Background: Neuroinflammation plays an important role in the pathophysiological process of various neurodegenerative diseases. It is well known that curcumin has obvious anti-inflammatory effects in various neuroinflammation models. However, its effect on the modulation of microglial polarization is largely unknown.

Objective: This study aimed to investigate whether curcumin changed microglia to an anti-inflammatory M2-phenotype by activating the AMP-activated protein kinase (AMPK) signaling pathway.

Methods: LPS treatment was used to establish BV2 cells and primary microglia neuroinflammation models. The neuroinflammation mouse model was established by an intracerebroventricular (ICV) injection of lipopolysaccharide (LPS) in the lateral septal complex region of the brain. TNF-α was measured by ELISA, and cell viability was measured by Cell Counting Kit-8 (CCK-8). The expression of proinflammatory and anti-inflammatory cytokines was examined by Q-PCR and Western blot analysis. Phenotypic polarization of BV2 microglia was detected by immunofluorescence.

Results: Curcumin enhanced AMPK activation in BV2 microglial cells in the presence and absence of LPS. Upon LPS stimulation, the addition of curcumin promoted M2 polarization of BV2 cells, as evidenced by suppressed M1 and the elevated M2 signature protein and gene expression. The effects of curcumin were inhibited by an AMPK inhibitor or AMPK knockdown. Calmodulin-dependent protein kinase kinase β (CaMKKβ) and liver kinase B1 (LKB1) are upstream kinases that activate AMPK. Curcumin can activate AMPK in Hela cells, which do not express LKB1. However, both the CaMKKβ inhibitor and siRNA blocked curcumin activation of AMPK in LPS-stimulated BV2 cells. Moreover, the CaMKKβ inhibitor and siRNA weaken the effect of curcumin suppression on M1 and enhancement of M2 protein and gene expression in LPS-stimulated BV2 cells. Finally, curcumin enhanced AMPK activation in the brain area where microglia were over-activated upon LPS stimulation in an in vivo neuroinflammation model. Moreover, curcumin also suppressed M1 and promoted M2 signature protein and gene expression in this in vivo model.

Conclusion: Curcumin enhances microglia M2 polarization via the CaMKKβ-dependent AMPK signaling pathway. Additionally, curcumin treatment was found to be neuroprotective and thus might be considered as a novel therapeutic agent to treat the neurodegenerative disease such as Alzheimer‘s disease, Parkinson's disease, etc.

背景：神经炎症在各种神经退行性疾病的病理生理过程中起着重要作用。众所周知，姜黄素在多种神经炎症模型中具有明显的抗炎作用。然而，它对小胶质细胞极化调制的影响在很大程度上是未知的。

目的：本研究旨在探讨姜黄素是否通过激活 AMP 活化蛋白激酶 (AMPK) 信号通路将小胶质细胞转变为抗炎 M2 表型。

方法：采用LPS处理建立BV2细胞和原代小胶质细胞神经炎症模型。神经炎症小鼠模型是通过脑室 (ICV) 内注射脂多糖 (LPS) 在大脑的侧隔复合体区域建立的。TNF-α通过ELISA测量，细胞活力通过Cell Counting Kit-8 (CCK-8)测量。通过 Q-PCR 和蛋白质印迹分析检查促炎和抗炎细胞因子的表达。通过免疫荧光检测 BV2 小胶质细胞的表型极化。

结果：姜黄素在 LPS 存在和不存在的情况下增强了 BV2 小胶质细胞中的 AMPK 活化。在 LPS 刺激下，姜黄素的添加促进了 BV2 细胞的 M2 极化，这可以通过抑制 M1 和升高的 M2 特征蛋白和基因表达来证明。姜黄素的作用受到 AMPK 抑制剂或 AMPK 敲低的抑制。钙调蛋白依赖性蛋白激酶激酶 β (CaMKKβ) 和肝激酶 B1 (LKB1) 是激活 AMPK 的上游激酶。姜黄素可以激活不表达 LKB1 的 Hela 细胞中的 AMPK。然而，CaMKKβ 抑制剂和 siRNA 都阻断了 LPS 刺激的 BV2 细胞中 AMPK 的姜黄素活化。此外，CaMKKβ 抑制剂和 siRNA 减弱了姜黄素对 M1 的抑制作用以及 LPS 刺激的 BV2 细胞中 M2 蛋白和基因表达的增强。最后，在体内神经炎症模型中，姜黄素增强了小胶质细胞在 LPS 刺激下过度激活的大脑区域的 AMPK 激活。此外，姜黄素还在该体内模型中抑制 M1 并促进 M2 特征蛋白和基因表达。

结论：姜黄素通过 CaMKKβ 依赖性 AMPK 信号通路增强小胶质细胞 M2 极化。此外，姜黄素治疗被发现具有神经保护作用，因此可能被认为是治疗阿尔茨海默病、帕金森病等神经退行性疾病的新型治疗剂。