Epigenetics explained: a topic “primer” for the epilepsy community by the ILAE Genetics/Epigenetics Task Force

Epigenetics refers broadly to processes that influence medium to long‐term gene expression by changing the readability and accessibility of the genetic code. The Neurobiology Commission of the International League Against Epilepsy (ILAE) recently convened a Task Force to explore and disseminate advances in epigenetics to better understand their role and intersection with genetics and the neurobiology of epilepsies and their co‐morbidities, and to accelerate translation of these findings into the development of better therapies. Here, we provide a topic primer on epigenetics, explaining the key processes and findings to date in experimental and human epilepsy. We review the growing list of genes with epigenetic functions that have been linked with epilepsy in humans. We consider potential practical applications, including using epigenetic signals as biomarkers for tissue‐ and biofluid‐based diagnostics and the prospects for developing epigenetic‐based treatments for epilepsy. We include a glossary of terms, FAQs and other supports to facilitate a broad understanding of the topic for the non‐expert. Last, we review the limitations, research gaps and the next challenges. In summary, epigenetic processes represent important mechanisms controlling the activity of genes, providing opportunities for insight into disease mechanisms, biomarkers and novel therapies for epilepsy.     

Differential role of RIP1 in Smac mimetic-mediated chemosensitization of neuroblastoma cells

We explored the potential of Smac mimetics, which antagonize Inhibitor of Apoptosis (IAP) proteins, for chemosensitization of neuroblastoma (NB). Here, we report that Smac mimetics, e.g. BV6, prime NB cells for chemotherapeutics including the topoisomerase II inhibitor doxorubicin (DOX) and vinca alkaloids such as Vincristine (VCR), Vinblastine (VBL) and Vinorelbine (VNR). Additionally, BV6 acts in concert with DOX or VCR to suppress long-term clonogenic growth. While BV6 causes rapid downregulation of cellular IAP (cIAP)1 protein and nuclear factor-kappaB (NF-κB) activation, DOX/BV6- or VCR/BV6-induced apoptosis occurs independently of NF-κB or TNFα signaling, since overexpression of dominant-negative IκBα superrepressor or the Tumor Necrosis Factor (TNF)α-blocking antibody Enbrel fail to block cell death. Mechanistic studies reveal that Receptor-interacting protein (RIP)1 is required for DOX/BV6-, but not for VCR/BV6-induced apoptosis, since transient or stable knockdown of RIP1 or the pharmacological RIP1 inhibitor necrostatin-1 significantly reduce apoptosis. By comparison, VCR/BV6-mediated apoptosis critically depends on the mitochondrial pathway. VCR/BV6 cotreatment causes phosphorylation of BCL-2 during mitotic arrest, enhanced activation of BAX and BAK and loss of mitochondrial membrane potential (MMP). Additionally, overexpression of BCL-2 profoundly suppresses VCR/BV6-induced apoptosis. Thus, BV6 sensitizes NB cells to chemotherapy-induced apoptosis via distinct initial signaling mechanisms depending on the chemotherapeutic drug. These findings provide novel mechanistic insights into Smac mimetic-mediated chemosensitization of NB.     

Fatal neonatal nephrocutaneous syndrome in 18 Roma children with EGFR deficiency

Epidermal growth factor receptor (EGFR) is a transmembrane glycoprotein with tyrosine-kinase signaling activity, involved in many cellular functions including cell growth and differentiation. Germ line loss-of-function mutations in EGFR lead to a severe neonatal skin disorder (Online Mendelian Inheritance in Man #131550). We report 18 premature Roma children from 16 families with birthweights ranging 440–1470 g and multisystem diseases due to the homozygous mutation c.1283G˃A (p.Gly428Asp) in EGFR. They presented with thin, translucent, fragile skin (14/15), skin desquamation (10/17), ichthyosis (9/17), recurrent skin infections and sepsis (9/12), nephromegaly (10/16) and congenital heart defects (7/17). Their prognosis was poor, and all died before the age of 6 months except one 13-year-old boy with a severe skin disorder, dentinogenesis imperfecta, Fanconi-like syndrome and secondary hyperaldosteronism. Management of ion and water imbalances and extremely demanding skin care may improve the unfavorable outcome of such patients.     

