Bone Tissue Engineering
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1 Bone Tissue Engineering Miqin Zhang University of Washington Dept of Materials Science and Engineering Funded by UWEB: A National Science Foundation Engineering Research Center (NSF-EEC ) Bone Defects -- Major Health Concern Accidents, sports, tumors, osteoporosis. Osteoporosis ~250,000 hip fractures ( $10 billion) ~70,000 spine fractures ($1 billion) 1
2 Treatment of Bone Defects Material Autograft Allograft Xenograft Tissue substitutes engineering Patient Or patient Animal Biomaterial Promising rapy 1. Patient pain 2. Infection 3. High cost 1. Donor shortage 2. Infection 3. Pathogens 4 Immune Rejection 1. Immune rejection 2. Pathogens Metals polymers bioceramics do not have good properties History of Tissue Engineering 4~ 25 years 4Yannnas et al: artificial skin from collagen and glycosaminoglycan (1980) Cima et al: chondrocytes from PLGA (1991) 4Langer and Vacanti (1993) define tissue engineering L. Bonassar, CA Vacanti, Journal of Cellular Biochemistry Supplements, 30/31: (1998) 2
3 Possible Tissue-Engineered Devices Bone Tissue Engineering Scaffold Cultivated cells Cells on scaffold Scaffold with cells implanted in bone ipatient-derived bone cells onto a macroporous man-made scaffolds to create completely natural new tissues iscaffolds gradually degraded and eventually eliminated 3
4 Criteria for Selecting Scaffold Materials Excellent mechanical strength Cancerous bone (compressive stress MPa) Three dimensional (3D) interconnected macroporous microstructures Controllable biodegradation and bioresorption Suitable surface chemistry Good biocompatibility and biofunctionality Host Response 4
5 Material Response Swelling and leaching Corrosion Particulates Deformation and failure Material/device-Living System Interaction + Biological System Material/device Cytoskeletal connection (1) Protein adsorption (2) Cell Integrin α β RGD RGD RGD Adhesion Protein Cell membrane adhesion Material/device Material/device Material/device 5
6 COOH Effects of Surface Properties on Living Systems Cell Cell Protein OH PEG Surface topography Hydrophilic/ Surface hydrophobic Crystallinity Character Surface Roughness Surface Composition Biointerfaces Bioinert Biofunctional Most implants Tissue engineering Resist non-specific protein adsorption Promote protein adsorption 6
7 Biocompatibility Tests Biomaterial First-level Biosafty Cytotoxicity Hemotoxicity Genotoxicity Histotoxicity cell morphlogy & viability in vitro hemolysis Ames test intramuscular implantations Second-level Biofunctionality Cytocompatibility Immunocompatibility Hemocompatibility Histocompatibility Infectability specialized funtions of primary cells hypersensitivity and immune depression platelet activation, coagulation process, fibrinolysis tissue reaction to implant ability of promoting bacterial adhesion and growth Biocompatibility of Porous Materials for Bone Tissue Engineering Bioinert Resist non-specific protein adsorption Biofunctional Bioactive: form bone between tissues and materials Osteoconduction: stimulate cell attachment and migration Osteoinduction: stimulate proliferation and differentiation of stem cells Osteogenisis: produce bone independently 7
8 Existing Porous Scaffolds for Load-bearing Bone Tissue Engineering Metals Stainless Steel Titanium High mechanical strength Not bioactive and osteoconductive need surface coatings Ceramics HA & β-tcp Composition similar to bone β-tcp > HA in degradation bioactive, osteoconductive ~ 2MPa Coral Can be converted into HA High mechanical strength ~ 14 MPa limited resources, expensive uncontrollable microstructures Hydroxyapatite (HA), tricalcium phosphate (TCP) Polymeric Porous Materials Syntic polymers Polyglycolic acid (PGA) Polylactic acid (PLA) Poly- latic-co-glycolic (PLGA) Biocompatible Biodegradable Bioresobable Hydrophobic Lack cell recognition signals Natural polymer Chitosan More biocompatible Minimal foreign body reaction Hydrophilic Suitable surface chemistry Osteogenic 8
9 Existing Composite Scaffolds Polymer as a matrix Ceramic a second phase Ceramics as matrix Polymer as a second phase Our approach Reinforced by ceramic powder and fibers Low mechanical strength MPa in compressive stress Polymer fills voids of ceramic High mechanical strength5-10mpa 5-10MPa two level controllable degradation easy to incorporate drugs Porous Ceramic Scaffolds for Load-bearing Bone Tissue Engineering Gel-polymer methods: integrating polymer and gel casting methods Ceramic suspension + monomer+ initiator Foaming agent + polymer sponge High mechanical strength Controllable microstructure Uniform structure H. Ramay and M. Zhang, Biomaterials 24 (19),
