Objective: Recent estimates by the International Diabetes Federation (IDF) indicate that nearly 589 million adults are living with diabetes, which is expected to increase to 853 million by 2050. Such high rates in incidence of diabetes warrant an immediate need for not only understanding the mechanisms underlying the metabolic dysregulation and demise of the islet β-cell, but also for increased efforts to develop novel therapeutics for halting β-cell defects and the onset of diabetes and its associated complications. Compelling experimental evidence implicates intracellular generation of ceramide (CER), a biologically active sphingolipid, as a trigger in the onset of β-cell demise under conditions of chronic glucolipotoxic (GLT) stress. Herein, using quantitative lipidomics approach we quantified specific CERs in pancreatic β-cells exposed to GLT conditions. Relative contributory roles of de novo biosynthesis of CERs were assessed using myriocin, a known inhibitor of de novo CER biosynthetic pathway. Methods: Insulin secreting clonal (INS-1 832/13) β-cells were cultured under basal (2.5mM glucose) or GLT (20mM glucose plus 0.5mM palmitate) in the absence or presence of myriocin (5µM) for 24 hrs. Cellular sphingolipids were extracted with isopropanol-ethyl acetate and analyzed for ceramides using precursor ion scans, m/z 264 following HPLC (Prominence XR system, Shimadzu) separation using Targa C8 column. The data were collected using Analyst 1.6.2 software, and the MRM transition chromatograms were quantified by MultiQuant software. The internal standard (C17 ceramide) signal in each chromatogram was used for normalization for recovery as well as relative quantitation of each analyte. Results: Quantitative lipidomics analysis indicated a significant increase in specific CERs, namely dihydroCERs (18:0/24:0 and 18:1/26:0) and hexosylCERs (18:1/16:0; 18:1/18:0; 18:1/22:0; and 18:1/24:0) in pancreatic β-cells exposed to GLT conditions. Co-provision of myriocin, a known inhibitor of serine palmitoyl transferase (de novo biosynthesis of CERs), markedly attenuated biosynthesis of dihydro- and hexosylCERs in pancreatic β-cells seen under the duress of GLT stress. Conclusions: GLT stress induces de novo biosynthesis of specific dihydro- and hexosylCERs in pancreatic β-cells. Targeting specific enzymes involved in biosynthesis or degradation might be useful in modulating levels of these ceramide species and improving β-cell function in in vitro and in vivo models of GLT stress. Funding: AK is supported by MERIT (I01BX004663; I01BX006377) and Senior Research Career Scientist (K6 BX005383) awards from the U.S. Department of Veterans Affairs. KRM is supported in part by grants from NIH S10RR027926 and S10OD032292 to the Lipidomics Core Facility of Wayne State University. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.
Kowluru et al. (Fri,) studied this question.