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The histopathological neurons in the brain tissue of drug-resistant epilepsy exhibit aberrant cytoarchitecture and imbalanced synaptic circuit function. However, the gene expression changes of these neurons remain unknown, making it difficult to determine the diagnosis or to dissect the mechanism of drug-resistant epilepsy. By integrating whole-cell patch clamp recording and single-cell RNA-seq approaches, we identified a transcriptionally distinct subset of cortical pyramidal neurons. These neurons highly expressed genes CDKN1A (P21), CCL2, and NFKBIA, which associate with mTOR pathway, inflammatory response, and cellular senescence. We confirmed the expression of senescent marker genes in a subpopulation of cortical pyramidal neurons with enlarged soma size in the brain tissue of drug-resistant epilepsy. We further revealed the expression of senescent cell markers P21, P53, COX2, γ-H2AX, and β-Gal, and reduction of nuclear integrity marker Lamin B1 in histopathological neurons in the brain tissue of patients with drug-resistant epilepsy with different pathologies, but not in control brain tissue with no history of epilepsy. Additionally, chronic, but not acute, epileptic seizures induced senescent marker expression in cortical neurons in mouse models of drug-resistant epilepsy. These results provide important molecular markers for histopathological neurons and what we believe to be new insights into the pathophysiological mechanisms of drug-resistant epilepsy.
Introduction
Patients with medically intractable epilepsy respond poorly to currently available antiepileptic drugs (1). Histopathological examinations of brain tissue surgically removed from the epileptic focus revealed various anatomical signatures of neuropathologies (2, 3). For example, dysmorphic neurons in the gray matter and balloon cells in the white matter were pathological characteristics in patients with drug-resistant epilepsy with focal cortical dysplasia (FCD) (4, 5), while neuronal loss and hippocampal sclerosis were characteristic in the epileptic focus of patients with drug-resistant epilepsy with temporal lobe epilepsy (TLE) (2, 3, 6). Understanding the gene expression changes in these cortical pyramidal neurons will be important both for the diagnosis and dissection of the mechanism of seizure genesis in these patients (2, 7). Bulk gene expression studies of the brain tissue in the epileptic focus of drug-resistant epilepsy using microarray or RNA -seg showed upregulation of genes associated with neuroinflammation, restructuring of neuronal networks (8-11), and activation of the mTOR pathway (12). However, representing a rare cell population within the cortex, the histopathological neurons are difficult to...