Aberrant PRDM2 methylation as an early event in serrated lesions destined to evolve into microsatellite-instable colorectal cancers.

Up to 30% of colorectal cancers (CRCs) develop from sessile serrated lesions (SSLs). Within the serrated neoplasia pathway, at least two principally distinct oncogenetic routes exist generating microsatellite-stable and microsatellite-instable CRCs, respectively. Aberrant DNA methylation (DNAm) is found early in the serrated pathway and might play a role in both oncogenetic routes. We studied a cohort of 23 SSLs with a small focus (<10 mm) of dysplasia or cancer, 10 of which were MLH1 deficient and 13 MLH1 proficient. By comparing, for each SSL, the methylation status of (1) the region of dysplasia or cancer (SSL-D), (2) the nondysplastic SSL (SSL), and (3) adjacent normal mucosa, differentially methylated probes (DMPs) and regions (DMRs) were assessed both genome-wide as well as in a tumor-suppressor gene-focused approach. By comparing DNAm of MLH1-deficient SSL-Ds with their corresponding SSLs, we identified five DMRs, including those annotating for PRDM2 and, not unexpectedly, MLH1. PRDM2 gene promotor methylation was associated with MLH1 expression status, as it was largely hypermethylated in MLH1-deficient SSL-Ds and hypomethylated in MLH1-proficient SSL-Ds. Significantly increased DNAm levels of PRDM2 and MLH1, in particular at 'critical' MLH1 probe sites, were to some extent already visible in SSLs as compared to normal mucosa (p = 0.02, p = 0.01, p < 0.0001, respectively). No DMRs, nor DMPs, were identified for SSLs destined to evolve into MLH1-proficient SSL-Ds. Our data indicate that, within both arms of the serrated CRC pathway, the majority of the epigenetic alterations are introduced early during SSL formation. Promoter hypermethylation of PRDM2 and MLH1 on the other hand specifically initiates in SSLs destined to transform into MLH1-deficient CRCs suggesting that the fate of SSLs may not necessarily result from a stochastic process but possibly is already imprinted and predisposed.

DNA methylation episignature, extension of the clinical features, and comparative epigenomic profiling of Hao-Fountain syndrome caused by variants in USP7.

PURPOSE: Hao-Fountain syndrome (HAFOUS) is a neurodevelopmental disorder caused by pathogenic variants in USP7. HAFOUS is characterized by developmental delay, intellectual disability, speech delay, behavioral abnormalities, autism spectrum disorder, seizures, hypogonadism, and mild dysmorphic features. We investigated the phenotype of 18 participants with HAFOUS and performed DNA methylation (DNAm) analysis, aiming to generate a diagnostic biomarker. Furthermore, we performed comparative analysis with known episignatures to gain more insight into the molecular pathophysiology of HAFOUS. METHODS: We assessed genomic DNAm profiles of 18 individuals with pathogenic variants and variants of uncertain significance (VUS) in USP7 to map and validate a specific episignature. The comparison between the USP7 cohort and 56 rare genetic disorders with earlier reported DNAm episignatures was performed with statistical and functional correlation. RESULTS: We mapped a sensitive and specific DNAm episignature for pathogenic variants in USP7 and utilized this to reclassify the VUS. Comparative epigenomic analysis showed evidence of HAFOUS similarity to a number of other rare genetic episignature disorders. CONCLUSION: We discovered a sensitive and specific DNAm episignature as a robust diagnostic biomarker for HAFOUS that enables VUS reclassification in USP7. We also expand the phenotypic spectrum of 9 new and 5 previously reported individuals with HAFOUS.

Long-term Temporal Stability of Peripheral Blood DNA Methylation Profiles in Patients With Inflammatory Bowel Disease.

