用于ADSCs增殖分化的海藻酸钙/骨粉微胶珠的制备与检测

Preparation and detection of calcium alginate/bone powder microbeads for expansion and differentiation of ADSCs

海藻酸钙微胶珠具有良好的生物相容性、可降解、机械强度高、韧性好等优点,已被广泛应用于组织工程研究。骨粉中含有大量的羟基磷灰石及BMP等生长因子,具有良好的骨诱导活性。结合上述两种材料的优点,制备了一种复合海藻酸钙/骨粉微胶珠,并系统研究了人源脂肪干细胞(adipose-derived stem cells,ADSCs)在其中的最佳包埋密度。通过正交实验综合考虑海藻酸钠溶液浓度、氯化钙溶液浓度和骨粉密度对海藻酸钙/骨粉微胶珠机械强度及传质性能的影响,并采用液滴法制备了最优化的用于ADSCs体外增殖分化的微胶珠。结果表明,海藻酸钠溶液的浓度对微胶珠传质能力和机械强度的影响最大,CaCl2溶液浓度其次,骨粉密度的影响最小。当海藻酸钠溶液浓度、CaCl2溶液浓度和骨粉密度分别为2.5%,4.5%和5.0mg/mL时,微胶珠的性能最好。对细胞扩增倍数的比较和死活染色等检测表明,当ADSCs的初始包埋密度为5×106 cells/mL时,更适于ADSCs的扩增与分化。

Calcium alginate/bone-powder microbeads have been widely used in tissue engineering research due to their excellent biocompatibility,biodegradability,enhanced mechanical strength and toughness.Bone-powder containing abundant hydroxylapatite and growth factors such as BMP,possess an excellent osteoinductive activity.Herein,a hybrid calcium alginate/bone-powder microbeads combining both advantages of the abovementioned materials was prepared,afterwards the optimal inoculating density of the Adipose-derived stem cells(ADSCs)embedded in hybrid microbeads was explored systematically.The optimized microbeads suitable to the in vitro expansion and differentiation of ADSCs,were prepared using the droplet method under overall considering the concentration of calcium alginate and calcium chloride,the density of bone powder,also by the orthogonal experiment.It demonstrates that the concentration of the sodium alginate has the strongest influence on the mass transfer and mechanical strength of the microbeads,secondly the calcium chloride,then the density of bone powder.The microbeads perform best when the concentrations of the sodium alginate and calcium chloride are 2.5%and 4.5%respectively,as well as the density of bone powder was 5.0mg/mL.Additionally,the comparison about cell expanding fold and live/dead staining are carried out,which shows the ADSCs expand and differentiate well with the initial embedding density of 5×106 cells/mL.