Rapid and quantitative detection of CpG-methylation status using TaqMan PCR combined with methyl-binding-domain polypeptide

ObjectivesTo assess the medical applicability of CpG methylation as molecular markers for cancer diagnosis, we established a new system to determine DNA methylation based on TaqMan PCR combined with a methyl-binding-domain polypeptide 2.Design and methodsWe evaluated the diagnostic applicability of this approach by examining the methylation status of two tumor suppressor genes, RASSF1A and APC, in 10 paired hepatocellular carcinoma (HCC) and the corresponding non-tumor liver tissues.ResultsMethylation levels of total 20 clinical samples measured by the TaqMan PCR assay showed a significantly positive correlation (R = 0.814, P < 0.0005 for RASSF1A, R = 0.736, P < 0.00001 for APC) with those calculated by bisulfite sequencing. The methylated DNA amount measured by our TaqMan PCR system precisely replicated the methylation status estimated by direct sequencing.ConclusionsThis suggests our method may serve as a reliable and easy-to-use tool for cancer diagnosis using methylated genes as biomarkers.     

Clinical significance of Stratifin,ERα and PR promoter methylation in tumor and serum DNA in Indian breast cancer patients

Objectives:The objective of this study was to determine the concordance of promoter methylation of stratifin, ERα and PR in tumor and circulating DNA in breast cancer patients and their association with clinicopathological parameters and disease prognosis.Design and methods:Methylation specific PCR were carried out to investigate the promoter methylation status of stratifin, ERα and PR in tumor and circulating DNA in 100 breast cancer patients in a prospective study. The effect of promoter methylation on protein expression was evaluated by immunohistochemistry.Results:Significant association was observed between promoter methylation of stratifin in tumors (61%) and paired sera (56%) (r = 0.78; p ≤ 0.001). Loss of stratifin expression was observed in 47% tumors and was associated with poor overall survival (p = 0.05). Significant correlation was observed between methylation status of ERα with PRB (p < 0.0001, OR = 20.8, 95% CI = 7.4–58.0) and stratifin (p = 0.003, OR = 2.0, 95% CI = 0.8–4.4).Conclusion:This study underscores the potential utility of serum DNA methylation of these genes as surrogate for tumor DNA methylation as a promising tool for cancer diagnosis.     

Aberrant hypomethylation of SALL4 gene is associated with intermediate and poor karyotypes in acute myeloid leukemia

ObjectiveSALL4 gene has been identified to stimulate the expansion of hematopoietic stem cell (HSCs) and enhance the self-renewal of HSCs. Overexpression of SALL4 has been found in several cancers. The present study was aimed to investigate the methylation status of SALL4 promoter region in acute myeloid leukemia (AML).Designs and methodsThe methylation status of SALL4 promoter was analyzed in 84 patients with AML using methylation-specific PCR (MSP) and its clinical significance was evaluated.ResultsAberrant hypomethylation of SALL4 gene, which was correlated with SALL4 expression, was found in 17.8% (15/84) cases. The patients with SALL4 hypomethylation had significantly older age and higher WBCs than those without SALL4 hypomethylation. The incidence of SALL4 hypomethylation was higher in M1 subtype than in M2 and other subtypes (50%, 26% and 6%, respectively, P = 0.001). SALL4 hypomethylation was associated with cytogenetically intermediate and poor groups. Although survival time of the SALL4-hypomethylated AML was shorter than that of SALL4-methylated group (4 months vs 9 months), the difference was not statistically significant (P = 0.356).ConclusionsHypomethylation of SALL4 promoter is a common event and is associated with the intermediate and poor karyotypes in AML.     

