牙周膜干细胞论文：牙周膜干细胞的培养和鉴定.doc

牙周膜干细胞论文：牙周膜干细胞的培养和鉴定【中文摘要】本研究从离体牙的牙周膜上分离出牙周膜细胞群,并对其进行纯化；在此基础上,对纯化后的细胞进行鉴定。证实该细胞为“干细胞性”细胞,为后续牙周组织工程研究提供实验基础。方法：1.分离纯化人牙周膜干细胞收集临床因正畸或阻生拔出的无龋病、无牙周病的恒牙,取根中1/3的牙周膜,采用酶解组织块法分离到牙周膜细胞群,然后有限稀释克隆法对牙周膜细胞群进行纯化,通过MTT法分析牙周膜细胞群与纯化后的牙周膜细胞的生长情况。2.牙周膜来源干细胞的初步鉴定采用流式细胞仪分析细胞周期和细胞表面标记物,经成骨诱导液诱导细胞,对细胞进行初步鉴定。结果：1.有限稀释克隆纯化后的细胞呈长梭形,通过MTT检测,证明该细胞是慢增殖周期的细胞。2.流式细胞仪检测纯化后的细胞的周期特点和细胞表面标志物,证实其符合干细胞特性。3.诱导纯化后的细胞成骨分化,细胞出现单极化,其间有针尖大小的钙结节形成,证实纯化后的细胞具有分化潜能。结论：1.酶解组织块法有效分离到人牙周膜细胞群,有限稀释克隆纯化获得的牙周膜干细胞保持了其生物学特性。该细胞具有干细胞周期特点,且有成骨分化潜能。2.培养出牙周膜干细胞,可为组织工程提供基础。【英文摘要】:Periodontal ligament cell populations were isolated from the periodontal membrane of extracted teeth and were purified; On this basis, the purified cells were identified. This paper confirmed that the cells were “stem cells” cells, to provide a basis for follow-up study of periodontal tissue engineering experimental.Methods:1. Separation and purification of human periodontal ligament stem cells Collecting the permanent teeth removal, without caries and periodontal disease, was indicated in the clinical cases due to orthodontic or impacted. Take the periodontal membrane on the middle 1/3 of the root surface. We separated cells within the periodontal tissue membrane by enzymatic, purified cells by limiting dilution cloning. The growth of periodontal ligament cell populations and purified periodontal ligament cells were analyzed by MTT.2. preliminary identification of Periodontal ligament stem cells:Cell cycle and cell surface markers were detected by flow cytometry. Osteogenic medium induced cells to identified cells.Results:1. Cells purified by limiting dilution cloning appeared Spindle. The results show that the cell is a slow proliferation cycle stem cells by MTT detection.2. The cells purified were detected cell surface markers and cell cycle characteristics by flow cytometry. The result demonstration that the cells accordance with characteristics of stem cell.3. Purified cells were induced by osteogenic medium, cells were unipolar, there have been needle the size of calcium nodules, confirmed that the purified cells possess differentiation potential.Conclusion:1. Cells which maintain their biological characteristics were effectively isolated by Enzymatic and purified by limiting dilution cloning. And the cells have characteristics of stem cell cycle and potential of Osteogenic differentiation.2. Cultured periodontal ligament stem cells provide the basis for tissue engineering.【关键词】牙周膜干细胞 组织工程 细胞鉴定【英文关键词】Periodontal ligament stem cells Tissue Engineering Cell identification【目录】牙周膜干细胞的培养和鉴定摘要3-4ABSTRACT4-5缩略词表7-8第1章 引言8-9第2章 人牙周膜干细胞的分离培养和鉴定9-18第一部分 分离纯化人牙周膜细胞9-131 材料和方法9-111.1 主要试剂和仪器9-101.2 细胞的分离和纯化10-112 结果11-122.1 细胞生长的情况112.2 细胞的克隆形成情况112.3 细胞生长曲线11-123 讨论12-13第二部分 筛选纯化后的人牙周膜干细胞的鉴定13-181 材料与方法13-151.1