Leukocyte differential gene expression prognostic value for high versus low seizure frequency in temporal lobe epilepsy

Abstract

Background: This study was performed to test the hypothesis that systemic leukocyte gene expression has prognostic value differentiating low from high seizure frequency refractory temporal lobe epilepsy (TLE).

Methods: A consecutive series of patients with refractory temporal lobe epilepsy was studied. Based on a median baseline seizure frequency of 2.0 seizures per month, low versus high seizure frequency was defined as ≤ 2 seizures/month and > 2 seizures/month, respectively. Systemic leukocyte gene expression was analyzed for prognostic value for TLE seizure frequency. All differentially expressed genes were analyzed, with Ingenuity® Pathway Analysis (IPA®) and Reactome, to identify leukocyte gene expression and biological pathways with prognostic value for seizure frequency.

Results: There were ten males and six females with a mean age of 39.4 years (range: 16 to 62 years, standard error of mean: 3.6 years). There were five patients in the high and eleven patients in the low seizure frequency cohorts, respectively. Based on a threshold of twofold change (p < 0.001, FC > 2.0, FDR < 0.05) and expression within at least two pathways from both Reactome and Ingenuity® Pathway Analysis (IPA®), 13 differentially expressed leukocyte genes were identified which were all over-expressed in the low when compared to the high seizure frequency groups, including NCF2, HMOX1, RHOB, FCGR2A, PRKCD, RAC2, TLR1, CHP1, TNFRSF1A, IFNGR1, LYN, MYD88, and CASP1. Similar analysis identified four differentially expressed genes which were all over-expressed in the high when compared to the low seizure frequency groups, including AK1, F2R, GNB5, and TYMS.

Conclusions: Low and high seizure frequency TLE are predicted by the respective upregulation and downregulation of specific leukocyte genes involved in canonical pathways of neuroinflammation, oxidative stress and lipid peroxidation, GABA (γ-aminobutyric acid) inhibition, and AMPA and NMDA receptor signaling. Furthermore, high seizure frequency-TLE is distinguished prognostically from low seizure frequency-TLE by differentially increased specific leukocyte gene expression involved in GABA inhibition and NMDA receptor signaling. High and low seizure frequency patients appear to represent two mechanistically different forms of temporal lobe epilepsy based on leukocyte gene expression.

Keywords: Gene expression; Leukocyte; Seizure frequency; Temporal lobe epilepsy.

Fig. 1

Multidimensional scaling plot (MDS) generated using edgeR showing segregation of the low and high seizure frequency patients’ leukocyte transcriptional profile. The three lowest and three highest seizure frequency samples were used to generate plot. Numbered sample IDs indicate patients from list in Table 1

Fig. 2

Heatmap generated in edgeR using the 13 most variable genes across samples. Unsupervised clustering showing the grouping of low (#5, #3, #2) and high (#6, #15, #11) seizure frequency patients. X-axis indicates sample IDs from subject list in Table 1. Red indicates a higher level of comparative expression while blue indicates a lower level of expression

Fig. 3

Leukocyte differentially expressed genes (DEGs) involved in neuroinflammation are upregulated in low but downregulated in high temporal lobe epilepsy seizure frequency. Figure prepared with BioRender.com

Fig. 4

Leukocyte differentially expressed genes (DEGs) involved in oxidative stress and lipid peroxidation are upregulated in low but downregulated in high temporal lobe epilepsy seizure frequency. Figure prepared with BioRender.com

Fig. 5

Leukocyte differentially expressed genes (DEGs) involved in glutamate/GABA-mediated excitotoxicity in temporal lobe epileptogenicity which are upregulated in low but downregulated in high temporal lobe epilepsy seizure frequency. Figure prepared with BioRender.com

Fig. 6

Leukocyte differentially expressed genes (DEGs) involved in NMDA facilitation and GABA inhibition which are upregulated in high compared to low temporal lobe epilepsy seizure frequency patients. Figure prepared with BioRender.com

Fig. 7

Upstream regulators predicted to drive the temporal lobe epilepsy response in leukocytes with an overlap p-value < 0.05. Blue cells indicate predicted inhibition of the pathway, orange shows predicted activation. The shades for colored cells represent z-scores values. Color of nodes indicates predicted activation (orange) or deactivation (blue). The NFKB complex is a key node in this network. Figure prepared using Ingenuity Pathway Analysis (IPA)

Fig. 8

This schematic depicts the intricate process of leukocyte infiltration into the brain from the systemic bloodstream. Initially, the leukocytes engage with Vascular Adhesion Molecule 1 (VCAM-1) and Intercellular Adhesion Molecule 1 (ICAM-1), facilitating trans-endothelial diapedesis, a crucial event in the breaching of the blood–brain barrier (BBB). This breach contributes significantly to epilepsy-associated neuroinflammation. This process leads to the activation of pro-inflammatory cytokines, instigating a feed-forward cascade that further stimulates microglial cells. This exacerbated neuroinflammatory response augments epileptogenicity in patients with Temporal Lobe Epilepsy (TLE), implying a complex interplay between the immune response and the neurological disorder. Created with BioRender.com