Inflammatory reactions in human medial temporal lobe epilepsy with hippocampal sclerosis

Abstract

Many experimental studies suggest that NFκB, a transcription factor involved in acute inflammation, and cytokines participate in neuronal excitability and/or glial scar formation in epilepsy. In this report, we looked for the expression of NFκB in hippocampi surgically removed in patients with medial temporal lobe epilepsy (MTLE) and hippocampal sclerosis (HS) who had an history of febrile convulsions. We analyzed 18 hippocampi from epileptic patients with MTLE and HS, and we used as control specimens three hippocampi from non-epileptic patients and four hippocampi from patients with cryptogenic MTLE without HS. We used antibodies raised against the NFκB-p65 subunit and we identified glial cells with specific antibodies. Hippocampi from patients with MTLE and HS displayed severe neuronal loss surrounded by gliosis in CA1 area and more or less in CA3/CA4 areas. Double immunolabeling showed that reactive astrocytes of lesioned areas over-expressed NFκB-p65 (significantly when compared to control specimens). Moreover, some surviving pyramidal neurons in these areas and numerous dentate granule cells were strongly positive for NFκB-p65 in cytoplasm and nucleus, whereas control hippocampi showed a faint basal cytoplasmic staining in neurons. These results suggest that in epileptic hippocampi with typical sclerosis, inflammatory processes are chronically active or transiently re-induced by recurrent seizures. Whether NFκB over-expression reflects protective or deleterious mechanisms in the epileptic focus remains to be elucidated.

Introduction

Epilepsy is a frequent disease with a prevalence in population-based study estimated at 0.7% (Picot et al., submitted). Partial epilepsy and particularly medial temporal lobe epilepsy is the most widespread and constitutes a large part of intractable epilepsies that can benefit from surgery with anterior temporal lobectomy and amygdalo-hippocampectomy. The most common etiological lesion of medial temporal lobe epilepsy is hippocampal sclerosis, characterized by neuronal loss in CA1 area and gliosis [6], [27]. The analysis of human epileptic hippocampi removed for epilepsy surgery confirmed the existence of CA1 lesions, surrounded by astrogliosis [3] and accounted for the dispersion of dentate granule cells [12]. According to the ‘epileptogenic glial scar’ hypothesis, reactive astrocytes in lesion areas are supposed to release trophic factors that support axonal sprouting and neosynaptogenesis, probably involved in the further development of a chronic hyperexcitability [25].

Currently, the role of inflammatory molecules, like cytokines, in neuronal excitability and in glial scar formation is suggested by various studies obtained in human and in experimental epilepsies [15]. A correlation between the frequency of the homozygote form of interleukine-1beta (IL-1β) and the susceptibility to develop hippocampal sclerosis after febrile seizures was demonstrated in Japanese patients [16], but not in Caucasian subjects [11]. An elevated level of interleukine-6 (IL-6) was reported in the cerebro-spinal fluid of epileptic patients [23]. In experimental models, kindling seizures were shown to transiently up-regulate the transcription of IL-1β, IL-1Receptor1, Tumor Necrosis Factor alpha (TNFα) and TGF-β1 [24], while, after lesional experimental status epilepticus, an increase of TNFα and IL-6 release was reported [21], [10]. Moreover, a direct pro-convulsant effect of IL-1β was demonstrated [31], [32]. These pro-inflammatory cytokines may act as signals, promptly released between the ictal event and glial reaction. It is now well admitted that the transcription factor Nuclear Factor kappa B (NFκB) plays a crucial role in many acute inflammatory reactions. NFκB is present in cytosol under an inactive form, due to IκB, an inhibitory subunit linked to the active subunits p50 and p65, that are associated in homo- or hetero-dimers. In response to various factors, like oxidative stress, kinase activation or stimulation of receptors belonging to the ‘death receptor family’, IκB is phosphorylated, allowing the release of p50 and p65. Both active monomers enter the nucleus, where they bind to specific consensus sequences. These subunits activate the transcription of numerous genes encoding cytokines, complement proteins, major histocompatibility complex glycoproteins, neurotrophines, cell adhesion molecules and various enzymes [4], [22]. In experimental limbic status epilepticus in rat, an over-expression of NFκB was reported in neurons and in reactive astrocytes, concomitant with microglial activation [18]. These reactions were still obvious 2 weeks after status epilepticus, suggesting that inflammatory processes participate in post-ictal events involved in secondary epileptogenesis. However, whether such inflammatory reactions are present in human chronic epileptic focus remains to be established.

Therefore, in this study, we looked for the expression of NFκB in the chronic epileptic focus, surgically removed in patients presenting medial temporal lobe epilepsy with hippocampal sclerosis.