Advanced imaging reveals enhanced malignancy in glioblastomas involving the subventricular zone: evidence of increased infiltrative growth and perfusion.

BACKGROUND: Glioblastoma's infiltrative growth and heterogeneity are influenced by neural, molecular, genetic, and immunological factors, with the precise origin of these tumors remaining elusive. Neurogenic zones might serve as the tumor stem cells' nest, with tumors in contact with these zones exhibiting worse outcomes and more aggressive growth patterns. This study aimed to determine if these characteristics are reflected in advanced imaging, specifically diffusion and perfusion data. METHODS: In this monocentric retrospective study, 137 glioblastoma therapy-naive patients (IDH-wildtype, grade 4) with advanced preoperative MRI, including perfusion and diffusion imaging, were analyzed. Tumors and neurogenic zones were automatically segmented. Advanced imaging metrics, including cerebral blood volume (CBV) from perfusion imaging, tissue volume mask (TVM), and free water corrected fractional anisotropy (FA-FWE) from diffusion imaging, were extracted. RESULTS: SVZ infiltration positively correlated with CBV, indicating higher perfusion in tumors. Significant CBV differences were noted between high and low SVZ infiltration cases at specific percentiles. Negative correlation was observed with TVM and positive correlation with FA-FWE, suggesting more infiltrative tumor growth. Significant differences in TVM and FA-FWE values were found between high and low SVZ infiltration cases. DISCUSSION: Glioblastomas with SVZ infiltration exhibit distinct imaging characteristics, including higher perfusion and lower cell density per voxel, indicating a more infiltrative growth and higher vascularization. Stem cell-like characteristics in SVZ-infiltrating cells could explain the increased infiltration and aggressive behavior. Understanding these imaging and biological correlations could enhance the understanding of glioblastoma evolution.

Exploring molecular glioblastoma: Insights from advanced imaging for a nuanced understanding of the molecularly defined malignant biology.

Background: Molecular glioblastoma (molGB) does not exhibit the histologic hallmarks of a grade 4 glioma but is nevertheless diagnosed as glioblastoma when harboring specific molecular markers. MolGB can easily be mistaken for similar-appearing lower-grade astrocytomas. Here, we investigated how advanced imaging could reflect the underlying tumor biology. Methods: Clinical and imaging data were collected for 7 molGB grade 4, 9 astrocytomas grade 2, and 12 astrocytomas grade 3. Four neuroradiologists performed VASARI-scoring of conventional imaging, and their inter-reader agreement was assessed using Fleiss κ coefficient. To evaluate the potential of advanced imaging, 2-sample t test, 1-way ANOVA, Mann-Whitney U, and Kruskal-Wallis test were performed to test for significant differences between apparent diffusion coefficient (ADC) and relative cerebral blood volume (rCBV) that were extracted fully automatically from the whole tumor volume. Results: While conventional VASARI imaging features did not allow for reliable differentiation between glioma entities, rCBV was significantly higher in molGB compared to astrocytomas for the 5th and 95th percentile, mean, and median values (P < .05). ADC values were significantly lower in molGB than in astrocytomas for mean, median, and the 95th percentile (P < .05). Although no molGB showed contrast enhancement initially, we observed enhancement in the short-term follow-up of 1 patient. Discussion: Quantitative analysis of diffusion and perfusion parameters shows potential in reflecting the malignant tumor biology of molGB. It may increase awareness of molGB in a nonenhancing, "benign" appearing tumor. Our results support the emerging hypothesis that molGB might present glioblastoma captured at an early stage of gliomagenesis.

Epigenome-850K-wide profiling reveals peripheral blood differential methylation in term low birth weight.