Hippocampal volumetry for lateralization of temporal lobe epilepsy: Automated versus manual methods

The hippocampus has been the primary region of interest in the preoperative imaging investigations of mesial temporal lobe epilepsy (mTLE). Hippocampal imaging and electroencephalographic features may be sufficient in several cases to declare the epileptogenic focus. In particular, hippocampal atrophy, as appreciated on T1-weighted (T1W) magnetic resonance (MR) images, may suggest a mesial temporal sclerosis. Qualitative visual assessment of hippocampal volume, however, is influenced by head position in the magnet and the amount of atrophy in different parts of the hippocampus. An entropy-based segmentation algorithm for subcortical brain structures (LocalInfo) was developed and supplemented by both a new multiple atlas strategy and a free-form deformation step to capture structural variability. Manually segmented T1-weighted magnetic resonance (MR) images of 10 non-epileptic subjects were used as atlases for the proposed automatic segmentation protocol which was applied to a cohort of 46 mTLE patients. The segmentation and lateralization accuracies of the proposed technique were compared with those of two other available programs, HAMMER and FreeSurfer, in addition to the manual method. The Dice coefficient for the proposed method was 11% (p < 10^?5) and 14% (p < 10^?4) higher in comparison with the HAMMER and FreeSurfer, respectively. Mean and Hausdorff distances in the proposed method were also 14% (p < 0.2) and 26% (p < 10^?3) lower in comparison with HAMMER and 8% (p < 0.8) and 48% (p < 10^?5) lower in comparison with FreeSurfer, respectively. LocalInfo proved to have higher concordance (87%) with the manual segmentation method than either HAMMER (85%) or FreeSurfer (83%). The accuracy of lateralization by volumetry in this study with LocalInfo was 74% compared to 78% with the manual segmentation method. LocalInfo yields a closer approximation to that of manual segmentation and may therefore prove to be more reliable than currently published automatic segmentation algorithms.     

Rapid determination of AKAP12 promoter methylation levels in peripheral blood using methylation-sensitive high resolution melting (MS-HRM) analysis: Application in colorectal cancer

BackgroundColorectal cancer is the third most common form of cancer and hypermethylation has been shown to increase the risk of developing this disease. DNA hypermethylation in the A kinase anchor protein 12 (AKAP12/Gravin) promoter region and the accompanied underexpression of it has been noted in a variety of human cancers.MethodsWe applied methylation-specific high resolution melting (MS-HRM) technology to detect quantitatively A kinase anchor protein 12 (AKAP12/Gravin) methylation in peripheral blood from 100 colorectal cancer patients and 50 healthy volunteers and in 3 colorectal cancer cell lines.ResultsIn this study 48 of the 100 colorectal cancer samples (48%) were found to be methylated at the AKAP12 promoter region. AKAP12 methylation was significantly higher in the colorectal cancer samples with differentiation (p = 0.03). We also compared the results generated by MS-HRM with a traditional methylation-specific PCR (MSP) assay. We found that intra-assay variability ranged from 6.14 to 9.90% and inter-assay variability ranged from 14.5 to 17.2%. The AKAP12 MS-HRM assay was able to reproducibly detect 1% methylated DNA, whereas the MSP method was unable to detect less than 5% methylation.ConclusionsWe demonstrate the utility of quantitative AKAP12 MS-HRM analysis of promoter methylation in peripheral blood samples. AKAP12 MS-HRM quantitative methods with excellent detection capabilities have many promising applications in the research and diagnosis of colorectal cancer.     

Methylated cyclin D2 gene circulating in the blood as a prognosis predictor of hepatocellular carcinoma

