Comprehensive mapping of the effects of azacitidine on DNA methylation, repressive/permissive histone marks and gene expression in primary cells from patients with MDS and MDS-related disease

Azacitidine (Aza) is first-line treatment for patients with high-risk myelodysplastic syndromes (MDS), although its precise mechanism of action is unknown. We performed the first study to globally evaluate the epigenetic effects of Aza on MDS bone marrow progenitor cells assessing gene expression (RNA seq), DNA methylation (Illumina 450k) and the histone modifications H3K18ac and H3K9me3 (ChIP seq). Aza induced a general increase in gene expression with 924 significantly upregulated genes but this increase showed no correlation with changes in DNA methylation or H3K18ac, and only a weak association with changes in H3K9me3. Interestingly, we observed activation of transcripts containing 15 endogenous retroviruses (ERVs) confirming previous cell line studies. DNA methylation decreased moderately in 99% of all genes, with a median beta-value reduction of 0.018; the most pronounced effects seen in heterochromatin. Aza-induced hypomethylation correlated significantly with change in H3K9me3. The pattern of H3K18ac and H3K9me3 displayed large differences between patients and healthy controls without any consistent pattern induced by Aza. We conclude that the marked induction of gene expression only partly could be explained by epigenetic changes, and propose that activation of ERVs may contribute to the clinical effects of Aza in MDS.Peer reviewe

A cytomorphological and immunohistochemical profile of aggressive B-cell lymphoma: high clinical impact of a cumulative immunohistochemical outcome predictor score

We analyzed morphological and immunohistochemical features in 174 aggressive B-cell lymphomas of nodal and extranodal origin. Morphological features included presence or absence of a follicular component and cytologic criteria according to the Kiel classification, whereas immunohistochemical studies included expression of CD10, BCL-2, BCL-6, IRF4/MUM1, HLA-DR, p53, Ki-67 and the assessment of plasmacytoid differentiation. Patients were treated with a CHOP-like regimen. While the presence or absence of either CD10, BCL-6 and IRF4/MUM1 reactivity or plasmacytoid differentiation did not identify particular cytomorphologic or site-specific subtypes, we found that expression of CD10 and BCL-6, and a low reactivity for IRF4/MUM1 were favourable prognostic indicators. In contrast, BCL-2 expression and presence of a monotypic cytoplasmic immunoglobulin expression was associated with an unfavourable prognosis in univariate analyses. Meta-analysis of these data resulted in the development of a cumulative immunohistochemical outcome predictor score (CIOPS) enabling the recognition of four distinct prognostic groups. Multivariate analysis proved this score to be independent of the international prognostic index. Such a cumulative immunohistochemical scoring approach might provide a valuable alternative in the recognition of defined risk types of aggressive B-cell lymphomas

Prolonged Mechanical Ventilation Induces Cell Cycle Arrest in Newborn Rat Lung

Rationale: The molecular mechanism(s) by which mechanical ventilation disrupts alveolar development, a hallmark of bronchopulmonary dysplasia, is unknown. Objective: To determine the effect of 24 h of mechanical ventilation on lung cell cycle regulators, cell proliferation and alveolar formation in newborn rats. Methods: Seven-day old rats were ventilated with room air for 8, 12 and 24 h using relatively moderate tidal volumes (8.5 mL.kg-1). Measurement and Main Results: Ventilation for 24 h (h) decreased the number of elastin-positive secondary crests and increased the mean linear intercept, indicating arrest of alveolar development. Proliferation (assessed by BrdU incorporation) was halved after 12 h of ventilation and completely arrested after 24 h. Cyclin D1 and E1 mRNA and protein levels were decreased after 8-24 h of ventilation, while that of p27Kip1 was significantly increased. Mechanical ventilation for 24 h also increased levels of p57Kip2, decreased that of p16INK4a, while the levels of p21Waf/Cip1 and p15INK4b were unchanged. Increased p27Ki

Is protein deuteration beneficial for proton detected solid-state NMR at and above 100 kHz magic-angle spinning?

1H-detection in solid-state NMR of proteins has been traditionally combined with deuteration for both resolution and sensitivity reasons, with the optimal level of proton dilution being dependent on MAS rate. Here we present 1H-detected 15N and 13C CP-HSQC spectra on two microcrystalline samples acquired at 60 and 111 kHz MAS and at ultra-high field. We critically compare the benefits of three labeling schemes yielding different levels of proton content in terms of resolution, coherence lifetimes and feasibility of scalar-based 2D correlations under these experimental conditions. We observe unexpectedly high resolution and sensitivity of aromatic resonances in 2D 13C-1H correlation spectra of protonated samples. Ultrafast MAS reduces or even removes the necessity of 1H dilution for high-resolution 1H-detection in biomolecular solid-state NMR. It yields 15N,1H and 13C,1H fingerprint spectra of exceptional resolution for fully protonated samples, with notably superior 1H and 13C lineshapes for side-chain resonances

Is protein deuteration beneficial for proton detected solid-state NMR at and above 100 kHz magic-angle spinning?

We thank the members of technical staff of ISA for assistance with NMR spectrometers.International audience1H-detection in solid-state NMR of proteins has been traditionally combined with deuteration for both resolution and sensitivity reasons, with the optimal level of proton dilution being dependent on MAS rate. Here we present 1H-detected 15N and 13C CP-HSQC spectra on two microcrystalline samples acquired at 60 and 111 kHz MAS and at ultra-high field. We critically compare the benefits of three labeling schemes yielding different levels of proton content in terms of resolution, coherence lifetimes and feasibility of scalar-based 2D correlations under these experimental conditions. We observe unexpectedly high resolution and sensitivity of aromatic resonances in 2D 13C-1H correlation spectra of protonated samples. Ultrafast MAS reduces or even removes the necessity of 1H dilution for high-resolution 1H-detection in biomolecular solid-state NMR. It yields 15N,1H and 13C,1H fingerprint spectra of exceptional resolution for fully protonated samples, with notably superior 1H and 13C lineshapes for side-chain resonances

Impaired mucosal homeostasis in short-term fiber deprivation is due to reduced mucus production rather than overgrowth of mucus-degrading bacteria

The gut mucosal environment is key in host health; protecting against pathogens and providing a niche for beneficial bacteria, thereby facilitating a mutualistic balance between host and microbiome. Lack of dietary fiber results in erosion of the mucosal layer, suggested to be a result of increased mucus-degrading gut bacteria. This study aimed to use quantitative analyses to investigate the diet-induced imbalance of mucosal homeostasis. Seven days of fiber-deficiency affected intestinal anatomy and physiology, seen by reduced intestinal length and loss of the colonic crypt-structure. Moreover, the mucus layer was diminished, muc2 expression decreased, and impaired mucus secretion was detected by stable isotope probing. Quantitative microbiome profiling of the gut microbiota showed a diet-induced reduction in bacterial load and decreased diversity across the intestinal tract, including taxa with fiber-degrading and butyrate-producing capabilities. Most importantly, there was little change in the absolute abundance of known mucus-degrading bacteria, although, due to the general loss of taxa, relative abundance would erroneously indicate an increase in mucus degraders. These findings underscore the importance of using quantitative methods in microbiome research, suggesting erosion of the mucus layer during fiber deprivation is due to diminished mucus production rather than overgrowth of mucus degraders

Deletions in 14q24.1q24.3 are associated with congenital heart defects, brachydactyly, and mild intellectual disability

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