Aim: We investigate the genome-wide DNA methylation (DNAm) patterns of term low birth weight (TLBW) neonates.Methods: In the discovery phase, we assayed 32 samples (TLBW/control:16/16) using the EPIC 850k BeadChip Array. Targeted pyrosequencing of in 60 samples (TLBW/control:28/32) using targeted pyrosequencing during the replication phase.Results: The 850K array identified TLBW-associated 144 differentially methylated positions (DMPs) and 149 DMRs. Nearly 77% DMPs exhibited hypomethylation, located in the opensea and gene body regions. The most significantly enriched pathway in KEGG is sphingolipid metabolism (hsa00600), and the genes GALC and SGMS1 related to this pathway both show hypomethylation.Conclusion: Our analysis provides evidence of genome-wide DNAm alterations in TLBW. Further investigations are needed to elucidate the functional significance of these DNAm changes.

This study looked at the DNA of babies born after 37 weeks of pregnancy but weighing less than 2500 grams. We found that these babies had lower levels of DNA methylation, which might change how their bodies handle fats.

TCERG1L hypermethylation is a risk factor of diabetic retinopathy in Chinese children with type 1 diabetes.

AIM: To identify the differential methylation sites (DMS) and their according genes associated with diabetic retinopathy (DR) development in type 1 diabetes (T1DM) children. METHODS: This study consists of two surveys. A total of 40 T1DM children was included in the first survey. Because no participant has DR, retina thinning was used as a surrogate indicator for DR. The lowest 25% participants with the thinnest macular retinal thickness were included into the case group, and the others were controls. The DNA methylation status was assessed by the Illumina methylation 850K array BeadChip assay, and compared between the case and control groups. Four DMS with a potential role in diabetes were identified. The second survey included 27 T1DM children, among which four had DR. The methylation patterns of the four DMS identified by 850K were compared between participants with and without DR by pyrosequencing. RESULTS: In the first survey, the 850K array revealed 751 sites significantly and differentially methylated in the case group comparing with the controls (|Δß|>0.1 and Adj.P<0.05), and 328 of these were identified with a significance of Adj.P<0.01. Among these, 319 CpG sites were hypermethylated and 432 were hypomethylated in the case group relative to the controls. Pyrosequencing revealed that the transcription elongation regulator 1 like (TCERG1L, cg07684215) gene was hypermethylated in the four T1DM children with DR (P=0.018), which was consistent with the result from the first survey. The methylation status of the other three DMS (cg26389052, cg25192647, and cg05413694) showed no difference (all P>0.05) between participants with and without DR. CONCLUSION: The hypermethylation of the TCERG1L gene is a risk factor for DR development in Chinese children with T1DM.

Comparative epigenomics indicate a common origin of ectopic and intrasellar corticotroph pituitary neuroendocrine tumors/adenomas: a case report.

Ectopic pituitary neuroendocrine tumors (PitNET)/adenomas are rare and diagnostically challenging extra-sellar tumors. Previous studies have demonstrated the impact of epigenomic analyses in the diagnostics of sellar neoplasms and characterized the close relationship of epigenomic signatures and cellular origins of PitNET/adenomas. As of today, little is known about the pathogenesis of ectopic PitNET/adenomas, and epigenomic analyses have not been performed in these rare tumors. We report on the clinical course of an 81-year-old patient with sphenoid ectopic sparsely granulated corticotroph PitNET/adenoma and deploy genome-wide DNA methylation analysis to compare its methylation profile to a reference cohort of sellar neoplasms. Genome-wide methylation analysis revealed an epigenomic profile analogous to reference sellar corticotroph PitNET/adenomas, and the copy number variation profile showed loss of chromosomes 18 and 22. The methylation profile shows concordance with sellar corticotroph PitNET/adenomas suggesting a common cellular origin and confirming the reliability of methylation analyses as a diagnostic method in these rare tumors. This is the first data suggesting that epigenetic profiles of ectopic PitNET/adenoma do not differ from their sellar counterparts.

Genome-wide DNA Methylation Profiling in Lyme Neuroborreliosis Reveals Altered Methylation Patterns of HLA Genes.