BackgroundPrognosis of hepatocellular carcinoma (HCC) remains poor because of high recurrence rate. We examined preoperatively the methylated CCND2 gene levels present in the serum following release from HCC cells as a prognosis predictor in patients undergoing curative hepatectomy.MethodsQuantitative real-time RT-PCR and quantitative methylation-specific PCR were used to measure methylated CCND2 gene and its mRNA levels.ResultsThe CCND2 mRNA levels were down-regulated in HCC with early intrahepatic recurrence (IHR) within 1 year of curative hepatectomy. We also identified that this down-regulation was due to promoter hypermethylation. In 70 HCC patients who underwent curative hepatectomy, 39 patients sero-positive for the methylated CCND2 gene (> 70 pg/ml serum) exhibited a significantly shorter disease-free survival (DFS) period (P = 0.02) than the 31 patients who were sero-negative for the methylated CCND2 gene. None of the sero-negative patients demonstrated early IHR, and this method of serum testing did not produce any false-negative predictions for early IHR. Multivariate analysis showed that the serum level of methylated CCND2 was an independent risk factor for DFS (hazard ratio of 1.866, 95% CI: 1.106–3.149).ConclusionMethylated CCND2 gene in the serum serves as a prognosis predictor of HCC after curative hepatectomy.     

Prognostic significance of RASSF1A promoter methylation in operable breast cancer

ObjectivesThe aim of our study was to evaluate the prognostic significance of RASSF1A promoter methylation status in operable breast cancer.Design and methodsBy using Methylation Specific PCR, we evaluated the specificity of RASSF1A promoter methylation in 10 breast tumors and matching normal tissues, 10 breast fibroadenomas and 11 normal breast tissues. The prognostic significance of RASSF1A methylation was validated in 93 formalin fixed paraffin-embedded (FFPE) tissues obtained from patients with operable breast cancer.ResultsMethylation of RASSF1A promoter was observed in 1/31 (3.2%) non-cancerous breast tissues and 53/93 (57.0%) early stage breast tumors. The only positive sample in the non-cancerous breast tissues group was found in a histological normal tissue surrounding the tumor. During the follow-up period, 24/93 (25.8%) patients relapsed and 19/93 (20.4%) died. Disease-Free-Interval (DFI) was significantly associated with RASSF1A methylation (p = 0.028).ConclusionsRASSF1A promoter methylation provides important prognostic information in early stage breast cancer patients.     

MP2RAGE,a self bias-field corrected sequence for improved segmentation and T1-mapping at high field

The large spatial inhomogeneity in transmit B₁ field (B₁⁺) observable in human MR images at high static magnetic fields (B₀) severely impairs image quality. To overcome this effect in brain T₁-weighted images, the MPRAGE sequence was modified to generate two different images at different inversion times, MP2RAGE. By combining the two images in a novel fashion, it was possible to create T₁-weigthed images where the result image was free of proton density contrast, T₂? contrast, reception bias field, and, to first order, transmit field inhomogeneity.MP2RAGE sequence parameters were optimized using Bloch equations to maximize contrast-to-noise ratio per unit of time between brain tissues and minimize the effect of B₁⁺ variations through space. Images of high anatomical quality and excellent brain tissue differentiation suitable for applications such as segmentation and voxel-based morphometry were obtained at 3 and 7 T.From such T₁-weighted images, acquired within 12 min, high-resolution 3D T₁ maps were routinely calculated at 7 T with sub-millimeter voxel resolution (0.65–0.85 mm isotropic). T₁ maps were validated in phantom experiments. In humans, the T₁ values obtained at 7 T were 1.15 ± 0.06 s for white matter (WM) and 1.92 ± 0.16 s for grey matter (GM), in good agreement with literature values obtained at lower spatial resolution. At 3 T, where whole-brain acquisitions with 1 mm isotropic voxels were acquired in 8 min, the T₁ values obtained (0.81 ± 0.03 s for WM and 1.35 ± 0.05 for GM) were once again found to be in very good agreement with values in the literature.     

