Evidence of a conserved functional role for DNA methylation in termites

Many organisms are capable of developing distinct phenotypes from the same genotype. This developmental plasticity is particularly prevalent in insects, which can produce alternate adaptive forms in response to distinct environmental cues. The ability to develop divergent phenotypes from the same genotype often relies on epigenetic information, which affects gene function and is transmitted through cell divisions. One of the most important epigenetic marks, DNA methylation, has been lost in several insect lineages, yet its taxonomic distribution and functional conservation remain uninvestigated in many taxa. In the present study, we demonstrate that the signature of high levels of DNA methylation exists in the expressed genes of two termites, Reticulitermes flavipes and Coptotermes formosanus. Further, we show that DNA methylation is preferentially targeted to genes with ubiquitous expression among morphs. Functional associations of DNA methylation are also similar to those observed in other invertebrate taxa with functional DNA methylation systems. Finally, we demonstrate an association between DNA methylation and the long‐term evolutionary conservation of genes. Overall, our findings strongly suggest DNA methylation is present at particularly high levels in termites and may play similar roles to those found in other insects.     

Epigenome‐wide DNA methylation changes with development of arsenic‐induced skin lesions in Bangladesh: A case–control follow‐up study

Studies have found an association between aberrant DNA methylation and arsenic‐induced skin lesions. However, little is known about DNA methylation changes over time in people who develop arsenic‐induced skin lesions. We sought to investigate epigenome‐wide changes of DNA methylation in people who developed arsenic‐induced skin lesions in a 10‐year period. In 2009–2011, we conducted a follow‐up study of 900 skin lesion cases and 900 controls and identified 10 people who developed skin lesions since a baseline survey in 2001–2003. The 10 cases (“New Cases”) were matched with 10 controls who did not have skin lesions at baseline or follow‐up (“Persistent Controls”). Drinking water and blood samples were collected, and skin lesion was diagnosed by the same physician at both time points. We measured DNA methylation in blood using Infinium HumanMethylation450K BeadChip, followed by quantitative validation using pyrosequencing. Two‐sample t‐tests were used to compare changes in percent methylation between New Cases and Persistent Controls. Six CpG (cytosine‐phosphate‐guanine) sites with greatest changes of DNA methylation over time among New Cases were further validated with a correlation of 93% using pyrosequencing. One of the validated CpG site (cg03333116; change of %methylation was 13.2 in New Cases versus ?0.09 in Persistent Controls; P < 0.001) belonged to the RHBDF1 gene, which was previously reported to be hypermethylated in arsenic‐exposed cases. We examined DNA methylation changes with the development of arsenic‐induced skin lesions over time but nothing was statistically significant given the small sample size of this exploratory study and the high dimensionality of data. Environ. Mol. Mutagen. 55:449–456, 2014. © 2014 Wiley Periodicals, Inc.     

Method for Detecting Toxin‐Induced Gamete DNA Damage in Male Rats

Despite the vast amount of literature on studies about the use of reproductive toxicants in laboratory rats as models, there are no reports documenting the detection of direct sperm DNA damage in rats as a result of exposure to a reproductive toxicant. These studies were initiated to develop methodology for detecting and scoring oxidative sperm DNA damage in Wistar rats. The first of four experiments used the basic Comet Assay to compare sperm collected from xanthotoxin‐treated rats with sperm collected from untreated rats to determine if oxidative sperm DNA damage was detectable. No differences were found between images of sperm from treated and non‐treated samples. During Experiment 2, fresh sperm samples from untreated rats were stored in PBS and then reacted with a 100mg/ml solution of xanthotoxin/acetone. Post‐reaction cells were subjected to mechanical sample grinding or incubation with trypsin in an attempt to disrupt the cell capsule. Again, no differences were recorded. Experiment 3 exposed treated and untreated sperm samples to proteinase K for varying time intervals (30 min – 3 hr) in another effort to disrupt the cell capsule. These proved to be promising, although at the time intervals used in Experiment 3, the entire cell was dissolved. A series of tests were then initiated during Experiment 4 to determine the best buffer and reaction time combination. All things were kept identical to Experiment 3 with the exception that samples were taken at more frequent intervals and a different buffer was used with each sample. The buffers used were PBS, TES, and PBS containing Chelex. The best results were obtained with TES at a 55‐minute time interval. The capsule surrounding the spermatatids proved to be very resilient to most digestive and mechanical agents. The enzyme proteinase K proved to be the best means for disrupting the cell capsule. Proteinase K in a TES buffer worked most effectively for detecting and scoring oxidative sperm DNA damage in Wistar rats.     

Oxidative stress and DNA damage responses in rat and mouse lung to inhaled carbon nanoparticles

Abstract

We have investigated whether short-term nose-only inhalation exposure to electric spark discharge-generated carbon nanoparticles (～60 nm) causes oxidative stress and DNA damage responses in the lungs of rats (152 μg/m³; 4 h) and mice (142 μg/m³; 4 h, or three times 4 h). In both species, no pulmonary inflammation and toxicity were detected by bronchoalveolar lavage or mRNA expression analyses. Oxidative DNA damage (measured by fpg-comet assay), was also not increased in mouse whole lung tissue or isolated lung epithelial cells from rat. In addition, the mRNA expressions of the DNA base excision repair genes OGG1, DNA Polβ and XRCC1 were not altered. However, in the lung epithelial cells isolated from the nanoparticle-exposed rats a small but significant increase in APE-1 mRNA expression was measured. Thus, short-term inhalation of carbon nanoparticles under the applied exposure regimen, does not cause oxidative stress and DNA damage in the lungs of healthy mice and rats.     

