Molecular diagnostics for verification of pleural tuberculosis in Morocco

Pleural tuberculosis (pTB) is a very common form of extrapulmonary tuberculosis (TB). pTB diagnostics represents a major burning challenge worldwide due to the limitations of available conventional diagnostic tools. These latter include microscopic examination of the pleural fluid for acid-fast bacilli, mycobacterial culture of pleural fluid in solid or liquid media, sputum or pleural tissue, and histopathological examination of pleural tissue; these tests have recognized limitations for clinical use. Hence, to overcome these limitations, attention has been devoted to new nucleic acid amplification (NAA) diagnostic tests such as the polymerase chain reaction (PCR) and real-time PCR (RT-PCR), owing to their accuracy, rapidity, high sensitivity and specificity. Within this context, this prospective study was conducted to evaluate the performance of molecular diagnosis methods for differentiation between tuberculosis and non-tuberculosis pleural effusions. Fifty patients with pleural effusion were enrolled in this prospective study in Rabat, Morocco. The efficacy of conventional polymerase chain reaction (PCR) in the diagnostics of tuberculous pleurisy by targeting IS6110 and mycobacterial internal transcribed spacer (MYITS) was evaluated compared to histopathologic examination and culture data. Our results showed that IS6110 PCR could “rule in” pTB, the sensitivity and specificity being 41.6% and 85.7%, respectively. Therefore, the findings confirmed that molecular tests exert a relatively high specificity in EPTB but lower sensitivity, thus a positive test is considered as a pTB case whereas negative one cannot exclude the disease. Although the study was limited by a small sample size, it adds to the body of evidence of usefulness of molecular testing as adjuncts to histopathologic examination for accurate diagnosis of pTB, to treat timely and to avoid the emergence and spread of drug resistant pTB. However, further efforts should be made to increase the sensitivity of NAA methods and to identify the best molecular targets to be useful in clinical practice

GFI1 proteins regulate stem cell formation in the AGM

In vertebrates, the first haematopoietic stem cells (HSCs) with multi-lineage and long-term repopulating potential arise in the AGM (aorta-gonad-mesonephros) region. These HSCs are generated from a rare and transient subset of endothelial cells, called haemogenic endothelium (HE), through an endothelial-to-haematopoietic transition (EHT). Here, we establish the absolute requirement of the transcriptional repressors GFI1 and GFI1B (growth factor independence 1 and 1B) in this unique trans-differentiation process. We first demonstrate that Gfi1 expression specifically defines the rare population of HE that generates emerging HSCs. We further establish that in the absence of GFI1 proteins, HSCs and haematopoietic progenitor cells are not produced in the AGM, revealing the critical requirement for GFI1 proteins in intra-embryonic EHT. Finally, we demonstrate that GFI1 proteins recruit the chromatin-modifying protein LSD1, a member of the CoREST repressive complex, to epigenetically silence the endothelial program in HE and allow the emergence of blood cells.We thank the staff at the Advanced Imaging, animal facility, Molecular Biology Core facilities and Flow Cytometry of CRUK Manchester Institute for technical support and Michael Lie-A-Ling and Elli Marinopoulou for initiating the DamID-PIP bioinformatics project. We thank members of the Stem Cell Biology group, the Stem Cell Haematopoiesis groups and Martin Gering for valuable advice and critical reading of the manuscript. Work in our laboratory is supported by the Leukaemia and Lymphoma Research Foundation (LLR), Cancer Research UK (CRUK) and the Biotechnology and Biological Sciences Research Council (BBSRC). SC is the recipient of an MRC senior fellowship (MR/J009202/1).This is the author accepted manuscript. The final version is available from NPG via http://dx.doi.org/10.1038/ncb327

Murine hematopoietic stem cell activity is derived from pre-circulation embryos but not yolk sacs.

