Rakeshlal Kapuge

Cardiovascular diseases often arise in anatomically specialised regions of the heart that remain poorly represented in existing single-cell and spatial atlas references. Here we profiled 28 regions of the adult human heart from 36 donors, combining paired single-nucleus RNA and chromatin accessibility assays with spatial transcriptomics. The resulting atlas integrates 1,059,175 expression profiles and 452,614 chromatin accessibility profiles, and defines spatial niches across the heart. In cardiac valves, we identify a valve fibroblast–macrophage niche that is polarised to the non-fibrosa surface and expands with age. Aortic stenosis genetic risk was enriched in inflammatory regulatory programmes of the Valve Fibroblast Immune cell state. In pulmonary veins, we define PITX ⁺ Myocardial Sleeve Cells, a region-restricted cardiomyocyte population in which a fine-mapped 4q25 atrial fibrillation variant intersects a cell type-specific open chromatin region. In coronary arteries, we resolve a RUNX1 -marked synthetic smooth muscle layer in the subintima that provides a molecular identity for diffuse intimal thickening, a constitutive feature of the healthy vessel wall and a substrate for atherosclerosis, and show by perturbation in human smooth muscle cells that RUNX1 promotes cell-cycle entry while restraining inflammatory cytokine gene expression. Finally, allele-specific accessibility together with deep learning models fine-tuned to predict regulatory activity from DNA sequence identify cell type-specific regulatory effects at GWAS loci for atrial fibrillation, coronary artery disease and calcific aortic valve stenosis, including a variant whose Valve Fibroblast-specific effect on the polyamine transporter ATP13A3 is opposite in direction to the effect seen in bulk-tissue eQTL references that lack valve cells. Together with these cardiac sequence models and CardioSleuth, an AI-powered exploration engine for cardiac regulatory effects, this atlas links cardiac anatomy, gene regulation and disease genetics at single-cell resolution. By placing genetic risk in its precise cellular and anatomical context, it provides a foundation for mechanistic disease understanding, cell-state-specific therapeutic target nomination, and improved interpretation of inherited cardiovascular risk to inform future therapy and management.

Hormones act across tissues and organs to coordinate physiological functions. Drawing inspiration from the Human Cell Atlas, we analyzed expression of 379 hormone and receptor genes in a transcriptomic dataset comprising 14 million single cells and nuclei across 47 human tissues. Using hormone2cell, we mapped putative hormone-producing and hormone-receiving cell types, defining tissue-specific and cross-tissue endocrine signatures. We predicted non-classical sites of hormone expression, including secretin in plasmacytoid dendritic cells, inferred convergent hormone action and endocrine feedback loops, and implicated cell populations in monogenic endocrine disorders. In a cross-tissue integration of adipocyte datasets, we uncovered dynamic endocrine programs across depots, within adipocyte subtypes and through adipogenic differentiation. Cumulatively, the Hormone Cell Atlas ( hormonecellatlas.org.uk ) provides a comprehensive framework for dissecting hormonal impact on health and disease.

The gastrointestinal tract is a multi-organ system crucial for efficient nutrient uptake and barrier immunity. Advances in genomics and a surge in gastrointestinal diseases^1,2 has fuelled efforts to catalogue cells constituting gastrointestinal tissues in health and disease³. Here we present systematic integration of 25 single-cell RNA sequencing datasets spanning the entire healthy gastrointestinal tract in development and in adulthood. We uniformly processed 385 samples from 189 healthy controls using a newly developed automated quality control approach (scAutoQC), leading to a healthy reference atlas with approximately 1.1 million cells and 136 fine-grained cell states. We anchor 12 gastrointestinal disease datasets spanning gastrointestinal cancers, coeliac disease, ulcerative colitis and Crohn’s disease to this reference. Utilizing this 1.6 million cell resource (gutcellatlas.org), we discover epithelial cell metaplasia originating from stem cells in intestinal inflammatory diseases with transcriptional similarity to cells found in pyloric and Brunner’s glands. Although previously linked to mucosal healing⁴, we now implicate pyloric gland metaplastic cells in inflammation through recruitment of immune cells including T cells and neutrophils. Overall, we describe inflammation-induced changes in stem cells that alter mucosal tissue architecture and promote further inflammation, a concept applicable to other tissues and diseases.

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.

