Keloid fibroblasts have elevated and dysfunctional mechanotransduction signaling that is independent of TGF-β

• Published in: Journal of dermatological science 2021-10, Vol.104 (1), p.11-20
• Main Authors: Deng, Zhenjun, Subilia, Manon, Chin, Ian L., Hortin, Nicole, Stevenson, Andrew W., Wood, Fiona M., Prêle, Cecilia M., Choi, Yu Suk, Fear, Mark W.
• Format: Article
• Language: English
• Subjects: Actins - metabolism; Adolescent; Adult; Aged; Cells, Cultured; Female; Fibroblasts - metabolism; Humans; Keloid - pathology; Keloid fibroblasts; Male; Matrix stiffness; Mechanotransduction, Cellular; Middle Aged; Normal fibroblasts; Primary Cell Culture; Signal Transduction; Skin - cytology; Skin - pathology; Transforming Growth Factor beta1 - metabolism; Transforming growth factor-β; YAP mechanotransduction; YAP-Signaling Proteins - metabolism
• ISSN: 0923-1811; 1873-569X
• DOI: 10.1016/j.jdermsci.2021.09.002

•Keloid fibroblasts are stiffer than normal fibroblasts when cultured on hydrogels that mimic tissue stiffness.•YAP signaling in normal fibroblasts but not keloid fibroblasts is sensitive to both biomechanical and biochemical stimuli in vitro.•Constitutive elevated YAP signalling in keloid fibroblasts alters their phenotype to increase α-SMA expression. Fibroblasts found in keloid tissues are known to present an altered sensitivity to microenvironmental stimuli. However, the impact of changes in extracellular matrix stiffness on phenotypes of normal fibroblasts (NFs) and keloid fibroblasts (KFs) is poorly understood. Investigation the impact of matrix stiffness on NFs and KFs mainly via detecting yes-associated protein (YAP) expression. We used fibronectin-coated polyacrylamide hydrogel substrates with a range from physiological to pathological stiffness values with or without TGF-β (fibrogenic inducer). Atomic force microscopy was used to measure the stiffness of fibroblasts. Cellular mechanoresponses were screened by immunocytochemistry, Western blot and Luminex assay. KFs are stiffer than NFs with greater expression of α-SMA. In NFs, YAP nuclear translocation was induced by increasing matrix stiffness as well as by stimulation with TGF-β. In contrast, KFs showed higher baseline levels of nuclear YAP that was not responsive to matrix stiffness or TGF-β. TGF-β1 induced p-SMAD3 in both KFs and NFs, demonstrating the pathway was functional and not hyperactivated in KFs. Moreover, blebbistatin suppressed α-SMA expression and cellular stiffness in KFs, linking the elevated YAP signaling to keloid phenotype. These data suggest that whilst normal skin fibroblasts respond to matrix stiffness in vitro, keloid fibroblasts have elevated activation of mechanotransduction signaling insensitive to the microenvironment. This elevated signaling appears linked to the expression of α-SMA, suggesting a direct link to disease pathogenesis. These findings suggest changes to keloid fibroblast phenotype related to mechanotransduction contribute to disease and may be a useful therapeutic target.