Lyme neuroborreliosis (LNB) is a complex neuroinflammatory disorder caused by Borrelia burgdorferi, which is transmitted through tick bites. Epigenetic alterations, specifically DNA methylation (DNAm), could play a role in the host immune response during infection. In this study, we present the first genome-wide analysis of DNAm in peripheral blood mononuclear cells from patients with LNB and those without LNB. Using a network-based approach, we highlighted HLA genes at the core of these DNAm changes, which were found to be enriched in immune-related pathways. These findings shed light on the role of epigenetic modifications in the LNB pathogenesis that should be confirmed and further expanded upon in future studies.

Preliminary study on whole genome methylation and transcriptomics in age-related cataracts.

DNA methylation plays an important role in the occurrence and development of age-related cataracts (ARC). This study aims to reveal potential epigenetic biomarkers of ARC by detecting modifications to the DNA methylation patterns of genes shown to be related to ARC by transcriptomics. The MethylationEPIC BeadChip (850 K) was used to analyze the DNA methylation levels in ARC patients and unaffected controls, and the Pearson correlation test was used to perform genome-wide integration analysis of DNA methylation and transcriptome data. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases were used to perform functional analysis of the whole genome, promoter regions (TSS1500/TSS200), and the associated differentially methylated genes (DMG). Pyrosequencing was used to verify the methylation levels of the selected genes. The results showed that, compared with the control group, a total of 52,705 differentially methylated sites were detected in the ARC group, of which 13,858 were hypermethylated and 38,847 were hypomethylated. GO and KEGG analyses identified functions related to the cell membrane, the calcium signaling pathway, and their possible molecular mechanisms. Then, 57 DMGs with negative promoter methylation correlations were screened by association analysis. Pyrosequencing verified that the ARC group had higher methylation levels of C3 and CCKAR and lower methylation levels of NLRP3, LEFTY1, and GPR35 compared with the control group. In summary, our study reveals the whole-genome DNA methylation patterns and gene expression profiles in ARC, and the molecular markers of methylation identified herein may aid in the prevention, diagnosis, treatment, and prognosis of ARC.

HepaClear, a blood-based panel combining novel methylated CpG sites and protein markers, for the detection of early-stage hepatocellular carcinoma.

BACKGROUND: Early screening and detection of hepatocellular carcinoma (HCC) can efficiently improve patient prognosis. We aimed to identify a series of hypermethylated DNA markers and develop a blood-based HCC diagnosis panel containing DNA methylation sites and protein markers with improved sensitivity for early-stage HCC detection. RESULTS: Overall, 850K methylation arrays were performed using paired tissue DNA samples from 60 HCC patients. Ten candidate hypermethylated CpG sites were selected for further evaluation by quantitative methylation-specific PCR with 60 pairs of tissue samples. Six methylated CpG sites, along with α-fetoprotein (AFP) and des-gamma-carboxyprothrombin (DCP), were assayed in 150 plasma samples. Finally, an HCC diagnosis panel, named HepaClear, was developed in a cohort consisting of 296 plasma samples and validated in an independent cohort consisting of 198 plasma samples. The HepaClear panel, containing 3 hypermethylated CpG sites (cg14263942, cg12701184, and cg14570307) and 2 protein markers (AFP and DCP), yielded a sensitivity of 82.6% and a specificity of 96.2% in the training set and a sensitivity of 84.7% and a specificity of 92.0% in the validation set. The HepaClear panel had higher sensitivity (72.0%) for early-stage HCC than AFP (≥ 20 ng/mL, 48.0%) and DCP (≥ 40 mAU/mL, 62.0%) and detected 67.5% of AFP-negative HCC patients (AFP ≤ 20 ng/mL). CONCLUSIONS: We developed a multimarker HCC detection panel (HepaClear) that shows high sensitivity for early-stage HCC. The HepaClear panel exhibits high potential for HCC screening and diagnosis from an at-risk population.

Stable methylation loci are associated with systolic blood pressure in a Croatian island population.

