Stem Cell-Based Self-Assembled Tissues Cultured on a Nano-Periodic-Structured Surface Patterned Using Femtosecond Laser Processing
Hiromichi Fujie*1,*2, Kei Oya*3, Yuki Tani*1, Kenji Suzuki*4, and Norimasa Nakamura*5
*1Graduate School of System Design, Tokyo Metropolitan University
6-6 Asahigaoka, Hino, Tokyo 191-0065, Japan
*2Research Institute for Science and Technology, Kogakuin University, Tokyo, Japan
*3Department of Materials and Life Science, Faculty of Science and Technology, Seikei University, Tokyo, Japan
*4Department of Mechanical Systems Engineering, Kogakuin University, Tokyo, Japan
*5Graduate School of Medicine, Osaka University, Osaka, Japan
The present study was conducted to determine the effects of a nano-periodic-structured surface on the morphological and mechanical properties of a stem-cell-based self-assembled tissue (scSAT) developed for biological tissue repair. Nano-periodic groove structures were patterned on a pure titanium surface using femtosecond laser processing, and the structure was replicated on polydimethylsiloxane (PDMS). The depth, periodic pitch, and surface roughness (Ra) of the PDMS grooves were 48 ± 21 nm, 522 ± 9 nm, and 17 ± 5 nm, respectively. Human synovial cells, including mesenchymal stem cells, were subjected to 4-time cell passage, and then cultured on the PDMS surface at a density of 4.0 × 105 cells/cm2 in a growth medium with 0.2 mM ascorbic acid 2-phosphate to produce scSATs (nano-scSAT). For comparison, some of the cells subjected to 4-time cell passage were cultured on either a flat PDMS substrate with 6 ± 1 nm of surface roughness (Ra) (flat-scSAT) or a commercially available cell culture plate of polystyrene (normal-scSAT), at a cell density identical to that in the nano-scSAT group. At 28 days of cell culture, the scSATs were gently detached from the culture plates and subjected to morphological observation and mechanical testing. Microscopic observation revealed that the nano-scSATs exhibited a dense tissue of cells and an extracellular matrix with an anisotropic structure, while the flat- and normal-scSATs exhibited a sparse and isotropic structure. The tangent modulus and tensile strength were significantly higher in the nano-scSATs than in the flat- and normal-scSATs. These results suggest that a nano-periodic-structured surface improves the morphological and mechanical properties of scSATs.
-  W. Ando, K. Tateishi, D. A. Hart, D. Katakai, Y. Tanaka, K. Nakata, J. Hashimoto, H. Fujie, K. Shino, H. Yoshikawa, and N. Nakamura, “Cartilage Repair Using an In Vitro Generated Scaffold-Free Tissue-Engineered Construct Derived from Porcine Synovial Mesenchymal Stem Cells,” Biomaterials, Vol.28, pp. 5462-5470, 2007.
-  A. I. Teixeira, G. A. Abrams, P. J. Bertices, C. J. Murphy, and P. F. Nealey, “Epithelial Contact Guidance on Well-Defined Micro- and Nanostructured Substrates,” Journal of Cell Science, Vol.116, pp. 1881-1192, 2003.
-  L. Zhang, V. M. Menendez-Flores, N. Murakami, and T. Ohno, “Improvement of Photocatalytic Activity of Brookite Titanium Dioxide Nanorods by Surface Modification Using Chemical Etching,” Applied Surface Science, Vol.258, pp. 5803-5809, 2012.
-  S. Ban, Y. Iwaya, H. Kono, and H. Sato, “Surface Modification of Titanium by Etching in Concentrated Sulfuric Acid,” Dental Materials, Vol.22, pp. 1115-1120, 2006.
-  H. Asoh, K. Uchibori, and S. Ono, “Anisotropic Chemical Etching of Silicon Through Anodic Oxide Films Formed on Silicon Coated with Microspheres,” Semiconductor Science Technology, Vol.26, 102001, 2011.
-  H. Asoh, F. Arai, and S. Ono, “Effect of Noble Metal Catalyst Species on Morphology of Macroporous Silicon Formed by Metal-Assisted Chemical Etching,” Electrochimica Acta, Vol.54, pp. 5142-5148, 2009.
-  D. Z. Xie, B. K. A. Ngoi, Y. Q. Fu, A. S. Ong, and B. H. Lim, “Etching Characteristics of Tini Thin Film by Focused Ion Beam,” Applied Serface Science, Vol.225, pp. 54-58, 2004.
-  S. Rennon, L. Bach, H. Köonig, J. P. Reithmaier, A. Forchel, J. L. Gentner, and L. Goldstein, “Nanoscale Patterning by Focused Ion Beam Enhanced Etching for Optoelectronic Device Fabrication,” Microelectronic Engineering, Vol.57-58, pp. 891-896, 2001.
-  Y. B. Xiao, E. H. Kim, S. M. Kong, J. H. Park, B. C. Min, and C. W. Chung, “Inductively Coupled Plasma Reactive Ion Etching of Titanium Thin Films Using A Cl2/Ar Gas,” Vacuum, Vol.85, pp. 434-438, 2010.
-  K-H. Leitz, B. Redlingshöofer, Y. Reg, A. Otto, and M. Schmidt, “Metal Ablation with Short and Ultrashort Laser Pulses,” Physical Procedia, Vol.12, pp. 230-238, 2011.
