This study aimed to delineate the mechanistic target of the rapamycin (mTOR) pathway in the brain tissue of patients with Rasmussen encephalitis (RE) compared to individuals without epilepsy and those with focal cortical dysplasia (FCD) to identify unique pathogenic mechanisms and potential therapeutic targets. Experimental analysis was conducted using RE, control and FCD tissue samples obtained through surgical resection. Western blotting was performed to quantify the expression of established markers of mTOR upstream or downstream signaling. Moreover, immunohistochemistry (IHC) and immunofluorescence (IF) were used to assess cortical and white matter abnormalities and the cell-specific expression of distinct biomarkers. Samples from patients with FCD were utilized as positive controls. We found significantly increased levels of phospho-S6 (Ser240/244), phospho-AKT (Ser473), phospho-p44/42 MAPK (ERK1/2) and phospho-Stat3 (Tyr705) in RE samples compared to those in controls, consistent with the activation of both mTOR complex 1 (mTORC1) and mTORC2. Based on the results of the IHC and IF analyses, we observed strong expression of p-S6 and p-AKT in ectopic neurons and giant neurons. Additionally, we noted expression in perivascular microglia, astrocytes, and microglial nodules. p-MAPK was primarily expressed in astrocytes and blood vessels but was occasionally expressed in neurons
p-MAPK was not coexpressed in microglia. Phospho-ULK1 (Ser757) was expressed in apoptotic neurons, while beclin-1 was predominantly present in microglial nodules and atypical neurons, with no expression in astrocytes. P-Stat3 exhibited positive nuclear expression, while cytoplasmic positivity was observed in cortical cells with a morphology resembling that of astrocytes. The expression level of p-MAPK was significantly correlated with the progression of RE. Our experimental results demonstrate aberrant activation of mTORC1 and mTORC2 in RE patients. These findings offer novel insights into the pathogenic mechanisms of RE and might reveal new therapeutic targets for drug intervention in the treatment of RE.