EFFECT OF GROWTH FACTORS ON DNA LABELING AND CYTOSKELETAL PROTEIN EXPRESSION IN 17-β-ESTRADIOL AND BASIC FIBROBLAST GROWTH FACTOR PRE-TREATED ASTROCYTE CULTURES

Astrocytes react to all noxae which damage neurons. Their reactions include degeneration, hypertrophy, hyperplasia and fibre formation. Growth factors inducing proliferation and differentiation of both neurons and astrocytes in culture play a pivotal role in the dynamic flow of signaling molecules between neurons and astroglia. Estrogens as well influence astroglia and are neuroprotectants. This study has investigated the interactions between growth factors and estrogens on DNA labeling and cytoskeletal protein [glial fibrillary acidic protein (GFAP) and vimentin] expression in 22 DIV astrocyte cultures treated for 24 or 36?h under different experimental conditions.

Contemporary addition of 17-β-estradiol (E₂) with two or three growth factors for 24?h, significantly stimulated methyl-[³H]thymidine incorporation into DNA from 22 days in vitro (DIV) astrocyte cultures.

This effect reached a peak when E₂ was co-added with epidermal growth factor (EGF), basic fibroblast growth factor (bFGF) and insulin. In astrocyte cultures treated for 36?h with E₂ and EGF+insulin or bFGF+insulin added in the last 12?h, DNA labeling was remarkably increased. The parallel cyclin D1 expression positively correlated with ERK2 activation. Western blot analysis for cytoskeletal proteins showed also changes of both GFAP and vimentin expression. The above data suggest the occurrence of a scheduled interaction between “competence” or “progression” growth factors and estrogens on DNA labeling and cytoskeletal protein expression during astroglial cell proliferation and differentiation in culture. A better understanding of the mechanisms of these interactions may contribute to develop strategies for controlling astroglial reaction in cerebrovascular disease including stroke and hypertensive brain damage.     

Gene mutations in Wilson disease in Egyptian children: Report on two novel mutations

Background and study aimsWilson disease (WD) is an autosomal recessive disorder, caused by defects in copper-transporting P-type adenosine triphosphatase (ATPase) encoded by the ATP7B gene, resulting in the deposition of copper in the liver and brain with significant disability or death if left untreated. An available regimen of treatment gives hope to those predisposed to the disease if diagnosed early. The objective of this study was to determine the frequency of the most common European mutation (p.H1069Q) in Egyptian children with WD, in addition to screening for previously reported mutations in the Egyptian patients in our selected group.Patients and methodsDirect DNA sequencing was applied to exons (13, 14, 18, and 19) of the ATP7B gene for 19 patients previously diagnosed with WD. Then DNA sequencing and pedigree analysis were performed in the families of the patients showing variations in their results for the purpose of family screening and carrier detection. Six out of 19 patients were studied with their families (three families).ResultsWe identified five variants of which two were novel among the studied patients. One of the novel variants was synonymous substitution (p.A1074A) in 16% of patients and the other was predicted to be missense disease-causing mutations (p.T1076I) in 16% of patients, and three previously published mutations p.H1069Q were detected in 5% of patients, p.P1273Q in 10% of patients, and a silent variant p.A1003A in 26% of patients.ConclusionScreening for the two exons 14 and 18 of the ATP7B gene is important in Egyptian patients especially in suspected patients without hepatic manifestations.     

The rediscovery of smallpox

Smallpox is an infectious disease that is unique to humans, caused by a poxvirus. It is one of the most lethal of diseases; the virus variant Variola major has a mortality rate of 30%. People surviving this disease have life-long consequences, but also assured immunity. Historically, smallpox was recognized early in human populations. This led to prevention attempts—variolation, quarantine, and the isolation of infected subjects—until Jenner’s discovery of the first steps of vaccination in the 18th century. After vaccination campaigns throughout the 19th and 20th centuries, the WHO declared the eradication of smallpox in 1980. With the development of microscopy techniques, the structural characterization of the virus began in the early 20th century. In 1990, the genomes of different smallpox viruses were determined; viruses could be classified in order to investigate their origin, diffusion, and evolution. To study the evolution and possible re-emergence of this viral pathogen, however, researchers can only use viral genomes collected during the 20th century. Cases of smallpox in ancient periods are sometimes well documented, so palaeomicrobiology and, more precisely, the study of ancient smallpox viral strains could be an exceptional opportunity. The analysis of poxvirus fragmented genomes could give new insights into the genetic evolution of the poxvirus. Recently, small fragments of the poxvirus genome were detected. With the genetic information obtained, a new phylogeny of smallpox virus was described. The interest in conducting studies on ancient strains is discussed, in order to explore the natural history of this disease.     

Total neoadjuvant therapy: Fact,fantasy, or fallacy?

Colorectal cancer is the second leading cause of cancer death in the United States for men and women, with an estimated 146,000 new cases per year - a staggering 53,000 patients die each year. Rectal cancer comprises a third of these patients, with a 5-year survival rate of 67%. Management of locally advanced rectal cancer in the U.S. had remained stagnant for more than a decade, with most of these patients being treated with long-course chemoradiotherapy, surgery, followed by adjuvant chemotherapy; adjuvant chemotherapy being administered despite lacking a high level of evidence. Over the past few years, with rectal cancer death rates exceeding 30% from metastatic disease, growing interest focused on the attributes of induction chemotherapy to eradicate minimal residual disease and purportedly increase survival. This led to the development of total neoadjuvant therapy (TNT). We now have high-quality data from randomized prospective trials to review the facts, fantasies, and fallacies of TNT.