Abstracts

Rationale: Tuberous sclerosis complex (TSC) is an autosomal dominant genetic disorder that arises from mutations in the TSC1 or TSC2 genes. Brain lesions (tubers) occur in 90-95% of TSC patients and contribute to a range of neurological disorders, including epilepsy. Up to 90% of TSC patients suffer from epileptic seizures, and many are refractory to pharmacological therapy, thus requiring surgical resection of epileptogenic tubers. The molecular and cellular events contributing to epileptogenic potential in TSC tubers are poorly understood. MicroRNAs are endogenous non-coding nucleotides that provide post-transcriptional regulation of many genes and most cell processes. Aberrant microRNA expression has been implicated in a wide range of diseases. However, the role of microRNAs in the epileptogenesis of TSC tubers has not previously been explored.Methods: Expression of microRNAs was examined in cortical tubers resected for the treatment of intractable epilepsy in TSC patients. Expression levels in tubers were compared to patient-matched adjacent non-tuber tissue that was concurrently resected due to epileptiform EEG activity. Analysis was performed on brain tissue obtained from five patients who underwent surgery for medically-refractory epilepsy at Children's Hospital of Michigan, Detroit (CHM). Agilent Human miRNA V3 microarrays were used, with probes for more than 800 human microRNAs. Stringent statistical filtering identified five microRNAs with a p-value < 0.05, corrected using the Westfall and Young family-wise error rate.Results: Four microRNAs exhibited higher expression in epileptogenic tubers compared with matched non-tuber tissue. Using a consensus of multiple target prediction methods, we investigated putative target genes of the microRNAs and found the over-expressed microRNAs exert combinatorial targeting of genes associated with synaptic signal transmission. The target gene set was enriched with genes known to confer risk of epilepsy. We utilized an in vitro luciferase reporter assay to confirm that miR-34a directly targets reelin (RELN), a gene where mutation and repression are associated with epilepsy and defects of cortical lamination.Conclusions: We report aberrant microRNA expression in tubers resected from five TSC patients to treat intractable seizures. Expression profiles compared between subject-matched pairs of epileptogenic tubers and adjacent non-tuber tissue revealed coordinate upregulation of microRNAs targeting genes involved in synaptic development and function, as well as epilepsy risk genes. We demonstrated that miR-34a targets RELN, a gene previously implicated in epilepsy. These data suggest that aberrant microRNA expression contributes to the mechanism of epileptogenesis in cortical tubers of TSC.