10 Techniques for Fabricating Porous Ceramics Polymer sponge Ceramic slurry + Polymer sponge Organic burn out and sintering Controllable microstructure Low mechanical strength Ununiform structure Gel-casting Ceramic suspension + monomer+ initiator Foaming agents High mechanical strength Poor microstructure HA Scaffolds by a Gel-Polymer Method C P P 100µm 15KV C 100 µm 15 KV Compressive yield stress: 8 MPa Compressive elastic modulus: 7 GPa 10
11 β-tcp and HA Nanofiber Nanocomposite Scaffolds (a) (b) 100 µm 200 µm 80 nm Compressive stress 9.8 MPa Human cancellous bone 2-10MPa H. Ramay and M. Zhang, Biomaterials, 25 (21), (2004). Mechanical Properties Cancellous bone modulus GPa Human Lumbar bone strength 1.86 MPa Compressive Strength of HA Scaffolds: 5 MPa (Ramay H, Zhang M,Biomaterials,2003) Compressive strength 9.6 MPa Compressive modulus 4.5 GPa Mechanical properties of biphasic ceramic increased with concentration of HA nanofibers 11
12 Toughness of Biphasic Ceramic Toughness (MPa.mm) Concentration of nano HA (wt%) Human Tibia: MPa.mm HA scaffolds: 1.0 MPa.mm (Lorenzo LMR, et al, J Biomed Mater Res, 2002) Toughness of biphasic ceramic increased with concentration of nano HA fibers Mechanisms for Increased Toughness Wavy fracture surface Area of crack surface is increased Clinching at crack tip Clinching reduces applied stress intensity factor 10 µm 12
13 Chitosan-based Scaffolds for Bone Tissue Engineering and Drug Carrier Chitosan: main organic component of shellfish like shrimp, crabs and squid. Advantages of polymers Biodegradable and biocompatible Cell signaling and immune recognition Control over biodegradation and bioactivity Form hydrogel Hydrophilic Molecular structure of chitosan. Fabrication of Porous Polymer Scaffolds Solid liquid phase separation Controllable pore size and shape ph % Chitosan-alginate solution at ph 7.4 Polymer solution Phase separation Porous polymer scaffold 13
14 Microstructure of Chitosan-based Scaffolds B Chitosan composite Chitosan Mechanical and Swelling Properties Yield Strength (Mpa) Chitosan Composite Chitosan Composite Young's Modules (MPa) 3~4 times increase in mechanical strength and compressive modulus C Chitosan composite Chitosan Chitosan composite scaffolds Do not swell in PBS solution Stable in neutral solution Incorporation of proteins without denaturization 14
15 In Vitro Studies of Chitosan Composite Scaffolds 3 day osteoblast on chitosan composite scaffolds 3 day osteoblast on pure chitosan scaffold In Vitro Studies of Chitosan Composite Scaffolds Chitosan Chitosan -composite 3 days 7 days 15
16 Porous and Non-porous Scaffold Non-porous Non-porous chitosan chitosan composites composites (Air (Air dried) dried) Porous Porous chitosan chitosan composite composite (Freeze-dried) (Freeze-dried) Harvest Bone Marrow Suck Suck bone bone marrow marrow with with a pediatric pediatric spinal spinal need need from from lower lower end end of of femur femur 16
17 Ectopic Inplant Scaffolds Remove Remove hair hair of of thigh,cut thigh,cut skin skin with with carpel carpel and and exposure exposure muscle. muscle. Make Make a a pouch pouch with with a a cut cut to to muscle. muscle. Embed Embed scaffold scaffold toger toger with with bone bone marrow marrow in in muscle muscle pouch pouch Implant Scaffolds 17
18 Ceramics Nesting Chitosan Sponges Large pores of HA/β-TCP Small pores of chitosan Hard bones High mechanical High surface/volume ratio High toughness HA/β-TCP Ceramics Nesting Chitosan Sponges In simulated body solution 5 days In cell culture 5 days 18
19 Cellular Compatibilty A. HA surface and B. Chitosan of HA nesting chitosan C. Chitosan/10%HA; D. Chitosan/5%HA/5% glass Ceramics Nesting Chitosan Sponges 6 5 Zhang Y and Zhang M, Journal of Biomedical Materials Research 61 (1), 1-8 (2002). Yield Stress (MPa) a b c a. Chitosan; b. HA; c. HA/chitosan 19
20 Acknowlodgements > Research team on scaffolds Hassna Ramay, Zhensheng Li, Narayan Bhattarai, Conroy Sun, Nathan Kohler > UWEB: A National Science Foundation Engineering Research Center (NSF-EEC ) > NIH BRP > Facilities at Center for Nanotechnology > Cell culture facilities at UWEB 20
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