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P2X7 Receptor Antagonist A804598 Inhibits Inflammation in Brain and Liver in C57BL/6J Mice Exposed to Chronic Ethanol and High Fat Diet

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Abstract

Chronic low-grade neuroinflammation is increasingly implicated in organ damage caused by alcohol abuse. Purinergic P2X7 receptors (P2X7Rs) play an important role in the generation of inflammatory responses during a number of CNS pathologies as evidenced from studies using pharmacological inhibition approach. P2X7Rs antagonism has not been tested during chronic alcohol abuse. In the present study, we tested the potential of P2X7R antagonist A804598 to reduce/abolish alcohol-induced neuroinflammation using chronic intragastric ethanol infusion and high-fat diet (Hybrid) in C57BL/6J mice. We have previously demonstrated an increase in neuroinflammatory response in 8 weeks of Hybrid paradigm. In the present study, we found neuroinflammatory response to 4 weeks of Hybrid exposure. A804598 treatment reversed the changes in microglia and astrocytes, reduced/abolished increases in mRNA levels of number of inflammatory markers, including IL-1β, iNOS, CXCR2, and components of inflammatory signaling pathways, such as TLR2, CASP1, NF-kB1 and CREB1, as well in the protein levels of pro-IL-1β and Nf-kB1. The P2X7R antagonist did not affect the increase in mRNA levels of fraktalkine (CX3CL1) and its receptor CX3CR1, an interaction that plays a neuroprotective role in neuron-glia communication. P2X7R antagonism also resulted in reduction of the inflammatory markers but did not alter steatosis in the liver. Taken together, these findings demonstrate how P2X7R antagonism suppresses inflammatory response in brain and liver but does not alter the neuroprotective response caused by Hybrid exposure. Overall, these findings support an important role of P2X7Rs in inflammation in brain and liver caused by combined chronic alcohol and high-fat diet.

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Abbreviations

P2X7R:

purinergic P2X7 receptor

BEC:

blood ethanol concentration

iG:

intragastric

HFD:

high fat diet

HCFD:

high cholesterol high fat diet

RT-qPCR:

reverse transcriptase – quantitative polymerase chain reaction

Gapdh:

glyceraldehyde 3-phosphate dehydrogenase

GFAP:

glial fibrillary acidic protein

Iba1:

ionized calcium binding adaptor molecule 1

References

  • Able SL, Fish RL, Bye H, Booth L, Logan YR, Nathaniel C, Hayter P, Katugampola SD (2011) Receptor localization, native tissue binding and ex vivo occupancy for centrally penetrant P2X7 antagonists in the rat. Br J Pharmacol 162:405–414

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Alfonso-Loeches S, Guerri C (2011) Molecular and behavioral aspects of the actions of alcohol on the adult and developing brain. Crit Rev Clin Lab Sci 48:19–47

    Article  CAS  PubMed  Google Scholar 

  • Alfonso-Loeches S, Pascual-Lucas M, Blanco AM, Sanchez-Vera I, Guerri C (2010) Pivotal role of TLR4 receptors in alcohol-induced neuroinflammation and brain damage. J Neurosci 30:8285–8295

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Asatryan L, Khoja S, Rodgers KE, Alkana RL, Tsukamoto H, Davies DL (2015) Chronic ethanol exposure combined with high fat diet up-regulates P2X7 receptors that parallels neuroinflammation and neuronal loss in C57BL/6J mice. J Neuroimmunol 285:169–179

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Asatryan L, Ostrovskaya O, Lieu D, Davies DL (2018) Ethanol differentially modulates P2X4 and P2X7 receptor activity and function in BV2 microglial cells. Neuropharmacol 128:11–21

    Article  CAS  Google Scholar 

  • Barbera-Cremades M, Gomez AI, Baroja-Mazo A, Martinez-Alarcon L, Martinez CM, de Torre-Minguela C, Pelegrin P (2017) P2X7 receptor induces tumor necrosis factor-alpha converting enzyme activation and release to boost TNF-alpha production. Front Immunol 8:862

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bartlett R, Stokes L, Sluyter R (2014) The P2X7 receptor channel: recent developments and the use of P2X7 antagonists in models of disease. Pharmacol Rev 66:638–675

    Article  CAS  PubMed  Google Scholar 

  • Catanzaro JM, Hueston CM, Deak MM, Deak T (2014) The impact of the P2X7 receptor antagonist A-804598 on neuroimmune and behavioral consequences of stress. Behav Pharmacol 25:582–598

