title: Discrete and complementary mechanisms of protection of beta-cells against cytokine-induced and oxidative damage achieved by bcl-2 overexpression and a cytokine selection strategy.
abstract: We have been investigating the potential utility of engineered cell lines as surrogates for primary islet cells in treatment of type 1 diabetes. To this end, two strategies that have emerged for procuring cell lines with resistance to immune-mediated damage are 1) selection of cytokine-resistant cell lines by growth of INS-1 insulinoma cells in iteratively increasing concentrations of interleukin (IL)-1beta + gamma-interferon (IFN-gamma), and 2) stable overexpression of the anti-apoptotic gene bcl-2 in INS-1 cells. Herein, we show that bcl-2-overexpressing cells are resistant to the cytotoxic effects of reactive oxygen and nitrogen species (ROS/RNS), but are only modestly protected against high concentrations of IL-1beta + INF-gamma, whereas the converse is true in cytokine selected cells. We also found that the combination of bcl-2 expression and cytokine selection confers a broader spectrum of resistance than either procedure alone, such that the resultant cells are highly resistant to cytokines and ROS/RNS, with no impairment in glucose-stimulated insulin secretion. INS-1-derived cells with combined bcl-2 expression and cytokine selection are also more resistant to damage induced by coculture with mitogen-activated peripheral blood mononuclear cells. Surprisingly, application of the cytokine selection procedure to bcl-2-overexpressing cells does not result in impairment of nuclear factor-kappaB translocation, iNOS expression, and NO production, as clearly occurs upon application of the selection procedure to cells without bcl-2 overexpression. Further investigation of the diverse pathways involved in the development of cytokine and ROS/RNS resistance may define simplified and specific strategies for preservation of beta-cell mass.
authors.affiliation: Touchstone Center for Diabetes Research, Department of Biochemistry and Biomedical Engineering Program, University of Texas Southwestern Medical Center, Dallas, TX, USA.
authors.forename: Veronique Vien
authors.initials: VV
authors.lastname: Tran
authors.forename: Guoxun
authors.initials: G
authors.lastname: Chen
authors.forename: Christopher B
authors.initials: CB
authors.lastname: Newgard
authors.forename: Hans E
authors.initials: HE
authors.lastname: Hohmeier
mesh_terms: D000818:Animals; D001483:Base Sequence; D016207:Cytokines / Q000494:pharmacology* / Q000502:physiology; D017931:DNA Primers; D003922:Diabetes Mellitus, Type 1 / Q000601:surgery; D019254:Genes, bcl-2 / Q000235:genetics*; D006801:Humans; D007340:Insulinoma; D007371:Interferon-gamma / Q000494:pharmacology; D007375:Interleukin-1 / Q000494:pharmacology; D007515:Islets of Langerhans / Q000187:drug effects / Q000276:immunology* / Q000502:physiology*; D008070:Lipopolysaccharides / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism; D020030:Nitric Oxide Donors / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects; D010190:Pancreatic Neoplasms; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D026423:S-Nitroso-N-Acetylpenicillamine / Q000494:pharmacology; D023822:Tissue Engineering; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: D016207:Cytokines; D017931:DNA Primers; D007375:Interleukin-1; D008070:Lipopolysaccharides; D020030:Nitric Oxide Donors; D019253:Proto-Oncogene Proteins c-bcl-2; D011994:Recombinant Proteins; D009569:Nitric Oxide; D026423:S-Nitroso-N-Acetylpenicillamine; D007371:Interferon-gamma