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M.E. Marin–Castano, O. Alcazar, G. Striker, S. Cousins; Effect of Hypertension–Associated Hormones Endothelin–1 and Angiotensin II on Regulation of Endothelin Receptors in RPE: Influence on Extracellular Matrix Turnover . Invest. Ophthalmol. Vis. Sci. 2006;47(13):1381.
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The pathogenesis of age–related macular degeneration is multifactorial, and hypertension (HTN) has been implicated. We postulate that HTN might contribute to RPE injury and subretinal deposit formation through the actions of HTN–associated hormones like Endothelin 1 (ET–1) and Angiotensin II (Ang–II). ET–1 and Ang–II through interactions with its cell surface receptors can regulate the synthesis of molecules important in the maintenance of the extracellular matrix (ECM), especially MMPs. Previously, we have demonstrated that human and murine RPE express both Ang–II receptors and that Ang–II increases expression of MMP–2 and MMP–9 activity at physiological concentrations. In this study we sought to determine: a) whether ET receptors are expressed and regulated by ET–1 alone and in combination with Ang II in RPE; b) the impact of both HTN–associated hormones ET–1 and Ang II together on molecules important in ECM turnover. Here, we first confirmed the expression of ET receptors (ETA and ETB) and its regulation by ET–1 alone or in combination with Ang–II, and then characterized the regulation of RPE expression of MMPs by these two HTN– associated hormones.
Expression of endothelin receptors was examined in isolated human RPE and in cultured human ARPE–19 cells, by using total RNA for RT–PCR. Regulation of these receptors by ET–1 and Ang II was evaluated by both RT–PCR and Real Time PCR. Confluent cultures of human ARPE–19 cells were incubated with ET–1 (5 ng/ml or 10 ng/ml) alone and in combination with physiologic concentrations of Ang–II (10–6 and 10–5 M) for 24 hours. Cells were collected to perform RT–PCR and Real Time PCR analysis. Supernatants were used to assess for MMPs activity.
We confirmed that both ARPE–19 and human fresh isolated RPE express ETA and ETB endothelin receptor subtypes. ET–1(10 ng/ml) alone for 24 hrs dowregulated ETA receptor. This decrease was blocked by Ang II. ETB receptor was upregulated by ET–1 (5ng/ml and 10 ng/ml) alone for 24 hrs, and this effect was increased by Ang II. MMPs activity analysis demonstrated that ET–1 alone upregulated MMP–2 and MMP–9 activities and that Ang II potentated this effect.
This study demonstrates the presence of functional endothelin receptors in human RPE. In addition, data support the hypothesis that ET1 and Ang II together may regulate ECM molecule expression in hypertensive patients who have age–related macular degeneration. CR: N.
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