Cytokines and proteoglycans

Cytokines play an important regulatory role in the metabolism of proteoglycans. Proteoglycans are found in plasma membranes, but predominantly in the extra-cellular matrix. In the latter they are quantitatively and qualitatively essential components. Especially in a tissue like cartilage without any blood vessels, the cells are dependent on cytokines for the communication among themselves in the extra-cellular matrix and also for communication with the outside world. Various cytokines have been found to be able to penetrate the extra-cellular matrix and inhibit, respectively stimulate the proteoglycan synthesis. Also, the degradation of proteoglycans can be stimulated, respectively inhibited by several cytokines. In addition, some cytokines have been found which regulate the effects of the other cytokines. With respect to proteoglycan metabolism a complex cytokine network is emerging.Furthermore it is becoming increasingly clear that proteoglycans are connected to the cytokine network by their own bioactive functions. First, they possibly possess cytokine activities themselves. Second, they can function as receptors, protectors, inactivators and storage ligands for cytokines. So the proteoglycans are clearly involved in the feedback signalling from the extra-cellular matrix to the cells that are synthesizing this extra-cellular matrix. Together with agonistic or antagonistic cytokines they are involved in the regulation of proteoglycan turnover during balanced or unbalanced metabolism in normal, respectively pathological situations.     

A discoidin domain receptor 1 knock-out mouse as a novel model for osteoarthritis of the temporomandibular joint

Discoidin domain receptor 1 (DDR-1)-deficient mice exhibited a high incidence of osteoarthritis (OA) in the temporomandibular joint (TMJ) as early as 9 weeks of age. They showed typical histological signs of OA, including surface fissures, loss of proteoglycans, chondrocyte cluster formation, collagen type I upregulation, and atypical collagen fibril arrangements. Chondrocytes isolated from the TMJs of DDR-1-deficient mice maintained their osteoarthritic characteristics when placed in culture. They expressed high levels of runx-2 and collagen type I, as well as low levels of sox-9 and aggrecan. The expression of DDR-2, a key factor in OA, was increased. DDR-1-deficient chondrocytes from the TMJ were positively influenced towards chondrogenesis by a three-dimensional matrix combined with a runx-2 knockdown or stimulation with extracellular matrix components, such as nidogen-2. Therefore, the DDR-1 knock-out mouse can serve as a novel model for temporomandibular disorders, such as OA of the TMJ, and will help to develop new treatment options, particularly those involving tissue regeneration.     

Extracellular matrix components associated with remodeling processes in brain

In the central nervous system, various extracellular matrix components have been identified which are strongly expressed during development and in most areas of the brain down-regulated during maturation. Examples are tenascin-C, neurocan and hyaluronan. While tenascin-C is well known to be associated with morphogenic events and the active contribution of hyaluronan to various physiological processes is increasingly acknowledged, neurocan belongs to a class of molecules thought to be generally more associated with barrier functions: chondroitin sulfate proteoglycans. Consideration of these and related molecules and their processing in the context of the general organization of the brain extracellular matrix, their changes during brain maturation and their implication in different types of remodeling processes in adult brain, like normal and pathological synaptic plasticity, inflammatory and dementia-associated diseases and gliomas, may indicate that components of the extracellular matrix could provide valuable early information about the pathological state of the brain.Received 29 January 2004; received after revision 25 March 2004; accepted 2 April 2004     

The extracellular matrix of the hematopoietic microenvironment

The bone marrow microenvironment plays an important role in promoting hematopoietic progenitor cell proliferation and differentiation and the controlled egress of these developing hematopoietic cells. The establishment of long-term bone marrow cultures, which are thought to mimic hematopoiesis in vitro, and various stromal cell lines has greatly facilitated the analysis of the functions of this microenvironment. Extracellular matrix (ECM) molecules of all three categories (collagens, proteoglycans and glycoproteins) have been identified as part of this microenvironment and have been shown to be involved in, different biological functions such as cell adhesion and anti-adhesion, binding and presentation of various cytokines and regulation of cell growth. It is suggested that these matrix molecules in combination with cytokines are crucial for compartmentalization of the bone marrow. Although many cell adhesion molecules have been characterized on the surface of hematopoietic progenitor cells, the nature of cellular receptors for the ECM components is less well defined. During leukemia, many immature blood cells are released from bone marrow, but it is not yet known whether these abnormalities in hematopoiesis are also caused by an altered microenvironment or altered composition of its extracellular matrix. The elucidation of the involvement of specific ECM-isoforms and as yet not characterized ECM components and their receptors in the bone marrow will certainly help towards a better understanding of these phenomena.     

