Sex, Epilepsy, and Epigenetics

     When referring to epilepsy, we are referring to a heterogeneous group of disorders, including both those of genetic origin and those acquired. They are associated with a number of pathogenic mechanisms, seizure manifestations, comorbidity profiles and therapeutic responses. It has been observed through clinical and translation research that many of these epileptic features are closely affect by sex differences, especially hormones. That is, there is emerging evidence that common pathological features in epilepsy syndrome are linked with sex differences with males exhibiting a greater incidence than females. Through this paper, Qureshi et al, explain the primary epigenetic mechanisms and how these are now being used to integrate hormonal and genetic influences at molecular, cellular and network levels in epileptic disorders and the process of epilptogenesis (described in previous post).

“The foremost epigenetic mechanisms include DNA methylation (and hydroxymethylation), histone protein post-translational modifications (PTMs) and higher-order chromatin remodeling, and noncoding RNA (ncRNA) regulation. These multilayered processes are highly interconnected and exert their regulatory effects through coordinate actions.”

     Epigenetic mechanisms are mediator of the brains form and function and it believed to be a source in the promotion of dimorphism in the brain and body. They help establish and maintain sex differences in gene expression, for example the X inactivation-specific transcript (XIST) and genomic imprinting (more details related to its function in paper). Various epigenetic factors are expressed in sex-specific patterns in the breain known to be dimorphic, however these factors and there mechanisms are sensitive to sex steroid hormone pathways and exposure. Sex modulation has also been observed in autosomally encoded factors. “These observations suggest that epigenetic regulators in brain are deployed in a sex-specific manner, consistent with other evidence from expression quantitative trait loci analyses revealing sex-biased gene regulatory architectures in human brain.”

     Qureshi et al. describe various non-mutually exclusive paradigms relating epigenetic factors and neurological diseases including: : mutations in genes encoding epigenetic factors that cause disease, genetic variation in genes encoding epigenetic factors modifying disease risk, and the expression and function of epigenetic factors targeting disease-associated genomic loci, gene products, and cellular pathways. The first has proven to be true linking DNA methylation and histone modifying enzymes with the onset of the disease. Emerging data on the second paradigm has demonstrated variability in the vulnerability to epileptic disorders, like for example polymorphisms of the bromodomain-containing protein 2 (BRD2) gene which confers susceptibility to common forms of myoclonic epilepsy. This has also been studied in mice, demonstrating a sex-specific decrease in seizure thresholds. Lastly, evidence indicating that an increase in DNA methylation in the hippocampus are associated with epileptogenesis and that adenosine exhibit inhibition of DNA methylation (described in previous post), supports the third paradigm.

     The observations detailed throughout the paper suggest that the epigenetic factors and mechanisms in males and females underlie sex differences associated with risk, onset, and progression of epileptic disorders. However, seeing as many bodily functions work in coaction it is important to study other pathways and how these interact with both epigenetic factors and epilepsy. In this way, it might be possible to generate a novel therapeutic drug to treat these and other diseases more efficiently.

Reference: Qureshi, I.A., Mehler, M., (July 4, 2014). Sex, Epilepsy and Epigenetics. Elsevier, Retrieved from http://ac.els-cdn.com/S0969996114001831/1-s2.0-S0969996114001831-main.pdf?_tid=7ae5413c-bdc7-11e6-a331-00000aacb361&acdnat=1481257694_2ef9836c0fdfbf1053537c6b34baff56

New Differentially Expressed Genes and Differential DNA Methylation Underlying Refractory Epilepsy

Over 65 million people are affected with epilepsy worldwide. A variety of genetic and environmental factors have been associated with epilepsy and seizures. In the case of epilepsy, DNA methylation has been deemed one of the principal epigenetic mechanisms leading to epilepsy. These affect genomic reprograming, tissue-specific gene expression and global gene silencing without affecting the sequence. 

