Differentiation of the mononuclear phagocyte system during mouse embryogenesis: The role of transcription factor PU.1

During mouse embryogenesis, macrophage-like cells arise first in the yolk sac and are produced subsequently in the liver. The onset of liver hematopoiesis is associated with the transition from primitive to definitive erythrocyte production. This report addresses the hypothesis that a similar transition in phenotype occurs in myelopoiesis. We have used whole mount in situ hybridization to detect macrophage-specific genes expressed during mouse development. The mouse c-fms mRNA, encoding the receptor for macrophage colony-stimulating factor (CSF-1), was expressed on phagocytic cells in the yolk sac and throughout the embryo before the onset of liver hematopoiesis, Similar cells were detected using the mannose receptor, the complement receptor (CR3), or the Microphthalmia transcription factor (MITF) as mRNA markers. By contrast, other markers including the F4/80 antigen, the macrophage scavenger receptor, the S-100 proteins, S100A8 and S100A9, and the secretory product lysozyme appeared later in development and appeared restricted to only a subset of c-fms-positive cells. Two-color immunolabeling on disaggregated cells confirmed that CR3 and c-fms proteins are expressed on the same cells. Among the genes appearing later in development was the macrophage-restricted transcription factor, PU.1, which has been shown to be required for normal adult myelopoiesis. Mice with null mutations in PU.1 had normal numbers of c-fms-positive phagocytes at 11.5dpc. PU.1(-/-) embryonic stem cells were able to give rise to macrophagelike cells after cultivation in vitro. The results support previous evidence that yolk sac-derived fetal phagocytes are functionally distinct from those arising in the liver and develop via a different pathway. (C) 1999 by The American Society of Hematology.

The mononuclear phagocyte system revisited

The mononuclear phagocyte system (MPS) was defined as a family of cells comprising bone marrow progenitors, blood monocytes, and tissue macrophages. In this review, we briefly consider markers for cells of this lineage in the mouse, especially the F4/80 surface antigen and the receptor for macrophage colony-stimulating factor. The concept of the MPS is challenged by evidence that there is a separate embryonic phagocyte lineage, the blurring of the boundaries between macrophages and other cells types arising from phenotypic plasticity and transdifferentiation, and evidence of local renewal of tissue macrophage populations as opposed to monocyte recruitment. Nevertheless, there is a unity to cells of the MPS suggested by their location, morphology, and shared markers. We discuss the origins of macrophage heterogeneity and argue that macrophages and antigen-representing dendritic cells are closely related and part of the MPS.

A macrophage colony-stimulating factor receptor-green fluorescent protein transgene is expressed throughout the mononuclear phagocyte system of the mouse

The c-fms gene encodes the receptor for macrophage colony-stimulating factor (CSF-1). The gene is expressed selectively in the macrophage and trophoblast cell lineages. Previous studies have indicated that sequences in intron 2 control transcript elongation in tissue-specific and regulated expression of c-fms. In humans, an alternative promoter was implicated in expression of the gene in trophoblasts. We show that in mice, c-fms transcripts in trophoblasts initiate from multiple points within the 2-kilobase (kb) region flanking the first coding exon. A reporter gene construct containing 3.5 kb of 5' flanking sequence and the down-stream intron 2 directed expression of enhanced green fluorescent protein (EGFP) to both trophoblasts and macrophages. EGFP was detected in trophoblasts from the earliest stage of implantation examined at embryonic day 7.5. During embryonic development, EGFP highlighted the large numbers of c-fms-positive macrophages, including those that originate from the yolk sac. In adult mice, EGFP location Was consistent with known F4/80-positive macrophage populations, including Langerhans cells of the skin, and permitted convenient sorting of isolated tissue macrophages from disaggregated tissue. Expression of EGFP in transgenic mice was dependent on intron 2 as no lines with detectable EGFP expression were obtained where either all of intron 2 or a conserved enhancer element FIRE (the Fms intronic regulatory element) was removed. We have therefore defined the elements required to generate myeloid- and trophoblast-specific transgenes as well as a model system for the study of mononuclear phagocyte development and function. (C) 2003 by The American Society of Hematology.

