Mycophenolate mofetil prevents the delayed T cell response after pilocarpine-induced status epilepticus in mice

Abstract

Growing clinical and laboratory evidence corroborates a role for the immune system in the pathophysiology of epilepsy. In order to delineate the immune response following pilocarpine-induced status epilepticus (SE) in the mouse, we monitored the kinetics of leukocyte presence in the hippocampus over the period of four weeks. SE was induced following a ramping protocol of pilocarpine injection into 4–5 weeks old C57BL/6 mice. Brains were removed at days 1–4, 14 or 28 after SE, and the hippocampi were analyzed via flow cytometry, via quantitative reverse transcriptase PCR (qRT-PCR) and via immunohistochemistry. Epileptogenesis was confirmed by Timm staining of mossy fiber sprouting in the inner molecular layer of the dentate gyrus. The flow cytometry data revealed a biphasic immune response following pilocarpine-induced SE with a transient increase in activated CD11b+ and F4/80+ macrophages within the first four days replaced by an increase in CD3+ T-lymphocytes around day 28. This delayed T cell response was confirmed via qRT-PCR and via immunohistochemistry. In addition, qRT-PCR data could show that the delayed T cell response was associated with an increased CD8/CD4 ratio indicating a cytotoxic T cell response after SE. Intriguingly, early intervention with mycophenolate mofetil administration on days 0–3 after SE prevented this delayed T cell response. These results show an orchestrated immunological sequela and provide evidence that the delayed T cell response is sensitive to early immunomodulatory intervention.

The evolution of the pilocarpine animal model of status epilepticus

“…As animals differ in their tolerance to pilocarpine, this method ensures that animals are not unnecessarily given high doses and improves the induction probability. These results were also observed in several other studies using both pilocarpine and LiCl-pilocarpine models ( Brandt et al., 2010 ; Leung et al., 2015 ; Neumann et al., 2017 ; Wu and Wang, 2018 ).…”

“…This however was not fruitful in all animal models. For example, studies have administered single injections of 100 mg/kg of pilocarpine intraperitoneally to induce SE in mice, with the total dosage reaching up to 800 mg/kg ( Neumann et al., 2017 ). Even with such a high overall dose, only 54% of the animals survived the procedure and developed SE.…”

Reporting animal research: Explanation and elaboration for the ARRIVE guidelines 2.0

“…. due to overt behavioral seizure activity the experimenter could not be blinded to whether the animal was injected with pilocarpine or with saline' [86].…”

Crosstalk between peripheral and the brain-resident immune components in epilepsy

“…A recent experiment showed a biphasic increase of CD45 + immune cells in the hippocampus was implicated in innate macrophages during the first four days as well as CD3 + T-lymphocytes at 28 days following pilocarpine-induced SE. Correspondingly, one study has suggested pilocarpineinduced SE of C57BL/6 mice leads to either the infiltration of activated macrophages into the hippocampus or activation of resident glial cells, which are predicted to induce a proinflammatory immune response that may attract more immune cells and allow T-lymphocytes to enter the CNS by affecting the permeability of the BBB [52]. Infiltrated T-cell then an attack on distinct hippocampal neurons sufficient to make a contribution to cellular and structural dynamics that displayed signs of limbic encephalitis that induced TLEhippocampal sclerosis (HS) [53].…”