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Deregulation of ocular nucleotide homeostasis in patients with diabetic retinopathy

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Abstract

Clear signaling roles for ATP and adenosine have been established in all tissues, including the eye. The magnitude of signaling responses is governed by networks of enzymes; however, little is known about the regulatory mechanisms of purinergic signaling in the eye. By employing thin-layer chromatographic assays with 3H-labeled substrates, this study aimed to evaluate the role of nucleotide homeostasis in the pathogenesis of vitreoretinal diseases in humans. We have identified soluble enzymes ecto-5′-nucleotidase/CD73, adenylate kinase-1, and nucleoside diphosphate kinase in the vitreous fluid that control active cycling between pro-inflammatory ATP and anti-inflammatory adenosine. Strikingly, patients with proliferative form of diabetic retinopathy (DR) had higher adenylate kinase activity and ATP concentration, when compared to non-proliferative DR eyes and non-diabetic controls operated for rhegmatogenous retinal detachment, macular hole, and pucker. The non-parametric correlation analysis revealed positive correlations between intravitreal adenylate kinase and concentrations of ATP, ADP, and other angiogenic (angiopoietins-1 and -2), profibrotic (transforming growth factor-β1), and proteolytic (matrix metalloproteinase-9) factors but not erythropoietin and VEGF. Immunohistochemical staining of postmortem human retina additionally revealed selective expression of ecto-5′-nucleotidase/CD73 on the rod-and-cone-containing photoreceptor cells. Collectively, these findings provide novel insights into the regulatory mechanisms that influence purinergic signaling in diseased eye and open up new possibilities in the development of enzyme-targeted therapeutic approaches for prevention and treatment of DR.

Key message

  • Ecto-5′-nucleotidase/CD73 and adenylate kinase-1 circulate in human vitreous fluid.

  • Adenylate kinase activity is high in diabetic eyes with proliferative retinopathy.

  • Diabetic eyes display higher intravitreal ATP/ADP ratio than non-diabetic controls.

  • Soluble adenylate kinase maintains resynthesis of inflammatory ATP in diabetic eyes

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Acknowledgments

This work was supported by grants from the Academy of Finland, the Sigrid Juselius Foundation, the European Community’s Seventh Framework Program (FP7/2007–2013; grant agreement no. 602200) (SJ and GGY), the Finnish Eye Foundation, the Eye and Tissue Bank Foundation, the Mary and Georg C. Ehrnrooth Foundation, the Nissi Foundation, the Friends of the Blind, and HUCH Clinical Research Grants (TYH2016230 after TYH1325) (SL). We are grateful to Professors Be Wieringa and Jean Sevigny for providing the antibodies. We thank Professor Pier Enrico Gallenga and Drs. Giuseppe Lattanzio, Markku Kallajoki, and Maria Gardber for their expert help in examining the histological samples. We also thank Sari Mäki and Teija Kanasuo for their technical assistance, Seija Rusanen for drawing the schematic eye images, and Ruth Fair-Mäkelä for the revision of the text.

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Authors and Affiliations

  1. Unit of Vitreoretinal Surgery, Department of Ophthalmology, Helsinki University Central Hospital and University of Helsinki, Helsinki, Finland

    Sirpa Loukovaara

  2. Cell Imaging Core, Turku Centre for Biotechnology, University of Turku and Åbo Akademi University, Turku, Finland

    Jouko Sandholm

  3. MediCity Research Laboratory and Department of Medical Microbiology and Immunology, University of Turku, Tykistökatu 6A, 20520, Turku, Finland

    Kristiina Aalto, Sirpa Jalkanen & Gennady G. Yegutkin

  4. Department of Ophthalmology, Turku University Hospital, Turku, Finland

    Janne Liukkonen

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  1. Sirpa Loukovaara
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Correspondence to Gennady G. Yegutkin.

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Loukovaara, S., Sandholm, J., Aalto, K. et al. Deregulation of ocular nucleotide homeostasis in patients with diabetic retinopathy. J Mol Med 95, 193–204 (2017). https://doi.org/10.1007/s00109-016-1472-6

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