Tissue Inhibitor of Metalloproteinase-1 Prevents Cytokine-Mediated Dysfunction and Cytotoxicity in Pancreatic Islets and β-cells

Abstract

In addition to inhibiting matrix metalloproteinase-2 and matrix metalloproteinase-9 activity, recent studies suggest that tissue inhibitor of metalloproteinase (TIMP)-1 may inhibit apoptosis in various cell lines. To address this question in pancreatic islets and β-cells, we treated rat pancreatic islets and INS-1 cells with a high-dose combination of the cytokines interleukin (IL)-1β, tumor necrosis factor-α, and interferon-γ with or without the addition of TIMP-1 and TIMP-2 protein. Using flow cytometry, we quantitated DNA fragmentation to assess cellular apoptosis and confirmed these observations with DNA laddering experiments. Next, we transfected the mouse TIMP-1 gene into INS-1 cells and performed Western immunoblotting to demonstrate expression of TIMP-1 protein. We treated TIMP-1–expressing INS-1 cells with high-dose cytokines and again used flow cytometry to assess DNA fragmentation. We also evaluated the effect of TIMP-1 on IL-1β–induced inhibition of glucose-stimulated insulin secretion (GSIS) in freshly isolated rat pancreatic islets. Finally, we evaluated the effect of TIMP-1 on inducible nitric oxide synthase (iNOS) gene expression and nuclear factor (NF)-κB activity in INS-1 cells stimulated with high-dose cytokines. TIMP-1 but not TIMP-2 prevented cytokine-induced apoptosis and cytokine-mediated inhibition of GSIS in rat islets and β-cells. TIMP-1 mediated these effects by inhibiting cytokine activation of NF-κB, but it did not affect nitric oxide production or iNOS gene expression. Therefore, TIMP-1 may be an ideal gene to prevent cytokine-mediated β-cell destruction and dysfunction in models of type 1 diabetes and islet transplantation rejection.