Memantine, Simvastatin, and Epicatechin Inhibit 7-Ketocholesterol-induced Apoptosis in Retinal Pigment Epithelial Cells But Not Neurosensory Retinal Cells In Vitro

Abstract

Purpose: 7-ketocholesterol (7kCh), a natural byproduct of oxidation in lipoprotein deposits, is implicated in the pathogenesis of diabetic retinopathy and age-related macular degeneration (AMD). This study was performed to investigate whether several clinical drugs can inhibit 7kCh-induced caspase activation and mitigate its apoptotic effects on retinal cells in vitro.
 Method: Two populations of retinal cells, human retinal pigment epithelial cells (ARPE-19) and rat neuroretinal cells (R28), were exposed to 7kCh in the presence of the following inhibitors: Z-VAD-FMK (pan-caspase inhibitor), simvastatin, memantine, epicatechin, and Z-IETD-FMK (caspase-8 inhibitor) or Z-ATAD-FMK (caspase-12 inhibitor). Caspase-3/7, -8, and -12 activity levels were measured by fluorochrome caspase assays to quantify cell death. IncuCyte live-cell microscopic images were obtained to quantify cell counts.
 Results: Exposure to 7kCh for 24 hours significantly increased caspase activities for both ARPE-19 and R28 cells (P < 0.05). In ARPE cells, pretreatment with various drugs had significantly lower caspase-3/7, -8, and -12 activities, reported in % change in mean signal intensity (msi): Z-VAD-FMK (48% decrease, P < 0.01), memantine (decreased 47.8% at 1 μM, P = 0.0039 and 81.9% at 1 mM, P < 0.001), simvastatin (decreased 85.3% at 0.01 μM, P < 0.001 and 84.8% at 0.05 μM , P < 0.001) or epicatechin (83.6% decrease, P < 0.05), Z-IETD-FMK (68.1% decrease, P < 0.01), and Z-ATAD-FMK (47.7% decrease, P = 0.0017). In contrast, R28 cells exposed to 7kCh continued to have elevated caspase- 3/7, -8, and -12 activities (between 25.7% decrease and 17.5% increase in msi, P > 0.05) regardless of the pretreatment.
 Conclusion: Several current drugs protect ARPE-19 cells but not R28 cells from 7kChinduced apoptosis, suggesting that a multiple-drug approach is needed to protect both cells types in various retinal diseases.

The retina-brain axis and diabetic retinopathy

“…163,164 Deviation of glutamate signaling occurs in the diabetic brain 165 and diabetic retina. 166 Interestingly, memantine has been able to protect RGCs, reduce vitreous VEGF, and reduce BRB breakdown in models of DR. 167 Meanwhile, it could protect RPE cells in in-vitro studies on DR. 168 Some other neuroprotective factors can protect the neural retina against glutamate; “Pituitary adenylate cyclase activating polypeptide (PACAP)” is one of them. 169 So more investigations on the efficacy of memantine and PACAP in the retina-brain axis are needed.…”

7-Ketocholesterol Promotes Retinal Pigment Epithelium Senescence and Fibrosis of Choroidal Neovascularization via IQGAP1 Phosphorylation-Dependent Signaling

“…Evidence links 7KC with AMD [ 9 , 10 , 11 ]. Compared to younger donor eyes, 7KC levels in RPE-associated drusen were significantly higher in older donor eyes [ 4 ].…”

“…In cultured ARPE-19 cells, 7KC induced an inflammatory response involving toll-like receptor 4 (TLR4)-mediated endoplasmic reticulum stress [ 13 ]. 7KC was also reported to induce apoptosis through caspase-dependent signaling in ARPE-19 [ 10 ]. Counteracting the effects of 7KC in several age-related diseases has been considered, including through neutraceutical intervention [ 14 ].…”