Cytokine-Induced Impairment of Mitochondrial Respiration is Partly Dependent on Nitric Oxide Production

Pro-inflammatory cytokines contribute to beta-cell dysfunction in type 1 diabetes (T1D) by inhibiting mitochondrial respiration and causing cell death by apoptosis. Previous studies in rodent models have demonstrated that nitric oxide (NO), generated by inducible NO synthase (iNOS), is a crucial mediator of cytokine-induced beta cell death. Moreover, it has been speculated that NO plays a role in the respiratory defects caused by cytokines. It remains elusive, however, whether cytokines also cause respiratory defects in human islets and if so, whether NO is involved. In this study, we investigated the role of NO for cytokine-induced mitochondrial dysfunction in rat INS-1E cells and examined if cytokines and NO affect human islet mitochondrial respiration.
Human pancreatic islets and rat INS-1E cells were exposed for 48h and 24h, respectively, to IL-1β and IFN-γ to mimic early beta-cell destruction in T1D. Mitochondrial bioenergetics were analysed by Seahorse XFe24 technology and gene expression of metabolic relevant genes was measured by real-time PCR.
Human islets exposed to cytokines for 48h showed a significant reduction in glucose-stimulated respiration. Further, higher basal mitochondrial oxygen consumption was observed, which was not associated with increased ATP production. The NO synthase inhibitor NG-Methyl-L-Arginine acetate salt (L-NMMA) alone tended to decrease the respiratory response of human islets to glucose, but partly counteracted the effects of cytokines. Cytokine-induced changes in gene expression were independent of NO in human islets.
A 24-h cytokine exposure strongly inhibited mitochondrial respiration in INS-1E cells. Cytokine treatment reduced the coupling efficiency of respiration, which was mirrored by impaired glucose-induced ATP production. Inhibition of cytokine-induced NO production by L-NMMA restored the coupling efficiency of mitochondrial respiration, whereas it did not prevent impairments in glucose-induced respiration. Acute cytokine injection did not affect INS-1E oxygen consumption up to 8h post treatment, proposing that cytokine-induced inhibition of mitochondrial respiration is a later event triggered by the cytokine signalling pathway. Cytokines reduced the expression of metabolic enzymes in an NO-dependent manner in INS-1E cells.
In conclusion, cytokines reduce glucose-induced respiration in both human islets and INS-1E cells, suggesting impaired mitochondrial respiration, which may contribute to islet failure in T1D. In both model systems, NO seems to play a role for mitochondrial dysfunction. Further studies are needed to fully clarify the possible pathogenic role of mitochondrial dysfunction in T1D and the underlying mechanisms.