O. Braissant¹, H. Henry¹, P. Jafari², C. Roux Petronelli¹, D. Ballhausen²

¹Biomedecine, Centre Hospitalier Universitaire Vaudois, Lausanne, Switzerland, ²Div Mol Ped, Centre Hospitalier Universitaire Vaudois, Lausanne, Switzerland

Introduction: Neurological damage is a common feature in methylmalonic aciduria (MMA). Cerebral accumulation of toxic metabolites upstream of the metabolic block is considered to be the main cause of neuronal damage, while the pathomechanism of neurodegeneration is still poorly understood. MMA manifests usually early in life. Cerebral maturation seems to play an important role in this age-dependant vulnerability. Methods: We treated rat 3D primary reaggregated brain cell cultures with the metabolites accumulating in MMA namely 2-methylcitric acid, methylmalonic acid and propionic acid. We analyzed the biochemical profile in the medium. Experiments were performed at three different time points of the 3D cultures reflecting a time window between the neonatal period and childhood. Results: 2-methylcitric acid treatment induced a high concentration of ammonia up to 2.8 mmol/l in the media at day 8 compared to 0.75 mmol/l in controls; 2.1 and 2.0 mmol/l versus 0.1 mmol/l respectively at day 14 and day 21. None of the other potential toxins produced an effect of equal amplitude. Concomitant with the increase of ammonia there was a significant decrease in glutamine from 1.5 mM for controls to 0.7 mM at day 8; and from 2.7 mmol/l to 1.5 and 1.7 mmol/l respectively at day 14 and 21. Conclusion: We found that 2-methylcitrate is the most toxic metabolite in MMA and that its toxicity is mediated by ammonia, with glutamine involved in its production. We have previously observed in the same culture system that ammonia above 1 mmol/l in the medium impaired axonal growth. The observed production of ammonia might be restricted to the brain and not be visible in the blood of patients. High cerebral ammonia may point to a not yet described pathomechanism in MMA and is a possible therapeutic target. We also confirm the clinical observation that toxicity seems to decrease with brain cell maturation. Outlook: Investigations on respiratory chain function, metabolomics and gene expression profiling are ongoing and might help us to further determine the pathways involved in neurotoxicity.