The persistence of Mfs in wounded tissue correlates with fibrotic disease,
34 and thus, uncovering mechanisms that promote Mf differentiation is important to our fundamental understanding and treatment of fibrosis. Cell surface receptors such as integrins, thrombospondin, and cation-independent mannose-6-phosphate receptor with and without proteases play a role in the activation of matrix-bound latent TGFβ complex into active TGFβ.
10,35–38 Although it has been established that the activation of TGFβ is a key to the generation of fibrosis, the mechanisms that promote the persistence of Mfs versus Mf apoptosis or reversal of the Mf phenotype remain unclear.
39 Toward the goal of preventing persistent Mfs, researchers have focused on the matricellular connective tissue growth factor (CTGF/CCN2) protein that surrounds Mfs in fibrotic tissue.
40,41 Although the precise role of CTGF in fibrosis is unknown, silencing of the CTGF gene expression has produced antifibrotic effects in many tissues, including the eye,
41–43 and reagents that target CTGF (antibodies and RNAi) are in clinical trials for several fibrotic diseases (Fibrogen
44 ; RXi Pharmaceuticals [Byrne MJ, et al.
IOVS 2012;53:ARVO E-Abstract 897]). Other strategies for reducing or preventing fibrosis include an inhibitory peptide that prevents incorporation of α-SMA into stress fibers, which reduces collagen type I synthesis in vitro and wound contraction in vivo.
45 A more general approach is to promote apoptosis of Mfs by protein kinase inhibitors, which interferes with activation of focal adhesion kinase (FAK) and phosphatidylinositol 3-kinase (PI3K)-AKT (pathways that protect Mfs from apoptosis).
15,16 A recent study demonstrating that FGF-2 selectively induced Mf but not fibroblast apoptosis suggests that FGF-2 may be a potential antifibrotic therapy as well.
46
In this study, we focused on the expression and secretion of uPA and PAI-1 that are increased in fibrotic disease.
29–31 How this imbalance contributes to fibrosis has been intensively studied; however, the connection between their increased expression and the generation of fibrotic disease remains elusive. PAI-1 and VN coordinate to detach cells through a disruption in integrin-mediated adhesion,
28 thus, it is paradoxical that Mfs persist in a matrix that contains VN and PAI-1.
29 Here we report that addition of uPA in the presence of TGFβ-stimulated PAI-1 promoted Mf differentiation on VN. Our data support the hypothesis that uPA “removes” PAI-1 from VN, permitting integrins αvβ3 and αvβ5 access to binding VN (presumably at the RGD domain), leading to significantly increased cell adhesion and Mf differentiation. PAI-1R, a dominant negative PAI-1 mutant,
32 decreased β3/β5-mediated cell adhesion on VN, even in the presence of excess uPA. We hypothesize that by decreasing cell adhesion, PAI-1R could combat the persistence of Mf in healing tissue and make PAI-1R a candidate for therapy that prevents corneal scarring.