Abstract
Purpose: :
In our previous studies, we have demonstrated that green tea polyphenols (GTPs) attenuate the early changes in the retina from experimental diabetic rats. As the Müller cell is the predominant type of glial cell in the retina and has a central role in the homeostasis in the retina and ARPE-19 is a human retinal pigment epithelium and has been used to evaluate not only the outer BRB but also the retinal neuroprotection, the aims of this study were to investigate the possible mechanisms involved into the protective effects of GTPs in diabetic milieu condition in in vitro studies.
Methods: :
For this, Müller and ARPE-19 cells were used and were exposed to normal glucose (NG; 5mM), high glucose (HG; 25mM) in presence or absence of GTPs treatment (1, 10 and 100 µg/ml) and HG with epigallocatechin gallate (EGCG) (for ARPE-19) for 24 hours. The total intracellular reactive species (ROS) was determined in both cells. In Müller cells, we analyzed the expression of glial fibrillary acidic protein (GFAP) by imunofluorescence, glutamate transporter (Glast), glutamate receptor (NMDAR1) and glutamine synthetase by Western blot and reduced glutathione (GSH) levels by colorimetric assays. For ARPE-19 cells, we analyzed the protein expressions of claudin-1, a tight junction protein, and the EAAC1, a glutamate transporter present in ARPE-19 cells.
Results: :
In Müller cells, under HG conditions, the expressions of GFAP and glutamine synthetase increased, glutamate transporter and receptor levels decreased accompanied by a reduction in glutathione content and increasing in ROS production. In ARPE-19 cells under HG medium, there was an increased ROS production accompanied by decreasing in claudin-1 and glutamate transporter expressions. The treatment with GTPs reestablished all of the above-mentioned parameters. Similar findings were observed in ROS production in presence of EGCG under HG condition in ARPE-19 cells.
Conclusions: :
In the Müller cells, there was a significant reestablishment of glutamatetransporter/NMDAR1 status and oxidative status. GTPs also prevented the ROS and EAAC1 increments and claudin-1 alteration in ARPE-19 cells. The involvement of glutamate cycle either in Müller as in ARPE-19 cells was described herein demonstrating the neuroprotection of GTPs in the retinal cells in diabetic milieu mimetic conditions.
Keywords: retinal culture • diabetic retinopathy • neuroprotection