Early and Delayed STAT1-Dependent Responses Drive Local Trained Immunity of Macrophages in the Spleen [bulk_RNA_seq_MS4A3]

Trained immunity (TI) is the process wherein innate immune cells gain functional memory upon exposure to specific ligands or pathogens, leading to augmented inflammatory responses and pathogen clearance upon secondary exposure. While the differentiation of hematopoietic stem cells (HSCs) and reprogramming of bone marrow progenitors are well-established mechanisms underpinning durable TI protection, the characteristics of effector differentiated cells within the tissue remains underexplored. In this study, we delve into the nature of TI in splenic myeloid cells, both recruited and local, responsible for tissue mediated protection. Our findings emphasize the centrality of the JAK-STAT1 pathway, activated by the presence of BCG in peripheral immune tissues, in driving TI. Utilizing single cell RNA-sequencing and flow cytometry, we discerned STAT1-regulated genes in TI-associated resident and recruited splenic myeloid populations. The temporal dynamics of TI were further elucidated, revealed both early and late classical and non-classical monocyte subsets with time-dependent, STAT1-specific signatures. Tissue-resident red pulp macrophages (RPM), initially depleted by BCG exposure, are restored from both a pool of tissue-trained, self-renewing macrophages and from bone marrow-derived progenitors, fostering long lasting local defense. Lastly, upon transient application of specific JAK-STAT inhibitors acting upstream of STAT1 activation, we were able to pinpoint a pivotal temporal window soon after BCG vaccination that anticipates the onset of training; the interference of which obstructs the TI phenotype. As a whole, our study unravels the diverse manifestations of TI as it emerges in mature immune cells, demonstrating its role within the functional tissue milieu of the spleen.