Isolation and characterization of human gingiva-derived mesenchymal stem cells using limiting dilution method
Isolation and characterization of human gingiva-derived mesenchymal stem cells using limiting dilution method
复制标题
使用有限稀释法分离和表征人牙龈来源的间充质干细胞
DOI：
10.1016/j.jds.2016.03.010
复制
发表时间：
2016-09-01
影响因子：
3.5
通讯作者：
Ge, Shaohua
中科院分区：
文献类型：
--
作者：
Du, Lingqian;Yang, Pishan;Ge, Shaohua
背景/目的：牙龈源性间充质干细胞（GMSC）是有吸引力的替代MSC来源，因为它们的来源相对丰富和易于获得。然而，用于窝藏GMSC的分离方法仍在讨论中。本研究的目的是分离、培养人牙龈间充质干细胞（GMSCs），并与原代培养的牙龈成纤维细胞（GFs）进行比较。结果：GMSCs的集落形成单位-成纤维细胞数量显著高于GFs（P < 0.001），但GMSCs的群体倍增能力降低。两种类型的细胞对MSC相关标记物CD 44、CD 73、CD 90、CD 105和CD 166均呈阳性，而对造血标记物CD 14、CD 34和CD 45呈阴性。唯一不同的标志物是STRO-1，其在GMSC中的表达（13.4%）比在大量培养的GF中的表达（0.02%）更高。在诱导后，GMSC显示出进行成骨、成脂和成软骨分化的能力。实时荧光定量聚合酶链反应显示，相关基因表达水平显著上调（P < 0.001）。相比之下，大量培养的GFs缺乏进行多向分化的能力，相关基因水平与对照组相比无显著差异。结论：数据验证了有限稀释法分离GMSCs的有效性。与大量培养的GF相比，GMSC具有干细胞特性，可以作为组织工程的替代细胞来源。版权所有（C）2016中华民国口腔医学会由爱思唯尔台湾有限公司出版。
Background/purpose: Gingiva-derived mesenchymal stem cells (GMSCs) are attractive alternative MSC sources because of their relative abundance of sources and ease of accessibility. However, the isolation method for harboring GMSCs remains under discussion. The aim of the study was to isolate and explore in vitro characterization of human GMSCs, and compare stem cell properties with bulk-cultured gingival fibroblasts (GFs).Materials and methods: GMSCs were isolated with limiting dilution method. Tissue-matched bulk-cultured GFs and GMSCs were evaluated in terms of their colony-forming abilities, population doubling capacities, cell surface epitopes, and multilineage differentiation potentials.Results: GMSCs showed a significantly higher number of colony-forming units-fibroblast (P < 0.001) than bulk-cultured GFs, while the population doubling capacity of GMSCs reduced. Both types of cells were uniformly positive for MSC-associated makers CD44, CD73, CD90, CD105, and CD166, and were negative for hematopoietic markers CD14, CD34, and CD45. The only distinct marker was STRO-1, which was more highly expressed in GMSCs (13.4%) than in bulk-cultured GFs (0.02%). Upon induction, GMSCs displayed the capacity to undergo osteogenic, adipogenic, and chondrogenic differentiation. Real-time polymerase chain reaction showed related gene levels were significantly upregulated (P < 0.001). By contrast, bulk-cultured GFs lacked the capacity to undergo multilineage differentiation, and related gene levels showed no significant difference when compared with control groups.Conclusion: The data validate the effectiveness of limiting dilution method for GMSCs isolation. GMSCs, in contrast to bulk-cultured GFs, harbor stem cell characteristics and can act as alternative cell sources for tissue engineering. Copyright (C) 2016, Association for Dental Sciences of the Republic of China. Published by Elsevier Taiwan LLC.