Myofibroblast transdifferentiation: The dark force in ocular wound healing and fibrosis

Abstract

Wound healing is one of the most complex biological processes to occur in life. Repair of tissue following injury involves dynamic interactions between multiple cell types, growth factors, inflammatory mediators and components of the extracellular matrix (ECM). Aberrant and uncontrolled wound healing leads to a non-functional mass of fibrotic tissue. In the eye, fibrotic disease disrupts the normally transparent ocular tissues resulting in irreversible loss of vision. A common feature in fibrotic eye disease is the transdifferentiation of cells into myofibroblasts that can occur through a process known as epithelial-mesenchymal transition (EMT). Myofibroblasts rapidly produce excessive amounts of ECM and exert tractional forces across the ECM, resulting in the distortion of tissue architecture. Transforming growth factor-beta (TGFβ) plays a major role in myofibroblast transdifferentiation and has been implicated in numerous fibrotic eye diseases including corneal opacification, pterygium, anterior subcapsular cataract, posterior capsular opacification, proliferative vitreoretinopathy, fibrovascular membrane formation associated with proliferative diabetic retinopathy, submacular fibrosis, glaucoma and orbital fibrosis. This review serves to introduce the pathological functions of the myofibroblast in fibrotic eye disease. We also highlight recent developments in elucidating the multiple signaling pathways involved in fibrogenesis that may be exploited in the development of novel anti-fibrotic therapies to reduce ocular morbidity due to scarring.

Keywords: Epithelial-mesenchymal transition (EMT); Fibrosis; Myofibroblast; Ocular; Transforming growth factor-beta (TGFβ); Wound healing.

Fig. 1. Smad and Smad-independent signaling pathways downstream of TGFβ

TGFβ binds to its receptors and the activated receptor complexes relay the signal to the cytoplasm by phosphorylating receptor-regulated Smad proteins (Smad2/3) that hetero-oligomerize with Smad4. This complex then translocates to the nucleus and regulates the transcription of respective target genes. TGFβ can also activate non-Smad signaling pathways including ERK1/2, JNK, p38, Rho/ROCK and PI3K/Akt/mTOR.

Fig. 2. Schematic diagram of epithelial-mesenchymal transition (EMT)

During epithelial-mesenchymal transition, epithelial cells lose their polarity and normal cell-cell and cell-basement membrane adhesions, and transdifferentiate into motile, spindle-shaped mesenchymal cells (myofibroblasts) that secrete extracellular matrix (grey).

Fig. 3. Schematic diagram of the corneal wound healing process

Corneal injury involving disruption to Bowman’s membrane results in the increased exposure of cytokines and growth factors including TGFβ and PDGF into the anterior stroma. TGFβ induces the normally quiescent keratocytes and/or bone-marrow derived cells to transdifferentiate into myofibroblasts to facilitate corneal repair. During physiological wound healing, myofibroblasts disappear by apoptosis and tissue transparency is maintained. However, in pathological conditions, myofibroblasts persist and secrete excessive amounts of aberrant extracellular matrix proteins (fibrosis) resulting in corneal scarring.

Fig. 4. A novel human lens epithelial explant model for studying cataractogenesis

Following 5 days of culture, phase contrast microscopy shows that untreated human lens epithelial explants (A, anterior capsulorhexis tissue obtained in surgery) remain as a normal cuboidal layer of epithelial cells, with membranous localization of β-catenin and minimal α-SMA (B) viewed with confocal microscopy. Immunofluorescence labeling was performed as described previously (Shu et al., 2016). Treatment with 10 ng/ml TGFβ2 displayed an elongate, spindle-shaped morphology (C). Although β-catenin remained localized to the membrane in TGFβ2-treated explants, the label no longer highlighted the normal cobblestone-packed arrangement as in control explants (D). TGFβ-treated explants exhibited strong immunoreactivity for α-SMA, co-labeling to stress fibers (D). Scale bar: 100 µm (A, C), 25 µm (B, D).