The role of LSD1 in regulating periodontal induced bone loss - Project Summary Osteoclasts are large multinucleated cells that secrete acid and proteases to resorb bone. Understanding the molecular events that regulate osteoclast activity during periodontal disease is critical to treating bone loss during inflammatory diseases. Reversible modifications to DNA such as histone acetylation, methylation, phosphorylation and ubiquitylation alter the access of transcriptional machinery to DNA and regulate gene expression. Lysine-specific demethylase 1 (LSD1) confers transcriptional repression by demethylating H3K4 or activating transcription by demethylating H3K9. The role and targets of LSD1 during osteoclast differentiation are unknown. Results from our work with LSD1;LysM-Cre (LSD1cKO) conditional mice begins to fill this knowledge gap. LSD1cKO mice have smaller osteoclasts and increased BV/TV compared to LSD1WT mice. Bulk RNA-SEQ of femur-derived osteoclast precursors from LSD1cKO mice indicate that genes in the IFN-b pathway are upregulated compared to LSD1WT cells. Unlike those from the femur, osteoclasts generated from the LSD1cKO mandible were not significantly different in size compared to their wild type littermates suggesting a difference in skeletal site regulation by LSD1. Our data demonstrates significant bone loss in LSD1WT mice but no significant bone loss in LSD1cKO mice in a ligature induced periodontitis model. In support of this, TNF- a, an inflammatory cytokine that induces osteoclast differentiation after priming by TGF-b, was unable to induce osteoclast differentiation in mandible derived cells suggesting a mechanism for the inability of LSD1cKO mice to lose bone in ligature induced periodontal disease. The central hypothesis is that LSD1 demethylates residues at H3K4 to inhibit expression of genes involved in the IFN-b pathway thus allowing inflammatory cytokines to induce osteoclast differentiation. In the first aim we will gain mechanistic insights by conducting an in-depth investigation of ligature induced periodontitis in mandible derived osteoclast precursors from LSD1cKO mice. We will confirm our initial data that LSD1cKO mice are resistant to bone loss during ligature induced periodontitis disease through micro-CT, histology, and ELISA of various cytokines. Additionally, we will perform bulk RNA- and ATAC-SEQ of mandible-derived preosteoclasts from un-ligated and ligated LSD1WT and -cKO mice. This data will allow us to determine skeletal site-specific changes in mandible derived osteoclasts during ligature induced periodontitis. In our second aim we will determine if LSD1 expression is necessary for innate immune training resulting from ligature induced periodontitis by performing single cell sequencing of bone marrow cells from LSD1WT and LSD1cKO mice. To complement our single cell sequencing we will also perform single cell ATAC-SEQ of bone marrow cells from LSD1WT and cKO mice who have undergone innate immunity training. Epigenetic modifications are modifiable, potentially reversible and represent a promising group of treatments for periodontal disease.
