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目的 :建立人嗅黏膜间充质干细胞的获取、分离、培养方法。方法 :通过人嗅黏膜组织贴壁法体外分离、培养嗅黏膜间充质干细胞,观察细胞形态学,绘制其生长曲线,观察其细胞周期分析,进行流式细胞仪检测来观察其细胞表面标记物。结果 :培养的人嗅黏膜间充质干细胞形态上以梭形为主,细胞排列呈放射状,细胞增殖迅速。生长曲线及细胞周期结果提示人嗅黏膜间充质干细胞符合细胞正常生长特征且细胞增殖较活跃。第5代人嗅黏膜间充质干细胞表达间充质干细胞阳性标记物CD73,CD90,不表达造血干细胞标记物CD34,CD45。细胞进行成脂肪组织、成骨组织诱导后,细胞油红O染色可见红色脂滴,茜素红染色后可见钙盐沉积。结论 :人嗅黏膜组织块贴壁培养法是一种有效的具有简便操作性的方法,通过此方法培养出的细胞具有间充质干细胞的一般生物学特性,细胞经诱导后具有多向的分化潜能,为其今后的临床应用提供了充足的种子细胞来源具有重大意义。
Objective: To establish a method for obtaining, isolating and culturing human olfactory mucosa-derived mesenchymal stem cells. Methods: Human olfactory mucosa-derived mesenchymal stem cells (MSCs) were isolated and cultured in vitro by human olfactory mucosa tissue adherent method. Morphology was observed and their growth curves were drawn. Cell cycle analysis was performed. Flow cytometry was used to observe the cell surface markers . Results: The cultured human olfactory mucosa-derived mesenchymal stem cells were spindle-shaped with radial arrangement of cells and rapid cell proliferation. Growth curve and cell cycle results suggest that human olfactory mucosa-derived mesenchymal stem cells conform to the normal growth characteristics of cells and cell proliferation is more active. The 5th generation human olfactory mucosa-derived mesenchymal stem cells express mesenchymal stem cell positive markers CD73, CD90, but do not express hematopoietic stem cell markers CD34, CD45. Cells into adipose tissue, osteogenic tissue after induction, the cells can be seen red oil droplets O red lipid droplets, alizarin red staining of calcium deposition. CONCLUSION: Human olfactory mucosa tissue adherent culture method is an effective and easy-to-operate method. The cells cultured by this method have the general biological characteristics of mesenchymal stem cells, and the cells are induced to have multi-directional differentiation Potential for its future clinical application provides ample source of seed cells of great significance.