Background: The objective was to identify stable and dynamic DNA methylation loci associated with cardiometabolic traits among an adult population from the Croatian island of Hvar. Materials & methods: An epigenome-wide association study was conducted using peripheral blood longitudinally collected at two time points 10 years apart via Infinium MethylationEPIC beadarray (n = 112). Stable and dynamic loci were identified using linear mixed models. Associations between cardiometabolic traits and loci were assessed using linear models. Results: 22 CpG loci were significantly associated with systolic blood pressure. Twenty were stable and two were dynamic. Conclusion: Multiple genes may be involved in the determination of systolic blood pressure level via stable epigenetic programming, potentially established earlier in life.

Cardiovascular disease is the leading cause of death worldwide. Previous studies have found that genetics incompletely explain susceptibility to cardiovascular disease. To find new potential risk factors, the authors investigated the possible contribution of DNA methylation (modifications to DNA that can affect gene expression but do not alter the underlying genetic code) in an adult population on the Croatian island of Hvar, which has a high number of people with cardiovascular and metabolic disease. By examining DNA methylation in blood collected at two time points, 10 years apart, the authors were able to identify DNA methylation that either stayed the same over time (stable) or changed the most over time (dynamic). These were then compared with clinical test results related to cardiovascular or metabolic diseases to determine if they are associated. Twenty-two methylation sites were found to be associated with systolic blood pressure. Of those, 20 were considered stable and two were dynamic. Additionally, there was one stable methylation site associated with serum calcium and one with C-reactive protein. These findings suggest that systolic blood pressure may be regulated through stable DNA methylation that is potentially established earlier in life.

Improving Infinium MethylationEPIC data processing: re-annotation of enhancers and long noncoding RNA genes and benchmarking of normalization methods.

Illumina Infinium DNA Methylation (5mC) arrays are a popular technology for low-cost, high-throughput, genome-scale measurement of 5mC distribution, especially in cancer and other complex diseases. After the success of its HumanMethylation450 array (450k), Illumina released the MethylationEPIC array (850k) featuring increased coverage of enhancers. Despite the widespread use of 850k, analysis of the corresponding data remains suboptimal: it still relies mostly on Illumina's default annotation, which underestimates enhancerss and long noncoding RNAs. Results: We have thus developed an approach, based on the ENCODE and LNCipedia databases, which greatly improves upon Illumina's default annotation of enhancers and long noncoding transcripts. We compared the re-annotated 850k with both 450k and reduced-representation bisulphite sequencing (RRBS), another high-throughput 5mC profiling technology. We found 850k to cover at least three times as many enhancers and long noncoding RNAs as either 450k or RRBS. We further investigated the reproducibility of the three technologies, applying various normalization methods to the 850k data. Most of these methods reduced variability to a level below that of RRBS data. We then used 850k with our new annotation and normalization to profile 5mC changes in breast cancer biopsies. 850k highlighted aberrant enhancer methylation as the predominant feature, in agreement with previous reports. Our study provides an updated processing approach for 850k data, based on refined probe annotation and normalization, allowing for improved analysis of methylation at enhancers and long noncoding RNA genes. Our findings will help to further advance understanding of the DNA methylome in health and disease.

Evaluation of cross-platform compatibility of a DNA methylation-based glucocorticoid response biomarker.

