MYCN-transformed neuroblasts from TH-MYCN mice are selectively isolated and maintained as spheres in vitro. Mus musculus

It has been already reported that there are undifferentiated/proliferating neuroblasts in the postnatal sympathetic ganglia in TH-MYCN mice. We established suitable spheroid culture condition that selectively isolates undifferentiated neuroblasts from superior mesenteric ganligon (SMG) of TH-MYCN mice. In the present study, in order to investigate the transcriptomic differences among 3-week-old WT SMG, TH-MYCN (hemizygote) SMG, and TH-MYCN (hemizygote) spheres, we carried out microarray gene expression analysis. We investigated whether or not undifferentiated cells observed in TH-MYCN SMG were selectively isolated and maintaned as spheres. Overall design: SMG tissue were dissected from 3-week-old WT and TH-MYCN (hemizygote) mice (biological replicate is eight). Primary spheres were cultured from 3-week-old TH-MYCN (hemizygote) mice (biological replicate is eight). And the primary spheres were passaged 5 times (biological replicate is four, and array replicate is two for each biological replicate). They were subjected to micorarray analyis.

已有研究报道，在TH-MYCN小鼠（TH-MYCN mouse）的出生后交感神经节中存在未分化/增殖性成神经细胞。本研究建立了适宜的球体培养体系，可从TH-MYCN小鼠的肠系膜上神经节（superior mesenteric ganglion, SMG）中选择性分离未分化成神经细胞。为探究3周龄野生型（wild type, WT）肠系膜上神经节、TH-MYCN杂合子小鼠肠系膜上神经节，以及TH-MYCN杂合子小鼠来源的球体这三类样本间的转录组差异，本研究开展了基因芯片表达谱分析，旨在明确TH-MYCN小鼠肠系膜上神经节中观察到的未分化细胞是否可被选择性分离并以球体形式稳定维持培养。实验整体设计如下：从3周龄野生型与TH-MYCN杂合子小鼠体内分离肠系膜上神经节组织，每组生物学重复数为8；从3周龄TH-MYCN杂合子小鼠体内培养原代球体，原代培养的生物学重复数为8；将原代球体传代5次后，取传代样本进行基因芯片分析，其中每个生物学重复对应2次芯片技术重复，共设置4个生物学重复，所有受试样本均完成基因芯片分析。