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Exosomal TNF-α mediates voltage-gated Na+ channel 1.6 overexpression and contributes to brain tumor–induced neuronal hyperexcitability
Cesar Adolfo Sanchez Trivino, … , Fabrizia Cesca, Vincent Torre
Cesar Adolfo Sanchez Trivino, … , Fabrizia Cesca, Vincent Torre
Published August 1, 2024
Citation Information: J Clin Invest. 2024;134(18):e166271. https://doi.org/10.1172/JCI166271.
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Research Article Neuroscience Oncology

Exosomal TNF-α mediates voltage-gated Na+ channel 1.6 overexpression and contributes to brain tumor–induced neuronal hyperexcitability

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Abstract

Patients affected by glioma frequently experience epileptic discharges; however, the causes of brain tumor–related epilepsy (BTRE) are still not completely understood. We investigated the mechanisms underlying BTRE by analyzing the effects of exosomes released by U87 glioma cells and by patient-derived glioma cells. Rat hippocampal neurons incubated for 24 hours with these exosomes exhibited increased spontaneous firing, while their resting membrane potential shifted positively by 10–15 mV. Voltage clamp recordings demonstrated that the activation of the Na+ current shifted toward more hyperpolarized voltages by 10–15 mV. To understand the factors inducing hyperexcitability, we focused on exosomal cytokines. Western blot and ELISAs showed that TNF-α was present inside glioma-derived exosomes. Remarkably, incubation with TNF-α fully mimicked the phenotype induced by exosomes, with neurons firing continuously, while their resting membrane potential shifted positively. Real-time PCR revealed that both exosomes and TNF-α induced overexpression of the voltage-gated Na+ channel Nav1.6, a low-threshold Na+ channel responsible for hyperexcitability. When neurons were preincubated with infliximab, a specific TNF-α inhibitor, the hyperexcitability induced by exosomes and TNF-α was drastically reduced. We propose that infliximab, an FDA-approved drug to treat rheumatoid arthritis, could ameliorate the conditions of glioma patients with BTRE.

Authors

Cesar Adolfo Sanchez Trivino, Renza Spelat, Federica Spada, Camilla D’Angelo, Ivana Manini, Irene Giulia Rolle, Tamara Ius, Pietro Parisse, Anna Menini, Daniela Cesselli, Miran Skrap, Fabrizia Cesca, Vincent Torre

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Figure 5

Increased spontaneous firing is associated with a shift of the activation curve of the inward voltage-dependent Na+ current.

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Increased spontaneous firing is associated with a shift of the activatio...
(A) Representative traces of voltage-dependent Na+ current activation in the presence of 140 mM (gray) and 70 mM (purple) NaCl. (B) Normalized currents in A show similar voltage activation dependence. Kruskal-Wallis test, P > 0.05, n = 5. (C) Replacement of NaCl by NMDG-Cl shows a residual small inward current (orange) blocked by addition of CdCl2 100 μM (brown); n = 5. (D) Representative traces of voltage-dependent Na+ currents obtained with CdCl 100 μM in the extracellular solution and internal solution with CsCl 135 mM plus NaCl 5 mM. Currents were recorded with the protocols used for activation and inactivation in control (black: n = 7–10) and U87 exosome–treated (blue: n = 12–14) neurons. (E) I-V curve comparing current density as in D, showing increased current in treated neurons (Kruskal-Wallis test, **P < 0.01). (F) Dependence of normalized conductance G/max G as a function of voltage for control and treated neurons, showing an average V1/2 shift of –6.4 mV (values in main text; *P < 0.05, Mann-Whitney U test). (G) Representative traces of Na+ currents activated under control conditions (black, n = 7) and in neurons treated for 24 hours with exosomes from patient S479 (green, n = 7). (H) Average I-V curves of Na+ currents for control and patient exosome–treated neurons, showing higher Na+ current density in cells treated with patients’ exosomes (**P < 0.01, Kruskal-Wallis test). (I) Normalized conductance curve for the conditions in H showing early activation of the Na+ conductance of neurons treated with patients’ exosomes (V1/2 shift of –12 ± 7.2 mV, values in main text; **P < 0.01, Mann-Whitney U test). Voltage clamp protocols to test Na+ current activation and inactivation are shown on the left of each panel.

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