It has been seen that many of these DNA methylations are present at the promoters of genes, resulting in a decrease of gene expression. Besides promotors, an inverse relationship between gene expression and DNA methylation has been seen in exons and introns. Not only has it been discovered that selective changes in genome-wide DNA methylation and increased DNA methyltransferase are associated with temporal lobe epilepsy (TLE), but also that ketogenic diets could attenuate seizure progression though DNA methylation. 

The work of Xi Liu et al. was focused on analyzing the pattern of genome wide DNA methylation and gene expression using methylated DNA immunoprecipitation linked with sequencing. As a result, they were able to distinguish a new pattern of DNA methylation associated with refractory epilepsy patients. They also found that there was no significant difference between epileptic samples and controls in genome, CpG, CHG, and CHH coverage distribution and that differentially methylated regions were discovered on all genes except for the male Y chromosome. Generally, no significant relationship in modulation was found between DNA methylation and gene expression. Xi Liu et al, also working towards generating DNA methylation and gene expression profiles to prove the relationship between DNA methylation and gene expression via distribution of hyper-, hypo- and unmethylated gene expression levels in different elements.

     The importance of this paper is that is the first genome wide report on DNA methylation and gene expression in refractory epilepsy patients. Over 62 differentially expressed genes were found to be correlated with epilepsy and seizures. However, the similarity in results between epileptic samples and the controls indicated no significant difference in global DNA methylation and gene expression. That is, the change in DNA methylation in the study is no corresponded with alterations in gene expression.

Reference: Liu, X., Ou, S., Xu, T., Liu, S. Yuan, J., Huang, Y., Qin, L., Yang, H., Chen, L., Tan, X., Chen, Y., (November 26,2016), New Differentially Expressed Genes and Differential DNA Methylation Underlying Refractory Epilepsy. Oncotarget. Retrieved from http://www.impactjournals.com/oncotarget/index.php?journal=oncotarget&page=article&op=view&path%5B%5D=13642

“MicroRNA epigenetic signatures” (Piletic & Kunej, 2016) in epilepsy

        Post-translational modifications in epigenetics, such as methylation and acetylation, have always been discussed, but on the other hand, the active role of microRNAs (miRNAs) has not been explained in such detail. miRNAs are short non-coding RNAs that participate as regulators of gene expression. Klara Piletic and Tanja Kunej describe 63 miRNA genes that are epigenetically regulated in association with 21 diseases, such as cardiovascular disease, rheumatoid arthritis, autism, gastric, cervical, ovarian, prostate and bladder cancer. Temporal lope epilepsy (TLE) is included in these diseases.

          MiRNAs are approximately 22 nucleotides long, and there are currently more than 460 human miRNAs known.  They are transcribed by RNA polymerase II (Pol II). miRNAs target messenger RNAs (mRNAS) and degrade them. miRNA is non coding RNA, that is transcribed and is exported from the nucleus. It is then cut by a Dicer protein down to 22 nucleotides and then loaded into the Argonaut complex, where it can inhibit gene translation and act to degrade other RNAs.

http://www.nature.com/leu/journal/v26/n3/images/leu2011344f1.jpg

In temporal lobe epilepsy, the following miRNAs genes are epigenetically regulated: miR-27a, miR-193a-5p, miR-486, miR-618, miR-133a-1, miR-151, miR-191, miR-375, miR-411, miR-342, miR-34a, miR-627 and miR-576. Patients that suffer from TLE, “can display hippocampal sclerosis, which is a histopathologic abnormality including segmental neuron loss as well as other changes” (2016). DNA hyper methylation plays a major role in this condition, but so does miRNA, that “plays a role in the pathophysiology of TLE” (2016).  

           

References:

Piletic, K. & Kunej, T. (2016). MicroRNA epigenetic signatures in human disease. Archives of Toxicology, 90(2405-2419). 

Epileptogenesis: Can the science of epigenetics give us answers?

