Glutathione-deficiency promotes basal hyperinsulinemia in the insulin secreting cell line INS-1 (832/13)

Glutathione (GSH) is an abundant antioxidant which maintains intracellular redox homeostasis by scavenging excess reactive oxygen species to abate oxidative stress. Previous studies have reported lowered GSH synthesis enzyme expression and GSH content in type 2 diabetic (T2D) islets. However, whether impairment to endogenous GSH synthesis and subsequent lowering of GSH content can be implicated in the progressive loss of β cell function in type 2 diabetes, independent of glucolipotoxicity remains unclear. This study sought to determine the impact of endogenous GSH synthesis on β cell function. Glutamate-cysteine ligase catalytic subunit (GCLC) is essential for GSH synthesis. To induce GSH deficiency, GCLC was inhibited in INS-1 (832/13) cells using buthionine sulfoximine (BSO). The 24-h treatment of INS-1 cells with BSO (250 μM) reduced GSH content to 33% of untreated, however, GSIS was unchanged. INS-1 cells require culture medium which contains 2-Mercaptoethanol (BME) to artificially bolster the cellular redox state. We removed BME from the cell culture medium to determine if further lowering of GSH content altered insulin secretion. The 24-h treatment of INS-1 cells with BME-free medium containing BSO further reduced GSH levels to 9% of untreated and elicited significantly higher basal insulin secretion. As expected, the concentration of reduced GSH, concentration of oxidized GSH (GSSG), the total GSH pool, redox state of the GSH/GSSG redox couple, and relative redox potential were significantly lower. We find that >90% GSH-deficiency induced basal hyperinsulinemia, independent of significant transcriptomic or metabolomic alteration. These results suggest endogenous GSH synthesis is directly related to β cell function. Additionally, basal hyperinsulinemia is a poorly-understood early change to β cell function in T2D, we provide a useful model for further investigation regarding the control that cellular redox state has over basal hyperinsulinemia.