Multiscale analysis and functional validation of the cellular and genetic determinants of skeletal disease [Mouse scRNA-seq]

Musculoskeletal diseases are a major global health burden. Development of new bone-active therapies is hindered by limited understanding of the complex interactions between the cells and genes that regulate the skeleton. To unravel this complexity, we systematically annotated all cells in bone and defined the genes that control their function at single-cell resolution. Integration with data from human gene-mapping studies of rare skeletal disorders and common skeletal disease traits identified novel disease genes, which we validated by functional analysis in more than one thousand genetic mouse models. This multiscale approach expands the repertoire of cells that regulate bone to include endothelial and vascular smooth muscle cells. This also revealed hundreds of skeletal disease-associated genes in this landscape of cells as potential drug targets. The cellular and genetic mechanisms revealed by this approach overcomes knowledge gaps and helps to accelerate development of next generation therapies to treat skeletal diseases. Overall design: Single-cell RNA sequencing was performed on bone microenvironment cells isolated from endosteal compartments of the metaphysis and diaphysis, as well as from bone marrow from femurs of adult male C57BL/6J mice. Cells from each of the 3 bone compartments were pooled into 5 replicates (5 mice per replicate) prior to sequencing, resulting in 15 samples total. One sample (Marrow, replicate 2) failed capture.