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Biological Effects of Functionalizing Copolymer Scaffolds with Nanodiamond Particles
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2013 (English)In: Tissue Engineering. Part A, ISSN 1937-3341, E-ISSN 1937-335X, Vol. 19, no 15-16, 1783-1791 p.Article in journal (Refereed) Published
Abstract [en]

Significant evidence has indicated that poly(L-lactide)-co-(epsilon-caprolactone) [(poly(LLA-co-CL)] scaffolds could be one of the suitable candidates for bone tissue engineering. Oxygen-terminated nanodiamond particles (n-DP) were combined with poly(LLA-co-CL) and revealed to be positive for cell growth. In this study, we evaluated the influence of poly(LLA-co-CL) scaffolds modified by n-DP on attachment, proliferation, differentiation of bone marrow stromal cells (BMSCs) in vitro, and on bone formation using a sheep calvarial defect model. BMSCs were seeded on either poly(LLA-co-CL)-or n-DP-coated scaffolds and incubated for 1 h. Scanning electron microscopy (SEM) and fluorescence microscopy were used in addition to protein and DNA measurements to evaluate cellular attachment on the scaffolds. To determine the effect of n-DP on proliferation of BMSCs, cell/scaffold constructs were harvested after 3 days and evaluated by Bicinchoninic Acid (BCA) protein assay and SEM. In addition, the osteogenic differentiation of cells grown for 2 weeks on the various scaffolds and in a dynamic culture condition was evaluated by real-time RT-PCR. Unmodified and modified scaffolds were implanted into the calvaria of six-year-old sheep. The expression of collagen type I (COL I) and bone morphogenetic protein-2 (BMP-2) after 4 weeks as well as the formation of new bone after 12 and 24 weeks were analyzed by immunohistochemistry and histology. Scaffolds modified with n-DP supported increased cell attachment and the mRNA expression of osteopontin (OPN), bone sialoprotein (BSP), and BMP-2 were significantly increased after 2 weeks of culture. The BMSCs had spread well on the various scaffolds investigated after 3 days in the study with no significant difference in cell proliferation. Furthermore, the in vivo data revealed more positive staining of COLI and BMP-2 in relation to the n-DP-coated scaffolds after 4 weeks and presented more bone formation after 12 and 24 weeks. n-DP modification significantly increased cell attachment and differentiation of BMSCs on poly(LLA-co-CL) scaffolds in vitro and enhanced bone formation in vivo.

Place, publisher, year, edition, pages
Mary Ann Liebert, 2013. Vol. 19, no 15-16, 1783-1791 p.
Keyword [en]
Bone marrow stromal cells (BMSCs), Bone morphogenetic protein-2, Bone tissue engineering, Cellular attachments, Immunohistochemistry, Nano-diamond particles, Osteogenic differentiation, Significant differences
National Category
Biological Sciences
URN: urn:nbn:se:kth:diva-125552DOI: 10.1089/ten.tea.2012.0336ISI: 000321039800013ScopusID: 2-s2.0-84879622713OAI: diva2:640066
EU, FP7, Seventh Framework Programme, 242175

