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Retinoic acid attenuates nuclear factor kappaB mediated induction of NLRP3 inflammasome

Pharmacological Reports volume 74, pages 189–203 (2022)Cite this article

Abstract

Background

Acetylcholine (ACh), a neurotransmitter and a part of the cholinergic system, can modify immune responses. Expression of acetylcholine receptors (AChR) in immune cells, including macrophages, leads to modulation of their function. Inflammasomes are part of the innate immune system and have been linked to a variety of inflammatory diseases. The NLRP3/ASC/caspase-1/IL-1 axis has emerged as a critical signaling pathway in inflammation process initiation. The role of ACh in modulating inflammasomes in macrophages remains relatively under-explored.

Methods

The effect of AChR agonist carbachol on inflammasome expression was investigated using murine and human macrophages. Cell lysates were assessed by western blot for protein analysis. Immunofluorescence studies were used to study the translocation of p65. The experiments were conducted in the presence of NF-ĸB inhibitor, AChR antagonists, and retinoic acid (RA) to study the role of NF-ĸB, ACh receptors, and RA, respectively.

Results

We found that carbachol increased the expression of NLRP3 inflammasome (NLRP3, ASC, cleaved caspase-1, IL-1β, and IL-18). The treated cells also showed an increase in NF-ĸB activation. The effect of carbachol was diminished by NF-ĸB inhibitor and atropine, a mAChR antagonist. The addition of RA also significantly reduced the effect of carbachol on NLRP3 inflammasomes.

Conclusions

Our current study suggests that carbachol induces NLRP3 inflammasome activation through mAChR and NF-ĸB, and that RA abolishes the inflammatory response. It reveals the potentials of co-administration of RA with cholinergic drugs to prevent inflammatory responses during cholinergic medications.

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References

  1. Beckmann J, Lips KS. The non-neuronal cholinergic system in health and disease. Pharmacol. 2013;92:286–302.

  2. Wessler I, Kirkpatrick CJ. Acetylcholine beyond neurons: the non-neuronal cholinergic system in humans. Br J Pharmacol. 2008;154(8):1558–71.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Eduardo CRC, Alejandra TIG, Guadalupe DRKJ, Herminia VRG, Lenin P, Enrique BV, et al. Modulation of the extraneuronal cholinergic system on main innate response leukocytes. J Neuroimmunol. 2019(327):22–35.

  4. Koarai A, Traves SL, Fenwick PS, Brown SM, Chana KK, Russell REK, et al. Expression of muscarinic receptors by human macrophages. Eur Respir J. 2012;39(3):698–704.

    Article  CAS  PubMed  Google Scholar 

  5. Kawashima K, Yoshikawa K, Fujii YX, Moriwaki Y, Misawa H. Expression and function of genes encoding cholinergic components in murine immune cells. Life Sci. 2007;80(24–25):2314–9.

    Article  CAS  PubMed  Google Scholar 

  6. Oenema TA, Kolahian S, Nanninga JE, Rieks D, Hiemstra PS, Zuyderduyn S, et al. Pro-inflammatory mechanisms of muscarinic receptor stimulation in airway smooth muscle. Respir Res. 2010;11:1–10.

    Article  Google Scholar 

  7. Guizzetti M, Moore NH, Vandemark KL, Giordano G, Costa LG. Muscarinic receptor-activated signal transduction pathways involved in the neuritogenic effect of astrocytes in hippocampal neurons. Eur J Pharmacol. 2011;659:102–7.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Gori S, Vermeulen M, Remes-Lenicov F, Jancic C, Scordo W, Ceballos A, et al. Acetylcholine polarizes dendritic cells toward a Th2-promoting profile. Allergy Eur J Allergy Clin Immunol. 2017;72(2):221–31.

    Article  CAS  Google Scholar 

  9. Xu ZP, Yang K, Xu GN, Zhu L, Hou LN, Zhang WH, et al. Role of M3 mAChR in in vivo and in vitro models of LPS-induced inflammatory response. Int Immunopharmacol. 2012;14(3):320–7.

