Jang Y K, Park J J, Lee M C, Yoon B H, Yang Y S, Yang S E, Kim S U
Brain Disease Research Center, Ajou University School of Medicine, Suwon, Korea.
J Neurosci Res. 2004 Feb 15;75(4):573-84. doi: 10.1002/jnr.10789.
Recent studies reporting trans-differentiation of mononucleated cells derived from human umbilical cord blood into neuronal cells aroused interest among investigators for their clinical implication and significance in regenerative medicine. In the present study, purified populations of hematopoietic stem cells were isolated via magnetic bead sorting and fluorescence-activated cell sorter (FACS) using a specific CD133 antibody, a cell type-specific marker for hematopoietic stem cells, and grown in culture in the presence of retinoic acid (RA). CD133+ hematopoietic stem cells expressed neuronal and glial phenotypes after RA treatment. RT-PCR analysis indicated that the RA treated CD133+ cells expressed mRNA transcripts for ATP-binding cassettes transporter ABCG2 (a universal stem cell marker), nestin (a specific cell type marker for neural stem cells), Musashi1 (a specific marker for neural stem cells) and RA receptors (RAR) including RAR-alpha, RAR-beta, and retinoid X receptor (RXR)-gamma. RA-treated CD133+ cells expressed mRNA transcripts for neuron-specific markers neurofilament proteins (NF-L, -M, -H) and synaptophysin as determined by RT-PCR, structural proteins characteristic of neurons including tubulin beta III and neuron specific enolase (NSE) by Western blot, and neuron-specific markers NeuN and microtubule-associated protein-2 (MAP2) by immunocytochemistry. RA-treated CD133+ cells also expressed the astrocyte-specific marker glial fibrillary acidic protein (GFAP), as demonstrated by RT-PCR, Western blot, and immunocytochemistry. In addition, RA-treated CD133+ cells expressed cell type-specific markers for oligodendrocytes including myelin basic protein (MBP) as shown by RT-PCR, proteolipid protein (PLP) by Western blot analysis, and cyclic nucleotide phosphodiesterase (CNPase) by immunostaining. Upregulated expression of several basic helix-loop-helix (bHLH) transcription factors important for early neurogenesis, including Otx2, Pax6, Wnt1, Olig2, Hash1 and NeuroD1, was also demonstrated in CD133+ cells after RA treatment. These results indicate that human cord blood-derived CD133+ hematopoietic stem cells could trans-differentiate into neural cell types of neuron-like cells, astrocytes, and oligodendrocytes by RA treatment.
最近的研究报道，源自人脐带血的单核细胞向神经元细胞的转分化因其在再生医学中的临床意义而引起了研究人员的兴趣。在本研究中，使用特异性CD133抗体（造血干细胞的细胞类型特异性标志物）通过磁珠分选和荧光激活细胞分选仪（FACS）分离纯化的造血干细胞群体，并在视黄酸（RA）存在的情况下进行培养。RA处理后，CD133 +造血干细胞表达神经元和神经胶质细胞表型。RT-PCR分析表明，经RA处理的CD133 +细胞表达ATP结合盒转运体ABCG2（一种通用干细胞标志物）、巢蛋白（神经干细胞的特异性细胞类型标志物）、Musashi1（神经干细胞的特异性标志物）以及视黄酸受体（RAR）包括RAR-α、RAR-β和类视黄醇X受体（RXR）-γ的mRNA转录本。通过RT-PCR测定，经RA处理的CD133 +细胞表达神经元特异性标志物神经丝蛋白（NF-L、-M、-H）和突触素的mRNA转录本；通过蛋白质印迹法检测到神经元特有的结构蛋白，包括微管蛋白βIII和神经元特异性烯醇化酶（NSE）；通过免疫细胞化学检测到神经元特异性标志物NeuN和微管相关蛋白-2（MAP2）。经RA处理的CD133 +细胞还表达星形胶质细胞特异性标志物胶质纤维酸性蛋白（GFAP），这通过RT-PCR、蛋白质印迹法和免疫细胞化学得到证实。此外，经RA处理的CD133 +细胞表达少突胶质细胞的细胞类型特异性标志物，包括RT-PCR显示的髓鞘碱性蛋白（MBP）、蛋白质印迹分析显示的蛋白脂蛋白（PLP）以及免疫染色显示的环核苷酸磷酸二酯酶（CNPase）。在RA处理后的CD133 +细胞中还证实了几种对早期神经发生重要的碱性螺旋-环-螺旋（bHLH）转录因子的表达上调，包括Otx2、Pax6、Wnt1、Olig2、Hash1和NeuroD1。这些结果表明，经RA处理后，源自人脐带血的CD133 +造血干细胞可转分化为神经元样细胞、星形胶质细胞和少突胶质细胞等神经细胞类型。