    CAS  PubMed  Google Scholar 

  • Chatterjee S, Rana R, Corbett J, Kadiiska MB, Goldstein J, Mason RP (2012) P2X7 receptor-NADPH oxidase axis mediates protein radical formation and Kupffer cell activation in carbon tetrachloride-mediated steatohepatitis in obese mice. Free Radic Biol Med 52:1666–1679

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Chavda S, Luthert PJ, Salt TE (2016) P2X7R modulation of visually evoked synaptic responses in the retina. Purinergic Signal 12:611–625

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Chen Q, Wu H, Qin S, Liu C, Chen Y, Yang Y, Xu C (2016) The P2X7 receptor involved in gp120-induced cell injury in BV2 microglia. Inflammation 39:1814–1826

    Article  CAS  PubMed  Google Scholar 

  • Chessell IP, Hatcher JP, Bountra C, Michel AD, Hughes JP, Green P, Egerton J, Murfin M, Richardson J, Peck WL (2005) Disruption of the P2X7 purinoceptor gene abolishes chronic inflammatory and neuropathic pain. Pain 114:386–396

    Article  CAS  PubMed  Google Scholar 

  • Crews FT, Sarkar DK, Qin L, Zou J, Boyadjieva N, Vetreno RP (2015) Neuroimmune function and the consequences of alcohol exposure. Alcohol Res 37:344–351

    Google Scholar 

  • Crews FT, Vetreno RP (2014) Neuroimmune basis of alcoholic brain damage. Int Rev Neurobiol 118:315–357

    Article  PubMed  PubMed Central  Google Scholar 

  • Crews FT, Vetreno RP (2016) Mechanisms of neuroimmune gene induction in alcoholism. Psychopharmacol (Berl) 233:1543–1557

    Article  CAS  Google Scholar 

  • Diaz-Hernandez M, Diez-Zaera M, Sanchez-Nogueiro J, Gomez-Villafuertes R, Canals JM, Alberch J, Miras-Portugal MT, Lucas JJ (2009) Altered P2X7-receptor level and function in mouse models of Huntington's disease and therapeutic efficacy of antagonist administration. FASEB J 23:1893–1906

    Article  CAS  PubMed  Google Scholar 

  • Donnelly-Roberts DL, Namovic MT, Surber B, Vaidyanathan SX, Perez-Medrano A, Wang Y, Carroll WA, Jarvis MF (2009) [3H]A-804598 ([3H]2-cyano-1-[(1S)-1-phenylethyl]-3-quinolin-5-ylguanidine) is a novel, potent, and selective antagonist radioligand for P2X7 receptors. Neuropharmacol 56:223–229

    Article  CAS  Google Scholar 

  • Ferrari D, Pizzirani C, Adinolfi E, Lemoli RM, Curti A, Idzko M, Panther E, Di Virgilio F (2006) The P2X7 receptor: a key player in IL-1 processing and release. J Immunol 176:3877–3883

    Article  CAS  PubMed  Google Scholar 

  • Graeber MB (2010) Changing face of microglia. Science 330:783–788

    Article  CAS  PubMed  Google Scholar 

  • He J, Crews FT (2008) Increased MCP-1 and microglia in various regions of the human alcoholic brain. Exp Neurol 210:349–358

    Article  CAS  PubMed  Google Scholar 

  • Idzko M, Ferrari D, Eltzschig HK (2014) Nucleotide signalling during inflammation. Nature 509:310–317

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Jung YH, Kim YO, Han JH, Kim YC, Yoon MH (2017) Isobolographic analysis of drug combinations with intrathecal BRL52537 (kappa-opioid agonist), Pregabalin (Calcium Channel modulator), AF 353 (P2X3 receptor antagonist), and A804598 (P2X7 receptor antagonist) in neuropathic rats. Anesth Analg 125:670–677

    Article  CAS  PubMed  Google Scholar 

  • Kane CJ, Phelan KD, Douglas JC, Wagoner G, Johnson JW, Xu J, Phelan PS, Drew PD (2014) Effects of ethanol on immune response in the brain: region-specific changes in adolescent versus adult mice. Alcohol Clin Exp Res 38:384–391

    Article  CAS  PubMed  Google Scholar 

  • Karmakar M, Katsnelson MA, Dubyak GR, Pearlman E (2016) Neutrophil P2X7 receptors mediate NLRP3 inflammasome-dependent IL-1beta secretion in response to ATP. Nat Commun 7:10555

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Khakh BS, North RA (2006) P2X receptors as cell-surface ATP sensors in health and disease. Nature 442:527–532

    Article  CAS  PubMed  Google Scholar 

  • Kneer K, Green MB, Meyer J, Rich CB, Minns MS, Trinkaus-Randall V (2018) High fat diet induces pre-type 2 diabetes with regional changes in corneal sensory nerves and altered P2X7 expression and localization. Exp Eye Res 175:44–55

