| The core of tissue engineering contains seed cells,scaffolds,growth factors.The function of scaffolds is to provide three-dimensional space and metabolic environment for cells growing,and the function of growth factors is to induce and stimulate cells growth and keep cells living and to promote tissue regeneration.But growth factors in medium are difficute to diffuse into scaffolds in vitro culture,so it is very important to realize growth factors's controlled release in scaffolds to success in tissue engineering.To solve the problem of nutrition absence in tissue engineering scaffolds,we incorporated microspheres containing the sustainably released nutrition into chitosan scaffolds,which can be controlled to release and maintain nutrition's concentration stable in scaffolds.First, microspheres containing BSA as a model protein were prepared with double emulsion method, PLGA concentration,stirring speed,external phase concentration,prepared temperature and BSA concentration to microspheres production were tested,and choosed four main effects to do orthogonal experiment to find optimum operation conditions of micropheres production, then mixed with different concentration chitosan solution,freeze-dried.The release profile of BSA from the microspheres and the chitosan scaffolds were monitored using ELIASA by BCA protein assay.The configuration of PLGA microspheres and any change of microspheres and chitosan scaffolds after dispersed in chitosan scaffolds were tested by scanning electron microscope.The results show that the optimized preparation conditions of microspheres are 100mg PLGA,15mg BSA,0.5%external phase concentration,and 400μl internal phase volume. PLGA microspheres were round morphology with 27～55μm in size range.The chitosan scaffolds represented uniform porous,and the porosity of 1%(w/w) chitosan scaffolds was higher than that of 2%'s.The porosity,water content and degradation of 1%chitosan scaffolds were(92.99±2.51)%,(89.66±0.66)%and(73.77±3.21)%,respectively.The tests for in vitro release of microspheres showed sustained release and little bursted.The cumulative release of BSA from PLGA microspheres at 288h were(28.26±0.85)%.PLGA microspheres can be uniformly embedded into scaffolds without any morphology change.The protein concentrations in 1%and 2%chitosan scaffolds at 192h were 13.26×10-2±1.14×10-2mg/ml and 12.43×10-2±3.83×10-2 mg/ml. This paper introduces a new controlled release scaffolds with drug loaded microspheres embedded,which can release protein slowly,keep the protein concentration sustaining stablility inside scaffolds.The experiment provides a base for further research on culturing cells in scaffolds. |
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