Abstract
BackgroundInsufficient clearance by microglial cells, prevalent in several neurological conditions and diseases, is intricately intertwined with MFG-E8 expression and inflammatory responses. Electromagnetic field (EMF) exposure can elicit the pro-inflammatory activation and may also trigger an alteration of the clearance function in microglial cells. Curcumin has important roles in the anti-inflammatory and phagocytic process. Here, we evaluated the ability of curcumin to ameliorate the phagocytic ability of EMF-exposed microglial cells (N9 cells) and documented relative pathways.MethodsN9 cells were pretreated with or without recombinant murine MFG-E8 (rmMFG-E8), curcumin and an antibody of toll-like receptor 4 (anti-TLR4), and subsequently treated with EMF or a sham exposure. Their phagocytic ability was evaluated using phosphatidylserine-containing fluorescent bioparticles. The pro-inflammatory activation of microglia was assessed via CD11b immunoreactivity and the production of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1β (IL-1β) and nitric oxide (NO) via the enzyme-linked immunosorbent assay or the Griess test. We evaluated the ability of curcumin to ameliorate the phagocytic ability of EMF-exposed N9 cells, including checking the expression of MFG-E8, αvβ3 integrin, TLR4, nuclear factor-κB (NF-κB) and signal transducer and activator of transcription 3 (STAT3) using Western blotting.ResultsEMF exposure dramatically enhanced the expression of CD11b and depressed the phagocytic ability of N9 cells. rmMFG-E8 could clearly ameliorate the phagocytic ability of N9 cells after EMF exposure. We also found that EMF exposure significantly increased the secretion of pro-inflammatory cytokines (TNF-α, IL-6 and IL-1β) and the production of NO; however, these increases were efficiently chilled by the addition of curcumin to the culture medium. This reduction led to the amelioration of the phagocytic ability of EMF-exposed N9 cells. Western blot analysis revealed that curcumin and naloxone restored the expression of MFG-E8 but had no effect on TLR4 and cytosolic STAT3. Moreover, curcumin significantly reduced the expression of NF-κB p65 in nuclei and phospho-STAT3 (p-STAT3) in cytosols and nuclei.ConclusionsThis study indicates that curcumin ameliorates the depressed MFG-E8 expression and the attenuated phagocytic ability of EMF-exposed N9 cells, which is attributable to the inhibition of the pro-inflammatory response through the NF-κB and STAT3 pathways.
Highlights
Insufficient clearance by microglial cells, prevalent in several neurological conditions and diseases, is intricately intertwined with milk-fat globule EGF factor-8 (MFG-E8) expression and inflammatory responses
Does Electromagnetic field (EMF) exposure depress microglial phagocytosis during inflammation? If EMF depresses phagocytosis, does curcumin directly regulate the depression of microglial phagocytosis or through its anti-inflammatory effects? We demonstrated that the inhibition of microglial pro-inflammatory responses by curcumin restores MFG-E8-mediated phagocytosis after EMF exposure
After the addition of curcumin to the culture medium, the expression of p-signal transducer and activator of transcription 3 (STAT3) and nuclear factor-κB (NF-κB) was chilled down to the baseline level of around 0.30 and 0.46 fold compared to the controls for EMF-exposed N9 cells (Figure 7A,B). These results suggested that curcumin may ameliorate defective microglial phagocytosis via the inhibition of NF-κB activation and STAT3 phosphorylation, but not through alteration of toll-like receptor 4 (TLR4) signaling
Summary
Insufficient clearance by microglial cells, prevalent in several neurological conditions and diseases, is intricately intertwined with MFG-E8 expression and inflammatory responses. Electromagnetic field (EMF) exposure can elicit the pro-inflammatory activation and may trigger an alteration of the clearance function in microglial cells. We previously observed that EMF exposure initiated the pro-inflammatory activation of microglial cells [11]. The MFG-E8-mediated phagocytosis of apoptotic cells results in an inhibition of inflammation via the MAPK and nuclear factor-κB (NF-κB) signaling pathways following the endotoxin response [18]. These findings indicate that the role of MFG-E8 in regulating the immune reaction of microglial phagocytosis and pro-inflammatory responses could depend on the types of stimulus
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