Cross‐talk Between MAPK Inhibitors and TGF‐β Signaling Results in Variable Activation of Human Dermal Fibroblasts

Differentiation of fibroblasts into myofibroblasts, a process known as fibroblast activation, is a key step in the wound healing process, but persistence of myofibroblasts through the resolution of wound remodeling can lead to fibroproliferative, pathophysiological disease states. Fibroblasts are activated by myriad extracellular cues including a multitude of growth factors which relay information from the extracellular environment and are integrated into a cellular differentiation response, but the rough contributions of particular signaling pathways, and the ways in which these pathways interact with each other, are not entirely understood. Here we demonstrate that small molecule‐mediated inhibition of the mitogen activated protein kinases ERK and JNK is sufficient to promote fibroblast activation, and that inhibition of p38 MAPK is sufficient to inhibit fibroblast activation, as determined by the expression of canonical myofibroblast markers assessed by qRT‐PCR, Western blot, immunofluorescence, and flow cytometry. The promotion of fibroblast activation by ERK and JNK inhibition and the antagonism of fibroblast activation by p38 MAPK inhibition results in expected changes in expression of extracellular matrix‐associated genes including those encoding type I collagen, type III collagen, matrix metalloproteinase 1, lysyl oxidase, myocardin, connective tissue growth factor, and the ED‐A isoform of fibronectin. Co‐treatment of human dermal fibroblasts with exogenous recombinant TGF‐β1 and ERK or JNK inhibitor leads to cooperative induction of fibroblast activation, and a higher degree of activation than that imparted by treatment of either of these factors individually. Conversely, treatment with p38 inhibitor attenuates ERK inhibitor‐mediated or JNK inhibitor‐mediated fibroblast activation, suggesting antagonism between these pathways. Treatment with ERK or JNK inhibitor results in relatively rapid activation of canonical TGF‐βR/SMAD signaling, likely contributing at least in part to the activation witnessed in these fibroblasts. Fibroblast activation as a result of JNK inhibition, but not of ERK inhibition, is partially attenuated by small molecule‐mediated inhibition of TGF‐βRI, suggesting variable mechanisms of induction of fibroblast activation by inhibition of these two pathways. These data demonstrate that multiple growth factor and MAPK signals are integrated to determine the extent of fibroblast activation, and they demonstrate evidence regarding the mechanisms of fibroblast activation induced by MAPK inhibition, providing valuable information for further development of much‐needed antifibrotic therapies. Support or Funding Information This research was supported by a National Institutes of Health award to Tanja Dominko (grant # R01GM85456) and a National Science Foundation Integrative Graduate Education and Research Traineeship (grant number DGE 1144804) awarded to David Dolivo. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .