Abstract
Purpose :
Myofibroblast transdifferentiation of retinal pigment epithelial (RPE) cells has been implicated as key event in fibrotic complications such as proliferative vitreoretinopathy or fibrosis associated with exudative age-related macular degeneration. We previously demonstrated that Rho GTPase plays a key role in this process. Interestingly, ROCK, one of the main downstream Rho effector, was not involved in this process. The purpose of this study is to examine the role of Arp2/3, another key downstream Rho effector involved in actin polymerization, in myofibroblast transdifferentiation of RPE cells.
Methods :
Primary cultured porcine RPE cells were used for experiments. Second passage RPE cells cultured on collagen hydrogels were stimulated with TGF-beta2 to induce myofibroblast transdifferentiation. Expression of myofibroblast markers, alpha-smooth muscle actin and tropomyosin-1, was examined by western blot analyses. Myofibroblast function was assessed by collagen hydrogel contraction. The cell permeable C3 transferase and CK-666 were used to inhibit Rho and Arp2/3, respectively.
Results :
As previously reported, TGF-beta2 stimulation of second passage RPE cells induced myofibroblast transdifferentiation, which was demonstrated by significant increase in expression of myofibroblast marker proteins as well as enhanced contraction of collagen hydrogel. Rho inhibitor and Arp2/3 inhibitor significantly reduced myofibroblast marker expression as well as collagen hydrogel contraction.
Conclusions :
Rho and its downstream effector Arp2/3 is involved in myofibroblast transdifferentiation of RPE cells. Actin polymerization is a key process in mechanosignaling that affects multiple downstream signaling molecules. Alteration of Rho-Arp2/3 pathway or its downstream targets may lead to therapeutic treatment of fibrotic complications such as proliferative vitreoretinopathy or fibrotic complications of exudative age-related macular degeneration in which RPE cell transdifferentiation to myofibroblasts play a significant role.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.