Noncollagenous, nonproteoglycan macromolecules of cartilage

Extracellular matrix comprises approximately 90% of cartilage, with collagens and proteoglycans making up the bulk of the tissue. In recent years, several abundant cartilage proteins that are neither collagens nor proteoglycans have been characterized in detail. The putative roles of these proteins range from involvement in matrix organization or matrix-cell signaling (PRELP, chondroadherin, cartilage oligomeric protein and cartilage matrix protein) through to molecules that are likely to be involved with modulation of the chondrocyte phenotype (CD-RAP, CDMPs, chondromodulin and pleiotrophin). Other molecules, such as the cartilage-derived C-type lectin and cartilage intermediate layer protein have no role as yet. Due to the difficulties associated with experimentally manipulating a tissue that is 90% extracellular matrix in a manner that can be readily transferred to the whole organism, many of these molecules have been focused on by a surprisingly small number of researchers. This review focuses on newly discovered proteins and glycoproteins in cartilage, with a bias towards those that have structural roles or that are unique to cartilage. Received 7 January 1999; accepted 11 March 1999     

Chromatin organization within nuclear blebs in leukocytes ofXenopus laevis

Summary The nuclei from leukocytes of peripheral blood, liver and spleen of an individual anaemicXenopus laevis have been found to possess numerous nuclear blebs or projections. These structures were found to be very variable in size and shape as viewed in electron micrographs, but commonly included an enclosed mass of cytoplasm bound on one side by a very thin section of nuclear material. Such sections are membrane bounded on each side and frequently display an interesting ordered array of chromatin.     

A method for distinction between RNA and DNA in aldehyde and osmiumtetroxide-fixed electron microscopic autoradiographs.

A simple method for distinction between RNA- and DNA-containing structures in aldehyde- and osmiumtextroxide-fixed electron microscopic autoradiographs (or ordinary thin sections) is described: the developer and the acetic acid used for processing autoradiographs extract selectively uranium acetate from DNA containing-structures which, after staining with lead citrate, leads to a characteristically 'bleached' appearance of the DNA.     

The extracellular matrix during heart development

The embryonic extracellular matrix, which is comprised of glycosaminoglycans, glycoproteins, collagens, and proteoglycans, is believed to play multiple roles during heart morphogenesis. Some of these ECM components appear throughout development, however, certain molecules exhibit an interesting transient spatial and temporal distribution. Due to significant new data that have been gathered predominantly in the past 10 years, a comprehensive review of the literature is needed. The intent of this review is to highlight work that addresses mechanisms by which extracellular matrix influences vertebrate heart development.     

Lecticans: organizers of the brain extracellular matrix

Lecticans are a family of chondroitin sulfate proteoglycans, encompassing aggrecan, versican, neurocan and brevican. These proteoglycans are characterized by the presence of ahyaluronan-binding domain and a C-type lectin domain in their core proteins. Through these domains, lecticans interact with carbohydrate and protein ligands in the extracellular matrix and act as linkers of these extracellular matrix molecules. In adult brain, lecticans are thought to interact with hyaluronan and tenascin-R to form a ternary complex. We propose that the hyaluronan-lectican-tenascin-R complex constitutes the core assembly of the adult brain extracellular matrix, which is found mainly in pericellular spaces of neurons as ‘perineuronal nets’. Received 27 September 1999; accepted 26 October 1999     

Independent modulation of collagen fibrillogenesis by decorin and lumican

The leucine-rich proteoglycans (also known as "small, leucine-rich proteoglycans," or SLRPs) lumican and decorin are thought to be involved in the regulation of collagen fibril assembly. Preparation of these proteoglycans in chemical amounts without exposure to denaturants has recently been achieved by infecting HT-1080 cells with vaccinia virus that contains an expression cassette for these molecules. Addition of lumican and decorin to a collagen fibrillogenesis assay based on turbidity demonstrated that lumican accelerated initial fibril formation while decorin retarded initial fibril formation. At the end of fibrillogenesis, both proteoglycans resulted in an overall reduced turbidity, suggesting that fibril diameter was lower. The presence of both proteoglycans had a synergistic effect, retarding fibril formation to a greater degree than either proteoglycan individually. Competitive binding studies showed that lumican did not compete for decorin-binding sites on collagen fibrils. Both proteoglycans increased the stability of fibrils to thermal denaturation to approximately the same degree. These studies show that lumican does not compete for decorin-binding sites on collagen, that decorin and lumican modulate collagen fibrillogenesis, and that, in the process, they also enhance collagen fibril stability.     

