For many years, HSCs were viewed as unbiased cells that initiated haematopoietic development and specialization. Enabled by new technologies, our understanding of haematopoiesis has been reframed to include the possibility that developmental biases may be present even in these most undifferentiated cells, such as with megakaryocyte-biased HSCs9,25. However, the mechanisms responsible for this early biasing or specialization are not clear26,27. Here, we propose that the traditional view of HSC residency in the BM should be reconsidered to include other tissues, such as the lung. Indeed, and remarkably, we found an equal frequency of HSPCs residing in the bone marrow and lung. This was surprising, especially given the absence of a distinct HSPC population despite extensive molecular profiling of the adult human lung16. We used approaches to enrich for the presence of HSPCs in our studies, which were not done in previous studies and likely enabled our detection of this rare subset of cells amongst the >30 different lung cell types.
It is clear from our studies that the lung HSPCs have unique features compared to their medullary counterparts, but also to HSPCs found at other extramedullary sites (peripheral blood and spleen28). Chief amongst these, and perhaps obvious from our understanding of lung biology, is that lung HSPCs, like spleen HSPCs, are less active in terms of cell division and the production of more differentiated and specialized hematopoietic cells than their BM counterparts. These results from in vitro experiments would suggest that perhaps lung HSPCs function as a reserve pool that could be mobilized in the setting of haematopoietic stress. Indeed, our xenotransplantation experiments indicated that lung HSPCs performed similarly to medullary HSPCs, at least in the short-term, when given the challenge of engraftment after sublethal irradiation. Our transcriptomics analysis revealed a distinct molecular program in lung HSPCs that was further distinguishing compared to medullary HSPCs. We found clear megakaryocyte biasing of lung vs. medullary HSPCs indicating that perhaps the lung is a source of these biased progenitors. This is an interesting finding given (1) the role of the lung in platelet biogenesis and (2) the presence of tissue resident immune-like megakaryocytes (of unclear ontogeny) in the lung7. We further found an erythroid bias of lung HSPCs, akin to that reported in peripheral blood and spleen HSPCs28. Erythroid bias therefore is a shared feature of steady-state extramedullary HSPCs, perhaps determined by distinct access to environmental oxygen versus the relatively hypoxic environment of the BM29.
Our spatial transcriptomic studies were essential in confirming the presence of HSPCs in the lung and defining their precise locations and niche. Given the prevailing dogma that HSCs widely circulate30-32, it was important to rule out that blood contamination was producing our results, although our results in mice indicated an extravascular location. We found a few intravascular HSPCs, confirming previous studies, but the vast majority were extravascular and predominately in vascular-rich zones of the lung alveoli. This anatomic location in the lung is similar to the location of HSPCs in the BM, which are closely positioned next to the vascular sinusoids33,34. In the lung, this positioning could be important for seeding of the lung with circulating HSCs and potentially also for the exiting the lung during haematopoietic stress. In this niche, it was notable that lung fibroblasts were in close proximity. The lung mesenchyme is well known to be a critical niche supporting epithelial cell development and repair and similar mechanisms could be operable influencing lung HSPCs35,36. Also, we identified a developmental trajectory between lung HSPCs and lung stromal cells. Subpopulations of lung fibroblasts are known to be CD34+ and previous lineage-tracing experiments in pulmonary fibrosis have shown a hematopoietic contribution to fibroblasts37,38. Future studies are needed in this area to understand how lung HSPCs could be involved in fibrotic lung diseases.
We have not addressed the ontogeny of lung HSPCs—something not possible given the restraints of our human tissue study. There is irrefutable evidence that HSCs commonly enter the circulation and microcirculatory beds, and given the functional and molecular similarities of lung HSPCs to other extramedullary HSPCs, this is perhaps the source of the tissue resident HSPCs in the lung31,39. There is precedence, however, for tissue residency to be endowed during development, such as with the yolk-sac derived macrophages40. Future studies will be needed to answer this question in mice, including the possibility that hemogenic endothelium in the lung could be the source.
Our findings reframe our understanding of the HSPC pool and its molecular diversity and should enable future studies that could potentially lead to therapeutic advances, such as for life-saving HSC transplantation for BM malignancies and failure. In the modern era, transplantation is mainly accomplished using mobilized HSCs obtained from the peripheral blood. We propose that these mobilized cells are sourced from diverse tissues, including the lung, and that the composition of this pool may be functionally heterogeneous, which could have important implications for treatment responses and complications. In this regard, our findings may also help to understand the mechanisms of leukemogenesis with the possibility that lung HSPCs are direct targets of environmental carcinogens41. Further, our findings add to our expanding understanding of rare cell types in the lung and their potential functions42, including the possibility of a lineage relationship with lung stromal cells that could be important in fibrosing lung conditions.