IFN-a Induces Heterogenous ROS Production in Human ß-Cells

Type 1 diabetes (T1D) is a multifactorial disease involving genetic and environmental factors, including viral infection. We investigated the impact of interferon alpha (IFN-a), a cytokine produced during the immune response to viral infection or the presence of un-edited endogenous double-stranded RNAs, on human beta cell physiology. Intravital microscopy on transplanted human islets using a beta cell-selective reactive oxygen species (ROS) biosensor (RIP1-GRX1-roGFP2), revealed a subset of human beta cells that acutely produce ROS in response to IFN-a. Comparison to Integrated Islet Distribution Program (IIDP) phenotypic data revealed that while all donors had functional islets, donors with lower BMI and HbA1c had more ROS accumulating cells. In vitro IFN-a treatment of human islets similarly elicited a heterogenous increase in superoxide production that originated in the mitochondria. To determine the unique molecular signature predisposing cells to IFN-a stimulated ROS production, we flow sorted human islets treated with IFN-a. RNA sequencing identified genes involved in inflammatory and immune response in the ROS-producing cells. Importantly, rapid stimulation of key antiviral response genes by IFN-a was dependent on mitochondrial ROS elevation. Comparison with single cell RNA-Seq datasets available through the Human Pancreas Analysis Program (HPAP) showed that genes upregulated in ROS-producing cells are enriched in beta cells from nondiabetic and autoantibody positive donors rather than T1D donors. Combined, these data suggest that IFN-a stimulates mitochondrial ROS production in healthy human beta cells, potentially predicting a more efficient antiviral response if the processes to respond to the ROS elevation are intact. Overall design: Human islets were obtained from organ donors and procured through either the Integrated Islet Distribution Program (IIDP) or the Alberta Diabetes Institute (ADI) then treated acutely with recombinant human IFN-? for 1h in the presence of dihydroethidium (DHE) to detect reactive oxygen species (ROS). Dispersed cells were flow sorted into ROS-positive and ROS-negative populations then RNA-sequencing was performed on the populations.