodrigues-Pinto, R., Richardson, S. M., and Hoyland, J. A., 2013, “Identification of Novel Nucleus Pulposus Markers: Interspecies Variations and Implications for Cell-Based Therapies for Intervertebral Disc Degeneration,” Bone Joint Res., 2(8), pp. 169–178. [CrossRef] [PubMed]
Tang, X., Jing, L., and Chen, J., 2012, “Changes in the Molecular Phenotype of Nucleus Pulposus Cells With Intervertebral Disc Aging,” PLoS One, 7(12), pp. e52020–e52020-7. [CrossRef] [PubMed]
Minogue, B. M., Richardson, S. M., Zeef, L. A., Freemont, A. J., and Hoyland, J. A., 2010, “Transcriptional Profiling of Bovine Intervertebral Disc Cells: Implications for Identification of Normal and Degenerate Human Intervertebral Disc Cell Phenotypes,” Arthritis Res. Ther., 12(1), pp. R22-1–R22-20. [CrossRef]
Lv, F., Leung, V. Y., Huwang, S., Huang, Y., Sun, Y., and Cheung, K. M., 2013, “In Search of Nucleus Pulposus-Specific Molecular Markers,” Rheumatology: Advance Access published online: September 18, 2013. [CrossRef]
Li, S., Duance, V. C., and Blain, E. J., 2008, “Zonal Variations in Cytoskeletal Element Organization, mRNA and Protein Expression in the Intervertebral Disc,” J. Anat., 213(6), pp. 725–732. [CrossRef] [PubMed]
Guilak, F., Ting-Beall, H. P., Baer, A. E., Trickey, W. R., Erickson, G. F., and Setton, L. A., 1999, “Viscoelastic Properties of Intervertebral Disc Cells Identification of Two Biomechanically Distinct Cell Populations,” Spine, 24(23), pp. 2475–2483. [CrossRef] [PubMed]
Gilson, A., Dreger, M., and Urban, J. P., 2010, “Differential Expression Level of Cytokeratin 8 in Cells of the Bovine Nucleus Pulposus Complicates the Search for Specific Intervertebral Disc Cell Markers,” Arthritis Res. Ther., 12(1), p. R24. [CrossRef] [PubMed]
Stosiek, P., Kasper, M., and Karsten, U., 1988, “Expression of Cytokeratin and Vimentin in Nucleus Pulposus Cells,” Differentiation, 39(1), pp. 78–81. [CrossRef] [PubMed]
Hunter, C. J., Matyas, J. R., and Duncan, N. A., 2004, “Cytomorphology of Notochordal and Chondrocytic Cells From the Nucleus Pulposus: A Species Comparison,” J. Anat.205(5), pp. 357–362. [CrossRef] [PubMed]
Brown, M. J., Hallam, J. A., Colucci-Guyon, E., and Shaw, S., 2001, “Rigidity of Circulating Lymphocytes is Primarily Conferred by Vimentin Intermediate Filaments,” J. Immunol., 166(11), pp. 6640–6646. [PubMed]
Herrmann, H., Bar, H., Kreplak, L., Strelkov, S.V., and Aebi, U., 2007, “Intermediate Filaments: From Cell Architecture to Nanomechanics,” Nat. Rev. Mol. Cell. Biol., 8(7), pp. 562–573. [CrossRef] [PubMed]
Benjamin, M., Archer, C. W., and Ralphs, J. R., 1994, “Cytoskeleton of Cartilage Cells,” Microsc. Res. Tech., 28(5), pp. 372–377. [CrossRef] [PubMed]
Eggli, P. S., Hunziker, E. B., and Schenk, R. K., 1988, “Quantitation of Structural Features Characterizing Weight- and Less-Weight-Bearing Regions in Articular Cartilage: A Stereological Analysis of Medial Femoral Condyles in Young Adult Rabbits,” Anat. Rec., 222(3), pp. 217–227. [CrossRef] [PubMed]
Chen, J., Yan, W., and Setton, L. A., 2004, “Static Compression Induces Zonal-Specific Changes in Gene Expression for Extracellular Matrix and Cytoskeletal Proteins in Intervertebral Disc Cells In Vitro,” Matrix Biol., 22(7), pp. 573–583. [CrossRef] [PubMed]