Flow calorimetry--a useful tool for determination of immobilized cis-epoxysuccinate hydrolase activity from Nocardia tartaricans

Bacterial cells Nocardia tartaricans with cis-epoxysuccinate hydrolase activity were entrapped in hardened calcium pectate gel by a commercial high performance encapsulator. This enzyme (in a single step reaction with no formation of side products) was used to hydrolyze disodium cis-epoxysuccinate to a pure enantiomer--disodium L-(+)-tartrate. Activities of this enzyme were determined using flow calorimetry. The validity of this method was corroborated by HPLC and isotachophoresis. The immobilized biocatalyst has activity (75.8 U/mgdry) able to convert disodium cis-epoxysuccinate to disodium tartrate at 94% yield in 5.5h. Immobilization of N. tartaricans in hardened calcium pectate gel beads had a positive effect on the activity of cis-epoxysuccinate hydrolase, storage stability, yield, and time of bioconversion.     

High incidences of DNA ploidy abnormalities in tongue squamous cell carcinoma of young patients: an international collaborative study

Santos‐Silva A R, Ribeiro A C P, Soubhia A M P, Miyahara G I, Carlos R, Speight P M, Hunter K D, Torres‐Rendon A, Vargas P A & Lopes M A (2011) Histopathology  58 , 1127–1135
High incidences of DNA ploidy abnormalities in tongue squamous cell carcinoma of young patients: an international collaborative study Aims: This multi‐centre analysis assessed the DNA content of TSCC in 37 young patients (<40 years) and 28 old patients (>50 years) and determined the correlation of DNA ploidy findings with clinicopathological data. Methods and results: Image cytometry was carried out using an automated cellular imaging system on Feulgen‐stained histological sections to obtain high‐fidelity DNA histograms. Among young patients, 37.8% were females compared to 18.7% in the older group (P = 0.002). In total, 48.6% patients were non‐smokers and 40.5% were non‐drinkers compared to 10.7% non‐smokers and non‐drinkers in the older group (P < 0.0001). TNM, clinical stage of disease and histological grade of differentiation did not differ between groups. Tumour aneuploidy was detected in 86.5% and tetraploidy in 24.3% young patients; this was significantly greater than in the older group where 64.3% were aneuploid (P < 0.0001) and 7.2% tetraploid (P < 0.0001). The mean values of DNA index (DI) and DNA heterogeneity index as well as the percentage of cells with DI exceeding 5N were higher in young patients (P < 0.0001). Conclusions: Young patients with TSCC represent a distinct clinical entity. The high incidence of DNA ploidy abnormalities suggest that they may have increased genomic instability and indicates underlying genetic differences between TSCC in young and older patients.     

Rai1 deficiency in mice causes learning impairment and motor dysfunction, whereas Rai1 heterozygous mice display minimal behavioral phenotypes

Smith-Magenis syndrome (SMS) is associated with an approximately 3.7 Mb common deletion in 17p11.2 and characterized by its craniofacial and neurobehavioral abnormalities. The reciprocal duplication leads to dup(17)(p11.2p11.2) associated with the Potocki-Lupski syndrome (PLS), a neurological disorder whose features include autism. Retinoic acid induced 1 (RAI1) appears to be responsible for the majority of clinical features in both SMS and PLS. Mouse models of these syndromes harboring an approximately 2 Mb chromosome engineered deletion and duplication, respectively, displayed abnormal locomotor activity and/or learning deficits. To determine the contribution of RAI1 in the neurobehavioral traits in SMS, we performed a battery of behavioral tests on Rai1 mutant mice and the Df(11)17-1/+ mice that have a small deletion of approximately 590 kb. The mice with the small deletion were hypoactive like the large deletion mice and they also showed learning deficits. The Rai1+/- mice exhibited normal locomotor activity. However, they had an abnormal electroencephalogram with overt seizure observed in a subset of mice. The few surviving Rai1-/- mice displayed more severe neurobehavioral abnormalities including hind limb clasping, overt seizures, motor impairment and context- and tone-dependant learning deficits. X-gal staining of the Rai1+/- mice suggests that Rai1 is predominantly expressed in neurons of the hippocampus and the cerebellum. Our results suggest that Rai1 is a critical gene in the central nervous system functioning in a dosage sensitive manner and that the neurobehavioral phenotype is modified by regulator(s) in the approximately 590 kb genomic interval, wherein the major modifier affecting the craniofacial penetrance resides.     

