Curcumin Prevents Neuroinflammation by Inducing Microglia to Transform into the M2-phenotype via CaMKKβ-dependent Activation of the AMP-Activated Protein Kinase Signal Pathway

Abstract

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.

Keywords: AMPK; Curcumin; Parkinson's disease; cytokines; microglia; neuroinflammation.