Identification of a novel IRGM promoter single nucleotide polymorphism associated with tuberculosis

BackgroundHuman immunity-related GTPase M (IRGM) is found to play an important role in defense against intracellular pathogen Mycobacterium tuberculosis in vitro by regulating autophagy. To verify whether single nucleotide polymorphisms (SNPs) in the promoter region of IRGM gene are associated with tuberculosis (TB) 1.7 kb IRGM promoter region was sequenced and SNP analysis was conducted in TB patients and healthy controls.MethodsA simple and rapid procedure for extracting DNA from clotted-blood was developed in this study. A 1.7 kb IRGM promoter region was amplified and sequenced for nucleotide polymorphism search. Then, 3 SNPs were selected and analyzed in 216 TB patients and 275 healthy subjects by ligase detection reaction technique.ResultsDNA extracted by our method was of high quality and suitable for PCR, sequencing, and genotyping. We identified 29 polymorphisms in the 1.7 kb IRGM promoter region, including 11 novel polymorphisms not yet reported. Large population analysis showed that frequencies of ? 1208A allele (P = 0.031), ? 1208AA genotype (P = 0.042), and ? 1208A/?1161C/?947C (P = 0.035) and ? 1208G/?1161C/?947C (P = 0.030) haplotypes in cases were significantly different from those in controls.ConclusionsIn 1.7 kb IRGM promoter region, only ? 1208A/G polymorphism is associated with susceptibility to TB.     

Mayo Registry for Telemetry Efficacy in Arrest (MR TEA) study: An analysis of code status change following cardiopulmonary arrest

IntroductionCode status discussions are important during a hospitalization, yet variation in its practice exists. No data have assessed the likelihood of patients to change code status following a cardiopulmonary arrest.MethodsA retrospective review of all patients that experienced a cardiopulmonary arrest between May 1, 2008 and June 30, 2014 at an academic medical center was performed. The proportion of code status modifications to do not resuscitate (DNR) from full code was assessed. Baseline clinical characteristics, resuscitation factors, and 24-h post-resuscitation, hospital, and overall survival rates were compared between the two subsets.ResultsA total of 157 patients survived the index event and were included. One hundred and fifteen (73.2%) patients did not have a change in code status following the index event, while 42 (26.8%) changed code status to DNR. Clinical characteristics were similar between subsets, although patients in the change to DNR subset were older (average age 67.7 years) compared to the full code subset (average age 59.2 years; p = 0.005). Patients in the DNR subset had longer overall resuscitation efforts with less attempts at defibrillation. Compared to the DNR subset, patients that remained full code demonstrated higher 24-h post-resuscitation (n = 108, 93.9% versus n = 32, 76.2%; p = 0.001) and hospital (n = 50, 43.5% versus n = 6, 14.3%; p = 0.001) survival rates. Patients in the DNR subset were more likely to have neurologic deficits on discharge and shorter overall survival.ConclusionsPatient code status wishes do tend to change during critical periods within a hospitalization, adding emphasis for continued code status evaluation.     

Multi-kernel graph embedding for detection,Gleason grading of prostate cancer via MRI/MRS