NOS1 ex1f-VNTR polymorphism influences prefrontal brain oxygenation during a working memory task

Nitric oxide (NO) synthase produces NO, which serves as first and second messenger in neurons, where the protein is encoded by the NOS1 gene. A functional variable number of tandem repeats (VNTR) polymorphism in the promoter region of the alternative first exon 1f of NOS1 is associated with various functions of human behavior, for example increased impulsivity, while another, non-functional variant was linked to decreased verbal working memory and a heightened risk for schizophrenia. We therefore investigated the influence of NOS1 ex 1f-VNTR on working memory function as reflected by both behavioral measures and prefrontal oxygenation. We hypothesized that homozygous short allele carriers exhibit altered brain oxygenation in task-related areas, namely the dorsolateral and ventrolateral prefrontal cortex and the parietal cortex. To this end, 56 healthy subjects were stratified into a homozygous long allele group and a homozygous short allele group comparable for age, sex and intelligence. All subjects completed a letter n-back task (one-, two-, and three-back), while concentration changes of oxygenated (O(2)Hb) hemoglobin in the prefrontal cortex were measured with functional near-infrared spectroscopy (fNIRS). We found load-associated O(2)Hb increases in the prefrontal and parts of the parietal cortex. Significant load-associated oxygenation differences between the two genotype groups could be shown for the dorsolateral prefrontal cortex and the parietal cortex. Specifically, short allele carriers showed a significantly larger increase in oxygenation in all three n-back tasks. This suggests a potential compensatory mechanism, with task-related brain regions being more active in short allele carriers to compensate for reduced NOS1 expression.     

NOS1 ex1f-VNTR polymorphism affects prefrontal oxygenation during response inhibition tasks

Impulsivity is a trait shared by many psychiatric disorders and therefore a suitable intermediate phenotype for their underlying biological mechanisms. One of the molecular determinants involved is the NOS1 ex1f‐VNTR, whose short variants are associated with a variety of impulsive behaviors. Fifty‐six healthy controls were stratified into homozygous long (LL) (30 probands) and short (SS) (26 probands) allele groups. Subjects completed a combined stop‐signal go/nogo task, while the oxygenation in the prefrontal cortex was measured with functional near‐infrared spectroscopy. Electromyography was recorded to control for differences in muscle activity in the two inhibition tasks. Two questionnaires on impulsive traits were completed. Differences between the two tasks are shown by distinct activation patterns within the prefrontal cortex. The nogo task resulted mainly in the activation of the dorsolateral prefrontal cortex (dlPFC), whereas successful and unsuccessful inhibition in the stop‐signal task elicited the predicted activity in the inferior frontal cortex (IFC). Although significant differences were found in neither the scores obtained on impulsivity‐related questionnaires nor the behavioral data, the LL group displayed increased dlPFC activity during nogo trials and the predicted activation in the IFC during successful inhibition in the stop‐signal task, while no significant activation was found in the SS group. Our data confirm an influence of NOS1 ex1f‐VNTR on impulsivity, as carriers of the short risk allele exhibited diminished activity of (pre‐)frontal brain regions during the inhibition in a stop‐signal task. Impairment of prefrontal control with consecutive failure of inhibitory processes might underlie association findings reported previously. Hum Brain Mapp, 2012. © 2011 Wiley Periodicals, Inc.     

The epilithon of a cooling tower of the power plant at Bełchatów, Poland

We studied the diversity of the photosynthetic microscopic epilithon overgrowing the concrete walls of a cooling tower of the power plant at Be?chatów, central Poland. Epilithon samples were collected from the open upper part of concrete walls of a cooling tower in March 2006 and examined in labs as dried material in September 2006, and again in March 2011. The aerophytic assemblages were strongly dominated by a nostocalean cyanobacterium, Scytonema myochrous C. Agardh ex Bornet et Flahault. Other cyanobacteria and algae occurred very sporadically and in low abundance. The trichome morphology of S. myochrous growing directly on the tower differs considerably from that of lab-cultured ones. Trichomes from field material were very long, conspicuously sheathed and dark brown, with relatively sparse ramification and with cylindrical cells and heterocytes. Trichomes cultivated in the laboratory were shorter and thicker, with thin, hyaline sheaths which were not layered; they were frequently ramified, and the cells and heterocytes were discoid. Besides Scytonema myochrous, the first revitalized cyanobacteria growing in laboratory conditions included Nostoc sp., Leptolyngbya gracillima (Zopf ex Hansgirg) Anagnostidis et Komárek, and Gloeothece rupestris (Lyngbye) Bornet in Witrock et Nordstedt.