BACKGROUND & AIMS: There is great current interest in the potential application of DNA methylation alterations in peripheral blood leukocytes (PBLs) as biomarkers of susceptibility, progression, and treatment response in inflammatory bowel disease (IBD). However, the intra-individual stability of PBL methylation in IBD has not been characterized. Here, we studied the long-term stability of all probes located on the Illumina HumanMethylation EPIC BeadChip array. METHODS: We followed a cohort of 46 adult patients with IBD (36 Crohn's disease [CD], 10 ulcerative colitis [UC]; median age, 44 years; interquartile range [IQR] 27-56 years; 50% female) that received standard care follow-up at the Amsterdam University Medical Centers. Paired PBL samples were collected at 2 time points with a median of 7 years (range, 2-9 years) in between. Differential methylation and intra-class correlation (ICC) analyses were used to identify time-associated differences and temporally stable CpGs, respectively. RESULTS: Around 60% of all EPIC array loci presented poor intra-individual stability (ICC <0.50); 78.114 (≈9%) showed good (ICC, 0.75-0.89), and 41.274 (≈5%) showed excellent (ICC ≥0.90) stability, between both measured time points. Focusing on previously identified consistently differentially methylated positions indicated that 22 CD-, 11 UC-, and 24 IBD-associated loci demonstrated high stability (ICC ≥0.75) over time; of these, we observed a marked stability of CpG loci associated to the HLA genes. CONCLUSIONS: Our data provide insight into the long-term stability of the PBL DNA methylome within an IBD context, facilitating the selection of biologically relevant and robust IBD-associated epigenetic biomarkers with increased potential for independent validation. These data also have potential implications in understanding disease pathogenesis.

Systematic Review and Meta-analysis of Peripheral Blood DNA Methylation Studies in Inflammatory Bowel Disease.

BACKGROUND AND AIMS: Over the past decade, the DNA methylome has been increasingly studied in peripheral blood of inflammatory bowel disease [IBD] patients. However, a comprehensive summary and meta-analysis of peripheral blood leukocyte [PBL] DNA methylation studies has thus far not been conducted. Here, we systematically reviewed all available literature up to February 2022 and summarized the observations by means of meta-analysis. METHODS: We conducted a systematic search and critical appraisal of IBD-associated DNA methylation studies in PBL using the biomarker-based cross-sectional studies [BIOCROSS] tool. Subsequently, we performed meta-analyses on the summary statistics obtained from epigenome-wide association studies [EWAS] that included patients with Crohn's disease [CD], ulcerative colitis [UC] and/or healthy controls [HC]. RESULTS: Altogether, we included 15 studies for systematic review. Critical appraisal revealed large methodological and outcome heterogeneity between studies. Summary statistics were obtained from four studies based on a cumulative 552 samples [177 CD, 132 UC and 243 HC]. Consistent differential methylation was identified for 256 differentially methylated probes [DMPs; Bonferroni-adjusted p ≤ 0.05] when comparing CD with HC and 103 when comparing UC with HC. Comparing IBD [CD + UC] with HC resulted in 224 DMPs. Importantly, several of the previously identified DMPs, such as VMP1/TMEM49/MIR21 and RPS6KA2, were consistently differentially methylated across all studies. CONCLUSION: Methodological homogenization of IBD epigenetic studies is needed to allow for easier aggregation and independent validation. Nonetheless, we were able to confirm previous observations. Our results can serve as the basis for future IBD epigenetic biomarker research in PBL.

Novel DNA methylome biomarkers associated with adalimumab response in rheumatoid arthritis patients.

Background and aims: Rheumatoid arthritis (RA) patients are currently treated with biological agents mostly aimed at cytokine blockade, such as tumor necrosis factor-alpha (TNFα). Currently, there are no biomarkers to predict therapy response to these agents. Here, we aimed to predict response to adalimumab (ADA) treatment in RA patients using DNA methylation in peripheral blood (PBL). Methods: DNA methylation profiling on whole peripheral blood from 92 RA patients before the start of ADA treatment was determined using Illumina HumanMethylationEPIC BeadChip array. After 6 months, treatment response was assessed according to the European Alliance of Associations for Rheumatology (EULAR) criteria for disease activity. Patients were classified as responders (Disease Activity Score in 28 Joints (DAS28) < 3.2 or decrease of 1.2 points) or as non-responders (DAS28 > 5.1 or decrease of less than 0.6 points). Machine learning models were built through stability-selected gradient boosting to predict response prior to ADA treatment with predictor DNA methylation markers. Results: Of the 94 RA patients, we classified 49 and 43 patients as responders and non-responders, respectively. We were capable of differentiating responders from non-responders with a high performance (area under the curve (AUC) 0.76) using a panel of 27 CpGs. These classifier CpGs are annotated to genes involved in immunological and pathophysiological pathways related to RA such as T-cell signaling, B-cell pathology, and angiogenesis. Conclusion: Our findings indicate that the DNA methylome of PBL provides discriminative capabilities in discerning responders and non-responders to ADA treatment and may therefore serve as a tool for therapy prediction.