Even though 1 in 6 men in the US, in their lifetime are expected to be diagnosed with prostate cancer (CaP), only 1 in 37 is expected to die on account of it. Consequently, among many men diagnosed with CaP, there has been a recent trend to resort to active surveillance (wait and watch) if diagnosed with a lower Gleason score on biopsy, as opposed to seeking immediate treatment. Some researchers have recently identified imaging markers for low and high grade CaP on multi-parametric (MP) magnetic resonance (MR) imaging (such as T2 weighted MR imaging (T2w MRI) and MR spectroscopy (MRS)). In this paper, we present a novel computerized decision support system (DSS), called Semi Supervised Multi Kernel Graph Embedding (SeSMiK-GE), that quantitatively combines structural, and metabolic imaging data for distinguishing (a) benign versus cancerous, and (b) high- versus low-Gleason grade CaP regions from in vivo MP-MRI. A total of 29 1.5 Tesla endorectal pre-operative in vivo MP MRI (T2w MRI, MRS) studies from patients undergoing radical prostatectomy were considered in this study. Ground truth for evaluation of the SeSMiK-GE classifier was obtained via annotation of disease extent on the pre-operative imaging by visually correlating the MRI to the ex vivo whole mount histologic specimens. The SeSMiK-GE framework comprises of three main modules: (1) multi-kernel learning, (2) semi-supervised learning, and (3) dimensionality reduction, which are leveraged for the construction of an integrated low dimensional representation of the different imaging and non-imaging MRI protocols. Hierarchical classifiers for diagnosis and Gleason grading of CaP are then constructed within this unified low dimensional representation. Step 1 of the hierarchical classifier employs a random forest classifier in conjunction with the SeSMiK-GE based data representation and a probabilistic pairwise Markov Random Field algorithm (which allows for imposition of local spatial constraints) to yield a voxel based classification of CaP presence. The CaP region of interest identified in Step 1 is then subsequently classified as either high or low Gleason grade CaP in Step 2. Comparing SeSMiK-GE with unimodal T2w MRI, MRS classifiers and a commonly used feature concatenation (COD) strategy, yielded areas (AUC) under the receiver operative curve (ROC) of (a) 0.89 ± 0.09 (SeSMiK), 0.54 ± 0.18 (T2w MRI), 0.61 ± 0.20 (MRS), and 0.64 ± 0.23 (COD) for distinguishing benign from CaP regions, and (b) 0.84 ± 0.07 (SeSMiK),0.54 ± 0.13 (MRI), 0.59 ± 0.19 (MRS), and 0.62 ± 0.18 (COD) for distinguishing high and low grade CaP using a leave one out cross-validation strategy, all evaluations being performed on a per voxel basis. Our results suggest that following further rigorous validation, SeSMiK-GE could be developed into a powerful diagnostic and prognostic tool for detection and grading of CaP in vivo and in helping to determine the appropriate treatment option. Identifying low grade disease in vivo might allow CaP patients to opt for active surveillance rather than immediately opt for aggressive therapy such as radical prostatectomy.     

Effects of a nurse-led transitional care program on clinical outcomes,health-related knowledge,physical and mental health status among Chinese patients with coronary artery disease: A randomized controlled trial

BackgroundCoronary artery disease is a major cause of morbidity and mortality among adults worldwide, including China. After a hospital stay, transitional care could help to ensure improved patient care and outcomes, and reduce Medicare costs. Nevertheless, the results of the existing transitional care are not always satisfactory and our knowledge of how to perform effective transitional care for patients with coronary artery disease is limited in mainland China.ObjectivesTo examine the effectiveness of a nurse-led transitional care program on clinical outcomes, health-related knowledge, and physical and mental health status among Chinese patients with coronary artery disease.DesignRandomized controlled trial.MethodsThe Omaha system and Pender’s health promoting model were employed in planning and implementing this nurse-led transitional care program. The sample was comprised of 199 Chinese patients with coronary artery disease. The experimental group (n = 100) received nurse-led transitional care intervention in addition to routine care. The nurse-led transitional care intervention included a structured assessment and health education, followed by 7 months of individual teaching and coaching (home visits, telephone follow-up and group activity). The control group (n = 99) received a comparable length routine care and follow-up contacts. Evaluations were conducted at baseline and completion of the interventions using the perceived knowledge scale for coronary heart disease, the medical outcomes study 36-item short-form health survey and clinical measures (blood pressure, blood glucose, lipids, body mass index). Data were collected between March and October 2014.ResultsCompared with the control group, participants in the experimental group showed significant better clinical outcomes (systolic blood pressure, t = 5.762, P = 0.000; diastolic blood pressure, t = 4.250, P = 0.000; fasting blood glucose, t = 2.249, P = 0.027; total cholesterol, t = 4.362, P = 0.000; triglyceride, t = 3.147, P = 0.002; low density lipoprotein cholesterol, t = 2.399, P = 0.018; and body mass index, t = 3.166, P = 0.002), higher knowledge scores for coronary artery disease (total knowledge score, t = −7.099, P = 0.000), better physical health status (t = −2.503, P = 0.014) and mental health status (t = −2.950, P = 0.004).ConclusionsThis study provides evidence for the value of a nurse-led transitional care program using both the Omaha system and Pender’s health promoting model as its theoretical framework. The structured interventions in this nurse-led transitional care program facilitate the use of this program in other settings.     

