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S. Morales, S. Mareninov, J. Braun, L. K. Gordon; Epithelial Membrane Protein 2 (EMP2) Controls in vitro Collagen Gel Contraction by ARPE-19 Through a FAK-Src Dependent Pathway. Invest. Ophthalmol. Vis. Sci. 2007;48(13):5729.
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© ARVO (1962-2015); The Authors (2016-present)
Proliferative vitreoretinopathy (PVR), which occurs in approximately 10% of patients with a retinal detachment, is believed to result from migration of the retinal pigment epithelium and other cell types with subsequent membrane formation and contraction in an aberrant wound-healing response. The in vitro correlate of PVR is the gel contraction assay. In this study we investigate the role of epithelial membrane protein 2 (EMP2), a member of the GAS3/PMP22 four transmembrane family, in collagengel contraction.
The human retinal pigment epithelium cell line ARPE-19 was used in these studies. Levels of EMP2 were modulated through stable infection of an EMP-2 overexpressing retrovirus construct (ARPE-RV-EMP2) or stable transfection with an EMP2 ribozyme to reduce expression levels (ARPE/ribo). An in vitro gel contraction assay was used in which RPE cells were seeded on a collagen gel and the percent contraction measured at specific time intervals. The areas of the gel were quantified using NIH Image software. In some experiments, cells were pretreated with multiple inhibitors of signal transduction pathways. Transient transfections with a FAK siRNA were used to decrease FAK expression.
EMP2 levels directly correlated with the cells ability to contract the collagen gels. Increasing EMP2 expression resulted in a 57% increase in contraction as compared to wildtype cells. Decreasing EMP2 expression resulted in a 55% decrease in gel contraction. EMP2 over expression resulted in a significant increase in FAK activation. EMP2 and FAK may physically interact as suggested by co-immunoprecipitation data. Inhibition of the FAK-Src complex in both wild type and EMP2 overexpressing ARPE-19 cells, significantly blocked contraction (P=.0001). FAK siRNA transient transfection significantly reduced FAK protein expression by 71% (P=.02) and concordantly decreased gel contraction by 78% (P=.0001)
RPE-mediated collagen gel contraction is a complex process. Activation of FAK-Src is necessary for collagen gel contraction produced by ARPE-19 cells. EMP2 facilitates gel contraction though increasing FAK activation. Additional studies are in progress to identify the relevance of this signal transduction pathway in PVR.
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