Fabrication of crosslinked carboxymethylchitosan microspheres and their incorporation into composite scaffolds for enhanced bone regeneration

Authors

Benjamin T. Reves, Joel D. Bumgardner, Warren O. Haggard

Abstract

Carboxymethylchitosan (CMCS) microspheres were prepared by the carboxymethylation of chitosan (CS) beads using monochloroacetic acid. The CMCS microspheres were crosslinked using two different methods: the amine-amine crosslinker genipin and carbodiimide chemistry, yielding Gen-X CMCS and X-CMCS beads, respectively. The Gen-X CMCS beads were found to have poor degradation and drug release profiles. The X-CMCS microspheres displayed good potential for use in tissue engineering applications in which degradation and local drug delivery are desired. The X-CMCS beads displayed enzymatic degradation of 82.7 ± 1.2% in 100 μg/mL lysozyme after 1 month. An extended release of rhBMP-2 for at least 45 days was also observed with the X-CMCS microspheres. Scaffolds were formed by fusing beads together, and the X-CMCS beads were successfully incorporated into composite X-CMCS/CS scaffolds. The composite scaffolds had increased degradation of 14.5 ± 6.6% compared to 0.5 ± 0.4% for CS-only scaffolds, and the X-CMCS/CS scaffolds released more rhBMP-2 at all timepoints. The composite scaffolds also supported the attachment and proliferation of SAOS-2 cells. The addition of X-CMCS beads resulted in fabrication of scaffolds with improved properties for use in bone tissue engineering.

Link to Article

http://dx.doi.org/10.1002/jbm.b.32865

Nano- to Macroscale Remodeling of Functional Tissue-Engineered Bone

Authors

Maria Ann Woodruff, Claudia Lange, Fulin Chen, Peter Fratzl, Dietmar Werner Hutmacher

Abstract

A higher degree of mineralization is found within scaffold groups implanted with cells compared to scaffold alone, demonstrating greater bone regenerative potential of cell-scaffold constructs Tissue engineered bone analysed using ESEM and SAXS demonstrates bone formation within the scaffold to be preferentially aligned around the scaffold struts. The mineral particles are not shown to orientate around the osteons within the native bone.

Link to Article

http://dx.doi.org/10.1002/adhm.201200289

Nano-crystalline diamond-coated titanium dental implants – A histomorphometric study in adult domestic pigs

Authors

Philipp Metzler, Cornelius von Wilmowsky, Bernd Stadlinger, Wolfgang Zemann, Karl Andreas Schlegel, Stephan Rosiwal, Stephan Rupprecht

Abstract

Promising biomaterial characteristics of diamond-coatings in biomedicine have been described in the literature. However, there is a lack of knowledge about implant osseointegration of this surface modification compared to the currently used sandblasted acid-etched Ti-Al6-V4 implants. The aim of this study was to investigate the osseointegration of microwave plasma-chemical-vapour deposition (MWP-CVD) diamond-coated Ti-Al6-V4 dental implants after healing periods of 2 and 5 months. Twenty-four MWP-CVD diamond-coated and 24 un-coated dental titanium-alloy implants (Ankylos®) were placed in the frontal skull of eight adult domestic pigs. To evaluate the effects of the nano-structured surfaces on bone formation, a histomorphometric analysis was performed after 2 and 5 months of implant healing. Histomorphometry analysed the bone-to-implant contact (BIC). No significant difference in BIC for the diamond-coated implants in comparison to reference implants could be observed for both healing periods. Scanning electron microscopy revealed an adequate interface between the bone and the diamond surface. No delamination or particle-dissociation due to shearing forces could be detected. In this study, diamond-coated dental titanium-alloy implants and sandblasted acid-etched implants showed a comparable degree of osseointegration.

Link to Article

http://dx.doi.org/10.1016/j.jcms.2012.11.020

Orthodontic mini-implant diameter does not affect in-situ linear microcrack generation in the mandible or the maxilla

Authors

Sean Shih-Yao Liu, Enrique Cruz-Marroquin, Jun Sun, Kelton T. Stewart, Matthew R. Allen

Abstract

Microdamage reduces bone mechanical properties and thus could contribute to implant failure. The objective of this study was to investigate whether the diameter of mini-implants affects linear microcrack generation and whether this differs between the mandible and the maxilla because of their contrasting cortical thicknesses. Maxillary and mandibular quadrants of 5 dogs were randomly assigned to receive, in situ, no pilot drilling or mini-implant insertion (control), pilot drilling only without mini-implants, or pilot drilling plus a mini-implant of 1 of 3 diameters: 1.4 mm (n = 18), 1.6 mm (n = 18), and 2.0 mm (n = 18). Linear microcracks were assessed on basic fuchsin-stained sections by using epifluorescence microscopy. Pilot drilling without mini-implant insertion produced significantly higher linear microcrack burdens in the mandible compared with the maxilla. In the both the mandible and the maxilla, all implants produced higher linear microcrack burdens than did the controls, yet there were no differences between the 3 implant diameters. Neither the diameter of the mini-implant nor the site of insertion (mandible vs maxilla) had a significant effect on the amount of linear microdamage adjacent to the implant when the implants were inserted after pilot drilling in situ.

