A new mouse mutant with cleavage-resistant versican and isoform-specific versican mutants demonstrate that proteolysis at the Glu-Ala peptide bond in the V1 isoform is essential for interdigital web regression.

Two inherent challenges in the mechanistic interpretation of protease-deficient phenotypes are defining the specific substrate cleavages whose reduction generates the phenotypes and determining whether the phenotypes result from loss of substrate function, substrate accumulation, or loss of a function(s) embodied in the substrate fragments. Hence, recapitulation of a protease-deficient phenotype by a cleavage-resistant substrate would stringently validate the importance of a proteolytic event and clarify the underlying mechanisms. Versican is a large proteoglycan required for development of the circulatory system and proper limb development, and is cleaved by ADAMTS proteases at the Glu-Ala peptide bond located in its alternatively spliced GAGβ domain.

Determinants of versican-V1 proteoglycan processing by the metalloproteinase ADAMTS5.

Proteolysis of the Glu(441)-Ala(442) bond in the glycosaminoglycan (GAG) β domain of the versican-V1 variant by a disintegrin-like and metalloproteinase domain with thrombospondin type 1 motif (ADAMTS) proteases is required for proper embryo morphogenesis. However, the processing mechanism and the possibility of additional ADAMTS-cleaved processing sites are unknown. We demonstrate here that if Glu(441) is mutated, ADAMTS5 cleaves inefficiently at a proximate upstream site but normally does not cleave elsewhere within the GAGβ domain.

Imbalanced expression of Vcan mRNA splice form proteins alters heart morphology and cellular protein profiles.

The fundamental importance of the proteoglycan versican to early heart formation was clearly demonstrated by the Vcan null mouse called heart defect (hdf). Total absence of the Vcan gene halts heart development at a stage prior to the heart's pulmonary/aortic outlet segment growth. This creates a problem for determining the significance of versican's expression in the forming valve precursors and vascular wall of the pulmonary and aortic roots.

Nogo receptor homolog NgR2 expressed in sensory DRG neurons controls epidermal innervation by interaction with Versican.

Primary sensory afferents of the dorsal root ganglion (DRG) that innervate the skin detect a wide range of stimuli, such as touch, temperature, pain, and itch. Different functional classes of nociceptors project their axons to distinct target zones within the developing skin, but the molecular mechanisms that regulate target innervation are less clear. Here we report that the Nogo66 receptor homolog NgR2 is essential for proper cutaneous innervation.

Three mechanisms assemble central nervous system nodes of Ranvier.

Rapid action potential propagation in myelinated axons requires Na⁺ channel clustering at nodes of Ranvier. However, the mechanism of clustering at CNS nodes remains poorly understood. Here, we show that the assembly of nodes of Ranvier in the CNS involves three mechanisms: a glia-derived extracellular matrix (ECM) complex containing proteoglycans and adhesion molecules that cluster NF186, paranodal axoglial junctions that function as barriers to restrict the position of nodal proteins, and axonal cytoskeletal scaffolds (CSs) that stabilize nodal Na⁺ channels.

A novel strategy for a splice-variant selective gene ablation: the example of the versican V0/V2 knockout.

The complete knockout of genes that give rise to alternative splice products can often provide only an integral view of the dominant function(s) of all the isoforms they encode. If one of these isoforms is indispensable for life, a constitutive and complete inactivation may even preclude any in vivo studies of later expressed splice-variants in mice. To explore function of the tissue-restricted versican V2 isoform during central nervous system maturation, for instance, we had to circumvent the early embryonic lethality of the complete knockout by employing a novel splice-variant-specific gene ablation approach.

Versican V0 and V1 direct the growth of peripheral axons in the developing chick hindlimb.

Peanut agglutinin-binding disaccharides and chondroitin sulfate mark transient mesenchymal barriers to advancing motor and sensory axons innervating the hindlimbs during chick development. Here we show that the vast majority of these carbohydrates are at the critical stage and location attached to the versican splice variants V0 and V1. We reveal that the isolated isoforms of this extracellular matrix proteoglycan suppress axon extension at low concentrations and induce growth cone collapse and rapid retraction at higher levels.

Oligodendrocyte myelin glycoprotein does not influence node of ranvier structure or assembly.

Oligodendrocyte myelin glycoprotein (OMgp) is expressed by both neurons and oligodendrocytes in the CNS. It has been implicated in growth cone collapse and neurite outgrowth inhibition by signaling through the Nogo receptor and paired Ig-like receptor B (PirB). OMgp was also reported to be an extracellular matrix (ECM) protein surrounding CNS nodes of Ranvier and proposed to function as (1) an inhibitor of nodal collateral sprouting and (2) an important contributor to proper nodal and paranodal architecture.

