MicroRNA Alterations and Associated Aberrant DNA Methylation Patterns across Multiple Sample Types in Oral Squamous Cell Carcinoma

Background: MicroRNA (miRNA) expression is broadly altered in cancer, but few studies have investigated miRNA deregulation in oral squamous cell carcinoma (OSCC). Epigenetic mechanisms are involved in the regulation of .30 miRNA genes in a range of tissues, and we aimed to investigate this further in OSCC. Methods: TaqManH qRT-PCR arrays and individual assays were used to profile miRNA expression in a panel of 25 tumors with matched adjacent tissues from patients with OSCC, and 8 control paired oral stroma and epithelium from healthy volunteers. Associated DNA methylation changes of candidate epigenetically deregulated miRNA genes were measured in the same samples using the MassArrayH mass spectrometry platform. MiRNA expression and DNA methylation changes were also investigated in FACS sorted CD44high oral cancer stem cells from primary tumor samples (CSCs), and in oral rinse and saliva from 15 OSCC patients and 7 healthy volunteers. Results: MiRNA expression patterns were consistent in healthy oral epithelium and stroma, but broadly altered in both tumor and adjacent tissue from OSCC patients. MiR-375 is repressed and miR-127 activated in OSCC, and we confirm previous reports of miR-137 hypermethylation in oral cancer. The miR-200 s/miR-205 were epigenetically activated in tumors vs normal tissues, but repressed in the absence of DNA hypermethylation specifically in CD44high oral CSCs. Aberrant miR-375 and miR-200a expression and miR-200c-141 methylation could be detected in and distinguish OSCC patient oral rinse and saliva from healthy volunteers, suggesting a potential clinical application for OSCC specific miRNA signatures in oral fluids. Conclusions: MiRNA expression and DNA methylation changes are a common event in OSCC, and we suggest miR-375, miR- 127, miR-137, the miR-200 family and miR-205 as promising candidates for future investigations. Although overall activated in OSCC, miR-200/miR-205 suppression in oral CSCs indicate that cell specific silencing of these miRNAs may drive tumor expansion and progression

MiR-137 Targets Estrogen-Related Receptor Alpha and Impairs the Proliferative and Migratory Capacity of Breast Cancer Cells

ERRα is an orphan nuclear receptor emerging as a novel biomarker of breast cancer. Over-expression of ERRα in breast tumor is considered as a prognostic factor of poor clinical outcome. The mechanisms underlying the dysexpression of this nuclear receptor, however, are poorly understood. MicroRNAs (miRNAs) regulate gene expression at the post-transcriptional level and play important roles in tumor initiation and progression. In the present study, we have identified that the expression of ERRα is regulated by miR-137, a potential tumor suppressor microRNA. The bioinformatics search revealed two putative and highly conserved target-sites for miR-137 located within the ERRα 3′UTR at nt 480–486 and nt 596–602 respectively. Luciferase-reporter assay demonstrated that the two predicted target sites were authentically functional. They mediated the repression of reporter gene expression induced by miR-137 in an additive manner. Moreover, ectopic expression of miR-137 down-regulated ERRα expression at both protein level and mRNA level, and the miR-137 induced ERRα-knockdown contributed to the impaired proliferative and migratory capacity of breast cancer cells. Furthermore, transfection with miR-137mimics suppressed at least two downstream target genes of ERRα–CCNE1 and WNT11, which are important effectors of ERRα implicated in tumor proliferation and migration. Taken together, our results establish a role of miR-137 in negatively regulating ERRα expression and breast cancer cell proliferation and migration. They suggest that manipulating the expression level of ERRα by microRNAs has the potential to influence breast cancer progression

Diet folate, DNA methylation and genetic polymorphisms of MTHFR C677T in association with the prognosis of esophageal squamous cell carcinoma