The embryonic site of definitive hematopoietic stem cell (dHSC) origination has been debated for decades. Although an intra-embryonic origin is well supported, the yolk sac (YS) contribution to adult hematopoiesis remains controversial. The same developmental origin makes it difficult to identify specific markers that discern between an intraembryonic versus YS-origin using a lineage trace approach. Additionally, the highly migratory nature of blood cells and the inability of pre-circulatory embryonic cells (i.e., 5-7 somite pairs (sp)) to robustly engraft in transplantation, even after culture, has precluded scientists from properly answering these questions. Here we report robust, multi-lineage and serially transplantable dHSC activity from cultured 2-7sp murine embryonic explants (Em-Ex). dHSC are undetectable in 2-7sp YS explants. Additionally, the engraftment from Em-Ex is confined to an emerging CD31+CD45+c-Kit+CD41- population. In sum, our work supports a model in which the embryo, not the YS, is the major source of lifelong definitive hematopoiesis

Intra-Aortic Clusters Undergo Endothelial to Hematopoietic Phenotypic Transition during Early Embryogenesis

Intra-aortic clusters (IACs) attach to floor of large arteries and are considered to have recently acquired hematopoietic stem cell (HSC)-potential in vertebrate early mid-gestation embryos. The formation and function of IACs is poorly understood. To address this issue, IACs were characterized by immunohistochemistry and flow cytometry in mouse embryos. Immunohistochemical analysis revealed that IACs simultaneously express the surface antigens CD31, CD34 and c-Kit. As embryos developed from 9.5 to 10.5 dpc, IACs up-regulate the hematopoietic markers CD41 and CD45 while down-regulating the endothelial surface antigen VE-cadherin/CD144, suggesting that IACs lose endothelial phenotype after 9.5 dpc. Analysis of the hematopoietic potential of IACs revealed a significant change in macrophage CFC activity from 9.5 to 10.5 dpc. To further characterize IACs, we isolated IACs based on CD45 expression. Correspondingly, the expression of hematopoietic transcription factors in the CD45(neg) fraction of IACs was significantly up-regulated. These results suggest that the transition from endothelial to hematopoietic phenotype of IACs occurs after 9.5 dpc

Decoding the regulatory network of early blood development from single-cell gene expression measurements.

Reconstruction of the molecular pathways controlling organ development has been hampered by a lack of methods to resolve embryonic progenitor cells. Here we describe a strategy to address this problem that combines gene expression profiling of large numbers of single cells with data analysis based on diffusion maps for dimensionality reduction and network synthesis from state transition graphs. Applying the approach to hematopoietic development in the mouse embryo, we map the progression of mesoderm toward blood using single-cell gene expression analysis of 3,934 cells with blood-forming potential captured at four time points between E7.0 and E8.5. Transitions between individual cellular states are then used as input to develop a single-cell network synthesis toolkit to generate a computationally executable transcriptional regulatory network model of blood development. Several model predictions concerning the roles of Sox and Hox factors are validated experimentally. Our results demonstrate that single-cell analysis of a developing organ coupled with computational approaches can reveal the transcriptional programs that underpin organogenesis.We thank J. Downing (St. Jude Children's Research Hospital, Memphis, TN, USA) for the Runx1-ires-GFP mouse. Research in the authors' laboratory is supported by the Medical Research Council, Biotechnology and Biological Sciences Research Council, Leukaemia and Lymphoma Research, the Leukemia and Lymphoma Society, Microsoft Research and core support grants by the Wellcome Trust to the Cambridge Institute for Medical Research and Wellcome Trust - MRC Cambridge Stem Cell Institute. V.M. is supported by a Medical Research Council Studentship and Centenary Award and S.W. by a Microsoft Research PhD Scholarship.This is the accepted manuscript for a paper published in Nature Biotechnology 33, 269–276 (2015) doi:10.1038/nbt.315

Inductive interactions mediated by interplay of asymmetric signalling underlie development of adult hematopoietic stem cells

During embryonic development, adult haematopoietic stem cells (HSCs) emerge preferentially in the ventral domain of the aorta in the aorta-gonad-mesonephros (AGM) region. Several signalling pathways such as Notch, Wnt, Shh and RA are implicated in this process, yet how these interact to regulate the emergence of HSCs has not previously been described in mammals. Using a combination of ex vivo and in vivo approaches, we report here that stage-specific reciprocal dorso-ventral inductive interactions and lateral input from the urogenital ridges are required to drive HSC development in the aorta. Our study strongly suggests that these inductive interactions in the AGM region are mediated by the interplay between spatially polarized signalling pathways. Specifically, Shh produced in the dorsal region of the AGM, stem cell factor in the ventral and lateral regions, and BMP inhibitory signals in the ventral tissue are integral parts of the regulatory system involved in the development of HSCs

