A106-10 Spatial Topographic Atlas of the Human Trisomy 21 Lungs

Rationale Trisomy 21 (T21), resulting in Down Syndrome (DS), is the most common human chromosomal anomaly. Although DS can affect many organ systems, pulmonary complications are one of the leading causes of morbidity and mortality. Using advanced transcriptomic approaches, we have previously constructed a comprehensive cellular atlas of the fetal human lung to elucidate the molecular mechanisms associated with T21. However, a significant knowledge gap exists since scRNA-Seq in isolation does not preserve spatial information and enable studies which examine molecular alterations in the context of cell differentiation and morphogenesis. The goal of this study was to identify aberrations in the spatial organization in T21 lungs associated with altered developmental trajectories. Methods All studies and procedures were completed with IRB approval. Spatial transcriptomics data was generated using human lung tissues obtained from T21 (n = 4) and non-T21 (n = 3) prenatal lungs samples ranging in gestational age from 15 - 20 weeks, using 10X Genomics Visium platform. Sequence data demultiplexing and pre-processing was completed using CellRanger and aligned to GRCh38, and initial quality control assessment, and normalization, were performed by Squidpy and SpotClean. Data analyses, including normalization, dimensional reduction, clustering, spatially variable feature detection, differential expression assessment, and visualization were performed using Seurat. Cell type annotation and pathway analyses were performed using Toppfun. Results Unsupervised clustering of spatial transcriptomic data identified nine distinct prenatal lung tissue spot types, each comprising unique proportions of epithelial, mesenchymal, endothelial, and immune cells with corresponding transcriptional profiles (Fig. 1). Within individual spot types, differential expression analysis revealed cluster-specific gene sets ranging from 177 to 1,104 genes exhibiting significant alterations in trisomy 21 (T21) lungs compared to age-matched controls. Pseudobulk analysis across all spot types identified 82 genes (28 upregulated, 54 downregulated) showing consistent differential expression in T21 lungs. Pathway enrichment analysis of these genes indicated activation of heparin-binding and protein-folding processes, accompanied by suppression of cell cycle-related pathways, including DNA methylation, DNA replication, and meiotic recombination. Summary We have developed a preliminary spatial atlas of the prenatal human trisomy 21 lung, offering novel insights into aberrant lung development.Figure 1: Identification of Spot Clusters in Human Fetal T21 and Control Lungs Uniform Manifold Approximation and Projection (UMAP) displays the 9 unique spot clusters in T21 and control lungs. Spots are colored by the tissue type annotation presented in the UMAP display in the legend. This abstract is funded by: LungMAP Consortium