Silencing PKM2 Attenuates Brain Injury Induced by Status Epilepticus by Inhibiting the AKT/mTOR Pathway and the NLRP3 Inflammasome.

PKM2 is a glycolytic pyruvate kinase isoenzyme, and its role in neurological diseases has been published. However, the role and mechanism of PKM2 in the process of status epilepticus have not been reported. The purpose of this study is to explore the role and mechanism of PKM2 in epilepsy. Quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting were used to explore the expression of PKM2 in cells. Enzyme-linked immunosorbent assay kits were used to evaluate the level of inflammatory factors. An epilepsy model was established by intraperitoneal injection of lithium chloride in rats. Various behavioural assays were conducted to explore the learning ability and cognitive level of rats. PKM2 expression was upregulated in Mg2+-induced hippocampal neurons. PKM2 inhibition ameliorated Mg2+-induced hippocampal neuronal inflammation and reduced neuronal apoptosis. In addition, PKM2 silencing inhibited the metabolic dysfunction of Mg2+-induced hippocampal neurons. Subsequent experiments showed that the Akt/mTOR pathway and NLRP3 inflammasome are involved in PKM2-mediated neuronal regulation. More importantly, PKM2 inhibition could alleviate status epilepticus in rats. PKM2 inhibition attenuates Mg2+-induced hippocampal neuronal inflammation, apoptosis and metabolic dysfunction and improves the cognitive ability of rats. Therefore, PKM2 may be an important target for epilepsy treatment.

Hydrogen Alleviates Necroptosis and Cognitive Deficits in Lithium-Pilocarpine Model of Status Epilepticus.

Status epilepticus without prompt seizure control always leads to neuronal death and long-term cognitive deficits, but effective intervention is still absent. Here, we found that hydrogen could alleviate the hippocampus-dependent spatial learning and memory deficit in lithium-pilocarpine model of status epilepticus in rats, as evidenced by the results in Morris water maze test. Hydrogen treatment downregulated the expression of necroptosis-related proteins, such as MLKL, phosphorylated-MLKL, and RIPK3 in hippocampus, and further protected neurons and astrocytes from necroptosis which was here first verified to occur in status epilepticus. Hydrogen also protected cells from apoptosis, which was indicated by the decreased cleaved-Caspase 3 expression. Meanwhile, Iba1+ microglial activation by status epilepticus was reduced by hydrogen treatment. These findings confirm the utility of hydrogen treatment in averting cell death including necroptosis and alleviating cognitive deficits caused by status epilepticus. Therefore, hydrogen may provide a potential and powerful clinical treatment for status epilepticus-related cognitive deficits.

Early Predictors of Drug-Resistant Epilepsy Development after Convulsive Status Epilepticus.

BACKGROUND: Drug-resistant epilepsy (DRE) is a common and serious consequence of convulsive status epilepticus (CSE). Little is known on the early prediction of DRE development after CSE. Our aim was to identify independent DRE predictors in patients with CSE. METHODS: One hundred and forty consecutive patients identified with CSE in a tertiary academic hospital between March 2008 and January 2015 were reviewed. Demographics, clinical features, serum albumin neuroimaging, and electroencephalogram characteristics were collected and analyzed. Independent predictors of DRE were identified using multivariate logistic regression. The receiver operating characteristic (ROC) curve was used to quantify the predictive validity of all the risk factors. RESULTS: After a median 62-month observation period, 91 patients were enrolled into this study. Thirty-seven (40.7%) patients did not have DRE, 22 (24.2%) developed DRE, and 32 (35.2%) were dead. History of epilepsy (OR 9.17, 95% CI 1.77-49.22, p = 0.010), status epilepticus duration ≥24 h (OR 4.82, 95% CI 1.04-22.37, p = 0.044), and cortical or hippocampal abnormalities on neuroimaging (OR 9.49, 95% CI 1.90-47.50, p = 0.006) were independent predictors of DRE after CSE. A combination of these 3 variables yielded an area under the ROC curve of 0.77 (0.65-0.89). CONCLUSIONS: History of epilepsy, longer SE duration, and cortical or hippocampal abnormalities on neuroimaging are early predictors for the development of DRE after CSE. Further studies are needed to assess whether a more aggressive treatment will reduce the likelihood of DRE development in these high-risk patients.

Melatonin Alleviates the Epilepsy-Associated Impairments in Hippocampal LTP and Spatial Learning Through Rescue of Surface GluR2 Expression at Hippocampal CA1 Synapses.

