Results
1.A transcriptomic profile of the developing human spinal cord
F1a. used 10x Chromium to profile single cells and single nuclei from four samples at gestational week (GW) 17 and GW18. 10x测序孕17周和18周的细胞和细胞核
F1b. used uniform manifold approximation and projection (UMAP) dimensionality reduction to visualize and cluster cells following the integration of single cells and single nuclei. 降维聚类
F1c. found a group of cells. 细胞类型展示
F1d. A heat map showing normalized expression of the top four to six unique markers per cell type. 在两种分离技术（细胞、细胞核）下，每种细胞类型的前4-6个的高表达且唯一标记的基因marker展示。（如何分配的唯一基因？第六个基因EBF1分配给了Neurons，而在Pericycytes中更高表达，可能是该基因算不上Pericycytes的前几基因，看图可知Pericycytes的第六基因FN1比EBF1更高）
F1e. Representative immunohistochemistry images of AQP4, NeuN (RBFOX3) and MBP in a coronal thoracic spinal cord cryosection at GW19. Hoechst shows nuclei. Hoechst染色，标志细胞核DNA，对孕19周的冠状胸段脊髓冰冻切片，对AQP4、Neun(RBFOX3)和MBP的代表性免疫组织化学。AQP4是星形胶质细胞Astrocytes的marker，MBP是少突胶质细胞谱系中的细胞OPC/oligo的marker，但Neun(RBFOX3)没在F1d中。（不太了解这里选这几个基因染色的目的，是为了显示脊髓的形状？）
2.Pseudotime analysis in the OPC/oligo lineage
F2a. Highlight of the OPC/oligo lineage in the main UMAP, showing the cells selected for subclustering. 突出研究少突胶质细胞谱系（少突胶质前体细胞(OPC)/少突胶质细胞(oligo)）
F2b. Subclustering identified cell types along the oligodendrocyte lineage. 少突胶质细胞谱系中的细胞类型鉴定，从上到下应该是依次成熟的，mature oligos（MOL）就是成熟少突胶质细胞
F2c. Diffusion pseudotime analysis revealed putative origins of the trajectory in cycling cells. 拟时序分析，推测该谱系可能起源于周期性少突胶质前体细胞Cycling OPCs
F2d. grouped variable genes along this pseudotime into ten clusters. 使用k均值将基因聚为10组(啥技术？)，可能是突出基因表达的时序性
F2e. A heat map showing scaled expression of cell type dynamic genes. For each cell type the five highest-ranking genes according to velocity are shown. 每种细胞类型，显示了根据速度排名最高的五个基因（速度是个拟时序分析中重要概念，后续还要了解一下）
F2f. UMAPs showing examples of dynamic genes with cell-type-specific velocity along the trajectory showing computed velocity (top) and normalized expression (bottom). 细胞特定基因的速度(上)和表达(下)，有对应关系，应该是为了说明时序性
3.Diversity of astrocytes in the developing human spinal cord
F3a. Highlight of astroglia in the main UMAP, showing the cells selected for subclustering. 研究星型突质细胞亚群
F3b. UMAP of the astroglia subcluster colored by type. 细胞分类
F3c. representative immunohistochemistry image for the astroglia marker GFAP. 星型突质细胞marker的免疫组化，四个不同区域有不同的星型突质细胞类型
F3d. UMAP plots showing gene expression of markers associated with dorsal and ventral identities in the astroglia subcluster. 星型突质细胞背腹轴基因差异表达
F3e. UMAP plots showing gene expression of markers associated with fibrous and protoplasmic astrocyte subtypes. 第二轴——白质和灰质，或者说纤维状和原质星形胶质细胞的基因差异表达
F3f. Representative immunohistochemistry images showing the fibrous astrocyte marker CRYAB and the protoplasmic astrocyte markers GLUL and SLC1A2. 免疫组化纤维状胶质细胞marker CRYAB，原质星形胶质细胞markers GLUL、SLC1A2
F3g. Quantification of the normalized mean fluorescence intensity for CRYAB, GLUL and SLC1A2 in the spinal cord at GW19. 归一化平均荧光强度定量说明F3f
