Isolation and characterization of human mesenchymal stem cells from gingival connective tissue
Isolation and characterization of human mesenchymal stem cells from gingival connective tissue
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DOI：
10.1111/jre.12228
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发表时间：
2015-08-01
影响因子：
3.5
通讯作者：
Park, J. B.
中科院分区：
文献类型：
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作者：
Jin, S. H.;Lee, J. E.;Park, J. B.
本研究的主要目的是分离和鉴定牙龈结缔组织来源的间充质干细胞（GMSCs）。第二个目的是提出一种改良的分离方法为GMSCs.Material和MethodsCollected健康牙龈组织样本进行deepithelialized和切碎成小碎片。组织用分散酶和胶原酶IV消化30分钟。弃去第一次消化的细胞悬浮液，然后在相同溶液中对剩余细胞进行额外消化90分钟。从牙龈分离的细胞在37 ℃潮湿条件下孵育，并通过倒置显微镜观察。鬼笔环肽免疫荧光法评价细胞骨架形态。通过集落形成单位成纤维细胞测定来测试细胞的效力。GMSCs的特点是成骨，成脂和成软骨分化，流式细胞仪，免疫荧光analysis.ResultsGMSCs显示梭形，成纤维细胞样形态，集落形成能力，粘附塑料和多向分化（成骨，成脂，成软骨）的潜力。流式细胞仪检测GMSCs表达CD 44、CD 73、CD 90和CD 105，不表达CD 14、CD 45、CD 34和CD 19。表达干细胞标志物（SSEA-4，STRO-1，CD 146，CD 166和CD 271）和间充质标志物（波形蛋白）观察通过免疫荧光。GMSCs具有多向分化潜能，增殖能力强，具有间充质干细胞的特征。GMSC是组织工程的有前途的来源，并且可以在局部麻醉下的常规程序中获得。GSMCs的增殖和冻存潜力有待进一步研究。
Background and ObjectiveThe main purpose of this study was to isolate and characterize gingival connective tissue-derived mesenchymal stem cells (GMSCs). The secondary purpose was to present a modified isolation method for the GMSCs.Material and MethodsCollected healthy gingival tissue samples were de-epithelialized and minced into small fragments. The tissues were digested by dispase and collagenase IV for 30min. The first digested cell suspension was discarded, and then additional digestion was performed to the remaining cells in the same solution for 90min. The isolated cells from gingiva was incubated in 37 degrees C humidified condition and observed by inverted microscope. Cytoskeletal morphology was evaluated by phalloidin immunofluorescence. Potency of the cells was tested by colony-forming unit fibroblast assay. GMSCs were characterized by osteogenic, adipogenic and chondrogenic differentiation, and flow cytometric, immunofluorescence analysis.ResultsGMSCs showed spindle-shaped, fibroblast-like morphology, colony-forming abilities, adherence to plastic and multilineage differentiation (osteogenic, adipogenic, chondrogenic) potency. GMSCs expressed CD44, CD73, CD90 and CD105, but did not express CD14, CD45, CD34 and CD19 in flow cytometry. Expression of stem cell markers (SSEA-4, STRO-1, CD146, CD166 and CD271) and a mesenchymal marker (vimentin) were observed by immunofluorescence.ConclusionsIn conclusion, we isolated and characterized stem cells from human gingival connective tissue with modified protocol. GMSCs showed multipotency with high proliferation and characteristics of mesenchymal stem cells. GMSCs are promising sources for tissue engineering and may be obtained during routine procedures under local anesthesia. Further research is needed to evaluate the potential of GSMCs' proliferation and cryopreservation.