Endothelial cell-specific TGM2 deficiency promotes progression of temporal lobe epilepsy by disrupting S1P metabolism and intercellular communication.

Abstract

Background: Temporal lobe epilepsy (TLE) is a common and medically refractory neurological disorder with a complex pathogenesis. Blood-brain barrier (BBB) dysfunction is a key pathogenic factor. As a multifunctional protein, TGM2 has been shown to participate in vascular homeostasis, metabolic pathway regulation, and immune-inflammatory responses. This study aimed to systematically investigate the expression pattern, core functions, and molecular regulatory mechanisms of TGM2 in TLE, and to clarify its clinical value.

Methods: Temporal cortex tissue and plasma samples from drug-resistant TLE patients and control subjects were collected, alongside kainic acid (KA)-induced epileptic mouse models. Single-cell RNA sequencing was adopted to identify TGM2's cell-specific expression pattern, and qPCR, Western blot and ELISA validated its expression levels. With endothelial cell-specific TGM2 conditional knockout mice, in vivo experiments assessed seizure severity and BBB permeability through behavioral observation, intracranial local field potential (LFP) recording and Evans blue (EB) assay. In vitro siRNA-mediated TGM2 knockdown in brain microvascular endothelial cells verified endothelial functional changes. Metabolomics, RNA-seq, nucleocytoplasmic separation, immunofluorescence and molecular docking were used to clarify regulatory mechanisms, and intercellular communication networks were constructed to analyze ligand-receptor interactions.

Results: Results indicated that endothelial TGM2 was significantly downregulated in TLE patients and epileptic mice. Endothelial TGM2 deficiency aggravated recurrent seizures, increased BBB permeability, impaired endothelial function, and reduced the expression of tight junction protein CLDN5. Mechanistically, TGM2 positively regulated sphingosine-1-phosphate (S1P) synthesis. Reduced S1P production abolished the inhibition of RelA nuclear translocation, thereby activating downstream inflammatory genes and triggering endothelial inflammation. Molecular docking confirmed the direct binding between S1P and RelA. Furthermore, TGM2 deficiency remodels endothelial-neuron-microglia communication and disrupts neural circuit homeostasis.

Conclusions: This study establishes a link between TGM2 and BBB function, metabolic pathways, and intercellular communication in TLE, providing an experimental basis for developing TLE therapeutic strategies targeting this pathway.