Survival or death: a dual role of autophagy in stress-induced pericyte loss in diabetic retinopathy

Dongxu Fu, Jeremy Y. Yu, Shihe Yang, Mingyuan Wu, Samar M Hammad, Anna R. Connell, Mei Du, Junping Chen, Timothy J. Lyons

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Abstract

Aims/hypothesis
Intra-retinal extravasation and modification of LDL have been implicated in diabetic retinopathy: autophagy may mediate these effects.
Methods
Immunohistochemistry was used to detect autophagy marker LC3B in human and murine diabetic and non-diabetic retinas. Cultured human retinal capillary pericytes (HRCPs) were treated with in vitro-modified heavily-oxidised glycated LDL (HOG-LDL) vs native LDL (N-LDL) with or without autophagy modulators: green fluorescent protein–LC3 transfection; small interfering RNAs against Beclin-1, c-Jun NH(2)-terminal kinase (JNK) and C/EBP-homologous protein (CHOP); autophagy inhibitor 3-MA (5 mmol/l) and/or caspase inhibitor Z-VAD-fmk (100 μmol/l). Autophagy, cell viability, oxidative stress, endoplasmic reticulum stress, JNK activation, apoptosis and CHOP expression were assessed by western blots, CCK-8 assay and TUNEL assay. Finally, HOG-LDL vs N-LDL were injected intravitreally to STZ-induced diabetic vs control rats (yielding 50 and 200 mg protein/l intravitreal concentration) and, after 7 days, retinas were analysed for ER stress, autophagy and apoptosis.
Results
Intra-retinal autophagy (LC3B staining) was increased in diabetic vs non-diabetic humans and mice. In HRCPs, 50 mg/l HOG-LDL elicited autophagy without altering cell viability, and inhibition of autophagy decreased survival. At 100–200 mg/l, HOG-LDL caused significant cell death, and inhibition of either autophagy or apoptosis improved survival. Further, 25–200 mg/l HOG-LDL dose-dependently induced oxidative and ER stress. JNK activation was implicated in autophagy but not in apoptosis. In diabetic rat retina, 50 mg/l intravitreal HOG-LDL elicited autophagy and ER stress but not apoptosis; 200 mg/l elicited greater ER stress and apoptosis.
Conclusions
Autophagy has a dual role in diabetic retinopathy: under mild stress (50 mg/l HOG-LDL) it is protective; under more severe stress (200 mg/l HOG-LDL) it promotes cell death.
Original languageEnglish
Pages (from-to)2251-2261
Number of pages11
JournalDiabetologia
Volume59
Issue number10
DOIs
Publication statusPublished - 30 Jul 2016

Keywords

  • Apoptosis . Autophagy . Diabetic retinopathy .

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    Fu, D., Yu, J. Y., Yang, S., Wu, M., Hammad, S. M., Connell, A. R., Du, M., Chen, J., & Lyons, T. J. (2016). Survival or death: a dual role of autophagy in stress-induced pericyte loss in diabetic retinopathy. Diabetologia, 59(10), 2251-2261. https://doi.org/10.1007/s00125-016-4058-5