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Dongxu Fu, Shihe Yang, Mingyuan Wu, Mei Du, Junping Chen, Kenneth Wilson, Timothy Lyons; Autophagy in Modified LDL-induced Pericyte Loss in Diabetic Retinopathy. Invest. Ophthalmol. Vis. Sci. 2013;54(15):272. doi: https://doi.org/.
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We hypothesise that extravasation of plasma lipoproteins through damaged blood retinal barriers and their subsequent modification (glycation, oxidation) are more important in the propagation of diabetic retinopathy (DR) than circulating lipoproteins. We previously showed that extravasated, modified LDL (modeled by highly-oxidized glycated LDL, HOG-LDL) was associated with human retinal capillary pericyte (HRCP) apoptosis and autophagy mediated by oxidative and ER Stress. Little is known about role of autophagy in modified LDL-induced pericyte loss.
HRCP was exposed to HOG-LDL at different concentrations (up to 200mg/l) and times (up to 24h); native LDL (N-LDL) was used as control. Cells were transfected with plasmid GFP-LC3, si-RNA (si-CHOP or si-JNK) ,for 24-36h or pre-treated with inhibitors of autophagy (3-methyladenine and chloroquine) and caspase (z-VAD-fm), ER stress (4-phenyl butyric acid), or JNK (sp600125), each for 1h before LDL treatment. Autophagy, apoptosis, JNK activity, cell viability, oxidative stress and ER stress were determined by methods including immunocytochemistry, western blots, CCK-8 assay, TUNEL assay and cellular ROS measurement assay. To define the relevance of the results in vivo, immunohistochemistry was performed to detect LC3 and p-JNK from (a) mouse model of DR that mimics exposure of the retina to elevated glucose and elevated LDL levels, and (b) human subjects with and without diabetes and DR. Markers above were also measured by Western blot in human retinas.
Compared with N-LDL, HOG-LDL induced autophagy in pericytes, pJNK played an essential role. With lower concentrations of HOG-LDL (0-50mg/l), autophagy inhibited apoptosis promoting cell survival, but with higher concentrations (50-200mg/l), autophagy induced autophagic death and apoptosis. In diabetic hyperlipidemic mice (vs. animals with either condition alone), retinal autophagy was enhanced. Autophagy was also enhanced in diabetic human retina, but was not correlated with severity of DR.
In DR, extravasated and modified plasma lipoproteins play an important role once blood retinal barrier (BRB) leakage is established, and autophagy is implicated. The findings may lead to new treatments to inhibit the development of DR.
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