Anchorage of collagen-tailed acetylcholinesterase to the extracellular matrix is mediated by heparan sulfate proteoglycans

Heparan sulfate and heparin, two sulfated glycosaminoglycans (GAGs), extracted collagen-tailed acetylcholinesterase (AChE) from the extracellular matrix (ECM) of the electric organ of Discopyge tschudii. The effect of heparan sulfate and heparin was abolished by protamine; other GAGs could not extract the esterase. The solubilization of the asymmetric AChE apparently occurs through the formation of a soluble AChE-GAG complex of 30S. Heparitinase treatment but not chondroitinase ABC treatment of the ECM released asymmetric AChE forms. This provides direct evidence for the vivo interaction between asymmetric AChE and heparan sulfate residues of the ECM. Biochemical analysis of the electric organ ECM showed that sulfated GAGs bound to proteoglycans account for 5% of the total basal lamina. Approximately 20% of the total GAGs were susceptible to heparitinase or nitrous acid oxidation which degrades specifically heparan sulfates, and approximately 80% were susceptible to digestion with chondroitinase ABC, which degrades chondroitin-4 and -6 sulfates and dermatan sulfate. Our experiments provide evidence that asymmetric AChE and carbohydrate components of proteoglycans are associated in the ECM; they also indicate that a heparan sulfate proteoglycan is involved in the anchorage of the collagen-tailed AChE to the synaptic basal lamina.     

Immunological characterization of basement membrane types of heparan sulfate proteoglycan

Antibodies were raised against a small high-density and a large low-density form of heparan sulfate proteoglycan from a basement membrane-producing mouse tumor and were characterized by radioimmunoassays, immunoprecipitation and immunohistological methods. Antigenicity was due to the protein cores and included epitopes unique to the low density form as well as some shared by both proteoglycans. The antibodies did not cross-react with other basement membrane proteins or with chondroitin sulfate proteoglycans from interstitial connective tissues. The heparan sulfate proteoglycans occurred ubiquitously in embryonic and adult basement membranes and could be initially detected at the 2-4 cell stage of mouse embryonic development. Low levels were also found in serum. Biosynthetic studies demonstrated identical or similar proteoglycans in cultures of normal and carcinoembryonic cells and in organ cultures of fetal tissues. They could be distinguished from liver cell membrane heparan sulfate proteoglycan, indicating that the basement membrane types of proteoglycans represent a unique class of extracellular matrix proteins.     

Matrix-associated heparan sulfate proteoglycan: core protein-specific monoclonal antibodies decorate the pericellular matrix of connective tissue cells and the stromal side of basement membranes

Cultured human lung fibroblasts produce a large, nonhydrophobic heparan sulfate proteoglycan that accumulates in the extracellular matrix of the monolayer (Heremans, A., J. J. Cassiman, H. Van den Berghe, and G. David. 1988. J. Biol. Chem. 263: 4731-4739). A panel of four monoclonal antibodies, specific for four distinct epitopes on the 400-kD core protein of this extracellular matrix heparan sulfate proteoglycan, detects similar proteoglycans in human epithelial cell cultures. Immunohistochemistry of human tissues with the monoclonal antibodies reveals that these proteoglycans are concentrated at cell-matrix interfaces. Immunogold labeling of ultracryosections of human skin indicates that the proteoglycan epitopes are nonhomogeneously distributed over the width of the basement membrane. Immunochemical investigations and amino acid sequence analysis indicate that the proteoglycan from the fibroblast matrix shares several structural features with the large, low density heparan sulfate proteoglycan isolated from the Engelbreth-Holm-Swarm sarcoma. Thus, both epithelial cell sheets and individual mesenchymal cells accumulate a large heparan sulfate proteoglycan(s) at the interface with the interstitial matrix, where the proteoglycan may adopt a specific topological orientation with respect to this matrix.     

