CEREBROSPINAL FLUID FROM NEWLY DIAGNOSED RELAPSING-REMITTING MULTIPLE SCLEROSIS PATIENTS INDUCES GLUTAMATE-MEDIATED TOXICITY IN PRIMARY HIPPOCAMPAL NEURONAL CULTURES

ARMONELLI S; MARCHESSINI N; VILCAES AA; PATRUCCO L; CRISTIANO E; BISBAL M; ROJAS JI; DEGANO AL; LOPEZ, P.

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Congreso; LXVII REUNIÓN ANUAL DE LA SOCIEDAD ARGENTINA DE INVESTIGACIÓN CLÍNICA (SAIC); 2022

SOCIEDAD ARGENTINA DE INVESTIGACIÓN CLÍNICA (SAIC)

Glutamate (Glu) toxicity has been postulated as a factor contributingto neurodegeneration in Multiple Sclerosis (MS), and is currentlyunder study as a biomarker for neurodegeneration. We recently describeda correlation between high Glu concentration in cerebrospinalfluid (CSF) and different imaging parameters of neural atrophyin newly diagnosed Relapsing-Remitting MS (RRMS) patients. Theaim of this work was to demonstrate a direct neurotoxic effect ofGlu in the CSF from MS patients on rat primary hippocampal neuronal(HN) cultures. CSF samples from 18 newly diagnosed RRMSpatients free of treatment and 18 control subjects were collected,and their toxicity tested by acute exposure on HN cultures previouslygrown for 8 days in vitro, then cells were further cultured foran additional 24h. The HN axodendritic network was visualized byimmunofluorescence using the neuron-specific marker β-3 tubulin.Glu-mediated neurotoxicity in CSF was identified by pre-treatmentof cultures with MK-801, a non-competitive NMDA receptor antagonist;or pre-treatment of CSF with the enzyme Glu dehydrogenase(GDH), which converts Glu in α-ketoglutarate. Acute exposure ofCSF from control subjects to HN did not induce significant neurotoxicitywhen compared to control (untreated) cultures. On the contrary,exposure of CSF from RRMS to HN resulted in a robust axodendriticnetwork loss (t-test; p<0,0001; mean 40 ± 0,06 %). Pre-treatmentwith MK-801 fully protected cultures from RRMS-derived CSF neurotoxicity(p=0,004, n=6, two way ANOVA). Similarly, GDH-mediated degradation of Glu present in CSF from RRMS prevented neurotoxicity(p=0,0008; n=6). Overall, our results confirm that Glu present inCSF from RRMS patients exerts axodendritic damage in HN. Thisdata support the role of Glu as a biomarker to predict and monitordisease progression of RRMS patients, and identify a therapeutictarget to prevent neurodegeneration associated with clinical progression.