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Apigenin and luteolin modulate microglial activation via inhibition of STAT1-induced CD40 expression

Kavon Rezai-Zadeh, Jared Ehrhart, Yun Bai, Paul R. Sanberg, Paula Bickford, Jun Tan, and R. Douglas Shytle

Published: 25 September 2008
Journal of Neuroinflammation 2008, 5:41

It is well known that most neurodegenerative diseases are associated with microglia-mediated inflammation. Our previous research demonstrates that the CD40 signaling is critically involved in microglia-related immune responses in the brain. We and others have previously reported that microglial CD40 expression is significantly induced by IFN-γ and amyloid-β (Aβ) peptide. Recent studies have shown that certain flavonoids possess anti-inflammatory and neuroprotective properties distinct from their well-known anti-oxidant effects. In particular, flavonoids, apigenin and luteolin have been found to be effective CD40 immunomodulators.

Cultured microglia, both N9 and primary derived lines, were treated with flavonoids in the presence of IFN-γ and/or CD40 ligation to assess any anti-inflammatory effects and/or mechanisms. Apigenin and luteolin concentration-dependently suppressed IFN-γ-induced CD40 expression. In addition, apigenin and luteolin markedly inhibited IFN-γ-induced phosphorylation of STAT1 with little impact on cell survival.

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Figure 1
Apigenin and luteolin reduce microglial CD40 expression induced by IFN-γ in a concentration dependent manner. FACS analysis showed significant concentration dependent decreases in IFN-γ-induced CD40 expression by both apigenin and luteolin following 8 hrs. of co-treatment in N9 cells seeded in 6-well tissue-culture plates (5 × 105/well). Data were represented as mean % of CD40 expressing cells (+/- SD). Results are representative of three independent experiments.

Figure 2
Apigenin and luteolin inhibit microglial CD40 expression induced by IFN-γ. N9 and murine-derived primary microglial cells were seeded in 6-well tissue-culture plates (5 × 105/well) for FACS analysis and 24-well tissue-culture plates (1 × 105/ well) for LDH analysis in parallel. Cultured cells were co-treated with IFN-γ (100 U/mL) in the presence or absence of apigenin and luteolin (25 μM) or treated with vehicle (1% DMSO; control) for 8 hrs. For A and B, FACS analysis showed significant decreases by both apigenin and luteolin (25 μM) in IFN-γ-induced CD40 expression in N9 cells and primary microglia. Data are represented as mean % of CD40 expressing cells (+/- SD). For C and D, cultured supernatants were collected and subjected to LDH assay as indicated. Data showed no significant increase in cell toxicity below a 50 μM concentration of either flavonoid in both N9 or primary microglial cells. Data were represented as mean % of total cell death as determined by LDH present during complete lysis (+/- SD). For A, B, C and D. Results are representative of three independent experiments. (*p < 0.05; **p <0.01).

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