Age-related differences in human skin proteoglycans.

Previous work has shown that versican, decorin and a catabolic fragment of decorin, termed decorunt, are the most abundant proteoglycans in human skin. Further analysis of versican indicates that four major core protein species are present in human skin at all ages examined from fetal to adult. Two of these are identified as the V0 and V1 isoforms, with the latter predominating.

Versican V2 assembles the extracellular matrix surrounding the nodes of ranvier in the CNS.

The CNS-restricted versican splice-variant V2 is a large chondroitin sulfate proteoglycan incorporated in the extracellular matrix surrounding myelinated fibers and particularly accumulating at nodes of Ranvier. In vitro, it is a potent inhibitor of axonal growth and therefore considered to participate in the reduction of structural plasticity connected to myelination. To study the role of versican V2 during postnatal development, we designed a novel isoform-specific gene inactivation approach circumventing early embryonic lethality of the complete knock-out and preventing compensation by the remaining versican splice variants.

Extracellular matrix of the central nervous system: from neglect to challenge.

The basic concept, that specialized extracellular matrices rich in hyaluronan, chondroitin sulfate proteoglycans (aggrecan, versican, neurocan, brevican, phosphacan), link proteins and tenascins (Tn-R, Tn-C) can regulate cellular migration and axonal growth and thus, actively participate in the development and maturation of the nervous system, has in recent years gained rapidly expanding experimental support. The swift assembly and remodeling of these matrices have been associated with axonal guidance functions in the periphery and with the structural stabilization of myelinated fiber tracts and synaptic contacts in the maturating central nervous system. Particular interest has been focused on the putative role of chondroitin sulfate proteoglycans in suppressing central nervous system regeneration after lesions.

Inter-laboratory validation of PCR-based HPV detection in pathology specimens.

The detection and typing of human papilloma virus (HPV) in pathology specimens is gaining increasingly in importance. In the context of the initiative for quality assurance in pathology (QuIP) of the German Society of Pathology and the Professional Association of German Pathologists, four panel laboratories with experience and expertise in polymerase chain reaction (PCR)-based HPV detection were selected to establish an inter-laboratory trial. In a first step, these laboratories performed an internal testing of their own methodologies, which comprised DNA sequencing, multiplex nested PCR and hybridization techniques.

Changes in steroid receptors and proteoglycan expression in the guinea pig prostate stroma during puberty and hormone manipulation.

Background: Proteoglycans are structural and informational molecules important during embryogenesis and organ maturation. Maturation of the prostate is influenced by androgens and estrogens, but changes in the relative spatiotemporal expression of steroid receptors and proteoglycans during hormonal change are unexplored.

Methods: Guinea pig prostate was used to define hormone-induced changes in the expression of androgen (AR) and estrogen (ER(alpha)) receptors, chondroitin sulfate (CS) glycosaminoglycan and core proteins of versican and syndecan-1.

Neural crest-derived cells with stem cell features can be traced back to multiple lineages in the adult skin.

Given their accessibility, multipotent skin-derived cells might be useful for future cell replacement therapies. We describe the isolation of multipotent stem cell-like cells from the adult trunk skin of mice and humans that express the neural crest stem cell markers p75 and Sox10 and display extensive self-renewal capacity in sphere cultures. To determine the origin of these cells, we genetically mapped the fate of neural crest cells in face and trunk skin of mouse.

Lesion-induced differential expression and cell association of Neurocan, Brevican, Versican V1 and V2 in the mouse dorsal root entry zone.

Lack of regeneration in the CNS has been attributed to many causes, including the presence of inhibitory molecules such as chondroitin sulfate proteoglycans (CSPGs). However, little is known about the contribution of CSPGs to regeneration failure in vivo, in particular at the dorsal root entry zone (DREZ), a unique CNS region that blocks regeneration of sensory fibers following dorsal root injury without glial scar formation. The goal of the present study was to evaluate the presence, regulation, and cellular identity of the proteoglycans Brevican, Neurocan, Versican V1 and Versican V2 in the DREZ using CSPG-specific antibodies and nucleic acid probes.

High frequency of t(14;18)-translocation breakpoints outside of major breakpoint and minor cluster regions in follicular lymphomas: improved polymerase chain reaction protocols for their detection.