    Article  CAS  PubMed  Google Scholar 

  10. Iho S, Tanaka Y, Takauji R, Kobayashi C, Muramatsu I, Iwasaki H, et al. Nicotine induces human neutrophils to produce IL-8 through the generation of peroxynitrite and subsequent activation of NF- ĸB. J Leukoc Biol. 2003;74(5):942–51.

    Article  CAS  PubMed  Google Scholar 

  11. Totti N 3rd, McCusker KT, Campbell EJ, Griffin GL, Senior RM. Nicotine is chemotactic for neutrophils and enhances neutrophil responsiveness to chemotactic peptides. Science. 1984;223(4632):169–71.

    Article  CAS  PubMed  Google Scholar 

  12. Hoover DB. Cholinergic modulation of the immune system presents new approaches for treating inflammation. Pharmacol Ther. 2017;179:1–16.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Yoshikawa H, Kurokawa M, Ozaki N, Nara K, Atou K, Takada E, et al. Nicotine inhibits the production of pro-inflammatory mediators in human monocytes by suppression of I-κB phosphorylation and nuclear factor-κB transcriptional activity through nicotinic acetylcholine receptor α7. Clin Exp Immunol. 2006;146(1):116–23.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Schroder K, Tschopp J. The inflammasomes. Cell. 2010;140(6):821–32.

    Article  CAS  PubMed  Google Scholar 

  15. Lamkanfi M, Dixit VM. Mechanisms and functions of inflammasomes. Cell. 2014;157(5):1013–22.

    Article  CAS  PubMed  Google Scholar 

  16. Latz E, Xiao TS, Stutz A. Activation and regulation of the inflammasomes. Nat Rev Immunol. 2013;13(6):397–411.

    Article  CAS  PubMed  Google Scholar 

  17. Guo H, Callaway JB, Ting JPY. Inflammasomes: mechanism of action, role in disease, and therapeutics. Nat Med. 2015;21(7):677–87.

    Article  PubMed  PubMed Central  Google Scholar 

  18. Lu F, Lan Z, Xin Z, He C, Guo Z, Xia X, et al. Emerging insights into molecular mechanisms underlying pyroptosis and functions of inflammasomes in diseases. J Cell Physiol. 2020;235(4):3207–21.

    Article  CAS  PubMed  Google Scholar 

  19. Song N, Liu ZS, Xue W, Bai ZF, Wang QY, Dai J, et al. NLRP3 phosphorylation is an essential priming event for inflammasome activation. Mol Cell. 2017;68(1):185-197.e6.

    Article  CAS  PubMed  Google Scholar 

  20. Abderrazak A, Syrovets T, Couchie D, El Hadri K, Friguet B, Simmet T, et al. NLRP3 inflammasome: from a danger signal sensor to a regulatory node of oxidative stress and inflammatory diseases. Redox Biol. 2015;4:296–307.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Shen HH, Yang YX, Meng X, Luo XY, Li XM, Shuai ZW, et al. NLRP3: a promising therapeutic target for autoimmune diseases. Autoimmun Rev. 2018;17(7):694–702.

    Article  CAS  PubMed  Google Scholar 

  22. Duong V, Rochette-Egly C. The molecular physiology of nuclear retinoic acid receptors. From health to disease. Biochim Biophys Acta Mol Basis Dis. 2011;1812(8):1023–31.

    Article  CAS  Google Scholar 

  23. Napoli JL. Physiological insights into all-trans-retinoic acid biosynthesis. Biochim Biophys Acta Mol Cell Biol Lipids. 2012;1821(1):152–67.