Heparin releasable and nonreleasable forms of heparan sulfate proteoglycan are found on the surfaces of cultured porcine aortic endothelial cells

Evidence suggests that endothelial cell layer heparan sulfate proteoglycans include a variety of different sized molecules which most likely contain different protein cores. In the present report, approximately half of endothelial cell surface associated heparan sulfate proteoglycan is shown to be releasable with soluble heparin. The remaining cell surface heparan sulfate proteoglycan, as well as extracellular matrix heparan sulfate proteoglycan, cannot be removed from the cells with heparin. The heparin nonreleasable cell surface proteoglycan can be released by membrane disrupting agents and is able to intercalate into liposomes. When the heparin releasable and nonreleasable cell surface heparan sulfate proteoglycans are compared, differences in proteoglycan size are also evident. Furthermore, the intact heparin releasable heparan sulfate proteoglycan is closer in size to proteoglycans isolated from the extracellular matrix and from growth medium than to that which is heparin nonreleasable. These data indicate that cultured porcine aortic endothelial cells contain at least two distinct types of cell surface heparan sulfate proteoglycans, one of which appears to be associated with the cells through its glycosaminoglycan chains. The other (which is more tightly associated) is probably linked via a membrane intercalated protein core.Abbreviations ECM extracellular matrix - HSPG heparan sulfate proteoglycan - PAE porcine aortic endothelial - PBS phosphate buffered saline     

Synthesis by Schwann cells of basal lamina and membrane-associated heparan sulfate proteoglycans

Primary cultures that contain only Schwann cells and sensory nerve cells synthesize basal lamina. The assembly of this basal lamina appears to be essential for normal Schwann cell development. In this study, we demonstrate that Schwann cells synthesize two major heparan sulfate-containing proteoglycans. Both proteoglycans band in dissociative CsCl gradients at densities less than 1.4 g/ml, and therefore, presumably, have relatively low carbohydrate-to-protein ratios. The larger of these proteoglycans elutes from Sepharose CL-4B in 4 M guanidine hydrochloride (GuHCl) at a Kav of 0.21 and contains heparan sulfate and chondroitin sulfate chains of Mr 21,000 in a ratio of approximately 3:1. This proteoglycan is extracted from cultures by 4 M GuHCl but not Triton X-100 and accumulates only when Schwann cells are actively synthesizing basal lamina. The smaller proteoglycan elutes from Sepharose CL-4B at a Kav of 0.44 and contains heparan sulfate and chondroitin sulfate chains of Mr 18,000 in a ratio of approximately 4:1. This proteoglycan is extracted by 4 M GuHCl or by Triton X-100. The accumulation of this proteoglycan is independent of basal lamina production.     

Structure and properties of an under-sulfated heparan sulfate proteoglycan synthesized by a rat hepatoma cell line

A rat hepatoma cell line was shown to synthesize heparan sulfate and chondroitin sulfate proteoglycans. Unlike cultured hepatocytes, the hepatoma cells did not deposit these proteoglycans into an extracellular matrix, and most of the newly synthesized heparan sulfate proteoglycans were secreted into the culture medium. Heparan sulfate proteoglycans were also found associated with the cell surface. These proteoglycans could be solubilized by mild trypsin or detergent treatment of the cells but could not be displaced from the cells by incubation with heparin. The detergent-solubilized heparan sulfate proteoglycan had a hydrophobic segment that enabled it to bind to octyl- Sepharose. This segment could conceivably anchor the molecule in the lipid interior of the plasma membrane. The size of the hepatoma heparan sulfate proteoglycans was similar to that of proteoglycans isolated from rat liver microsomes or from primary cultures of rat hepatocytes. Ion-exchange chromatography on DEAE-Sephacel indicated that the hepatoma heparan sulfate proteoglycans had a lower average charge density than the rat liver heparan sulfate proteoglycans. The lower charge density of the hepatoma heparan sulfate can be largely attributed to a reduced number of N-sulfated glucosamine units in the polysaccharide chain compared with that of rat liver heparan sulfate. Hepatoma heparan sulfate proteoglycans purified from the culture medium had a considerably lower affinity for fibronectin-Sepharose compared with that of rat liver heparan sulfate proteoglycans. Furthermore, the hepatoma proteoglycan did not bind to the neoplastic cells, whereas heparan sulfate from normal rat liver bound to the hepatoma cells in a time-dependent reaction. The possible consequences of the reduced sulfation of the heparan sulfate proteoglycan produced by the hepatoma cells are discussed in terms of the postulated roles of heparan sulfate in the regulation of cell growth and extracellular matrix formation.     