Butler, W. F., 1988, “Comparative Anatomy and Development of the Mammalian Disc,” The Biology of the Intervertebral Disc, P.Gosh, ed., CRC, Boca Raton, pp. 39–82.
Lee, C. R, Sakai, D., Nakai, T., Toyama, K., Mochia, J., Alini, M., and Grad, S., 2007, “A Phenotypic Comparison of Intervertebral Disc and Articular Cartilage Cells in the Rat,” Eur. Spine J., 16(12), pp. 2174–2185. [CrossRef] [PubMed]
Sakai, D., Nakai, T., Mochia, J., Alini, M., and Grad, S., 2009, “Differential Phenotype of Intervertebral Disc Cells: Microarray and Immunohistochemical Analysis of Canine Nucleus Pulposus and Anulus Fibrosus,” Spine, 34(14), pp. 1448–1456. [CrossRef] [PubMed]
Risbud, M. V., Schaer, T. P., and Shapiro, I. M., 2010, “Toward an Understanding of the Role of Notochordal Cells in the Adult Intervertebral Disc: From Discord to Accord,” Dev. Dyn., 239(8), pp. 2141–-2148. [CrossRef] [PubMed]
Roberts, S., Ayad, S., and Menage, P. J., 1991, “Immunolocalisation of Type VI Collagen in the Intervertebral Disc,” Ann. Rheum. Dis., 50(11), pp. 787–791. [CrossRef] [PubMed]
Trout, J. J., Buckwalter, J. A., and Moore, K. C., 1982, “Ultrastructure of the Human Intervertebral Disc: II. Cells of the Nucleus Pulposus,” Anat. Rec., 204(4), pp. 307–314. [CrossRef] [PubMed]
Cao, L., Guilak, F., and Setton, L. A., 2007, Three-Dimensional Morphology of the Pericellular Matrix of Intervertebral Disc Cells in the Rat,” J. Anat., 211(4), pp. 444–452. [CrossRef] [PubMed]
Errington, R. J., Puustjarvi, K., White, I. R., Roberts, S., Urban, J. P., 1998, “Characterisation of Cytoplasm-Filled Processes in Cells of the Intervertebral Disc,” J. Anat., 192(3), pp. 369–378. [CrossRef] [PubMed]
Iatridis, J. C., Setton, L. A., Weidenbaum, M., and Mow, V. C., 1997, “The Viscoelastic Behavior of the Non-Degenerate Human Lumbar Nucleus Pulposus in Shear,” J. Biomech., 30(10), pp. 1005–1013. [CrossRef] [PubMed]
Iatridis, J. C, Weidenbaum, M., Setton, L. A., and Mow, V. C., 1996, “Is the Nucleus Pulposus a Solid or a Fluid? Mechanical Behaviors of the Nucleus Pulposus of the Human Intervertebral Disc,” Spine, 21(10), pp. 1174–1184. [CrossRef] [PubMed]
Cloyd, J. M., Malhotra, N. R., Weng, L., Chen, W., Mauck, R. L., and Elliott, D. M., 2007, Material Properties in Unconfined Compression of Human Nucleus Pulposus, Injectable Hyaluronic Acid-Based Hydrogels and Tissue Engineering Scaffolds,” Eur Spine J., 16(11), pp. 1892–1898, [CrossRef] [PubMed]
Eyre, D. R., and Muir, H., 1977, “Quantitative Analysis of Types I and II Collagens in Human Intervertebral Discs at Various Ages,” Biochim. Biophys. Acta.492(1), pp. 29–42. [CrossRef] [PubMed]
Gower, W. E., and Pedrini, V., 1969, “Age-Related Variations in Proteinpolysaccharides From Human Nucleus Pulposus, Annulus Fibrosus, and Costal Cartilage,” J Bone Joint Surg. Am., 51(6), pp. 1154–1162. [PubMed]
Taylor, J. R., and Twomey, L. T., 1988, “The Development of the Human Intervertebral Disc,” The Biology of the Intervertebral Disc, P.Gosh, ed., CRC, Boca Raton, FL., pp. 39–82.
