【作者】 谭晓华；

【导师】 李玉林； 刘晋宇；

【作者基本信息】 吉林大学 ， 病理学与病理生理学， 2010， 硕士

【摘要】 疾病对人类的健康和生存构成重大威胁、是世界各国普遍面临的最重要的社会问题。干细胞移植技术的问世和进展为传统治疗手段认为是“不治之症”的患者带来了新的生机和希望。但干细胞的来源、自我更新潜能的维持、向功能细胞分化的潜能以及移植治疗的安全性、有效性一直是干细胞基础理论和实际应用研究中急待解决的关键科学问题。毛囊是皮肤的附属器之一,起源于表皮与间充质间的相互作用,终生自我更新,是存在于成体内的胚胎样器官。新近研究发现,毛囊中除了含有表皮干细胞和黑色素干细胞外,还含有具有多向分化潜能的间充质干细胞。这些多潜能干细胞具有自我更新和定向分化成其它细胞的能力。与其他类型间充质干细胞相比,毛囊干细胞具有来源丰富,获取方便,不受年龄性别限制,不涉及伦理问题等优点。首先,本研究探索了一种简便可行的获取毛囊多潜能干细胞的方法,通过应用组织块培养法,从拔取的人毛发中分离培养出多潜能毛囊干细胞,并应用免疫细胞化学法结合流式细胞仪技术,对毛囊多潜能干细胞的表型进行了分析鉴定,结果显示多潜能毛囊干细胞表达间充质干细胞相对特异的表面标记物CD29、CD44、CD90、CD105(SH-2)、CD73(SH-4)表面抗原,不表达CD31、CD34、CD45、HLA-DR、K15等表面抗原。其次,本实验初步建立了毛囊多潜能干细胞传代、扩增、冻存与复苏的方法。最后,本实验成功诱导多潜能毛囊干细胞向成骨细胞、脂肪细胞分化。探索了多潜能毛囊干细胞向神经样细胞诱导分化的条件,通过免疫细胞化学法检测诱导前后神经元特异性烯醇化酶(NSE)、神经丝蛋白(NF-L)、胶质纤维酸性蛋白(GFAP)、神经巢蛋白(nestin)、神经微管蛋白Ⅲ(β-tubulinⅢ)、S100钙结合蛋白B(S100B)及环核苷磷酸二酯酶(CNPase)的表达水平。认为双丁酰环磷腺苷(Dibutyryl Cyclic AMP)联合3-异丁基-1-甲基黄嘌呤(IBMX)是目前本实验条件下诱导人毛囊干细胞向神经元细胞分化最为有效的方法。为后期利用多潜能毛囊干细胞移植修复损伤神经、帕金森及其它神经系统病变奠定了必要的前期工作基础。更多还原

【Abstract】 The optimal source of stem cells for regenerative medicine is a major question. Embryonic stem (ES) cells have shown promise for pluripotency but have ethical issues and potential to form teratomas. Pluripotent stem cells have been produced from skin cells by either viral-, plasmid- or transposon-mediated gene transfer. These stem cells have been termed induced pluripotent stem cells or iPS cells. iPS cells may also have malignant potential and are inefficiently produced. Embryonic stem cells may not be suited for individualized therapy, since they can undergo immunologic rejection.Recent researches found that hair follicle contains a variety of stem cells e×cepts epidermal stem cells and melanoma cells . hair follicle pluripotent stem cells present in the hair follicle dermal papilla and dermal sheath with self-renewal ability and differentiation potential. under certain condition cells can differentiate into a variety of cells. The cells in the adult dermal papilla and dermal sheath are well-characterised particularly because of their remarkable inductive powers.Directly isolated and cultured dermal cells from mature adult follicles have been shown to be capable of inducing a range of epithelia to form follicles.multipotent human follicular stem cells in the body with slow periodic, but have high proliferative capacity specific in vitro conditions. multipotent human follicular stem cells have self-renewal ability and differentiation potential, under certain conditions cells can differentiate into a variety of functions. multipotent differentiation capacity is the most important characteristic to the identification of stem cells. More and more research have proved that mesenchymal stem cells can different into a variety of cell by given the cultivation of different cell differentiation conditions.Our e×periment is studying on the multipotent human follicular stem cells’segregation, culture and identification. In e×periment, we obtained the human hair follicle-derived mesenchymal-like stem cells simply by plucking the adults’hair . The flow cytometry analysis shows their high e×pression of mesenchymal stem cells related markers , including: CD29、CD44、CD73、CD90、CD105;rare e×pression of epidermal stem cells marker CK15, hematopoietic stem cells marker CD34,leukocyte marker CD45,endothelial cell marker CD31 and immune response-related major histocompatibility antigen HLA-DR.All the above indicates that these cells are rich in mesenchymal cells. The multipotent human follicular stem cells showed adipogenic and osteogenic capacity and can different into neuronal-like cells with db-cAMP and IBMX. So multipotent human follicular stem cells may treat spinal cord injury, Parkinson’s disease and other nervous system disease one day.更多还原