The mononuclear phagocyte system

The mononuclear phagocyte system (MPS) has been defined as a family of cells comprising bone marrow progenitors, blood monocytes and tissue macrophages. Macrophages are a major cell population in most of the tissues in the body, and their numbers increase further in inflammation, wounding and malignancy. Their trophic roles for other cell types in development and homeostasis are becoming increasingly evident. The receptor for macrophage colony-stimulating factor (CSF-1R) is expressed in a large proportion of cells considered to be mononuclear phagocytes, including antigen-presenting dendritic cells, which can be considered a specialized adaptive state rather than a separate lineage. The unity of the MPS is challenged by evidence that there is a separate embryonic phagocyte lineage, by the transdifferentiation and fusion of MPS cells with other cell types, and by evidence of local renewal of tissue macrophage populations as opposed to monocyte recruitment. The concept of the MPS may have partly outlived its usefulness.

Continued discovery of transcriptional units expressed in cells of the mouse mononuclear phagocyte lineage

The current RIKEN transcript set represents a significant proportion of the mouse transcriptome but transcripts expressed in the innate and acquired immune systems are poorly represented. In the present study we have assessed the complexity of the transcriptome expressed in mouse macrophages before and after treatment with lipopolysaccharide, a global regulator of macrophage gene expression, using existing RIKEN 19K arrays. By comparison to array profiles of other cells and tissues, we identify a large set of macrophage-enriched genes, many of which have obvious functions in endocytosis and phagocytosis. In addition, a significant number of LPS-inducible genes were identified. The data suggest that macrophages are a complex source of mRNA for transcriptome studies. To assess complexity and identify additional macrophage expressed genes, cDNA libraries were created from purified populations of macrophage and dendritic cells, a functionally related cell type. Sequence analysis revealed a high incidence of novel mRNAs within these cDNA libraries. These studies provide insights into the depths of transcriptional complexity still untapped amongst products of inducible genes, and identify macrophage and dendritic cell populations as a starting point for sampling the inducible mammalian transcriptome.

Apoptotic cell removal in development and tissue homeostasis

Cell deletion is a physiological process for the development and maintenance of tissue homeostasis in metazoa. This is mainly achieved by the induction of various forms of programmed cell death followed by the recognition and removal of the targeted cells by phagocytes. In this review, we will discuss cell deletion in relation to the development and function of the innate immune system, particularly of the mononuclear phagocyte system (MPS), its ontogeny and potential role in tissue remodeling in the embryo and adult. Ongoing studies are addressing the roles of professional phagocytes of the MPS and neighboring tissue cells in dying cell removal, and candidate molecules that might attract mononuclear phagocytes to the dying cells. The potential phagocyte must discriminate between living and dying cells; current concepts for this discrimination derive from the observation of newly exposed ligands on the dying cells and new evidence for direct inhibition of uptake by viable cells.

Transcriptional network dynamics in macrophage activation

Transcriptional regulatory networks govern cell differentiation and the cellular response to external stimuli. However, mammalian model systems have not yet been accessible for network analysis. Here, we present a genome-wide network analysis of the transcriptional regulation underlying the mouse macrophage response to bacterial lipopolysaccharide (LPS). Key to uncovering the network structure is our combination of time-series cap analysis of gene expression with in silico prediction of transcription factor binding sites. By integrating microarray and qPCR time-series expression data with a promoter analysis, we find dynamic subnetworks that describe how signaling pathways change dynamically during the progress of the macrophage LPS response, thus defining regulatory modules characteristic of the inflammatory response. In particular, our integrative analysis enabled us to suggest novel roles for the transcription factors ATF-3 and NRF-2 during the inflammatory response. We believe that our system approach presented here is applicable to understanding cellular differentiation in higher eukaryotes. (c) 2006 Elsevier Inc. All rights reserved.