<p>Abstract</p> <p>Background</p> <p>Folic acid may affect the development of human cancers. However, few studies have evaluated the consumption of diet folate in the prognosis of patients with esophageal squamous cell carcinoma (ESCC).</p> <p>Methods</p> <p>One hundred and twenty five ESCC patients underwent esophagectomy between January 2005 and March 2006 in the Yangzhong People's Hospital were recruited and followed up. The effects of diet folate, aberrant DNA methylation of selected genes and methylenetetrahydrofolate reductase (<it>MTHFR</it>) C677T genetic polymorphisms on the prognosis of ESCC were evaluated by using Cox proportional hazard regression models.</p> <p>Results</p> <p>Our analysis showed an inverse association between diet folate intake and the risk of death after esophagectomy. The median survival time was 3.06 years for low or moderate folate consumption and over 4.59 years for high folate consumption. After adjusting for potential confounders, the hazard ratios (95% confidence interval) [HRs (95% CI)] were 0.72 (0.36-1.46) for moderate and 0.39 (0.20-0.78) for high folate intake, respectively (<it>P </it>for trend = 0.007). This preventive effect was more evident in patients carrying <it>MTHFR </it>677CC genotype. No significant relation was observed between aberrant DNA methylation of <it>P16</it>, <it>MGMT </it>and <it>hMLH1 </it>gene, as well as <it>MTHFR </it>C677T genetic polymorphisms and the prognosis of ESCC.</p> <p>Conclusions</p> <p>Our research indicated that diet folate intake may have benefits on the prognosis of ESCC after esophagectomy. From a practical viewpoint, the findings of our study help to establish practical intervention and surveillance strategies for managements of ESCC patients and can finally decrease the disease burden.</p

Measuring the morphological characteristics of thoracolumbar fascia in ultrasound images: an inter-rater reliability study

BACKGROUND: Chronic lower back pain is still regarded as a poorly understood multifactorial condition. Recently, the thoracolumbar fascia complex has been found to be a contributing factor. Ultrasound imaging has shown that people with chronic lower back pain demonstrate both a significant decrease in shear strain, and a 25% increase in thickness of the thoracolumbar fascia. There is sparse data on whether medical practitioners agree on the level of disorganisation in ultrasound images of thoracolumbar fascia. The purpose of this study was to establish inter-rater reliability of the ranking of architectural disorganisation of thoracolumbar fascia on a scale from ‘very disorganised’ to ‘very organised’. METHODS: An exploratory analysis was performed using a fully crossed design of inter-rater reliability. Thirty observers were recruited, consisting of 21 medical doctors, 7 physiotherapists and 2 radiologists, with an average of 13.03 ± 9.6 years of clinical experience. All 30 observers independently rated the architectural disorganisation of the thoracolumbar fascia in 30 ultrasound scans, on a Likert-type scale with rankings from 1 = very disorganised to 10 = very organised. Internal consistency was assessed using Cronbach’s alpha. Krippendorff’s alpha was used to calculate the overall inter-rater reliability. RESULTS: The Krippendorf’s alpha was .61, indicating a modest degree of agreement between observers on the different morphologies of thoracolumbar fascia.The Cronbach’s alpha (0.98), indicated that there was a high degree of consistency between observers. Experience in ultrasound image analysis did not affect constancy between observers (Cronbach’s range between experienced and inexperienced raters: 0.95 and 0.96 respectively). CONCLUSIONS: Medical practitioners agree on morphological features such as levels of organisation and disorganisation in ultrasound images of thoracolumbar fascia, regardless of experience. Further analysis by an expert panel is required to develop specific classification criteria for thoracolumbar fascia

Cervical radiculopathy: Study protocol of a randomised clinical trial evaluating the effect of mobilisations and exercises targeting the opening of intervertebral foramen [NCT01500044]

<p>Abstract</p> <p>Background</p> <p>Cervical radiculopathy is a common form of neck pain and has been shown to lead to severe disability. Clinical rehabilitation approaches for cervical radiculopathies commonly include exercise and manual therapy interventions targeting the opening of intervertebral foramen, but evidence regarding their effectiveness is scarce. The primary objective of this randomised clinical trial is to compare, in terms of pain and disability, a rehabilitation program targeting the opening of intervertebral foramen to a conventional rehabilitation program, for patients presenting acute or subacute cervical radiculopathies. The hypothesis is that the rehabilitation program targeting the opening of intervertebral foramen will be significantly more effective in reducing pain and disability than the conventional rehabilitation program.</p> <p>Methods/Design</p> <p>This study is a double-blind (participants and evaluators blinded) randomised clinical trial that will allow the comparison of patients with a cervical radiculopathy randomly assigned to one of two groups: one group will receive a 4-week rehabilitation program targeting the opening of intervertebral foramen, and the second group will receive a 4-week conventional rehabilitation program. Thirty-six subjects with cervical radiculopathy will be recruited from participating medical and physiotherapy clinics and will be evaluated at baseline, at the end of the 4-week program and four weeks following the end of the program. The primary outcome measure will be the validated Neck Disability Index questionnaire. Secondary outcome measures will include the short version of the Disabilities of the Arm, Shoulder and Hand questionnaire, a numerical pain rating scale, cervicothoracic mobility and patients' perceived global rating of change. During the 4-week rehabilitation program, each participant will take part in eight physiotherapy treatment sessions (2 session/week) and will perform a home exercise program. A mixed-model, 2-way ANOVA will be used to analyze the effects of the rehabilitation programs.</p> <p>Discussion</p> <p>Control trials are needed to define ideal intervention approaches in rehabilitation for this population. This randomised clinical trial will be the first study that directly compares a rehabilitation program targeting the opening of intervertebral foramen to a conventional rehabilitation program for patients with cervical radiculopathy. The results of this study may help to establish best clinical practice guidelines for this patient population.</p> <p>Trial Registration</p> <p>ClinicalTrials.gov: <a href="http://www.clinicaltrials.gov/ct2/show/NCT01500044">NCT01500044</a></p

