Histone Methylation Directs Myeloid TLR4 Expression and Regulates Wound Healing following Cutaneous Tissue Injury

Abstract

Myeloid cells are critical for orchestrating regulated inflammation during wound healing. TLRs, particularly TLR4, and its downstream-signaling MyD88 pathway play an important role in regulating myeloid-mediated inflammation. Because an initial inflammatory phase is vital for tissue repair, we investigated the role of TLR4-regulated, myeloid-mediated inflammation in wound healing. In a cutaneous tissue injury murine model, we found that TLR4 expression is dynamic in wound myeloid cells during the course of normal wound healing. We identified that changes in myeloid TLR4 during tissue repair correlated with increased expression of the histone methyltransferase, mixed-lineage leukemia 1 (MLL1), which specifically trimethylates the histone 3 lysine 4 (H3K4me3) position of the TLR4 promoter. Furthermore, we used a myeloid-specific Mll1 knockout (Mll1 f/f Lyz2 Cre+) to determine MLL1 drives Tlr4 expression during wound healing. To understand the critical role of myeloid-specific TLR4 signaling, we used mice deficient in Tlr4 (Tlr4 2/2), Myd88 (Myd88 2/2), and myeloid-specific Tlr4 (Tlr4 f/f Lyz2 Cre+) to demonstrate delayed wound healing at early time points postinjury. Furthermore, in vivo wound myeloid cells isolated from Tlr4 2/2 and Myd88 2/2 wounds demonstrated decreased inflammatory cytokine production. Importantly, adoptive transfer of monocyte/macrophages from wildtype mice trafficked to wounds with restoration of normal healing and myeloid cell function in Tlr4-deficient mice. These results define a role for myeloid-specific, MyD88-dependent TLR4 signaling in the inflammatory response following cutaneous tissue injury and suggest that MLL1 regulates TLR4 expression in wound myeloid cells.

TNF-α regulates diabetic macrophage function through the histone acetyltransferase MOF

“…This switch is significantly impaired in obesity/T2D and results in chronic inflammation and nonhealing wounds; however, the etiology is unclear. We and others have previously identified that chronic inflammation in diabetic wounds is governed by epigenetic regulation of macrophages (12)(13)(14)(15)(16). In this study, we identify that the HAT MOF regulates inflammation in macrophages during normal and diabetic wound repair.…”

“…Wounds were then carefully minced with sharp scissors and digested by incubating in a solution of 50 mg/mL Liberase TM (Roche) and 20 U/mL DNase I (Milli-poreSigma). Because the final cell number is limited, wounds from the same genotype/treatment were pooled to maximize cell yield, as previously published (11,12,14,16,54,(57)(58)(59). Wound cell suspensions were then gently plunged and filtered through a 100-μm filter to yield a single-cell suspension.…”

“…Sustained inflammation due to the inability of wound macrophages to transition to the regeneration stage necessary for normal skin repair is a hallmark of impaired wound healing associated with diseases such as type 2 diabetes (T2D) (6)(7)(8)(9)(10)(11). The mechanisms regulating this failed macrophage transition are not completely clear, but recent evidence suggests that epigenetic signatures can drive prolonged inflammation in diabetic wounds (12)(13)(14)(15)(16). Our group and others have identified that diabetic wound macrophages exhibit changes in histone methylation that promote a proinflammatory phenotype (12,14,16).…”

“…The mechanisms regulating this failed macrophage transition are not completely clear, but recent evidence suggests that epigenetic signatures can drive prolonged inflammation in diabetic wounds (12)(13)(14)(15)(16). Our group and others have identified that diabetic wound macrophages exhibit changes in histone methylation that promote a proinflammatory phenotype (12,14,16). Specifically, we have shown that MLL1 and JMJD3; a histone methyltransferase and histone demethylase, respectively, are altered in T2D macrophages, drive production of proinflammatory cytokines, and establish a proinflammatory feedback loop leading to uncontrolled inflammation (12,14,16).…”

A review of epigenetic regulation in wound healing: Implications for the future of wound care

“…Major surgery induces rapid changes in DNA methylation in genes associated with the inflammatory response and wound healing, including C‐reactive protein and IL‐6 99,100 . It is recognized that these shifts in methylation may contribute to surgical complications including infection and delirium 99,101,102 …”