Material properties and osteogenic differentiation of marrow stromal cells on fiber-reinforced laminated hydrogel nanocomposites.

The fibrils in the bone matrix are glued together by extracellular matrix proteins to form laminated structures (osteons) to provide elasticity and a supportive substrate for osteogenesis. The objective of this work was to investigate material properties and osteogenic differentiation of bone marrow stromal (BMS) cells seeded on osteon-mimetic fiber-reinforced hydrogel/apatite composites. Layers of electrospun poly(l-lactide) fiber mesh coated with a poly(lactide-co-ethylene oxide fumarate) (PLEOF) hydrogel precursor solution were stacked and pressed together, and crosslinked to produce a laminated fiber-reinforced composite. Hydroxyapatite (HA) nanocrystals were added to the precursor solution to produce an osteoconductive matrix for BMS cells. Acrylamide-terminated Arg-Gly-Asp (RGD) peptide (Ac-GRGD) was conjugated to the PLEOF/HA hydrogel phase to promote focal point adhesion of BMS cells. Laminates were characterized with respect to the Young's modulus, degradation kinetics and osteogenic differentiation of BMS cells. The moduli of the laminates under dry and wet conditions were significantly higher than those of the fiber mesh and PLEOF/HA hydrogel, and within the range of values reported for wet human cancellous bone. At days 14 and 21, alkaline phosphatase (ALPase) activity of the laminates was significantly higher than those of the fiber mesh and hydrogel. Lamination significantly increased the extent of mineralization of BMS cells and laminates with HA and conjugated with RGD (Lam-RGD-HA) had 2.7-, 3.5- and 2.8-fold higher calcium content (compared to laminates without HA or RGD) after 7, 14 and 21days, respectively. The Lam-RGD-HA group had significantly higher expression of osteopontin and osteocalcin compared to the hydrogel or laminates without HA or RGD, consistent with the higher ALPase activity and calcium content of Lam-RGD-HA. Laminated osteon-mimetic structures have the potential to provide mechanical strength to the regenerating region as well as supporting the differentiation of progenitor cells to the osteogenic lineage.

[1]  J. Buckwalter,et al.  Bone biology. I: Structure, blood supply, cells, matrix, and mineralization. , 1996, Instructional course lectures.

[2]  S. Boden,et al.  Use of Recombinant Human Bone Morphogenetic Protein-2 to Achieve Posterolateral Lumbar Spine Fusion in Humans: A Prospective, Randomized Clinical Pilot Trial 2002 Volvo Award in Clinical Studies , 2002, Spine.

[3]  M. Kotaki,et al.  Electrospun P(LLA-CL) nanofiber: a biomimetic extracellular matrix for smooth muscle cell and endothelial cell proliferation. , 2004, Biomaterials.

[4]  Martijn van Dijk,et al.  The Effect of Cage Stiffness on the Rate of Lumbar Interbody Fusion: An In Vivo Model Using Poly(L-Lactic Acid) and Titanium Cages , 2002, Spine.

[5]  E. Kaiser,et al.  Color test for detection of free terminal amino groups in the solid-phase synthesis of peptides. , 1970, Analytical biochemistry.

[6]  K. Kusumoto,et al.  Bone tissue engineering using human adipose-derived stem cells and honeycomb collagen scaffold. , 2008, Journal of biomedical materials research. Part A.

[7]  Henrik Birkedal,et al.  Influence of the degradation of the organic matrix on the microscopic fracture behavior of trabecular bone. , 2004, Bone.

[8]  D. Beruto,et al.  Role of the water matric potential (psi(M)) and of equilibrium water content (EWC) on the water self-diffusion coefficient and on the oxygen permeability in hydrogel contact lenses. , 2004, Biomaterials.

[9]  F. Wehrli,et al.  Water Content Measured by Proton‐Deuteron Exchange NMR Predicts Bone Mineral Density and Mechanical Properties , 2003, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[10]  Sheng Lin-Gibson,et al.  Systematic investigation of porogen size and content on scaffold morphometric parameters and properties. , 2007, Biomacromolecules.

[11]  E. Jabbari,et al.  Modeling and experimental investigation of rheological properties of injectable poly(lactide ethylene oxide fumarate)/hydroxyapatite nanocomposites. , 2006, Biomacromolecules.

[12]  P. Ma,et al.  Engineering new bone tissue in vitro on highly porous poly(alpha-hydroxyl acids)/hydroxyapatite composite scaffolds. , 2001, Journal of biomedical materials research.

[13]  Masashi Neo,et al.  In vivo evaluation of a porous hydroxyapatite/poly-DL-lactide composite for bone tissue engineering. , 2007, Journal of biomedical materials research. Part A.

[14]  David L Kaplan,et al.  Stem cell- and scaffold-based tissue engineering approaches to osteochondral regenerative medicine. , 2009, Seminars in cell & developmental biology.

[15]  S. K. Agrawal,et al.  Nanoparticle-reinforced associative network hydrogels. , 2008, Langmuir : the ACS journal of surfaces and colloids.

