Vascular Endothelial Growth Factor (VEGF) Induced Downstream Responses to Transient Receptor Potential Vanilloid 1 (TRPV1) and 3-Iodothyronamine (3-T1AM) in Human Corneal Keratocytes

Ersal Türker; Fabian Garreis; Noushafarin Khajavi; Peter S Reinach; Pooja Joshi; Tobias Brockmann; Alexander Lucius; Nina Ljubojevic; Elizabeth Turan; Drew Cooper; Felix Schick; Rob Reinholz; Uwe Pleyer; Josef Köhrle; Stefan Mergler

doi:10.3389/fendo.2018.00670

Vascular Endothelial Growth Factor (VEGF) Induced Downstream Responses to Transient Receptor Potential Vanilloid 1 (TRPV1) and 3-Iodothyronamine (3-T₁AM) in Human Corneal Keratocytes

Front Endocrinol (Lausanne). 2018 Nov 22:9:670. doi: 10.3389/fendo.2018.00670. eCollection 2018.

Authors

Ersal Türker¹, Fabian Garreis², Noushafarin Khajavi^{3

4}, Peter S Reinach⁵, Pooja Joshi¹, Tobias Brockmann^{1

6}, Alexander Lucius¹, Nina Ljubojevic¹, Elizabeth Turan¹, Drew Cooper¹, Felix Schick¹, Rob Reinholz¹, Uwe Pleyer¹, Josef Köhrle⁷, Stefan Mergler¹

Affiliations

¹ Klinik für Augenheilkunde, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Berlin, Germany.
² Department of Functional and Clinical Anatomy, Friedrich Alexander University Erlangen-Nürnberg, Erlangen, Germany.
³ Institut für Experimentelle Pädiatrische Endokrinologie, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Berlin, Germany.
⁴ Walter Straub Institute of Pharmacology and Toxicology, Ludwig Maximilian University of Munich, Munich, Germany.
⁵ School of Ophthalmology and Optometry, Wenzhou Medical University, Wenzhou, China.
⁶ Berlin Institute of Health (BIH), Berlin, Germany.
⁷ Institut für Experimentelle Endokrinologie, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Berlin, Germany.

Abstract

This study was undertaken to determine if crosstalk among the transient receptor potential (TRP) melastatin 8 (TRPM8), TRP vanilloid 1 (TRPV1), and vascular endothelial growth factor (VEGF) receptor triad modulates VEGF-induced Ca²⁺ signaling in human corneal keratocytes. Using RT-PCR, qPCR and immunohistochemistry, we determined TRPV1 and TRPM8 gene and protein coexpression in a human corneal keratocyte cell line (HCK) and human corneal cross sections. Fluorescence Ca²⁺ imaging using both a photomultiplier and a single cell digital imaging system as well as planar patch-clamping measured relative intracellular Ca²⁺ levels and underlying whole-cell currents. The TRPV1 agonist capsaicin increased both intracellular Ca²⁺ levels and whole-cell currents, while the antagonist capsazepine (CPZ) inhibited them. VEGF-induced Ca²⁺ transients and rises in whole-cell currents were suppressed by CPZ, whereas a selective TRPM8 antagonist, AMTB, increased VEGF signaling. In contrast, an endogenous thyroid hormone-derived metabolite 3-Iodothyronamine (3-T₁AM) suppressed increases in the VEGF-induced current. The TRPM8 agonist menthol increased the currents, while AMTB suppressed this response. The VEGF-induced increases in Ca²⁺ influx and their underlying ionic currents stem from crosstalk between VEGFR and TRPV1, which can be impeded by 3-T₁AM-induced TRPM8 activation. Such suppression in turn blocks VEGF-induced TRPV1 activation. Therefore, crosstalk between TRPM8 and TRPV1 inhibits VEGFR-induced activation of TRPV1.

Keywords: 3-iodothyronamine; human corneal keratocytes; intracellular Ca2+; planar patch-clamp technique; thyronamine; transient receptor potential channel melastatin 8; transient receptor potential channel vanilloid 1; vascular endothelial growth factor.