Oncostatin M is a Master Regulator of an Inflammatory Network in Dnmt3a-Mutant Hematopoietic Stem Cells

Age-associated clonal hematopoiesis (CH) occurs due to somatic mutations accrued in hematopoietic stem cells (HSCs) that confer a selective advantage in the context of aging. The mechanisms by which CH-mutant HSCs gain this advantage with aging are not comprehensively understood. Using unbiased transcriptomic approaches, we identify Oncostatin M (OSM) signaling as a candidate contributor to aging-driven Dnmt3a-mutant CH. We find that Dnmt3a-mutant HSCs from young mice do not functionally respond to acute OSM stimulation with respect to proliferation, apoptosis, hematopoietic engraftment, or myeloid differentiation. However, young Dnmt3a-mutant HSCs transcriptionally upregulate an inflammatory cytokine network in response to acute OSM including genes encoding IL-6, IL-1b and TNFa. In addition, OSM-stimulated Dnmt3a-mutant HSCs upregulate the anti-inflammatory genes Socs3 and Nr4a1, creating a negative feedback loop limiting sustained activation of the inflammatory network. In the context of an aged BM microenvironment with chronically elevated levels of OSM, Dnmt3a-mutant HSCs upregulate pro-inflammatory genes but do not upregulate Socs3 and Nr4a1. Together, our work suggests that chronic inflammation with aging exhausts the regulatory mechanisms in young CH-mutant HSCs that resolve inflammatory states, and that OSM is a master regulator of an inflammatory network that contributes to age-associated CH. Overall design: We sought to test the transcriptional consequences of OSM-STAT3 signaling in a more purified control and R878H/+ HSC population. We prospectively isolated control and R878H/+ HSCs from young adult mice, stimulated ex vivo with 500ng/mL of recombinant murine OSM or vehicle control for 60min and immediately flash froze cell pellets for RNA extraction and RNA-seq. We identified significantly differentially expressed genes comparing OSM- vs. vehicle-treated control HSCs, and OSM- vs. vehicle-treated R878H/+ HSCs, using P < 0.05 and fold change (FC) ± 1.5 cutoffs. Differential gene expression of 1 hour stimulation vehicle vs. OSM in 1000 control HSCs or R878H/+ HSCs respectively. N= 6 mice per group, experiment done two independent times.