Autophagy in retinal pigment epithelium cells exposed to an in vitro inflammatory model

BERMUDEZ, V.; TENCONI, P.E.; GIUSTO, N.M.; MATEOS, M.V.

Buenos Aires

Workshop; Buenos Aires Research Conference on Autophagy 2017. Molecular Mechanisms in Biology and Disease.; 2017

Lipopolysaccharide (LPS) can reach the retinal pigment epithelium (RPE) in patients with bacterial endophthalmitis, an unusual but serious ocular pathology. Our previous studies demonstrated the participation of classical phospholipases D (PLDs) in the LPS-induced inflammatory response of RPE cells. The aim of the present work is to study the autophagy process in RPE cells exposed to LPS.D407 and ARPE-19 human RPE cells were exposed to LPS (25 μg/ml) for 24 and 48 h. To study the role of the PLD pathway, cells were pre-incubated for 1 h with selective PLD1 (VU0359595) or PLD2 (VU0285655-1) inhibitors prior to LPS addition. Our results demonstrate that LPS reduced RPE cell viability (MTT reduction assay) after 48 h treatment. Western blot showed that LPS increased LC3II and reduced SQSTM1/p62 content and immunofluorescence assays showed an increment in LC3-positive punctate structures in cells exposed to LPS. In addition, PLD1 and PLD2 inhibition highly increased LC3II content and restored cell viability in LPS-exposed cells. On the contrary, autophagy inhibitors (3-MA and LY294002) worsened LPS-exposed cell viability. In summary, our results suggest that LPS treatment induces autophagy in RPE cells and that PLDs inhibitors could restore cell viability, possibly through the modulation of LPS-triggered autophagy.