Roberts, S., Menage, J., Duance, V., Wotton, S., and Ayad, S., 1991, “Volvo Award in Basic Sciences Collagen Types Around the Cells of the Intervertebral Disc and Cartilage End Plate: An Immunolocalization Study,” Spine, 16(9), pp. 1030–1038. [CrossRef] [PubMed]
Melrose, J., Ghosh, P., and Taylor, T. K., 2001, “A Comparative Analysis of the Differential Spatial and Temporal Distributions of the Large (Aggrecan, Versican) and Small (Decorin, Biglycan, Fibromodulin) Proteoglycans of the Intervertebral Disc,” J. Anat., 198(1), pp. 3–15. [CrossRef] [PubMed]
Yu, J., 2002, “Elastic Tissues of the Intervertebral Disc,” Biochem. Soc. Trans, 30(6), pp. 848–852. [CrossRef] [PubMed]
Chen, J., Jing, L., Gilchrist, C. L., Richardson, W. J., Fitch, R. D., and Setton, L. A., 2009, “Expression of Laminin Isoforms, Receptors and Binding Proteins Unique to Nucleus Pulposus Cells of Immature Intervertebral Disc,” Connect Tissue Res., 50, pp. 294–306. [PubMed]
Nettles, D. L., Richardson, W. J., and Setton, L. A., 2004, “Integrin Expression in Cells of the Intervertebral Disc,” J. Anat.204(6), pp. 515–520. [CrossRef] [PubMed]
Natarajan, R. N., Williams, J. R., and Andersson, G. B., 2004, “Recent Advances in Analytical Modeling of Lumbar Disc Degeneration,” Spine, 29(23), pp. 2733–2741. [CrossRef] [PubMed]
Discher, D. E., Janmey, P., and Wang, Y. L., 2005, “Tissue Cells Feel and Respond to the Stiffness of Their Substrate,” Science, 310(5751), pp. 1139–1143. [CrossRef] [PubMed]
Pelham, R. J., Jr., and Wang, Y., 1997, “Cell Locomotion and Focal Adhesions are Regulated by Substrate Flexibility,” Proc. Natl. Acad. Sci. U.S.A, 94(25), pp. 13661–5. [CrossRef] [PubMed]
Guo, W. H., Frey, M. T., Burnham, N. A., and Wang, Y. L., 2006, “Substrate Rigidity Regulates the Formation and Maintenance of Tissues,” Biophys. J., 90(6), pp. 2213–2220. [CrossRef] [PubMed]
Gilchrist., C. L., Darling, E. M., Chen, J., and Setton, L. A., 2011, “Extracellular Matrix Ligand and Stiffness Modulate Immature Nucleus Pulposus Cell-Cell Interactions,” PLoS One, 6(11), pp. e27170-1–e27170-9. [CrossRef]
Johannessen, W., and Elliott, D. M., 2005, “Effects of Degeneration on the Biphasic Material Properties of Human Nucleus Pulposus in Confined Compression,” Spine, 30(24), pp. E724–E729. [CrossRef] [PubMed]
Iatridis, J. C., Setton, L. A., Weidenbaum, M., and Mow, V. S., 1997, “Alterations in the Mechanical Behavior of the Human Lumbar Nucleus Pulposus With Degeneration and Aging,” J. Orthop. Res., 15(2), pp. 318–322. [CrossRef] [PubMed]
Miner, J. H., and Yurchenco, P. D., 2004, “Laminin Functions in Tissue Morphogenesis,” Annu. Rev. Cell. Dev. Biol., 20, pp. 255–284. [CrossRef] [PubMed]
Zaidel-Bar, R,. Shalev, I., Ma'ayan A., Iyengar, R., Geiger, B., 2007, “Functional Atlas of the Integrin Adhesome,” Nat. Cell. Biol., 9(8), pp. 858–867. [CrossRef] [PubMed]
Hoffman, B. D., Grashoff, C., and Schwartz, M. A., 2011, “Dynamic Molecular Processes Mediate Cellular Mechanotransduction,” Nature, 475(7356), pp. 316–323. [CrossRef] [PubMed]
Hynes, R. O., 1992, “Integrins: Versatility, Modulation, and Signaling in Cell Adhesion,” Cell, 69(1), pp. 11–25. [CrossRef] [PubMed]