The gastrointestinal (GI) tract consists of connected organs, from the oral cavity to rectum, which function to ensure efficient nutrient uptake and barrier immunity. Diseases of the GI tract affect millions worldwide and as such there are now over 25 published single cell RNA-sequencing (scRNAseq) datasets surveying the GI tract, profiling specific anatomical regions, cell lineages, ages and diseases. To consolidate these efforts, we harmonised and integrated scRNAseq datasets across the whole GI tract from developing and adult human tissues, as well as newly generated data from preterm gut. We uniformly processed 385 samples from 189 healthy controls using a newly developed automated QC approach (scAutoQC). In total, our healthy reference contains ~1.1 million cells which we annotated to a total of 137 fine-grained cell states. We anchor 13 published and 1 unpublished GI disease datasets covering gastric and colorectal (CRC) cancers, celiac disease, ulcerative colitis (UC) and Crohn's disease (CD) to this reference, taking our atlas to a total of 1.6 million cells. We provide our atlas as a valuable resource to the community (available at gutcellatlas.org). Using this resource, we discover epithelial cell metaplasia arising from stem cells across intestinal inflammatory diseases (celiac, UC and CD) and CRC with transcriptional similarity to cells of the gastric and Brunner's glands. Whilst previously linked to mucosal healing, we now implicate these cells in inflammation through recruitment of immune cells including T cells and neutrophils, and through direct interactions with T cells. Overall, we discover a shift in paradigm whereby changes in stem cells during inflammation lead to altered mucosal tissue architecture, which in turn contributes to ongoing inflammation. These findings highlight that in addition to barrier function, epithelial cells actively contribute to progression of inflammation which may be a function applicable to other tissues and diseases.

The COVID-19 pandemic is an ongoing global health threat, yet our understanding of the dynamics of early cellular responses to this disease remains limited¹. Here in our SARS-CoV-2 human challenge study, we used single-cell multi-omics profiling of nasopharyngeal swabs and blood to temporally resolve abortive, transient and sustained infections in seronegative individuals challenged with pre-Alpha SARS-CoV-2. Our analyses revealed rapid changes in cell-type proportions and dozens of highly dynamic cellular response states in epithelial and immune cells associated with specific time points and infection status. We observed that the interferon response in blood preceded the nasopharyngeal response. Moreover, nasopharyngeal immune infiltration occurred early in samples from individuals with only transient infection and later in samples from individuals with sustained infection. High expression of HLA-DQA2 before inoculation was associated with preventing sustained infection. Ciliated cells showed multiple immune responses and were most permissive for viral replication, whereas nasopharyngeal T cells and macrophages were infected non-productively. We resolved 54 T cell states, including acutely activated T cells that clonally expanded while carrying convergent SARS-CoV-2 motifs. Our new computational pipeline Cell2TCR identifies activated antigen-responding T cells based on a gene expression signature and clusters these into clonotype groups and motifs. Overall, our detailed time series data can serve as a Rosetta stone for epithelial and immune cell responses and reveals early dynamic responses associated with protection against infection.

The function of a cell is defined by its intrinsic characteristics and its niche: the tissue microenvironment in which it dwells. Here we combine single-cell and spatial transcriptomics data to discover cellular niches within eight regions of the human heart. We map cells to microanatomical locations and integrate knowledge-based and unsupervised structural annotations. We also profile the cells of the human cardiac conduction system¹. The results revealed their distinctive repertoire of ion channels, G-protein-coupled receptors (GPCRs) and regulatory networks, and implicated FOXP2 in the pacemaker phenotype. We show that the sinoatrial node is compartmentalized, with a core of pacemaker cells, fibroblasts and glial cells supporting glutamatergic signalling. Using a custom CellPhoneDB.org module, we identify trans-synaptic pacemaker cell interactions with glia. We introduce a druggable target prediction tool, drug2cell, which leverages single-cell profiles and drug–target interactions to provide mechanistic insights into the chronotropic effects of drugs, including GLP-1 analogues. In the epicardium, we show enrichment of both IgG⁺ and IgA⁺ plasma cells forming immune niches that may contribute to infection defence. Overall, we provide new clarity to cardiac electro-anatomy and immunology, and our suite of computational approaches can be applied to other tissues and organs.

The COVID-19 pandemic is an ongoing global health threat, yet our understanding of the cellular disease dynamics remains limited. In our unique COVID-19 human challenge study we used single cell genomics of nasopharyngeal swabs and blood to temporally resolve abortive, transient and sustained infections in 16 seronegative individuals challenged with preAlpha-SARS-CoV-2. Our analyses revealed rapid changes in cell type proportions and dozens of highly dynamic cellular response states in epithelial and immune cells associated with specific timepoints or infection status. We observed that the interferon response in blood precedes the nasopharynx, and that nasopharyngeal immune infiltration occurred early in transient but later in sustained infection, and thus correlated with preventing sustained infection. Ciliated cells showed an acute response phase, upregulated MHC class II while infected, and were most permissive for viral replication, whilst nasal T cells and macrophages were infected non-productively. We resolve 54 T cell states, including acutely activated T cells that clonally expanded while carrying convergent SARS-CoV-2 motifs. Our novel computational pipeline (Cell2TCR) identifies activated antigen-responding clonotype groups and motifs in any dataset. Together, we show that our detailed time series data ( covid19cellatlas.org ) can serve as a “Rosetta stone” for the epithelial and immune cell responses, and reveals early dynamic responses associated with protection from infection.