Effect of Acute IFN-α Exposure on β-cell Survival and Function in the Absence of Functional Autophagy

Abstract

Background and Hypothesis: It has been demonstrated that the protective degradative process of autophagy is defective in the context of human T1D. Additionally, a type I interferon-inducible transcriptional signature has been detected in blood cells of children genetically at risk for T1D prior to the appearance of autoantibodies. Preliminary data demonstrate that autophagy can be stimulated by IFN-α in β-cells of non-diabetic mice, but this response is defective in β-cells of nonobese diabetic (NOD) mice, a mouse model for autoimmune diabetes. We hypothesize that impaired autophagy and exposure to IFN-α will lead to reduced β-cell survival and impaired function.

Methods:
Bafilomycin A, a lysosomal inhibitor, was used to inhibit autophagy in INS-1 832/13 cells and pancreatic human islets. We then utilized western blot analysis, Caspase-Glo® 3/7 Assay, and Insulin HTRF Assay to assess β-cell survival and function in response to IFN-α exposure.

Results: Western blot analysis was used to monitor inhibition of autophagy and stimulation of IFN-α pathway in INS-1 832/13 cells (n=4) and pancreatic human islets (n=1). An increase in LC3II/LC3I ratio was observed after treatment with Baf A, indicating accumulation of autophagosomes. Phosphorylation of STAT2 was observed in IFN-α treated cells, indicating stimulation of the IFN-α pathway. Impaired autophagy and IFN-α exposure did not induce apoptosis in INS-1 832/13 cells (n=2). Following Baf A and IFN-α treatment, glucose stimulated insulin secretion was measured. Glucose stimulated insulin secretion is decreased in INS-1 832/13 cells following Baf A and IFN-α treatment (n=2).

Conclusions: These data suggest that inhibition of autophagy followed by an acute treatment with IFN-α does not induce significant cell death, but modestly impairs β-cell function. Future studies will determine the effects of prolonged IFN-α exposure on β-cell survival and function as well as investigate other potential mechanisms of cell death.