Microwave-induced production of boron-doped HAp (B-HAp) and B-HAp coated composite scaffolds.

The aim of the present study is to produce boron (B) doped hydroxyapatite (B-HAp), which has an osteoinductive property, and investigate in-vitro osteogenesis potential of B-HAp coated chitosan (B-HAp/Ch) scaffolds. At first, B-HAp was produced by the interaction of ions within the concentrated synthetic body fluid containing boron (B-SBF) with microwave energy. Boron incorporation into HAp structure was performed by the substitution of borate ions with phosphate and hydroxyl ions. Experiments were carried out with different microwave powers and exposure times, and optimum conditions for the production of B-HAp were determined. B-HAp precipitated from B-SBF by 600W microwave power has 1.15±0.11% (w/w) B, 1.40 (w/w) Ca/P ratio, 4.30±0.07% (w/w) carbonate content, 30±4nm rod-like morphology and bone-like amorphous structure. Then, chitosan scaffolds that were prepared by freeze-drying were coated with B-HAp by performing microwave-assisted precipitation in the presence of scaffolds to improve their bioactivities and mechanical properties. The formation of apatite layer and the penetration of apatites into the pores were observed by scanning electron microscopy (SEM). Fourier Transform Infrared spectroscopy (ATR-FTIR) and X-ray diffraction (XRD) analysis also confirmed the presence of B-HAp layer. As control, hydroxyapatite coated chitosan scaffolds (HAp/Ch) produced at the same conditions were used. The results of cell culture studies indicated that B releasing from scaffolds enhances proliferation and osteoblastic differentiation of MC3T3-E1 cells. This work emphasized the importance of the use of B within the scaffolds for enhancing in-vitro bone tissue engineering applications.

Microwave-induced biomimetic approach for hydroxyapatite coatings of chitosan scaffolds.

Simulated body fluid (SBF) can form calcium phosphates on osteoinductive materials, so it is widely used for coating of bone scaffolds to mimic natural extracellular matrix (ECM). However, difficulties of bulk coating in 3D scaffolds and the necessity of long process times are the common problems for coating with SBF. In the present study, a microwave-assisted process was developed for rapid and internal coating of chitosan scaffolds. The scaffolds were fabricated as superporous hydrogel (SPH) by combining microwave irradiation and gas foaming methods. Then, they were immersed into 10x SBF-like solution and homogenous bone-like hydroxyapatite (HA) coating was achieved by microwave treatment at 600W without the need of any nucleating agent. Cell culture studies with MC3T3-E1 preosteoblasts showed that microwave-assisted biomimetic HA coating process could be evaluated as an efficient and rapid method to obtain composite scaffolds for bone tissue engineering.

Preparation of bioactive and antimicrobial PLGA membranes by magainin II/EGF functionalization.

Development of dual functional materials that are capable of both reducing bacterial interaction and encouraging host tissue integration has gained importance in design of biomaterials. In this study, we prepared a bilayer poly (lactide co-glycolide) fibrous membrane with antibacterial and bioactive properties by electrospinning. The antibacterial layer was produced by covalent immobilization of antimicrobial peptide, Magainin II. The bioactive layer incorporating epidermal growth factor (EGF) molecules was subsequently electrospun on the antibacterial layer. The membranes were characterized by X-ray photoelectron spectroscopy, scanning electron microscopy and fluorescence microscopy. EGF release was detected by enzyme-linked immunosorbent assay. The antibacterial activity was tested against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus). The ability to support tissue cell integration was detected by using L-929 mouse fibroblasts. The dual functional membranes established enhanced antibacterial properties and increased tissue cell compatibility. This combined approach suggests a promising strategy for wound dressings, vascular grafts and dental membranes as well as catheters and fixation devices.

Combined delivery of PDGF-BB and BMP-6 for enhanced osteoblastic differentiation.

