CXCL1-CXCR1/2 signaling is induced in human temporal lobe epilepsy and contributes to seizures in a murine model of acquired epilepsy

Abstract CXCL1, a functional murine orthologue of the human chemokine CXCL8 (IL-8), and its CXCR1 and CXCR2 receptors were investigated in a murine model of acquired epilepsy developing following status epilepticus (SE) induced by intra-amygdala kainate. CXCL8 and its receptors were also studied in human temporal lobe epilepsy (TLE). The functional involvement of the chemokine in seizure generation and neuronal cell loss was assessed in mice using reparixin (formerly referred to as repertaxin), a non-competitive allosteric inhibitor of CXCR1/2 receptors. We found a significant increase in hippocampal CXCL1 level within 24 h of SE onset that lasted for at least 1 week. No changes were measured in blood. In analogy with human TLE, immunohistochemistry in epileptic mice showed that CXCL1 and its two receptors were increased in hippocampal neuronal cells. Additional expression of these molecules was found in glia in human TLE. Mice were treated with reparixin or vehicle during SE and for additional 6 days thereafter, using subcutaneous osmotic minipumps. Drug-treated mice showed a faster SE decay, a reduced incidence of acute symptomatic seizures during 48 h post-SE, and a delayed time to spontaneous seizures onset compared to vehicle controls. Upon reparixin discontinuation, mice developed spontaneous seizures similar to vehicle mice, as shown by EEG monitoring at 14 days and 2.5 months post-SE. In the same epileptic mice, reparixin reduced neuronal cell loss in the hippocampus vs vehicle-injected mice, as assessed by Nissl staining at completion of EEG monitoring. Reparixin administration for 2 weeks in mice with established chronic seizures, reduced by 2-fold on average seizure number vs pre-treatment baseline, and this effect was reversible upon drug discontinuation. No significant changes in seizure number were measured in vehicle-injected epileptic mice that were EEG monitored in parallel. Data show that CXCL1-IL-8 signaling is activated in experimental and human epilepsy and contributes to acute and chronic seizures in mice, therefore representing a potential new target to attain anti-ictogenic effects

Biomarkers of Epileptogenesis and Novel Therapeutic Approaches in Animal Models of Acquired Epilepsy

Epilepsy is a chronic neurological disorder that affects over 50 million people worldwide. The current therapy is mainly based on symptomatic drugs acting on seizures but these drugs do not affect the underlying mechanisms of the disease. Moreover, despite more than 20 available antiseizure drugs, seizures remain uncontrolled in ~30% of patients. Hence, there is an urgent need to find novel therapeutics to improve the management of seizures and the neurological comorbidities. Epilepsy may occur after an acquired epileptogenic insult (e.g., traumatic brain injury, status epilepticus, CNS infections) and the risk of developing the disease varies among individuals exposed to similar injuries. Prevention of epilepsy is hampered by the lack of sensitive and specific prognostic biomarkers that may enable the identification of the individuals at risk of developing epilepsy. This prediction would greatly facilitate clinical trials of anti-epileptogenic treatments. Animal models are required to investigate the pathological mechanisms driving the transition from the epileptogenic insult to clinical onset and progression of epilepsy. These models are instrumental for discovering novel targets and developing effective treatments and biomarkers of epileptogenesis. Based on these clinical needs, we characterized two clinically relevant rodent models of epileptogenesis of differing etiology (traumatic brain injury, status epilepticus) with a 50-60% epilepsy incidence to identify potential biomarkers by performing EEG, MRI and behavioural measurements. Moreover, we studied the activation and the role of the IL-8 signaling in pharmacoresistant epilepsy using human temporal lobe epilepsy brain specimens and a wellestablished mouse model. This chemokine was reported to be increased in biospecimens of patients with drug-resistat seizures. Our studies provided evidence of prognostic biomarkers of epileptogenesis with clinical translatability, and a new druggable target for controlling pharmacoresistant seizures

Reorganization of Thalamic Inputs to Lesioned Cortex Following Experimental Traumatic Brain Injury

