Interleukin-1 beta-induced nitric oxide synthase expression by rat pancreatic beta-cells: evidence for the involvement of nuclear factor kappa B in the signaling mechanism.
Interleukin-1 beta-induced nitric oxide synthase expression by rat pancreatic beta-cells: evidence for the involvement of nuclear factor kappa B in the signaling mechanism.
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DOI：
10.1210/endo.136.11.7588208
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发表时间：
1995-11
期刊：
影响因子：
4.8
通讯作者：
G. Kwon;J. A. Corbett;Charles P. Rodi;Patrick Sullivan;M. L. McDaniel
中科院分区：
文献类型：
--
作者：
G. Kwon;J. A. Corbett;Charles P. Rodi;Patrick Sullivan;M. L. McDaniel
最近的证据表明，过量产生一氧化氮介导细胞因子诱导的胰岛胰岛素分泌抑制。目前的研究旨在描述白细胞介素-1 （IL-1）诱导的诱导型一氧化氮合酶（iNOS）在原代和转化大鼠胰腺β细胞中的表达过程中涉及转录因子NFkappaB的信号事件。由于纯化的原代β细胞数量的限制，主要使用胰岛素瘤细胞系RINm5F进行生化和分子研究。NFkappaB、二乙基二硫代氨基甲酸酯、吡啶二硫代氨基甲酸酯和n -乙酰半胱氨酸抑制剂在信使RNA、蛋白质和亚硝酸盐生成水平上阻止il -1诱导的iNOS表达。IL-1诱导NFkappaB从细胞质到细胞核的时间依赖性易位，在IL-1处理后约15-30分钟观察到最大易位，这是通过电泳迁移量转移试验确定的。含有NF κ B dna -蛋白复合物的条带的特异性通过与超过150倍的非放射性标记NF κ B寡核苷酸的竞争显示出来。利用免疫球蛋白G对NF κ b亚基p50和p65进行超移检测表明，该蛋白复合物含有p50和p65的异源二聚体。IL-1诱导的NF κ B转位被100微米（100微米）二乙基二硫氨基甲酸酯或100微米（100微米）吡咯烷二硫氨基甲酸酯阻断，进一步证实NF κ B在IL-1诱导大鼠胰腺β细胞iNOS中的关键作用。酪氨酸激酶的激活似乎先于NF κ B的激活，因为酪氨酸激酶抑制剂染料木素（100微米）可阻断il -1诱导的NF κ B易位。了解细胞因子诱导β细胞产生一氧化氮的信号转导途径，将为进一步评估一氧化氮在介导β细胞功能障碍中的作用提供干预策略。
Recent evidence indicates that overproduction of nitric oxide mediates cytokine-induced inhibition of insulin secretion by pancreatic islets. The current studies were designed to characterize signaling events involving the transcriptional factor NFkappaB in interleukin-1 (IL-1)-induced expression of inducible nitric oxide synthase (iNOS) by primary and transformed rat pancreatic beta-cells. Due to limitations of cell numbers of purified primary beta-cells, biochemical and molecular studies were performed primarily using the insulinoma cell line, RINm5F. Inhibitors of NFkappaB, diethyldithiocarbamate, pyrrolidine dithiocarbamate, and N-acetyl cysteine prevent IL-1-induced iNOS expression at the level of messenger RNA, protein, and nitrite generation. IL-1 induces a time-dependent translocation of NFkappaB from cytosol to nucleus, with maximal translocation observed approximately 15-30 min after IL-1 treatment, as determined by electrophoretic mobility shift assays. The specificity of the band containing the NF kappa B DNA-protein complex was shown by competition with a 150-fold excess of nonradiolabeled NF kappa B oligonucleotide. Supershift assays using immunoglobulins G against NF kappa b subunits p50 an p65 indicate that the protein complex contains a heterodimer of p50 and p65. IL-1-induced translocation of NF kappa B was blocked by 100 microns 100 microM diethyldithiocarbamate or 100 microM pyrrolidine dithiocarbamate, further establishing a critical role for NF kappa B in the induction of iNOS by IL-1 in rat pancreatic beta-cells. Activation of tyrosine kinase appears to precede NF kappa B activation, as the tyrosine kinase inhibitor genistein (100 microM) blocks IL-1-induced translocation of NF kappa B. An understanding of the signal transduction pathway of cytokine-induced nitric oxide generation by beta-cells will provide strategies of intervention to further evaluate the role of nitric oxide in mediating beta-cell dysfunction.