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Shigeo Tamiya, Kevin McDonald, Henry J Kaplan; PYK2 plays a role in matrix contraction by RPE-derived cells. Invest. Ophthalmol. Vis. Sci. 2016;57(12):246. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Contractile fibroblastic/myofibroblastic cells derived from retinal pigment epithelial (RPE) cells have been implicated in the fibrotic complications of proliferative vitreoretinopathy. We have previously demonstrated that matrix contraction by RPE-derived cells can be inhibited by dasatinib, an FDA-approved cancer medication that inhibits multiple tyrosine kinases. The purpose of this project was to identify molecular targets affected by dasatinib that contribute to matrix contraction.
Primary cultured porcine RPE cells were used between passages 3 to 5. Cells were cultured for 3 days in 25% vitreous fluid supplemented DMEM in the presence or absence of tyrosine kinase inhibitors for the final 24 hours. Western blot analyses were used to determine the phosphorylation status of phospho-tyrosine proteins. A type I collagen contraction assay was utilized to examine matrix contraction by cells.
Dasatinib reduced tyrosine phosphorylation of multiple proteins. Interestingly, while FAK auto-phosphorylation on Tyr397 was unaffected, auto-phosphorylation on Tyr402 of PYK2, also known as FAK2, was significantly reduced. In agreement with the Western blot data, PF431396, a dual inhibitor of PYK2 and FAK, significantly reduced matrix contraction while PF573228, an inhibitor of FAK but not PYK2, was without effect.
The inhibitory effect of dasatinib on matrix contraction by RPE-derived cells is, at least in part, due to the prevention of PYK2 activation, which plays a role in the contractile process.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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