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| Author(s): |
Jaqueline Cruz Geraldis
Total Authors: 1
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| Document type: | Master's Dissertation |
| Press: | Campinas, SP. |
| Institution: | Universidade Estadual de Campinas (UNICAMP). Faculdade de Ciências Médicas |
| Defense date: | 2020-07-08 |
| Examining board members: |
Íscia Teresinha Lopes Cendes;
Andre Schwambach Vieira;
Esper Abrão Cavalheiro
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| Advisor: | Íscia Teresinha Lopes Cendes; Danyella Barbosa Dogini |
| Abstract | |
Mesial temporal lobe epilepsy (MTLE) with hippocampal sclerosis (HS) is one of the most frequent and severe types of epilepsy. For patients with refractory MTLE, epilepsy surgery may be a useful therapeutic alternative. DNA methylation is one of the several epigenetic mechanisms and may modulate gene expression. Thus, alterations in DNA methylation may be involved in the underlying mechanisms of disease. This work aims to investigate whether there are differences in the whole-genome DNA methylation pattern, also called methylome, in tissue obtained from patients with refractory MTLE who underwent epilepsy surgery. In addition, we want to investigate if the DNA methylation pattern changes with the increased duration of the disease. To achieve these objectives, we performed whole-genome bisulfite sequencing (WGBS), which is currently considered is the gold-standard approach to determine global DNA methylation. We included tissue samples from 11 patients with MTLE+HS, five belonging to patients with less than 20 years of disease duration (Group 1), and six samples from patients with more than 20 years of disease (Group 2). Also, we used four control samples from autopsy material. DNA samples were extracted from the tissue samples (hippocampus and dentate gyrus) using the phenol-chloroform protocol. DNA was converted by sodium bisulfite, and the samples were prepared for whole-genome bisulfite sequencing. Sequencing data were analyzed using bioinformatics and statistics tools. We performed three different comparations: Group 1 versus Controls, Group 2 versus Controls, and finally, Group 1 versus Group 2. Overall, we found a total of 3,772 differentially methylated regions (DMRs). We selected the 20 DMRs displaying the greats biological relevance for each comparison and performed further analysis. First, we searched for candidate genes presenting functions that have been previously associated with epilepsy in each of the 60 DMRs selected, and we found four candidate genes: GRIN2D, FAM168B, GRP88, and PDCD7. Subsequently, we performed intergenic network analysis and identified three additional candidate genes: MAP2K7, KCND3, and RGS11. In conclusion, i) we found significant differences in the genomic methylation pattern of brain tissue from patients with MTLE+HS, compared to controls, ii) we also observed that the methylome pattern changed with an increased duration of the disease, suggesting that the tissue injury in patients with MTLE+HS progresses over the time, iii) we found a total of 3,772 DMRs and observed a predominance of hypermethylation in patients as compared to controls, iv) we identified four candidate genes which mapped within the DMRs identified, and v) in silico gene interaction analysis identified three additional candidate genes. Additionally, we performed a preliminary analysis integrating the methylome and RNA Sequencing data and found correlations that need further confirmation by quantitative RT-PCR performed in specific transcripts. Altogether, we have found evidence that differential DNA methylation is a relevant mechanism that should be further explored in future studies aiming to unravel the underlying mechanisms leading to disease as well as the progression of the tissue lesion in MTLE+HS (AU) | |
| FAPESP's process: | 17/23954-0 - Methylation pattern of Genomic DNA in mesial temporal lobe epilepsy associated with hippocampal sclerosis |
| Grantee: | Jaqueline Cruz Geraldis |
| Support Opportunities: | Scholarships in Brazil - Master |