Page 88 - Development of Functional Scaffolds for Bone Tissue Engineering Using 3D-Bioprinting of Cells and Biomaterials - Yasaman Zamani
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pre-osteoblasts. This should provide solid insights in which type of scaffold is preferred for future bone tissue engineering applications.
MATERIALS AND METHODS
Fabrication of porous scaffolds by solvent casting-porogen leaching
Scaffolds were fabricated using a previously described method [13] with slight modifications. Briefly, sugar crystals (300-500 μm) were used as the porogen. A 10 mm diameter cylindrical glass mold filled with the sugar crystals was placed in a humidified sealed container and incubated at 37°C for 16 h to partly fuse the crystals, after which drying was performed at room temperature for 24 h. PLGA (75/25, PURAC Biomaterials, Netherlands) was dissolved in dimethyl sulfoxide (DMSO, MP Biomedicals, France) to 12.5% (w/w) and thoroughly mixed with β-TCP powder (particle size: 0.5-1 μm, Nik Ceram Razi, Isfahan, Iran) in a 2:1 (w/w) ratio. This PLGA/β-TCP mixture was then added dropwise to the sugar mold allowing diffusion throughout, and transferred to -20°C for 2 h to set the scaffold. The sugar crystals were subsequently leached out of the precipitated PLGA/β-TCP mixture in deionized water at room temperature for 3 days (3 refreshments per day). Scaffolds were UV sterilized for 30 min, and immersed in 70% ethanol for 1 h prior to cell seeding. Scaffolds fabricated by this method are referred to as “porous” scaffolds.
Fabrication of scaffolds by 3D-printing
PLGA granules were melted in a container on a hot plate at 150°C. β-TCP powder was subsequently added to the molten PLGA in a 2:1 ratio, and mixing occurred for 5 min. The PLGA/β-TCP mixture was transferred to the heating tank of a 3D Discovery® bioprinter (RegenHU, Villaz-St-Pierre, Switzerland). All scaffolds were printed using the HM-300H thermos polymer extruder (RegenHU Villaz-St-Pierre, Switzerland) at 125°C and 29 revs/min, equipped with a needle with an inner diameter of 300 μm. The mixture was extruded through the pre-heated needle at 0.4 MPa (4 Bar), and the struts of PLGA/β-TCP were plotted layer-by-layer with alternating 0°/90° lay-down pattern. Cylindrical scaffolds with a diameter of 10 mm and height of 6 mm (total volume 0.471 cm3) were produced. Scaffolds were UV sterilized for 30 min, and immersed in 70% ethanol for 1 h prior to cell seeding.
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