A multi-omic atlas of human embryonic skeletal development
Nature volume 635, pages 657–667 (2024 )Cite this article
Human embryonic bone and joint formation is determined by coordinated differentiation of progenitors in the nascent skeleton. The cell states, epigenetic processes and key regulatory factors that underlie lineage commitment of these cells remain elusive. Here we applied paired transcriptional and epigenetic profiling of approximately 336,000 nucleus droplets and spatial transcriptomics to establish a multi-omic atlas of human embryonic joint and cranium development between 5 and 11 weeks after conception. Using combined modelling of transcriptional and epigenetic data, we characterized regionally distinct limb and cranial osteoprogenitor trajectories across the embryonic skeleton and further described regulatory networks that govern intramembranous and endochondral ossification. Spatial localization of cell clusters in our in situ sequencing data using a new tool, ISS-Patcher, revealed mechanisms of progenitor zonation during bone and joint formation. Through trajectory analysis, we predicted potential non-canonical cellular origins for human chondrocytes from Schwann cells. We also introduce SNP2Cell, a tool to link cell-type-specific regulatory networks to polygenic traits such as osteoarthritis. Using osteolineage trajectories characterized here, we simulated in silico perturbations of genes that cause monogenic craniosynostosis and implicate potential cell states and disease mechanisms. This work forms a detailed and dynamic regulatory atlas of bone and cartilage maturation and advances our fundamental understanding of cell-fate determination in human skeletal development.
Human bone development begins between 6 and 8 weeks after conception (post-conception weeks, PCW) during the transition from embryonic to fetal stages. In the cranium, calvarial progenitors differentiate into osteoblasts through intramembranous ossification and continue to house osteoprogenitors postnatally1,2. In the nascent synovial joint, an interzone condensation appears in the limb bud at 5–6 PCW3 and forms a joint cavity between 7 and 8 PCW, varying in timing across joints, within which fibrous and ligamentous structures develop4,5 (Fig. 1b,c). Cartilage scaffolds form on either side of synovial joints to facilitate development of the body plane until they are replaced by bone tissue as endochondral ossification ensues from 8 PCW6,7. These regionally distinct modes of ossification govern osteogenesis throughout the human skeleton. To our knowledge, the cellular basis by which they form and mature remain incompletely described in human development at single-cell resolution. To address this, we applied single-nucleus paired RNA (snRNA) and assay for transposase-accessible chromatin (snATAC) sequencing (seq), and spatial methods, to decipher the regulatory landscape that mediates maturation of the distinct bone-forming and joint-forming niches in the embryonic cranium and limbs from 5 to 11 PCW. Through this, we uncovered previously undescribed cellular diversity in the osteogenic and chondrogenic lineages. We developed ISS-Patcher, a tool to impute cell labels from the droplet data on our high-resolution 155-plex in situ sequencing (ISS) datasets, which facilitated insights into spatially defined niches within the embryonic synovial joint. Applying OrganAxis8, a new spatial transcriptomics annotation tool, we also define the spatial trajectory of the developing cranial bone. We characterized novel cell states of the craniofacial region and additionally delineated processes of human Schwann cell and fibroblast development. Our resource and new computational toolset, including SNP2Cell, enabled predictions of the mechanisms of developmental conditions, such as craniosynostosis9,10,11, and allowed association of gene-regulatory networks (GRNs) in region-specific mesenchymal clusters to ageing diseases of the human skeleton, such as osteoarthritis12,13.