Investigation of the microstructure of kidney stones (oxalate type) by high voltage electron microscopy and electron diffraction

Summary The organic matrix of this type of urinary calculus contains 3 components, which differ in form and in amorphous/crystalline content. Deposition of crystalline material in the early stages of mineralisation seems to be epitaxially related to the orientation of the organic matrix.Acknowledgments. The authors are indebted to Ass. Prof. N. Allouba of Ein-Shams University (Cairo) for supplying the kidney stones and medical information, and to the Physiology and Zoology Departments of Cambridge University for assistance in cutting the sections. The work was performed during the tenure by K. E. S. of a research bursary from the British Council.     

Protein and lysozyme content of adult human nucleus pulposus

A radiologically normal human nucleus pulposus was extracted with 4 M guanidinium chloride and the non-collagenous proteins separated from the proteoglycans by dissociative density gradient centrifugation. Lysozyme was identified as a matrix constituent of the normal, mature human nucleus pulposus.     

Syndecan family of cell surface proteoglycans: Developmentally regulated receptors for extracellular effector molecules

Syndecans are a family of integral membrane proteoglycans with conserved membrane-spanning and intracellular domains but with structurally distinct extracellular domains (ectodomains). They are known to function as heparan sulphate co-receptors in fibroblast growth factor signalling as well as to link cells directly to the extracellular matrix. These and other biological activities of syndecans involve specific interactions of the heparan sulphate side chains of syndecans with cytokines and extracellular matrix proteins. Four different vertebrate syndecans, designated as syndecans 1–4 (or syndecan, fibroglycan, N-syndecan and amphiglycan, respectively), are known. During embryonic development, syndecans have specific and highly regulated expression patterns that are distinct from the expression in adult tissue, suggesting an active role in morphogenetic processes. The developmental expression of syndecans is particularly intense in mesenchymal condensates and at epithelium mesenchyme interfaces, where a number of heparan sulphate-binding cytokines and matrix components are also expressed in a regulated manner, ofter spatially and temporally co-ordinated with the syndecan expression. Recent evidence indicates that the regulation of heparan sulphate fine structure (mainly the number and arrangement of sulphate groups along the polymer) provides a mechanism for the cellular control of syndecan-protein interactions. Furthermore, morphogenetically active cytokines such as fibroblast growth factor-2 and transforming growth factor-β participate in the regulation of syndecan expression and glycosaminoglycan structure. This review discusses the developmental expression and binding functions of syndecans as well as the molecular regulation of specific heparan sulphate-protein interactions.     

Regulation of glycosaminoglycan structure and atherogenesis

Cardiovascular disease is the major cause of premature death in modern society, and its impact is increasing due to rising rates of obesity and type 2diabetes. Clinical studies based on targeting metabolic abnormalities and biomarkers demonstrate significant benefits, but always an element of disease remains which is resistant to treatment. Recent evidence has strongly implicated an early interaction of atherogenic lipoproteins with vascular matrix proteoglycans as the initiating step in atherogenesis. Expert commentary has pointed to the need for vascular directed therapies to provide reductions in the residual disease component. We propose that the regulation of synthesis and thus structure of glycosaminoglycans on proteoglycans provides a potential pathway to this reduction. We review existing evidence that the vascular synthesis of glycosaminoglycan chains can be regulated in a manner which reduces lipoprotein binding and the potential application of this strategy to attenuation of the current cardiovascular disease pandemic.Received 21 October 2003; received after revision 16 December 2003; accepted 29 December 2003     

Protein and lysozyme content of adult human nucleus pulposus

Summary A radiologically normal human nucleus pulposus was extracted with 4 M guanidinium chloride and the non-collagenous proteins separated from the proteoglycans by dissociative density gradient centrifugation. Lysozyme was identified as a matrix constituent of the normal, mature human nucleus pulposus.Supported by Public Health Service Training Grant ES-07088.     

