Abstracts

Rationale: The abundance of peripherally derived immune cells in resected epileptic tissue suggests the immune system as a potential target for anti-epileptogenic therapy. Vascular blood brain barrier (BBB) breakdown and chronic inflammation can lower seizure threshold, and initiate and propagate epileptogenesis.   We aim to modify epileptogenesis by modulating immune cell trafficking through intravenous (I.V.) administration of naked poly (lactic-co-glycolic acid) nanoparticle (PLG) in the intrahippocampal kainic acid (IHKA) model of temporal lobe epilepsy (TLE). We quantified brain infiltrating immune cells using flow cytometry and detected seizures to determine therapeutic efficacy of PLG using continuous video electroencephalogram (vEEG). Methods: Prolonged status epilepticus (SE) was induced  in 5-6 week old C57BL/6J mice by stereotaxic injection of 150nl of kainic acid (1 mg/ml)  into the right CA3 / dorsal hippocampus (coordinates: AP -1.7, ML -1.7, DV -1.8). Some animals were implanted with both CA1 depth electrodes (AP -1.7, ML -1.0, DV 1.3), and cortical electrodes and monitored for spontaneous recurrent seizures. Following IHKA injection, we treated one group with daily PLG for 5 days (IHKA-nano, n=11) and the other group with PBS (IHKA-PBS, n=18). Lymphocytes were isolated from the tissue 7 days (D7) after IHKA and quantified with flow cytometry using a T cell panel including the following markers: CD3, CD4, CD44, gd-TCR, CD69, CD28, CD25, NK1.1, CD11b, and CD8. Continuous (48h/week) vEEG monitoring was used to verify the existence of nonconvulsive seizures (>270mv, >20 spikes/10 seconds) on D7. Results: Flow cytometric analysis of infiltrating lymphocytes and leukocytes in the brain on D7 showed that PLG administration reduced infiltration of all peripheral immune cells in the brain, while maintaining the regulatory T cell count. IHKA-PBS animals exhibited high levels of infiltrating antigen presenting cells, especially macrophages. Conversely, PLG treated spleens exhibited increased sequestration of leukocytes compared to controls, thereby restricting their migration to the periphery and away from the site of inflammation, the brain. By D7, 70% (9/13) of IHKA-PBS developed electrographic seizures while only 33% (2/6) IHKA-nano demonstrated electrographic seizures. Conclusions: The increase in cytotoxic and activated T cells, in addition to gd T cells, natural killer cells, and natural killer T cells in the hippocampus after the IHKA injection demonstrated the heightened inflammatory milieu after SE prior to occurrence of spontaneous recurrent seizures in this model of TLE. Repeated PLG nanoparticle treatments afterIHKA induced-SE decreased immune cell infiltration into the brain and demonstrated a remarkable decrease in electrographic seizures.  Funding: Supported by NIH/NINDS R01 NS073768