聚丙交酯-乙交酯支架结合脂肪源性干细胞构建组织工程尿道

Poly(lactide-co-glycolide)scaffold combined with adipose derived stem cells in tissue-engineered urethral reconstruction

背景:生物电喷雾与静电纺丝技术的结合能促进活细胞和支架材料的直接整合,可将细胞均匀地分布在支架纤维间,是制备含细胞支架的一种很有前景的替代方法。目的:分析多方法结合制备富含脂肪源性干细胞(adipose derived stem cells,ASCs)的聚丙交酯-乙交酯[(poly(lactide-co-glycolide)),PLGA]三维支架作为尿道组织重建材料的可行性。方法:采用静电纺丝和细胞生物电喷雾相结合的方法将ASCs整合到PLGA中得到支架PLGA-ASCs,采用静电纺丝技术制备单纯PLGA支架,检测两种支架的微观结构、体外降解性能、机械性能与残留溶剂含量,MTT法检测PLGA-ASCs支架中的细胞活性。将PLGA-ASCs支架培养于37℃细胞培养箱内,培养1,7,15 d后,采用MTT法检测细胞活性,扫描电镜观察和共聚焦显微镜观察支架上的细胞生长与扩散。结果与结论:①扫描电镜显示,两种支架的纤维表面光滑,其中ASCs随机分布在PLGA-ASCs支架上,PLGA-ASCs支架的纤维平均直径、支架厚度大于PLGA支架;PLGA-ASCs支架内的细胞存活率为(87.0±4.4)%,支架中的细胞整合效率为28%;②体外降解实验显示,PLGA支架的重均分子质量在前15 d下降较快,PLGA-ASCs的重均分子质量在15-45 d下降更快,至45 d时两组支架的重均分子质量无差异;③PLGA-ASCs支架的杨氏模量、最大载荷及最大延伸率低于PLGA支架;④在培养的1-7 d内,PLGA-ASCs支架内的细胞数量逐渐增加,在7-15 d内细胞数量无明显增加;⑤扫描电镜观察和共聚焦显微镜显示,随着培养时间的延长,PLGA-ASCs支架内的细胞数量逐渐增加并与支架整合;⑥结果表明,PLGA-ASCs具有良好的理化性能与生物活性,可作为尿道组织重建材料。

BACKGROUND:The combination of bioelectrospray and electrospinning can promote the direct integration of living cells and scaffold materials,and can distribute the cells evenly among scaffold fibers,which is a promising alternative to the preparation of scaffold containing cells.OBJECTIVE:To investigate the feasibility using multiple methods to prepare poly(lactide-co-glycolide)(PLGA)three-dimensional scaffolds rich in adipose derived stem cells(ASCs)as urethral tissue reconstruction materials.METHODS:The ASCs were integrated into PLGA by electrospinning and cell bio-electrospray method.The cell scaffold PLGA-ASCs was obtained.The pure PLGA scaffold was prepared by electrospinning.The microstructure,in vitro degradation,mechanical properties and residual solvent content of the two scaffolds were detected.The cell viability of PLGA-ASCs was detected by MTT assay.PLGA-ASCs were cultured in 37°C cell incubator for 1,7 and 15 days.MTT assay was used to detect cell viability.Scanning electron microscopy and confocal microscopy were used to observe the growth and diffusion of ASCs on the scaffolds.RESULTS AND CONCLUSION:(1)Scanning electron microscopy showed that the fiber surface of PLGA-ASCs and PLGA scaffolds was smooth,and ASCs were randomly distributed on PLGA-ASCs scaffolds.The average fiber diameter and thickness of PLGA-ASCs scaffolds were larger than those of PLGA scaffolds.The cell survival rate in PLGA-ASCs scaffolds was(87.0±4.4)%,and the cell integration efficiency in PLGA-ASCs scaffolds was 28%.(2)In vitro degradation experiment showed that weight average molecular weight of PLGA scaffolds decreased rapidly in the first 15 days.The weight average molecular weight of PLGA-ASCs scaffolds decreased rapidly on 15-45 days.There was no difference in weight average molecular weight between the two groups at 45 days.(3)Young’s modulus,maximum load,and maximum elongation of PLGA-ASCs scaffolds were lower than those of PLGA scaffolds.(4)During 1-7 days of culture,the number of cells in PLGA-ASCs scaffolds gradually increased,and the number of cells in PLGA-ASCs scaffolds increased gradually from 7 to 15 days.(5)The results of scanning electron microscope observation and confocal microscope showed that the number of cells in PLGA-ASCs scaffolds increased gradually and integrated with the scaffolds with the extension of culture time.(6)The results showed that PLGA-ASCs had good physical and chemical properties and biological activity,and could be used as urethral tissue reconstruction materials.