Preparation and surface characterization of poly-L-lysine-coated PLGA microsphere scaffolds containing retinoic acid for nerve tissue engineering: in vitro study.

The pluripotent nature and proliferative capacity of embryonic stem cells make them an attractive cell source for tissue engineering. In this study, the poly-L-lysine-coated PLGA microspheres which contain retinoic acid (RA) as an inducer factor were prepared by using a water-in-oil-in-water emulsion/solvent evaporation technique. Then, pluripotent P19 embryonic carcinoma cells were seeded on them for differentiating into neural cells. Size and surface morphology of PLGA microspheres were evaluated by scanning electron microscope (SEM). For in vitro examinations, SEM, MTT assay, immunofluorescent staining, histology and reverse transcriptase-polymerase chain reaction (RT-PCR) analyses were carried out. SEM micrographs of the scaffolds showed a diameter in range of 13-100 microm. Based on the release profiles obtained, the concentration of RA released from microspheres reached 10(-6) to 10(-7) mg/ml. MTT assay showed that the number of cells attached on coated microspheres were more in comparison with uncoated microspheres. Immunoflourescent staining and RT-PCR analyses for MapII, beta-tubulin III, Nestin and Pax6 indicated differentiation of P19 cells into neural cells on all of the samples. Finally, the counting of positive cells showed 80+/-8.8% and 72+/-6.9% of the cells expressed beta-tubulin III on the surface of coated and uncoated RA-loaded PLGA microspheres, respectively, while the 64+/-1.1% (P < 0.05) cells expressed tubulin III in group with soluble.