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低氧对人牙周膜干细胞增殖和分化的影响

Effect of Hypoxia on Proliferation and Differentiation of Human Periodontal Ligament Stem Cells.
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摘要 目的:观察低氧预处理对人牙周膜干细胞(human periodontal ligament stem cells,hPDLSCs)干性的影响。方法:收集hPDLSCs并进行鉴定后,分别在常氧(20%O2)和低氧(5%O2)环境下培养5d,噻唑蓝(methyl thiazol tetrazolium,MTT)法检测细胞的增殖能力。分别在低氧环境培养1d和5d,实时定量聚合酶链反应(Real-time quantitative polymerase chain reaction,Real-time PCR)检测hPDLSCs中干性相关基因Oct4、Sox2、Nanog的mRNA表达水平。细胞增殖核抗原Ki-67染色法和结晶紫染色观察细胞自我更新能力。分别用成骨和成脂诱导hPDLSCs 21d,观察低氧对人牙周膜干细胞多项分化能力的影响。结果:MTT结果显示低氧预处理能显著增强hPDLSCs增殖能力(P<0.05)。Real-time PCR结果显示低氧在第1天和第5天时能够提高细胞的干性基因表达(P<0.05),其中在第5天的时候SOX2的提高最为显著(P<0.05)。结晶紫染色结果表明,低氧环境能够增加细胞克隆形成率(P<0.05)。低氧环境还能增强干细胞的成骨分化能力,成骨相关基因RUNX2的表达显著提高(P<0.05),但是对成脂相关基因的增强不明显,对成脂标志基因PPARγ2的影响不明显。结论:低氧预处理hPDLSCs能够提高细胞的增殖和干性相关基因的表达。另外,低氧环境也能提高干细胞的分化,其中对成骨相关基因的促进作用较为显著,但对成脂分化影响不大。 Objective: To examine the effect of hypoxia on human periodontal ligament stem cells (hPDLSCs).Methods: HPDLSCs were collected and validated.Cells were cultured in normoxia (20% O 2) or hypoxia (5%O 2) for 5 days.MTT was applied to detect the cell proliferation ability in each day.OCT4,SOX2,and NANOG genes were analyzed by RT-PCR in cells culture in hypoxia at day 1 and day 5.Ki67 staining and crystal violet staining assay were used to detect the self-renewal ability of hPDLSCs.Osteogenic and adipogenic differentiation abilities were investigated after 21-day-induction.Results: The results from MTT assay showed that the proliferation ability of hPDLSCs treated with hypoxia was higher (P<0.05).Hypoxia treatment enhanced the related gene expression at both day 1 and day 5 (P<0.05).SOX2 was enhanced strongly at day 5 ( P <0.01).Crystal violet staining assay indicated that colony-forming efficiency was increased in hypoxia group ( P < 0.05).In addition,hypoxia also enhanced the osteogenic differentiation capacity of hPDLSCs.Osteogenic gene RUNX2 was significantly increased (P<0.05).However,although hypoxia increased the expression of adipogenic gene PPAR γ2 and adipogenic differentiation,the results was not statistically significant.Conclusion: Hypoxic preconditioning of hPDLSCs can increase cell proliferation and expression of stemness-related genes.In addition,hypoxia environment can also improve the osteogenesis differentiation of hPDLSCs,but has little effect on adipogenic differentiation.
作者 郑树灿 邹晖 黄旭瑶 邵海宾 周勇 张瑞芳 李朝晖 ZHENG Shu-can;ZOU Hui;HUANG Xu-yao;SHAO Hai-bin;ZHOU Yong;ZHANG Rui-fang;LI Chao-hui(Stomatology Center, Guangzhou Huadu District Maternal and Child Health Hospital,Affiliated Guangzhou Huadu Hospital, Guangdong Medical University, Guangzhou 510800, China.)
出处 《口腔医学研究》 CAS 北大核心 2019年第1期38-41,共4页 Journal of Oral Science Research
基金 广东省自然科学基金(编号:2013B021800076).
关键词 低氧 人牙周膜干细胞 增殖 分化 Hypoxia Human periodontal ligamentstem cells Proliferation Differentiation
作者简介 郑树灿(1983~),男,广东潮州人,硕士,主治医师,主要从事口腔修复学和口腔种植学研究;通讯作者:李朝晖,E-mail: gdlzh1013@163.com
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