Apolipoprotein M promoter polymorphisms alter promoter activity and confer the susceptibility to the development of type 1 diabetes

ObjectivesApolipoprotein M plays an important role in the formation of preβ-HDL and cholesterol efflux to HDL. In the present study, we investigate the potential association between the ApoM promoter polymorphisms and type 1 diabetes.Design and methodsThe study was conducted in Peking Union Medical College, Beijing, China and Karolinska Institutet, Stockholm, Sweden. Two populations, including 493 Han Chinese subjects (177 T1D patients/316 controls) and 225 Swedish (124/101), are enrolled in the present study. Three single nucleotide polymorphisms (SNP) C-1065A, T-855C and T-778C in the promoter region of the ApoM gene are genotyped using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) protocol. Promoter activity was measured by reporter gene assay.ResultsSNP T-778C was strongly associated with T1D in both Han Chinese (p = 0.002, OR = 2.188, CI 95% = 1.338–3.581) and Swedish (p = 0.021, OR = 2.865, CI 95% = 1.128–7.278) populations. The luciferase activity of ?778C promoter was 1.41 times as high as that of ?778T promoter (9.90 ± 1.92 vs. 7.04 ± 0.76, p = 0.001).ConclusionsAllele C of SNP T-778C may increase promoter activity and confer the risk susceptibility to the development of T1D.     

A promoter polymorphism − 2122C>T of CHI3L1 is associated with serum low density lipoprotein cholesterol level in Korean subjects

ObjectivesThis study assessed the effects of 10 tagging single nucleotide polymorphisms (SNPs) of the CHI3L1 gene on serum LDL cholesterol levels in 290 Korean subjects.Design and methodsGenotyping analyses of SNPs were conducted by TaqMan® method. The effects of the promoter SNP on mRNA expression and nuclear factor binding were measured by real-time PCR and electrophoretic mobility shift assay, respectively.ResultsAmong 10 tagging SNPs, ? 2122C>T SNP (rs946261) in the promoter region was significantly associated with serum LDL cholesterol level (P = 0.005). The T allele of ? 2122C>T was associated with significantly increased mRNA expressions in peripheral blood cells of the subjects, and also increased a nuclear factor binding measured by an electrophoretic mobility shift assay.Conclusions? 2122C>T of CHI3L1, a promoter SNP which affects the mRNA expression and nuclear factor binding, is significantly associated with serum LDL cholesterol levels in Korean subjects.     

Impacto de una intervención educacional sobre la calidad de la dieta mediterránea,la actividad física y el estado ponderal en adolescentes: ensayo clínico aleatorizado

ObjectiveTo analyse the impact of an educational intervention on the quality of the Mediterranean diet, physical activity and weight status in adolescents.MethodRandomised clinical trial (RCT), controlled with a multimodal educational intervention (control group [n = 36] and experimental group [n = 46]). Data collection at the beginning and end of the study, in teenagers from Cáceres, Spain. In both groups anthropometric measurements and sociodemographic data were determined. The quality of the Mediterranean diet was assessed through the KIDMED test, the degree of physical activity through the PAQ-A questionnaire and weight status with the growth charts of the Faustino Orbegozo Eizaguirre Foundation.ResultsWe obtained a significant increase in the experimental group in the PAQ-A questionnaire that assesses physical activity (P = .029). No significant differences were observed between groups in the weight status (P = .916). When comparing the quality of the Mediterranean diet (high vs moderate or low quality) with physical activity (P control = .730; experimental P = .495) and with weight status (P control = .838; experimental P = .372), No significant differences are observed.ConclusionsThe educational intervention did not improve the quality of the Mediterranean diet or physical activity, although most of our sample had normal weight and acceptable physical activity. We must continue to improve the healthy eating pattern of our adolescents, to ensure an adequate state of health in the future.     

