不同比例的聚乳酸-聚乙醇酸共聚物与羟基磷灰石复合支架对人牙髓干细胞增殖及矿化能力影响研究

Effects of different proportions of polylactic-co-glycolic acid and hydroxyapatite composite scaffold on the proliferation and mineralization of human dental pulp stem cells

目的探讨不同比例的聚乳酸-聚乙醇酸共聚物与羟基磷灰石(PLGA/HA)复合支架的降解速率及其对人牙髓干细胞(human dental pulp stem cells,h DPSCs)增殖及矿化能力的影响。方法采用溶液浇筑/颗粒沥析技术构建含质量分数分别为10%、20%及30%HA的PLGA/HA复合支架的3个实验组,单纯PLGA支架作为对照组。采用降解实验检测支架的降解速率。将h DPSCs接种于不同比例的PLGA/HA复合支架进行培养,扫描电镜观察细胞形态,应用MTT法检测细胞增殖能力。h DPSCs接种于各组支架培养72 h后,将复合支架植入裸鼠体内,分别于饲养1个月及3个月后处死裸鼠取出支架,应用HE染色观察组织学形态,应用牙本质基质蛋白1(DMP1)免疫组化染色观察细胞的矿化程度。结果 10%HA组和20%HA组具有较适宜的降解速率。各组均具有较高的促hDPSCs增殖能力,促进效果随HA含量的增加而加强。HE染色和免疫组化染色结果显示,HA可促进hDPSCs在体内的成牙本质向分化,分化程度与HA含量成正比,20%HA组及30%HA组矿化程度均较佳。结论含20%HA的PLGA/HA复合支架具有较佳的降解速率及促细胞增殖和矿化能力,是比较理想的细胞支架材料。

Objective To investigate the degradation rate of different proportion of polylactic-co-glycolic acid(PLGA)/hydroxylapatite(HA)composite scaffolds and the effects on proliferation and mineralization of human dental pulp stem cells(hDPSCs).Methods PLGA/HA composite scaffolds with different HA contents(10%,20%and 30%)were fabricated by solvent casting/particulate leaching technology,and PLGA without HA was used as the control group.Degradation rate of scaffolds was measured by degradation experiment.The cells were seeded in PLGA/HA composite scaffolds with different proportions.Cell morphology was observed by scanning electron microscopy,and proliferation ability were measured by MTT assay.After hDPSCs were preincubated with scaffolds for 72 h,the composite scaffolds were implanted into nude mice.One month and 3 months later,the scaffolds were removed from the sacrificed nude mice.HE staining was used to observe the histological morphology.Dentin matrix protein 1(DMP1)expression was detected by immunohistochemical staining to further investigate the mineralization degree of hDPSCs.Results PLGA/HA composite scaffolds containing 10%,20%and 30%HA could all promote the proliferation of hDPSCs compared with PLGA group,and promotion effect increased with increasing HA content.The 10%HA and 20%HA group showed suitable degradation rates.The results of HE staining and immunohistochemical staining showed that HA could promote the dentinogenesis of hDPSCs,and the degree of differentiation was proportional to the content of HA.The 20%HA and 30%HA showed a better degree of mineralization.Conclusion PLGA/HA composite scaffolds with 20%HA exhibits a better degradation rate and ability to promote proliferation and mineralization of hDPSCs,which can be used as an ideal biomaterial for cell scaffold.