Ali Tawbeh ¹ Catherine GONDCAILLE ^1,2 Fatima-Ezzhara Saih ¹ Quentin Raas ¹ Damien Loichot ¹ Yannick HAMON ³ Celine Keime ⁴ Alexandre Benani ^1,2 Francesca Di Cara ⁵ Mustapha CHERKAOUI-MALKI ^1,2 Pierre Andreoletti ^1,2 Stéphane Savary ^1,2*

• ¹ Université de Bourgogne, Dijon, France
• ² UMR6265 Centre des Sciences du Goût et de l'Alimentation (CSGA), Dijon, Burgundy, France
• ³ INSERM U1104 Centre d'immunologie de Marseille-Luminy (CIML), Marseille, Provence-Alpes-Côte d'Azur, France
• ⁴ Université de Strasbourg, Strasbourg, Alsace, France
• ⁵ Dalhousie University, Halifax, Nova Scotia, Canada

Microglia, the immune cells of the central nervous system, activate neuroinflammatory pathways in response to homeostatic disturbances, a process implicated in the pathogenesis of various neurodegenerative diseases. Emerging evidences identify abnormal microglial activation as a causal factor at the onset of peroxisomal leukodystrophies, including X-linked adrenoleukodystrophy (X-ALD). This study investigates how primary peroxisomal deficiencies influence oxidative properties of microglia, and examines the subsequent impact on neurons and oligodendrocytes. Using BV-2 microglial cells lacking ABCD1, ABCD2, or ACOX1, peroxisomal proteins that play key roles in the very-long chain fatty acid beta-oxidation, we analyzed their response under basal condition and after stimulation by lipopolysaccharide (LPS). Transcriptomic analysis of the mutant microglial cells revealed numerous differentially expressed genes, particularly in redox-related pathways following LPS exposure. These changes are consistent with the increased production of reactive oxygen species (ROS) and nitric oxide (NO). Conditioned media (CM) from the mutant cells were then applied to cultures of neuron and oligodendrocyte cell lines. Exposure to CM from LPS-stimulated mutant microglial cells significantly increased apoptosis in both cell types. Furthermore, treated neurons exhibited a reduction in cell complexity and an increased ability to secrete neuropeptides. These findings demonstrate that peroxisomal impairments in microglia exacerbate inflammatory response and ROS/NO production, affecting the survival of neurons and oligodendrocytes, as well as neuronal morphology and function. This dysfunction might contribute to the early neurodegenerative events in X-ALD by triggering and sustaining neuroinflammatory cascades. Therapeutic strategies that target microglial activation and secretion profiles could hold promise in managing peroxisomal disorders such as X-ALD.a supprimé: Recombinant mouse TNF (R&D Systems 410-MT-01/CF) was used at concentrations of 0.447 ng/mL or 1.26 ng/mL, the NO donor S-Nitro-N-acetylpenicillamine (SNAP -Sigma-Aldrich N3398) at 6 μM or 50 µM, and CTSB (Acro Biosystems CTB-M52H9) at 155 ng/mL or 460 ng/mL or 2518 ng/mL.