Identification of gene expression profile of neural crest-derived cells isolated from submandibular glands of adult mice.
Identification of gene expression profile of neural crest-derived cells isolated from submandibular glands of adult mice.
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鉴定从成年小鼠下颌下腺分离的神经嵴衍生细胞的基因表达谱。
DOI：
10.1016/j.bbrc.2014.02.130
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发表时间：
2014
期刊：
影响因子：
--
通讯作者：
Kamijo R
中科院分区：
文献类型：
--
作者：
Takahashi M;Suzawa T;Yamada A;Yamaguchi T;Mishima K;Osumi N;Maki K;Kamijo R
胚胎中的神经嵴细胞作为神经嵴衍生细胞（NCDC）迁移到靶位点，在那里它们分化成各种衍生物。一些NCDC在动物的整个生命过程中保持未分化状态，被认为是再生医学的有用细胞来源。然而，没有建立的方法，以获得足够的再生医学从成人具有高纯度，因为它们在成人组织中的分布还没有完全了解。在成人中鉴定可靠的NCDC标记物是至关重要的，因为最可靠的NCDC标记物P0和Wnt 1的表达在胚胎阶段被关闭。为了分析成年组织中NCDC的特征，我们利用双转基因小鼠品系P0-Cre/CAG-CAT-EGFP转基因小鼠（P0小鼠），其中NCDC显示表达EGFP，并且我们能够识别其中的GFP阳性细胞。我们专注于颌下腺（SMG），这是已知的来自神经嵴。GFP阳性细胞呈岛状散在分布于成年P0小鼠的SMG中。我们通过手术从成年小鼠中取出SMG，并将样品消化成单细胞悬液。用流式细胞术分离的GFP阳性细胞表达高水平的Sox 10，一种胚胎神经嵴细胞的标志物，表明成功分离NCDC。为了确定候选标记基因在孤立的NCDC，我们进行了DNA微阵列分析和实时荧光PCR分析的GFP阳性和阴性细胞分离P0小鼠，然后选择基因显示差异基因表达模式。与GFP阴性细胞相比，GFP阳性细胞表达Gpr 4和Ednrb的水平较高，而Pdgfra和Pdgfrb的表达水平较低。此外，DNA微阵列分析显示GFP阳性细胞对腺泡细胞的标志物水通道蛋白5呈阳性。总之，我们的研究结果表明，成年SMG中的NCDC具有特征性的基因表达谱，特别是其细胞表面分子。使用这些特异性细胞表面蛋白的组合进行细胞分选将是用于从SMG中以高纯度分离NCDC的有用策略。
Neural crest cells in the embryo migrate to reach target sites as neural crest-derived cells (NCDCs) where they differentiate into a variety of derivatives. Some NCDCs are maintained in an undifferentiated state throughout the life of the animal and are considered to be a useful cell source for regenerative medicine. However, no established method to obtain NCDCs sufficient for regenerative medicine from adults with high purity has been presented, since their distribution in adult tissues is not fully understood. It is critical to identify reliable markers for NCDCs in adults, as the expressions ofP0andWnt1, the most reliable NCDC markers, are shut off in the embryonic stage. To analyze the characteristics of NCDCs in adult tissues, we utilized a double transgenic mouse strain, P0-Cre/CAG-CAT-EGFP transgenic mice (P0 mice), in which NCDCs were shown to express EGFP and we were able to recognize GFP-positive cells in those. We focused on the submandibular glands (SMGs), which are known to be derived from the neural crest. GFP-positive cells were shown to be scattered like islands in the SMGs of adult P0 mice. We surgically removed SMGs from adult mice and digested samples into single cell suspensions. GFP-positive cells separated using flow cytometry expressed a high level ofSox10, a marker of embryonic neural crest cells, suggesting successful isolation of NCDCs. To identify candidate marker genes in isolated NCDCs, we performed DNA microarray analyses and real-time PCR analysis of GFP-positive and -negative cells isolated from P0 mice, then selected genes showing differential gene expression patterns. As compared to GFP-negative cells, GFP-positive cells expressedGpr4andEdnrbat higher levels, whereasPdgfraandPdgfrbwere expressed at lower levels. Furthermore, DNA microarray analysis showed that GFP-positive cells were positive for aquaporin 5, a marker for acinar cells. Together, our results indicate that NCDCs in adult SMGs have characteristic gene expression profiles specially their cell surface molecules. Cell sorting using a combination of these specific cell surface proteins would be a useful strategy for isolation of NCDCs from SMGs with high purity.