Diabetes Alters Sphingolipid Metabolism in the Retina

Diabetes Alters Sphingolipid Metabolism in the Retina A Potential Mechanism of Cell Death in Diabetic Retinopathy Todd E. Fox 1 , Xianlin Han 2 , Samuel Kelly 3 , Alfred H. Merrill, Jr. 3 , Rex E. Martin 4 , Robert E. Anderson 4 , Thomas W. Gardner 5 and Mark Kester 1 1 Department of Pharmacology and Cellular and Molecular Physiology, Penn State College of Medicine, Hershey, Pennsylvania 2 Division of Bioorganic Chemistry and Molecular Pharmacology, Department of Medicine, Washington University School of Medicine, St. Louis, Missouri 3 School of Biology, Georgia Institute of Technology, Atlanta, Georgia 4 Departments of Cell Biology and Ophthalmology, University of Oklahoma Health Sciences Center, Dean A. McGee Eye Institute, Oklahoma City, Oklahoma 5 Departments of Ophthalmology and Cellular and Molecular Physiology, Penn State College of Medicine, Hershey, Pennsylvania Address correspondence and reprint requests to Mark Kester, Department of Pharmacology H078, Penn State College of Medicine, Hershey, PA 17033. E-mail: mxk38{at}psu.edu Abstract Dysregulated sphingolipid metabolism causes neuronal cell death and is associated with insulin resistance and diseases. Thus, we hypothesized that diabetes-induced changes in retinal sphingolipid metabolism may contribute to neuronal pathologies in diabetic retinopathy. ESI-MS/MS was used to measure ceramide content and ceramide metabolites in whole retinas after 2, 4, and 8 weeks of streptozotocin-induced diabetes. After 4 and 8 weeks of diabetes, a ∼30% decrease in total ceramide content was observed, concomitant with a significant ∼30% increase in glucosylceramide levels in fed diabetic rats compared with their age-matched controls. Acute insulin therapy as well as a short-term lowering of glucose via fasting did not affect the increase in glucosylceramide composition. To assess the putative biological consequences of the increase in glucosylceramide composition, R28 retinal neurons were treated with glucosylceramide synthase inhibitors. Inhibiting glycosphingolipid metabolism increased insulin sensitivity in retinal neurons. Glycosphingolipid inhibitors augmented insulin-stimulated p70 S6kinase activity in the presence of inhibitory concentrations of high glucose or glucosamine. Inhibition of glycosphingolipid synthesis also suppressed glucosamine- and interleukin-1β–induced death. Consistent with these inhibitor studies, pharmacological accumulation of glycosphingolipids increased activation of the endoplasmic