Differential involvement of phosphoinositide 3-kinase/Akt in human RPE MCP-1 and IL-8 expression
Differential involvement of phosphoinositide 3-kinase/Akt in human RPE MCP-1 and IL-8 expression
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DOI：
10.1167/iovs.03-0608
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发表时间：
2004-06-01
影响因子：
4.4
通讯作者：
Elner, VM
中科院分区：
文献类型：
--
作者：
Bian, ZM;Elner, SG;Elner, VM
目的。目的探讨磷脂酰肌醇3-激酶(PI3K)信号转导通路及其信号介质AP-1在人视网膜色素上皮(HRPE)细胞单核细胞趋化蛋白-1(MCP-1)和白介素8(IL-8)基因表达中的作用。用ELISA法和RT-PCR法分别检测MCP-1和IL-8蛋白和mRNA的诱导作用。结果：在刺激诱导趋化因子表达的同时，PI3K及其下游靶标Akt、GSK和FKHR也被磷酸化。PI3K特异性抑制剂Ly294002呈时间和剂量依赖性地阻断MCP-1mRNA的表达和蛋白的产生。抑制IL-1β、TNF-α和hRPE-单核细胞结合诱导的MCP-1分泌的IC50分别为16、12和小于3微米。相反，Ly294002不能抑制任何刺激诱导的IL-8表达。MEK、p38和JNK的选择性抑制剂Ly294002和U0126、SB202190和SP600125分别强烈抑制c-fos的诱导表达，而Ly294002不抑制MEK、p38和JNK的诱导。阻断PI3K/Akt可阻断IL-1β诱导的AP-1核转位，但不改变对IkappaB降解的诱导。结论Ly294002敏感的PI3K/Akt通路可调节hRPE细胞MCP-1的表达，但不依赖于MAPK和IkappaB调节IL-8的表达。依赖PI3K诱导hRPE、c-fos和AP-1核转位可能是调控MCP-1治疗视网膜疾病的靶点。
PURPOSE. To investigate the role of the phosphatidylinositol 3-kinase (PI3K) pathway and the signal mediator AP-1 in monocyte chemotactic protein (MCP)-1 and interleukin (IL)-8 gene expression in human retinal pigment epithelial (hRPE) cells.METHODS. hRPE cells were stimulated with IL-1beta and TNF-alpha and by coculturing with monocytes in the presence or absence of a series of kinase inhibitors. The induction of MCP-1 and IL-8 protein and mRNA was determined by ELISA and RT-PCR, respectively. Western blot analysis, kinase assays, and electrophoretic mobility shift assays were used to detect the activation of signaling mediators and transcription factors.RESULTS. Concomitant with the induction of chemokine expression by the stimuli, there was phosphorylation of PI3K and its downstream targets-namely, Akt, GSK, and FKHR. Ly294002, a specific inhibitor of PI3K, resulted in time- and dose-dependent blockade of MCP-1 mRNA expression and protein production. The IC50 for inhibition of MCP-1 secretion induced by IL-1beta, TNF-alpha, and hRPE-monocyte binding was 16, 12, and less than 3 muM, respectively. In contrast, Ly294002 did not inhibit the IL-8 expression induced by any of the stimuli. Ly294002 as well as U0126, SB202190, and SP600125, the selective inhibitors of MEK, p38, and JNK, respectively, strongly inhibited induced c-fos expression, whereas Ly294002 did not inhibit induction of MEK, p38, or JNK. Blockade of PI3K/Akt abolished IL-1beta-induced nuclear translocation of AP-1, whereas the induction Of IkappaB degradation was unchanged.CONCLUSIONS. The Ly294002-sensitive PI3K/Akt pathway regulates MCP-1, but not IL-8 expression in hRPE cells independent of MAPK and IkappaB. PI3K-dependent induction of hRPE c-fos and AP-1 nuclear translocation may be a target for therapies aimed at modulating MCP-1 in retinal diseases.