Traumatic brain injury (TBI) disrupts thalamic and cortical integrity. The effect of post-injury reorganization and plasticity in thalamocortical pathways on the functional outcome remains unclear. We evaluated whether TBI causes structural changes in the thalamocortical axonal projection terminals in the primary somatosensory cortex (S1) that lead to hyperexcitability. TBI was induced in adult male Sprague Dawley rats with lateral fluid-percussion injury. A virus carrying the fluorescent-tagged opsin channel rhodopsin 2 transgene was injected into the ventroposterior thalamus. We then traced the thalamocortical pathways and analyzed the reorganization of their axonal terminals in S1. Next, we optogenetically stimulated the thalamocortical relays from the ventral posterior lateral and medial nuclei to assess the post-TBI functionality of the pathway. Immunohistochemical analysis revealed that TBI did not alter the spatial distribution or lamina-specific targeting of projection terminals in S1. TBI reduced the axon terminal density in the motor cortex by 44% and in S1 by 30%. A nematic tensor-based analysis revealed that in control rats, the axon terminals in layer V were orientated perpendicular to the pial surface (60.3°). In TBI rats their orientation was more parallel to the pial surface (5.43°, difference between the groups p < 0.05). Moreover, the level of anisotropy of the axon terminals was high in controls (0.063) compared with TBI rats (0.045, p < 0.05). Optical stimulation of the sensory thalamus increased alpha activity in electroencephalography by 312% in controls (p > 0.05) and 237% (p > 0.05) in TBI rats compared with the baseline. However, only TBI rats showed increased beta activity (33%) with harmonics at 5 Hz. Our findings indicate that TBI induces reorganization of thalamocortical axonal terminals in the perilesional cortex, which alters responses to thalamic stimulation

Misure critiche. Nuova Serie. A. 14, n. 2 (2015)

N.S. A. 14, n. 2 (2015) : Di Lorenzo E., Aspetti della prosa artistica del “De re rustica” di Columella, P. 5 ; Galli Q., Dalla tradizione alla rivolta protoromantica, P. 26 ; Beniscelli A., Teatro e allegoria: un percorso settecentesco, P. 46 ; Pascale V., Mario Puzo. “The Fortunate Pilgrim”: sogni incondizionati, P. 64 ; Mauro C., Sulla poesia di Antonio Spagnuolo (con una lettera inedita di Umberto Saba), p. 85 ; Reina L., “CartaStraccia”. Sulle tracce dell’AntiPoesia, P. 107 ; Martelli S., “Per civile conversazione”. Sugli studi per Amedeo Quondam, P. 123 ; Sapia G., Un gratuito tormentone dantesco ‘E ’l modo ancor m’offende’ (Inf. V, 102), P. 134 ; Salsano R., Una poetica di ombra e luce: “Asfalto” di Valerio Mello, P. 141 ; Aruta Stampacchia A., “Footsophie. Le foot comme identité et fictionnement médiatique en Italie et ailleurs” di Gius Gargiulo: tra tragedia moderna e terminologia di confronto con il mondo, P. 150 ; Mezzasalma C., “La finzione il nulla” di Luigi Martellini, p. 161 ; Tenzone G., Le dolci rime d’amor ch’io solea, P. 169 ; Cesaro R., recensione a, Valorosa vipera gentile. Poesia e letteratura in volgare attorno ai Visconti fra Trecento e primo Quattrocento, a cura di Simone Albonico, Marco Limongelli, Barbara Pagliari, Roma, Viella 2014, P. 173 ; Terracciano R., recensione a, Lorenzo Da Ponte, Lettere a Guglielmo Piatti (1826-1836), a cura di Laura Paolino, Stony Brook (New York), Forum Italicum Publishing 2013, P. 176 ; di Brigida M., recensione a, Georges Hérelle, Gabriele d’Annunzio, ou théorie et pratique de la surhumanité, a cura di Mario Cimini, Berna, Peter Lang 2015, P. 179 ; Farina M., recensione a, Idolina Landolfi, “Il piccolo vascello solca i mari”. Tommaso Landolfi e i suoi editori. Bibliografia degli scritti di e su Landolfi (1929-2006), Fiesole, Cadmo 2015, P. 182

