Effect of N-acetylaspartate and FDA-approved drugs on microglial cells

Neuroinflammation is a key feature shared by several neurodegenerative conditions. While it initially serves as a protective mechanism against external insults, sustained or chronic neuroinflammation can become detrimental, impeding neuronal regeneration. Within the central nervous system (CNS), microglia, the resident immune cells, play a pivotal role in chronic inflammatory processes, particularly due to an imbalance between their neurotoxic (M1) and neuroprotective (M2) phenotypes. The M1 phenotype drives pro-inflammatory responses, relying on glycolytic metabolism for rapid ATP production, whereas M2 microglia are involved in tissue repair and anti-inflammatory functions, utilizing oxidative metabolism to sustain long-term activities4–6 . CNS homeostasis is tightly regulated by intricate communication between neurons and glial cells, with neurons releasing signal metabolites into the extracellular environment that can modulate glial cell functions. N-acetylaspartate (NAA) is the second most abundant amino acid in the human brain, reaching very high concentrations even close to 10 mM7. NAA is synthesized from aspartate and acetyl-CoA via aspartate N-acetyltransferase (NAT8L) in neuronal mitochondria and is subsequently catabolized in acetate and aspartate by cells expressing the enzyme aspartoacylase (ASPA). The acetate can then be converted into acetylCoA, serving as a precursor for lipid synthesis8. Although NAA has long been recognized as one of the most abundant metabolites in the CNS, our understanding of its physiological role in the brain remains limited, with most studies focusing on its interactions with oligodendrocytes and astrocytes. Alterations in NAA metabolism are typical of neurodegenerative disorders, particularly demyelinating diseases such as Multiple Sclerosis (MS), where low levels of brain NAA are observed. Despite the established link between neurodegenerative disorders, NAA dysmetabolism, and neuroinflammatory processes, the role of NAA in microglial cells has not yet been explored. Here, we present evidence of an anti-inflammatory effect of NAA in BV2 cells, a finding also confirmed in primary microglial cells. Upon exposure to NAA, microglial cells undergo metabolic reprogramming characterized by an upregulation in lipid synthesis and degradation turnover. Functionally, this reprogramming enhances the cells' migratory capacity and phagocytic activity. Notably, NAA reduces iNOS protein levels, intracellular NO content, and mRNA levels of TNF-α and IL-6 by modulating the levels and translocation of transcription factors NF-κB and STAT1. Investigating the molecular mechanisms underlying this signaling process, we suggest that histone deacetylases (HDACs) could play a critical role, as we observed increased HDAC1 mRNA and protein levels following NAA treatment. Moreover, inhibiting these enzymes counteracted the effects of NAA. To further explore therapeutic strategies for neurodegenerative diseases, the analysis of FDA-approved drugs originally developed for nonneurodegenerative purposes helped identify those with potential therapeutic effects in mitigating neuroinflammation. Among the drugs evaluated, PARP inhibitors demonstrated the ability to modulate microglial polarization. This finding underscores the potential of drug repurposing as a strategy to develop effective treatments for neurodegenerative diseases, offering a promising avenue for therapeutic intervention.