Complement activation by RPE cells preexposed to TNFα and IFNγ

Area of Science:

• Ophthalmology
• Immunology
• Cell Biology

Background:

• Age-related macular degeneration (AMD) is linked to complement system activation and elevated cytokine levels.
• Retinal pigment epithelial (RPE) cells are crucial for retinal health and are implicated in AMD pathogenesis.

Purpose of the Study:

• To investigate if pre-exposure of RPE cells with inflammatory cytokines enhances complement activation and membrane attack complex (MAC) deposition.
• To explore the role of specific cytokines, interferon-gamma (IFNγ) and tumor necrosis factor-alpha (TNFα), in this process.

Main Methods:

• Primary human RPE and ARPE19 cells were pre-exposed to IFNγ and TNFα.
• Cells were subsequently exposed to normal or complement factor B deficient (CFBd) serum.
• Membrane attack complex (MAC) deposition was quantified using ELISA and immunofluorescence.
• Cellular viability and metabolism were assessed using LDH and MTT assays.
• The effect of eculizumab (anti-C5) on MAC deposition was evaluated.

Main Results:

• MAC deposition occurred exclusively on RPE cells pre-exposed to both IFNγ and TNFα.
• MAC deposition was dependent on complement factor B and C5.
• Reconstitution of CFBd serum with complement factor B restored MAC deposition.
• MAC deposition led to RPE cell damage (LDH release) but did not alter mitochondrial activity (MTT).

Conclusions:

• Pre-exposure of RPE cells with inflammatory cytokines promotes alternative complement pathway activation and MAC deposition.
• Circulating inflammatory mediators may increase RPE susceptibility to complement activation.
• This cytokine-induced MAC deposition could be an early event in the pathogenesis of AMD.