Authors :
Presenting Author: Samantha Reed, PhD – Emory University
Yilin Wang, MS – Emory University
Yangping Li, PhD – Emory University
Guoku Hu, PhD – University of Nebraska Medical Center
Bing Yao, PhD – Emory University
Jingjing Yang, PhD – Emory University
Andrew Escayg, PhD – Emory University
Rationale:
Mesial temporal lobe epilepsy (MTLE) a common form of treatment-resistant epilepsy that is often associated with a preceding neurological injury. Emerging evidence suggests that the miRNA content of brain derived EVs (BDEVs) is altered in mouse models of MTLE. However, the functional effects of these alterations is unknown and thus, it is not clear if BDEVs play a role in epileptogenesis. To address this, we examined the functional properties of BDEVs following status epilepticus (SE) in the pilocarpine mouse model of MTLE.
Methods:
Mice were subjected to pilocarpine-induced SE and hippocampi were collected 24 hours post-SE for BDEV isolation. We then treated N2a neuron-like and BV2 microglia-like cells with BDEVs isolated from control mice (CON-BDEVs), BDEVs from mice 24 hours after SE (SE-BDEVs) or vehicle. We then conducted transcriptomics on treated cells to determine how BDEVs altered gene expression. In order to determine if alterations in the miRNA content of the BDEVs could cause any observed changes, we also conducted miRNA-sequencing on BDEVs 24 hours and 10-days post SE.
Results:
When N2a cells were treated with CON-BDEVs, we observed altered expression of genes involved in processes such as neurotransmitter signaling and amino acid regulation of mTORC1 and TOR signaling. However, N2a cells treated with SE-BDEVs did not show altered expression of these genes but instead showed altered expression of genes involved in TGF-β signaling. In BV2 cells, both CON- and SE-BDEVs altered the expression of genes related to immune processes, but SE-BDEVs showed an increased ability to alter expression of genes involved in inflammatory cytokine production. When the miRNA expression was compared, we identified 8 differentially expressed (DE) miRNAs in SE-BDEVs. Subsequent target analysis revealed that many of the DE genes that were observed in BV2 and N2a cells were potential direct targets of these DE miRNAs. We also examined miRNA expression in BDEVs collected 10 days after SE to determine if any miRNAs showed sustained changes. We identified 7 DE miRNAs, two of which were also differentially expressed 24 hours post SE.
Conclusions:
SE-BDEVs showed dysregulation in their ability to regulate gene expression in both N2a and BV2 cells. Notably, the increased regulation of genes related to the production of cytokines by SE-BDEVs suggests they have pro-inflammatory properties. As increased inflammation is known to the pathogenetic after SE, it is possible that BDEVs contribute to these processes and impact epileptogenesis. Furthermore, we observed changes in the miRNA content in BDEVs suggesting that these changes could underly the dysfunction in BDEVs.Funding:
This project was supported by NIH Training Grant 5T32NS096050-24 and a predoctoral fellowship from the American Epilepsy Society.