-  V. Oliveira, S. Ausset, and R. Vilar, “Surface Micro/Nanostructuring of Titanium under Stationary and Non-Stationary Femtosecond Laser Irradiation,” Applied Surface Science, Vol.255, pp. 7556-7560, 2009.
-  J. Reif, O. Varlamova, and F. Costache, “Femtosecond Laser Induced Nanostructure Formation: Self-Organization Control Parameters,” Journal of Applied Physics, Vol.92, pp. 1019-1024, 2008.
-  K. Oya, S. Aoki, K. Shimomura, N. Sugita, K. Suzuki, N. Nakamura, and H. Fujie, “Morphological Observations of Mesenchymal Stem Cell Adhesion to a Nano-Periodic Structured Titanium Surface Patterned Using Femtosecond Laser Processing,” Japanese Journal of Applied Physics, Vol.51, 125203, 2012.
-  W. Ando, K. Tateishi, D. Katakai, D. A. Hart, C. Higuchi, K. Nakata, J. Hashimoto, H. Fujie, K. Shino, H. Yoshikawa, and N. Nakamura, “In Vitro Generation of a Scaffold-Free Tissue-Engineered Construct (TEC) Derived from Human Synovial Mesenchymal Stem Cells: Biological and Mechanical Properties and Further Chondrogenic Potential,” Tissue Engineering Part A, Vol.14, pp. 2041-2049, 2008.
-  K. Shimomura, W. Ando, K. Tateishi, R. Nansai, H. Fujie, D. A. Hart, H. Kohda, K. Kita, T. Kanamoto, T. Mae, K. Nakata, K. Shino, H. Yoshikawa, and N. Nakamura, “The Influence of Skeletal Maturity on Allogenic Synovial Mesenchymal Stem Cell-Based Repair of Cartilage in a Large Animal Model,” Biomaterials, Vol.31, pp. 8004-8011, 2010.
-  M. J. Karnovsky, “A Formaldehyde-Glutaraldehyde Fixative of High Osmolality for Use in Electron Microscopy,” Journal of Cell Biology, Vol.27, pp. 137-138, 1965.
-  K. Saito, A. Ogawa, W. Ando, N. Nakamura, and H. Fujie, “Effects of Cyclic Tensioning Duration on the Mechanical Property of a Stem Cell-Based Self-Assembled Tissue (scSAT) Derived from Synovim,” Transactions of the Bioengineering Conference (ASME), 204339, 2009.
-  S. Sakabe, M. Hashida, S. Tokita, S. Namba, and K. Okamuro, “Mechanism for Self-Formation of Periodic Grating Structures on a Metal Surface by a Femtosecond Laser Pulse,” Physical Review B, Vol.79, 033409, 2009.
-  F. Costache, M. Henyk, and J. Reif, “Surface Patterning on Insulators upon Femtosecond Laser Ablation,” Applied Surface Science, Vol.208-209, pp. 486-491, 2003.
-  D. V. Tran, H. Y. Zheng, Y. C. Lam, V. M. Murukeshan, J. C. Chai, and D. E. Hardt, “Femtosecond Laser-Induced Damage Morphologies of Crystalline Silicon by Sub-Threshold Pulses,” Optics and Lasers in Engineering, Vol.43, pp. 977-986, 2005.
-  B. Alberts, A. Johnson, J. Lewis, M. Raff, K. Roberts, and P. Walter, “Molecular Biology of the Cell 5th Edition,” Garland Science, Abingdon, UK, 2008.
-  S. Fujita, D. Ono, M. Ohshima, and H. Iwata, “Supercritical CO2-Assisted Embossing for Studying Cell Behaviour on Microtextured Surfaces,” Biomaterials, Vol.29, pp. 4494-4500, 2008.
-  J. H. Wang, F. Jia, T. W. Gilbert, and S. LY. Woo, “Cell Orientation Determines the Alignment of Cell-Produced Collagenous Matrix,” Journal of Biomechanics, Vol.36, pp. 97-102, 2003.
-  K. Shimomura, Y. Moriguchi, W. Ando, R. Nansai, H. Fujie, D. A. Hart, A. Gobbi, K. Kita, S. Horibe, K. Shino, H. Yoshikawa, and N. Nakamura, “Osteochondral Repair Using a Scaffold-Free Tissue-Engineered Construct Derived from Synovial Mesenchymal Stem Cells and a Hydroxyapatite-Based Artificial Bone,” Tissue Engineering Part A, Vol.20, pp. 2291-2304, 2014.
-  D. Katakai, M. Imura, W. Ando, K. Tateishi, H. Yoshikawa, N. Nakamura, and H. Fujie, “Compressive Properties of Cartilage-Like Tissues Repaired in Vivo With Scaffold-Free, Tissue Engineered Constructs,” Clinical Biomechanics, Vol.24, pp. 110-116, 2009.
-  R. Nansai, T. Suzuki, K. Shimomura, W. Ando, N. Nakamura, and H. Fujie, “Surface Morphology and Stiffness of Cartilage-Like Tissue Repaired with a Scaffold-Free Tissue Engineered Construct,” Journal of Biomechanical Science and Engineering, Vol.6, pp. 40-48, 2011.
-  W. Ando, H. Fujie, Y. Moriguch, R. Nansa, K. Shimomura, D. A. Hart, H. Yoshikawa, and N. Nakamur, “Detection of Abnormalities in The Superficial Zone of Cartilage Repaired Using a Tissue Engineered Construct Derived from Synovial Stem Cells,” European Cells and Materials Journal, Vol.24, pp. 292-307, 2012.