Progressive Polycomb Assembly on H3K27me3 Compartments Generates Polycomb Bodies with Developmentally Regulated Motion

Polycomb group (PcG) proteins are conserved chromatin factors that maintain silencing of key developmental genes outside of their expression domains. Recent genome-wide analyses showed a Polycomb (PC) distribution with binding to discrete PcG response elements (PREs). Within the cell nucleus, PcG proteins localize in structures called PC bodies that contain PcG-silenced genes, and it has been recently shown that PREs form local and long-range spatial networks. Here, we studied the nuclear distribution of two PcG proteins, PC and Polyhomeotic (PH). Thanks to a combination of immunostaining, immuno-FISH, and live imaging of GFP fusion proteins, we could analyze the formation and the mobility of PC bodies during fly embryogenesis as well as compare their behavior to that of the condensed fraction of euchromatin. Immuno-FISH experiments show that PC bodies mainly correspond to 3D structural counterparts of the linear genomic domains identified in genome-wide studies. During early embryogenesis, PC and PH progressively accumulate within PC bodies, which form nuclear structures localized on distinct euchromatin domains containing histone H3 tri-methylated on K27. Time-lapse analysis indicates that two types of motion influence the displacement of PC bodies and chromatin domains containing H2Av-GFP. First, chromatin domains and PC bodies coordinately undergo long-range motions that may correspond to the movement of whole chromosome territories. Second, each PC body and chromatin domain has its own fast and highly constrained motion. In this motion regime, PC bodies move within volumes slightly larger than those of condensed chromatin domains. Moreover, both types of domains move within volumes much smaller than chromosome territories, strongly restricting their possibility of interaction with other nuclear structures. The fast motion of PC bodies and chromatin domains observed during early embryogenesis strongly decreases in late developmental stages, indicating a possible contribution of chromatin dynamics in the maintenance of stable gene silencing

DNA methylation levels are highly correlated between pooled samples and averaged values when analysed using the Infinium HumanMethylation450 BeadChip array.

BACKGROUND: DNA methylation is a heritable and stable epigenetic mark implicated in complex human traits. Epigenome-wide association studies (EWAS) using array-based technology are becoming widely used to identify differentially methylated sites associated with complex diseases. EWAS studies require large sample sizes to detect small effects, which increases project costs. In the present study we propose to pool DNA samples in methylation array studies as an affordable and accurate alternative to individual samples studies, in order to reduce economic costs or when low amounts of DNA are available. For this study, 20 individual DNA samples and 4 pooled DNA samples were analysed using the Illumina Infinium HumanMethylation450 BeadChip array to evaluate the efficiency of the pooling approach in EWAS studies. Statistical power calculations were also performed to discover the minimum sample size needed for the pooling strategy in EWAS. RESULTS: A total of 485,577 CpG sites across the whole genome were assessed. Comparison of methylation levels of all CpG sites between individual samples and their related pooled samples revealed highly significant correlations (rho > 0.99, p-val  0.98, p-val < 10(-16)). Also, it was calculated that n = 43 is the minimum sample size required to achieve a 95 % statistical power and a 10(-06) significance level in EWAS, when using a DNA pool strategy. CONCLUSIONS: DNA pooling strategies seems to accurately provide estimations of averaged DNA methylation state using array based EWAS studies. This type of approach can be applied to the assessment of disease phenotypes, reducing the amount of DNA required and the cost of large-scale epigenetic analyses

Exploring genetic resistance to infectious salmon anaemia virus in Atlantic salmon by genome-wide association and RNA sequencing

Funding The authors gratefully acknowledge funding from BBSRC (BB/R008612/1, BB/R008973/1), in addition to BBSRC Institute Strategic Programme Grants to the Roslin Institute (BB/P013759/1 and BB/P013740/1).Peer reviewedPublisher PD