Cross-platform single cell analysis of kidney development shows stromal cells express Gdnf.
Cross-platform single cell analysis of kidney development shows stromal cells express Gdnf.
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DOI：
10.1016/j.ydbio.2017.11.006
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发表时间：
2018-02-01
影响因子：
2.7
通讯作者：
Potter SS
中科院分区：
文献类型：
--
作者：
Magella B;Adam M;Potter AS;Venkatasubramanian M;Chetal K;Hay SB;Salomonis N;Potter SS
发育中的肾脏为研究器官发生原理提供了一个有用的模型。在本报告中，我们使用三个独立的平台，即Drop - Seq、Chromium 10x Genomics和Fluidigm C1，对胚胎期14.5天的小鼠肾脏进行单细胞RNA测序（scRNA - Seq）分析。利用AltAnalyze软件，结合无监督方法ICGS，我们无法在这个活跃的肾发生阶段识别并确认16种不同细胞群的存在。采用一种新的综合有监督计算策略，我们能够成功地协调并比较所有三个技术平台的细胞图谱。对可能的跨区室受体/配体相互作用的分析确定肾发生区基质是胶质细胞源性神经营养因子（GDNF）的一个来源。这是出乎意料的，因为帽状间充质肾单位祖细胞一直被认为是GDNF的唯一来源，而GDNF是集合管系统分支形态发生的关键驱动因素。基质细胞中Gdnf的表达通过多种方式得到验证，包括Gdnf原位杂交结合肾单位祖细胞标志物SIX2以及基质细胞标志物MEIS1的免疫组织化学检测。最后，本研究中产生的单细胞基因表达图谱证实并扩展了先前的研究成果，即肾脏发育过程中存在多谱系启动。肾单位祖细胞显示出与多种潜在分化谱系相关基因的随机表达。
The developing kidney provides a useful model for study of the principles of organogenesis. In this report we use three independent platforms, Drop-Seq, Chromium 10x Genomics and Fluidigm C1, to carry out single cell RNA-Seq (scRNA-Seq) analysis of the E14.5 mouse kidney. Using the software AltAnalyze, in conjunction with the unsupervised approach ICGS, we were unable to identify and confirm the presence of 16 distinct cell populations during this stage of active nephrogenesis. Using a novel integrative supervised computational strategy, we were able to successfully harmonize and compare the cell profiles across all three technological platforms. Analysis of possible cross compartment receptor/ligand interactions identified the nephrogenic zone stroma as a source of GDNF. This was unexpected because the cap mesenchyme nephron progenitors had been thought to be the sole source of GDNF, which is a key driver of branching morphogenesis of the collecting duct system. The expression of Gdnf by stromal cells was validated in several ways, including Gdnf in situ hybridization combined with immunohistochemistry for SIX2, and marker of nephron progenitors, and MEIS1, a marker of stromal cells. Finally, the single cell gene expression profiles generated in this study confirmed and extended previous work showing the presence of multilineage priming during kidney development. Nephron progenitors showed stochastic expression of genes associated with multiple potential differentiation lineages.
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