Extracellular matrix heparan sulfate modulates endothelial cell susceptibility to Staphylococcus aureus

The ability of extracellular matrix heparan sulfate to alter the susceptibility of human endothelial cells to S. aureus was investigated. Endothelial cells grown on extracellular matrix synthesized by S. aureus-infected endothelial cells were more susceptible to subsequent staphylococcal infection than endothelial cells grown on the extracellular matrix synthesized by untreated endothelial cells. Endothelial cells were more susceptible to S. aureus infection when (1) grown on heparitinase-treated extracellular matrix that removed heparan sulfate chains, (2) grown on extracellular matrix produced by chlorate-treated endothelial cells that reduced sulfation in the matrix heparan sulfate proteoglycans, (3) grown on heparan sulfate purified from extracellular matrix elaborated by infected endothelial cells, and (4) endothelial cells were chlorate-treated and therefore expressed desulfated cellular heparan sulfate proteoglycans. Extracellular matrix produced by S. aureus-infected endothelial cells contained heparan sulfate proteoglycans with reduced sulfation. The altered extracellular matrix with reduced sulfated heparan sulfate proteoglycans signalled the uninfected endothelial cells to produce under sulfated cellular heparan sulfate proteoglycans that increased S. aureus adherence to the endothelial cells. J. Cell. Physiol. 173:102–109, 1997. © 1997 Wiley-Liss, Inc.     

Transformed mouse mammary epithelial cells synthesize undersulfated basement membrane proteoglycan

Proteoglycans deposited in the basal lamina of [14C] glucosamine-labeled normal and [3H]glucosamine-labeled transformed mouse mammary epithelial cells grown on type I-collagen gels, were extracted in 4 M guanidinium chloride and cofractionated over Sepharose CL 4B. The heparan sulfate chains carried by these proteoglycans were isolated by treatment with alkaline borohydride, protease K, chondroitinase ABC, and cetylpyridinium chloride precipitation. Heparan sulfate isolated from transformed cell cultures consistently eluted from DEAE-cellulose at lower salt concentrations and was of smaller apparent Mr when chromatographed over Sepharose CL 6B, than heparan sulfate of normal cell cultures. Experiments using doubly labeled cultures ([3H]glucosamine and [35S]sulfate) demonstrated an approximately 30% reduction in the sulfate/hexosamine ratio in heparan sulfate derived from transformed cultures. Both N- and O-sulfate were decreased. The decreased Mr and decreased sulfation of heparan sulfate upon transformation appear sufficient to explain the altered heparan sulfate/chondroitin sulfate ratios previously observed in these cells. These changes may have implications for the molecular interactions in which these proteoglycans are normally engaged during basal lamina assembly, and cause the poor basal lamina formation displayed by these transformed cells.     

Heparan sulfate proteoglycans from mouse mammary epithelial cells. Basal extracellular proteoglycan binds specifically to native type I collagen fibrils

