Chronic alcohol-induced neuroinflammation involves CCR2/5-dependent peripheral macrophage infiltration and microglia alterations

Patrick P Lowe,Star Seyedkazemi,Arvin Iracheta-Vellve,Gyongyi Szabo,Erica Kwiatkowski,Laurent Fischer,Eric Lefebvre,Aditya Ambade,Donna Catalano,Abhishek Satishchandran,Istvan Furi,Dorothy P Schafer,Caroline Morel,Benedek Gyongyosi,Yeonhee Cho

doi:10.1186/s12974-020-01972-5

Abstract

BackgroundChronic alcohol consumption is associated with neuroinflammation, neuronal damage, and behavioral alterations including addiction. Alcohol-induced neuroinflammation is characterized by increased expression of proinflammatory cytokines (including TNFα, IL-1β, and CCL2) and microglial activation. We hypothesized chronic alcohol consumption results in peripheral immune cell infiltration to the CNS. Since chemotaxis through the CCL2-CCR2 signaling axis is critical for macrophage recruitment peripherally and centrally, we further hypothesized that blockade of CCL2 signaling using the dual CCR2/5 inhibitor cenicriviroc (CVC) would prevent alcohol-induced CNS infiltration of peripheral macrophages and alter the neuroinflammatory state in the brain after chronic alcohol consumption.MethodsC57BL/6J female mice were fed an isocaloric or 5% (v/v) ethanol Lieber DeCarli diet for 6 weeks. Some mice received daily injections of CVC. Microglia and infiltrating macrophages were characterized and quantified by flow cytometry and visualized using CX3CR1eGFP/+ CCR2RFP/+ reporter mice. The effect of ethanol and CVC treatment on the expression of inflammatory genes was evaluated in various regions of the brain, using a Nanostring nCounter inflammation panel. Microglia activation was analyzed by immunofluorescence. CVC-treated and untreated mice were presented with the two-bottle choice test.ResultsChronic alcohol consumption induced microglia activation and peripheral macrophage infiltration in the CNS, particularly in the hippocampus. Treatment with CVC abrogated ethanol-induced recruitment of peripheral macrophages and partially reversed microglia activation. Furthermore, the expression of proinflammatory markers was upregulated by chronic alcohol consumption in various regions of the brain, including the cortex, hippocampus, and cerebellum. Inhibition of CCR2/5 decreased alcohol-mediated expression of inflammatory markers. Finally, microglia function was impaired by chronic alcohol consumption and restored by CVC treatment. CVC treatment did not change the ethanol consumption or preference of mice in the two-bottle choice test.ConclusionsTogether, our data establish that chronic alcohol consumption promotes the recruitment of peripheral macrophages into the CNS and microglia alterations through the CCR2/5 axis. Therefore, further exploration of the CCR2/5 axis as a modulator of neuroinflammation may offer a potential therapeutic approach for the treatment of alcohol-associated neuroinflammation.

Highlights

Alcohol use disorder (AUD) impacts millions of people across the world, including at least 17 million US patients [1]
Together, our data establish that chronic alcohol consumption promotes the recruitment of peripheral macrophages into the central nervous system (CNS) and microglia alterations through the CCR2/5 axis
Alcohol induces recruitment of peripheral macrophages into the CNS In order to investigate the role of peripheral infiltrating macrophages (IMs) in chronic alcohol, we used a common model [20] of chronic alcohol consumption in mice (Fig. 1a)

Summary

Introduction

Alcohol use disorder (AUD) impacts millions of people across the world, including at least 17 million US patients [1]. Recent studies using animal models have shown that inflammatory signaling contributes to neurodegeneration and to alcohol addiction [5,6,7], making targeting of neuroinflammation a critical approach in the treatment of AUD. Both infiltrating macrophages (IMs) and microglia become activated in response to tissue damage and can release proinflammatory cytokines, which may contribute to neuroinflammation and blood-brain barrier breakdown [8, 9]. Since chemotaxis through the CCL2-CCR2 signaling axis is critical for macrophage recruitment peripherally and centrally, we further hypothesized that blockade of CCL2 signaling using the dual CCR2/5 inhibitor cenicriviroc (CVC) would prevent alcohol-induced CNS infiltration of peripheral macrophages and alter the neuroinflammatory state in the brain after chronic alcohol consumption

Methods

Results

Conclusion