Epilepsy-associated cognitive impairment is common, and negatively impacts patients' quality of life. However, most antiepileptic drugs focus on the suppression of seizures, and fewer emphasize treatment of cognitive dysfunction. Melatonin, an indolamine synthesized primarily in the pineal grand, is reported to be neuroprotective against several central nervous system disorders. In this study, we investigated whether melatonin could reverse cognitive dysfunction in lithium-pilocarpine treated rats. Chronic treatment with melatonin (8 mg/kg daily for 15 days) after induction of status epilepticus significantly alleviated seizure severity, reduced neuronal death in the CA1 region of the hippocampus, improved spatial learning (as measured by the Morris water maze test), and reversed LTP impairments, compared to vehicle treatment. Furthermore, we found that melatonin rescued the decreased surface levels of GluR2 in the CA1 region observed in epilepsy, which might be the underlying mechanism of the neuroprotective and synapse-modulating function of melatonin. Our study provides experimental evidence for the possible clinical utility of melatonin as an adjunctive therapy to prevent epilepsy-associated cognitive impairments.

Hydrogen treatment reduces electroencephalographic activity and neuronal death in rats with refractory status epilepticus by inhibiting membrane NR2B phosphorylation and oxidative stress.

OBJECTIVE: To investigate the effects of hydrogen therapy on epileptic seizures in rats with refractory status epilepticus and the underlying mechanisms. METHODS: Status epilepticus was induced using pilocarpine. The effects of hydrogen treatment on epilepsy severity in model rats were then monitored using Racine scores and electroencephalography (EEG), followed by western blot of plasma membrane N-methyl-D-aspartate receptor subtype 2B (NR2B) and phosphorylated NR2B expression. We also generated a cellular epilepsy model using Mg2+-free medium and used polymerase chain reaction to investigate the neuroprotective effects of hydrogen. RESULTS: There were no significant differences in Racine scores between the hydrogen and control groups. EEG amplitudes were lower in the hydrogen treatment group than in the control group. In epilepsy model rats, hippocampal cell membrane NR2B expression and phosphorylation increased gradually over time. Although hippocampal cell membrane NR2B expression was not significantly different between the two groups, NR2B phosphorylation levels were significantly lower in the hydrogen group. Hydrogen treatment also increased superoxide dismutase, mitochondrial (SOD2) expression. CONCLUSIONS: Hydrogen treatment reduced EEG amplitudes and NR2B phosphorylation; it also decreased neuronal death by reducing oxidative stress. Hydrogen may thus be a potential treatment for refractory status epilepticus by inhibiting membrane NR2B phosphorylation and oxidative stress.

The anticonvulsant and neuroprotective effects of baicalin on pilocarpine-induced epileptic model in rats.

Baicalin, a flavonoid compound purified from plant Scutellaria baicalensis Georgi, has been reported to possess a wide variety of pharmacological properties including anti-oxidative, anti-apoptotic and neuroprotective properties. Oxidative stress can dramatically alter neuronal function and has been linked to status epilepticus (SE). However, the neuroprotective effect of baicalin on epilepsy is unclear. In this study we investigated whether Baicalin could exert anticonvulsant and neuroprotective effects in the pilocarpine-induced epileptic model in rats. To this end, we recorded the latency to first limbic seizure and SE and observed the incidence of SE and mortality. The changes of oxidative stress were measured 24 h after pilocarpine-induced SE. Nissl staining, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling and Fluoro-Jade B staining were performed to detect the neuronal loss, apoptosis and degeneration in hippocampus 72 h after pilocarpine-induced seizure. Pretreatment with baicalin significantly delayed the onset of the first limbic seizures and SE, reduced the mortality rate, and attenuated the changes in the levels of lipid peroxidation, nitrite content and reduced glutathione in the hippocampus of pilocarpine-treated rats. Furthermore, we also found that baicalin attenuated the neuronal cell loss, apoptosis, and degeneration caused by pilocarpine-induced seizures in rat hippocampus. Collectively, these results indicated remarkable anticonvulsant and neuroprotective effects of baicalin and should encourage further studies to investigate baicalin as an adjuvant in epilepsy both to prevent seizures and to protect against seizure induced brain injury.

Magnolol upregulates CHRM1 to attenuate Amyloid-ß-triggered neuronal injury through regulating the cAMP/PKA/CREB pathway.

