Sphingolipids, emerging mediators in proliferative retinopathies?

Abstract

Müller glial cells (MGC) and retinal pigment epithelium (RPE) cells are crucial for preserving retina homeostasis but their reactive response contributes to the progress of retina proliferative diseases, as diabetic retinopathy. We demonstrated that sphingosine-1-phosphate (S1P) and ceramide-1-phosphate (C1P) regulate MGC and RPE cell migration. We now investigated whether they regulate viability and fibrotic and inflammatory changes in these cells. Incubation of RPE cell cultures with 5 µM S1P or 10 µM C1P for 24 h increased mRNA levels of IL-6 and IL-8, inflammatory interleukins, and α-smooth muscle actin (α-SMA), an epithelial mesenchymal transition marker. C1P-induced migration of RPE cells was not affected by inhibiting C1P endogenous synthesis with NVP-231 (NVP), a ceramide kinase inhibitor, but was markedly reduced when S1P synthesis was blocked with an inhibitor of sphingosine kinase 1 (SphK1), the enzyme involved in S1P synthesis. Interestingly, C1P addition enhanced SphK1 transcription. These results imply S1P and C1P promote RPE cell migration, pro-inflammatory and pro-fibrotic changes, and S1P endogenous synthesis is essential for RPE cell migration. To evaluate the role of C1P in MGC and RPE cells in an in vitro model of high glucose (HG)-induced damage, we pre-treated primary MGC cultures, obtained from rat retinas, and D407 cells, a RPE cell line, with NVP or its vehicle, and then exposed them to HG (30 mM), normal glucose (NG, 5 mM) or an osmotic control (25 mM Mannitol + NG) for 24-72 h. NVP pre-treatment induced morphological changes both in MGC and RPE cells exposed to HG and affected their viability, decreasing the amount of cell nuclei, compared to NG and Mannitol-treated cultures, suggesting C1P synthesis is required to protect MGC and RPE cells from the oxidative stress induced by HG. As a whole, these results suggest S1P and C1P play multiple roles in proliferative retinopathies, promoting inflammatory changes but also preventing cell death.