Identification of therapeutic targets for treating fibrotic diseases and cancer remains challenging. Our study aimed to investigate the effects of TGF-β1 and TGF-β3 on myofibroblast differentiation and extracellular matrix deposition in different types of fibroblasts, including normal/dermal, cancer-associated, and scar-derived fibroblasts. When comparing the phenotype and signaling pathways activation we observed extreme heterogeneity of studied markers across different fibroblast populations, even within those isolated from the same tissue. Specifically, the presence of myofibroblast and deposition of extracellular matrix were dependent on the origin of the fibroblasts and the type of treatment they received (TGF-β1 vs. TGF-β3). In parallel, we detected activation of canonical signaling (pSMAD2/3) across all studied fibroblasts, albeit to various extents. Treatment with TGF-β1 and TGF-β3 resulted in the activation of canonical and several non-canonical pathways, including AKT, ERK, and ROCK. Among studied cells, cancer-associated fibroblasts displayed the most heterogenic response to TGF-β1/3 treatments. In general, TGF-β1 demonstrated a more potent activation of signaling pathways compared to TGF-β3, whereas TGF-β3 exhibited rather an inhibitory effect in keloid- and hypertrophic scar-derived fibroblasts suggesting its clinical potential for scar treatment. In summary, our study has implications for comprehending the role of TGF-β signaling in fibroblast biology, fibrotic diseases, and cancer. Future research should focus on unraveling the mechanisms beyond differential fibroblast responses to TGF-β isomers considering inherent fibroblast heterogeneity.

BIOCEV 1st Faculty of Medicine Charles University 252 50 Vestec Czech Republic

Department Dermatovenereology 1st Faculty of Medicine Charles University and General University Hospital 128 08 Prague Czech Republic

Department of Biomedical Research East Slovak Institute of Cardiovascular Diseases Inc Ondavská 040 11 Košice Slovak Republic

Department of Heart Surgery East Slovak Institute of Cardiovascular Diseases Inc 040 11 Košice Slovak Republic

Department of Pharmacognosy and Botany Faculty of Pharmacy Comenius University 832 32 Bratislava Slovak Republic

Department of Pharmacology Faculty of Medicine Pavol Jozef Šafárik University in Košice 040 11 Košice Slovak Republic

Institute of Anatomy 1st Faculty of Medicine Charles University U Nemocnice 2 128 00 Prague Czech Republic

Institute of Neurobiology Biomedical Research Center of the Slovak Academy of Sciences 040 01 Košice Slovak Republic

Prague Burn Center 3rd Faculty of Medicine Charles University and University Hospital Královské Vinohrady 100 00 Prague Czech Republic

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