Link to Article

http://dx.doi.org/10.1016/j.ajodo.2012.07.014

Balancing mechanical strength with bioactivity in chitosan–calcium phosphate 3D microsphere scaffolds for bone tissue engineering: air- vs. freeze-drying processes

Authors

D.T. Nguyen, J.D. McCanless, M.M. Mecwan, A.P. Noblett, W.O. Haggard, R.A. Smith & J.D. Bumgardner

Abstract

The objective of this study was to evaluate the potential benefit of 3D composite scaffolds composed of chitosan and calcium phosphate for bone tissue engineering. Additionally, incorporation of mechanically weak lyophilized microspheres within those air-dried (AD) was considered for enhanced bioactivity. AD microsphere, alone, and air- and freeze-dried microsphere (FDAD) 3D scaffolds were evaluated in vitro using a 28-day osteogenic culture model with the Saos-2 cell line. Mechanical testing, quantitative microscopy, and lysozyme-driven enzymatic degradation of the scaffolds were also studied. FDAD scaffold showed a higher concentration (p < 0.01) in cells per scaffold mass vs. AD constructs. Collagen was 31% greater (p < 0.01) on FDAD compared to AD scaffolds not evident in microscopy of microsphere surfaces. Alternatively, AD scaffolds demonstrated a superior threefold increase in compressive strength over FDAD (12 vs. 4 MPa) with minimal degradation. Inclusion of FD spheres within the FDAD scaffolds allowed increased cellular activity through improved seeding, proliferation, and extracellular matrix production (as collagen), although mechanical strength was sacrificed through introduction of the less stiff, porous FD spheres.

Link to Article

http://dx.doi.org/10.1080/09205063.2012.735099

Osteogenesis and angiogenesis induced by porous β-CaSiO3/PDLGA composite scaffold via activation of AMPK/ERK1/2 and PI3K/Akt pathways

Authors

Chen Wang, Kaili Lin, Jiang Chang, Jiao Sun

Abstract

As a potential bioactive material, β-calcium silicate (β-CS) has attracted particular attention in the field of bone regeneration. In this study, porous β-CS/Poly-d,l-Lactide-Glycolide (PDLGA) composite scaffolds were developed with the goals of controlling the degradation rate and improving the mechanical and biological properties. The compressive strength and toughness were significantly enhanced by PDLGA modification of porous β-CS ceramic scaffolds. The effects of the ionic extract from β-CS/PDLGA composite scaffolds on osteogenic differentiation of rat bone marrow-derived mesenchymal stem cells (rBMSCs), proliferation of human umbilical vein endothelial cells (HUVECs) and the related mechanisms were investigated. It was shown that bioactive ions from β-CS/PDLGA scaffolds could enhance cell viability, alkaline phosphatase (ALP) activity, calcium mineral deposition, and mRNA expression levels of osteoblast-related genes of rBMSCs without addition of extra osteogenic reagents. The activation in AMP-activated protein kinase (AMPK), extracellular signal-related kinases (ERK) 1/2 and RUNX-2 were observed in rBMSCs cultured in the extract of β-CS/PDLGA, and these effects could be blocked by AMPK inhibitor Compound C. The extracts of β-CS/PDLGA composites stimulated HUVECs proliferation that was associated with phosphorylation of protein kinase B (Akt) and endothelial nitric oxide synthase (eNOS) as well as an increase in nitric oxide (NO) production and secretion of vascular endothelial growth factor (VEGF). The inductions were abolished by the addition of phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002. The composite scaffolds were implanted in critical sized rabbit femur defects (6 × 10 mm) for 4, 12 and 20 weeks with β-tricalcium phosphate (β-TCP) as controls. Sequential histological evaluations and radiographs revealed that β-CS/PDLGA dramatically stimulated new bone formation and angiogenesis. The biodegradation rate of the β-CS/PDLGA scaffolds was lower than that of β-TCP at each time point examined, and matched the new bone formation rates. These data suggest that β-CS/PDLGA could promote bone regeneration in vivo, which might be ascribed to the enhanced osteogenic differentiation of mesenchymal stem cells (MSCs) and increased angiogenic activity of endothelial cells (ECs).

Link to Article

http://dx.doi.org/10.1016/j.biomaterials.2012.09.021