Glucocorticoids induce retinal toxicity through mechanisms mainly associated with paraptosis.
OBJECTIVE
Corticosteroids have recorded beneficial clinical effects and are widely used in medicine. In ophthalmology, besides their treatment benefits, side effects, including ocular toxicity have been observed especially when intraocular delivery is used. The mechanism of these toxic events remains, however, poorly understood. In our present study, we investigated the mechanisms and potential pathways of corticosteroid-induced retinal cell death.


METHODS
Rats were sacrificed 24 h and 8 days after an intravitreous injection of 1 microl (40 microg) of Kenacort Retard. The eyes were processed for ultra structure analysis and detection of activated caspase-3, cytochrome-C, apoptosis-inducing factor (AIF), LEI-L-Dnase II, terminal transferase dUTP nick end labeling (TUNEL), and microtubule-associated protein 1-light chain 3 (MAP-LC3). In vitro, rat retinal pigment epithelial cells (RPE), retinal Müller glial cells (RMG) and human ARPE-19 cells were treated with triamcinolone acetonide (TA) or other glucocorticoids. Cell viability was quantified by 3-(4,5-dimethylthiazol-2-yl)-2,5 phenyltetrazolium bromide test (MTT) assay and cell counts. Nuclei staining, TUNEL assay, annexin-V binding, activated caspase-3 and lactate dehydrogenase (LDH) production characterized cell death. Localization of cytochrome-C, AIF, LEI-and L-Dnase II, and staining with MAP-LC3 or monodansylcadaverine were also carried out. Finally, ARPE-19 cells transfected with AIP-1/Alix were exposed to TA.


RESULTS
In vitro incubation of retinal cell in the presence of corticosteroids induced a specific and dose-dependent reduction of cell viability. These toxic events were not associated with the anti-inflammatory activity of these compounds but depended on the hydro solubility of their formulation. Before cell death, extensive cytoplasmic vacuolization was observed in the retinal pigment epithelial (RPE) cells in vivo and in vitro. The cells however, did not show known caspase-dependent or caspase-independent apoptotic reactions. These intracellular vacuoles were negative for MAP-LC3 but some stained positive for monodansylcadaverine. Furthermore, over expression of AIP-1/Alix inhibited RPE cell death.


CONCLUSIONS
These observations suggest that corticosteroid-induced retinal cell death may be carried out mainly through a paraptosis pathway.
Valamanesh
Fatemeh
F
INSERM, Physiopathology of ocular diseases: Therapeutic innovations, Institut des Cordeliers, Université René Descartes, Paris, France.
Torriglia
Alicia
A
Savoldelli
Michéle
M
Gandolphe
Christelle
C
Jeanny
Jean-Claude
JC
BenEzra
David
D
Behar-Cohen
Francine
F
D000305:Adrenal Cortex Hormones / Q000008:administration & dosage / Q000737:chemistry / Q000506:poisoning*; D000818:Animals; D017209:Apoptosis; D001343:Autophagy; D016923:Cell Death; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003593:Cytoplasm / Q000473:pathology; D004305:Dose-Response Relationship, Drug; D006801:Humans; D007267:Injections; D008854:Microscopy, Electron; D009336:Necrosis; D009457:Neuroglia / Q000187:drug effects; D010857:Pigment Epithelium of Eye / Q000187:drug effects / Q000473:pathology; D011136:Polysorbates / Q000506:poisoning; D051381:Rats; D011917:Rats, Inbred Lew; D012160:Retina / Q000187:drug effects* / Q000473:pathology* / Q000503:physiopathology; D012995:Solubility; D014222:Triamcinolone Acetonide / Q000506:poisoning; D014617:Vacuoles / Q000473:pathology; D014822:Vitreous Body; D014867:Water
D000305:Adrenal Cortex Hormones; D011136:Polysorbates; D014867:Water; D014222:Triamcinolone Acetonide