Efficacy of anti-inflammatory therapy in a model of acute seizures and in a population of pediatric drug resistant epileptics

Abstract

Targeting pro-inflammatory events to reduce seizures is gaining momentum. Experimentally, antagonism of inflammatory processes and of blood-brain barrier (BBB) damage has been demonstrated to be beneficial in reducing status epilepticus (SE). Clinically, a role of inflammation in the pathophysiology of drug resistant epilepsies is suspected. However, the use anti-inflammatory drug such as glucocorticosteroids (GCs) is limited to selected pediatric epileptic syndromes and spasms. Lack of animal data may be one of the reasons for the limited use of GCs in epilepsy. We evaluated the effect of the CG dexamethasone in reducing the onset and the severity of pilocarpine SE in rats. We assessed BBB integrity by measuring serum S100β and Evans Blue brain extravasation. Electrophysiological monitoring and hematologic measurements (WBCs and IL-1β) were performed. We reviewed the effect of add on dexamethasone treatment on a population of pediatric patients affected by drug resistant epilepsy. We excluded subjects affected by West, Landau-Kleffner or Lennox-Gastaut syndromes and Rasmussen encephalitis, known to respond to GCs or adrenocorticotropic hormone (ACTH). The effect of two additional GCs, methylprednisolone and hydrocortisone, was also reviewed in this population. When dexamethasone treatment preceded exposure to the convulsive agent pilocarpine, the number of rats developing status epilepticus (SE) was reduced. When SE developed, the time-to-onset was significantly delayed compared to pilocarpine alone and mortality associated with pilocarpine-SE was abolished. Dexamethasone significantly protected the BBB from damage. The clinical study included pediatric drug resistant epileptic subjects receiving add on GC treatments. Decreased seizure frequency (≥ 50%) or interruption of status epilepticus was observed in the majority of the subjects, regardless of the underlying pathology. Our experimental results point to a seizure-reducing effect of dexamethasone. The mechanism encompasses improvement of BBB integrity. Our results also suggest that add on GCs could be of efficacy in controlling pediatric drug resistant seizures.

Figure 1. Effects of dexamethasone on pilocarpine-induced SE.

(A) The number of rats experiencing SE was reduced by anti-inflammatory treatments. Data are compared to IL-RA treatments . (B) When SE developed, its onset was delayed (control vs. IL1ra p = 0.03; control vs. Dexa p = 0.04). (C) Twelve hours mortality associated with pilocarpine seizures was decreased by IL-RA and abolished by dexamethasone (p = 0.02). To attest the efficacy of treatment on survival, seizures were not stopped using barbiturates. (D-E) Examples of EEG recordings show that SE in treated animals was of lesser intensity compared to pilocarpine alone (see also Figure S2). Data are relative to n = 15 rats / group. Asterisks: p<0.05, by paired t-test and Fisher test.

Figure 2. Time-joint frequency analysis of EEG recordings.

(A–C) Single asterisk refers to the first seizure episode. The double asterisk shows the maximal electrographic and behavioral seizures observed under any given condition. The actual EEG recordings are also shown. Time-joint frequency plots show a reduction of seizure intensity (frequency and amplitude domains, color coded) in treated animals compared to pilocarpine alone. Data shown refer to 2 hours of EEG recordings. See also Figure S2 for peak area distribution and instantaneous frequency analysis.

Figure 3. Dexamethasone reduces BBB damage in pilocarpine-treated rats.

BBB integrity was assessed by Evans blue (A) and serum S100β (B) measurements. Evans blue is an indicator of paracellular leakage while S100β is a surrogate serum marker of the integrity of the cerebrovascular endothelial interface. Both methods revealed a reduction of pilocarpine-induced BBB damage in dexamethasone-pretreated animals (DEXA-PILO). Similar efficacy was previously reported for IL1-RA (see [29]). The asterisks refers to p<0.05 by paired t-test, n = 5 rats per group.

Figure 4. Hematologic and serologic changes after dexamethasone treatment in rats.

Figure 5. Efficacy of dexamethasone and ACTH in drug resistant pediatric epilepsy.

Data relative to the efficacy of methyl-prednisolone and hydrocortisone are presented in Figure S3. (A) A total of 53 treatments were evaluated. Treatments were administered as described in the Methods and Table S1. Seizures were assessed by behavioral and EEG observations. The values reported refer to decrease in seizure burden compared to baseline EEG seizure quantification. T-test was used to assess significance. (B) Mosaic plot showing the correlation between etiology of epilepsy and likelihood of a response ≥50%. Note that etiology did not always predict response. Noteworthy, dysplasia and other non-encephalopathic diseases responded to the treatments. Cryptogenic seizures were least affected. Bar width is proportional to the number of observations. Colors refer to the response as indicated in the inset.

Figure 6. Radiologic indices of successful treatment with dexamethasone.

The seizure reducing effect of dexamethasone (B) was paralleled by a decrease in FLAIR hyperintensity.