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lazaro R, Wu R, Lee S, Zhu NL, Chen CL, French SW, Xu J, Machida K, Tsukamoto H (2015) Osteopontin deficiency does not prevent but promotes alcoholic neutrophilic hepatitis in mice. Hepatol 61:129–140

    Article  CAS  Google Scholar 

  • Liu X, Zhao Z, Ji R, Zhu J, Sui QQ, Knight GE, Burnstock G, He C, Yuan H, Xiang Z (2017) Inhibition of P2X7 receptors improves outcomes after traumatic brain injury in rats. Purinergic Signal 13:529–544

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lovinger DM (2008) Communication networks in the brain: neurons, receptors, neurotransmitters. and alcohol Alcohol Res Health 31:196–214

    PubMed  Google Scholar 

  • Marshall SA, McClain JA, Kelso ML, Hopkins DM, Pauly JR, Nixon K (2013) Microglia activation is not equivalent to neuroinflammation in alcohol-induced neurodegeneration: the importance of mciroglia phenotype. Neurobiol Dis 54:239–251

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Matute C, Torre I, Perez-Cerda F, Perez-Samartin A, Alberdi E, Etxebarria E, Arranz AM, Ravid R, Rodriguez-Antiguedad A, Sanchez-Gomez M, Domercq M (2007) P2X(7) receptor blockade prevents ATP excitotoxicity in oligodendrocytes and ameliorates experimental autoimmune encephalomyelitis. J Neurosci 27:9525–9533

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • McClain JA, Morris SA, Deeny MA, Marshall SA, Hayes DM, Kiser ZM, Nixon K (2011) Adolescent binge alcohol exposure induces long-lasting partial activation of microglia. Brain Behav Immun 25(Suppl 1):S120–S128

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • McLarnon JG, Ryu JK, Walker DG, Choi HB (2006) Upregulated expression of purinergic P2X7 receptor in Alzheimer disease and amyloid-[beta] peptide-treated microglia and in peptide-injected rat hippocampus. J Neuropathol Exp Neurol 65:1090–1097

    Article  CAS  PubMed  Google Scholar 

  • Monif M, Burnstock G, Williams DA (2010) Microglia: proliferation and activation driven by the P2X7 receptor. Int J Biochem Cell Biol 42:1753–1756

    Article  CAS  PubMed  Google Scholar 

  • North RA, Jarvis MF (2013) P2X receptors as drug targets. Mol Pharmacol 83:759–769

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Pandolfi JB, Ferraro AA, Sananez I, Gancedo MC, Baz P, Billordo LA, Fainboim L, Arruvito L (2016) ATP-induced inflammation drives tissue-resident Th17 cells in metabolically unhealthy obesity. J Immunol 196:3287–3296

    Article  CAS  PubMed  Google Scholar 

  • Panenka W, Jijon H, Herx LM, Armstrong JN, Feighan D, Wei T, Yong VW, Ransohoff RM, MacVicar BA (2001) P2X7-like receptor activation in astrocytes increases chemokine monocyte chemoattractant protein-1 expression via mitogen-activated protein kinase. J Neurosci 21:7135–7142

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Pascual M, Balino P, Aragon CM, Guerri C (2015) Cytokines and chemokines as biomarkers of ethanol-induced neuroinflammation and anxiety-related behavior: role of TLR4 and TLR2. Neuropharmacol 89:352–359

    Article  CAS  Google Scholar 

  • Peng W, Cotrina ML, Han X, Yu H, Bekar L, Blum L, Takano T, Tian GF, Goldman SA, Nedergaard M (2009) Systemic administration of an antagonist of the ATP-sensitive receptor P2X7 improves recovery after spinal cord injury. PNAS USA 106:12489–12493

    Article  PubMed  PubMed Central  Google Scholar 

  • Sacks JJ, Gonzales KR, Bouchery EE, Tomedi LE, Brewer RD (2015) 2010 national and state costs of excessive alcohol consumption. Am J Prev Med 49:e73–e79

    Article  PubMed  Google Scholar 

  • Schmittgen TD, Livak KJ (2008) Analyzing real-time PCR data by the comparative C(T) method. Nat Protoc 3:1101–1108

    Article  CAS  PubMed  Google Scholar 

  • Sheridan GK, Murphy KJ (2013) Neuron-glia crosstalk in health and disease: fractalkine and CX3CR1 take Centre stage. Open Biol 3:130181

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Skaper SD, Debetto P, Giusti P (2010) The P2X7 purinergic receptor: from physiology to neurological disorders. FASEB J 24:337–345