The detection of t(14;18) translocations is widely used for the diagnosis and monitoring of follicular lymphomas displaying a high prevalence for this aberration. Cytogenetics, Southern blotting, and polymerase chain reaction (PCR) are commonly used techniques. It is generally believed that the vast majority of the breakpoints occurs on chromosome 18 in the major breakpoint region (mbr) and the minor cluster region (mcr).

Brain derived versican V2 is a potent inhibitor of axonal growth.

In this paper, we identify the chondroitin sulfate proteoglycan versican V2 as a major inhibitor of axonal growth in the extracellular matrix of the mature central nervous system. In immunohistochemical and in situ hybridization experiments we show that this tissue-specific splice variant of versican is predominantly present in myelinated fiber tracts of the brain and in the optic nerve, most likely being expressed by oligodendrocytes. We demonstrate that isolated versican V2 strongly inhibits neurite outgrowth of central and peripheral neurons in stripe-choice assays using laminin-1 as permissive substrate.

Versican V2 is a major extracellular matrix component of the mature bovine brain.

We have isolated and characterized the proteoglycan isoforms of versican from bovine brain extracts. Our approach included (i) cDNA cloning and sequencing of the entire open reading frame encoding the bovine versican splice variants; (ii) preparation of antibodies against bovine versican using recombinant core protein fragments and synthetic peptides; (iii) isolation of versican isoforms by ammonium sulfate precipitation followed by anion exchange and hyaluronan affinity chromatography; and (iv) characterization by SDS-polyacrylamide gel electrophoresis and Coomassie Blue staining or immunoblotting. Our results demonstrate that versican V2 is, together with brevican, a major component of the mature brain extracellular matrix.

[Molecular pathologic detection of mycobacteria].

In recent years, significant advances in the diagnosis of mycobacterial infections have been made by the introduction of direct pathogen detection methods. These techniques are usually based on the polymerase chain reaction (PCR) or on a transcription-mediated amplification (TMA) process. The majority of the protocols have been optimized for the detection of mycobacterial nucleic acids in fresh fluid or fresh tissue specimen.

Differential expression of versican isoforms in brain tumors.

Versican is a member of the family of large aggregating chondroitin sulfate proteoglycans which are one of the major constituents of brain extracellular matrix (ECM). We examined the expression of versican splice variants at mRNA and protein levels in normal human brain and in gliomas, medulloblastomas, schwannomas, neurofibromas, and meningiomas. RT-PCR revealed transcripts for V0 and V1 in all tissues.

Clonal disease in extracutaneous compartments in cutaneous T-cell lymphomas. A comparative study between cutaneous T-cell lymphomas and pseudo lymphomas.

Extracutaneous involvement is a sign of poor prognosis in cutaneous T-cell lymphomas (CTCL). Unfortunately it becomes clinically and histologically manifest only late in the course of the disease. It was the purpose of this study to detect clonality in peripheral blood, lymph nodes and bone marrow samples at times when extracutaneous involvement cannot otherwise be demonstrated.

Distribution of the large aggregating proteoglycan versican in adult human tissues.

We studied the distribution of the large hyaluronan-binding proteoglycan versican (also known as PG-M) in human adult tissues using affinity-purified polyclonal antibodies that recognize the core protein of the prominent versican splice variants VO and V1. Versican was present in the loose connective tissues of various organs and was often associated with the elastic fiber network. Furthermore, it was localized in most smooth muscle tissues and in fibrous and elastic cartilage.

Primary cutaneous T-cell-rich B-cell lymphoma. A case report with a 13-year follow-up.

Cutaneous B-cell lymphomas constitute approximately 20% of primary cutaneous lymphomas. Most histologic subtypes of nodal B-cell lymphomas also occur primarily in the skin. The recently described T-cell-rich B-cell lymphomas (TCRBCLs) manifest mainly in the lymph nodes.

Immunohistochemical and mutation analyses demonstrate that procollagen VII is processed to collagen VII through removal of the NC-2 domain.

Collagen VII is the major structural constituent of anchoring fibrils in the skin. It is synthesized as a procollagen that is larger than the collagen deposited in the tissue. In this study, we investigated the conversion of procollagen VII to collagen VII in human skin and in cutaneous cells in vitro and identified the propeptide using domain-specific antibodies.

Occult hepatosplenic T-gamma delta lymphoma. Value of genotypic analysis in the differential diagnosis.

We report on a patient with a rare hepatosplenic gamma delta T-cell lymphoma (gamma delta TCL) presenting clinically with B-symptoms, hepatosplenomegaly and pancytopenia. During the initial stage of the disease the sparse malignant cells could not be detected histologically. Furthermore, their identification was obscured by massive macrophage proliferation with haemophagocytosis in the spleen.