Peripheral Blood DNA Methylation Profiles Do Not Predict Endoscopic Post-Operative Recurrence in Crohn's Disease Patients.

Prediction of endoscopic post-operative recurrence (POR) in Crohn's disease (CD) patients following ileocolonic resection (ICR) using clinical risk factors alone has thus far been inadequate. While peripheral blood leukocyte (PBL) DNA methylation has shown promise as a tool for predicting recurrence in cancer, no data in CD patients exists. Therefore, this study explored the association and predictive value of PBL DNA methylation in CD patients following ICR. From a cohort of 117 CD patients undergoing ICR, epigenome-wide PBL methylation profiles from 25 carefully selected patients presenting either clear endoscopic remission (n = 12) or severe recurrence (n = 13) were assessed using the Illumina MethylationEPIC (850K) array. No statistically significant differentially methylated positions (DMPs) or regions (DMRs) associated with endoscopic POR were identified (FDR p ≤ 0.05), further evidenced by the low accuracy (0.625) following elastic net classification analysis. Nonetheless, interrogating the most significant differences in methylation suggested POR-associated hypermethylation in the MBNL1, RAB29 and LEPR genes, respectively, which are involved in intestinal fibrosis, inflammation and wound healing. Notably, we observed a higher estimated proportion of monocytes in endoscopic POR compared to remission. Altogether, we observed limited differences in the genome-wide DNA methylome among CD patients with and without endoscopic POR. We therefore conclude that PBL DNA methylation is not a feasible predictive tool in post-operative CD.

Modulation of macrophage inflammatory function through selective inhibition of the epigenetic reader protein SP140.

BACKGROUND: SP140 is a bromodomain-containing protein expressed predominantly in immune cells. Genetic polymorphisms and epigenetic modifications in the SP140 locus have been linked to Crohn's disease (CD), suggesting a role in inflammation. RESULTS: We report the development of the first small molecule SP140 inhibitor (GSK761) and utilize this to elucidate SP140 function in macrophages. We show that SP140 is highly expressed in CD mucosal macrophages and in in vitro-generated inflammatory macrophages. SP140 inhibition through GSK761 reduced monocyte-to-inflammatory macrophage differentiation and lipopolysaccharide (LPS)-induced inflammatory activation, while inducing the generation of CD206+ regulatory macrophages that were shown to associate with a therapeutic response to anti-TNF in CD patients. SP140 preferentially occupies transcriptional start sites in inflammatory macrophages, with enrichment at gene loci encoding pro-inflammatory cytokines/chemokines and inflammatory pathways. GSK761 specifically reduces SP140 chromatin binding and thereby expression of SP140-regulated genes. GSK761 inhibits the expression of cytokines, including TNF, by CD14+ macrophages isolated from CD intestinal mucosa. CONCLUSIONS: This study identifies SP140 as a druggable epigenetic therapeutic target for CD.

Metabolic profile changes in serum of migraine patients detected using 1H-NMR spectroscopy.

