Abstract
Purpose: :
To elucidate potential role of hyaluronic acid (HA) in epithelial-mesenchymal transition (EMT) in retinal pigment epithelial (RPE) cells and angiogenesis.
Methods: :
ARPE-19 cells were cultured and stimulated with tumor necrosis factor-α (TNF-α) with or without hyaluronidase (HAase) treatments. Morphological changes were photographed by phase-contrast microscopy. Status of adherens junctions was observed by immunofluorescence. Endothelial cell-attachment assay was performed between human umbilical vein endothelial cells (HUVECs) and ARPE-19 cells.
Results: :
In cultured ARPE-19 cells, EMT-associated fibrotic changes resulted from interplay between TNF-α and transforming growth factor-β (TGF-β) signaling, as reported in our previous study (Takahashi et al., J Biol Chem, 2010). Immunostaining experiments demonstrated abundant expression of HA in the TNF-α-induced cell aggregations (tentatively designated as EMT-associated fibrotic foci, EAFFs) in ARPE-19 cells. After the occurrence of EAFFs, additional experiments with HAase treatment and removal of TNF-α from the culture media induced the reverse changes in cell types, mesenchymal to epithelial transition (MET). In this experimental condition, the number of TNF-α-induced EAFFs was significantly reduced, and cadherin-madiated cell-cell interaction was re-established. Moreover, in further experiments by using co-cultured ARPE-19 with HUVECs, our studies showed that vascular endothelial cells could adhere onto the cultured RPE cells predominantly in TNF-α-induced HA deposits.
Conclusions: :
It is well known that, in chronic inflammatory lesions, inflammatory cells migrate, and produce numerous cytokines, contributing to the occurrence of EMT. Subsequently, EMT elicits over-production of extracellular matrix in the msenchymal cells (originated from epithelial cells), resulting in organ fibrosis. Our results showed that HA may play not only a key role of maintenance of EMT in RPE cells, but also can be a critical contributing factor to regulate RPE-associated angiogenesis. Our hypothesis implies that regulation of HA-associated phenomena may allow us to create new strategy for the treatment of choroidal neovascularization and other inflammatory disorders in the outer retina.
Keywords: EMT (epithelial mesenchymal transition) • extracellular matrix • neovascularization