Expression of microRNAs miR21, miR146a, and miR155 in tuberous sclerosis complex cortical tubers and their regulation in human astrocytes and SEGA-derived cell cultures.

Tuberous sclerosis complex (TSC) is a genetic disease presenting with multiple neurological symptoms including epilepsy, mental retardation, and autism. Abnormal activation of various inflammatory pathways has been observed in astrocytes in brain lesions associated with TSC. Increasing evidence supports the involvement of microRNAs in the regulation of astrocyte-mediated inflammatory response. To study the role of inflammation-related microRNAs in TSC, we employed real-time PCR and in situ hybridization to characterize the expression of miR21, miR146a, and miR155 in TSC lesions (cortical tubers and subependymal giant cell astrocytomas, SEGAs). We observed an increased expression of miR21, miR146a, and miR155 in TSC tubers compared with control and perituberal brain tissue. Expression was localized in dysmorphic neurons, giant cells, and reactive astrocytes and positively correlated with IL-1ß expression. In addition, cultured human astrocytes and SEGA-derived cell cultures were used to study the regulation of the expression of these miRNAs in response to the proinflammatory cytokine IL-1ß and to evaluate the effects of overexpression or knockdown of miR21, miR146a, and miR155 on inflammatory signaling. IL-1ß stimulation of cultured glial cells strongly induced intracellular miR21, miR146a, and miR155 expression, as well as miR146a extracellular release. IL-1ß signaling was differentially modulated by overexpression of miR155 or miR146a, which resulted in pro- or anti-inflammatory effects, respectively. This study provides supportive evidence that inflammation-related microRNAs play a role in TSC. In particular, miR146a and miR155 appear to be key players in the regulation of astrocyte-mediated inflammatory response, with miR146a as most interesting anti-inflammatory therapeutic candidate.

Differential expression and clinical significance of three inflammation-related microRNAs in gangliogliomas.

PURPOSE: miR21, miR146, and miR155 represent a trio of microRNAs which has been shown to play a key role in the regulation of immune and inflammatory responses. In the present study, we investigated the differential expression and clinical significance of these three miRNAs in glioneuronal tumors (gangliogliomas, GGs) which are characterized by prominent activation of the innate immune response. METHODS: The expression levels of miR21, miR146, and miR155 were evaluated using Taqman PCR in 34 GGs, including 15 cases with sufficient amount of perilesional cortex. Their expression was correlated with the tumor features and the clinical history of epilepsy. In addition, in situ hybridization was used to evaluate their cellular distribution in both tumor and peritumoral cortex. RESULTS: Increased expression of miR146a was observed in both tumor and peritumoral cortex compared to control samples. miR146a was detected in both neuronal and astroglial cells. Tumor and peritumoral miR146a expression was negatively correlated with frequency of seizures and the density of activated microglial cells. Neuronal and astroglial expression was observed for both miR21 and miR155 with increased expression of miR21 within the tumor and miR155 in the peritumoral region. Negative correlations were observed between the miRNA levels and the expression of putative targets within the astroglial component of the tumor. CONCLUSION: We report a differential regulation of three miRNAs, known to be related to inflammation, in both tumor and peritumoral cortex of patients with GG. Moreover, our findings suggest a functional relationship between miR146a expression and epilepsy, either directly in epileptogenesis or as modulation of seizure activity.

Expression of neurodegenerative disease-related proteins and caspase-3 in glioneuronal tumours.

AIMS: Recent evidence supports the activation of mechanisms underlying cellular ageing and neurodegeneration in developmental lesions associated with epilepsy. The present study examined the ongoing cell injury and vulnerability to neuronal degeneration in glioneuronal tumours (GNT). METHODS: We evaluated a series of GNT (n = 31 gangliogliomas, GG and n = 30 dysembryoplastic neuroepithelial tumours, DNT). Sections were processed for immunohistochemistry using markers for the evaluation of caspase-3 and neurodegeneration-related proteins/pathways and their expression was correlated with the tumour features and the clinical history of epilepsy. RESULTS: Both GG and DNT specimens contained caspase-3-positive cells. In GG, expression of activated caspase-3 was negatively correlated the with the BRAF V600E mutation status. We also observed an abnormal expression of death receptor-6 and ß-amyloid precursor protein (APP). Moreover, dysplastic neurones expressed p62, phosphorylated (p)TDP43 and pTau. Double labelling experiments showed colocalization of phosphorylated S6 (marker of mammalian target of rapamycin, mTOR, pathway activation) with pTau and p62. In GG, neuronal p62 expression was positively correlated with pS6. The immunoreactivity score (IRS) of caspase-3, APP, DR6, p62 and pTDP43 were found to be significantly higher in GG than in DNT. Expression of APP, DR6, pTau (in GG and DNT) and caspase-3 (in GG) positively correlated with duration of epilepsy. In GG, the expression of neuronal caspase-3, DR6 and glial p62 was associated with a worse postoperative seizure outcome. CONCLUSIONS: Our observations in GNT provide evidence of premature activation of mechanisms of neurodegeneration which are associated with the clinical course of epilepsy in patient with GG.