BACKGROUND: Migraine is a common brain disorder but reliable diagnostic biomarkers in blood are still lacking. Our aim was to identify, using proton nuclear magnetic resonance (1H-NMR) spectroscopy, metabolites in serum that are associated with lifetime and active migraine by comparing metabolic profiles of patients and controls. METHODS: Fasting serum samples from 313 migraine patients and 1512 controls from the Erasmus Rucphen Family (ERF) study were available for 1H-NMR spectroscopy. Data was analysed using elastic net regression analysis. RESULTS: A total of 100 signals representing 49 different metabolites were detected in 289 cases (of which 150 active migraine patients) and 1360 controls. We were able to identify profiles consisting of 6 metabolites predictive for lifetime migraine status and 22 metabolites predictive for active migraine status. We estimated with subsequent regression models that after correction for age, sex, BMI and smoking, the association with the metabolite profile in active migraine remained. Several of the metabolites in this profile are involved in lipid, glucose and amino acid metabolism. CONCLUSION: This study indicates that metabolic profiles, based on serum concentrations of several metabolites, including lipids, amino acids and metabolites of glucose metabolism, can distinguish active migraine patients from controls.

Novel Insights Into Rheumatoid Arthritis Through Characterization of Concordant Changes in DNA Methylation and Gene Expression in Synovial Biopsies of Patients With Differing Numbers of Swollen Joints.

In this study, we sought to characterize synovial tissue obtained from individuals with arthralgia and disease-specific auto-antibodies and patients with established rheumatoid arthritis (RA), by applying an integrative multi-omics approach where we investigated differences at the level of DNA methylation and gene expression in relation to disease pathogenesis. We performed concurrent whole-genome bisulphite sequencing and RNA-Sequencing on synovial tissue obtained from the knee and ankle from 4 auto-antibody positive arthralgia patients and thirteen RA patients. Through multi-omics factor analysis we observed that the latent factor explaining the variance in gene expression and DNA methylation was associated with Swollen Joint Count 66 (SJC66), with patients with SJC66 of 9 or more displaying separation from the rest. Interrogating these observed differences revealed activation of the immune response as well as dysregulation of cell adhesion pathways at the level of both DNA methylation and gene expression. We observed differences for 59 genes in particular at the level of both transcript expression and DNA methylation. Our results highlight the utility of genome-wide multi-omics profiling of synovial samples for improved understanding of changes associated with disease spread in arthralgia and RA patients, and point to novel candidate targets for the treatment of the disease.

Recursive ensemble feature selection provides a robust mRNA expression signature for myalgic encephalomyelitis/chronic fatigue syndrome.

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a chronic disorder characterized by disabling fatigue. Several studies have sought to identify diagnostic biomarkers, with varying results. Here, we innovate this process by combining both mRNA expression and DNA methylation data. We performed recursive ensemble feature selection (REFS) on publicly available mRNA expression data in peripheral blood mononuclear cells (PBMCs) of 93 ME/CFS patients and 25 healthy controls, and found a signature of 23 genes capable of distinguishing cases and controls. REFS highly outperformed other methods, with an AUC of 0.92. We validated the results on a different platform (AUC of 0.95) and in DNA methylation data obtained from four public studies on ME/CFS (99 patients and 50 controls), identifying 48 gene-associated CpGs that predicted disease status as well (AUC of 0.97). Finally, ten of the 23 genes could be interpreted in the context of the derailed immune system of ME/CFS.

Epigenetic age acceleration in the emerging burden of cardiometabolic diseases among migrant and non-migrant African populations: the population based cross-sectional RODAM study.