    Article  CAS  Google Scholar 

  24. Erkelens MN, Mebius RE. Retinoic acid and immune homeostasis: a balancing act. Trends Immunol. 2017;38(3):168–80.

    Article  CAS  PubMed  Google Scholar 

  25. Nurrahmah Q, Madhyastha R, Madhyastha H, Purbasari B, Nakajima Y, Maruyama M. Retinoic acid abrogates LPS-induced inflammatory response via negative regulation of NF-kappa B/mir-21 signaling. Immunopharmacol Immunotoxicol. 2021;43 (3):299–308.

  26. Pappano AJ. Cholinoceptor-activating and cholinesterase-inhibiting drugs. In: Katzung BG, editor. Basic and clinical pharmacology, 12th ed. New York: McGraw-Hill; 2012. p. 97–113.

  27. Sava C, Iancu R, Corbu C. Intraocular pressure dynamics after carbachol administration during cataract surgery. Farmacia. 2014;62(3):564–69.

  28. Kawashima K, Fujii T. Extraneuronal cholinergic system in lymphocytes. Pharmacol Ther. 2000;86(1):29–48.

    Article  CAS  PubMed  Google Scholar 

  29. Shiroma  LO,  Costa  VP.  Parasympatomimetic.  In:  Shaarawy  TM,  Sherwood MB, Hitchings RA, Crowston JG, editors. Glaucoma, 2nd ed. London, England, UK: Elsevier/ Saunders; 2015. p. 577–582.

  30. Dorrington MG, Fraser IDC. NF-κB signaling in macrophages: Dynamics, crosstalk, and signal integration. Front Immunol. 2019;10:705.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Fujii T, Mashimo M, Moriwaki Y, Misawa H, Ono S, Horiguchi K, et al. Physiological functions of the cholinergic system in immune cells. J Pharmacol Sci. 2017;134(1):1–21.

    Article  CAS  PubMed  Google Scholar 

  32. Tartey S, Kanneganti TD. Differential role of the NLRP3 inflammasome in infection and tumorigenesis. Immunology. 2019;156(4):329–38.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. de la Torre E, Genaro AM, Ribeiro ML, Pagotto R, Pignataro OP, Sales ME. Proliferative actions of muscarinic receptors expressed in macrophages derived from normal and tumor bearing mice. Biochim Biophys Acta Mol Basis Dis. 2008;1782(2):82–9.

    Article  Google Scholar 

  34. Milara J, Cervera A, de Diego A, Sanz C, Juan G, Gavaldà A, et al. Non-neuronal cholinergic system contributes to corticosteroid resistance in chronic obstructive pulmonary disease patients. Respir Res. 2016;17(1):1–14.

    Article  Google Scholar 

  35. Salamone G, Lombardi G, Gori S, Nahmod K, Jancic C, Amaral MM, et al. Cholinergic modulation of dendritic cell function. J Neuroimmunol. 2011;236(1–2):47–56.

    Article  CAS  PubMed  Google Scholar 

  36. Singh GB, Kshirasagar N, Patibandla S, Puchchakayala G, Koka S, Boini KM. Nicotine instigates podocyte injury via NLRP3 inflammasomes activation. Aging (Albany NY). 2019;11(24):12810–21.

    Article  CAS  Google Scholar 

  37. Wu X, Zhang H, Qi W, Zhang Y, Li J, Li Z, et al. Nicotine promotes atherosclerosis via ROS-NLRP3-mediated endothelial cell pyroptosis. Cell Death Dis. 2018;9:171.

    Article  PubMed  PubMed Central  Google Scholar 

  38. Yu P, Zhang X, Liu N, Tang L, Peng C, Chen X. Pyroptosis: mechanisms and diseases. Signal Transduct Target Ther. 2021;6(1):1–21.

    PubMed  PubMed Central  Google Scholar 

  39. Conos SA, Lawlor KE, Vaux DL, Vince JE, Lindqvist LM. Cell death is not essential for caspase-1-mediated interleukin-1β activation and secretion. Cell Death Differ. 2016;23(11):1827–38.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Carty M, Kearney J, Shanahan KA, Hams E, Sugisawa R, Connolly D, et al. Cell survival and cytokine release after inflammasome activation is regulated by the Toll-IL-1R protein SARM. Immunity. 2019;50(6):1412-1424.e6.

