FGF2-mediated attenuation of myofibroblast activation is modulated by distinct MAPK signaling pathways in human dermal fibroblasts

Abstract

Background Previous human and animal studies have demonstrated the ability of exogenously administered basic fibroblast growth factor (FGF2) to act as an antifibrotic agent in the skin. Though the activity of FGF2 as an anti-scarring agent is well-established for fibrotic skin wounds, the mechanisms by which FGF2 exerts these actions are not entirely understood. Canonical FGF2 signaling proceeds in part via FGFR/MAPK pathways in human dermal fibroblasts, and FGF2 has been described to prevent or reverse the fibroblast-to-myofibroblast transition, which is driven by TGFβ signaling and understood to be an important step in the formation of a fibrotic scar in vivo. Thus, we set out to investigate the antagonistic effects of FGF2 on TGFβ signaling as well as the broader effects of MAPK inhibition on the TGFβ-mediated induction of myofibroblast gene expression. Objective To better understand the effects of FGF2 signaling pathways on myofibroblastic gene expression and cell phenotypes. Methods Human dermal fibroblasts were cultured in vitro in the presence of FGF2, TGFβ, and/or MAPK inhibitors, and the effects of these agents were investigated by molecular biology techniques including qRT-PCR, immunofluorescence, Western blot, and flow cytometry. Results FGF2 inhibited TGFβ-mediated fibroblast activation, resulting in more rapidly proliferating, spindle-shaped cells, compared to the more slowly proliferating, flatter TGFβ-treated cells. Treatment with FGF2 also attenuated TGFβ-mediated increase in expression of myofibroblast markers smooth muscle α-actin, calponin, transgelin, connective tissue growth factor, ED-A fibronectin, and collagen I. FGF2-mediated antagonism of the TGFβ-mediated fibroblast-to-myofibroblast transition was reversed by small molecule inhibition of ERK or JNK, and it was potentiated by inhibition of p38. MAPK inhibition was demonstrated to have qualitatively similar effects even in the absence of exogenous FGF2, and small molecule inhibition of p38 MAPK was sufficient to attenuate TGFβ-mediated fibroblast activation. Conclusions Inhibition of select MAPK signaling pathways can reverse or potentiate anti-fibrotic FGF2 effects on human dermal fibroblasts, as well as exert their effects independently of exogenous FGF2 supplementation.

Artesunate inhibits myofibroblast formation via induction of apoptosis and antagonism of pro‐fibrotic gene expression in human dermal fibroblasts

“…Samples were processed and Western blot was performed using standard protocols as described previously (Dolivo et al, ).…”

“…Cells were cultured in VisiPlate 24‐well plates (Perkin Elmer) until analysis. Cells were fixed and analyzed by immunofluorescence as described previously (Dolivo et al, ) and imaged on an Axiovert 200 M (Zeiss). Alternatively, cells were stained for FITC‐annexin V/PI according to manufacturer's protocols and analyzed on an Axiovert 200 M or an Accuri C6 flow cytometer (BD Biosciences).…”

FGF2 engineered SPIONs attenuate tumor stroma and potentiate the effect of chemotherapy in 3D heterospheroidal model of pancreatic tumor

“…FGF2 is an interesting growth factor, which is known to inhibit fibroblasts activation in vitro by distinct signaling pathways such as Smad2/3 and MAPK pathways [15,16,18]. The differentiation and activation of hPSCs into CAF-like myofibroblasts is mainly induced by TGF-β as a result of the interaction with cancer cells within the tumor microenvironment.…”

“…Activation and differentiation of PSCs into myofibroblastic CAFs is commonly regulated by Transforming growth factor beta (TGF-β) [13]. In literature, fibroblast growth factor 2 (FGF2, 17.2 kDa) has been shown to exert inhibitory effects against TGF-β [ [14][15][16][17][18]. The activation of extracellularregulated kinase 1/2 (ERK1/2) pathway by FGF2 has been shown to counteract the TGF-β mediated activation in endothelial cells or fibroblasts [14][15][16][17][18].…”

Fibroblast Growth Factor 2 as an Antifibrotic: Antagonism of Myofibroblast Differentiation and Suppression of Pro-Fibrotic Gene Expression

“…Our lab recently demonstrated that FGF2 was able to inhibit TGF-β1-mediated activation of several lines of primary adult and neonatal human dermal fibroblasts, and that this inhibition could be abrogated by treatment with small molecule inhibitors of ERK and JNK but not of p38. We also demonstrated that treatment with ERK or JNK inhibitor in the absence of exogenous FGF2 was sufficient to induce fibroblast activation, potentially implicating basal activation of these pathways in the promotion of myofibroblast phenotypes, or implicating well-established cross-talk between MAPK and TGF-β signaling more generally in this process [69]. Taken together, these data suggest that FGF2 has the ability to prevent or reverse fibroblast activation via various cellular pathways and mechanisms that are likely context-dependent.…”