Metabolic stress remodels the Golgi-sphingolipid-lipid droplet axis in INS-1E β-cells: an integrated molecular signature of early β-cell vulnerability.

backgroundGlucotoxicity, lipotoxicity, and glucolipotoxicity contribute to pancreatic β-cell dysfunction in type 2 diabetes. Although endoplasmic reticulum stress has been widely studied, the response of the Golgi apparatus to metabolic stress and its relationship with sphingolipid metabolism, lipid droplets, and insulin granule-associated pathways remain poorly understood. We examined these processes together in INS-1E β-cells exposed to different forms of metabolic stress. METHODS AND

resultsINS-1E cells were exposed to high glucose (33 mM), palmitic acid (0.5 mM), or both for 24 h. Cellular metabolic activity was measured by MTT assay. The expression of CREB3, ARF4, SMPD1, SMPD2, perilipin-3, ATGL, PC1/3, Syntaxin 6, and Syntaxin 12/13 was assessed by Western blotting, while NBD-ceramide fluorescence was examined by fluorescence microscopy. Metabolic stress affected Golgi stress-associated proteins and increased SMPD1 and SMPD2 expression. Increased NBD-ceramide fluorescence was consistent with changes in ceramide handling. Perilipin-3 increased without a corresponding change in ATGL, suggesting a shift toward lipid sequestration. High glucose alone largely preserved metabolic activity, whereas palmitate and combined glucose-palmitate exposure reduced both metabolic activity and PC1/3 expression. Syntaxin 6 and Syntaxin 12/13 remained unchanged.

conclusionsMetabolic stress was associated with concurrent changes in Golgi stress-associated proteins, sphingolipid metabolism, lipid droplet regulation, and PC1/3 expression in INS-1E β-cells. These findings demonstrate coordinated molecular alterations involving Golgi stress-associated proteins, sphingolipid metabolism, lipid droplet regulation, and PC1/3 expression during metabolic stress. Functional and lipidomic studies are now needed to determine how these molecular changes contribute to β-cell dysfunction.