摘要
Objective： To deliver cells deep into injectable calcium phosphate cement（CPC） through alginate-chitosan（AC） microcapsules and investigate the biological behavior of the cells released from microcapsules into the CPC.Methods： Mouse osteoblastic MC3T3-E1 cells were embedded in alginate and AC microcapsules using an electrostatic droplet generator.The two types of cell-encapsulating microcapsules were then mixed with a CPC paste.MC3T3-E1 cell viability was investigated using a Wst-8 kit,and osteogenic differentiation was demonstrated by an alkaline phosphatase（ALP） activity assay.Cell attachment in CPC was observed by an environment scanning electron microscopy.Results： Both alginate and AC microcapsules were able to release the encapsulated MC3T3-E1 cells when mixed with CPC paste.The released cells attached to the setting CPC scaffolds,survived,differentiated,and formed mineralized nodules.Cells grew in the pores concomitantly created by the AC microcapsules in situ within the CPC.At Day 21,cellular ALP activity in the AC group was approximately four times that at Day 7 and exceeded that of the alginate microcapsule group（P0.05）.Pores formed by the AC microcapsules had a diameter of several hundred microns and were spherical compared with those formed by alginate microcapsules.Conclusions： AC microcapsule is a promising carrier to release seeding cells deep into an injectable CPC scaffold for bone engineering.
研究目的:本研究应用海藻酸钠-壳聚糖微囊保护成骨细胞,接种到β-磷酸三钙/磷酸钙骨水泥(β-TCP/CPC)浆料中,使β-TCP/CPC骨修复材料具有一定的细胞活性,同时提高固化后材料的孔隙率和孔径,以最终实现提高β-TCP/CPC骨水泥的降解速度,加快成骨和骨修复。创新要点:本研究首次应用海藻酸钠-壳聚糖微胶囊包封成骨细胞与CPC浆料复合,复合后实现自动细胞释放,释放出的细胞具有良好的生物学活性。研究方法:(1)高压静电成囊法制备载小鼠成骨前体细胞(MC3T3-E1)的海藻酸钙和海藻酸钠-壳聚糖微胶囊;(2)微囊化MC3T3-E1细胞,进行体外培养,使用细胞计数试剂盒(CCK-8)检测细胞活性,并用钙黄绿素-AM(Calcein-AM)和碘化丙啶(PI)进行活死细胞双重染色;(3)微囊化MC3T3-E1细胞与β-TCP/CPC浆料复合培养后,激光共聚焦扫描显微镜和环境扫描电子显微镜观测细胞在材料上的释放、粘附,CCK-8法检测材料上细胞的活力,碱性磷酸酶(ALP)检测观察细胞的分化状况,茜素红染色观察释放细胞的矿化能力。重要结论:海藻酸钠-壳聚糖微胶囊可作为可注射磷酸钙骨水泥内部接种成骨细胞并实现细胞释放的良好载体,释放出的成骨细胞具有良好的生物学活性。
基金
supported by the National Natural Science Foundation of China(No.30772447)
the Talent Introduction Project of Peking University Health Science Center(No.bmu2009139),China
