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Osmotic induction of placental growth factor in retinal pigment epithelial cells in vitro: contribution of NFAT5 activity

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Abstract

One risk factor of neovascular age-related macular degeneration is systemic hypertension; hypertension is mainly caused by extracellular hyperosmolarity after consumption of dietary salt. In retinal pigment epithelial (RPE) cells, high extracellular osmolarity induces vascular endothelial growth factor (VEGF)-A (Hollborn et al. in Mol Vis 21:360–377, 2015). The aim of the present study was to determine whether extracellular hyperosmolarity and chemical hypoxia trigger the expression of further VEGF family members including placental growth factor (PlGF) in human RPE cells. Hyperosmotic media were made up by addition of 100 mM NaCl or sucrose. Chemical hypoxia was induced by CoCl2. Gene expression was quantified by real-time RT-PCR, and secretion of PlGF-2 was investigated with ELISA. Nuclear factor of activated T cell 5 (NFAT5) was depleted using siRNA. Extracellular hyperosmolarity triggered expression of VEGF-A, VEGF-D, and PlGF genes, and secretion of PlGF-2. Hypoosmolarity decreased PlGF gene expression. Hypoxia induced expression of VEGF-A, VEGF-B, VEGF-D, and PlGF genes. Extracellular hyperosmolarity and hypoxia produced additive PlGF gene expression. Both hyperosmolarity and hypoxia induced expression of KDR and FLT-4 receptor genes, while hyperosmolarity caused neuropilin-2 and hypoxia neuropilin-1 gene expression. The hyperosmotic, but not the hypoxic, PlGF gene expression was in part mediated by NFAT5. The expression of PlGF in RPE cells depends on the extracellular osmolarity. The data suggest that high consumption of dietary salt may exacerbate the angiogenic response of RPE cells in the hypoxic retina via transcriptional activation of various VEGF family member genes.

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Acknowledgments

The authors thank Ute Weinbrecht for excellent technical assistance. This work was supported by grants from the Deutsche Forschungsgemeinschaft (KO 1547/7-1 to L.K.) and the Geschwister Freter Stiftung (Hannover, Germany).

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    Authors and Affiliations

    1. Department of Ophthalmology and Eye Hospital, Faculty of Medicine, University of Leipzig, Liebigstrasse 10-14, 04103, Leipzig, Germany

      Margrit Hollborn, Konrad Reichmuth, Philipp Prager, Peter Wiedemann, Andreas Bringmann & Leon Kohen

    2. Helios Klinikum Aue, Aue, Germany

      Leon Kohen

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    1. Margrit Hollborn
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    Correspondence to Margrit Hollborn.

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    Margrit Hollborn and Konrad Reichmuth have contributed equally to the work.

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    Hollborn, M., Reichmuth, K., Prager, P. et al. Osmotic induction of placental growth factor in retinal pigment epithelial cells in vitro: contribution of NFAT5 activity. Mol Biol Rep 43, 803–814 (2016). https://doi.org/10.1007/s11033-016-4016-9

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