AUGMENTED LEVELS OF Ap₄A AND ECTONUCLEOTIDASES IN GLAUCOMATOUS MICE RETINAS

Abstract

Purpose : The dinucleotide Ap₄A, diadenosine tetraphosphate, has been claimed as naturally occurring substance with neuroprotective properties. The aim of this study was to investigate the levels of Ap₄A as well as the pattern of ectonucleotidase activity expression in glaucomatous retinas during the progress the pathology.

Methods : Ap₄A levels were analyzed by HPLC in glaucomatous retinas from the DBA/2J mice at 3, 9, 15 and 23 months of age, with a mobile phase consisting of 10 mM KH₂PO₄, 2 mM tetrabutylammonium bisulfate (TBA) and 20% acetonitrile adjusted to pH 7.5. The retinas were dissected as flattened whole-mounts and stimulated in Ringer buffer with or without 59 mM KCl. NPP1 expression was analyzed by RT-PCR and western blot and their distribution was assessed by immunohistochemistry studies examined under confocal microscopy.

Results : Glaucomatous mice exhibited Ap₄A values, which changed in stimulated retinas as long as the pathology progressed varying from 0.73 ± 0.04 (3 months) to 0.170 ± 0.05 pmol/mg retina (23 months). Concomitantly, NPP1 expression was significantly increased (82.15 %) in the DBA/2J mice at 15 months. Furthermore, immunohistochemical studies showed that NPP1 labelling was stronger in OPL and IPL labelling tangentially in the vitreal part of the retina and was up-regulated at 15 months of age.

Conclusions : Our findings demonstrate that Ap₄A decreased levels may be related with exacerbated activity of NPP1 protein in glaucomatous degeneration and in this way contributing to elucidate different mechanisms involved in retinal impairment in glaucomatous degeneration.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.