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背景:人脐带沃顿胶间充质干细胞满足了国际细胞治疗协会规定的间充质干细胞的特点,能够向骨、软骨、脂肪、肌肉、神经细胞诱导分化并支持其他干细胞的扩增,对免疫系统有良好的耐受性,对肿瘤有定向迁徙性。目的:观察人脐带沃顿胶间充质干细胞与脑肿瘤干细胞体外共培养后,脑肿瘤干细胞的生物学变化。方法:以原位法培养人脐带沃顿胶间充质干细胞,以酶消化法培养人脑肿瘤组织脑肿瘤干细胞,以细胞传代法获取第3代细胞。应用不添加任何生长因子的无血清培养基将两种细胞在24孔板中进行直接共培养。第3,7天应用流式细胞术检测细胞CD133表达;应用免疫荧光法检测贴壁细胞巢蛋白和胶质纤维酸性蛋白表达;将第3天离心所得的共培养上清液重悬第3代脑肿瘤干细胞并与正常培养悬浮的第3代脑肿瘤干细胞置入96孔板中,应用酶标仪检测两组细胞生长曲线的差异。结果与结论:两种细胞共培养后在倒置显微镜下可见脑肿瘤干细胞球随着培养时间的增加出现分解、贴壁、分化现象;贴壁的脑肿瘤干细胞免疫荧光染色胶质纤维酸性蛋白和巢蛋白均阳性。恶性程度高的脑肿瘤组织培养的脑肿瘤干细胞表达CD133量越高,而与沃顿胶间充质干细胞共培养后随着时间的变化均出现CD133表达量降低。共培养3d的上清液培养的脑肿瘤干细胞与正常培养基培养的脑肿瘤干细胞相比,增殖明显受到抑制。结果显示沃顿胶间充质干细胞与脑肿瘤干细胞体外共培养后可限制脑肿瘤干细胞表面标记物CD133的阳性率以及细胞增殖能力并促使其分化。
Background: Human umbilical cordon mesenchymal stem cells (MSCs) fulfill the characteristics of mesenchymal stem cells regulated by International Association for Cell Therapy. They can induce differentiation into bone, cartilage, fat, muscle and nerve cells and support expansion of other stem cells. The system is well tolerated and has targeted migration to the tumor. OBJECTIVE: To observe the biological changes of brain tumor stem cells after human umbilical cord blood mesenchymal stem cells and brain tumor stem cells co-cultured in vitro. Methods: Human umbilical cord blood Wound-mesenchymal stem cells were cultured by in situ method and brain tumor stem cells were cultured by enzyme digestion. The third passage cells were obtained by cell passage method. Both cells were directly co-cultured in 24-well plates with serum-free medium without addition of any growth factors. Flow cytometry was used to detect the expression of CD133 on days 3 and 7; the expression of adherent cells nestin and glial fibrillary acidic protein was detected by immunofluorescence; and the co-culture supernatant was resuspended on the third day for the third generation Brain tumor stem cells and the third generation of brain tumor stem cells suspended in normal culture were placed in 96-well plates. The difference of cell growth curve between the two groups was detected by microplate reader. RESULTS AND CONCLUSION: After co-culture of two kinds of cells, under inverted microscope, the brain tumor stem cells showed decomposition, adherent and differentiation as the culture time increased. The adherent brain tumor stem cells immunofluorescence staining glial fibrillary acidic protein and nest Protein positive. The expression of CD133 was higher in brain tumor stem cells cultured in malignant brain tumors than that in control group. However, the expression of CD133 was decreased with the passage of time after co-cultured with Wharton ’s Dayton mesenchymal stem cells. The proliferation of brain tumor stem cells cultured in the supernatants of three days co-cultured with brain tumor stem cells cultured in normal medium was significantly inhibited. The results showed that the co-culture of Wharton’s cell mesenchymal stem cells and brain tumor stem cells in vitro could limit the positive rate of CD133 on the surface of brain tumor stem cells as well as the ability of cell proliferation and differentiation.