不同原代培养方法培养人牙髓细胞的研究.pdfVIP

天津医药2011 年2 月第39 卷第2 期 139 不同原代培养方法培养人牙髓细胞的研究 沈晴昳 摘 建 方 选 要 目的： 立一种操作简单方便的人牙髓细胞原代培养方法。 法： 择因正畸拔除的完整无龋双尖牙45 颗，无菌条件下取出牙髓，分别采用组织块法、酶消化法和改良组织块法（每法15颗）进行人牙髓细胞的体外原代培 养，比较观察3 种不同培养方法的成活率、细胞形态和数量，测定生长曲线并进行细胞来源鉴定。结果：3 种方法均可 从人牙髓组织分离培养获得外胚间充质来源的牙髓细胞，且呈成纤维细胞样，抗波形丝蛋白染色阳性，抗角蛋白染色 阴性。其中，组织块法耗时较长，4 例培养成功（26.7% ）；酶消化法培养成活率较低,且细胞数量较少,2 例培养成功 结 改良组织块法解决了传统组织块法 （13.3%）；改良组织块法中未发现有漂浮的组织块，12例培养成功（80.0%）。 论： 中组织块难以贴壁的问题，简化了操作步骤并具有较高的成功率,是一种科学可行的人牙髓细胞原代培养方法。 关键词 牙髓 细胞 双尖牙 细胞培养技术 体外研究 StudyonDifferentMethodsinPrimaryCulturesofHumanDentalPulpCellsinVitro SHEN Qingyi DepartmentofProsthodontics,ShanghaiStomatologicalDiseaseCenter,Shanghai200031,China Abstract Objective: To establish a simple primary culture technique of human dental pulp cells in vitro. Methods: The pulp tissue was separated from 45 healthy young human teeth extracted for orthodontic purpose. The pulp explants were cultured by three methods, tissue explants method, tissue-explants collagenase/dispase digestion method and modified tissue culture techniques respectively. The cell culture efficiency ，cell morphology and quantity were observed and evaluated by phase-contract microscopy and growth curres of human pulptissue cells. Immunocytochemical staining method was used to characterize the dental pulp cell lineage. Results:The cultured cells appeared to be fibroblast-like cells, which was positive for anti-vimentin and negative for anti-cytokeratin by immunohistochemical stain. The primary dental pulp cells cultured by the tissue explants method could reach confluence after 4 weeks and the culture success