包埋PLGA微球的可控释壳聚糖支架材料的研究

Sustained Releasable Chitosan Scaffolds Embedded with PLGA Microspheres

为解决组织工程支架材料内部营养供应不足的问题,将营养物质包埋在聚合物微球内再植入支架,通过微球内营养物质的控制释放保持支架内部营养物质浓度的持续均匀性。以牛血清白蛋白(BSA)为模型蛋白,乳酸-羟基乙酸共聚物(PLGA)为外包被材料,采用复乳法(w/o/w)制备聚合物微球,然后将该微球与壳聚糖溶液混合,冻干形成支架。以BCA法检测载药微球自身的释放情况以及其植入到支架后的释放情况,通过扫描电镜观察载药微球的结构以及其植入支架材料后结构变化。结果表明,微球形态圆整,粒径范围在27～55μm之间。壳聚糖支架呈多孔状结构,1%壳聚糖支架孔隙率、吸水率和降解率分别为(92.99±2.51)%、(89.66±0.66)%和(73.77±3.21)%。体外释放显示:微球可以持续释放所包埋的蛋白,突释较小,168h后PLGA微球的累积释放量为(20.24±0.83)%。电镜照片显示,微球植入支架后,与支架结合紧密,微球结构没有发生明显改变。168h后支架内部蛋白浓度为(11.44±1.81)×10-2mg·mL-1。相比传统靠外部培养基自由扩散到支架内部为支架内细胞供养的方法,营养物质或细胞因子包埋在控释微球中并与支架材料复合成可控释支架,可以长时间维持支架中这些因子的浓度均匀,从而为组织工程提供理想的支架材料。

In order to solve the problem of nutrition insufficiency in the tissue engineering scaffolds,the microspheres containing sustained releasable nutrition were incorporated into the chitosan-based scaffolds to make that a stable nutrition concentration can be maintained within the scaffolds by the sustained release of the nutrition in the microspheres.The PLGA microspheres containing BSA used as model protein were prepared by double emulsion method,then through the processes of mixing them with chitosan solution and freeze-drying,the chitosan scaffolds were formed.The BSA release profiles of the microspheres before and after incorporated into the chitosan scaffolds were monitored by ELIASA（enzyme linked immunosobent assay） with BCA protein assay.The configuration of PLGA microspheres and the configuration changes of the microspheres and chitosan scaffolds after the microspheres were incorporated into the scaffolds were observed by scanning electron microscope.The results show that the PLGA microspheres can be uniformly embedded into the scaffolds without any morphology change and they have a size range of 27～55 μm and the chitosan scaffolds present the construction with uniform porous.It was found that the porosity,water content and degradation ratios of the chitosan scaffolds are（92.99±2.51）%,（89.66±0.66）% and（73.77±3.21）%,respectively.The in vitro BSA release tests of the microspheres show that the BSA release is sustained and the bursting release is very few.The cumulative release amount of BSA from PLGA microspheres within 168 h is（20.24±0.83）%,and the protein concentration in the chitosan scaffolds after 168 h is（11.44±1.81）×10-2 mg·mL-1.Comparing with the traditional nutrition supply method of providing the nutrition to cells locating inside the scaffolds by free diffusion of the external culture medium,the proposed method of using the novel chitosan-based scaffolds embedded with nutrition containing PLGA microspheres is better,for it has the sustained release ability and can maintain a homogeneous concentration of the cells growth factors inside the scaffolds for a long time.In other words,the proposed method provides the promising ideal scaffolds for the tissue engineering.