甲氨蝶呤对人牙髓间充质干细胞生物学特性的影响
The effects of methotrexate on the biological properties of dental pulp stem cells
汤颖 1朱琦 2顾永春 1沈忆芬 1冯兴梅3
作者信息
- 1. 吴江区第一人民医院中心实验室,江苏苏州215200
- 2. 吴江区第一人民医院口腔科,江苏苏州215200
- 3. 南通大学附属医院口腔科,江苏南通226000
- 折叠
摘要
目的:探讨不同浓度甲氨蝶呤(MTX)对人牙髓间充质干细胞(hDPSCs)生物学特性的影响.方法:从新鲜拔除的健康年轻恒牙中分离培养并鉴定hDPSCs;然后将细胞分为4组,分别用含0、0.1、1.0、10.0 μmol/L的MTX进行培养,检测不同浓度MTX对hDPSCs的集落形成、增殖(CCK-8法)、迁移(划痕法)、成骨分化能力(茜素红染色及Western blot实验)以及细胞周期(流式细胞术分析)影响.结果:随着MTX浓度增加,hDPSCs的细胞集落形成能力及迁移速度逐渐降低,呈浓度依赖性;1.0、10.0 μmol/L组较0.1μmol/L组对细胞增殖的抑制更显著(P＜0.05),但1.0、10.0 μmol/L组间相比无统计学差异(P＞0.05);细胞周期分析显示,加入MTX后细胞被阻滞于S期,S相细胞从35.4％增加到60.3％ ～64.5％(P＜ 0.05),但3个实验组S相细胞百分数相比P＞ 0.05;茜红素染色显示,1.0、10.0 μmol/L组明显抑制矿化结节的形成;Western blot结果显示,各组间RUNX2、OSX、ALP的表达水平相比P＞0.05.结论:MTX对hDPSCs的各项生物学特性均会产生不利作用,但影响程度会受药物浓度、作用时间的影响.
Abstract
AIM:To investigate the effects of methotrexate (MTX) on the biological properties of dental pulp stem cells (hDPSCs).METHODS:hDPSCs were isolated,cultured and identified from dental pulp of young permanent teeth extracted for orthodontic reasons.The hDPSCs were divided into 4 groups,and were cultivated in media containing 0 (control),0.1,1.0 and 10.0 μmol/L MTX respectively.The effects of MTX on the percentage of colony-forming unit (CFU-F),cell proliferation (tested by CCK-8 method),migration (scratch assay),cell circle phase (flow cytometric analysis) and osteogenic potential (Alizarin red staining and Western blot assay of expression of RUNX2,OSX and ALP) were examined.RFSULTS:The capability of CFU-F and migration was significantly reduced by the treatment of MTX concentration in a dose-related manner.The middle and high concentration groups exhibited more inhibition on cell proliferation than the low concentration group (P ＜ 0.05),while there was no statistical difference between 1.0 and 10.0 μmol/L MTX groups (P ＞ 0.05).Flow cytometric analysis showed that MTX treatment inhibited hDPSCs proliferation,and the percentage of S-phase cells increased from 35.4％ to 60.3％-64.5％ (P ＜ 0.05).Whereas there was no statistical difference among the 3 MTX groups (P ＞ 0.05).Alizarin red staining test indicated a decrease of calcified nodules with increase of MTX concentration.However,Western blot analysis showed the expression level of RUNX2,OSX,and ALP had no statistical difference among the 4 groups (P ＞ 0.05).CONCLUSION:MTX has adverse effects on multiple biological properties of hDPSCs,which can be influenced by the drug concentration and time of the treatment.
关键词
甲氨蝶呤/牙髓间充质干细胞/细胞增殖/成骨分化Key words
methotrexate/dental pulp stem cells/cell proliferation/osteogenic potential引用本文复制引用
出版年
2017
NETL
NSTL
维普
万方数据