    Article  CAS  PubMed  Google Scholar 

  • Stahre M, Roeber J, Kanny D, Brewer RD, Zhang X (2014) Contribution of excessive alcohol consumption to deaths and years of potential life lost in the United States. Prev Chronic Dis 11:E109

    Article  PubMed  PubMed Central  Google Scholar 

  • Suzuki T, Hide I, Ido K, Kohsaka S, Inoue K, Nakata Y (2004) Production and release of neuroprotective tumor necrosis factor by P2X7 receptor-activated microglia. J Neurosci 24:1–7

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Szabo G, Lippai D (2014) Converging actions of alcohol on liver and brain immune signaling. Int Rev Neurobiol 118:359–380

    Article  PubMed  Google Scholar 

  • Takenouchi T, Sekiyama K, Sekigawa A, Fujita M, Waragai M, Sugama S, Iwamaru Y, Kitani H, Hashimoto M (2010) P2X7 receptor signaling pathway as a therapeutic target for neurodegenerative diseases. Arch Immunol Therap Exp 58:91–96

    Article  CAS  Google Scholar 

  • Takenouchi T, Sugama S, Iwamaru Y, Hashimoto M, Kitani H (2009) Modulation of the ATP-induced release and processing of IL-1beta in microglial cells. Crit Rev Immunol 29:335–345

    Article  CAS  PubMed  Google Scholar 

  • Tewari M, Seth P (2015) Emerging role of P2X7 receptors in CNS health and disease. Ageing Res Rev 24:328–342

    Article  CAS  PubMed  Google Scholar 

  • Ueno A, Lazaro R, Wang PY, Higashiyama R, Machida K, Tsukamoto H (2012) Mouse intragastric infusion (iG) model. Nat Protoc 7:771–781

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Vengeliene V, Bilbao A, Molander A, Spanagel R (2008) Neuropharmacology of alcohol addiction. Br J Pharmacol 154:299–315

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Vetreno RP, Crews FT (2014) Current hypotheses on the mechanisms of alcoholism. Handb Clin Neurol 125:477–497

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Volonte C, Apolloni S, Carri MT, D'Ambrosi N (2011) ALS: focus on purinergic signalling. Pharmacol Ther 132:111–122

    Article  CAS  PubMed  Google Scholar 

  • Volonte C, Apolloni S, Skaper SD, Burnstock G (2012) P2X7 receptors: channels, pores and more. CNS & neurological disorders drug targets 11:705–721

    Article  CAS  Google Scholar 

  • Wieser V, Adolph TE, Enrich B, Kuliopulos A, Kaser A, Tilg H, Kaneider NC (2017) Reversal of murine alcoholic steatohepatitis by pepducin-based functional blockade of interleukin-8 receptors. Gut 66:930–938

    Article  CAS  PubMed  Google Scholar 

  • Xu J, Lai KK, Verlinsky A, Lugea A, French SW, Cooper MP, Ji C, Tsukamoto H (2011) Synergistic steatohepatitis by moderate obesity and alcohol in mice despite increased adiponectin and p-AMPK. J Hepatol 55:673–682

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

We would like to thank the Animal Core of the Southern California Research Center for ALPD and Cirrhosis (Director Dr. H. Tsukamoto) for providing us with the experimental animals, Elliott Cheung for Western blots and Lisa Walter for proof reading the manuscript and valuable recommendations. This work was conducted as partial fulfilment of the requirements for the MS. degree in Pharmaceutical Sciences, University of Southern California (D.F.). Sources of support: NIH/NIAAA AA017243 and Zumberge Individual Research Fund (L.A.), NIH/NIAAA A022448 (D.L.D.), NIH P50AA011999 (H.T.).

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Authors and Affiliations

  1. Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California, 1985 Zonal Avenue, Los Angeles, CA, 90033, USA

    Daniel Freire

  2. Department of Neurology, Keck School of Medicine, HCT 1520 San Pablo St, Los Angeles, CA, 90033, USA

    Daniel Freire

  3. Titus Family Department of Clinical Pharmacy, School of Pharmacy, University of Southern California, 1985 Zonal Avenue, Los Angeles, CA, 90033, USA

    Rachel E. Reyes, Ared Baghram, Daryl L. Davies & Liana Asatryan

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  1. Daniel Freire
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Correspondence to Liana Asatryan.

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Freire, D., Reyes, R.E., Baghram, A. et al. P2X7 Receptor Antagonist A804598 Inhibits Inflammation in Brain and Liver in C57BL/6J Mice Exposed to Chronic Ethanol and High Fat Diet. J Neuroimmune Pharmacol 14, 263–277 (2019). https://doi.org/10.1007/s11481-018-9816-3

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