Tissue engineering employs a variety of different cell friendly platforms capable of supporting and enhancing cell growth and proliferation . Bone and endothelial tissue interaction is closely connected since bone formation depends on vascularization . This work reports on the fabrication of microspheres as biocompatible carriers for the controlled delivery of the growth factors bone morphogenetic protein 2 (BMP-2) and vascular endothelial growth factor (VEGF), responsible for bone and endothelial tissue formation respectively. Polyvinyl alcohol (PVA) and gelatin were blended in different compositions to determine the concentration with the optimal biocompatibility, degradation rate, and protein release using appropriate cell types.
The microspheres were fabricated following the emulsion cross-linking technique. PVA solutions in deionized water were prepared at final concentrations of 5, 7 and 10% w/v. Gelatin (10% w/v) was dissolved in the aqueous PVA solutions under stirring. The mixtures were added dropwise through a syringe to sunflower oil to create a water-in-oil emulsion. 1 ml of 50% v/v glutaraldehyde, and then 30 ml of pure acetone were added. The microspheres were collected through centrifugation at 3500 rpm, washed with acetone and lyophilized. Their morphology was investigated by means of scanning electron microscopy (SEM). Four samples with different % w/v PVA concentrations (0, 5, 7, and 10% w/v) were used to encapsulate bovine serum albumin (BSA) as a model protein. The microspheres were evaluated for % erosion, loading capacity (LC%) and encapsulation efficiency (EE%). The biocompatibility of the different PVA-gelatin microspheres concentrations (0.125, 0.25, 0.5, 1, and 2 mg/ml) was assessed using L929 ﬁbroblasts. Ongoing experiments with BMP-2- and VEGF-loaded microspheres are in progress.
The fabricated microspheres have a diameter ranging from 20-200 μm depending on the concentration of PVA. Microsphere size distribution measurements reveal a positive correlation between PVA concentration and microsphere diameter, which is expected to affect the rate of drug release. The EE% ranged from 80% to 96%, and the % LC from 129% to 155%. Protein release studies depict that the initial burst release occurs during the first 24 h, followed by a steady release rate of protein during a 350 h incubation period. The 0%PVA composition demonstrated the fastest release kinetics with almost 80% release of total encapsulated protein during the first 24 h. 5% and 7% PVA microspheres displayed comparable release kinetics with values of 20% after 24 h and 35% during the first two weeks. An increase of PVA concentration up to 10% led to an even slower protein release profile. The investigated concentrations from 0.5 mg/ml and lower present similar or higher cell viability values compared to the TCPS control.
The PVA-gelatin microspheres are highly biocompatible, demonstrating a low initial burst release and a prolonged cumulative release up to 14 days, thus they function as effective growth factor carriers.
This research was funded by the Hellenic Foundation for Research and Innovation (H.F.R.I.) project number HFRI-FM17-1999.
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