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 IC(50) for inhibition of MCP-1 secretion induced by IL-1beta, TNF-alpha, and hRPE-monocyte binding was 16, 12, and less than 3 micro M, 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.
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 IC(50) for inhibition of MCP-1 secretion induced by IL-1beta, TNF-alpha, and hRPE-monocyte binding was 16, 12, and less than 3 micro M, 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.
Authors: Khaliq H Kurji; Jing Z Cui; Tony Lin; David Harriman; Shiv S Prasad; Ljuba Kojic; Joanne A Matsubara Journal: Invest Ophthalmol Vis Sci Date: 2009-09-24 Impact factor: 4.799
Authors: Dongli Yang; Susan G Elner; Zong-Mei Bian; Gerd O Till; Howard R Petty; Victor M Elner Journal: Exp Eye Res Date: 2007-06-27 Impact factor: 3.467
Authors: Wenbo Zhang; Modesto Rojas; Brenda Lilly; Nai-Tse Tsai; Tahira Lemtalsi; Gregory I Liou; Robert W Caldwell; Ruth B Caldwell Journal: Invest Ophthalmol Vis Sci Date: 2009-02-21 Impact factor: 4.799