Signaling pathways for glycated human serum albumin-induced IL-8 and MCP-1 secretion in human RPE cells.
Signaling pathways for glycated human serum albumin-induced IL-8 and MCP-1 secretion in human RPE cells.
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发表时间：
2001-06
影响因子：
4.4
通讯作者：
Z. Bian;V. Elner;Ayako Yoshida;S. L. Kunkel;S. Elner
中科院分区：
文献类型：
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作者：
Z. Bian;V. Elner;Ayako Yoshida;S. L. Kunkel;S. Elner
目的探讨糖化人血清白蛋白（GHSA）刺激人视网膜色素上皮（hRPE）细胞分泌白细胞介素（IL）-8和单核细胞趋化蛋白（MCP）-1的信号通路。方法在一系列激酶抑制剂存在或不存在的情况下，用GHSA刺激hRPE细胞。采用逆转录-聚合酶链反应（RT-PCR）和酶联免疫吸附试验（ELISA）检测IL-8和MCP-1的mRNA和蛋白表达。Western印迹分析、电泳迁移率变动分析和免疫组织化学染色用于分析信号介质和转录因子的活化。结果：GHSA能迅速激活Raf-1、细胞外信号调节蛋白激酶（ERK）1/2、p38和转录因子核因子（NF）-kappaB。将hRPE细胞与促分裂原活化蛋白（MAP）激酶（MEK）抑制剂U 0126、NF-κ B抑制剂BAY 11 -7085、咖啡酸苯乙酯（CAPE）、小白菊素和姜黄素、蛋白激酶（PK）C抑制剂Ro 318220和蛋白酪氨酸激酶（PTK）抑制剂染料木黄酮共孵育，可在很大程度上消除大部分IL-8和MCP-1的刺激产生。通过U 0126、SB 202190和AG 490对MEK、p38和Janus激酶（jak）的组合抑制揭示了GHSA刺激IL-8产生主要由MEK介导，并且在较小程度上由p38途径介导，而MEK、p38和jak的激活是最大MCP-1诱导所需的。此外，GHSA刺激的IL-8分泌对U 0126（50%抑制浓度[IC（50）] = 0.5 μ M）比MCP-1（IC（50）= 10 μ M）更敏感。结论：GHSA通过不同的和重叠的，但不相同的细胞内信号级联刺激hRPE IL-8和MCP-1的产生。GHSA诱导一系列激酶的活化，包括PKC、PTK、MAPK、p38和jak以及转录因子NF-κ B。Raf/MAPK通路在GHSA信号转导中起着重要作用。
PURPOSE To determine the signal mediators involved in glycated human serum albumin (GHSA) stimulation of interleukin (IL)-8 and monocyte chemotactic protein (MCP)-1 secretion in human retinal pigment epithelium (hRPE) cells. METHODS hRPE cells were stimulated by GHSA in the presence or absence of a series of kinase inhibitors. The induced IL-8 and MCP-1 mRNA and proteins were determined by reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). Western blot analysis, electrophoretic mobility shift assay, and immunohistochemical staining were used to analyze activation of signaling mediators and transcription factors. RESULTS Incubation of hRPE cells with GHSA resulted in rapid activation of Raf-1, extracellular signal-regulated protein kinases (ERK) 1/2, p38, and the transcription factor nuclear factor (NF)-kappaB. Coincubation of hRPE cells with the mitogen-activated protein (MAP) kinase (MEK) inhibitor U0126; NF-kappaB inhibitors BAY11-7085, caffeic acid phenethyl ester (CAPE), parthenolide, and curcumin; protein kinase (PK)C inhibitor Ro318220; and protein tyrosine kinase (PTK) inhibitor genistein largely eliminated most of the stimulated production of IL-8 and MCP-1. Combined inhibition of MEK by U0126, p38 by SB202190, and Janus kinase (jak) by AG490 revealed that GHSA stimulation of IL-8 production was predominately mediated by MEK and to a lesser extent by p38 pathways, whereas activation of MEK, p38, and jak was required for maximal MCP-1 induction. Moreover, GHSA-stimulated IL-8 secretion was more sensitive to U0126 (50% inhibitory concentration [IC(50)] = 0.5 microM) than MCP-1 (IC(50) = 10 microM). CONCLUSIONS GHSA stimulates hRPE IL-8 and MCP-1 production through divergent and overlapping, but not identical, intracellular signaling cascades. GHSA induces activation of a series of kinases including PKC, PTK, MAPK, p38, and jak and the transcription factor NF-kappaB. The Raf/MAPK pathway plays an essential role in GHSA signaling.