The extracellular matrix (ECM) provides structural support for tissue architecture and is a major effector of cell behavior during skin repair and inflammation. Macrophages are involved in all stages of skin repair but only limited knowledge exists about macrophage-specific expression and regulation of ECM components. In this study, we used transcriptome profiling and bioinformatic analysis to define the unique expression of ECM-associated genes in cultured macrophages. Characterization of the matrisome revealed that most genes were constitutively expressed and that several genes were uniquely regulated upon interferon gamma (IFNγ) and dexamethasone stimulation. Among those core matrisome and matrisome-associated components transforming growth factor beta (TGFβ)-induced, matrix metalloproteinase 9 (MMP9), elastin microfibril interfacer (EMILIN)-1, netrin-1 and gliomedin were also present within the wound bed at time points that are characterized by profound macrophage infiltration. Hence, macrophages are a source of ECM components in vitro as well as during skin wound healing, and identification of these matrisome components is a first step to understand the role and therapeutic value of ECM components in macrophages and during wound healing.

Background: During vertebrate embryogenesis the initial stages of bone formation by endochondral ossification involve the aggregation and proliferation of mesenchymal cells into condensations. Continued growth of the condensations and differentiation of the mesenchymal cells into chondrocytes results in the formation of cartilage templates, or anlagen, which prefigure the shape of the future bones. The chondrocytes in the anlagen further differentiate by undergoing a complex sequence of maturation and hypertrophy, and are eventually replaced by mineralized bone. Regulation of the onset of chondrogenesis is incompletely understood, and would be informed by comprehensive analyses of in vivo gene expression. Results: Tibial and fibular pre-condensed mesenchyme was microdissected from mouse hind limbs at 11.5 dpc, and the corresponding condensations at 12.5 dpc and cartilage anlagen at 13.5 dpc. Total RNA was isolated, and cRNA generated by linear amplification was interrogated using mouse whole genome microarrays. Differential expression was validated by quantitative PCR for Agc1, Bmp8a, Col2a1, Fgfr4, Foxa3, Gdf5, Klf2, Klf4, Lepre1, Ncad, Sox11, and Trpv4. Further, independent validation of the microarray data was achieved by in situ hybridization to analyse the expression of Lepre1, Pcdh8, Sox11, and Trpv4 from 11.5 dpc to 13.5 dpc during mouse hind limb development. We found significant differential expression of 931 genes during these early stages of chondrogenesis. Of these, 380 genes were down-regulated and 551 up-regulated. Our studies characterized the expression pattern of gene families previously associated with chondrogenesis, such as adhesion molecules, secreted signalling molecules, transcription factors, and extracellular matrix components. Gene ontology approaches identified 892 differentially expressed genes not previously identified during the initiation of chondrogenesis. These included several Bmp, Gdf, Wnt, Sox and Fox family members. Conclusion: These data represent the first global gene expression profiling analysis of chondrogenic tissues during in vivo development. They identify genes for further study on their functional roles in chondrogenesis, and provide a comprehensive and important resource for future studies on cartilage development and disease.

Identification and clearance of apoptotic cells prevents the release of harmful cell contents thereby suppressing inflammation and autoimmune reactions. Highly conserved annexins may modulate the phagocytic cell removal by acting as bridging molecules to phosphatidylserine, a characteristic phagocytosis signal of dying cells. In this study five members of the structurally and functionally related annexin family were characterized for their capacity to interact with phosphatidylserine and dying cells. The results showed that AnxA3, AnxA4, AnxA13, and the already described interaction partner AnxA5 can bind to phosphatidylserine and apoptotic cells, whereas AnxA8 lacks this ability. Sequence alignment experiments located the essential amino residues for the recognition of surface exposed phosphatidylserine within the calcium binding motifs common to all annexins. These amino acid residues were missing in the evolutionary young AnxA8 and when they were reintroduced by site directed mutagenesis AnxA8 gains the capability to interact with phosphatidylserine containing liposomes and apoptotic cells. By defining the evolutionary conserved amino acid residues mediating phosphatidylserine binding of annexins we show that the recognition of dying cells represent a common feature of most annexins. Hence, the individual annexin repertoire bound to the cell surface of dying cells may fulfil opsonin-like function in cell death recognition.

Axial growth of long bones occurs through a coordinated process of growth plate chondrocyte proliferation and differentiation. This maturation of chondrocytes is reflected in a zonal change in gene expression and cell morphology from resting to proliferative, prehypertrophic, and hypertrophic chondrocytes of the growth plate followed by ossification. A major experimental limitation in understanding growth plate biology and pathophysiology is the lack of a robust technique to isolate cells from the different zones, particularly from small animals. Here, we report on a new strategy for separating distinct chondrocyte populations from mouse growth plates. By transcriptome profiling of microdissected zones of growth plates, we identified novel, zone-specific cell surface markers and used these for flow cytometry and immunomagnetic cell separation to quantify, enrich, and characterize chondrocytes populations with respect to their differentiation status. This approach provides a novel platform to study cartilage development and characterize mouse growth plate chondrocytes to reveal unique cellular phenotypes of the distinct subpopulations within the growth plate. © 2010 American Society for Bone and Mineral Research