BACKGROUND: Identifying blood-based DNA methylation patterns is a minimally invasive way to detect biomarkers in predicting age, characteristics of certain diseases and conditions, as well as responses to immunotherapies. As microarray platforms continue to evolve and increase the scope of CpGs measured, new discoveries based on the most recent platform version and how they compare to available data from the previous versions of the platform are unknown. The neutrophil dexamethasone methylation index (NDMI 850) is a blood-based DNA methylation biomarker built on the Illumina MethylationEPIC (850K) array that measures epigenetic responses to dexamethasone (DEX), a synthetic glucocorticoid often administered for inflammation. Here, we compare the NDMI 850 to one we built using data from the Illumina Methylation 450K (NDMI 450). RESULTS: The NDMI 450 consisted of 22 loci, 15 of which were present on the NDMI 850. In adult whole blood samples, the linear composite scores from NDMI 450 and NDMI 850 were highly correlated and had equivalent predictive accuracy for detecting DEX exposure among adult glioma patients and non-glioma adult controls. However, the NDMI 450 scores of newborn cord blood were significantly lower than NDMI 850 in samples measured with both assays. CONCLUSIONS: We developed an algorithm that reproduces the DNA methylation glucocorticoid response score using 450K data, increasing the accessibility for researchers to assess this biomarker in archived or publicly available datasets that use the 450K version of the Illumina BeadChip array. However, the NDMI850 and NDMI450 do not give similar results in cord blood, and due to data availability limitations, results from sample types of newborn cord blood should be interpreted with care.

Genome-wide methylation profiling of early colorectal cancer using an Illumina Infinium Methylation EPIC BeadChip.

BACKGROUND: DNA methylation is a part of epigenetic modification, that is closely related to the growth and development of colorectal cancer (CRC). Specific methylated genes and methylated diagnostic models of tumors have become current research focuses. The methylation status of circulating DNA in plasma might serve as a potential biomarker for CRC. AIM: To investigate genome-wide methylation pattern in early CRC using the Illumina Infinium Human Methylation 850K BeadChip. METHODS: The 850K Methylation BeadChip was used to analyze the genome-wide methylation status of early CRC patients (n = 5) and colorectal adenoma patients (n = 5). Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways enrichment analyses were performed on the selected differentially methylated sites to further discover candidate methylation biomarkers in plasma. RESULTS: A total of 1865 methylated CpG sites with significant differences were detected, including 676 hypermethylated sites and 1189 hypomethylated sites. The distribution of these sites covered from the 1st to 22nd chromosomes and are mainly distributed on the gene body and gene promoter region. GO and KEGG enrichment analysis showed that the functions of these genes were related to biological regulation, molecular binding, transcription factor activity and signal transduction pathway. CONCLUSION: The study demonstrated that the Illumina Infinium Human Methylation 850K BeadChip can be used to investigate genome-wide methylation status of plasma DNA in early CRC and colorectal adenoma patients.

Epigenetic Signatures Discriminate Patients With Primary Sclerosing Cholangitis and Ulcerative Colitis From Patients With Ulcerative Colitis.

Background: Primary sclerosing cholangitis (PSC) is a chronic inflammatory liver disease affecting the intra- and extrahepatic bile ducts, and is strongly associated with ulcerative colitis (UC). In this study, we explored the peripheral blood DNA methylome and its immune cell composition in patients with PSC-UC, UC, and healthy controls (HC) with the aim to develop a predictive assay in distinguishing patients with PSC-UC from those with UC alone. Methods: The peripheral blood DNA methylome of male patients with PSC and concomitant UC, UC and HCs was profiled using the Illumina HumanMethylation Infinium EPIC BeadChip (850K) array. Differentially methylated CpG position (DMP) and region (DMR) analyses were performed alongside gradient boosting classification analyses to discern PSC-UC from UC patients. As observed differences in the DNA methylome could be the result of differences in cellular populations, we additionally employed mass cytometry (CyTOF) to characterize the immune cell compositions. Results: Genome wide methylation analysis did not reveal large differences between PSC-UC and UC patients nor HCs. Nonetheless, using gradient boosting we were capable of discerning PSC-UC from UC with an area under the receiver operator curve (AUROC) of 0.80. Four CpG sites annotated to the NINJ2 gene were found to strongly contribute to the predictive performance. While CyTOF analyses corroborated the largely similar blood cell composition among patients with PSC-UC, UC and HC, a higher abundance of myeloid cells was observed in UC compared to PSC-UC patients. Conclusion: DNA methylation enables discerning PSC-UC from UC patients, with a potential for biomarker development.