BACKGROUND: African populations are experiencing health transitions due to rapid urbanization and international migration. However, the role of biological aging in this emerging burden of cardiometabolic diseases (CMD) among migrant and non-migrant Africans is unknown. We aimed to examine differences in epigenetic age acceleration (EAA) as measured by four clocks (Horvath, Hannum, PhenoAge and GrimAge) and their associations with cardiometabolic factors among migrant Ghanaians in Europe and non-migrant Ghanaians. METHODS: Genome-wide DNA methylation (DNAm) data of 712 Ghanaians from cross-sectional RODAM study were used to quantify EAA. We assessed correlation of DNAmAge measures with chronological age, and then performed linear regressions to determine associations of body mass index (BMI), fasting blood glucose (FBG), blood pressure, alcohol consumption, smoking, physical activity, and one-carbon metabolism nutrients with EAA among migrant and non-migrants. We replicated our findings among 172 rural-urban sibling pairs from India migration study and among 120 native South Africans from PURE-SA-NW study. FINDINGS: We found that Ghanaian migrants have lower EAA than non-migrants. Within migrants, higher FBG was positively associated with EAA measures. Within non-migrants, higher BMI, and Vitamin B9 (folate) intake were negatively associated with EAA measures. Our findings on FBG, BMI and folate were replicated in the independent cohorts. INTERPRETATION: Our study shows that migration is negatively associated with EAA among Ghanaians. Moreover, cardiometabolic factors are differentially associated with EAA within migrant and non-migrant subgroups. Our results call for context-based interventions for CMD among transitioning populations that account for effects of biological aging. FUNDING: European Commission.

Differential DNA methylation in familial hypercholesterolemia.

BACKGROUND: Familial hypercholesterolemia (FH) is a monogenic disorder characterized by elevated low-density lipoprotein cholesterol (LDL-C). A FH causing genetic variant in LDLR, APOB, or PCSK9 is not identified in 12-60% of clinical FH patients (FH mutation-negative patients). We aimed to assess whether altered DNA methylation might be associated with FH in this latter group. METHODS: In this study we included 78 FH mutation-negative patients and 58 FH mutation-positive patients with a pathogenic LDLR variant. All patients were male, not using lipid lowering therapies and had LDL-C levels >6 mmol/L and triglyceride levels <3.5 mmol/L. DNA methylation was measured with the Infinium Methylation EPIC 850 K beadchip assay. Multiple linear regression analyses were used to explore DNA methylation differences between the two groups in genes related to lipid metabolism. A gradient boosting machine learning model was applied to investigate accumulated genome-wide differences between the two groups. FINDINGS: Candidate gene analysis revealed one significantly hypomethylated CpG site in CPT1A (cg00574958) in FH mutation-negative patients, while no differences in methylation in other lipid genes were observed. The machine learning model did distinguish the two groups with a mean Area Under the Curve (AUC)±SD of 0.80±0.17 and provided two CpG sites (cg26426080 and cg11478607) in genes with a possible link to lipid metabolism (PRDM16 and GSTT1). INTERPRETATION: FH mutation-negative patients are characterized by accumulated genome wide DNA methylation differences, but not by major DNA methylation alterations in known lipid genes compared to FH mutation-positive patients. FUNDING: ZonMW grant (VIDI no. 016.156.445).

HDAC3 Mediates the Inflammatory Response and LPS Tolerance in Human Monocytes and Macrophages.

Histone deacetylases (HDACs) are a group of enzymes that control histone deacetylation and bear potential to direct expression of large gene sets. We determined the effect of HDAC inhibitors (HDACi) on human monocytes and macrophages, with respect to their polarization, activation, and their capabilities of inducing endotoxin tolerance. To address the role for HDACs in macrophage polarization, we treated monocytes with HDAC3i, HDAC6i or pan-HDACi prior to polarization into M1 or M2 macrophages using IFNγ or IL-4 respectively. To study the HDAC inhibition effect on cytokine expression, macrophages were treated with HDACi prior to LPS-stimulation. TNFα, IL-6, and p40 were measured with ELISA, whereas modifications of Histone 3 and STAT1 were assessed using western blot. To address the role for HDAC3 in repeated LPS challenge induction, HDAC3i or HDAC3 siRNA was added to monocytes prior to incubation with IFNγ, which were then repeatedly challenged with LPS and analyzed by means of protein analyses and transcriptional profiling. Pan-HDACi and HDAC3i reduced cytokine secretion in monocytes and M1 macrophages, whereas HDAC6i yielded no such effect. Notably, neither pan-HDACi nor HDAC3i reduced cytokine secretion in M2 macrophages. In contrast to previous reports in mouse macrophages, HDAC3i did not affect macrophage polarization in human cells. Likewise, HDAC3 was not required for IFNγ signaling or IFNß secretion. Cytokine and gene expression analyses confirmed that IFNγ-treated macrophages consistently develop a cytokine response after LPS repeated challenge, but pretreatment with HDAC3i or HDAC3 siRNA reinstates a state of tolerance reflected by general suppression of tolerizable genes, possibly through decreasing TLRs expression, and particularly TLR4/CD14. The development of endotoxin tolerance in macrophages is important to reduce exacerbated immune response and limit tissue damage. We conclude that HDAC3 is an attractive protein target to mediate macrophage reactivity and tolerance induction in inflammatory macrophages.