Mouse mammary epithelial cells (NMuMG cells) deposit at their basal surfaces an extracellular heparan sulfate-rich proteoglycan that binds to type I collagen. The binding of the purified proteoglycan to collagen was studied by (i) a solid phase assay, (ii) a suspension assay using preformed collagen fibrils, and (iii) a collagen fibril affinity column. The binding interaction occurs at physiological pH and ionic strength and can be inhibited only by salt concentrations that greatly exceed those found physiologically. Binding requires the intact proteoglycan since the protein-free glycosaminoglycan chains will not bind under the conditions of these assays. However, binding is mediated through the heparan sulfate chains as it can be inhibited by block-sulfated polysaccharides, including heparin. Binding requires native collagen structure which may be optimal when the collagen is in a fibrillar configuration. Binding sites on collagen fibrils are saturable, high affinity (Kd approximately 10(-10) M), and selective for heparin-like glycosaminoglycans. Because a culture substratum of type I collagen fibrils causes NMuMG cells to accumulate heparan sulfate proteoglycan into a basal lamina-like layer, binding of heparan sulfate proteoglycans to type I collagen may lead to the formation of a basal lamina and may link the basal lamina to the connective tissue matrix, an association found in basement membranes.     

Ovarian carcinoma cells synthesize both chondroitin sulfate and heparan sulfate cell surface proteoglycans that mediate cell adhesion to interstitial matrix

Metastatic ovarian carcinoma metastasizes by intra-peritoneal, non-hematogenous dissemination. The adhesion of the ovarian carcinoma cells to extracellular matrix components, such as types I and III collagen and cellular fibronectin, is essential for intra-peritoneal dissemination. The purpose of this study was to determine whether cell surface proteoglycans (a class of matrix receptors) are produced by ovarian carcinoma cells, and whether these proteoglycans have a role in the adhesion of ovarian carcinoma cells to types I and III collagen and fibronectin. Proteoglycans were metabolically labeled for biochemical studies. Both phosphatidylinositol-anchored and integral membrane-type cell surface proteoglycans were found to be present on the SK-OV-3 and NIH:OVCAR-3 cell lines. Three proteoglycan populations of differing hydrodynamic size were detected in both SK-OV-3 and NIH:OVCAR-3 cells. Digestions with heparitinase and chondroitinase ABC showed that cell surface proteoglycans of SK-OV-3 cells had higher proportion of chondroitin sulfate proteoglycans (75:25 of chondroitin sulfate:heparan sulfate ratio), while NIH:OVCAR-3 cells had higher proportion of heparan sulfate proteoglycans (10:90 of chondroitin sulfate:heparan sulfate ratio). RT-PCR indicated the synthesis of a unique assortment of syndecans, glypicans, and CD44 by the two cell lines. In adhesion assays performed on matrix-coated titer plates both cell lines adhered to types I and III collagen and cellular fibronectin, and cell adhesion was inhibited by preincubation of the matrix with heparin, heparan sulfate, chondroitin sulfate, dermatan sulfate, or chondroitin glycosaminoglycans. Treatment of the cells with heparitinase, chondroitinase ABC, or methylumbelliferyl xyloside also interfered with adhesion confirming the role of both heparan sulfate and chondroitin sulfate cell surface proteoglycans as matrix receptors on ovarian carcinoma cells.     

Proteoheparan sulfate from human skin fibroblasts. Evidence for self-interaction via the heparan sulfate side chains

We have studied the affinity between fibroblast proteoheparan sulfate (medium- and cell surface-derived species) and heparan sulfate-agaroses by affinity chromatography. The evidence for an interaction between the heparan sulfate side chains of the proteoglycans and the immobilized heparan sulfate are as follows: (a) the individual side chains released from the proteoglycan by papain bind to the affinity matrix, (b) the bound proteoglycans are desorbed by a solution of cognate heparan sulfate chains, and (c) the core protein obtained by heparan sulfate-lyase digestion of the proteoglycan does not bind to the affinity matrix. The proteoglycans interact only with one subtype of heparan sulfate. The binding of free heparan sulfate chains to the affinity matrix is completely abolished by heparan sulfate oligosaccharides provided they are composed of both iduronate- and glucuronate-containing disaccharide sequences.     

