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Invest Ophthalmol Vis Sci. 2001 Jun;42(7):1660-8.

Signaling pathways for glycated human serum albumin-induced IL-8 and MCP-1 secretion in human RPE cells.

Author information

1
Departments of Ophthalmology, University of Michigan, Ann Arbor 48105, USA.

Abstract

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.

PMID:
11381075
[Indexed for MEDLINE]

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