Exposure to parental verbal abuse is associated with increased gray matter volume in superior temporal gyrus

ObjectiveExposure to parental verbal aggression (PVA) during childhood increases risk for the development of psychopathology, particularly mood and anxiety disorders. Other forms of childhood abuse have been found to be associated with alterations in brain structure. The aim of this study was to ascertain whether exposure to PVA was associated with discernible effects on brain morphology.MethodsOptimized voxel-based morphometry was performed on 21 unmedicated, right-handed subjects (18–25 years) with histories of PVA and 19 psychiatrically healthy controls of comparable age and gender. Group differences in gray matter volume (GMV) – covaried by age, gender, parental education, financial stress, and total GMV – were assessed using high-resolution, T1-weighted, volumetric MRI data sets (Siemens 3T trio scanner).ResultsGMV was increased by 14.1% in the left superior temporal gyrus (STG, BA 22) (P = 0.004, corrected cluster level). GMV in this cluster was associated most strongly with levels of maternal (ß = 0.544, P < 0.0001) and paternal (ß = 0.300, P < 0.02) verbal aggression and inversely associated with parental education (ß = ? 0.577, P < 0.0001).ConclusionPrevious studies have demonstrated an increase in STG GMV in children with abuse histories, and found a reduction in fractional anisotropy in the arcuate fasciculus connecting Wernicke's and frontal areas in young adults exposed to PVA. These findings and the present results suggest that the development of auditory association cortex involved in language processing may be affected by exposure to early stress and/or emotionally abusive language.     

Improved measurement of LINE-1 sequence methylation for cancer detection

BackgroundMethylation of long interspersed nuclear element-1 (LINE-1) sequences varies among normal cells and it is often decreased in cancer genomes and white blood cells (WBC) of cancer patients. Current measurement techniques of genome-wide level are inadequate because LINE-1 methylation is distinctive at each locus. Here, we improved the detection of cancer by combining information of LINE-1 methylation pattern and level.MethodsCombined bisulfite restriction analysis (COBRA) of LINE-1, COBRA LINE-1, was used to test cancer cell lines, two oral rinse cohorts, and WBC from normal and cancer patients. COBRA LINE-1 separated LINE-1 sequences into 4 products depending on the methylation statuses of 2 CpG dinucleotides, as follows: 2 unmethylated CpGs (^uC^uC), partial methylation (^mC^uC), 1 methylated CpG (^mC), and 1 unmethylated CpG (^uC).ResultsThe association between ^mC^uC and ^uC^uC was directly correlated in normal cells (r = 0.4895, p = 0.0009) but inversely correlated in cancer (r = ? 0.8979, p = 0.0002). Oral rinse AUC values of ^uC^uC were 0.763 and 0.926 and methylation levels were 0.707 and 0.621, respectively. ^uC^uC, but not overall methylation level, differentiated cancer WBC from normal (p = 0.0082 and p = 0.4830, respectively).ConclusionLINE-1 partial methylation represents hypomethylation in normal cells but hypermethylation in cancer cells. This information improves LINE-1 methylation detection in cancer.     

