Matrix Softness Induces an Afibrogenic Lipofibroblast Phenotype in Fibroblasts from IPF Patients

Contractile myofibroblasts immersed in stiffened remodelled extracellular matrix (ECM) characterise fibrotic lesions in idiopathic pulmonary fibrosis (IPF). Lipofibroblasts are lipid-droplet containing interstitial fibroblasts that support functional homeostasis of the developing and adult lung. We show that stiff substrates augment myofibroblast differentiation and ECM production in vitro under basal conditions and following transforming growth factor-β1 (TGF-β1) incubation when cultured on tissue culture plastic, whereas culture in soft microenvironments (as spheroids or on soft collagen-coated substrate) redirects myofibroblast to a lipofibroblast-like phenotype (identified by expression of adipose differentiation-related protein (ADRP) and intracellular lipid droplets), with reduced basal alpha-smooth muscle actin (α-SMA), collagen I, vimentin (VIM) and fibronectin (FN1) expression. The fibrogenic effects of TGF-β1 are prevented in fibroblasts cultured in soft settings. Global proteomics showed similar numbers of TGF-β1-induced differentially expressed proteins in stiff and soft settings (271 and 436, respectively). Of these, only 33 were similarly altered by TGF-β1; 200 were exclusively altered by TGF-β1 in stiff setting; 365 in soft setting, while 38 showed opposite responses. Reductions in YAP/TAZ, beta-catenin and SMAD expression and their limited nuclear levels in soft settings may explain the "afibrogenic" characteristic of these lipofibroblasts. Thus, in spheroids of lipofibroblasts TGF-β1 intracellular signalling is redirected and uncoupled from fibrogenesis, including YAP/TAZ, β-catenin and SMAD. Understanding the proximal causal mechano-transduction signalling networks that are differentially active in soft and stiff microenvironments may reveal novel drug targets for fibrosis treatment.