Ceramide in beta cells apoptosis induced by gluco-lipotoxicity:possible role of cer flow for the biosynthesis of complex sphingolipids

CERAMIDE IN BETA CELLS APOPTOSIS INDUCED BY GLUCO-LIPOTOXICITY: POSSIBLE ROLE OF CER FLOW FOR THE BIOSYNTHESIS OF COMPLEX SPHINGOLIPIDS P. Giussani1, H. Le Stunff2, E. Gjoni1, J. Veret2, L. Riboni1, P. Viani1 1Department of Medical Chemistry, Biochemistry and Biotechnology, University of Milano, LITA Segrate, Via Fratelli Cervi, 93, 20090 Segrate (MI) Italy 2Unité Biologie Fonctionnelle et Adaptative - EAC CNRS 4413, Laboratoire de Biologie et Pathologie du Pancréas Endocrine, Université PARIS- DIDEROT (7),4, rue Marie-Andrée Lagroua Weill-Halle, 75205 PARIS Cedex 13 France Background: In type 2 diabetes the detrimental effects of chronic exposure to NEFA, in particular palmitate, on beta cell function and viability have been correlated to hyperglycaemia. De novo synthesis of ceramide (Cer) and ER stress are among the molecular pathways and regulators involved in these negative effects. In order to gain insight into the molecular mechanism(s) of gluco-lipotoxicity we studied the effect of palmitate and high glucose concentrations on ceramide metabolism in INS-1 cells. Results: The results obtained demonstrated that in INS-1 cells palmitate and glucose taken separately did not induce cell death, whereas the combined treatment with both nutrients resulted in an extensive cell apoptosis and this was associated to a significant increase of Cer levels. The presence of fumonisin-B1 partially reversed the apoptosis induced by the combined treatment with palmitate and glucose thus suggesting a role for ER-associated Cer in gluco-lipotoxicity in INS-1 cells. Metabolic studies show that treatment with palmitate results in the inhibition of Cer utilization for SM biosynthesis. Fluorescence microscopy studies suggest a reduced ER to Golgi flow of Cer. To evaluate if Cer accumulation could be due to a defect in the vesicular and/or CERT-mediated transport of Cer, we studied Cer metabolism in INS-1 cells silenced for CERT incubated with glucose in the presence or not of palmitate. In downregulated cells palmitate inhibited Cer utilization for SM biosynthesis; these results indicated that the vesicle-mediated transport is involved in the reduced ER to Golgi flow of Cer. Conclusions: Altogether these data suggest that the accumulation of Cer can be due to a decrease in utilization of newly synthesized Cer for SM biosynthesis thus supporting a role of ER-associated Cer in the regulation of beta cell death induced by gluco-lipotoxicity.