Cholesterol 24-hydroxylase is a novel pharmacological target for anti-ictogenic and disease modification effects in epilepsy

Therapies for epilepsy mainly provide symptomatic control of seizures since most of the available drugs do not target disease mechanisms. Moreover, about one-third of patients fail to achieve seizure control. To address the clinical need for disease-modifying therapies, research should focus on targets which permit interventions finely balanced between optimal efficacy and safety. One potential candidate is the brain-specific enzyme cholesterol 24-hydroxylase. This enzyme converts cholesterol to 24S-hydroxycholesterol, a metabolite which among its biological roles modulates neuronal functions relevant for hyperexcitability underlying seizures. To study the role of cholesterol 24-hydroxylase in epileptogenesis, we administered soticlestat (TAK-935/OV935), a potent and selective brain-penetrant inhibitor of the enzyme, during the early disease phase in a mouse model of acquired epilepsy using a clinically relevant dose. During soticlestat treatment, the onset of epilepsy was delayed and the number of ensuing seizures was decreased by about 3-fold compared to vehicle-treated mice, as assessed by EEG monitoring. Notably, the therapeutic effect was maintained 6.5 weeks after drug wash-out when seizure number was reduced by about 4-fold and their duration by 2-fold. Soticlestat-treated mice showed neuroprotection of hippocampal CA1 neurons and hilar mossy cells as assessed by post-mortem brain histology. High throughput RNA-sequencing of hippocampal neurons and glia in mice treated with soticlestat during epileptogenesis showed that inhibition of cholesterol 24-hydroxylase did not directly affect the epileptogenic transcriptional network, but rather modulated a non-overlapping set of genes that might oppose the pathogenic mechanisms of the disease. In human temporal lobe epileptic foci, we determined that cholesterol 24-hydroxylase expression trends higher in neurons, similarly to epileptic mice, while the enzyme is ectopically induced in astrocytes compared to control specimens. Soticlestat reduced significantly the number of spontaneous seizures in chronic epileptic mice when was administered during established epilepsy. Data show that cholesterol 24-hydroxylase contributes to spontaneous seizures and is involved in disease progression, thus it represents a novel target for chronic seizures inhibition and disease-modification therapy in epilepsy

A systems-level analysis highlights microglial activation as a modifying factor in common epilepsies.

AIMS The causes of distinct patterns of reduced cortical thickness in the common human epilepsies, detectable on neuroimaging and with important clinical consequences, are unknown. We investigated the underlying mechanisms of cortical thinning using a systems-level analysis. METHODS Imaging-based cortical structural maps from a large-scale epilepsy neuroimaging study were overlaid with highly spatially resolved human brain gene expression data from the Allen Human Brain Atlas. Cell-type deconvolution, differential expression analysis and cell-type enrichment analyses were used to identify differences in cell-type distribution. These differences were followed up in post-mortem brain tissue from humans with epilepsy using Iba1 immunolabelling. Furthermore, to investigate a causal effect in cortical thinning, cell-type-specific depletion was used in a murine model of acquired epilepsy. RESULTS We identified elevated fractions of microglia and endothelial cells in regions of reduced cortical thickness. Differentially expressed genes showed enrichment for microglial markers and, in particular, activated microglial states. Analysis of post-mortem brain tissue from humans with epilepsy confirmed excess activated microglia. In the murine model, transient depletion of activated microglia during the early phase of the disease development prevented cortical thinning and neuronal cell loss in the temporal cortex. Although the development of chronic seizures was unaffected, the epileptic mice with early depletion of activated microglia did not develop deficits in a non-spatial memory test seen in epileptic mice not depleted of microglia. CONCLUSIONS These convergent data strongly implicate activated microglia in cortical thinning, representing a new dimension for concern and disease modification in the epilepsies, potentially distinct from seizure control