Abstract Body: Early after tissue injury, resident fibroblasts secrete pro-inflammatory factors that contribute to the inflammatory response. Several days following tissue injury, fibroblasts begin to transition to myofibroblasts and express classic myofibroblast genes (acta2, tagln, myl9, cald1) in order to contract and close wounds. In type 2 diabetes (T2D), myofibroblasts are decreased in number and function, leading to delayed wound healing. Our lab specializes in the epigenetic changes that lead to cell phenotype, and we hypothesized that reduced myofibroblast gene expression in T2D was due to differential expression of a chromatin modifying enzyme (CME). Using single cell RNA-sequencing of human T2D and non-T2D wounds, we screened multiple known CMEs and found that Mll-1, a H3K4 methyltransferase, was decreased in T2D fibroblasts compared to non-T2D fibroblasts. Next, we treated in-vitro wound fibroblasts with a small molecule inhibitor of MLL-1, which reduced myofibroblast gene expression. Further, murine wounds injected with the inhibitor of MLL-1 healed worse than sham injections. We developed a tamoxifen inducible, fibroblast-specific knockout of Mll-1 (Mll-1 ColCreERT^f/f); mice deficient in Mll-1 healed worse than controls, and fibroblasts isolated from Mll-1 deficient wounds expressed lower amounts of myofibroblast genes, which correlated with reduced H3K4me3, an activating mark, at the promoter sites of myofibroblast genes. ChIP on myofibroblast promoter sites of wound fibroblasts from diabetic mice showed vastly decreased H3K4me3 compared to control, suggesting additional contribution H3K4 demethylases. We found that Jarid1C, a H3K4me3 demethylase, was expressed at higher concentrations in murine diabetic wounds. We hypothesized that inhibition of JARID1C would improve wound healing and increase myofibroblast gene expression. Murine wounds injected with an inhibitor of JARID1C healed better than sham injections. We again developed a tamoxifen inducible, fibroblast-specific knockout of Jarid1C (Jarid1C ColCreERT^f/f); mice with fibroblasts deficient in Jarid1C healed better after wounding compared to control. Wound fibroblasts deficient in Jarid1C expressed myofibroblast genes at higher levels than control in vitro. This work has identified that Mll1 and Jarid1C regulate myofibroblast gene expression during diabetic wound healing, and that low levels of Mll1 and high levels of Jarid1C contribute to reduced fibroblast-to-myofibroblast transition in T2D.