Islet neogenesis associated protein (INGAP) protects pancreatic β cells from IL-1β and IFNγ-induced apoptosis

The goal of this study was to determine whether recombinant Islet NeoGenesis Associated Protein (rINGAP) and its active core, a pentadecapeptide INGAP(104-118) (Ingap-p), protect beta cells against cytokine-induced death. INGAP has been shown to induce islet neogenesis in diabetic animals, to stimulate beta-cell proliferation and differentiation, and to improve islet survival and function. Importantly, Ingap-p has shown promising results in clinical trials for diabetes (phase I/II). However, the full potential of INGAP and its mechanisms of action remain poorly understood. Using rat insulinoma cells RINm5F and INS-1 treated with interleukin-1 beta (IL-1 beta) and interferon-gamma (IFN-gamma), we demonstrate here that both rINGAP and Ingap-p inhibit apoptosis, Caspase-3 activation, inducible nitric oxide synthase (iNOS) expression and nitric oxide (NO) production, and explore the related signaling pathways. As expected, IL-1 beta induced nuclear factor kappa B (NF-kappa B), p38, and JNK signaling, whereas interferon-gamma (IFN-gamma) activated the JAK2/STAT1 pathway and potentiated the IL-1 beta effects. Both rINGAP and Ingap-p decreased phosphorylation of IKK alpha/beta, IkB alpha, and p65, although p65 nuclear translocation was not inhibited. rINGAP, used for further analysis, also inhibited STAT3, p38, and JNK activation. Interestingly, all inhibitory effects of rINGAP were observed for the cytokine cocktail, not IL-1 beta alone, and were roughly equal to reversing the potentiating effects of INF gamma. Furthermore, rINGAP had no effect on IL-1 beta/NF-kappa B-induced gene expression (e.g., Ccl2, Sod2) but downregulated several IFN gamma-stimulated (Irf1, Socs1, Socs3) or IFN gamma-potentiated (Nos2) genes. This, however, was observed again only for the cytokine cocktail, not IFN gamma alone, and rINGAP did not inhibit the IFN gamma-induced JAK2/STAT1 activation. Together, these intriguing results suggest that INGAP does not target either IL-1 beta or IFN gamma individually but rather inhibits the signaling crosstalk between the two, the exact mechanism of which remains to be investigated. In summary, our study characterizes the anti-inflammatory effects of INGAP, both protein and peptide, and suggests a new therapeutic utility for INGAP in the treatment of diabetes.

Automated summary

This study demonstrates that both rINGAP and Ingap-p can inhibit cytokine-induced cell apoptosis and related signaling pathways, suggesting that INGAP may act by influencing the crosstalk between cytokine signaling pathways.