Comparing genome-scale DNA methylation and CNV marks between adult human cultured ITGA6+ testicular cells and seminomas to assess in vitro genomic stability.

Autologous transplantation of spermatogonial stem cells is a promising new avenue to restore fertility in infertile recipients. Expansion of the initial spermatogonial stem cell pool through cell culturing is a necessary step to obtain enough cells for effective repopulation of the testis after transplantation. Since in vitro propagation can lead to (epi-)genetic mutations and possibly malignant transformation of the starting cell population, we set out to investigate genome-wide DNA methylation status in uncultured and cultured primary testicular ITGA6+ sorted cells and compare them with germ cell tumor samples of the seminoma subtype. Seminomas displayed a severely global hypomethylated profile, including loss of genomic imprinting, which we did not detect in cultured primary testicular ITGA6+ cells. Differential methylation analysis revealed altered regulation of gamete formation and meiotic processes in cultured primary testicular ITGA6+ cells but not in seminomas. The pivotal POU5F1 marker was hypomethylated in seminomas but not in uncultured or cultured primary testicular ITGA6+ cells, which is reflected in the POU5F1 mRNA expression levels. Lastly, seminomas displayed a number of characteristic copy number variations that were not detectable in primary testicular ITGA6+ cells, either before or after culture. Together, the data show a distinct DNA methylation patterns in cultured primary testicular ITGA6+ cells that does not resemble the pattern found in seminomas, but also highlight the need for more sensitive methods to fully exclude the presence of malignant cells after culture and to further study the epigenetic events that take place during in vitro culture.

DNA methylation abundantly associates with fetal alcohol spectrum disorder and its subphenotypes.

Aim: Fetal alcohol spectrum disorder (FASD) involves prenatal growth delay, impaired facial and CNS development and causes severe clinical, social-economic burdens. Here, we aim to detect DNA-methylation aberrations associated with FASD and potential FASD diagnostic and prognostic biomarkers. Patients & methods: The FASD diagnosis was established according to golden-standard protocols in a discovery and independent replication cohort. Genome-wide differential methylation association and replication analyses were performed. Results: We identified several loci that were robustly associated with FASD or one of its sub phenotypes. Our findings were evaluated using previously reported genome-wide surveys. Conclusion: We have detected robust FASD associated differentially methylated positions and differentially methylated regions for FASD in general and for FASD subphenotypes, in other words on growth delay, impaired facial and CNS development.

Meta-Analysis of in vitro-Differentiated Macrophages Identifies Transcriptomic Signatures That Classify Disease Macrophages in vivo.

Macrophages are heterogeneous leukocytes regulated in a tissue- and disease-specific context. While in vitro macrophage models have been used to study diseases empirically, a systematic analysis of the transcriptome thereof is lacking. Here, we acquired gene expression data from eight commonly-used in vitro macrophage models to perform a meta-analysis. Specifically, we obtained gene expression data from unstimulated macrophages (M0) and macrophages stimulated with lipopolysaccharides (LPS) for 2-4 h (M-LPSearly), LPS for 24 h (M-LPSlate), LPS and interferon-γ (M-LPS+IFNγ), IFNγ (M-IFNγ), interleukin-4 (M-IL4), interleukin-10 (M-IL10), and dexamethasone (M-dex). Our meta-analysis identified consistently differentially expressed genes that have been implicated in inflammatory and metabolic processes. In addition, we built macIDR, a robust classifier capable of distinguishing macrophage activation states with high accuracy (>0.95). We classified in vivo macrophages with macIDR to define their tissue- and disease-specific characteristics. We demonstrate that alveolar macrophages display high resemblance to IL10 activation, but show a drop in IFNγ signature in chronic obstructive pulmonary disease patients. Adipose tissue-derived macrophages were classified as unstimulated macrophages, but acquired LPS-activation features in diabetic-obese patients. Rheumatoid arthritis synovial macrophages exhibit characteristics of IL10- or IFNγ-stimulation. Altogether, we defined consensus transcriptional profiles for the eight in vitro macrophage activation states, built a classification model, and demonstrated the utility of the latter for in vivo macrophages.