Schwartz, M. A., 2010, “Integrins and Extracellular Matrix in Mechanotransduction,” Cold Spring Harb. Perspect. Biol., 2(12), pp. a005066-1–a005066-13. [CrossRef]
Bridgen, D. T., Gilchrist, C. L., Richardson, W. J., Isaacs, R. E., Brown, C. R., Yang, K. L., Chen, J., and Setton, L. A., 2013, “Integrin-Mediated Interactions With Extracellular Matrix Proteins for Nucleus Pulposus Cells of the Human Intervertebral Disc,”. J. Orthop. Res., 31(10), pp. 1661–1667. [CrossRef] [PubMed]
Le Maitre, C. L., Frain, J., Millward-Sadler, J., Fotheringham, A. P., Freemont, A. J., and Hoyland, J. A., 2009, “Altered Integrin Mechanotransduction in Human Nucleus Pulposus Cells Derived From Degenerated Discs,” Arthritis Rheum., 60(2), pp. 460–469. [CrossRef] [PubMed]
Anderson, D. G., Li, X., and Balian, G., 2005, “A Fibronectin Fragment Alters the Metabolism by Rabbit Intervertebral Disc Cells in vitro,” Spine, 30(11), pp. 1242–1246. [CrossRef] [PubMed]
Aota, Y., An, H. S., Homandberg, G., Thonar, E. J., Andersson, G. B., Pichika, R., and Masuda, K., 2005, “Differential Effects of Fibronectin Fragment on Proteoglycan Metabolism by Intervertebral Disc Cells: A Comparison With Articular Chondrocytes,” Spine, 30(7), pp. 722–728. [CrossRef] [PubMed]
Chen, J., Yan, W., and Setton, L. A., 2006, “Molecular Phenotypes of Notochordal Cells Purified From Immature Nucleus Pulposus,” Eur. Spine J., 15(3), pp. S303–S311. [CrossRef] [PubMed]
Gilchrist, C. L., Francisco, A. T., Plopper, G. E., Chen, J., and Setton, L. A., 2011, “Nucleus Pulposus Cell-Matrix Interactions With Laminins,” Eur. Cell. Mater, 21, pp. 523–532. [PubMed]
Wang, Y. L., and Pelham, R. J., Jr., 1998, “Preparation of a Flexible, Porous Polyacrylamide Substrate for Mechanical Studies of Cultured Cells,” Methods Enzymol., 298, pp. 489–496. [CrossRef] [PubMed]
Hassell, J. R., Robey, P. G., Barrach, H. J., Wilczek, J., Rennard, S. I., and Martin, G. F., 1980, “Isolation of a Heparan Sulfate-Containing Proteoglycan From Basement Membrane,” Proc. Natl. Acad. Sci. U.S.A., 77(8), pp. 4494–4498. [CrossRef] [PubMed]
Zhang, Y.H., Zhao, C.Q., Jiang, L.S., and Dai, L.Y., 2011, “Substrate Stiffness Regulates Apoptosis and the mRNA Expression of Extracellular Matrix Regulatory Genes in the Rat Annular Cells,” Matrix Biol., 30(2), pp. 135–144. [CrossRef] [PubMed]
Engler, A. J., Sen, S., Sweeney, H. L., and Discher, D. E., 2006, “Matrix Elasticity Directs Stem Cell Lineage Specification,” Cell, 126(4) pp. 677–689. [CrossRef] [PubMed]
Yeung, T., Georges, P. C., Flanagan, L. A., Marg, B., Ortiz, M., Funaki, M., Zahir, N., Ming, W., Weaver, V., and Janmey, P. A., 2005, “Effects of Substrate Stiffness on Cell Morphology, Cytoskeletal Structure, and Adhesion,” Cell. Motil. Cytoskeleton, 60(1), pp. 24–34. [CrossRef] [PubMed]
Wang, H. B., Dembo, M., and Wang, Y. L., 2000, “Substrate Flexibility Regulates Growth and Apoptosis of Normal but not Transformed Cells,” Am. J. Physiol. Cell. Physiol., 279(5), pp. C1345–C1350. [PubMed]
Weber, G. F., Bjerke, M. A., and DeSimone, D. W., 2011, “Integrins and Cadherins Join Forces to Form Adhesive Networks,” J. Cell. Sci., 124(8), pp. 1183–1193. [CrossRef] [PubMed]