Permeable Endothelium and the Interstitial Space of Brain

1. Fenestrated vessels can be reversibly induced in brain by agents that stimulate urokinase production. This plasminogen activator, like vascular endothelial growth factor and metalloproteinases, is secreted by tumor cells and may account for induction of fenestrated vessels. Why only some of the brain's barrier vessels are converted to fenestrated vessels is unknown.2. The structures responsible for the filtering of solutes by fenestrated vessels may be the same as those of continuous, less permeable vessels: the glycocalyx on the surfaces of the endothelial cells and the subendothelial basal lamina.3. Solutes leaving the cerebral ventricles immediately enter the interstitial clefts between the cells lining the ventricles. A fraction of a variety of solutes, injected into CSF compartments, is retained by subendothelial basal lamina, from which the solutes may be released in a regulated way.4. The brain's CSF and interstitial clefts are the conduits for nonsynaptic volume transmission of diffusible signals, e.g., ions, neurotransmitters, and hormones. This type of transmission could be abetted by a parallel, cell-to-cell volume transmission mediated by gap junctions between astrocytes bordering CSF compartments and parenchymal astrocytes.5. The width and contents of the interstitial clefts in fetal brain permit cell migration and outgrowth of neurites. The contents of the narrower and different interstitial clefts of mature brain permit solute convection but must be enzymatically degraded in order for cells to migrate through it.     

Glypican-1, phosphacan/receptor protein-tyrosine phosphatase-ζ/β and its ligand,tenascin-C,are expressed by neural stem cells and neural cells derived from embryonic stem cells

The heparan sulfate proteoglycan glypican-1, the chondroitin sulfate proteoglycan phosphacan/RPTP (receptor protein-tyrosine phosphatase)-ζ/β and the extracellular matrix protein tenascin-C were all found to be expressed by neural stem cells and by neural cells derived from them. Expression of proteoglycans and tenascin-C increased after retinoic acid induction of SSEA1-positive ES (embryonic stem) cells to nestin-positive neural stem cells, and after neural differentiation, proteoglycans and tenascin-C are expressed by both neurons and astrocytes, where they surround cell bodies and processes and in certain cases show distinctive expression patterns. With the exception of tenascin-C (whose expression may decrease somewhat), expression levels do not change noticeably during the following 2 weeks in culture. The significant expression, by neural stem cells and neurons and astrocytes derived from them, of two major heparan sulfate and chondroitin sulfate proteoglycans of nervous tissue and of tenascin-C, a high-affinity ligand of phosphacan/RPTP-ζ/β, indicates that an understanding of their specific functional roles in stem cell neurobiology will be important for the therapeutic application of this new technology in facilitating nervous tissue repair and regeneration.     

Glycosaminoglycans differentially bind HARP and modulate its biological activity

Heparin affin regulatory peptide (HARP) is a polypeptide belonging to a family of heparin binding growth/differentiation factors. The high affinity of HARP for heparin suggests that this secreted polypeptide should also bind to heparan sulfate proteoglycans derived from cell surface and extracellular matrix defined as extracellular compartments. Using Western blot analysis, we detected HARP bound to heparan sulfate proteoglycans in the extracellular compartments of MDA-MB 231 and MC 3T3-E1 as well as NIH3T3 cells overexpressing HARP protein. Heparitinase treatment of BEL cells inhibited HARP-induced cell proliferation, and the biological activity of HARP in this system was restored by the addition of heparin. We report that heparan sulfate, dermatan sulfate, and to a lesser extent, chondroitin sulfate A, displaced HARP bound to the extracellular compartment. Binding analyses with a biosensor showed that HARP bound heparin with fast association and dissociation kinetics (kass = 1.6 x 10(6) M-1 s-1; kdiss = 0.02 s-1), yielding a Kd value of 13 nM; the interaction between HARP and dermatan sulfate was characterized by slower association kinetics (kass = 0.68 x 10(6) M-1 s-1) and a lower affinity (Kd = 51 nM). Exogenous heparin, heparan sulfate, and dermatan sulfate potentiated the growth-stimulatory activity of HARP, suggesting that corresponding proteoglycans could be involved in the regulation of the mitogenic activity of HARP.     

