Dynamic DNA methylation reveals novel cis-regulatory elements in mouse hematopoiesis.

Differentiation of hematopoietic stem and progenitor cells to terminally differentiated immune cells is accompanied by large-scale remodeling of the DNA methylation landscape. Although significant insights into the molecular mechanisms of hematopoietic tissue regeneration were derived from mouse models, profiling of DNA methylation has been hampered by high cost or low resolution using available methods. The recent development of the Infinium Mouse Methylation BeadChip (MMBC) array facilitates methylation profiling of the mouse genome at a single CpG resolution at affordable cost. We extended the RnBeads package to provide a computational framework for the analysis of MMBC data. This framework was applied to a newly generated reference map of mouse hematopoiesis encompassing nine different cell types. Analysis of dynamically regulated CpG sites showed progressive and unidirectional DNA methylation changes from hematopoietic stem and progenitor cells to differentiated hematopoietic cells and allowed the identification of lineage- and cell type-specific DNA methylation programs. Comparison with previously published catalogs of cis-regulatory elements (CREs) revealed 12,856 novel putative CREs that were dynamically regulated by DNA methylation (mdCREs). These mdCREs were predominantly associated with patterns of cell type-specific DNA hypomethylation and could be identified as epigenetic control regions regulating the expression of key hematopoietic genes during differentiation. In summary, we established an analysis pipeline for MMBC data sets and provide a DNA methylation atlas of mouse hematopoiesis. This resource allowed us to identify novel putative CREs involved in hematopoiesis and will serve as a platform to study epigenetic regulation of normal and malignant hematopoiesis.

Measurement of IgE to hazelnut allergen components cannot replace hazelnut challenge in Dutch adults.

BACKGROUND: Component-resolved diagnostics (CRD) help predict hazelnut allergy (HA) in children, but are of unknown diagnostic value in adults. This study aimed to evaluate the diagnostic accuracy of IgE to hazelnut extract and components in adults. METHODS: A Dutch population of consecutively presenting adults suspected of HA, who underwent a double-blind placebo-controlled food challenge, were included. Serum IgE to hazelnut extract and Cor a 1, 8, 9, and 14 was measured on ImmunoCAP. Diagnostic accuracy was assessed by area under the curve (AUC) analysis. RESULTS: Of 89 patients undergoing challenge, 46 had challenge-confirmed HA: 17 based on objective and 29 based on subjective symptoms. At commonly applied cutoffs 0.1 and 0.35 kUA /L, high sensitivity was observed for IgE to hazelnut extract and Cor a 1 (range 85-91%), and high specificity for IgE to Cor a 8, 9 and 14 (range 77-95%). However, the AUCs for hazelnut extract and components were too low for accurate prediction of HA (range 0.50-0.56). Combining hazelnut extract and component IgE measurements did not significantly improve accuracy. Higher IgE levels to Cor a 9 and 14 were tentatively associated with HA with objective symptoms, but the corresponding AUCs still only reached 0.68 and 0.63, respectively. CONCLUSIONS: Although hazelnut allergic adults are generally sensitized to hazelnut extract and Cor a 1, and hazelnut tolerant adults are usually not sensitized to Cor a 8, 9, or 14, challenge testing is still needed to accurately discriminate between presence and absence of HA in adults from a birch-endemic country.

PARP1 regulates DNA damage-induced nucleolar-nucleoplasmic shuttling of WRN and XRCC1 in a toxicant and protein-specific manner.

The prime function of nucleoli is ribogenesis, however, several other, non-canonical functions have recently been identified, including a role in genotoxic stress response. Upon DNA damage, numerous proteins shuttle dynamically between the nucleolus and the nucleoplasm, yet the underlying molecular mechanisms are incompletely understood. Here, we demonstrate that PARP1 and PARylation contribute to genotoxic stress-induced nucleolar-nucleoplasmic shuttling of key genome maintenance factors in HeLa cells. Our work revealed that the RECQ helicase, WRN, translocates from nucleoli to the nucleoplasm upon treatment with the oxidizing agent H2O2, the alkylating agent 2-chloroethyl ethyl sulfide (CEES), and the topoisomerase inhibitor camptothecin (CPT). We show that after treatment with H2O2 and CEES, but not CPT, WRN translocation was dependent on PARP1 protein, yet independent of its enzymatic activity. In contrast, nucleolar-nucleoplasmic translocation of the base excision repair protein, XRCC1, was dependent on both PARP1 protein and its enzymatic activity. Furthermore, gossypol, which inhibits PARP1 activity by disruption of PARP1-protein interactions, abolishes nucleolar-nucleoplasmic shuttling of WRN, XRCC1 and PARP1, indicating the involvement of further upstream factors. In conclusion, this study highlights a prominent role of PARP1 in the DNA damage-induced nucleolar-nucleoplasmic shuttling of genome maintenance factors in HeLa cells in a toxicant and protein-specific manner.

Outcome of Infants with Therapeutic Hypothermia after Perinatal Asphyxia and Early-Onset Sepsis.

BACKGROUND: Animal models suggest that neuroprotective effects of therapeutic hypothermia (TH) after perinatal asphyxia are reduced in infants with early-onset sepsis. OBJECTIVES: To assess the outcome of infants with perinatal asphyxia, neonatal encephalopathy, and TH in the presence of early-onset sepsis. METHODS: In a retrospective cohort of 1,084 infants with perinatal asphyxia and TH, the outcome of 42 infants (gestational age 36.1-42.6 weeks and birth weight 2,280-5,240 g) with proven sepsis (n = 14) and probable sepsis (n = 28) was analyzed. Death, cerebral palsy, or a delayed development at 2 years was considered an adverse outcome. RESULTS: Sepsis was caused mostly by group B streptococci (n = 17), other Gram-positive bacteria (n = 5), and Candida albicans (n = 1). Of the 42 infants, 9 (21.4%) died, and 5 (11.9%) showed impairments on follow-up. The outcome is comparable to the previously reported outcome of infants with TH without early-onset sepsis. CONCLUSION: A good outcome was reported in the majority of infants with perinatal asphyxia, TH, and early-onset sepsis. Cooling should not be withheld from these infants.