Abstract
Purpose: :
Collagen gel contraction by ARPE-19 is an in vitro model for proliferative vitreoretinopathy (PVR), an aberrant wound healing response. Expression of the tetraspan protein epithelial membrane protein 2 (EMP2) controls gel contraction through FAK activation. Peripheral myelin protein 22 (PMP22), another member of the tetraspan web, is closely related to EMP2. The purpose of this study was to determine if PMP22 also controls the contractile phase associated with PVR.
Methods: :
PMP22 levels in ARPE-19 cells were increased through stable transfection (ARPE-19/PMP22). A PMP22 specific siRNA was used to decrease PMP22 expression in the cells. AKT activation was blocked using a small molecule inhibitor Ly294002. Integrin expression, adhesion, and protein expression were assessed respectively through flow cytometry, binding to collagen type I and IV, and Western Blot analysis. Collagen gel contraction was assessed using an in vitro assay.
Results: :
In comparison to the ARPE-19 cells, ARPE-19/PMP22 cells exhibited an increased collagen adhesion. Gel contraction, however, was reduced by greater than 50% in the PMP22 overexpressing cells (P<0.001). In contrast to the FAK activation observed by increasing EMP2 expression, PMP22 overexpression led to increased AKT activation and reduced FAK activation. The decrease in gel contraction by the ARPE-19/PMP22 cells was partially reversed through either PMP22 siRNA or by blockade of AKT.
Conclusions: :
Relative expression of EMP2 or PMP22 within the tetraspan web drives a cellular response towards a FAK or AKT dependent pathway, respectively. EMP2 and PMP22 differentially regulate collagen gel contraction in the ARPE-19 cell line, an in vitro model for PVR. The implication of this finding adds a new dimension to the concept of the tetraspan web that can drive the downstream response depending on both the presence and abundance of specific proteins.
Keywords: retinal pigment epithelium • proliferative vitreoretinopathy • signal transduction