Nicotinamide adenine dinucleotide treatment alleviates the symptoms of experimental autoimmune encephalomyelitis by activating autophagy and inhibiting the NLRP3 inflammasome

Int Immunopharmacol. 2021 Jan:90:107092. doi: 10.1016/j.intimp.2020.107092. Epub 2020 Dec 4.

Abstract

Nicotinamide adenine dinucleotide (NAD + ) is an essential cofactor in numerous metabolic pathways, and so may support protective and reparative processes against central nervous system diseases such as multiple sclerosis (MS). Here, we investigated the therapeutic potential of NAD + administration in the experimental autoimmune encephalomyelitis (EAE) mouse model of MS and the contributions of autophagic regulation and NLRP3 inflammasome activity. EAE was induced in female C57BL/6 mice by immunization with myelin oligodendrocyte glycoprotein (MOG) p35-55 and disease severity analyzed by neurological function score and histological scores of spinal cord sections stained with hematoxylin-eosin or luxol fast blue. Outcomes were compared among control mice and EAE groups receiving daily post-immunization vehicle injections, NAD + injections, injection of the autophagy inhibitor 3-methyladenine (3-MA), or co-injection of NAD + and 3-MA. Expression levels of autophagy-related proteins (Beclin1, LC3-II/I, and p62/SQSTM1) were assessed by Western blotting, the activated microglial cells were evaluated by immunohistochemistry, while mRNA expression levels of NOD-like receptor family pyrin domain containing 3 (NLRP3), interleukin (IL)-1β, IL-2, IL-17, IL-18, interferon-γ (IFN-γ) and IL-10 were detected by real-time PCR. The proportions of Th1 and Th17 cells in spleen were evaluated using flow cytometry. Treatment with NAD + alleviated demyelination, nerve injury, microglial activation and motor function abnormalities of EAE mice. In addition, NAD + increased the expressions of the autophagy-related proteins LC3-II/I and Beclin 1, and reduced the expression of p62. Treatment with NAD + also inhibited the expressions of NLRP3 and modulated the differentiation of Th1 and Th17 cells, reduced the expressions of the pro-inflammatory factors IL-1β, IL-2, IL-18, IFN-γ and IL-17, and increased the expression of anti-inflammatory IL-10. Conversely, 3-MA aggravated spinal cord inflammation and demyelination, and delayed spontaneous remission from EAE. Furthermore, the beneficial effects of NAD + were abolished by 3-MA cotreatment. Our results indicate that NAD + suppresses the NLRP3 inflammasome at least in part through the activation of autophagy to relieve the symptoms of EAE. Therefore, regulation of autophagy by NAD + treatment may be an effective therapeutic strategy for MS.

Keywords: 3-MA; Autophagy; Inflammatory; Multiple sclerosis; Myelin-Oligodendrocyte Glycoprotein; NLRP3, NAD+.

MeSH terms

  • Animals
  • Autophagy / drug effects*
  • Autophagy-Related Proteins / metabolism
  • Cytokines / genetics
  • Cytokines / metabolism
  • Encephalomyelitis, Autoimmune, Experimental / immunology
  • Encephalomyelitis, Autoimmune, Experimental / metabolism
  • Encephalomyelitis, Autoimmune, Experimental / pathology
  • Encephalomyelitis, Autoimmune, Experimental / prevention & control*
  • Female
  • Inflammasomes / antagonists & inhibitors*
  • Inflammasomes / genetics
  • Inflammasomes / metabolism
  • Inflammation Mediators / metabolism
  • Mice
  • Mice, Inbred C57BL
  • Microglia / drug effects*
  • Microglia / immunology
  • Microglia / metabolism
  • Microglia / pathology
  • NAD / pharmacology*
  • NLR Family, Pyrin Domain-Containing 3 Protein / antagonists & inhibitors*
  • NLR Family, Pyrin Domain-Containing 3 Protein / genetics
  • NLR Family, Pyrin Domain-Containing 3 Protein / metabolism
  • Neurons / drug effects*
  • Neurons / immunology
  • Neurons / metabolism
  • Neurons / pathology
  • Signal Transduction
  • Spinal Cord / drug effects*
  • Spinal Cord / immunology
  • Spinal Cord / metabolism
  • Spinal Cord / pathology

Substances

  • Autophagy-Related Proteins
  • Cytokines
  • Inflammasomes
  • Inflammation Mediators
  • NLR Family, Pyrin Domain-Containing 3 Protein
  • Nlrp3 protein, mouse
  • NAD