Monocyte Subsets with High Osteoclastogenic Potential and Their Epigenetic Regulation Orchestrated by IRF8 [RNA-Seq sorted cells]

Currently, it is unclear if all monocyte subsets exhibit osteoclastogenic potential. Furthermore, the role of lineage determining TFs in regulating OC differentiation on a genome-wide scale remains poorly understood. In this study, we utilized a novel Irf8 conditional knockout (Irf8 cKO) mouse model to characterize the importance of IRF8 in OC progenitor development and epigenetic regulation of OC differentiation. To identify global transcriptional program governing the osteoclastogenic potential of Ly6Chi, Ly6Cint, and Ly6C– monocytes, and how it may be further influenced by IRF8 deficiency, we performed RNA-seq on sorted Ly6Chi, Ly6Cint, and Ly6C– cells from WT and Irf8 cKO mice. Cells were analyzed prior to (BMMs) and 4 days after RANKL stimulation (OCs). Here we show that WT Ly6Chi and Ly6Cint monocytes developmentally contain OC-specific transcripts, which are augmented upon RANKL stimulation. Additionally, in the absence of IRF8, OC-specific transcripts in all three monocyte subsets are primed to robustly respond to RANKL stimulation. Overall design: We performed RNA-seq on sorted Ly6Chi, Ly6Cint, and Ly6C– cells from WT and Irf8 cKO mice. Results include RNA-seq analysis of transcriptional changes in BMMs stimulated with MCSF (day 0) and OCs stimulated with MCSF plus RANKL (day 4). Both male and female mice and cells obtained from both genders were analyzed in this study.