Alzheimer's disease (AD) is a common neurodegenerative disease characterized by neuronal degeneration and hyperphosphorylated Tau. Magnolol is an active component isolated from Magnolia officinalis with potential neuroprotection activity. However, the function and mechanism of magnolol in AD progression is largely uncertain. In present study, the biomarkers related to AD and magnolol were predicted by bioinformatics analyses. The key biomarker levels were predicted by GSE5281 and GSE36980 using AlzData. Cell viability was detected by CCK-8 assay. mRNA and protein levels were examined by qRT-PCR and western blotting assays. Cell apoptosis was investigated by caspase-3 activity and flow cytometry analyses. The cAMP/PKA/CREB signaling was evaluated by ELISA and western blotting analyses. The results showed that CHRM1 was a key biomarker for magnolol against AD progression. Magnolol attenuated Aß-induced viability inhibition, Tau hyperphosphorylation and apoptosis in SH-SY5Y cells by upregulating CHRM1. In addition, the cAMP signaling might be a potential pathway of CHRM1 in AD. Magnolol contributed to activation of the cAMP/PKA/CREB pathway through enhancing CHRM1 level. Inactivation of the cAMP/PKA/CREB signaling reversed the suppressive effect of magnolol on Tau hyperphosphorylation and apoptosis in Aß-treated SH-SY5Y cells. As a conclusion, magnolol mitigated Aß-induced Tau hyperphosphorylation and neuron apoptosis by upregulating CHRM1 and activating the cAMP/PKA/CREB pathway.

Multimodal Predictions of Super-Refractory Status Epilepticus and Outcome in Status Epilepticus Due to Acute Encephalitis.

Objective: Status epilepticus (SE) is one of the most critical symptoms of encephalitis. Studies on early predictions of progression to super-refractory status epilepticus (SRSE) and poor outcome in SE due to acute encephalitis are scarce. We aimed to investigate the values of neuroimaging and continuous electroencephalogram (EEG) in the multimodal prediction. Methods: Consecutive patients with convulsive SE due to acute encephalitis were included in this study. Demographics, clinical features, neuro-imaging characteristics, medical interventions, and anti-epileptic treatment responses were collected. All the patients had EEG monitoring for at least 24 h. We determined the early predictors of SRSE and prognostic factors of 3-month outcome using multivariate logistic regression analyses. Results: From March 2008 to February 2018, 570 patients with acute encephalitis were admitted to neurological intensive care unit (N-ICU) of Xijing hospital. Among them, a total of 94 patients with SE were included in this study. The percentage of non-SRSE and SRSE were 76.6 and 23.4%. Cortical or hippocampal abnormality on neuroimaging (p = 0.002, OR 20.55, 95% CI 3.16-133.46) and END-IT score (p < 0.001, OR 4.07, 95% CI 1.91-8.67) were independent predictors of the progression to SRSE. At 3 months after N-ICU discharge, 56 (59.6%) patients attained good outcomes, and 38 (40.4%) patients had poor outcomes. The recurrence of clinical or EEG seizures within 2 h after the infusion rate of a single anesthetic drug >50% proposed maximal dose (p = 0.044, OR 4.52, 95% CI 1.04-19.68), tracheal intubation (p = 0.011, OR 4.99, 95% CI 1.37-11.69) and emergency resuscitation (p = 0.040, OR 9.80, 95% 1.11-86.47) predicted poor functional outcome. Interpretation: Initial neuro-imaging findings assist early identification of the progression to SRSE. Continuous EEG monitoring contributes to outcome prediction in SE due to acute encephalitis.

MiR-134 blockade prevents status epilepticus like-activity and is neuroprotective in cultured hippocampal neurons.

Status epilepticus (SE) is a life-threatening neurological disorder associated with significant morbidity and mortality. MicroRNAs (miRNAs) are small, non-coding RNAs that act post-transcriptionally modulating messenger RNA (mRNA) translation or stability which may have important roles in the pathogenesis of epilepsy. It has been reported that silencing microRNA-134 in vivo has significant neuroprotective and prolonged seizure-suppressive effects. However, the mechanism by which miR-134 inhibition suppressed seizures and whether miR-134 inhibition works in an in vitro model of SE, is unknown. Compared to a complex in vivo system, in vitro models of SE-like electrographic activity can be powerful tools to study this miRNA. Using a cell culture model of low Mg(2+) treatment of rat hippocampal neurons, we found SE-like electrographic activity increased expression of miR-134. Inhibiting expression of miR-134 using an inhibitor lentivirus with two miR-134 binding sites reduced SE-like electrographic activity in the hippocampal neurons and reduced neuronal death. This study provides direct evidence that inhibition of miR-134 can block status epilepticus-like discharges and is neuroprotective in hippocampal neuronal cultures and implies that inhibiting miR-134 may be a potential candidate for the clinical treatment of SE.