TFEC is a macrophage-restricted member of the microphthalmia-TFE subfamily of basic helix-loop-helix leucine zipper transcription factors

The murine homologue of the TFEC was cloned as part of an analysis of the expression of the microphthalmia-TFE (MiT) subfamily of transcription factors in macrophages. TFEC, which most likely acts as a transcriptional repressor in heterodimers with other MiT family members, was identified in cells of the mononuclear phagocyte lineage, coexpressed,vith all other known MiT subfamily members (Mitf, TFE3, TFEB), Northern blot analysis of several different cell lineages indicated that the expression of murine TFEC (mTFEC) was restricted to macrophages. A 600-bp fragment of the TATA-less putative proximal promoter of TFEC shares features with many known macrophage-specific promoters and preferentially directs luciferase expression in the RAW264.7 macrophage cell line in transient transfection assays. Five of six putative Ets motifs identified in the TFEC promoter bind the macrophage-restricted transcription factor PU,I under in vitro conditions and in transfected 3T3 fibroblasts; the minimal luciferase activity of the TFEC promoter could be induced by coexpression of PU.1 or the related transcription factor Ets-2. The functional importance of the tissue-restricted expression of TFEC and a possible role in macrophage-specific gene regulation require further investigation, but are likely to be linked to the role of the other MiT family members in this lineage.

Regulation of CSF-1 receptor expression

Cells of the mononuclear phagocyte lineage possess receptors for macrophage colony-stimulating factor (CSF-1) encoded by the c-fms protooncogene and respond to CSF-1 with increased survival, growth, differentiation, and reversible changes in function. The c-fms gene is itself a macrophage differentiation marker. In whole mount analyses of mRNA expression in embryos, c-fms is expressed at very high levels on placental trophoblasts. It is detectable on individual cells in the yolk sac around 8.5 to 9 days postcoitus, appears on isolated cells in the head of the embryo around 9.5 dpc, and appears on numerous cells throughout the embryo by day 10.5. The extent of c-fms expression is much greater than for other macrophage-specific genes including lysozyme and a macrophage-specific protein tyrosine phosphatase. Our studies of the cis-acting elements of the c-fms promoter have indicated a key role for collaboration between the macrophage-specific transcription factor, Pu.1, which functions in determining the site of transcription initiation, and other members of the Ets transcription factor family. This is emerging as a common pattern in macrophage-specific promoters. We have shown that two PU box elements alone can function as a macrophage-specific promoter. The activity of both the artifical promoter and the c-fms promoter is activated synergistically by coexpression of Pu.1 and another Ets factor, c-Ets-2. A 3.5kb c-fms exon 2 promoter (but not the 300bp proximal promoter) is also active in a wide diversity of tumor cell lines. The interesting exception is the melanoma cell line K1735, in which the promoter is completely shut down and expression of c-fms causes growth arrest and cell death. The activity of the exon 2 promoter in these nonmacrophages is at least as serum responsive as the classic serum-responsive promoter of the c-fos gene. It is further inducible in nonmacrophages by coexpression of the c-fms product. Unlike other CSF-1/c-fms-responsive promoters, the c-fms promoter is not responsive to activated Ras even when c-Ets-2 is coexpressed. In most lines, production of full length c-fms is prevented by a downstream intronic terminator, but in Lewis lung carcinoma, read-through does occur, and expression of both c-fms and other macrophage-specific genes such as lysozyme and urokinase becomes detectable in conditions of serum deprivation. (C) 1997 Wiley-Liss, Inc.

Differentiated dendritic cells expressing nuclear RelB are predominantly located in rheumatoid synovial tissue perivascular mononuclear cell aggregates