Regulatory peptide immunocytochemistry at light- and electron microscopical levels

Summary Immunocytochemical techniques applied at both light- and electron microscopical levels are valuable in the study of regulatory peptide distribution in normal and diseased tissue, whether in the form of sections or whole cell preparations. Successful immunolocalisation depends on 1) adequate preservation of the peptide antigen and the tissue structure in which it resides; 2) a suitably specific and sensitive labelled antibody detecting system. In general, peptides are stable molecules, most of which retain their antigenicity after conventional cross-linking fixation and tissue processing, allowing standard immunocytochemical methods to be used for light- and electron microscopy. Regulatory peptides are derived from precursor molecules and several families of structurally similar peptides are now generally recognised. Region-specific antibodies may be needed to overcome problems of cross-reactivity or to identify a bioactive form in the presence of its precursor. Multiple co-localisation of different related and unrelated peptides in the same cell or even storage granule is now recognised and can be identified by immunocytochemistry.     

Isolation and purification of proteoglycans

Purification of a protein typically involves development of a quantitative assay to track protein integrity (e.g. enzyme activity) during subsequent isolation steps. The generalized procedure involves choosing the source of the protein, defining extraction conditions, developing bulk purification methods followed by refined, more selective methods. The purification of proteoglycans is often complicated by a) limited source quantities, b) necessity of chaotropic solvents for efficient extraction, c) their large molecular size and d) lack of defined functions to enable purity (i.e. activity, conformation) to be assessed. Because the usual goal of proteoglycan purification is physical characterization (intact molecular weight, core protein and glycosaminoglycan class and size), the problems of a suitable assay and/or native conformation are avoided. The assay for tracking proteoglycan isolation typically utilizes uronic acid content or radiolabel incorporation as a marker. Once extracted from their cellular/extracellular environment, proteoglycans can be isolated by density gradient centrifugation and/or column chromatography techniques. Recent advances in the composition of chromatographic supports have enabled the application of ion-exchange, gel permeation, hydrophobic interaction and affinity chromatography resins using efficient high-pressure liquid chromatography to proteoglycan purification.     

Permeable embryonic cells of sea urchins as a model for studying nucleus-cytoplasm interactions

Sea urchin embryonic cells were made permeable by treating them with glycerol solution for the purpose of exchanging cytoplasmic components. When proteoglycans prepared from embryos of advanced stages of development permeated into these glycerinated cells, these substances accumulated rapidly in the nucleus, being bound with chromatin.     

Circular proteoglycans from sponges: first members of the spongican family

Species-specific cell adhesion in marine sponges is mediated by a new family of modular proteoglycans whose general supramolecular structure resembles that of hyalectans. However, neither their protein nor their glycan moieties have significant sequence homology to other proteoglycans, despite having protein subunits equivalent to link proteins and to proteoglycan monomer core proteins, and glycan subunits equivalent to hyaluronan and to the glycosaminoglycans of hyalectans. In some species, these molecular components are assembled into a structure with a circular core formed by the link protein- and hyaluronan-like subunits. Besides their involvement in cell adhesion, these sponge proteoglycans, for which we propose the term spongicans, participate in signal transduction processes and are suspected to play a role in sponge self-nonself recognition. Their in vivo roles and the mild methods used to purify large amounts of functionally active spongicans make them ideal models to study the functions and possible new applications of proteoglycans in biomedical research. Received 21 May 2002; received after revision 5 July 2002; accepted 10 July 2002 RID="*" ID="*"Corresponding author.     

Multifunctionality of extracellular and cell surface heparan sulfate proteoglycans

Heparan sulfate proteoglycans are a remarkably diverse family of glycosaminoglycan-bearing protein cores that include the syndecans, the glypicans, perlecan, agrin, and collagen XVIII. Members of this protein class play key roles during normal processes that occur during development, tissue morphogenesis, and wound healing. As key components of basement membranes in organs and tissues, they also participate in selective filtration of biological fluids, in establishing cellular barriers, and in modulation of angiogenesis. The ability to perform these functions is provided both by the features of the protein cores as well as by the unique properties of heparan sulfate, which is assembled as a polymer of N-acetylglucosamine and glucuronic acid and modified by specific enzymes to generate specialized biologically active structures. This article discusses the structures and functions of this amazing family of proteoglycans and provides a platform for further study of the individual members.