A distinct epigenetic profile distinguishes stenotic from non-inflamed fibroblasts in the ileal mucosa of Crohn's disease patients.

BACKGROUND: The chronic remitting and relapsing intestinal inflammation characteristic of Crohn's disease frequently leads to fibrosis and subsequent stenosis of the inflamed region. Approximately a third of all Crohn's disease patients require resection at some stage in their disease course. As the pathogenesis of Crohn's disease associated fibrosis is largely unknown, a strong necessity exists to better understand the pathophysiology thereof. METHODS: In this study, we investigated changes of the DNA methylome and transcriptome of ileum-derived fibroblasts associated to the occurrence of Crohn's disease associated fibrosis. Eighteen samples were included in a DNA methylation array and twenty-one samples were used for RNA sequencing. RESULTS: Most differentially methylated regions and differentially expressed genes were observed when comparing stenotic with non-inflamed samples. By contrast, few differences were observed when comparing Crohn's disease with non-Crohn's disease, or inflamed with non-inflamed tissue. Integrative methylation and gene expression analyses revealed dysregulation of genes associated to the PRKACA and E2F1 network, which is involved in cell cycle progression, angiogenesis, epithelial to mesenchymal transition, and bile metabolism. CONCLUSION: Our research provides evidence that the methylome and the transcriptome are systematically dysregulated in stenosis-associated fibroblasts.

Peripheral insulin resistance rather than beta cell dysfunction accounts for geographical differences in impaired fasting blood glucose among sub-Saharan African individuals: findings from the RODAM study.

AIMS/HYPOTHESIS: The aim of this study was to assess the extent to which insulin resistance and beta cell dysfunction account for differences in impaired fasting blood glucose (IFBG) levels in sub-Saharan African individuals living in different locations in Europe and Africa. We also aimed to identify determinants associated with insulin resistance and beta cell dysfunction among this population. METHODS: Data from the cross-sectional multicentre Research on Obesity and Diabetes among African Migrants (RODAM) study were analysed. Participants included Ghanaian individuals without diabetes, aged 18-96 years old, who were residing in Amsterdam (n = 1337), Berlin (n = 502), London (n = 961), urban Ghana (n = 1309) and rural Ghana (n = 970). Glucose and insulin were measured in fasting venous blood samples. Anthropometrics were assessed during a physical examination. Questionnaires were used to assess demographics, physical activity, smoking status, alcohol consumption and energy intake. Insulin resistance and beta cell function were determined using homeostatic modelling (HOMA-IR and HOMA-B, respectively). Logistic regression analysis was used to study the contribution of HOMA-IR and inverse HOMA-B (beta cell dysfunction) to geographical differences in IFBG (fasting glucose 5.6-6.9 mmol/l). Multivariate linear regression analysis was used to identify determinants associated with HOMA-IR and inverse HOMA-B. RESULTS: IFBG was more common in individuals residing in urban Ghana (OR 1.41 [95% CI 1.08, 1.84]), Amsterdam (OR 3.44 [95% CI 2.69, 4.39]) and London (OR 1.58 [95% CI 1.20 2.08), but similar in individuals living in Berlin (OR 1.00 [95% CI 0.70, 1.45]), compared with those in rural Ghana (reference population). The attributable risk of IFBG per 1 SD increase in HOMA-IR was 69.3% and in inverse HOMA-B was 11.1%. After adjustment for HOMA-IR, the odds for IFBG reduced to 0.96 (95% CI 0.72, 1.27), 2.52 (95%CI 1.94, 3.26) and 1.02 (95% CI 0.78, 1.38) for individuals in Urban Ghana, Amsterdam and London compared with rural Ghana, respectively. In contrast, adjustment for inverse HOMA-B had very minor impact on the ORs of IFBG. In multivariate analyses, BMI (ß = 0.17 [95% CI 0.11, 0.24]) and waist circumference (ß = 0.29 [95%CI 0.22, 0.36]) were most strongly associated with higher HOMA-IR, whereas inverse HOMA-B was most strongly associated with age (ß = 0.20 [95% CI 0.16, 0.23]) and excess alcohol consumption (ß = 0.25 [95% CI 0.07, 0.43]). CONCLUSIONS/INTERPRETATION: Our findings suggest that insulin resistance, rather than beta cell dysfunction, is more important in accounting for the geographical differences in IFBG among sub-Saharan African individuals. We also show that BMI and waist circumference are important factors in insulin resistance in this population.

