细胞培养与干细胞技术.pptVIP

* Figure 5. Embryoid Body-Mediated Differentiation of Human iPS Cells (A) Floating culture of iPS cells at day 8. (B–E) Images of differentiated cells at day 16 (B), neuron-like cells (C), epithelial cells (D), and cobblestone-like cells (E). (F–K) Immunocytochemistry of a-fetoprotein (F), vimentin (G), a-smooth muscle actin (H), desmin (I), bIII-tubulin (J), and GFAP (K). Bars =200 mm (A and B) and 100 mm (C–K). Nuclei were stained with Hoechst 33342 (blue). (L) RT-PCR analyses of various differentiation markers for the three germ layers. * Figure 6. Directed Differentiations of Human iPS Cells (A) Phase-contrast image of differentiated iPS cells after 18 days cultivation on PA6. (B) Immunocytochemistry of the cells shown in (A) with bIII-tubulin (red) and tyrosine hydroxylase (green) antibodies. Nuclei were stained with Hoechst 33342 (blue). (C) RT-PCR analyses of dopaminergic neuron markers. (D) Phase-contrast image of iPS cells differentiated into cardiomyocytes. (E) RT-PCR analyses of cardiomyocyte markers. Bars = 200 mm (A and D) and 100 mm (B). * Figure 7. Teratoma Derived from Human iPS Cells. Hematoxylin and eosin staining of teratoma derived from iPS cells (clone 201B7). Cells were transplanted subcutaneously into four parts of a SCID mouse. A tumor developed from one injection site. * * * * * 通过造血干细胞分化而成的最早期祖细胞，即高增殖潜能集落形成单位（colony for ming unit-homotopoictic primitivc progenitor，CFU-HPP；培养28d）和长期培养启动细胞（long time culture-induced cell，LTC-IC；长期培养至少5周）可反映造血干细胞的存在及变化。当然，不做CFU-HPP和LTC-IC培养，而进行CD34+、CD33-、DR-或CD34+、Thy-l-、/Lin-等反映造血干细胞表面标志的多数流式细胞技术检测，同样可以反映造血干细胞的存在及变化，定量度也可靠。另外，培养CFU-GE MM 、pBFU-E、BFU-MK等均代表造血早期祖细胞，但不如最早期祖细胞CFU-HPP和LTC-IC那样直接地反映造血干细胞的存在及变化，而mBFU-E、CFU-GM、CFU-E等只能代表没有临床移植价值的晚期造血祖细胞。 * 二、造血干细胞的鉴别 迄今还没有公认的识别造血干细胞的形态学标准。 由于近代分化抗原研究的进展，促进了造血干细胞与造血祖细胞分离、纯化及鉴定的研究。 从早期造血祖细胞到晚期造血祖细胞，细胞表面的CD34抗原表达一直为阳性。CD34+细胞群中90%以上为造血祖细胞。造血干细胞的表面抗原还有Thy-1、c-kit以及Sca-1、Sca-2等。 * 在造血干细胞分化初期，最早期祖细胞出现了CD33、CD38、HLA-DR以及CD45RO等分化抗原。 在造血祖细胞定向分化时