Runx1 is required for the endothelial to haematopoietic cell transition but not thereafter

HSCs are the founder cells of the adult hematopoietic system, and thus knowledge of the molecular program directing their generation during development is important for regenerative hematopoietic strategies. Runx1 is a pivotal transcription factor required for HSC generation in the vascular regions of the mouse conceptus - the aorta, vitelline and umbilical arteries, yolk sac and placenta 1, 2. It is thought that HSCs emerge from vascular endothelial cells through the formation of intra-arterial clusters 3 and that Runx1 functions during the transition from ‘hemogenic endothelium’ to HSCs 4, 5. Here we show by conditional deletion that Runx1 activity in vascular endothelial cadherin (VEC) positive endothelial cells is indeed essential for intra-arterial cluster, hematopoietic progenitor, and HSC formation. In contrast, Runx1 is not required in cells expressing Vav, one of the first pan-hematopoietic genes expressed in HSCs. Collectively these data show that Runx1 function is essential in endothelial cells for hematopoietic progenitor and HSC formation from the vasculature, but its requirement ends once or before Vav is expressed

A characterization of polynomially Riesz strongly continuous semigroups

summary:In this paper we characterize the class of polynomially Riesz strongly continuous semigroups on a Banach space $X$. Our main results assert, in particular, that the generators of such semigroups are either polynomially Riesz (then bounded) or there exist two closed infinite dimensional invariant subspaces $X_0$ and $X_1$ of $X$ with $X=X_0\oplus X_1$ such that the part of the generator in $X_0$ is unbounded with resolvent of Riesz type while its part in $X_1$ is a polynomially Riesz operator

Fixed Point Theorems with Applications to the Solvability of Operator Equations and Inclusions on Function Spaces

Fixed point theory is an elegant mathematical theory which is a beautiful mixture of analysis, topology, and geometry. It is an interdisciplinary theory which provides powerful tools for the solvability of central problems in many areas of current interest in mathematics and other quantitative sciences, such as physics, engineering, biology, and economy. In fact, the existence of linear and nonlinear problems is frequently transformed into fixed point problems, for example, the existence of solutions to partial differential equations, the existence of solutions to integral equations, and the existence of periodic orbits in dynamical systems. This makes fixed point theory a topical area and a subject of active scientific research, constantly evolving and growing and in a perpetual progress. Fixed point theory has two main branches: on one hand, we can consider the results obtained by using metric properties; on the other hand, we can consider the results deduced from topological properties. Among the classical results, which are the basis for the metric branch, we retain the Banach contraction principle in complete metric spaces established by Banach in 1922. With respect to the topology branch, the main theorems are those of Brouwer and his infinite dimensional version, the Schauder fixed point theorem, which were proved in 1912 and 1930, respectively. In both, the compactness plays an essential role. A hybrid result combining metric and topological properties was established by Krasnoselskii in 1955. This result is exciting and has a very wide scope. In recent years, a number of authors have found this theorem a very satisfactory outcome for the study of stability and asymptotic behavior of solutions for certain differential equations which resisted the direct Lyapunov method. The development of such fixed point theorems remains a constant concern for many investigators who have continued to enrich this field by new quite interesting contributions following constantly the progress of applications to fully meet the needs of this dynamic and active field. Thus, fixed point theorems were developed for single-valued and multivalued mappings on topological vector spaces, metric spaces, Banach spaces, Banach algebras, posets, lattices, Banach lattices, and so forth. Due to the importance of fixed point theory and its applications, it is worthwhile to publish a special issue on this topic to highlight recent advances made by mathematicians actively working in this area. This special issue was originally elaborated to report the latest advances in fixed point theory in abstract spaces and their applications. It includes works on single-valued and multivalued mappings in normed and metric spaces and various applications to boundary value problems, equilibrium problems, and variational inequalities