解剖学报

解剖学报 ›› 2019, Vol. 50 ›› Issue (4): 527-532.doi: 10.16098/j.issn.0529-1356.2019.04.021

• 生物工程学 • 上一篇    下一篇

条件培养基诱导人牙髓干细胞分化为角膜上皮样细胞

葛芳 杜立群*   

  1. 山东大学齐鲁医院眼科，济南 250012
  • 收稿日期:2018-10-08 修回日期:2018-12-17 出版日期:2019-08-06 发布日期:2019-08-06
  • 通讯作者: 杜立群 E-mail:dlqzqt@163.com
  • 基金资助:
    山东省自然科学基金面上项目

Differentiation of human dental pulp stem cells into corneal epithelial-like cells by conditioned medium

GE Fang DU Li-qun*   

  1. Department of Ophthalmology, Qilu Hospital of Shandong University, Ji’nan 250012, China
  • Received:2018-10-08 Revised:2018-12-17 Online:2019-08-06 Published:2019-08-06
  • Contact: DU Li-qun E-mail:dlqzqt@163.com

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摘要：

目的 探讨条件培养基（CM）诱导人牙髓干细胞（DPSCs）分化为角膜上皮样细胞的可行性。 方法 体外分离培养DPSCs，通过流式细胞术鉴定DPSCs，细胞计数试剂盒-8（CCK-8）检测基础培养基（BM）和不同比例CM培养的DPSCs增殖活性，选用30%、60%、90% CM诱导DPSCs，BM培养的DPSCs设为阴性对照组，免疫荧光检测角膜上皮细胞标志物细胞角蛋白3（CK3）、细胞角蛋白12（CK12）的表达。 结果 DPSCs增殖活性的差异无统计学意义（P> 0.05）；诱导3 d时，30%、60%、90% CM组细胞不表达CK3，60%和90%CM组细胞开始表达CK12；7 d时，30%、60%、90%CM组细胞可见CK3、CK12的表达；11 d和14 d时，30%、60%、90%CM组细胞继续表达CK3、CK12。BM组细胞未见CK3、CK12表达。 结论 在一定诱导条件下，DPSCs可分化为角膜上皮样细胞。

关键词: 牙髓干细胞, 条件培养基, 流式细胞术, 免疫荧光, 人

Abstract:

Objective To investigate the feasibility of inducing human dental pulp stem cells (DPSCs) to differentiate into corneal epithelial-like cells by conditioned medium (CM). Methods DPSCs were isolated and identified by flow cytometry. The effect of basal medium (BM) and different CM on the proliferation activity of DPSCs was detected by cell counting kit-8(CCK-8) assay. DPSCs were induced by 30%CM, 60%CM, 90%CM. The cells cultured in BM were negative control group. Corneal epithelial cells markers cytokeratin 3 (CK3) and cytokeratin 12 (CK12) were detected by immunofluorescence assay. Results There was no significant difference in the proliferation activity of DPSCs between BM group and different CM group(P> 0.05). Cells in the 30%, 60%, 90%CM group did not express CK3 after 3 days induction, cells in the 60%and the 90%CM group began to express CK12; CK3 and CK12 were expressed in the 30%, 60%, 90%CM group after 7 days; At the 11th and 14th day, cells continued to express CK3 and CK12 in the 30%, 60%, and 90% CM groups. No expression of CK3 and CK12 was observed in the BM group. Conclusion DPSCs are capable of differentiating into corneal epithelial-like cells under the induction of CM.

Key words: Dental pulp stem cell, Conditioned medium, Flow cytometry, Immunofluorescence, Humen