Objective. Differentiated dendritic cells (DC) and other antigen-presenting cells are characterized by the nuclear location of RelB, a member of the nuclear factor kappa B/Rel family. To characterize and enumerate differentiated DC in rheumatoid arthritis (RA) peripheral blood (PB), synovial fluid (SF), and synovial tissue (ST), the expression and location of RelB were examined. Methods. RelB protein expression and cellular location were determined in RA PB, SF, and ST by flow cytometry and immunohistochemical analysis of purified cells or formalin-fixed tissue. DNA-binding activity of RelB was determined by electrophoretic: mobility shift-Western immunoblotting assays. Results. Circulating RA PBDC resembled normal immature PBDC in that they did not express intracellular RelB protein. In RA ST serial sections, cells containing nuclear RelB (nRelB) were enriched in perivascular regions. A mean +/- SD of 84 +/- 10% of these cells were DC. The remaining nRelB+,HLA-DR+ cells comprised B cells and macrophages. Only 3% of sorted SFDC contained nRelB, However, RelB present in the nucleus of these SFDC was capable of binding DNA, and therefore capable of transcriptional activity. Conclusion. Circulating DC precursors differentiate and express RelB after entry into rheumatoid ST. Differentiated DC can thus be identified by immunohistochemistry in formalin-fixed ST. Signals for DC maturation may differ between RA ST and SF, resulting in nuclear location of RelB predominantly in ST. This is likely to have functional consequences for the DC in these sites.

A synchronous multimedia annotation system for secure collaboratories

In this paper, we describe the Vannotea system - an application designed to enable collaborating groups to discuss and annotate collections of high quality images, video, audio or 3D objects. The system has been designed specifically to capture and share scholarly discourse and annotations about multimedia research data by teams of trusted colleagues within a research or academic environment. As such, it provides: authenticated access to a web browser search interface for discovering and retrieving media objects; a media replay window that can incorporate a variety of embedded plug-ins to render different scientific media formats; an annotation authoring, editing, searching and browsing tool; and session logging and replay capabilities. Annotations are personal remarks, interpretations, questions or references that can be attached to whole files, segments or regions. Vannotea enables annotations to be attached either synchronously (using jabber message passing and audio/video conferencing) or asynchronously and stand-alone. The annotations are stored on an Annotea server, extended for multimedia content. Their access, retrieval and re-use is controlled via Shibboleth identity management and XACML access policies.

Biomechanical consequences of impairment: A taxonomically valid basis for classification in a unified disability athletics system

Developing a unified classification system to replace four of the systems currently used in disability athletics (i.e., track and field) has been widely advocated. The diverse impairments to be included in a unified system require severed assessment methods, results of which cannot be meaningfully compared. Therefore, the taxonomic basis of current classification systems is invalid in a unified system. Biomechanical analysis establishes that force, a vector described in terms of magnitude and direction, is a key determinant of success in all athletic disciplines. It is posited that all impairments to be included in a unified system may be classified as either force magnitude impairments (FMI) or force control impairments (FCI). This framework would provide a valid taxonomic basis for a unified system, creating the opportunity to decrease the number of classes and enhance the viability of disability athletics.

DR-NEGOTIATE - A system for automated agent negotiation with defeasible logic-based strategies

This paper reports on a system for automated agent negotiation, based on a formal and executable approach to capture the behavior of parties involved in a negotiation. It uses the JADE agent framework, and its major distinctive feature is the use of declarative negotiation strategies. The negotiation strategies are expressed in a declarative rules language, defeasible logic, and are applied using the implemented system DR-DEVICE. The key ideas and the overall system architecture are described, and a particular negotiation case is presented in detail.

Probabilistic Action System Trace Semantics

Action systems are a construct for reasoning about concurrent, reactive systems, in which concurrent behaviour is described by interleaving atomic actions. Sere and Troubitsyna have proposed an extension to action systems in which actions may be expressed and composed using discrete probabilistic choice as well as demonic nondeterministic choice. In this paper we develop a trace-based semantics for probabilistic action systems. This semantics provides a simple theoretical base on which practical refinement rules for probabilistic action systems may be justified.

The emergence of the determiner system in Mauritian Creole: a syntax semantics mapping

This paper describes the emergence of new functional items in the Mauritian Creole noun phrase, following the collapse of the French determiner system when superstrate and substrate came into contact. The aim of the paper is to show how the new language strived to express the universal semantic contrasts of (in)definiteness and singular vs. plural. The process of grammaticalization of new functional items in the determiner system was accompanied by changes in the syntax from French to creole. An analysis within Chomsky’s Minimalist framework (1995, 2000, 2001) suggests that these changes were driven by the need to map semantic features onto the syntax.