CM-Dil标记人牙龈间充质干细胞在大鼠颅骨缺损修复模型中的示踪研究

口腔生物医学 ›› 2021, Vol. 12 ›› Issue (2): 82-86.

CM-Dil标记人牙龈间充质干细胞在大鼠颅骨缺损修复模型中的示踪研究

许璐¹,顾春宁¹,石安源²,王斌³,秦海燕⁴

1. 南京大学医学院附属口腔医院·南京市口腔医院种植科
2. 南京大学医学院附属口腔医院，南京市口腔医院
3. 南京大学医学院附属鼓楼医院
4. 南京大学医学院附属口腔医院

收稿日期:2020-12-31 修回日期:2021-05-25 出版日期:2021-06-25 发布日期:2021-06-25
通讯作者: 秦海燕 E-mail:haiyanandrew@163.com
基金资助:
国家自然科学基金;南京市卫生科技发展专项资金

CM-Dil labeled human gingival mesenchymal stem cells and its tracer study in rat skull defect repair model

Received:2020-12-31 Revised:2021-05-25 Online:2021-06-25 Published:2021-06-25

摘要/Abstract

摘要： 目的：探讨氯甲基苯甲酰氨（CM-Dil）荧光染料在体外标记人牙龈间充质干细胞（hGMSCs）后对其增殖、分化能力的影响以及在体内荧光标记持续时间。方法：组织块法原代培养hGMSCs，采用流式细胞术检测间充质干细胞表面标志物，然后以CM-Dil标记hGMSCs，CCK-8检测标记后细胞的增殖；标记及未标记的细胞进行成骨及成脂诱导培养21?d后，分别进行茜素红染色和油红O染色。大鼠颅骨缺损实验检测CM-Dil标记的hGMSCs体内标记效果。结果：原代培养的hGMSCs高表达CD105、CD90和CD73，而阴性表达CD14、CD34、CD19、CD45和人类白细胞DR抗原（HLA-DR），符合间充质干细胞表面特征。与对照组相比，CM-Dil并不影响hGMSCs的增殖和成骨、成脂分化能力（P＞0.05）。体内移植的CM-Dil标记hGMSCs在第8周时仍可检测到红色荧光。结论：CM-Dil在体外标记hGMSCs不会影响hGMSCs的干细胞特性，且体内标记时间长达8周，可作为今后干细胞体内标记示踪的方法之一。

关键词: CM-Dil, 牙龈间充质干细胞, 细胞标记, 颅骨缺损

Abstract: Objective:?To investigate the effect of chloromethyl-benzamidodialkylcarbocyanine(CM-Dil) on the proliferation and differentiation of human gingival mesenchymal stem cells(hGMSCs) in vitro and the duration of fluorescence of CM-Dil in vivo. Methods:?hGMSCs were primary cultured by tissue mass method, and surface markers were detected by flow cytometry. After hGMSCs were labeled by CM-Dil, the proliferation was analyzed using CCK-8 assay. Osteogenic differentiation of hGMSCs was investigated using Alizarin Red staining, lipid-forming ability of hGMSCs was detected by Oil Red O staining. The effect of CM-Dil-labeled hGMSCs in vivo was detected in rat skull defect experiment. Results:?The hGMSCs were positive for CD105, CD90 and CD73, but negative for CD14, CD34, CD19, CD45 and HLA-DR. Comparing with the control group, CM-Dil did not affect the proliferation of hGMSCs (P＞0.05), nor did it affect the osteogenic and lipogenic differentiation ability of hGMSCs. In vivo the CM-Dil-labeled hGMSCs could still be observed at week 8. Conclusions:?CM-Dil does not affect the characteristics of hGMSCs in vitro, and the CM-Dil-labeled hGMSCs can be observed for up to 8 weeks in vivo, and CM-Dil can be used as a tracking label of MSC in vivo in the future.

Key words: CM-Dil, gingival mesenchymal stem cells, cell lableing, skull defect model

许璐顾春宁石安源王斌秦海燕. CM-Dil标记人牙龈间充质干细胞在大鼠颅骨缺损修复模型中的示踪研究[J]. 口腔生物医学, 2021, 12(2): 82-86.

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