F3h. the ventral fibrous marker CD38 with the pan-astrocyte markers AQP4 and GFAP in a coronal spinal cord. 免疫组化显示腹侧纤维星形胶质细胞特异表达CD38，泛星形胶质细胞特异表达AQP4 and GFAP
F3i. A dot plot showing the expression of selected genes associated with astrocyte functions in fibrous and protoplasmic subtypes. 纤维状和原质星形胶质细胞功能基因表达具有差异，从而具有不同的生理功能；原质星形胶质细胞有较高表达的神经递质转运基因，等等一大段文献引用说明两类细胞的基因表达差异及其功能差异
4.Midline glia and mGPCs in the developing human spinal cord
F4a. Highlight of ventricular zone (VZ) cells in the astroglia subcluster, highlighting the cells selected for subclustering. 研究星形胶质细胞亚群中的脑室区(VZ)细胞，突出显示被选中进行亚群聚类的细胞
F4b. UMAP of the VZ cell subcluster, colored by cell type. 细胞分类
F4c. UMAP plots showing gene expression of markers specific to RP (GDF7 and GDF10) and FP (FOXA2 and SHH) cells. 星型胶质细胞在脑室区VZ的细胞谱系，有floor plate (FP), roof plate (RP), midplate (MP); 该图展示RP和FP的marker
F4d. expression of PPP1R17 in RP and FP cells. 磷酸酶抑制物PPP1R17是大脑皮层中间前体细胞的标志
F4e. Representative immunohistochemistry images showing PPP1R17, SOX9 and the cell cycle marker KI67 in VZ cells in a coronal upper thoracic spinal cord cryosection. 免疫组化发现PPP1R17在FP和RP细胞和中线细胞中表达（在扩展数据中，PPP1R17在小鼠脊髓脑室区细胞中有极低的表达。未来实验可确定PPP1R17是否在脊髓中发挥人类特有的作用。）；SOX9是星型胶质细胞marker；KI67表细胞增殖
5.Early MN identity specification in humans
F5a. Highlight of neurons in the main UMAP, highlighting the cells selected for subclustering. 研究脊髓神经元细胞多样性
F5b. UMAP of the neuron subcluster, colored by neurotransmitter identity. 以神经递质来分类脊髓神经元细胞
F5c. A bar plot showing the proportion of cells within the neuron subcluster in each neurotransmitter identity group. 细胞比例
F5d. Bubble plot showing expression of neurotransmitter identity-related genes as average scaled expression per group. 上述分类细胞相关高表达基因
F5e. Representative immunohistochemistry images showing neurons with different neurotransmitter identities. 免疫组化标志细胞位置，CHAT 标示胆碱能神经元，GLYT2（SLC6A5）标示甘氨酸能神经元，GABA 标示 GABA 能神经元，VGLUT2（SLC17A6）标示谷氨酸能神经元
F5f. used label transfer to compare our cell types with a harmonized atlas of cell types in the mouse spinal cord. 使用标记转移将人类脊髓细胞类型与小鼠脊髓细胞类型的协调图谱进行比较，存在不同类型的兴奋性、抑制性和胆碱能神经元，以及背侧、腹侧和中间神经元
F5g. Highlight of skeletal and visceral MNs in the neuron subcluster. 两种胆碱能神经元，即支配骨骼运动和内脏运动的神经元motor neuron (MN)
F5h. MN subcluster colored by identity. 运动神经元聚类，其中骨骼MN由两个亚群组成
F5i. Using label transfer of MNs from adult mouse spinal cord. 对成年小鼠脊髓MN的标记转移，人类内脏MN定位于小鼠的内脏MN，而骨骼MN分为α和γ亚群
F5j. Bar plot showing the percent of predicted identities in the MN subcluster (this study) on the basis of annotations from Blum et al. 借助小鼠的运动神经元注释，预测该研究中人类运动神经元的比例
F5k. A heat map of scaled gene expression across cells in the MN subcluster. 运动神经元中高表达基因热图
F5l. Representative in situ hybridization of CHAT, CHODL and NDNF in coronal lumbar spinal cord cryosections at GW18. CHODL是α-MN的特异高表达基因，NDNF是γ-MN的特异高表达基因，具有比较明显的分离，而CHAT与这两类细胞有一定的共定位，说明在孕18周的时候这两种神经元就分离了