HDL 2 Particles are associated with hyperglycaemia,lower PON1 activity and oxidative stress in type 2 diabetes mellitus patients

ObjectivesIn this study we examined the relationship of oxidative stress and hyperglycaemia to antioxidative capacity of high-density cholesterol (HDL-C) particles in type 2 diabetes mellitus (DM).Design and methodsOxidative stress status parameters (superoxide anion (O₂^?), superoxide dismutase (SOD) activity and paraoxonase (PON1) status were assessed in 114 patients with type 2 DM and 91 healthy subjects. HDL particle diameters were determined by non-denaturing polyacrylamide gradient (3–31%) gel electrophoresis.ResultsPatients had significantly higher concentrations of oxidative stress parameter O₂^?(p < 0.001) and antioxidative defence, SOD activity (p < 0.001). Paraoxonase activity was significantly lower in diabetics (p < 0.001). The PON1₁₉₂ phenotype distribution among study groups was not significantly different. HDL 3 phenotype was significantly prevalent among patients (p < 0.001). Paraoxonase activity was significantly lower in patients with predominantly HDL 2 particles than in controls.ConclusionsThe results of our current study indicate that the diabetic HDL 2 phenotype is associated with hyperglycaemia, lower PON1 activity and elevated oxidative stress.     

Intraoperative dynamic susceptibility contrast MRI (iDSC-MRI) is as reliable as preoperatively acquired perfusion mapping

DSC-MRI was applied intraoperatively during human brain tumor removal. Immediately after complete tumor resection was presumed, MRI including a dynamic susceptibility contrast T2?-weighted EPI sequence was performed in 30 patients while the skull was still open using a flexible two-channel coil system at an intraoperative 1.5-Tesla MR scanner. Maps of relative regional blood flow (rCBF), blood volume (rCBV), and mean transit time (MTT) were calculated, and ratios of these maps were compared to preoperatively acquired DSC-MRI data. The extent of the resection was compared with the postoperative MRI performed 24 h after the operation. In 8 of these patients residual tumor tissue was depicted at the time of intraoperative MRI. In corresponding regions ratios for rCBV and rCBF did not differ significantly between pre- and intraoperatively acquired data (two-tailed t-test). Furthermore, we found a high correlation between ratios created from pre- and intraoperatively measured data for both rCBV and rCBF, respectively (Pearson correlation; r²_rCBV = 0.86, p ≤ 0.01; r²_rCBF = 0.86, p ≤ 0.01). DSC-MRI is a powerful tool for the differential diagnosis of brain lesions. Its use intraoperatively provides pathophysiologic information that is up-to-date, independently of an intraoperative brain shift and also independently of the known leakage phenomenon caused by surgical manipulation. It can assist in the decision to depict residual tumor burden beyond conventional imaging. Our data demonstrate that iDSC-MRI is as reliable as preoperatively acquired data.     

Automatic MRI segmentation and morphoanatomy analysis of the vestibular system in adolescent idiopathic scoliosis

The vestibular system is the sensory organ responsible for perceiving head rotational movements and maintaining postural balance of human body. The objectives of this study are to propose an innovative computational technique capable of automatically segmenting the vestibular system and to analyze its geometrical features from high resolution T2-weighted MR images. In this study, the proposed technique was used to test the hypothesis that the morphoanatomy of vestibular system in adolescent idiopathic scoliosis (AIS) patients is different from healthy control subjects. The findings could contribute significantly to the understanding of the etiopathogenesis of AIS. The segmentation pipeline consisted of extraction of region of interest, image pre-processing, K-means clustering, and surface smoothing. The geometry of this high-genus labyrinth structure was analyzed through automatic partition into genus-0 units and approximation using the best-fit circle and plane for each unit. The metrics of the best-fit planes and circles were taken as shape measures. The proposed technique was applied on a cohort of 20 right-thoracic AIS patients (mean age 14.7 years old) and 20 age-matched healthy girls. The intermediate results were validated by subjective scoring. The result showed that the distance between centers of lateral and superior canals and the angle with vertex at the center of posterior canal were significantly smaller in AIS than in healthy controls in the left-side vestibular system with p = 0.0264 and p = 0.0200 respectively, but not in the right-side counterparts. The detected morphoanatomical changes are likely to be associated with subclinical postural, vestibular and proprioceptive dysfunctions reported frequently in AIS. This study has demonstrated that the proposed method could be applied in MRI-based morphoanatomy studies of vestibular system clinically.