Schwartz, M. A., and DeSimone, D. W., 2008, “Cell Adhesion Receptors in Mechanotransduction,” Curr. Opin. Cell. Biol., 20(5), pp. 551–556. [CrossRef] [PubMed]
Braga, V. M., 2002, “Cell-Cell Adhesion and Signalling,” Curr. Opin. Cell. Biol., 14(5), pp. 546–556. [CrossRef] [PubMed]
Arthur, W. T., Noren, N. K., and Burridge, K., 2002, “Regulation of Rho Family GTPases by Cell-Cell and Cell-Matrix Adhesion,” Biol. Res., 35(2), pp. 239–246. [CrossRef] [PubMed]
Amano, M., Ito, M., Kimura, K., Fukata, Y., Chihara, K., Nakano, T., Matsuura, Y., and Kaibuchi, K., 1996, “Phosphorylation and Activation of Myosin by Rho-Associated Kinase (Rho-Kinase),” J. Biol. Chem., 271(34), pp. 20246–20249. [CrossRef] [PubMed]
Amano, M., Chihara, K., Kazushi, K., Fukata, Y., Nakamura, N., Matsuura, Y., and Kaibuchi, K., 1997, “Formation of Actin Stress Fibers and Focal Adhesions Enhanced by Rho-Kinase,” Science, 275(5304), pp. 1308–1311. [CrossRef] [PubMed]
Kimura, K., Ito, M., Amano, M., Chihara, K., Fukata, Y., Nakafuku, M., Yamamori, B., Feng, J., Nakano, T., Okawa, K., Iwamatsu, A., and Kaibuchi, K., 1996, Regulation of Myosin Phosphatase by Rho and Rho-Associated Kinase (Rho-Kinase), Science, 273(5272), pp. 245–248. [CrossRef] [PubMed]
Hall, A., 2005, “Rho GTPases and the Control of Cell Behaviour,” Biochem. Soc. Trans., 33(5), pp. 891–895. [CrossRef] [PubMed]
Burridge, K., and Wennerberg, K., 2004, “Rho and Rac Take Center Stage,” Cell, 116(2), pp. 167–179. [CrossRef] [PubMed]
Yamada, S., and Nelson, W. J., 2007, “Localized Zones of Rho and Rac Activities Drive Initiation and Expansion of Epithelial Cell-Cell Adhesion,” J. Cell. Biol., 178(3), pp. 517–527. [CrossRef] [PubMed]
Noren, N. K., Niessen, C. M., Gumbiner, B. M., and Burridge, K., 2001, “Cadherin Engagement Regulates Rho Family GTPases,” J. Biol. Chem., 276(36), pp. 33305–33308. [CrossRef] [PubMed]
Fujita, N., Miyamoto, T., Imai, J., Hosogane, N., Suzuki, T., Yagi, M., Morita, K., Ninomiya, K., Miyamoto, K., Takaishi, H., Matsumoto, M., Morioka, H., Yabe, H., Chiba, K., Watanabe, S., Toyama, Y., and Suda, T., 2005, “CD24 is Expressed Specifically in the Nucleus Pulposus of Intervertebral Discs,” Biochem Biophys Res Commun, 338(4), pp. 1890–1896. [CrossRef] [PubMed]
Tang, X. J. L., Setton, L. A., Richardson, W. J., Isaacs, R. E., Fitch, R. D., Brown, C. R., and Chen, J., 2013, “Identifying the Molecular Phenotype of Cells in the Human Intervertebral Disc Reveals the Existence of a Unique Notochordal-Like Cell Population,” Trans. Orthop. Res. Soc., 38(859), p. 0859.
Weiler, C., Nerlich, A.G., Schaaf, R., Bachmeier, B.E., Wuertz, K., and Boos, N., 2010, “Immunohistochemical Identification of Notochordal Markers in Cells in the Aging Human Lumbar Intervertebral Disc,” Eur. Spine J., 19(10), pp. 1761–170. [CrossRef] [PubMed]
Nerlich, A. G., Schleicher, E. D., and Boos, N., 1997, “Volvo Award Winner in Basic Science Studies. Immunohistologic Markers for Age-Related Changes of Human Lumbar Intervertebral Discs,” Spine, 22(24), pp. 2781–2795. [CrossRef] [PubMed]