Preliminary Study on the Sequencing of Whole Genomic Methylation and Transcriptome-Related Genes in Thyroid Carcinoma.

Thyroid carcinoma is the most prevalent endocrine cancer globally and the primary cause of cancer-related mortality. Epigenetic modifications are progressively being linked to metastasis. This study aimed to examine whole-genome DNA methylation patterns and the gene expression profiles in thyroid cancer tissue samples using a MethylationEPIC BeadChip (850K), RNA sequencing, and a targeted bisulfite sequencing assay. The results of the Illumina Infinium human methylation kit (850K) analyses identified differentially methylated CpG locations (DMPs) and differentially methylated CpG regions (DMRs) encompassing nearly the entire genome with high resolution and depth. Gene ontology and KEGG pathway analyses revealed that the genes associated with DMRs belonged to various domain-specific ontologies, including cell adhesion, molecule binding, and proliferation. The RNA-Seq study found 1627 differentially expressed genes, 1174 of which that were up-regulated and 453 of which that were down-regulated. The targeted bisulfite sequencing assay revealed that CHST2, DPP4, DUSP6, ITGA2, SLC1A5, TIAM1, TNIK, and ABTB2 methylation levels were dramatically lowered in thyroid cancer patients when compared to the controls, but GALNTL6, HTR7, SPOCD1, and GRM5 methylation levels were significantly raised. Our study revealed that the whole-genome DNA methylation patterns and gene expression profiles in thyroid cancer shed new light on the tumorigenesis of thyroid cancer.

Association of environmental exposure to chromium with differential DNA methylation: An epigenome-wide study.

Introduction: Previous studies have reported that chromium (Cr)-induced epigenetic alterations and DNA methylation play a vital role in the pathogenesis of diseases induced by chromium exposure. Epigenomic analyses have been limited and mainly focused on occupational chromium exposure; their findings are not generalizable to populations with environmental Cr exposure. Methods: We identified the differential methylation of genes and regions to elucidate the mechanisms of toxicity related to environmental chromium exposure. DNA methylation was measured in blood samples collected from individuals in Cr-contaminated (n = 10) and unexposed areas (n = 10) by using the Illumina Infinium HumanMethylation850K array. To evaluate the relationship between chromium levels in urine and CpG methylation at 850 thousand sites, we investigated differentially methylated positions (DMPs) and differentially methylated regions (DMRs) by using linear models and DMRcate method, respectively. The model was adjusted for biologically relevant variables and estimated cell-type compositions. Results: At the epigenome-wide level, we identified five CpGs [cg20690919 (p FDR =0.006), cg00704664 (p FDR =0.024), cg10809143 (p FDR =0.043), cg27057652 (p FDR =0.047), cg05390480 (p FDR =0.024)] and one DMR (chr17: 19,648,718-19,648,972), annotated to ALDH3A1 genes (p < 0.05) as being significantly associated with log2 transformed urinary chromium levels. Discussion: Environmental chromium exposure is associated with DNA methylation, and the significant DMPs and DMR being annotated to cause DNA damage and genomic instability were found in this work. Research involving larger samples is required to further explore the epigenetic effect of environmental chromium exposure on health outcomes through DNA methylation.

Integrative Methylome and Transcriptome Characterization Identifies SERINC2 as a Tumor-Driven Gene for Papillary Thyroid Carcinoma.

