title: Osseointegrative properties of electrospun hydroxyapatite-containing nanofibrous chitosan scaffolds.
abstract: Our long-term goal is to develop smart biomaterials that can facilitate regeneration of critical-size craniofacial lesions. In this study, we tested the hypothesis that biomimetic scaffolds electrospun from chitosan (CTS) will promote tissue repair and regeneration in a critical size calvarial defect. To test this hypothesis, we first compared in vitro ability of electrospun CTS scaffolds crosslinked with genipin (CTS-GP) to those of mineralized CTS-GP scaffolds containing hydroxyapatite (CTS-HA-GP), by assessing proliferation/metabolic activity and alkaline phosphatase (ALP) levels of murine mesenchymal stem cells (mMSCs). The cells' metabolic activity exhibited a biphasic behavior, indicative of initial proliferation followed by subsequent differentiation for all scaffolds. ALP activity of mMSCs, a surrogate measure of osteogenic differentiation, increased over time in culture. After 3 weeks in maintenance medium, ALP activity of mMSCs seeded onto CTS-HA-GP scaffolds was approximately two times higher than that of cells cultured on CTS-GP scaffolds. The mineralized CTS-HA-GP scaffolds were also osseointegrative in vivo, as inferred from the enhanced bone regeneration in a murine model of critical size calvarial defects. Tissue regeneration was evaluated over a 3 month period by microCT and histology (Hematoxylin and Eosin and Masson's Trichrome). Treatment of the lesions with CTS-HA-GP scaffolds induced a 38% increase in the area of de novo generated mineralized tissue area after 3 months, whereas CTS-GP scaffolds only led to a 10% increase. Preseeding with mMSCs significantly enhanced the regenerative capacity of CTS-GP scaffolds (by ∼3-fold), to 35% increase in mineralized tissue area after 3 months. CTS-HA-GP scaffolds preseeded with mMSCs yielded 45% new mineralized tissue formation in the defects. We conclude that the presence of HA in the CTS-GP scaffolds significantly enhances their osseointegrative capacity and that mineralized chitosan-based scaffolds crosslinked with genipin may represent a unique biomaterial with possible clinical relevance for the repair of critical calvarial bone defects.
authors.affiliation: 1 School of Biomedical Engineering, Science and Health Systems, Drexel University , Philadelphia, Pennsylvania.
authors.forename: Michael E
authors.initials: ME
authors.lastname: Frohbergh
authors.forename: Anya
authors.initials: A
authors.lastname: Katsman
authors.forename: Mark J
authors.initials: MJ
authors.lastname: Mondrinos
authors.forename: Collin T
authors.initials: CT
authors.lastname: Stabler
authors.forename: Kurt D
authors.initials: KD
authors.lastname: Hankenson
authors.forename: Jeffrey T
authors.initials: JT
authors.lastname: Oristaglio
authors.forename: Peter I
authors.initials: PI
authors.lastname: Lelkes
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D001861:Bone Regeneration / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D048271:Chitosan / Q000494:pharmacology*; D017886:Durapatite / Q000494:pharmacology*; D005260:Female; D005453:Fluorescence; D039823:Iridoids / Q000494:pharmacology; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D051379:Mice; D057139:Nanofibers / Q000737:chemistry*; D016348:Osseointegration / Q000187:drug effects*; D011856:Radiographic Image Enhancement; D012886:Skull / Q000000981:diagnostic imaging / Q000187:drug effects; D023822:Tissue Engineering / Q000379:methods*; D054457:Tissue Scaffolds / Q000737:chemistry*; D014945:Wound Healing / Q000187:drug effects; D055114:X-Ray Microtomography
chemical_list: D039823:Iridoids; D048271:Chitosan; D017886:Durapatite; C007834:genipin; D000469:Alkaline Phosphatase