QC 20130812

Available from: 2013-08-12 Created: 2013-08-09 Last updated: 2014-09-29Bibliographically approved
In thesis
1. Engineering and Functionalization of Degradable Scaffolds for Medical Implant Applications
Open this publication in new window or tab >>Engineering and Functionalization of Degradable Scaffolds for Medical Implant Applications
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The treatment of bone defects is facing the situation of lacking donations for autotransplantation. As a valid approach, scaffold-based tissue engineering combines the construction of well-defined porous scaffolds with advanced cell culturing technology to guide tissue regeneration. The role for the scaffold is to provide a suitable environment with a sufficient mechanical stiffness, supports for cell attachment, migration, nutrients and metabolite transport and space for cell remodeling and tissue regeneration. The random copolymers poly(L-lactide-co-ɛ-caprolactone) (poly(LLA-co-CL)) and poly(L-lactide-co-1,5-dioxepan-2-one) (poly(LLA-co-DXO)) have been successfully incorporated into 3D porous scaffolds to induce specific interactions with cells and direct osteogenic cell differentiation. In this thesis, these scaffolds have been modified in chemical and physical ways to map and understand requirements for bone regeneration. Scaffold functionalities and properties, such as hydrophilicity, stiffness, size/shape, and reproducibility, were studied. The hydrophilicity was varied by adding 3–20 % (w/w) Tween 80 to poly(LLA-co-CL) and poly(LLA-co-DXO) respectively, which resulted in contact angles from 35° to 15°. With 3 % Tween 80, the resultant mechanical and thermal properties were similar to pristine polymer materials. Tween 80 did not significantly influence cell attachment or proliferation but did stimulate the mRNA expression of osteogenetic markers. The surface functionality and mechanical properties were altered by introducing nanodiamond particles (n-DP) into poly(LLA-co-CL) scaffolds by means of surface physisorption or hybrid blending. Scaffold with n-DP physisorbed showed improved cell attachment, differentiation, and bone reformation. Hybrid n-DP/poly(LLA-co-CL) composites were obtained by direct blending of polylactide modified n-DP (n-DP-PLA) with poly(LLA-coCL). The n-DP-PLA was prepared by sodium hydride-mediated anionic polymerization using n-DP as the initiator. Prepared n-DP-PLA could be dispersed homogenously in organic solvents and blended with poly(LLA-coCL) solution. The n-DP-PLA particles were homogenously distributed in the composite material, which significantly improved mechanical properties. For comparison, the addition of benzoquinone-modified n-DP (n-DP-BQ) did not reinforce poly(LLA-co-CL). This indicated the importance of specific surface grafting, which determined different particle-polymer interactions. For the treatment of critical size defects, a large porous poly(LLA-co-CL) scaffold (12.5 mm diameter × 25 mm thickness) was developed and produced by molding and salt-leaching methods. The large porous scaffolds were evaluated in a scaffold-customized perfusion-based bioreactor system. It was obvious that the scaffold could support improved cell distribution and support the stimulation of human mesenchymal stem cell (hMSC) especially with dynamic flow in a bioreactor. To improve the scaffolding technique, a three-dimensional fiber deposition (3DF) technique was employed to build layer-based scaffolds. Poly(LLA-coCL) scaffolds produced by the 3DF method showed enhanced mechanical properties and a homogeneous distribution of human osteoblasts (hOBs) in the scaffolds. Although poly(LLA-co-CL) was thermally degraded, the degradation did not influence the scaffold mechanical properties. Based on the computerized design, a 3DF scaffold of amorphous copolymer poly(LLAco-CL) provides high-precision control and reproducibility. In summary, the design of porous scaffolds is one of the essential factors in tissue engineering as to mimicking the intrinsic extracellular environment. For bone tissue engineering, an optimized scaffold can maintain a contact angle greater than 35 degrees. Pristine or modified n-DP, introduced as an additive by surface physisorption or direct blending, can improve scaffold mechanical properties and cell response. Various sizes of scaffolds can be easily produced by a mold-mediated salt-leaching method. However, when 100 % reproducibility is required, the 3DF method can be used to create customizable scaffolds.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2014. 73 p.
TRITA-CHE-Report, ISSN 1654-1081 ; 2014:36
Tissue engineering, nanodiamond, scaffold, bioreactor
National Category
Polymer Technologies
Research subject
Fibre and Polymer Science
urn:nbn:se:kth:diva-152605 (URN)978-91-7595-256-7 (ISBN)
Public defence
2014-10-17, Kollegiesalen, Brinellvägen 8, KTH, Stockholm, 13:44 (English)
EU, FP7, Seventh Framework Programme, Vascubone

QC 20140929

Available from: 2014-09-29 Created: 2014-09-29 Last updated: 2014-09-29Bibliographically approved

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