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Thomas Clahsen, Birgit Regenfuss, Christian Büttner, Marie-Luise Dreisow, Tim Gabriel, Felix Bock, André Reis, Claus Cursiefen; Tyrosinase: a novel endogenous regulator of (corneal) lymphangiogenesis. Invest. Ophthalmol. Vis. Sci. 2017;58(8):1009. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Lymphangiogenesis is critically involved in tissue fluid balance, tumor metastasis and immune responses. The normal cornea is an avascular and alymphatic tissue with a distinct limbal lymphatic vascular arcade. The genetic background significantly influences the architecture of this limbal lymphatic arcade and of corneal lymphangiogenesis. Here we analyzed the responsible genetic factors for the different vascular phenotype in limbal lymphatic vessel architecture in BALB/c (low endogenous limbal lymphatic vessels) and C57BL/6 (high endogenous limbal lymphatic vessels) mice.
Quantitative trait locus (QTL) analysis was used to combine phenotypic information (e.g. vessel area) and genotypic data (e.g. molecular markers) to reveal possible candidate genes contributing to the observed variation in the analyzed trait. Naïve corneas of BALB/c, C57BL/6N and B6N-TyrcBrd mice were excised and whole-mounts were stained with the lymphatic vessel marker LYVE-1. The vessel area, vessel length, number of branching points (BPs), endpoints (EPs), and sprouts were analyzed using cellF. Proliferation of human dermal lymphatic endothelial cells (HDLEC) was measured by using a BrdU based cell proliferation ELISA. The cell viability of HDLECs was determined by using MTT.
By using the BALB/c x C57BL/6 intercross for QTL analysis we identified a potential candidate gene responsible for the differences in the limbal lymphatic vessel architecture on chromosome 7, the tyrosinase gene. To further analyze if tyrosinase influences the limbal lymphatic vessel architecture, we compared C57BL/6 and B6N-TyrcBrd mice. The comparison of both strains showed a significant increase in lymph vessel area in B6N-TyrcBrd mice compared to C57BL/6N mice. Similarly, the relative number of EPs and BPs was significantly higher in B6N-TyrcBrd mice compared to C57BL/6N mice. Furthermore in vitro experiments showed that the stimulation of HDLECs with recombinant tyrosinase led to significant reduced viability, proliferation and tube formation compared to unstimulated HDLECs.
In this study we identified tyrosinase as a novel endogenous regulator of lymphangiogenesis and as a novel regulator of different limbal lymphatic vascular phenotypes in BALB/c and C57BL/6 mice. This opens new treatment avenues in diseases associated with pathologic lymphangiogenesis.
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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