电喷雾法制备海藻酸盐-明胶-脂肪干细胞微球及其修复关节软骨损伤的可行性

Preparation of alginate-gelatin-adipose-derived stem cells microspheres by electrospray and feasibility on repairing articular cartilage injury

背景:微球可作为细胞递送系统,通过注射或者与其他支架相结合的方式使细胞定位于软骨损伤处使其再生。目的:采用电喷雾法将脂肪源性干细胞负载至海藻酸盐-明胶微球中,评估微球中的细胞活性、增殖和成软骨分化能力。方法:使用电喷雾技术制备含不同质量浓度明胶(0,5,15,25 g/L)的海藻酸盐微球,光学显微镜下观察微球的外观,选择形成完整微球的明胶质量浓度用于以下实验。使用电喷雾技术制备含脂肪源性干细胞的海藻酸盐微球与海藻酸盐微球-明胶微球,光学显微镜下观察成球效果。将含细胞的两种微球培养于旋转细胞培养系统内,在设定的时间点检测微球内细胞的增殖能力、活性与成软骨能力。结果与结论:①加入0,5 g/L的明胶可制备出完整的微球,微球直径分别为(365.85±16.88),(358.85±23.97)μm,两种微球直径比较差异无显著性意义(P﹥0.05),后续实验选择明胶质量浓度为5 g/L;②加入脂肪源性干细胞不影响藻酸盐微球与海藻酸盐微球-明胶微球的成球性与微球直径;③含细胞海藻酸盐微球-明胶微球培养7,14 d的细胞增殖数量、活细胞比例均高于含细胞海藻酸盐微球(P<0.05);④成软骨诱导培养后,含细胞海藻酸盐微球-明胶微球诱导7,14,21 d的糖胺多糖含量高于含细胞海藻酸盐微球(P<0.05);⑤结果表明,海藻酸盐-明胶微球更易促进脂肪源性干细胞的增殖并保持细胞活性,是软骨组织工程领域中一种具有巨大潜力的生物材料。

BACKGROUND:Microsphere can be used as a cell delivery system to localize cells in cartilage injury site and regenerate them by injection or combined with other scaffolds.OBJECTIVE:To produce adipose-derived stem cells(ADSCs)-embedded alginate-gelatin microspheres for cartilage tissue engineering by electrospray,and to evaluate cell viability,proliferation and chondrogenic differentiation ability of microspheres.METHODS:The electrospray technology was used to produce alginate-gelatin microspheres with different gelatin concentrations(0,5,15,25 g/L).The morphology of microspheres could be observed under the optical microscope.The mass concentration of gelatin to form complete microspheres was selected for the following experiments.Alginate-gelatin-ADSCs microspheres and alginate-ADSCs microspheres were produced by electrospray.The spherical effect was observed under the optical microscope.Two kinds of microspheres containing cells were cultured in rotating cell culture system.The proliferation ability,viability and chondrogenic ability of the cells in the microspheres were detected at the set time point.RESULTS AND CONCLUSION:(1)Intact microspheres could be produced by adding 0,5 g/L gelatin.The diameter of the microspheres was(365.85±16.88)or(358.85±23.97)μm.There was no significant difference in the diameter of the two kinds of microspheres(P>0.05).The mass concentration of gelatin as 5 g/L was selected for subsequent experiments.(2)The addition of ADSCs did not affect the spheroidization and the diameter of alginate microspheres and alginate microspheres-gelatin microspheres.(3)The number of cell proliferation and the proportion of living cells at 7 and 14 days were higher in the alginate-gelatin-ADSCs microspheres than those without alginate-ADSCs microspheres(P<0.05).(4)The content of glycosaminoglycan was higher in alginate-gelatin-ADSCs microspheres than that in alginate-ADSCs microspheres at 7,14,and 21 days after chondrogenic differentiation(P<0.05).(5)It is concluded that alginategelatin-ADSCs microspheres are prone to promote proliferation and keep cell viability,and are a kind of biomaterials with great potential in the field of cartilage tissue engineering.