Cell-specific occupancy of an extended repertoire of CREM and CREB binding loci in male germ cells

Background: CREB and CREM are closely related factors that regulate transcription in response to various stress, metabolic and developmental signals. The CREMτ activator isoform is selectively expressed in haploid spermatids and plays an essential role in murine spermiogenesis.Results: We have used chromatin immunoprecipitation coupled to sequencing (ChIP-seq) to map CREM and CREB target loci in round spermatids from adult mouse testis and spermatogonia derived GC1-spg cells respectively. We identify more than 9000 genomic loci most of which are cell-specifically occupied. Despite the fact that round spermatids correspond to a highly specialised differentiated state, our results show that they have a remarkably accessible chromatin environment as CREM occupies more than 6700 target loci corresponding not only to the promoters of genes selectively expressed in spermiogenesis, but also of genes involved in functions specific to other cell types. The expression of only a small subset of these target genes are affected in the round spermatids of CREM knockout animals. We also identify a set of intergenic binding loci some of which are associated with H3K4 trimethylation and elongating RNA polymerase II suggesting the existence of novel CREB and CREM regulated transcripts.Conclusions: We demonstrate that CREM and CREB occupy a large number of promoters in highly cell specific manner. This is the first study of CREM target promoters directly in a physiologically relevant tissue in vivo and represents the most comprehensive experimental analysis of CREB/CREM regulatory potential to date

De hygiënehypothese onder de loep

The hygiene hypothesis states that as a result of improved hygiene and reduction of childhood diseases, the human immune system overreacts to relatively harmless stimuli, offering an explanation for the increase in the prevalence of eczema, asthma and allergies in the industrialised world. On the 20th of September 2018, a debate on the hygiene hypothesis took place in ARTIS-Micropia, Amsterdam. Experts in the fields of microbiology, immunology and infection prevention debated with each other and with the audience about 5 statements regarding the hygiene hypothesis. The first 3 statements dealt with the use of the name ‘hygiene hypothesis’, whether we can confirm or disprove the hypothesis and whether prevention of allergies by administration of micro-organisms is effective. The last 2 statements stated that intensive contact with micro-organisms prevents an overactive immune system and that playing outside and having a certain dietary pattern can prevent autoimmune and allergic diseases. The experts in the panel reached consensus about the importance of exposure to a high diversity of micro-organisms in early life to prevent an overactive immune system. They agreed that more research is needed in the coming years to be able to substantiate effective and practical interventions in our daily lives

A polymorphism in C-C Chemokine Receptor 5 (CCR5) associates with Lofgren's syndrome and alters receptor expression as well as functional response

C-C chemokine receptor 5 (CCR5) and polymorphisms in CCR5 gene are associated with sarcoidosis and Lofgren's syndrome. Lofgren's syndrome is an acute and usually self-remitting phenotype of sarcoidosis. We investigated whether the single nucleotide polymorphism (SNP) rs1799987 is associated with susceptibility for Lofgren's syndrome and has an effect on CCR5 expression on monocytes and function of CCR5. A total of 106 patients with Lofgren's syndrome and 257 controls were genotyped for rs1799987. Expression of CCR5 on monocytes was measured by flowcytometry. We evaluated calcium influx kinetics following stimulation upon N-formylmethionyl-leucyl-phenylalanine (fMLP) and macrophage inflammatory protein-1 alpha (MIP-1 alpha) on monocytes by measuring the median fluorescence intensity (MFI). The frequency of the G allele of rs1799987 was significantly higher in Lofgren's syndrome than in healthy controls (p = 0.0015, confidence interval (CI) 1.22-2.32, odds ratio (OR) 1.680). Patients with a GG genotype showed higher CCR5 expression on monocytes than patients with the AA genotype (p = 0.026). A significantly (p = 0.027) lower count of patients with the GG genotype showed a calcium influx reaction to simulation upon MIP-1 alpha, compared with patients with the AA genotype. The rs1799987 G allele in CCR5 gene is associated with susceptibility to Lofgren's syndrome and with quantitative and qualitative changes in CCR5, potentially effecting the inflammatory response.Pathophysiology and treatment of rheumatic disease

Collaboration with general practitioners: preferences of medical specialists – a qualitative study

BACKGROUND: Collaboration between general practitioners (GPs) and specialists has been the focus of many collaborative care projects during the past decade. Unfortunately, quite a number of these projects failed. This raises the question of what motivates medical specialists to initiate and continue participating with GPs in new collaborative care models. The following question is addressed in this study: What motivates medical specialists to initiate and sustain new models for collaborating with GPs? METHODS: We conducted semi-structured interviews with eighteen medical specialists in the province of Groningen, in the North of The Netherlands. The sampling criteria were age, gender, type of hospital in which they were practicing, and specialty. The interviews were recorded, fully transcribed, and analysed by three researchers working independently. The resulting motivational factors were grouped into categories. RESULTS: 'Teaching GPs' and 'regulating patient flow' (referrals) appeared to dominate when the motivational factors were considered. In addition, specialists want to develop relationships with the GPs on a more personal level. Most specialists believe that there is not much they can learn from GPs. 'Lack of time', 'no financial compensation', and 'no support from colleagues' were considered to be the main concerns to establishing collaborative care practices. Additionally, projects were often experienced as too complex and time consuming whereas guidelines were experienced as too restrictive. CONCLUSION: Specialists are particularly interested in collaborating because the GP is the gatekeeper for access to secondary health care resources. Specialists feel that they are able to teach the GPs something, but they do not feel that they have anything to learn from the GPs. With respect to professional expertise, therefore, specialists do not consider GPs as equals. Once personal relationships with the GPs have been established, an informal network with incidental professional contact seems to be sufficient to satisfy the collaborative needs of the specialist. The concerns seem to outweigh any positive motivational forces to developing new models of collaborative practice

