Aldose Reductase Inhibitor Fidarestat Counteracts Apoptosis, Nitrosative Stress and Parp Activation in Diabetic Retina and High Glucose-Exposed Cultured Retinal Capillary Cells

Abstract

Purpose:: Numerous findings suggest that aldose reductase inhibition counteracts oxidative-nitrosative stress, and its major consequences i.e., PARP activation and apoptosis, but the information has been contradictory. The purpose of our study was to assess the effect of the potent and specific aldose reductase inhibitor fidarestat on apoptosis, nitrosative stress and PARP activation in the retina of STZ-diabetic rats and high glucose-exposed bovine retinal pericytes and endothelial cells.

Methods:: Control (C) and STZ-diabetic (D) rats were treated with/without fidarestat (F, 16 mgkg^-1d^-1) for 10 wks after 2 wks without treatment. The rate of apoptosis was assessed in flat-mounted retinas by TUNEL assay with immunoperoxidase staining. Primary bovine retinal pericytes and endothelial cells were cultured in 5 mM glucose or 30 mM glucose with/without F (10 µM) for 14 d. Apoptosis was assessed by TUNEL assay, cell viability with Trypan blue, and nitrotyrosine (NT) and poly(ADP-ribose) (PAR) by immunocytochemistry.

Results:: The number of TUNEL-positive nuclei (Mean ± SEM) was increased ~4-fold in D (207 ± 33 vs 49 ± 4 in C, p < 0.01), and this increase was partially prevented by F (106 ± 34, p < 0.05 vs D). The apoptotic cell number, NT immunoreactivity, and PAR-positive nuclei counts were increased 5.1-, 2.7- and 6.2-fold in pericytes cultured in 30 mM vs those cultured in 5 mM glucose, and 5-, 2.5- , and 5-fold, respectively, in endothelial cells cultured in 30 mM glucose vs those cultured in 5 mM glucose. F treatment partially prevented enhanced nitrosative stress (to 1.2-fold of control) and PARP activation (to 2.3-fold of control), and essentially prevented increased apoptosis (to 1.03-fold of control) in high glucose-exposed bovine retinal pericytes, and partially prevented PARP activation and apoptosis (to 1.8- and 2.4-fold of controls) despite a complete prevention of nitrosative stress in high glucose-exposed endothelial cells.

Conclusions:: AR inhibition with F counteracts diabetes- and high glucose-induced apoptosis, nitrosative stress, and PARP activation in retina and retinal capillary cells. Evaluation of F on Bax and Bcl-2 levels and early retinal vascular abnormalities is in progress.

Keywords: oxidation/oxidative or free radical damage • apoptosis/cell death