Complement Stimulates Retinal Pigment Epithelial Cells to Undergo Pro-Inflammatory Changes.
OBJECTIVE
We examined the effect of human complement sera (HCS) on retinal pigment epithelial (RPE) cells with respect to pro-inflammatory mediators relevant in early age-related macular degeneration (AMD).


METHODS
RPE cells were treated with complement-containing HCS or with heat-inactivated (HI) HCS or C7-deficient HCS as controls. Cells were analysed for C5b-9 using immunocytochemistry and flow cytometry. Interleukin (IL)-6, IL-8, and monocyte chemoattractant protein-1 (MCP-1) were quantified by ELISA and RT-PCR. Tumour necrosis factor-α (TNF-α), intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), were analysed by Western blotting. The intracellular distribution of nuclear factor (NF)-x03BA;B was investigated by immunofluorescence.


RESULTS
A concentration-dependent increased staining for C5b-9 but no influence on cell viability was observed after HCS treatment. ELISA and RT-PCR analysis revealed elevated secretion and expression of IL-6, IL-8, and MCP-1. Western blot analysis showed a concentration-dependent increase in ICAM-1, VCAM-1, and TNF-α in response to HCS, and immunofluorescence staining revealed nuclear translocation of NF-x03BA;B.


CONCLUSIONS
This study suggests that complement stimulates NF-x03BA;B activation in RPE cells that might further create a pro-inflammatory environment. All these factors together may support early AMD development.
Ophtha-Lab, Department of Ophthalmology at St. Franziskus Hospital, Muenster, Germany.
Katharina
K
Lueck
Martin
M
Busch
Stephen E
SE
Moss
John
J
Greenwood
Maren
M
Kasper
Albrecht
A
Lommatzsch
Daniel
D
Pauleikhoff
Susanne
S
Wasmuth
D015153:Blotting, Western; D002478:Cells, Cultured; D015938:Complement Membrane Attack Complex / Q000378:metabolism*; D003165:Complement System Proteins / Q000502:physiology*; D016207:Cytokines / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D018836:Inflammation Mediators / Q000378:metabolism*; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D008268:Macular Degeneration / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D055213:Retinal Pigment Epithelium / Q000378:metabolism*; D019010:Vascular Cell Adhesion Molecule-1 / Q000378:metabolism
D015938:Complement Membrane Attack Complex; D016207:Cytokines; D018836:Inflammation Mediators; D016328:NF-kappa B; D019010:Vascular Cell Adhesion Molecule-1; D018799:Intercellular Adhesion Molecule-1; D003165:Complement System Proteins