Human oral isolate Lactobacillus fermentum AGR1487 induces a proinflammatory response in germ-free rat colons

Lactobacilli are thought to be beneficial for human health, with lactobacilli-associated infections being confined to immune-compromised individuals. However, Lactobacillus fermentum AGR1487 negatively affects barrier integrity in vitro so we hypothesized that it caused a pro-inflammatory response in the host. We compared germ-free rats inoculated with AGR1487 to those inoculated with another L. fermentum strain, AGR1485, which does not affect in vitro barrier integrity. We showed that rats inoculated with AGR1487 had more inflammatory cells in their colon, higher levels of inflammatory biomarkers, and increased colonic gene expression of pro-inflammatory pathways. In addition, our in vitro studies showed that AGR1487 had a greater capacity to activate TLR signaling and induce pro-inflammatory cytokines in immune cells. This study indicates the potential of strains of the same species to differentially elicit inflammatory responses in the host and highlights the importance of strain characterization in probiotic approaches to treat inflammatory disorders

A Functional Role for 4qA/B in the Structural Rearrangement of the 4q35 Region and in the Regulation of FRG1 and ANT1 in Facioscapulohumeral Dystrophy

The number of D4Z4 repeats in the subtelomeric region of chromosome 4q is strongly reduced in patients with Facio-Scapulo-Humeral Dystrophy (FSHD). We performed chromosome conformation capture (3C) analysis to document the interactions taking place among different 4q35 markers. We found that the reduced number of D4Z4 repeats in FSHD myoblasts was associated with a global alteration of the three-dimensional structure of the 4q35 region. Indeed, differently from normal myoblasts, the 4qA/B marker interacted directly with the promoters of the FRG1 and ANT1 genes in FSHD cells. Along with the presence of a newly identified transcriptional enhancer within the 4qA allele, our demonstration of an interaction occurring between chromosomal segments located megabases away on the same chromosome 4q allows to revisit the possible mechanisms leading to FSHD

Remodeling of the chromatin structure of the facioscapulohumeral muscular dystrophy (FSHD) locus and upregulation of FSHD-related gene 1 (FRG1) expression during human myogenic differentiation

<p>Abstract</p> <p>Background</p> <p>Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal dominant neuromuscular disorder associated with the partial deletion of integral numbers of 3.3 kb D4Z4 DNA repeats within the subtelomere of chromosome 4q. A number of candidate FSHD genes, adenine nucleotide translocator 1 gene (<it>ANT1</it>), FSHD-related gene 1 (<it>FRG1</it>), <it>FRG2 </it>and <it>DUX4c</it>, upstream of the D4Z4 array (FSHD locus), and double homeobox chromosome 4 (<it>DUX4</it>) within the repeat itself, are upregulated in some patients, thus suggesting an underlying perturbation of the chromatin structure. Furthermore, a mouse model overexpressing <it>FRG1 </it>has been generated, displaying skeletal muscle defects.</p> <p>Results</p> <p>In the context of myogenic differentiation, we compared the chromatin structure and tridimensional interaction of the D4Z4 array and <it>FRG1 </it>gene promoter, and <it>FRG1 </it>expression, in control and FSHD cells. The <it>FRG1 </it>gene was prematurely expressed during FSHD myoblast differentiation, thus suggesting that the number of D4Z4 repeats in the array may affect the correct timing of <it>FRG1 </it>expression. Using chromosome conformation capture (3C) technology, we revealed that the <it>FRG1 </it>promoter and D4Z4 array physically interacted. Furthermore, this chromatin structure underwent dynamic changes during myogenic differentiation that led to the loosening of the <it>FRG1</it>/4q-D4Z4 array loop in myotubes. The <it>FRG1 </it>promoter in both normal and FSHD myoblasts was characterized by H3K27 trimethylation and Polycomb repressor complex binding, but these repression signs were replaced by H3K4 trimethylation during differentiation. The D4Z4 sequences behaved similarly, with H3K27 trimethylation and Polycomb binding being lost upon myogenic differentiation.</p> <p>Conclusion</p> <p>We propose a model in which the D4Z4 array may play a critical chromatin function as an orchestrator of <it>in cis </it>chromatin loops, thus suggesting that this repeat may play a role in coordinating gene expression.</p

Genome-wide DNA methylation profiling of non-small cell lung carcinomas

Background: Non-small cell lung carcinoma (NSCLC) is a complex malignancy that owing to its heterogeneity and poor prognosis poses many challenges to diagnosis, prognosis and patient treatment. DNA methylation is an important mechanism of epigenetic regulation involved in normal development and cancer. It is a very stable and specific modification and therefore in principle a very suitable marker for epigenetic phenotyping of tumors. Here we present a genome-wide DNA methylation analysis of NSCLC samples and paired lung tissues, where we combine MethylCap and next generation sequencing (MethylCap-seq) to provide comprehensive DNA methylation maps of the tumor and paired lung samples. The MethylCap-seq data were validated by bisulfite sequencing and methyl-specific polymerase chain reaction of selected regions. Results: Analysis of the MethylCap-seq data revealed a strong positive correlation between replicate experiments and between paired tumor/lung samples. We identified 57 differentially methylated regions (DMRs) present in all NSCLC tumors analyzed by MethylCap-seq. While hypomethylated DMRs did not correlate to any particular functional category of genes, the hypermethylated DMRs were strongly associated with genes encoding transcriptional regulators. Furthermore, subtelomeric regions and satelli Conclusions: Collectively, we provide a resource containing genome-wide DNA methylation maps of NSCLC and their paired lung tissues, and comprehensive lists of known and novel DMRs and associated genes in NSCLC