体外培养人牙周膜细胞方法的探讨

An Investigation on Culturing Method of Human Periodontal Ligament Cells

目的:研究如何提高体外培养人牙周膜细胞(hPDL)的成功率,为简便、快速地获得大量成活率高的hPDL膜细胞,建立体外细胞模型提供一定的实验方法。方法:利用3种不同方法(组织块法、酶消化组织块法、全牙消化法)进行牙周膜细胞的原代培养,用形态学、免疫细胞化学染色等方法鉴定细胞来源;通过生长曲线的测定研究体外细胞生长特性。结果:3种方法获得的细胞形态和生物学行为大致相同;波丝蛋白抗体染色阳性,角蛋白抗体染色阴性。组织块法培养的细胞同源性好,但初代培养时间长;全牙消化法的原代培养成功率高于其他2种方法。结论:通过对牙周膜细胞培养方法的改良,可以显著提高hPDL原代培养的成功率。

Objective： To establish a simple, fast, and efficient technique for human periodontal ligament （hPDL） cell in vitro cultivation with higher success rate. Methord： Primary hPDL cells were cultured using three different methods （tissue explant, explant with enzymatic digestion, and tooth enzymatic digestion）. Morphological analysis and immunocytochemical staining were used to characterize the cell lineage, and growth curve assay was adopted to evaluate the biological features of hPDL cells. Results ： Morphological and biological characteristics of cells cultured with the 3 kinds of method were similar. They were spindle-shaped, and showed positive reaction to antibodies against vimentin, and negative reaction to antibodies against keratin. Cells obtained with tissue explant showed better homology, but need longer culture time ; The success rate of primary hPDL cell culture in tooth enzymatic digestion group was significantly higher than those in the other 2 groups. Conclusion： By making improvement in specimen collecting and culturing methods, the success rate of primary culture of hPDL cells can be noticeably increased.