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Ignacio Benedicto, Guillermo Lehmann-Mantaras, Olivier Elemento, Arvydas Maminishkis, Sheldon S Miller, Shahin Rafii, Enrique J. Rodriguez-Boulan; Tissue-specific expression of alpha-2-macroglobulin by choroid endothelial cells in response to RPE-secreted factors: implications for AMD. Invest. Ophthalmol. Vis. Sci. 2017;58(8):2261.
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© ARVO (1962-2015); The Authors (2016-present)
The onset and progression of AMD correlates with the dysfunction and atrophy of choroid blood vessels, as well as with structural and functional alterations of Bruch’s membrane and choroid extracellular matrix (ECM). However, whether and how these processes are connected remains unknown. Recent research suggests that, beyond their generic role as blood conduits, endothelial cells (ECs) secrete tissue-specific angiocrine factors that regulate organ differentiation and regeneration in response to local cues (Rafii et al., Nature 2016). Extension of this concept to the eye led us to test the hypothesis that choroid ECs perform unique retinal homeostasis roles in response to local signals (e.g. from RPE).
We isolated to high purity adult mouse choroid, neural retina, lung, liver and heart ECs and determined their transcriptome by RNAseq. We carried out qPCR and western blot assays to test in vitro whether incubation of ECs with human fetal RPE-conditioned medium induces the expression of choroid EC-enriched genes. We conducted chemical inhibition, gelatin zymography and lentiviral-mediated knockdown experiments to investigate molecular mechanisms underlying RPE-EC crosstalk. We assessed by qPCR the in vivo expression of mouse choroid EC-enriched genes after sodium iodate treatment, an established procedure to damage RPE.
Choroid ECs display high expression levels of genes encoding various secretory proteins including alpha-2-macroglobulin (A2M), a proteinase inhibitor that regulates ECM stability and turnover. RPE-conditioned medium induced A2M transcription specifically in ECs in a VEGF-dependent manner, which resulted in increased levels of secreted A2M. RPE-conditioned medium reduced the gelatinase activity of EC culture supernatants, which was partially restored after A2M knockdown in ECs. Induction of RPE damage in vivo by sodium iodate treatment resulted in decreased levels of A2M expression exclusively in the central part of the RPE/choroid tissue, where RPE damage is most severe.
Our results suggest that RPE-secreted factors (e.g. VEGF) promote increased expression of choroid EC-specific angiocrine factors (e.g. A2M) essential for normal tissue homeostasis. Disruption of this crosstalk likely perturbs choroid ECM turnover and Bruch’s membrane stability, contributing to the onset and progression of AMD.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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