Due to the complex progression of epilepsyand the multiple inherited and acquired factors that have influence on the onset and progression of this disease, developing cures for this disroder has proved to be challenging, with current treatment have focused on controlling seizure activity. Epigenetic mechanisms like DNA and histone modifications may contribute to epileptogenesis. Aberrant epigenetic patterns have already been identified in a number of central nervous system disorders like schizophrenia and Alzheimer disease, and this also includes epilepsy. It is still up to debate if the epigenetic changes are the cause or the result of many diseases. Here, we will discuss some recent findings suggesting that epileptogenesis alters the epigenetic landscape after seizures.

Seizure activity results in gene expression changes, including alterations in mRNA levels for GluR2 and bdnf (epileptogenesis-related genes). Following seizures in an animal model, histone acetyltransferases (HATs) mediated histone acetylation at the promoter regions of both genes, which could result in a higher expression of these genes. On the other hand, HDAC (histone deacetylase) inhibitors have been used as treatment for neurological disorders such as epilepsy. There has been evidence of hypermethylation  of the reelin promoter in association with temporal lobe epilepsy (TLE). Supporting this, there is increased expression of DNMT (DNA methyltransferase) in the neurons from the temporal neocortex of TLE patients. Histone methylation, like DNA methylation, may also play a role in epileptogenesis. The JARID1C histone demethylase is associated with X-linked mental retardation and in a minority of epilepsy patients. Also, as histone methylation is critical for memory formation, abnormal regulation of this methylation may lead to cogitive decline, which is associated with epilepsy. Some transcriptional activators recruit epigenetically related co-activators and repressors that influence chromatin restructuring. For example, the transciptional factor CREB and nuclear factor-kB are activated after a seizure and associate with HAT proteins to remodel chromatin. The transcription factor REST has also been implicated in the regulation of several epileptogenesis specific factors, like growth factors, neurotransmitter receptors, and ion channels. These finding suggest that there is a possibility of developing epigenetic based drug therapy. For example, VPA is a drug treatment used for epilepsy and was recently discovered to have HDAC inhibitory properties.

For more information, you can read the paper cited here:

Lubin, F. D. (2012). Epileptogenesis: can the science of epigenetics give us answers?. Epilepsy Currents, 12(3), 105-110.

“Epigenetics: Impacts on Disease Susceptibility and Pharmacotherapy” (Abubakar & Haque, 2016) in epilepsy

        Scientists have learned from epigenetics that the hereditary material that compromises all living beings, can be modified by simple and complex mechanisms, that can change completely the way it is expressed. Based on this, “genes that are linked to other diseases apart from cancer can be identified, silenced or activated using drugs or diets in order to treat the disease aetiology (or causes).” (2016)

        Since epigenetics is “involved in the control of the central nervous system and memory regulation” (2016), it has a tremendous impact on neurological disorders, such as epilepsy. This impact is made through regulation of ”genes responsible for signal transduction, inflammation, cell metabolism, ion transport, synaptic transmission, and stress” (2016). In the case of epilepsy, DNA methylation is increased in promoter regions found in temporal lobe epilepsy (TLE). This kind of epilepsy is the most common form with focal seizures. This may result in loss or trouble with memory, due to the function of the temporal lobe related to the creation of memories. The left temporal lobe is important for verbal memories, such as learning names or remembering facts for a test. Epilepsy in the left temporal lobe can cause problems remembering names and finishing sentences. The right temporal lobe is important for visual memories, such as remembering the face of another person or remembering how to get to a particular place (Epilepsy Society, 2015). Additional to hyper methylation in the promoter sequence of the temporal lobe, it has also been found in the Reelin DNA promoter. Reelin (RELN) is a large protein produced in the brain that triggers nerve cells through a signaling pathway in order to migrate them to their proper locations (Genetic Home References). “Reelin may also regulate synaptic plasticity, which is the ability of connections between neurons (synapses) to change and adapt over time in response to experience” (Genetic Home References). By over methylation of the gene that codes for the reelin protein, the proper connections between neurons cannot happen in the brain and therefore, communication is lost. “Once Reelin is silenced epigenetically, granule cell dispersion will be emerged causing epilepsy.” (Abubakar & Haque, 2016)