Heparan sulfate proteoglycans on the cell surface: versatile coordinators of cellular functions

Heparan sulfate proteoglycans are complex molecules composed of a core protein with covalently attached glycosaminoglycan chains. While the protein part determines localization of the proteoglycan on the cell surfaces or in the extracellular matrix, the glycosaminoglycan component, heparan sulfate, mediates interactions with a variety of extracellular ligands such as growth factors and adhesion molecules. Through these interactions, heparan sulfate proteoglycans participate in many events during cell adhesion, migration, proliferation and differentiation. We are determining the multitude of proteoglycan functions, as their intricate roles in many pathways are revealed. They act as coreceptors for growth factors, participate in signalling during cell adhesion, modulate the activity of a broad range of molecules, and partake in many developmental and pathological processes, including tumorigenesis and wound repair. This review concentrates on biological roles of cell surface heparan sulfate proteoglycans, namely syndecans and glypicans, and outlines the progress achieved during the last decade in unraveling the molecular interactions behind proteoglycan functions.     

Extracellular matrix organization in developing muscle: correlation with acetylcholine receptor aggregates

Monoclonal antibodies recognizing laminin, heparan sulfate proteoglycan, fibronectin, and two apparently novel connective tissue components have been used to examine the organization of extracellular matrix of skeletal muscle in vivo and in vitro. Four of the five monoclonal antibodies are described for the first time here. Immunocytochemical experiments with frozen-sectioned muscle demonstrated that both the heparan sulfate proteoglycan and laminin exhibited staining patterns identical to that expected for components of the basal lamina. In contrast, the remaining matrix constituents were detected in all regions of muscle connective tissue: the endomysium, perimysium, and epimysium. Embryonic muscle cells developing in culture elaborated an extracellular matrix, each antigen exhibiting a unique distribution. Of particular interest was the organization of extracellular matrix on myotubes: the build-up of matrix components was most apparent in plaques overlying clusters of an integral membrane protein, the acetylcholine receptor (AChR). The heparan sulfate proteoglycan was concentrated at virtually all AChR clusters and showed a remarkable level of congruence with receptor organization; laminin was detected at 70-95% of AChR clusters but often was not completely co-distributed with AChR within the cluster; fibronectin and the two other extracellular matrix antigens occurred at approximately 20, 8, and 2% of the AChR clusters, respectively, and showed little or no congruence with AChR. From observations on the distribution of extracellular matrix components in tissue cultured fibroblasts and myogenic cells, several ideas about the organization of extracellular matrix are suggested. (a) Congruence between AChR clusters and heparan sulfate proteoglycan suggests the existence of some linkage between the two molecules, possibly important for regulation of AChR distribution within the muscle membrane. (b) The qualitatively different patterns of extracellular matrix organization over myotubes and fibroblasts suggest that each of these cell types uses somewhat different means to regulate the assembly of extracellular matrix components within its domain. (c) The limited co-distribution of different components within the extracellular matrix in vitro and the selective immune precipitation of each antigen from conditioned medium suggest that each extracellular matrix component is secreted in a form that is not complexed with other matrix constituents.     

Immunocytochemical localization of laminin in hamster tracheal epithelial cell cultures

EM examination of 28 day cultures of enzymatically dissociated hamster tracheal epithelial (HTE) cells grown on collagen coated millipore filters reveals that fragments of basal lamina may be present at the basal plasmalemma. Since the basal lamina consists of several major components including type IV collagen, heparan sulfate proteoglycans, entactin/nidogen, and laminin, questions naturally arise concerning the presence of such a structure in this cell culture system. When immunocytochemical procedures utilizing anti-laminin antibody and PAP techniques are carried out with paraffin sections of HTE culture at 1,2,3, and 4 weeks in vitro, LM analysis reveals that a thin, dense line of reaction product is present between the basal surface of the HTE cells and the underlying collagen substrate. Immunoblotting evaluation carried out with supernatants of 7d HTE cell homogenates and HTE cell conditioned media also indicate that laminin is being produced by the tracheal cells. Thus, the presence of basal lamina-like fragments, the immunocytochemical localization of laminin, and immunoblot identification of laminin in hamster tracheal epithelial cell cultures, suggest that, although basal lamina components may be produced by HTE cells, at the time points tested, they are not yet being organized into a complete basal lamina.     

