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Cannabidiol prevents LPS-induced microglial inflammation by inhibiting ROS/NF-κB-dependent signaling and glucose consumption.

Glia (2019-10-28)
Mauricio Dos-Santos-Pereira, Franscisco S Guimarães, Elaine Del-Bel, Rita Raisman-Vozari, Patrick P Michel
ABSTRACT

We used mouse microglial cells in culture activated by lipopolysaccharide (LPS, 10 ng/ml) to study the anti-inflammatory potential of cannabidiol (CBD), the major nonpsychoactive component of cannabis. Under LPS stimulation, CBD (1-10 μM) potently inhibited the release of prototypical proinflammatory cytokines (TNF-α and IL-1β) and that of glutamate, a noncytokine mediator of inflammation. The effects of CBD were predominantly receptor-independent and only marginally blunted by blockade of CB2 receptors. We established that CBD inhibited a mechanism involving, sequentially, NADPH oxidase-mediated ROS production and NF-κB-dependent signaling events. In line with these observations, active concentrations of CBD demonstrated an intrinsic free-radical scavenging capacity in the cell-free DPPH assay. Of interest, CBD also prevented the rise in glucose uptake observed in microglial cells challenged with LPS, as did the inhibitor of NADPH oxidase apocynin and the inhibitor of IκB kinase-2, TPCA-1. This indicated that the capacity of CBD to prevent glucose uptake also contributed to its anti-inflammatory activity. Supporting this view, the glycolytic inhibitor 2-deoxy-d-glucose (2-DG) mimicked the antioxidant/immunosuppressive effects of CBD. Interestingly, CBD and 2-DG, as well as apocynin and TPCA-1 caused a reduction in glucose-derived NADPH, a cofactor required for NADPH oxidase activation and ROS generation. These different observations suggest that CBD exerts its anti-inflammatory effects towards microglia through an intrinsic antioxidant effect, which is amplified through inhibition of glucose-dependent NADPH synthesis. These results also further confirm that CBD may have therapeutic utility in conditions where neuroinflammatory processes are prominent.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Lactate Assay Kit, sufficient for 100 colorimetric or fluorometric tests
Sigma-Aldrich
TPCA-1, ≥95% (HPLC)
Sigma-Aldrich
2-Deoxy-D-glucose, ≥98% (GC), crystalline
Sigma-Aldrich
Lipopolysaccharides from Escherichia coli O26:B6, ≥10,000 EU/mg, purified by phenol extraction
Supelco
Poly(ethyleneimine) solution, analytical standard, 50 % (w/v) in H2O
Sigma-Aldrich
High Sensitivity NADPH Assay Kit, Sufficient for 100 Fluorometric tests
Sigma-Aldrich
2,2-Diphenyl-1-picrylhydrazyl