Abstract
Purpose:
Diabetic retinopathy (DR), the most frequently occurring microvascular complication of diabetes, is characterized by blood retinal barrier vascular leakage and pre-retinal neovascularization. High glucose concentrations inhibit PI3 kinase pro-survival signaling in retinal endothelial cells resulting in p38-dependent apoptosis. We have demonstrated that quinic acids promote REC survival under various genotoxic stresses by inhibiting p38 pro-apoptotic signaling. Thus, we hypothesized that the quinic acid derivative, KZ-41, would inhibit glucose-induced apoptosis in RECs.
Methods:
RECs were grown in M131 medium supplemented with normal (5mM, NG) or high (25mM, HG) glucose. After 3 days, KZ-41 (10 µM), +/- the PI3K antagonist LY294002, was added. Two hours later, cell lysates were collected for measurement of cleaved caspase-3 (ELISA) and phosphorylated and total levels of Akt and PI3K (p85 subunit) proteins (Western blot).
Results:
KZ-41 significantly reduced the cleaved caspase-3 levels in RECs exposed to high glucose (0. 37 ± 0.09 vs. 0.62 ± 0.04; P < 0.01). The ratio of phosphorylated to total Akt protein expression was significantly decreased between 24-96 hours of high glucose incubation. In addition, the ratio of phosphorylated to total p85 protein was significantly reduced. Following 2 hours incubation, KZ-41 (10 μM) reversed the inhibitory effect of high glucose on phosphorylated Akt and p85 expression. The PI3K inhibitor LY294002 abolished the effect of KZ-41 on p85 and Akt activation.
Conclusions:
High glucose promoted REC apoptosis by inhibiting pro-survival signaling through the PI3K/Akt pathway. KZ-41 prevented glucose-induced cleaved caspase-3 activity via a PI3K-dependent mechanism. Together, these data provide further insight into the pro-survival mechanisms of quinic acid derivatives. Based on these data, current work is focused on the hypothesis that quinic acids exert their beneficial effects through one or more of the receptor tyrosine kinase family members, which activate cellular PI3K activity.
Keywords: 700 retinal neovascularization •
499 diabetic retinopathy •
715 signal transduction: pharmacology/physiology