Immobilization of alkaline phosphatase on microporous nanofibrous fibrin scaffolds for bone tissue engineering |
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| Authors: | Thanaphum Osathanon Cecilia M. Giachelli Martha J. Somerman |
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| Affiliation: | 1. Department of Oral Biology, School of Dentistry, University of Washington, Seattle, WA 98195, USA;2. Department of Bioengineering, College of Engineering, University of Washington, Seattle, WA 98195, USA;3. Department of Periodontics, School of Dentistry, University of Washington, Box 357444, B-127 Health Sciences Building, Seattle, WA 98195, USA;1. Department of Chemical Engineering, Virginia Tech, Blacksburg, VA 24061, USA;2. Department of Materials Science and Engineering, Virginia Tech, Blacksburg, VA 24061, USA;3. School of Biomedical Engineering and Sciences, Virginia Tech, Blacksburg, VA 24061, USA;1. Departamento de Química, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, 14040-901, Ribeirão Preto, São Paulo, Brazil;2. Sanford Burnham Prebys Medical Discovery Institute, La Jolla, CA, 92037, USA;1. Department of Biomaterials, Faculty of Oral and Dental Medicine, Cairo University, 11562 Cairo, Egypt;2. Institute of Polymer Materials, University of Erlangen-Nuremberg, 91058 Erlangen, Germany;3. Department of Hand, Plastic and Reconstructive Surgery – Burn Center – BG Trauma Center Ludwigshafen, Plastic and Hand Surgery, University of Heidelberg, Ludwigshafen, Germany;4. Institute of Biomaterials, University of Erlangen-Nuremberg, 91058 Erlangen, Germany |
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| Abstract: | ![]()
Alkaline phosphatase (ALP) promotes bone formation by degrading inorganic pyrophosphate (PPi), an inhibitor of hydroxyapatite formation, and generating inorganic phosphate (Pi), an inducer of hydroxyapatite formation. Pi is a crucial molecule in differentiation and mineralization of osteoblasts. In this study, a method to immobilize ALP on fibrin scaffolds with tightly controllable pore size and pore interconnection was developed, and the biological properties of these scaffolds were characterized both in vitro and in vivo. Microporous, nanofibrous fibrin scaffolds (FS) were fabricated using a sphere-templating method. ALP was covalently immobilized on the fibrin scaffolds using 1-ethyl-3-(dimethylaminopropyl)carbodiimide hydrochloride (EDC). Scanning electron microscopic observation (SEM) showed that mineral was deposited on immobilized alkaline phosphatase fibrin scaffolds (immobilized ALP/FS) when incubated in medium supplemented with β-glycerophosphate, suggesting that the immobilized ALP was active. Primary calvarial cells attached, spread and formed multiple layers on the surface of the scaffolds. Mineral deposition was also observed when calvarial cells were seeded on immobilized ALP/FS. Furthermore, cells seeded on immobilized ALP/FS exhibited higher osteoblast marker gene expression compared to control FS. Upon implantation in mouse calvarial defects, both the immobilized ALP/FS and FS alone treated group had higher bone volume in the defect compared to the empty defect control. Furthermore, bone formation in the immobilized ALP/FS treated group was statistically significant compared to FS alone group. However, the response was not robust. |
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