Plasmalogens and cell-cell communication between retinal glial cells

Purpose: Plasmalogens (or ether-lipids) are a particular class of glycerophospholipids characterized by the presence of a vinyl-ether bond at the sn-1 position of the glycerol backbone and by the preferential esterification of polyunsaturated fatty acids (PUFAs) at the sn-2 position. Plasmalogens are found at high concentrations in retinal macroglial cells such as Müller cells and astrocytes. Müller cells and astrocytes are known to communicate by the way of intracellular calcium waves and gap junctions. Connexin 43 is the major protein of these communicating junctions. The aim of this study was to determine the effect of plasmalogens depletion on communication between astrocytes and Müller cells. Methods: Primary retinal Müller cells were isolated from eyes of Wistar rats aged from 9 days whereas astrocytes were obtained from cortex of 21-days old animals. SiRNA against DHAPA-AT -the key enzyme of plasmalogen biosynthesis- were used to decrease the cell content in plasmalogen. After lipid extraction, the concentrations of plasmalogens in cell membranes were monitored by gas chromatography coupled to flame ionization detection. Communication between astrocytes and Müller cells was studied by calcium imaging. Changes in the expression cellular proteins, including connexin 43, were evaluated by proteomics and western-blotting. Results: Lipidomic analyses confirmed that plasmalogens are concentrated in retinal macroglial cells (17% and 13% of total phospholipids for Müller cells and astrocytes, respectively) when compared to the entire retina (7% of total phospholipids). SiRNA against DHAP-AT led to a diminution of plasmalogen content of more than 50%. Decreasing the concentrations of plasmalogens in membranes of glial cells impacted cell-cell communication through decrease in the expression of connexion 43 by more than 50%. Western-blot analyses showed modifications in the expression/phosphorylation of MAPK proteins. Conclusions: Plasmalogens may influence cell-cell communication between retinal Müller cells and astrocytes. As retinal macroglial cells play crucial roles in preserving neuronal integrity and in proper vascular development, modifications of retinal plasmalogen levels are likely to be at the origin of many retinal diseases