Most papillary thyroid carcinomas (PTCs) can be diagnosed preoperatively by routine evaluation, such as thyroid ultrasonography and fine-needle aspiration biopsy. Nevertheless, understanding how to differentiate indolent thyroid tumors from aggressive thyroid cancers remains a challenge, which may cause overtreatment. This study aimed to identify papillary thyroid cancer-specific indicators with whole-genome DNA methylation and gene expression profiles utilizing Infinium Methylation EPIC BeadChip (850k) and RNA arrays. In this paper, we report SERINC2 as a potential tumor-driven indicator in PTC. The up-regulated expression levels of SERINC2 were verified in PTC cell lines via qPCR. Then, cell counting kit 8 (CCK-8), wound healing, and flow cytometric assays were performed to confirm the influence of SERINC2 on proliferation and apoptosis in PTC cell lines after intervention or overexpression. Moreover, the investigation of data from the Cancer Dependency Map (DepMap) provided a potential pathway targeted by SERINC2. The activation of the tryptophan metabolic pathway may reduce the dependency of SERINC2 in thyroid cancers. In conclusion, our results demonstrate the whole-genome DNA methylation and gene expression profiles of papillary thyroid carcinoma, identify SERINC2 as a potential tumor-driven biomarker, and preliminarily verify its function in PTC.

Combined prenatal Lactobacillus reuteri and ω-3 supplementation synergistically modulates DNA methylation in neonatal T helper cells.

BACKGROUND: Environmental exposures may alter DNA methylation patterns of T helper cells. As T helper cells are instrumental for allergy development, changes in methylation patterns may constitute a mechanism of action for allergy preventive interventions. While epigenetic effects of separate perinatal probiotic or ω-3 fatty acid supplementation have been studied previously, the combined treatment has not been assessed. We aimed to investigate epigenome-wide DNA methylation patterns from a sub-group of children in an on-going randomised double-blind placebo-controlled allergy prevention trial using pre- and postnatal combined Lactobacillus reuteri and ω-3 fatty acid treatment. To this end, > 866000 CpG sites (MethylationEPIC 850K array) in cord blood CD4+ T cells were examined in samples from all four study arms (double-treatment: n = 18, single treatments: probiotics n = 16, ω-3 n = 15, and double placebo: n = 14). Statistical and bioinformatic analyses identified treatment-associated differentially methylated CpGs and genes, which were used to identify putatively treatment-induced network modules. Pathway analyses inferred biological relevance, and comparisons were made to an independent allergy data set. RESULTS: Comparing the active treatments to the double placebo group, most differentially methylated CpGs and genes were hypermethylated, possibly suggesting induction of transcriptional inhibition. The double-treated group showed the largest number of differentially methylated CpGs, of which many were unique, suggesting synergy between interventions. Clusters within the double-treated network module consisted of immune-related pathways, including T cell receptor signalling, and antigen processing and presentation, with similar pathways revealed for the single-treatment modules. CpGs derived from differential methylation and network module analyses were enriched in an independent allergy data set, particularly in the double-treatment group, proposing treatment-induced DNA methylation changes as relevant for allergy development. CONCLUSION: Prenatal L. reuteri and/or ω-3 fatty acid treatment results in hypermethylation and affects immune- and allergy-related pathways in neonatal T helper cells, with potentially synergistic effects between the interventions and relevance for allergic disease. Further studies need to address these findings on a transcriptional level, and whether the results associate to allergy development in the children. Understanding the role of DNA methylation in regulating effects of perinatal probiotic and ω-3 interventions may provide essential knowledge in the development of efficacious allergy preventive strategies. Trial registration ClinicalTrials.gov, ClinicalTrials.gov-ID: NCT01542970. Registered 27th of February 2012-Retrospectively registered, https://clinicaltrials.gov/ct2/show/NCT01542970 .

Transcriptome and Differential Methylation Integration Analysis Identified Important Differential Methylation Annotation Genes and Functional Epigenetic Modules Related to Vitiligo.

