Tissue inhibitor of metalloproteinase-1 prevents cytokine-mediated dysfunction and cytotoxicity in pancreatic islets and beta-cells.
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 beta-cells, we treated rat pancreatic islets and INS-1 cells with a high-dose combination of the cytokines interleukin (IL)-1beta, tumor necrosis factor-alpha, and interferon-gamma 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-1beta-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)-kappaB 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 beta-cells. TIMP-1 mediated these effects by inhibiting cytokine activation of NF-kappaB, but it did not affect nitric oxide production or iNOS gene expression. Therefore, TIMP-1 may be an ideal gene to prevent cytokine-mediated beta-cell destruction and dysfunction in models of type 1 diabetes and islet transplantation rejection.
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Leslie and Susan Gonda (Goldschmeid) Diabetes and Genetics Research Center, Department of Diabetes, Endocrinology, and Metabolism, City of Hope National Medical Center, Duarte, California 91010, USA.
Sun
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Scott
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Bleich
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D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D016207:Cytokines / Q000037:antagonists & inhibitors / Q000633:toxicity*; D053938:DNA Fragmentation / Q000187:drug effects; D005434:Flow Cytometry; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D017930:Genes, Reporter; D005947:Glucose / Q000494:pharmacology; D066298:In Vitro Techniques; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007371:Interferon-gamma / Q000633:toxicity; D007375:Interleukin-1 / Q000633:toxicity; D007515:Islets of Langerhans / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D008156:Luciferases / Q000235:genetics; D051379:Mice; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000235:genetics; D052247:Nitric Oxide Synthase Type II; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000235:genetics / Q000494:pharmacology* / Q000502:physiology*; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000494:pharmacology*; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D016207:Cytokines; D007328:Insulin; D007375:Interleukin-1; D016328:NF-kappa B; D011994:Recombinant Proteins; D019715:Tissue Inhibitor of Metalloproteinase-1; D014409:Tumor Necrosis Factor-alpha; D019716:Tissue Inhibitor of Metalloproteinase-2; D007371:Interferon-gamma; D008156:Luciferases; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; C496318:Nos2 protein, rat; D005947:Glucose