Poster Abstracts

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High mobility group box 1 in the infl ammatory pathogenesis of epilepsy: profi ling circulating levels after experimental and clinical seizuresLauren Walker, Karen Tse, Emanuele Ricci, Thimmasettappa Thippeswamy, Graeme J Sills, Steve H White, Daniel J Antoine, Anthony Marson, Munir Pirmohamed

Abstract Background High mobility group box 1 (HMGB1) is a chromatin binding protein that is passively released by necrotic cells and actively secreted in response to infl ammatory stimuli in a hyperacetylated form. HMGB1 is upregulated in the brain after injury and can regulate localised infl ammatory reactions resulting in seizures. Antiepileptic drugs prevent seizures but have no disease modifying eff ect or infl uence on natural history. Immunomodulatory antiepileptic drugs have huge potential, but can have serious adverse eff ects. Patient stratifi cation is required to maximise the benefi t to risk ratio. We investigated serum and brain concentrations of HMGB1 in rodent models of seizures and epilepsy and in people with refractory epilepsy.

Methods We used three experimental models in this study. Adult male C57BL/6J mice received repeated intraperitoneal injections of kainic acid (KA) until the onset of convulsive status epilepticus; status epilepticus was terminated after 2 h with diazepam, with brain and blood samples obtained at 3, 6, 24, and 72 h and 7 and 14 days thereafter. Tonic seizures were induced in adult male CF1 mice by maximal electroshock (MES) delivered via corneal electrodes, with brain and blood samples obtained at 1, 4, 8, 16, and 24 h after seizure induction. Finally, serial samples were obtained at baseline, 1, 4, 8, and 12 h after an observed seizure from people with drug refractory epilepsy undergoing videoelectroencephalograph telemetry. Total HMGB1 expression was measured by western blot and immunohistochemistry in brain and by ELISA in serum. Expression of individual HMGB1 isoforms was determined by liquid-chromatography tandem mass-spectrometry (LC-MS/MS).

Findings HMGB1 expression was signifi cantly raised in hippocampus and cortex (p<0·001) at 24 h after KA-induced status epilepticus. Increased cytoplasmic HMGB1 staining was observed at 24 h and associated with an increase in the hyperacetylation of HMGB1 in the hippocampus. In the MES model, there was a signifi cant increase in hippocampal (p<0·05) and cortical (p<0·05) HMGB1 expression at 24 h after a single tonic seizure. Serum concentrations of HMGB1 peaked 3 h after KA-induced status epilepticus (baseline mean 5·0 ng/mL [SD 0·6 ] vs 3 h 16·5 [1·5], p<0·0001) and 4 h after MES seizures (baseline 9·2 [2·5] vs 4 h 19·7 [11·3], p<0·05). 14 days after status epilepticus, serum HMGB1 remained signifi cantly raised (18·0 [11·5], p<0·05) and the isoform was hyperacetylated. An analogous rise in the total HMGB1 serum concentration occurred after seizures in patients with epilepsy (baseline 7·1 [1·4] vs 4 h 15·2 [7·0], p<0·05). Mean serum HMGB1 was signifi cantly higher in patients with drug refractory epilepsy than in healthy controls (8·6 ng/mL [3·5] vs 0·7 [0·3], p<0·002) and in patients with epilepsy who had been seizure free for more than 6 months (8·6 [3·5] vs 1·25 [0·71], p<0·0001).

Interpretation These fi ndings suggest that blood and brain HMGB1 concentrations are increased as a result of seizures in both animal models and patients with epilepsy. Increases in HMGB1 might occur as a consequence of seizures. Patients with refractory epilepsy have higher baseline HMGB1, which might predispose them to recurrent seizure activity. The association between HMGB1 and seizures requires further exploration.

Funding UK Medical Research Council, ICON, GlaxoSmithKline, AstraZeneca, the Medical Evaluation Unit.

ContributorsLW wrote the abstract and collected the data. KT assisted with rodent epilepsy experiments. ER performed the animal pathology. TT is the principal investigator for the KA rodent study. SHW is the principal investigator for the maximal electroshock rodent study. LW, GJS, DJA, AM, and MP analysed, reviewed, and interpreted data. All authors reviewed the abstract.

Confl icts of interestWe declare that we have no confl icts of interest.

Published OnlineFebruary 26, 2014

Poster 97

Department of Molecular and Clinical Pharmacology, University of Liverpool, Liverpool, UK (L Walker MBChB, K Tse MSc, G J Sills PhD, D J Antoine PhD, Prof A Marson MBChB, Prof M Pirmohamed MBChB); School of Veterinary Science, University of Liverpool, Liverpool, UK (E Ricci PhD); Department of Biomedical Sciences, College of Veterinary Medicine, Iowa State University, Ames, IA, USA (Prof T Thippeswamy PhD); and Department of Pharmacology and Toxicology, University of Utah, Salt Lake City, UT, USA (Prof S H White PhD)

Correspondence to: Dr Lauren Walker, The Wolfson Centre for Personalised Medicine Waterhouse Block A, 1-5 Brownlow Street University of Liverpool, Liverpool L69 3GL, UKlauren.walker@liv.ac.uk