Grahic Jump Location
Fig. 1

The intervertebral disc is situated between vertebral bodies in the spinal column, and acts to support loads, provide flexibility, and dissipate energy in the spine. The disc is comprised of distinct anatomic zones: the anulus fibrosus (AF), nucleus pulposus (NP), and cartilage endplates. The AF consists of concentric lamella of highly-aligned collagen fibers, with cells typically aligned along the fiber direction. The NP is a gelatinous, highly-hydrated tissue, with cells typically exhibiting rounded, unaligned morphologies. Staining is safranin O and fast green. Images of specific cell morphology in each region were obtained via light microscopy.

Grahic Jump Location
Fig. 2

Porcine NP cells preferentially attach and spread upon laminin-containing substrates. (a) Fraction of adherent cells remaining attached to ECM substrates following application of centrifugal detachment force. Higher numbers of NP cells resist detachment when adherent to laminin ligands (isoforms LM-332, LM-511, LM-111), as compared to collagen and fibronectin ECM ligands ((b) and (c)) NP cell spreading and NP cell shape dynamics on ECM substrates. NP cells on laminin isoforms LM-332 and LM-511 spread rapidly and to a greater extent as compared to other matrix substrates NP cells on laminin isoforms. Additionally, NP cells lost their original shape factor as the cells spread on laminin isoforms (error bars omitted for clarity, significant effects of substrate and time were detected via two-way ANOVA, p < 0.05; substrates not labeled with same letter were statistically different. (LM = laminin, FN = fibronectin, BSA = bovine serum albumin, CM = cultured media) Specific methods described in detail and image adapted from Gilchrist et al. 2011 [83].

Grahic Jump Location
Fig. 3

Soft laminin-containing substrates promote immature NP cells to form multicell clusters, while retaining cell dimensions and rounded morphology. Actin immunostaining of immature porcine NP cell behavior on BME-functionalized polyacrylamide gel (BME-PAAm) (100 and 290 Pa), “soft” BME (300 Pa), and “stiff” BME (2900 Pa) substrates after 7 days of culture (green = actin (phalloidin), red = cell nuclei (propidium iodide), bar = 100 μm). Specific methods and image adapted from Gilchrist et al. 2011 [70].

Grahic Jump Location
Fig. 4

Changes in immature porcine NP cell morphology on substrates. (a) Immature porcine NP cells spread out on stiff BME but maintain rounded morphology on soft BME. (b) Immature porcine NP cells have significantly decreased cell velocity on soft BME upon formation of cell cluster. On stiff BME, NP cells continue to send out lamellipodia and filopodia as if sensing the underlying substrate. (c) Immature porcine NP cells transfected with GFP-actin display distinct actin fibers as the cell spreads and attaches to the underlying stiff BME substrate. On soft BME, NP cells remain rounded and do not have any actin stress fiber formation. Methods for substrate development were adapted from Gilchrist et al. 2011 [70]. Imaging and analysis performed using the Olympus VivaView Fluorescent Incubator Microscope and Metamorph Software, in collaboration with the Duke Light Microscopy Core Facility.

Grahic Jump Location
Fig. 5

Matrix production and changes in gene expression in immature porcine NP cells cultured upon various substrates (a) Matrix production in immature porcine NP cells on soft BME substrates is significantly higher (*p < 0.05, One-way ANOVA, with Tukey's post hoc analysis) than matrix production in NP cells on all other substrates. (b) Gene expression was calculated relative to values for 18 s mRNA and normalized by values for stiff BME. mRNA values for NP-specific and NP-matrix-related markers were higher in immature porcine NP cells on soft BME substrates compared to all other substrates Methods for biochemical assays are adapted from Gilchrist et al. 2011 [70], and methods for gene expression are adapted from Tang et al. 2012 [34]

Grahic Jump Location
Fig. 6

Treatment of immature porcine NP cells with Rho GTPase inhibitors, ROCK (Y27632) and Rac1 (NSC23766). (a) Immature porcine NP cells are unable to form cell clusters on soft BME substrates after treatment with ROCK inhibitor but not Rac1 inhibitor (green = phalloidin, red = propidium iodide, bar = 50 μm). (b) Decreased matrix production in NP cells after 4-day treatment with ROCK inhibitor on soft BME substrates (*p < 0.01, **p < 0.05, Two-way ANOVA with Tukey's post hoc analysis); matrix production is unaffected by ROCK inhibitor in NP cells on all other substrates. (c) Gene expression was calculated relative to values for 18 s mRNA and normalized by values for stiff BME. mRNA values for NP-specific and NP-matrix-related markers are decreased in immature porcine NP cells after 4-day treatment with ROCK inhibitor on soft BME substrates. Methods for biochemical assays are adapted from Gilchrist et al. 2011 [70], and methods for gene expression are adapted from Tang et al. 2012 [34].



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