High-resolution transcriptomics informs glial pathology in human temporal lobe epilepsy

Abstract

The pathophysiology of epilepsy underlies a complex network dysfunction between neurons and glia, the molecular cell type-specific contributions of which remain poorly defined in the human disease. In this study, we validated a method that simultaneously isolates neuronal (NEUN +), astrocyte (PAX6 + NEUN–), and oligodendroglial progenitor (OPC) (OLIG2 + NEUN–) enriched nuclei populations from non-diseased, fresh-frozen human neocortex and then applied it to characterize the distinct transcriptomes of such populations isolated from electrode-mapped temporal lobe epilepsy (TLE) surgical samples. Nuclear RNA-seq confirmed cell type specificity and informed both common and distinct pathways associated with TLE in astrocytes, OPCs, and neurons. Compared to postmortem control, the transcriptome of epilepsy astrocytes showed downregulation of mature astrocyte functions and upregulation of development-related genes. To gain further insight into glial heterogeneity in TLE, we performed single cell transcriptomics (scRNA-seq) on four additional human TLE samples. Analysis of the integrated TLE dataset uncovered a prominent subpopulation of glia that express a hybrid signature of both reactive astrocyte and OPC markers, including many cells with a mixed GFAP + OLIG2 + phenotype. A further integrated analysis of this TLE scRNA-seq dataset and a previously published normal human temporal lobe scRNA-seq dataset confirmed the unique presence of hybrid glia only in TLE. Pseudotime analysis revealed cell transition trajectories stemming from this hybrid population towards both OPCs and reactive astrocytes. Immunofluorescence studies in human TLE samples confirmed the rare presence of GFAP + OLIG2 + glia, including some cells with proliferative activity, and functional analysis of cells isolated directly from these samples disclosed abnormal neurosphere formation in vitro. Overall, cell type-specific isolation of glia from surgical epilepsy samples combined with transcriptomic analyses uncovered abnormal glial subpopulations with de-differentiated phenotype, motivating further studies into the dysfunctional role of reactive glia in temporal lobe epilepsy.

Aerobic exercise alters DNA hydroxymethylation levels in an experimental rodent model of temporal lobe epilepsy

“…The hippocampus is a well-studied brain region, but as we discover more about cell type diversity, hippocampus heterogeneity becomes apparent [34] . This fact is even more relevant in the context of the epileptic brain and its accompanying neuronal loss [35] , [36] . We therefore sought to measure DNA 5-hmC specifically in neurons and astrocytes.…”

Identification of hub genes significantly linked to temporal lobe epilepsy and apoptosis via bioinformatics analysis

“…GSE186334 includes 24 epilepsy patient cortical samples and 12 healthy control cortical samples ( Gomes-Duarte et al, 2022 ). GSE140393 validation set consists of 12 epilepsy patient cortical samples and 9 healthy control cortical samples ( Pai et al, 2022 ). Additionally, we downloaded a set of 580 genes associated with apoptosis from PMID36341760 ( Zou et al, 2022 ) ( Supplementary Table 2 ).…”

The Matrix Receptor CD44 Is Present in Astrocytes Throughout the Human CNS and Accumulates in Hypoxia and Seizures

“…Because of the reports of CD44 increase in epilepsy [14,15] and in tuberous sclerosis, which is often accompanied by seizures [13,16], we examined a group of neurosurgical specimens removed from patients with temporal lobe epilepsy, both the hippocampi and the adjacent temporal isocortex for CD44+ astrocytes. The hippocampi showed an increase in CD44 immunostaining in all sectors (Figure 12, and compare with Figure 4).…”

“…Seizures provoke an increase in CD44 in astrocytes at least in the isocortex and hippocampus, the two areas se examined. Furthermore, transcriptomics of the reactive astrocytosis accompanying temporal lobe epilepsy show upregulated CD44 [15]. The mechanisms underlying this are not known.…”