    Article  CAS  PubMed  Google Scholar 

  41. Sutterwala FS, Haasken S, Cassel SL. Mechanism of NLRP3 inflammasome activation. Ann NY Acad Sci. 2014;1319(1):82–95.

    Article  CAS  PubMed  Google Scholar 

  42. Liu T, Zhang L, Joo D, Sun SC. NF-κB signaling in inflammation. Signal Transduct Target Ther. 2017;2:1–9.

    CAS  Google Scholar 

  43. Giridharan S, Srinivasan M. Mechanisms of NF-κB p65 and strategies for therapeutic manipulation. J Inflamm Res. 2018;11:407–19.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Pengjam Y, Madhyastha H, Madhyastha R, Yamaguchi Y, Nakajima Y, Maruyama M. NF-κB pathway inhibition by anthrocyclic glycoside aloin is key event in preventing osteoclastogenesis in RAW264.7 cells. Phytomedicine. 2016;23(4):417–28.

    Article  CAS  PubMed  Google Scholar 

  45. Nurrahmah Q, Madhyastha R, Madhyastha H, Nakajima Y, Maruyama M. Nuclear factor-kappa B and JNK mediate macrophage polarization shift induced by C-phycocyanin. Indian J Exp Biol. 2019;57:381–9.

    CAS  Google Scholar 

  46. Madhyastha R, Madhyastha H, Nurrahmah QI, Purbasari B, Maruyama M, Nakajima Y. MicroRNA 21 elicits a pro-inflammatory response in macrophages, with exosomes functioning as delivery vehicles. Inflamm. 2021;44(4):1274–1287.

  47. Guizzetti M, Bordi F, Dieguez-Acuña FJ, Vitalone A, Madia F, Woods JS, et al. Nuclear factor κB activation by muscarinic receptors in astroglial cells: effect of ethanol. Neuroscience. 2003;120(4):941–50.

    Article  CAS  PubMed  Google Scholar 

  48. Guizzetti M, Thompson BD, Kim Y, VanDeMark K, Costa LG. Role of phospholipase D signaling in ethanol-induced inhibition of carbachol-stimulated DNA synthesis of 1321N1 astrocytoma cells. J Neurochem. 2004;90(3):646–53.

    Article  CAS  PubMed  Google Scholar 

  49. Guizzetti M, Costa LG. Possible role of protein kinase C ζ in muscarinic receptor-induced proliferation of astrocytoma cells. Biochem Pharmacol. 2000;60(10):1457–66.

    Article  CAS  PubMed  Google Scholar 

  50. Li X, Song L, Jope RS. Cholinergic stimulation of AP-1 and NFκB transcription factors is differentially sensitive to oxidative stress in SH-SY5Y neuroblastoma: relationship to phosphoinositide hydrolysis. J Neurosci. 1996;16(19):5914–22.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Todisco A, Ramamoorthy S, Pausawasdi N, Tacey K. Carbachol activates IκB kinase in isolated canine gastric parietal cells. Biochem Biophys Res Commun. 1999;261(3):877–84.

    Article  CAS  PubMed  Google Scholar 

  52. Ye RD. Regulation of nuclear factor kappaB activation by G-protein-coupled receptors. J Leukoc Biol. 2001;70(6):839–48.

    Article  CAS  PubMed  Google Scholar 

  53. Profita M, Bonanno A, Siena L, Ferraro M, Montalbano AM, Pompeo F, et al. Acetylcholine mediates the release of IL-8 in human bronchial epithelial cells by a NFkB/ERK-dependent mechanism. Eur J Pharmacol. 2008;582(1–3):145–53.