Natural microenvironment during bone tissue regeneration involves integration of multiple biological growth factors which regulate mitogenic activities and differentiation to induce bone repair. Among them platelet derived growth factor (PDGF-BB) and bone morphogenic protein-6 (BMP-6) are known to play a prominent role. The aim of this study was to investigate the benefits of combined delivery of PDGF-BB and BMP-6 on proliferation and osteoblastic differentiation of MC3T3-E1 preosteoblastic cells. PDGF-BB and BMP-6 were loaded in gelatin and poly (3-hydroxybutyric acid-co-3-hydroxyvaleric acid) particles, respectively. The carrier particles were then loaded into 3D chitosan matrix fabricated by freeze drying. The fast release of PDGF-BB during 7 days was accompanied by slower and prolonged release of BMP-6. The premising release of mitogenic factor PDGF-BB resulted in an increased MC3T3-E1 cell population seeded on chitosan scaffolds. Osteogenic markers of RunX2, Col 1, OPN were higher on chitosan scaffolds loaded with growth factors either individually or in combination. However, OCN expression and bone mineral formation were prominent on chitosan scaffolds incorporating PDGF-BB and BMP-6 as a combination.

Sustained release of 17ß-estradiol stimulates osteogenic differentiation of adipose tissue-derived mesenchymal stem cells on chitosan-hydroxyapatite scaffolds.

The aim of this study was to develop a 17ß-estradiol (E2)-releasing scaffold-nanoparticle system in order to promote osteogenic differentiation of rat adipose tissue-derived mesenchymal stem cells (AdMSCs) for bone tissue regeneration. E2-loaded poly(lactide-co-glycolide) (PLGA) nanoparticles with a diameter of ∼240 nm were produced via an emulsion-diffusion-evaporation method. Because of its higher encapsulation efficiency (54%), PLGA, which has a 65:35 composition, was chosen for the preparation of nanoparticles. Chitosan-hydroxyapatite (HA) scaffolds in macroporous structures with interconnected pores were prepared by combining microwave irradiation and gas-foaming techniques. PLGA nanoparticles were loaded onto scaffolds in 2 ways: via embedding after scaffold fabrication and during fabrication. While 100% of the loaded E2 was released during 55 days from scaffolds loaded by embedding, a controlled release behavior of E2 was observed over 135 days in scaffolds loaded during manufacture. The results of cell culture studies indicated that the controlled delivery of E2 from PLGA nanoparticles loaded on chitosan-HA scaffolds had a significant effect on the osteogenic differentiation of AdMSCs.

Superporous polyacrylate/chitosan IPN hydrogels for protein delivery.

In this study, poly(acrylamide), poly(AAm), and poly(acrylamide-co-acrylic acid), poly(AAm-co-AA) superporous hydrogels (SPHs) were synthesized by radical polymerization in the presence of gas blowing agent, sodium bicarbonate. In addition, ionically crosslinked chitosan (CH) superporous hydrogels were synthesized to form interpenetrating superporous hydrogels, i.e. poly(AAm)-CH and poly(AAm-co-AA)-CH SPH-IPNs. The hydrogels have a structure of interconnected pores with pore sizes of approximately 100-150 µm. Although the extent of swelling increased when AA were incorporated to the poly(AAm) structure, the time to reach the equilibrium swelling (~30 s) was not affected so much. In the presence of chitosan network mechanical properties significantly improved when compared with SPHs, however, equilibrium swelling time (~30 min) was prolonged significantly as due to the lower porosities and pore sizes of SPH-IPNs than that of SPHs. Model protein bovine serum albumin (BSA) was loaded into SPHs and SPH-IPNs by solvent sorption in very short time (<1 h) and very high capacities (~30-300 mg BSA/g dry gel) when compared to conventional hydrogels. BSA release profiles from SPHs and SPH-IPNs were characterized by an initial burst of protein during the first 20 min followed by a completed release within 1 h. However, total releasable amount of BSA from SPH-IPNs was lower than that of SPHs as due to the electrostatic interactions between chitosan and BSA.