Increased extracellular matrix stiffness regulates myofibroblast transformation through induction of autophagy-mediated Kindlin-2 cytoplasmic translocation

Abstract

Abstract The extracellular matrix (ECM) provides structural support for connective tissue and environmental cues for cells. Its mechanical properties regulate biological processes, such as fibroblast-myofibroblast transformation (FMT), which is a crucial component in pelvic organ prolapse (POP) development and leads to an increase in ECM stiffness. The ‘Kindlin-2’ protein, expressed by fibroblasts, plays an important role in the development of the mesoderm, which is responsible for smooth muscle, blood vessel, and connective tissue formation; however, the role of Kindlin-2 in FMT remains to be explored. In this study, we aimed to explore the role of Kindlin-2 in FMT during POP development. In our study, fibroblasts were cultured using gels of different stiffness. We also examined the expression of Kindlin-2 and genes related to the Hippo pathway and FMT, validating them in human tissues and animal models. We found that ECM stiffness induces autophagy to translocate Kindlin-2 to the cytoplasm of L929 cells, where it interacts with and degrades MOB1, thereby facilitating Yes-associated protein (YAP) entry into the nucleus and influencing FMT progression. Stiffness-induced autophagy was inhibited when using an autophagy inhibitor, which blocked the translocation of Kindlin-2 to the cytoplasm and partially reversed high-stiffness-induced FMT. We found that the fibrosis inducer TGF-β failed to induce FMT after Kindlin-2 knockdown, suggesting that Kindlin-2 is involved in the TGF-β-Smad fibrosis axis. In patients with POP, we observed an increase in cytoplasmic Kindlin-2 and nuclear YAP levels. Similar changes in vaginal wall-associated proteins were observed in a mouse model of acute vaginal injury. Overall, Kindlin-2 is a key gene in ECM stiffness, regulating FMT through autophagy induction. Thus, the inhibition of Kindlin-2 transfer to the cytoplasm could be a potential target for the treatment of POP.