CAJ | 학술논문

目的探讨新型壳聚糖基网络复合膜诱导骨髓间充质干细胞(MSCs)定向成骨分化的可行性.方法采用仿生学方法,壳聚糖、明胶、果胶按照一定比例制作成新型壳聚糖基仿生网络复合膜.设计4个组:实验组1(复合膜+常规培养基),对照组1(常规培养基),实验组2(复合膜+成骨诱导(OS)培养基),对照组2(OS培养基).通过倒置相差显微镜、四甲基偶氮唑盐(MTT)法、扫描电镜(SEM)检测细胞在复合膜上的生长和增殖情况;通过测定碱性磷酸酶(ALP)活性来评价MSCs在复合膜上的成骨分化能力;通过特殊染色和能谱分析(EDX)来评定钙盐的沉积.结果 MSCs在网络复合膜上贴附、生长良好且增殖旺盛.MTr法检测细胞活力显示实验组吸光度(OD)值与对照组比较无统计学意义(P＞0.05).SEM观察到细胞在支架材料表面呈聚集生长,分泌大量的细胞外基质,可见散在的结节形成.实验组1的ALP活性明显增高,与实验组2、对照组比较有统计学意义(P＜0.01).茜素红和Von Kossa染色可见实验组细胞分化后形成的钙化结节;ALP染色可见胞浆内蓝染颗粒;EDX检测到Ca、P沉积.结论新型壳聚糖基网络复合材料具有良好的生物相容性,在不添加诱导剂的条件下,可以诱导MSCs定向成骨分化.
목적 탐토신형각취당기망락복합막유도골수간충질간세포(MSCs)정향성골분화적가행성.방법 채용방생학방법,각취당、명효、과효안조일정비례제작성신형각취당기방생망락복합막.설계4개조:실험조1(복합막+상규배양기),대조조1(상규배양기),실험조2(복합막+성골유도(OS)배양기),대조조2(OS배양기).통과도치상차현미경、사갑기우담서염(MTT)법、소묘전경(SEM)검측세포재복합막상적생장화증식정황;통과측정감성린산매(ALP)활성래평개MSCs재복합막상적성골분화능력;통과특수염색화능보분석(EDX)래평정개염적침적.결과 MSCs재망락복합막상첩부、생장량호차증식왕성.MTr법검측세포활력현시실험조흡광도(OD)치여대조조비교무통계학의의(P＞0.05).SEM관찰도세포재지가재료표면정취집생장,분비대량적세포외기질,가견산재적결절형성.실험조1적ALP활성명현증고,여실험조2、대조조비교유통계학의의(P＜0.01).천소홍화Von Kossa염색가견실험조세포분화후형성적개화결절;ALP염색가견포장내람염과립;EDX검측도Ca、P침적.결론 신형각취당기망락복합재료구유량호적생물상용성,재불첨가유도제적조건하,가이유도MSCs정향성골분화.
Objective To study the osteogenic differentiation capacity of mesenchymal stem cells(MSCs)on chitosan based biomimetic network composites films.Methods The chitosan-based biomimetic network composites films were prepared using chitosan,gelatin and pectin in certain ratio by biomimetic approach.The experiment was designed as follow:experimental group 1 (composites films+routine medium),control group 1 (routine medium),experimental group 2(composites films+osteogenic medium),control group 2(osteogenic medium).The growth and proliferation of MSCs were observed by inverted phase contrast microscope and scanning electronmicroscopy(SEM) and detected through methyl thiazolyl tetrazolium(MTT) assay.Alkaline phosphatase(ALP) activity was used to investigate the osteogenic differentiation capacity.The formation of mineral was determined by special staining and the energy dispersive X-ray apparatus (EDX).Results MSCs could well adhere,grow and proliferate on the network composites films.The MTT assay demonstrated that there was no significant difference in cell viability between the experimental and control groups (P＞0.05).SEM observation showed that MSCs grew assembly on the surface of the films secreted a large number of extracelluar matrix,and formed scattered nodules.The cells on the experimental group 1 showed statistically higher ALP activity level than those cells on the other groups (P＜0.01).Moreover,the calcified nodules can be seen in the experimental groups by Alizarin red and Von Kossa staining.ALP staining showed blue-staining granules within the cytoplasm,and the deposition of Ca and P was detected on the films by SEM-EDX.Conclusion The novel chitosan-based biomimetic network composites films not only have good biocompatibility,but can also improve MSCs differentiation toward the osteogenic lineage.