[16]  D. Hak.  The Use of Osteoconductive Bone Graft Substitutes in Orthopaedic Trauma , 2007, The Journal of the American Academy of Orthopaedic Surgeons.

[17]  S. Sen,et al.  Matrix Elasticity Directs Stem Cell Lineage Specification , 2006, Cell.

[18]  K. V. Van Vliet,et al.  Tuning compliance of nanoscale polyelectrolyte multilayers to modulate cell adhesion. , 2005, Biomaterials.

[19]  E. Jabbari,et al.  Solid-phase synthesis of reactive peptide crosslinker by selective deprotection. , 2006, Protein and peptide letters.

[20]  A J Verbout,et al.  Design and fabrication of standardized hydroxyapatite scaffolds with a defined macro-architecture by rapid prototyping for bone-tissue-engineering research. , 2004, Journal of biomedical materials research. Part A.

[21]  A. Thor,et al.  Reconstruction of the severely resorbed maxilla with autogenous bone, platelet-rich plasma, and implants: 1-year results of a controlled prospective 5-year study. , 2005, Clinical implant dentistry and related research.

[22]  J A Planell,et al.  Fabrication of low temperature macroporous hydroxyapatite scaffolds by foaming and hydrolysis of an alpha-TCP paste. , 2004, Biomaterials.

[23]  E. Jabbari,et al.  Synthesis and characterization of bioresorbable in situ crosslinkable ultra low molecular weight poly(lactide) macromer , 2008, Journal of materials science. Materials in medicine.

[24]  A. Greiner,et al.  Characterization of a PLLA-Collagen I Blend Nanofiber Scaffold with Respect to Growth and Osteogenic Differentiation of Human Mesenchymal Stem Cells , 2009, TheScientificWorldJournal.

[25]  C. M. Agrawal,et al.  Fundamentals of biomechanics in tissue engineering of bone. , 2000, Tissue engineering.

[26]  G. Daculsi,et al.  Short-term effects of mineral particle sizes on cellular degradation activity after implantation of injectable calcium phosphate biomaterials and the consequences for bone substitution. , 1999, Bone.

[27]  S. Parikh.  Bone graft substitutes: past, present, future. , 2002, Journal of postgraduate medicine.

[28]  Tae Gwan Park,et al.  Biomimicking extracellular matrix: cell adhesive RGD peptide modified electrospun poly(D,L-lactic-co-glycolic acid) nanofiber mesh. , 2006, Tissue engineering.

[29]  M. Pfaffl,et al.  A new mathematical model for relative quantification in real-time RT-PCR. , 2001, Nucleic acids research.

[30]  K. Koval,et al.  Bone grafts and bone graft substitutes in orthopaedic trauma surgery. A critical analysis. , 2007, The Journal of bone and joint surgery. American volume.

[31]  P. Schaaf,et al.  Polyelectrolyte multilayers with a tunable Young's modulus: influence of film stiffness on cell adhesion. , 2006, Langmuir : the ACS journal of surfaces and colloids.

[32]  R Borojevic,et al.  Characterization of a bovine collagen-hydroxyapatite composite scaffold for bone tissue engineering. , 2003, Biomaterials.

[33]  F. Coe,et al.  Disorders of bone and mineral metabolism , 1992 .

[34]  Thomas J Webster,et al.  Arginine-glycine-aspartic acid modified rosette nanotube-hydrogel composites for bone tissue engineering. , 2009, Biomaterials.

[35]  Takashi Nakamura,et al.  Biodegradation behavior of ultra-high-strength hydroxyapatite/poly (L-lactide) composite rods for internal fixation of bone fractures. , 2000, Biomaterials.

[36]  E. Jabbari,et al.  Osteonectin-derived peptide increases the modulus of a bone-mimetic nanocomposite , 2008, European Biophysics Journal.

[37]  E. Jabbari,et al.  Viscoelastic characterization and modeling of gelation kinetics of injectable in situ cross-linkable poly(lactide-co-ethylene oxide-co-fumarate) hydrogels. , 2007, Biomacromolecules.

[38]  Yasuhiko Tabata,et al.  Tissue regeneration based on growth factor release. , 2003, Tissue engineering.

[39]  E. Jabbari,et al.  Gelation and degradation characteristics of in situ photo-crosslinked poly(l-lactide-co-ethylene oxide-co-fumarate) hydrogels , 2007 .

[40]  J. Buckwalter,et al.  Bone structure and function. , 1987, Instructional course lectures.

[41]  Esmaiel Jabbari,et al.  Effect of osteonectin-derived peptide on the viscoelasticity of hydrogel/apatite nanocomposite scaffolds. , 2007, Biopolymers.

[42]  Hideki Yoshikawa,et al.  Bone tissue engineering with porous hydroxyapatite ceramics , 2005, Journal of artificial organs : the official journal of the Japanese Society for Artificial Organs.

[43]  P. Prendergast,et al.  Gene expression by marrow stromal cells in a porous collagen–glycosaminoglycan scaffold is affected by pore size and mechanical stimulation , 2008, Journal of materials science. Materials in medicine.

