Alternatively activated macrophages derived from THP‐1 cells promote the fibrogenic activities of human dermal fibroblasts

Abstract

Macrophages play a key role in the wound healing process and can be divided into classically activated macrophages (M1) and alternatively activated macrophages (M2). Fibroblasts maintain the physical integrity of connective tissue, participate in wound closure as well as produce and remodel extracellular matrix. Macrophages have a close relationship with fibroblasts by increasing the production of matrix metalloproteinase-1 (MMP-1) for faster wound closure and remodeling and myofibroblast differentiation from fibroblasts. In this study, resting state (M0), M1 and M2 macrophages differentiated from the human monocytic THP-1 cell line were used to co-culture with human dermal fibroblasts (HDF) for 48, 96 and 144 hours to investigate the effect of macrophages subsets on the fibrogenic activity of fibroblasts. The differentiation and polarization from THP-1 cells to M0, M1 and M2 macrophages were characterized by flow cytometry and cell cycle analysis. Cell sorting was performed to purify M0 and M2 macrophages. Cell proliferation, collagen synthesis, myofibroblast formation, gene expression of anti-fibrotic and pro-fibrotic factors, MMP-1 activity, and cytokine concentration were investigated. Results showed differentiation of M0 and polarization of M1 and M2 macrophages. M2 macrophages promoted the fibrogenic activities of co-cultured HDF by facilitating cell proliferation, increasing the collagen content, alpha-smooth muscle actin expressed cells, expression of the pro-fibrotic genes and concentration of M2 macrophage related factors, as well as decreasing the expression of the anti-fibrotic genes and MMP-1 activity. These findings reinforce the pro-fibrotic role of M2 macrophages, suggesting therapeutic strategies in fibrotic diseases should target M2 macrophages in the future.

IL‐6 mediates ER expansion during hyperpolarization of alternatively activated macrophages

“…Additionally, future work should be performed with IRE1‐deficient macrophages, to identify a definitive mechanism between XBP1 and IL‐6. Interestingly, it is thought that soluble mediators produced by M2 programmed macrophages enhance fibroblasts fibrogenicity by promoting their proliferation, collagen production and fibroblasts‐to‐myofibroblast differentiation . It is difficult to speculate if any or all of these processes are impacted by targeting ER expansion in macrophages.…”

“…Interestingly, it is thought that soluble mediators produced by M2 programmed macrophages enhance fibroblasts fibrogenicity by promoting their proliferation, collagen production and fibroblasts-to-myofibroblast differentiation. 24,38 It is difficult to speculate if any or all of these processes are impacted by targeting ER expansion in macrophages. These experiments will yield novel and exciting data allowing for the better understanding of mechanisms underlying macrophage profibrotic activities.…”

Macrophages in wound healing: activation and plasticity

“…For instance, IL‐4/IL‐13‐activated human THP‐1 cells induced proliferation, collagen synthesis and α‐smooth muscle actin (α‐SMA) expression by human dermal fibroblasts, in a co‐culture assay . These M2‐differentiated THP‐1 cells also increased dermal fibroblast expression of α‐SMA, a feature of myofibroblasts, indicating that fibroblasts were differentiated into myofibroblasts . Furthermore, abrogation of IL‐4Rα signaling in macrophages impaired wound repair in in vivo models of wound healing by skin punch biopsy, chemical‐induced injury or invasive helminth infection‐induced injury .…”

“…The critical importance of M2 macrophages in mediating wound healing is demonstrated in numerous in vitro and in vivo studies. For instance, IL‐4/IL‐13‐activated human THP‐1 cells induced proliferation, collagen synthesis and α‐smooth muscle actin (α‐SMA) expression by human dermal fibroblasts, in a co‐culture assay . These M2‐differentiated THP‐1 cells also increased dermal fibroblast expression of α‐SMA, a feature of myofibroblasts, indicating that fibroblasts were differentiated into myofibroblasts .…”

Black, White, and Gray: Macrophages in Skin Repair and Disease

“…M2 macrophages and particularly a subset that is CD206+/CD301b+ increase fibroblast proliferation and collagen deposition through conversion of fibroblasts into a-smooth muscle actin depositing myofibroblasts 68,69 . Conversely, pro-inflammatory macrophages increase the production of inflammatory cytokines in fibroblasts and promote extracellular matrix breakdown 67 .…”