        Whenever someone’s been diagnosed with a health or neurological condition, the first question that arises is “can it be treated or cured?” Drugs used for treatment involving epigenetics, do not halt these epigenetic processes, but instead, correct the ketogenesis (Abubakar & Haque, 2016). There are various drugs for epigenetic pharmacotherapy, such as Vidaza, Zolinza and Vorinostat. They are divided by two classes: DNA methyltransferase inhibitors (Vidanza) and histone deacetylase inhibitors (Zolinza/Vorinostat) (Abubakar & Haque, 2016). As discussed previously on another post, a ketogenic diet with low fat, low carbohydrate, and high content, has been proven to decrease gene methylation. Similarly, these drugs also inhibit gene methylation, but on the other hand, they also promote acetylation. Another drug that works in the same way is imipramine. “DNAdemethylase removes the methyl group in the genome, and the methyl groups on the histone moiety are removed by histone demethylase. Therefore, induction of these enzymes through diet or pharmaco­therapy is likely to improve gene expression.” (Abubakar & Haque, 2016)

References:

(2015, August). How epilepsy can affect memory. Retrieved from https://www.epilepsysociety.org.uk/how-epilepsy-can-affect-memory

(2016, December). RELN gene. Retrieved from https://ghr.nlm.nih.gov/gene/RELN

Abubakar, A. & Haque, M. (2016) Epigenetics: Impact on Disease Susceptibility and Pharmacotherapy. Indian Journal of Phamaceutical Education and Research, 50(310-321). 

Study proves that “the administration of an anti-convulsive ketogenic diet is associated with gene regulating DNA methylation changes in rat TLE” (Kobow et al., 2013).

In epilepsy, it is shown that there is an increase, rather than a loss, of DNA methylation. The ketogenic diet (KD) is based on a high-fat and low-carbohydrate diet that acts as an anti-epileptic therapy.

It is recognized to work well in both humans and animal models, since it is well known that both, diet and environmental changes, play a very important role in the epigenome.  This diet was used by investigators, led by Katja Kobow, to examine methylation in the CpG islands of rats that are positive for temporal lobe epilepsy (TLE) (2013).  

In this study, the TLE rats were fed with a standard ketogenic diet, while strictly controlling body weight. Apart from the control group rats, the experimental group rats were prepared by inserting through brain surgery, a continuous video-electroencephalography monitor for studying of the brain before, during and after a seizure. DNA methylation profiling was made via examination of the extracted hippocampal tissue (2013). The investigators used Methyl-capture and massive parallel sequencing (Methyl-Seq) for analysis of this genomic DNA methylation. This was backed up by examination of mRNA (mRNA-Seq) sequencing from the same tissue.

The ketogenic diet was proven to regulate gene expression by modifying chromatin structure, by altering DNA methylation. Results showed that KD did not have a significant impact in the severity or duration of a clinical seizure, but it did however, affect the seizure frequency per week, by reducing those (2013). As mentioned before, epilepsy is related to hyper DNA methylation, but on the contrary, the KD diet was proven to “reduce DNA methylation at gene bodies as well as intronic and exonic regions.” (2013)

The KD, is “a well-recognized anti-epileptic treatment in children with severe and chronic epilepsy that delays chronification of the disease and partially rescues the DNA methylation and corresponding gene expression phenotype” (2013). This diet “interferes with aberrant seizure-related genomic and locus specific alterations in DNA methylation and gene expression” (2013). In this study, “both hyper- and hypomethylation events were detected with subsequent gene repression or activation” (2013).  It is not known exactly how the KD diet works to reduce DNA methylation and reduce the patient’s seizures. This study opens the doors for further epileptic analysis through epigenetics, in order to better understand and hopefully discover a new and better anti-seizure therapy.

References:

Kobow, K. et al. (2013) Deep sequencing reveals increased DNA methylation in chronic rat epilepsy. Acta Neuropathologica, 126(741-756). 

Access:

http://link.springer.com/article/10.1007/s00401-013-1168-8