Histone Demethylation and Toll‐like Receptor 8–Dependent Cross‐Talk in Monocytes Promotes Transdifferentiation of Fibroblasts in Systemic Sclerosis Via Fra‐2

Abstract

Objective To investigate whether epigenetic changes can modulate monocytes to produce tissue inhibitor of metalloproteinases 1 (TIMP‐1) via Fra‐2 (an activator protein 1 [AP‐1] family member), a novel downstream mediator that promotes fibrogenesis. Methods AP‐1 transcription factors and TIMP‐1 expression were measured in monocytes from systemic sclerosis (SSc) patients and healthy controls. Involvement of Fra‐2 in the regulation of TIMP‐1 following treatment with Toll‐like receptor 8 (TLR‐8) agonist was investigated using a luciferase activity assay and chromatin immunoprecipitation (ChIP) analysis. Expression of TIMP‐1 and Fra‐2 was determined in response to TLR‐8 treatment and to different histone modifications, including 3′‐deazaneplanocin (DZNep) and apicidin. Fibroblasts from healthy controls were cocultured with DZNep plus TLR‐8–treated healthy control monocytes. Results Up‐regulation of Fra‐2 was detected in bleomycin‐challenged mice and in skin biopsy samples from SSc patients. Enhanced expression of Fra‐2 and TIMP‐1 was correlated in SSc monocytes (P = 0.021). The expression of Fra‐1 was significantly reduced (P = 0.037) in SSc monocytes. Inhibiting AP‐1 activity reduced TIMP‐1 production in TLR‐8–stimulated monocytes from healthy controls and SSc patients. ChIP experiments revealed binding of Fra‐2 to the TIMP‐1 promoter. Stimulation with DZNep plus TLR‐8 enhanced Fra‐2 and TIMP‐1 expression in healthy control monocytes, whereas TLR‐8 plus apicidin repressed Fra‐2 and TIMP‐1 expression. Finally, healthy control monocytes treated with DZNep plus TLR‐8 induced strong production of α‐smooth muscle actin in dermal fibroblasts, which was inhibited by TIMP‐1–blocking antibody. Conclusion These data demonstrate a novel role of histone demethylation induced by DZNep on Fra‐2–mediated TIMP‐1 production by monocytes in the presence of TLR‐8 agonist. This consequently orchestrates the transdifferentiation of fibroblasts, a key event in the pathogenesis of SSc.

The role of microRNA‐5196 in the pathogenesis of systemic sclerosis

“…Apicidin is another histone modifier inducing histones hyperacetylation. We have previously validated that DZNep and apicidin concentrations, which were used to stimulate monocytes, did not affect cells viability . This suggests that only specific epigenetic agent (DZNep, but not apicidin) plus TLR8 stimulation can activate monocytes to produce profibrotic genes and changes cell morphology.…”

Toll Like Receptors in systemic sclerosis: An emerging target

“…Interestingly, single stranded RNA is found in many types of virus, suggesting that a viral origin is responsible. We have now demonstrated that the single stranded RNA mediates TIMP-1 via TLR8 and depends on trimethylation of histone27 which alters the expression of the AP-1 transcription factor Fos-Related Antigen-2 (FRA2) [58]. This links possibly viral activation of TLR8 to activation of FRA2 via epigenetic modulation by histone methylation [58].…”

Toll-like receptors in the pathogenesis of autoimmune diseases: recent and emerging translational developments

“…It has also recently been found that the AP-1 family member, Fos-related antigen 2 (Fra2), facilitates TLR8-mediated overexpression of TIMP-1, in ex vivo monocytes derived from patients with SSc 45. Skin-infiltrating monocytes from patients with SSc showed an increase in Fra2; treatment of these monocytes with a histone demethylation agent caused an increase in both Fra2 and TIMP-1 production when challenged with the TLR8 agonist ssRNA; and increased TIMP-1 can directly induce myofibroblast transdifferentiation 45.…”

“…Skin-infiltrating monocytes from patients with SSc showed an increase in Fra2; treatment of these monocytes with a histone demethylation agent caused an increase in both Fra2 and TIMP-1 production when challenged with the TLR8 agonist ssRNA; and increased TIMP-1 can directly induce myofibroblast transdifferentiation 45. The factors affecting transdifferentiation of myofibroblast is complex; once transdifferentiation is complete, the myofibroblasts are able to upregulate the expression of many fibrotic proteins, such as collagen 1.…”