Peripheral blood methylation profiling of female Crohn's disease patients.

BACKGROUND: Crohn's disease (CD) is a chronic inflammatory disorder belonging to the inflammatory bowel diseases (IBD). CD affects distinct parts of the gastrointestinal tract, leading to symptoms including diarrhea, fever, abdominal pain, weight loss, and anemia. The aim of this study was to assess whether the DNA methylome of peripheral blood cells can be associated with CD in women. METHODS: Samples were obtained from 18 female patients with histologically confirmed ileal or ileocolic CD and 25 healthy age- and gender-matched controls (mean age and standard deviation: 30.5 ± 6.5 years for both groups). Genome-wide DNA methylation was determined using the Illumina HumanMethylation 450k BeadChip. RESULTS: Our analysis implicated 4287 differentially methylated positions (DMPs; corrected p < 0.05) that are associated to 2715 unique genes. Gene ontology enrichment analysis revealed significant enrichment of our DMPs in immune response processes and inflammatory pathways. Of the 4287 DMPs, 32 DMPs were located on chromosome X with several hits for MIR223 and PABPC5. Comparison with previously performed (epi)genome-wide studies revealed that we replicated 33 IBD-associated genes. In addition to DMPs, we found eight differentially methylated regions (DMRs). CONCLUSIONS: CD patients display a characteristic DNA methylation landscape, with the differentially methylated genes being implicated in immune response. Additionally, DMPs were found on chromosome X suggesting X-linked manifestations of CD that could be associated with female-specific symptoms.

Radioprotection of IDH1-Mutated Cancer Cells by the IDH1-Mutant Inhibitor AGI-5198.

Isocitrate dehydrogenase 1 (IDH1) is mutated in various types of human cancer to IDH1(R132H), a structural alteration that leads to catalysis of α-ketoglutarate to the oncometabolite D-2-hydroxyglutarate. In this study, we present evidence that small-molecule inhibitors of IDH1(R132H) that are being developed for cancer therapy may pose risks with coadministration of radiotherapy. Cancer cells heterozygous for the IDH1(R132H) mutation exhibited less IDH-mediated production of NADPH, such that after exposure to ionizing radiation (IR), there were higher levels of reactive oxygen species, DNA double-strand breaks, and cell death compared with IDH1 wild-type cells. These effects were reversed by the IDH1(R132H) inhibitor AGI-5198. Exposure of IDH1 wild-type cells to D-2-hydroxyglutarate was sufficient to reduce IDH-mediated NADPH production and increase IR sensitivity. Mechanistic investigations revealed that the radiosensitivity of heterozygous cells was independent of the well-described DNA hypermethylation phenotype in IDH1-mutated cancers. Thus, our results argue that altered oxidative stress responses are a plausible mechanism to understand the radiosensitivity of IDH1-mutated cancer cells. Further, they offer an explanation for the relatively longer survival of patients with IDH1-mutated tumors, and they imply that administration of IDH1(R132H) inhibitors in these patients may limit irradiation efficacy in this setting.