6.Mapping disease-related genes
F6a. A heat map (left) showing the sum of gene counts normalized to total counts per cell cluster for genes associated with myelin-related disorders and UMAP plots (right) showing gene expression of selected genes (bold). 髓鞘相关疾病基因热图与分布，髓鞘少突胶质细胞或雪旺细胞(PLP1、FA2H和GJC2)中特异表达的基因，以及一组在不同类型的星形胶质细胞(GJA1、GFAP和EDNRB)或小胶质细胞(CSF1R和TREM2)中表达的基因，这表明不同类型的细胞之间的相互作用可能在发病中起作用
F6b. A heat map (left) showing the sum of gene counts normalized to total counts per cell cluster for genes associated with Charcot-Marie-Tooth (CMT) and UMAP plots (right) showing expression of selected genes (bold). 腓骨肌萎缩症疾病基因热图与分布，OPC/少突胶质细胞谱系或雪旺细胞(MPZ和PMP22)、星形胶质细胞(FGD4)和小胶质细胞(EGR2)中表达的基因，NFOL中的FA2H，MFOL中的HSPB1和COP中的GNB4
F6c. A heat map (left) showing the sum of gene counts normalized to total counts per cell cluster for genes associated with Amyotrophic Lateral Sclerosis (ALS) and UMAP plots (right) showing expression of selected genes (bold). 肌萎缩性侧索硬化症ALS相关的基因的表达，SOD1普遍表达；MAPT在神经元高表达；SQSTM1在VLMC，周细胞和小胶质细胞高表达；CENPV在OPC/少突胶质细胞高表达
7.Developing human spinal cord dataset integration
F7a. A schematic showing all the human developing spinal cord samples included in the dataset integration. 数据集与另外两个正在开发的人类脊髓数据集进行了整合。整合后的数据集包括跨越22 周的35个样本，经过质量控制(QC)和筛选后，总共有783408个细胞和166807细胞核
F7b. UMAP representation of integrated datasets. 整合图谱
F7c. Bar plot showing the percent of cells within the OPC/oligo lineage over gestational time. 拟时序分析，量化OPC/少突胶质细胞谱系随时间的细胞比例
embryonic (GW4–9), early fetal 1 (GW10–11), early fetal 2 (GW12–14), early mid-fetal 1 (GW15–17), early mid-fetal 2 (GW18–19) and late mid-fetal (GW20–25).
F7d. A bar plot showing the percent of cells within the astroglia subcluster over gestational time. 星形胶质细胞谱系的比例变化，前体星形胶质细胞群(例如周期、FP、RP和MP)的百分比减少，原质星形胶质细胞在发育早期的星形胶质细胞中所占比例更高，而纤维性星形胶质细胞在胎儿中期后期似乎更丰富
F7e. Representative immunohistochemistry images showing the neuronal marker NeuN, the astrocyte marker AQP4 and the myelinating oligo marker MBP in GW11 spinal cord cryosections. 免疫组化标记定位神经元细胞（NeuN），星形胶质细胞（AQP4），髓鞘少突胶质细胞（MBP）
F7f. Representative immunohistochemistry images showing the fibrous and protoplasmic astrocyte markers CRYAB and GLUL in GW11 spinal cord cryosections. 免疫组化标记定位纤维胶质细胞（CRYAB）和原质星形胶质细胞(GLUL)；结合上图表示，胎儿早期(GW11)，星形胶质细胞（AQP4）中的原质星形胶质细胞(GLUL)是要早于纤维胶质细胞（CRYAB）产生
F7g. Bar plots showing the percent of cells within the OPC/oligo lineage over gestational time and separated in cervical (C), thoracic (T) and lumbar (L) regions of the spinal cord. 研究吻尾轴的发育动力学，对颈部、胸部和腰部的样本进行分析，发现在OPC/少突胶质细胞谱系中没有发现明显的区域特异性分布
F7h. Bar plots showing the percent of cells within the astroglia subcluster over gestational time and separated in cervical (C), thoracic (T) and lumbar (L) regions. 在星形胶质细胞谱系中，我们发现与颈部和腰部相比，胸部区域的原质星形胶质细胞比例较低
F7i. A curve plot showing the cumulative frequency of alpha (α) and gamma (γ) MNs over time in the integrated dataset. 在发育早期，没有看到骨骼运动神经元的明显数量差异
