Key Laboratory of Medical Protection for Electromagnetic Radiation Ministry of Education, Third Military Medical University, 30 Gaotanyan Street, Chongqing 400038, PR China.
J Neuroinflammation. 2014 Mar 19;11:49. doi: 10.1186/1742-2094-11-49.
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 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.
N9 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.
EMF 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.
This 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.
在几种神经状况和疾病中普遍存在的小胶质细胞清除不足，与 MFG-E8 表达和炎症反应密切相关。电磁场（EMF）暴露会引起促炎激活，并且还可能触发小胶质细胞清除功能的改变。姜黄素在抗炎和吞噬过程中具有重要作用。在这里，我们评估了姜黄素改善 EMF 暴露的小胶质细胞（N9 细胞）吞噬能力的能力，并记录了相关途径。
N9 细胞用或不用重组鼠 MFG-E8（rmMFG-E8）、姜黄素和 Toll 样受体 4 抗体（抗 TLR4）预处理，然后用 EMF 或假暴露处理。使用含有磷脂酰丝氨酸的荧光生物颗粒评估其吞噬能力。通过 CD11b 免疫反应性以及通过酶联免疫吸附试验或格里斯试验测定肿瘤坏死因子-α（TNF-α）、白细胞介素-6（IL-6）、白细胞介素-1β（IL-1β）和一氧化氮（NO）的产生来评估小胶质细胞的促炎激活。我们评估了姜黄素改善 EMF 暴露的 N9 细胞吞噬能力的能力，包括通过 Western blot 检查 MFG-E8、αvβ3 整联蛋白、TLR4、核因子-κB（NF-κB）和信号转导和转录激活因子 3（STAT3）的表达。
EMF 暴露显著增强了 CD11b 的表达，并抑制了 N9 细胞的吞噬能力。rmMFG-E8 可明显改善 EMF 暴露后 N9 细胞的吞噬能力。我们还发现，EMF 暴露显著增加了促炎细胞因子（TNF-α、IL-6 和 IL-1β）的分泌和 NO 的产生；然而，在培养基中加入姜黄素可有效抑制这些增加。这种减少导致 EMF 暴露的 N9 细胞吞噬能力的改善。Western blot 分析显示，姜黄素和纳洛酮恢复了 MFG-E8 的表达，但对 TLR4 和胞质 STAT3 没有影响。此外，姜黄素显著降低了核内 NF-κB p65 和胞质及核内磷酸化 STAT3（p-STAT3）的表达。
本研究表明，姜黄素改善了 EMF 暴露的 N9 细胞中抑制的 MFG-E8 表达和减弱的吞噬能力，这归因于通过 NF-κB 和 STAT3 途径抑制促炎反应。
