A peninsular structure coordinates asynchronous differentiation with morphogenesis to generate pancreatic islets

This scRNA-seq data is an integral part of a manuscript with the above title. Using computational methods, we were able to reconstruct a detailed branched trajectory reflecting pancreatic endocrine differentiation in the mouse embryo. Analysis of the transcriptional changes occuring during the differentiation suggested that epithelial-to-mesenchymal transition likely plays no role in this process, contrary to the prevailing dogma. Our findings were corroborated with high-resolution imaging of the developing pancreas, revealing how differentiating endocrine progenitors migrate in cohesion, forming bud-like islet precursors, or "peninsulas", and that spatiotemporal collinearity during differentiation leads to the typical core-mantle architecture of the mature, spherical islet. This work led to a complete overhaul of our understanding of how pancreatic islets are developed, laying the ground for the generation of entire islets in vitro as a potential novel source of islet transplantation. Single-cell suspensions were prepared from pancreata of Neurogenin 3-eGFP mouse embryos sacrificed at different days of embryonic development. Single eGFP-positive cells were FACS-sorted into 96-well plates, and single-cell cDNA was prepared using the SMART-seq protocol. Single-cell sequencing libraries were generated using the Nextera XT DNA library preparation kit and sequenced on an Illumina HiSeq sequencer. Reads were aligned to the mouse reference genome build mm10 with TopHat, and single-cell gene expression profiles were computed using Cufflinks.本单细胞RNA测序（single-cell RNA sequencing, scRNA-seq）数据集为本标题对应手稿的核心组成部分。本研究通过计算方法，成功重构出反映小鼠胚胎胰腺内分泌细胞分化过程的精细分支轨迹。对分化过程中转录动态变化的分析显示，上皮间质转化（epithelial-to-mesenchymal transition）大概率不参与该过程，这与当前主流学术共识相悖。我们的研究结果经发育中胰腺的高分辨率成像实验验证：分化中的内分泌祖细胞以黏附方式集体迁移，形成芽状胰岛前体，即所谓“半岛”结构；分化过程中的时空共线性特征，最终造就了成熟球形胰岛典型的核心-包膜结构。本研究彻底革新了学界对胰岛发育机制的认知，为体外构建完整胰岛以作为胰岛移植的新型潜在来源奠定了坚实基础。 实验流程如下：我们从不同胚胎发育天数处死的Neurogenin 3-增强型绿色荧光蛋白（Neurogenin 3-eGFP）小鼠胚胎的胰腺组织中制备单细胞悬液；通过荧光激活细胞分选（Fluorescence-Activated Cell Sorting, FACS）将eGFP阳性单细胞分选至96孔板内，并采用SMART-seq技术制备单细胞cDNA；使用Nextera XT DNA文库制备试剂盒构建单细胞测序文库，随后在Illumina HiSeq测序仪上完成高通量测序；利用TopHat将测序读数比对至小鼠参考基因组mm10版本，并通过Cufflinks计算得到单细胞基因表达谱。