CAJ | 학술논문

组织工程软骨的体外构建被认为是一种有希望治疗关节软骨缺损的有效途径.为评估载脂肪干细胞(Adipose-derived stem cells,ADSCs)壳聚糖/明胶水凝胶支架,在体外动态构建组织工程软骨相对传统静态培养的优势,本研究用壳聚糖/明胶制备了软骨仿生支架,并检测其物理性质.在制备的水凝胶支架上以1×107 cells/mL 密度接种 ADSCs 后,分别置于转瓶及 T-瓶的软骨诱导基中培养两周,通过试剂染色、代谢检测和电镜观察,考察了细胞的软骨分化能力、活性、生长分布、渗透深度、增殖及胞外基质分泌情况.结果表明,壳聚糖/明胶支架的平均孔径为118．25±19．51μm,孔隙率为82．60±2．34％,吸水率为361．28±0．47％,弹性模量为61．2±0．16 kPa,具有良好生物相容性.ADSCs 生长状态良好,可向软骨细胞分化,适于作为组织工程软骨构建的种子细胞.表征结果显示,转瓶内水凝胶支架中细胞蛋白多糖的表达更显著,细胞生长分布更加均匀,细胞外基质分泌基本填满整个支架.因此,转瓶载壳聚糖/明胶支架所提供的三维动态环境,是体外构建组织工程软骨的良好方法.
조직공정연골적체외구건피인위시일충유희망치료관절연골결손적유효도경.위평고재지방간세포(Adipose-derived stem cells,ADSCs)각취당/명효수응효지가,재체외동태구건조직공정연골상대전통정태배양적우세,본연구용각취당/명효제비료연골방생지가,병검측기물이성질.재제비적수응효지가상이1×107 cells/mL 밀도접충 ADSCs 후,분별치우전병급 T-병적연골유도기중배양량주,통과시제염색、대사검측화전경관찰,고찰료세포적연골분화능력、활성、생장분포、삼투심도、증식급포외기질분비정황.결과표명,각취당/명효지가적평균공경위118．25±19．51μm,공극솔위82．60±2．34％,흡수솔위361．28±0．47％,탄성모량위61．2±0．16 kPa,구유량호생물상용성.ADSCs 생장상태량호,가향연골세포분화,괄우작위조직공정연골구건적충자세포.표정결과현시,전병내수응효지가중세포단백다당적표체경현저,세포생장분포경가균균,세포외기질분비기본전만정개지가.인차,전병재각취당/명효지가소제공적삼유동태배경,시체외구건조직공정연골적량호방법.
It was considered to be promising in treating articular cartilage defect by fabricating tissue engineered cartilage in vitro.To assess the advantages of chitosan/gelatin hydrogel scaffold seeded with ADSCs to construct tissue engineered cartilage dynamically in vitro in contrast to traditional static culture,chitosan/gelatin hybrid was chosen as cartilage biomimetic scaffold and its physical properties were subsequently tested.After that,adi-pose-derived stem cells (ADSCs)were inoculated into the chitosan/gelatin hydrogel scaffold at density of 1 × 10 7 cells/mL and cultured in a spinner flask and T-flask with chondroinductive media for two weeks,respective-ly.Chondrogenic differentiation ability of ADSCs within hydrogel scaffold was investigated with Toluidine Blue and Safranine O staining,the cell metabolism of glucose and lactic acid was analyzed through detecting kits, while the cell distribution,adhesion and extracellular matrix secretion within scaffold were observed by Dead/Live staining and scanning electron microscopy (SEM),respectively.The results showed that the average pore size,porosity,swelling rate and elasticity modulus of chitosan/gelatin hydrogel scaffold with good biocompati-bility were 1 18.25±1 9.5 1 μm,82.60±2.34%,361.28±0.47%,and 61.2±0.1 6 kPa,respectively.The ADSCs, in good conditions,could differentiate into chondrocytes,thus were suitable as seeding cells in tissue engineered cartilage construction.In addition,it was confirmed that the induced cells could express more proteoglycans in the dynamic spinner flask,where the cell distribution within the scaffold was more uniform and the scaffold could be filled mostly by extracellular matrix.The spinner flask with immobilized scaffold enhanced prolifera-tion and chondrogenic differentiation of ADSCs within chitosan/gelatin hydrogel scaffolds,thus accelerated the dynamic fabrication of cell-hydrogel constructs,which could be an excellent method to construct tissue engi-neered cartilage in vitro.