Polyclonal regeneration of mouse bone marrow endothelial cells after irradiative conditioning

Bone marrow endothelial cells (BM-ECs) are the essential components of the BM niche and support the function of hematopoietic stem cells (HSCs). However, conditioning for HSC transplantation causes damage to the recipients’ BM-ECs and may lead to transplantation-related morbidity. Here, we investigated the cellular and clonal mechanisms of BM-EC regeneration after irradiative conditioning. Using single-cell RNA sequencing, imaging, and flow cytometry, we revealed how the heterogeneous pool of BM-ECs changes during regeneration from irradiation stress. Next, we developed a single-cell in vitro clonogenic assay and demonstrated that all EC fractions hold a high potential to reenter the cell cycle and form vessel-like structures. Finally, we used Rainbow mice and a machine-learning-based model to show that the regeneration of BM-ECs after irradiation is mostly polyclonal and driven by the broad fraction of BM-ECs; however, the cell output among clones varies at later stages of regeneration.