[44]  Y. Nodasaka,et al.  Acidic amino acid-rich sequences as binding sites of osteonectin to hydroxyapatite crystals. , 1996, Biochimica et biophysica acta.

[45]  Gordana Vunjak-Novakovic,et al.  Bioactive hydrogel scaffolds for controllable vascular differentiation of human embryonic stem cells. , 2007, Biomaterials.

[46]  P. Supaphol,et al.  Osteoblastic phenotype expression of MC3T3-E1 cultured on electrospun polycaprolactone fiber mats filled with hydroxyapatite nanoparticles. , 2007, Biomacromolecules.

[47]  Zuzana Dobbie,et al.  Processing of gene expression data generated by quantitative real-time RT-PCR. , 2002, BioTechniques.

[48]  Natalia Juncosa-Melvin,et al.  Effect of scaffold material, construct length and mechanical stimulation on the in vitro stiffness of the engineered tendon construct. , 2008, Journal of biomechanics.

[49]  Byung-Soo Kim,et al.  Poly(lactide-co-glycolide)/hydroxyapatite composite scaffolds for bone tissue engineering. , 2006, Biomaterials.

[50]  M J Glimcher,et al.  Bone biology. II: Formation, form, modeling, remodeling, and regulation of cell function. , 1996, Instructional course lectures.

[51]  Fabrizio Gelain,et al.  Biological Designer Self-Assembling Peptide Nanofiber Scaffolds Significantly Enhance Osteoblast Proliferation, Differentiation and 3-D Migration , 2007, PloS one.

[52]  黄亚明.  MedScape , 2009 .

[53]  S. Weiner,et al.  Lamellar bone: structure-function relations. , 1999, Journal of structural biology.

[54]  E. Jabbari,et al.  Effect of a low-molecular-weight cross-linkable macromer on electrospinning of poly(lactide-co-glycolide) fibers , 2007, Journal of biomaterials science. Polymer edition.

[55]  Yen Chang,et al.  Bioengineered cardiac patch constructed from multilayered mesenchymal stem cells for myocardial repair. , 2008, Biomaterials.

[56]  M. Mochizuki,et al.  Long-term stability of bone tissues induced by an osteoinductive biomaterial, recombinant human bone morphogenetic protein-2 and a biodegradable carrier. , 2004, Biomaterials.

[57]  So-Jung Gwak,et al.  Orthotopic bone formation by implantation of apatite-coated poly(lactide-co-glycolide)/hydroxyapatite composite particulates and bone morphogenetic protein-2. , 2008, Journal of biomedical materials research. Part A.

[58]  E. Jabbari,et al.  Material properties and bone marrow stromal cells response to in situ crosslinkable RGD-functionlized lactide-co-glycolide scaffolds. , 2008, Journal of biomedical materials research. Part A.

[59]  E. Jabbari,et al.  Effect of grafting RGD and BMP-2 protein-derived peptides to a hydrogel substrate on osteogenic differentiation of marrow stromal cells. , 2008, Langmuir : the ACS journal of surfaces and colloids.

[60]  Shengshou Hu,et al.  A novel small‐diameter vascular graft: In vivo behavior of biodegradable three‐layered tubular scaffolds , 2008, Biotechnology and bioengineering.

[61]  E. Jabbari,et al.  A model for the viscoelastic behavior of nanofilled hydrogel composites under oscillatory shear loading , 2008 .

[62]  E. Jabbari,et al.  Effect of Composition on Gelation Kinetics of Unfilled and Nanoapatite-Filled Poly(lactide-ethylene oxide-fumarate) Hydrogels , 2007 .

[63]  G. Falini,et al.  Biologically inspired synthesis of bone-like composite: self-assembled collagen fibers/hydroxyapatite nanocrystals. , 2003, Journal of biomedical materials research. Part A.

[64]  Peng Zhao,et al.  Poly (d,l-lactide)/nano-hydroxyapatite composite scaffolds for bone tissue engineering and biocompatibility evaluation , 2008, Journal of materials science. Materials in medicine.

[65]  Hisatoshi Kobayashi,et al.  Osteogenic differentiation of mesenchymal stem cells in self-assembled peptide-amphiphile nanofibers. , 2006, Biomaterials.

[66]  K. Jacob,et al.  Characterization of Polymer Nanocomposite Interphase and Its Impact on Mechanical Properties , 2006 .

[67]  E. Jabbari,et al.  Material properties and cytocompatibility of injectable MMP degradable poly(lactide ethylene oxide fumarate) hydrogel as a carrier for marrow stromal cells. , 2007, Biomacromolecules.

[68]  Jennifer L West,et al.  Poly(ethylene glycol) hydrogel system supports preadipocyte viability, adhesion, and proliferation. , 2005, Tissue engineering.

[69]  V. Hasırcı,et al.  Bone tissue engineering on patterned collagen films: an in vitro study. , 2005, Biomaterials.