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

BERMÚDEZ, VICENTE; TENCONI, PAULA ESTEFANÍA; GIUSTO, NORMA MARÍA; MATEOS, MELINA VALERIA

Buenos Aires

Workshop; Buenos Aires research conferences on autophagy. Molecular mechanisms in biology and diseases?; 2017

International Centre for Genetic Engineering and Biotechnology

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.