Vitiligo is an pigmentation disorder caused by a variety of pathogenic factors; its main pathophysiological conditions include oxidative stress, immune activation, and genetic background. Additionally, DNA methylation is often associated with the pathogenesis of vitiligo; however, the underlying mechanism remains unknown. In the present study, we used the Human Methylation 850K BeadChip platform to detect DNA methylation changes in the vitiligo melanocytes. We then integrated the results with the transcriptome data of vitiligo melanocytes and lesions to analyse the correlation between differentially methylated levels and differentially expressed genes. The results showed that there was a significant negative correlation between methylation levels and differentially expressed genes. Subsequently, we enriched GO and KEGG based on methylated differentially expressed genes (MDEGs) using R package ClusterProfiler, and the results were closely related to the pathogenesis of vitiligo. In addition, we also constructed a PPI network of MDEGs and excavated three important functional epigenetic modules, involving a total of 12 (BCL2L1, CDK1, ECT2, HELLS, HSP90AA1, KIF23, MC1R, MLANA, PBK, PTGS2, SOX10, and TYRP1) genes. These genes affect melanocyte melanogenesis, cellular oxidative stress and other important biological processes. Our comprehensive analysis results support the significant contribution of the status of DNA methylation modification to vitiligo, which will help us to better understand the molecular mechanism of vitiligo and explore new therapeutic strategies.

Genome-wide DNA methylation profiling is able to identify prefibrotic PMF cases at risk for progression to myelofibrosis.

BACKGROUND: Patients suffering from the BCR-ABL1-negative myeloproliferative disease prefibrotic primary myelofibrosis (pre-PMF) have a certain risk for progression to myelofibrosis. Accurate risk estimation for this fibrotic progression is of prognostic importance and clinically relevant. Commonly applied risk scores are based on clinical, cytogenetic, and genetic data but do not include epigenetic modifications. Therefore, we evaluated the assessment of genome-wide DNA methylation patterns for their ability to predict fibrotic progression in PMF patients. RESULTS: For this purpose, the DNA methylation profile was analyzed genome-wide in a training set of 22 bone marrow trephines from patients with either fibrotic progression (n = 12) or stable disease over several years (n = 10) using the 850 k EPIC array from Illumina. The DNA methylation classifier constructed from this data set was validated in an independently measured test set of additional 11 bone marrow trephines (7 with stable disease, 4 with fibrotic progress). Hierarchical clustering of methylation ß-values and linear discriminant classification yielded very good discrimination between both patient groups. By gene ontology analysis, the most differentially methylated CpG sites are primarily associated with genes involved in cell-cell and cell-matrix interactions. CONCLUSIONS: In conclusion, we could show that genome-wide DNA methylation profiling of bone marrow trephines is feasible under routine diagnostic conditions and, more importantly, is able to predict fibrotic progression in pre-fibrotic primary myelofibrosis with high accuracy.

Genome-wide DNA methylation profile analysis in thoracic ossification of the ligamentum flavum.

Thoracic ossification of the ligamentum flavum (TOLF) causes serious spinal canal stenosis. The underlying aetiology may relate to genetic and inflammatory factors. DNA methylation plays a critical role in osteogenesis and inflammation, whereas there is no genome-wide DNA methylation analysis about TOLF. The two subtypes of TOLF (single-level and multiple-level) have distinct clinical features. Using micro-computed tomography (micro-CT), we showed the ossification arose from the joint between two vertebrae at one/both sides of ligament flavum. With Illumina Infinium Human Methylation 850 BeadChip arrays, genome-wide DNA methylation profile was measured in ligament flavum of eight healthy and eight TOLF samples. Only 65 of the differentially methylated cytosine-phosphate-guanine dinucleotides were found in both subtype groups. Principal component analysis and heat map analysis showed a different methylation pattern in TOLF samples, and methylation patterns of two subtypes are also distinct. The Gene Ontology enrichment analysis was significantly enriched in differentiation and inflammation. Pyrosequencing analysis and quantitative real-time polymerase chain reaction were performed to validate the arrays results and expression levels, to test six differentially methylated genes (SLC7A11, HOXA10, HOXA11AS, TNIK, homeobox transcript antisense RNA, IFITM1), using another independent samples (P < 0.05). Our findings first demonstrated an altered Genome-wide DNA methylation profile in TOLF, and implied distinct methylated features in two subtypes.