    Article  CAS  PubMed  Google Scholar 

  54. Kim CH. Retinoic Acid, Immunity, and Inflammation. In: Litwack G, editor. Vitamins and the immune system, volume 86. 1st ed. Amsterdam, The Netherlands: Academic Press-Elsevier; 2011. p. 83–101. 

  55. Hong K, Zhang Y, Guo Y, Xie J, Wang J, He X, et al. All-trans retinoic acid attenuates experimental colitis through inhibition of NF-κB signaling. Immunol Lett. 2014;162(1):34–40.

    Article  CAS  PubMed  Google Scholar 

  56. Behairi N, Belkhelfa M, Rafa H, Labsi M, Deghbar N, Bouzid N, et al. All-trans retinoic acid (ATRA) prevents lipopolysaccharide-induced neuroinflammation, amyloidogenesis and memory impairment in aged rats. J Neuroimmunol. 2016;300:21–9.

    Article  CAS  PubMed  Google Scholar 

  57. Priyanka SH, Syam Das S, Thushara AJ, Rauf AA, Indira M. All trans retinoic acid attenuates markers of neuroinflammation in rat brain by modulation of SIRT1 and NFκB. Neurochem Res. 2018;43(9):1791–801.

    Article  CAS  PubMed  Google Scholar 

  58. Egea J, Buendia I, Parada E, Navarro E, León R, Lopez MG. Anti-inflammatory role of microglial alpha7 nAChRs and its role in neuroprotection. Biochem Pharmacol. 2015;97(4):463–72.

    Article  CAS  PubMed  Google Scholar 

  59. Goverse G, Stakenborg M, Matteoli G. The intestinal cholinergic anti-inflammatory pathway. J Physiol. 2016;594(20):5771–80.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  60. Czarnewski P, Das S, Parigi SM, Villablanca EJ. Retinoic acid and its role in modulating intestinal innate immunity. Nutrients. 2017;9(1):68.

    Article  PubMed Central  Google Scholar 

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Acknowledgements

This study was supported by a Monbukagakusho research scholarship from the Ministry of Education. Culture, Sports, Science, and Technology, Japan to BP.

Funding

This research received no external funding.

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Authors and Affiliations

  1. Division of Cardiovascular Physiology, Department of Basic Medical Sciences, Faculty of Medicine, University of Miyazaki, Kihara 5200, Kiyotake cho, Miyazaki, 889-1692, Japan

    Bethasiwi Purbasari, Radha Madhyastha, Harishkumar Madhyastha, Queen Intan Nurrahmah, Masugi Maruyama, Yuichi Nakajima & Nozomi Watanabe

  2. Section of Oncopathology and Regenerative Biology, Department of Pathology, Faculty of Medicine, University of Miyazaki, Kihara 5200, Kiyotake cho, Miyazaki, 889-1692, Japan

    Hiroaki Kataoka

Authors
  1. Bethasiwi Purbasari
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  2. Radha Madhyastha
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  3. Harishkumar Madhyastha
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  4. Queen Intan Nurrahmah
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  5. Masugi Maruyama
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  6. Yuichi Nakajima
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  7. Hiroaki Kataoka
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  8. Nozomi Watanabe
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Contributions

Investigation, analysis, and manuscript preparation—BP; Conceptualization, methodology, investigation, and manuscript editing—MR and MH; Investigation, analysis—NQI; Supervision, review, and editing—MM, NY, KH, and NW.

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Correspondence to Nozomi Watanabe.

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Purbasari, B., Madhyastha, R., Madhyastha, H. et al. Retinoic acid attenuates nuclear factor kappaB mediated induction of NLRP3 inflammasome. Pharmacol. Rep 74, 189–203 (2022). https://doi.org/10.1007/s43440-021-00321-4

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  • DOI: https://doi.org/10.1007/s43440-021-00321-4

Keywords

  • Acetylcholine
  • Macrophage
  • NLRP3 inflammasomes
  • NF-ĸB
  • Muscarinic receptor