Transcriptomic Profiling of Medial Temporal Lobe Epilepsy

Abstract

Epilepsy is one of the most prevalent neurological disorders affecting ~1% of the population. Medial temporal lobe epilepsy (MTLE) is the most frequent type of epilepsy observed in adults who do not respond to pharmacological treatment. The reason for intractability in these patients has not been systematically studied. Further, no markers are available that can predict the subset of patients who will not respond to pharmacotherapy. To identify potential biomarkers of epileptogenicity, we compared the mRNA profiles of surgically resected tissue from seizure zones with non-seizure zones from cases of intractable MTLE. We identified 413 genes that exhibited ≥2-fold change that were statistically significant across these two groups. Several of these differentially expressed genes have not been previously described in the context of MTLE including claudin 11 (CLDN11) and bone morphogenetic protein receptor, type IB (BMPR1B). In addition, we found significant downregulation of a subset of gamma-aminobutyric acid (GABA) associated genes. We also identified molecules such as BACH2 and ADAMTS15, which are already known to be associated with epilepsy. We validated one upregulated molecule, serine/threonine kinase 31 (STK31) and one downregulated molecule, SMARCA4, by immunohistochemical labeling of tissue sections. These molecules need to be further confirmed in large-scale studies to determine their potential use as diagnostic as well as prognostic markers in intractable MTLE.

Keywords: DNA microarrays; GABA receptor; GeneSpring; Medial temporal sclerosis; Temporal lobe epilepsy; Transcriptome profile.

Figure 1. Workflow of transcriptomic studies for MTLE

A two color DNA microarray analysis based approach employed for the transcriptome profiling of spiking (seizure) against non-spiking (non-seizure) zones of MTLE. An unsupervised hierarchical clustering was performed on differentially expressed molecules and a selected set of novel molecules were validated by IHC.

Figure 2. Heat map of differentially expressed genes in MTLE

A heat map was generated by performing unsupervised hierarchical clustering of 413 differentially expressed genes with a p value cut-off of <0.05 and a fold value cut-off of ≥2. Euclidean distance and centroid linkage were used for generating the hierarchical cluster (A). A heat map of a selected set of differentially regulated genes is shown in B.

Figure 3. A statistically significant network identified by pathway analysis

The network was overlaid with GABAergic pathway and the color code of molecules represents the expression levels wherein red shows upregulated and green indicates downregulation.

Figure 4. Overexpression of STK31 in MTLE

Pyramidal neurons of temporal cortex (B) (Obj x 20) and Ammon’s horn (C) (Obj x 10) reveals strong, cytoplasmic and nuclear labeling from a case of hippocampal sclerosis in contrast to lack of expression in normal control hippocampus (A) (Obj x 10). Note the dendritic labeling in pyramidal neurons and axonal labeling in the alveus (C).

Figure 5. Downregualtion of SMARCA4 in MTLE

A–B: Control hippocampus showing diffuse labeling of dentate gyrus granule neurons (A) (Obj x 40), and pyramidal neurons of Ammon’s horn (B) (Obj x 40). C–D: Hippocampus from a case of mesial temporal sclerosis showing absent expression in dentate granule neuron (C) (Obj x 40) is compared to control. Light cytoplasmic labeling of pyramidal neurons and nuclear labeling of scattered glial cells seen in temporal cortex (D) (Obj x 20), markedly downregulated compared to controls.