Interleukin-1 beta induced changes in the protein expression of rat islets: A computerized database
Interleukin-1 beta induced changes in the protein expression of rat islets: A computerized database
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DOI：
10.1002/elps.1150181136
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发表时间：
1997-10-01
期刊：
影响因子：
2.9
通讯作者：
Nerup, J
中科院分区：
文献类型：
--
作者：
Andersen, HU;Fey, SJ;Nerup, J
胰岛素依赖型糖尿病是由胰岛中β细胞的自身免疫破坏引起的。细胞因子白细胞介素1抑制胰岛素释放，并对分离的大鼠胰岛中的β细胞具有选择性细胞毒性。触发免疫应答的抗原以及白细胞介素1介导的β细胞毒性的细胞内作用机制尚不清楚。然而，以前的研究发现β细胞破坏与蛋白质合成的改变有关。因此，胰岛蛋白质的二维（2-D）凝胶电泳可能是促进胰岛素依赖型糖尿病分子发病机制研究的重要工具。胰岛蛋白质的二维凝胶电泳可能导致（i）确定细胞因子诱导的特定胰岛蛋白质的定性和定量变化，（ii）确定调节细胞因子作用的药物的作用，以及（iii）鉴定启动免疫破坏的主要胰岛蛋白抗原β细胞。因此，本研究的目的是建立数据库（DB）的所有可重复检测的蛋白质斑点的10%和15%丙烯酰胺2-D凝胶的新生大鼠胰岛（10%和15%DB），在标准化的培养条件下标记。在15%等电聚焦（IEF）和非平衡pH梯度电泳（NEPHGE）DB中，5个凝胶中分别有1235和557个斑点，而在10% IEF和NEPHGE DB中，5个凝胶中分别有995和378个斑点，斑点检测的重现性在75.2%和91.7%之间。在两种DB中，所有凝胶中存在的斑点的积分光密度百分比（%IOD的CV%）的平均变异系数在42.4%和45.7%之间。当在不同日期在连续凝胶组中分析相同样品时（批间分析），%IOD的平均CV%为35.5%-36.1%。当在一组凝胶中重复分析相同样品时（试验内分析），IEF凝胶中%IOD的平均CV%为30.2%，而NEPHGE凝胶中%IOD的平均CV%为45.7%。向培养物中加入白细胞介素-1 β（IL-1 β）导致10%凝胶中105种蛋白质的统计学显著调节或从头合成。总之，我们提出了第一个10%和15%丙烯酰胺2-D凝胶蛋白质数据库的新生大鼠胰岛，并证明其用途，以确定蛋白质的表达改变的IL-1 β *。
Insulin-dependent diabetes mellitus is caused by an autoimmune destruction of the beta-cells in the islets of Langerhans. The cytokine interleukin 1 inhibits insulin release and is selectively cytotoxic to beta-cells in isolated pancreatic rat islets. The antigen(s) triggering the immune response as well as the intracellular mechanisms of action of interleukin 1-mediated beta-cell cytotoxicity are unknown. However, previous studies have found an association of beta-cell destruction with alterations in protein synthesis. Thus, two-dimensional (2-D) gel electrophoresis of pancreatic islet proteins may be an important tool facilitating studies of the molecular pathogenesis of insulin-dependent diabetes mellitus. 2-D gel electrophoresis of islet proteins may lead to (i) the determination of qualitative and quantitative changes in specific islet proteins induced by cytokines, (ii) the determination of the effects of agents modulating cytokine action, and (iii) the identification of primary islet protein antigen(s) initiating the immune destruction of the beta-cells. Therefore, the aim of this study was to create databases (DB) of all reproducibly detectable protein spots on 10% and 15% acrylamide 2-D gels of neonatal rat islets (10% and 15% DB), labeled under standardized culture conditions. 1235 and 557 spots were present in 5 of 5 gels in the 15% isoelectric focusing (IEF) and nonequilibrium pH gradient electrophoresis (NEPHGE) DB, respectively, whereas 995 and 378 spots were present in 5 of 5 gels in the 10% IEF and NEPHGE DB, respectively, yielding a reproducibility of spot detection between 75.2% and 91.7%. In both DBs, the average coefficient of variation of the percentage of integrated optical density (CV% of %IOD) for spots present in all gels was between 42.4% and 45.7%. When the same sample was analyzed in consecutive sets of gels on different days (interassay analysis), the average CV% of %IOD was 35.5%-36.1%. When the same sample was analyzed repeatedly in one set of gels (intra-assay analysis), the average CV% of %IOD was 30.2% in the IEF gels, while the average CV% of %IOD was 45.7% in the NEPHGE gels. Addition of interlenkin-1 beta (IL-1 beta) to the cultures resulted in statistically significant modulation or de novo synthesis of 105 proteins in the 10% gels. In conclusion, we present the first 10% and 15% acrylamide 2-D gel protein databases of neonatal rat islets of Langerhans and demonstrate its usage to identify proteins altered in expression by IL-1 beta*.