Fibroblast EXT1-Levels Influence Tumor Cell Proliferation and Migration in Composite Spheroids

Background

Stromal fibroblasts are important determinants of tumor cell behavior. They act to condition the tumor microenvironment, influence tumor growth, support tumor angiogenesis and affect tumor metastasis. Heparan sulfate proteoglycans, present both on tumor and stromal cells, interact with a large number of ligands including growth factors, their receptors, and structural components of the extracellular matrix. Being ubiquitously expressed in the tumor microenvironment heparan sulfate proteoglycans are candidates for playing central roles in tumor-stroma interactions. The objective of this work was to investigate the role of heparan sulfate expressed by stromal fibroblasts in modulating the growth of tumor cells and in controlling the interstitial fluid pressure in a 3-D model.

Methodology/Principal Findings

We generated spheroids composed of fibroblasts alone, or composite spheroids, composed of fibroblasts and tumor cells. Here we show that stromal fibroblasts with a mutation in the heparan sulfate elongating enzyme Ext1 and thus a low heparan sulfate content, formed composite fibroblast/tumor cell spheroids with a significant lower interstitial fluid pressure than corresponding wild-type fibroblast/tumor cell composite spheroids. Furthermore, immunohistochemistry of composite spheroids revealed that the cells segregated, so that after 6 days in culture, the wild-type fibroblasts formed an inner core and the tumor cells an outer layer of cells. For composite spheroids containing Ext1-mutated fibroblasts this segregation was less obvious, indicating impaired cell migration. Analysis of tumor cells expressing the firefly luciferase gene revealed that the changes in tumor cell migration in mutant fibroblast/tumor cell composite spheroids coincided with a lower proliferation rate.

Conclusions/Significance

This is the first demonstration that stromal Ext1-levels modulate tumor cell proliferation and affect the interstitial fluid pressure in a 3-D spheroid model. Learning how structural changes in stromal heparan sulfate influence tumor cells is essential for our understanding how non-malignant cells of the tumor microenvironment influence tumor cell progression.     

Heparan sulfate proteoglycans in invasion and metastasis

Because heparan sulfate proteoglycans mediate cell adhesion and control the activities of numerous growth and motility factors, they play a critical role in regulating the metastatic behavior of tumor cells. Due to their utilitarian nature, heparan sulfate proteoglycans may at times act as inhibitors of cell invasion and at other times as promoters of cell invasion, with their function being determined by their location (cell surface or extracellular matrix), the heparin-binding molecules they associate with, the presence of modifying enzymes (proteases, heparanases) and the precise structural characteristics of the proteoglycan. Also, the tissue type and pathophysiological state of the tumor influence proteogylcan function. This review summarizes our current knowledge of the role heparan sulfate proteoglycans play in regulating tumor cell metastasis, proposes mechanisms of how these molecules function and examines the potential for discovery of new therapeutic approaches designed to block metastatic cancer.     

Heterophilic NCAM-Mediated Cell Adhesion to Proteoglycans from Chick Embryonic Brain Membranes

The vertebrate neural cell adhesion molecule NCAM mediates adhesion by both homophilic and heterophilic mechanisms, with heparan sulfate proteoglycans (HSPGs) being likely heterophilic ligands. In this study, transfected chicken NCAM polypeptides expressed on mouse L cells mediated the adhesion of these cells to several different heparan sulfate proteoglycans in nonionic detergent extracts of Embryonic Day 10 chicken brain membranes. In addition, adhesion inhibition experiments suggested a hitherto-undetected role for chondroitin sulfate proteoglycans in the stimulation of NCAM-mediated adhesion to some, but not all, of the HSPG ligands. Our experiments support the view that NCAM is a multivalent adhesive molecule whose function is affected by interactions with extracellular matrix and cell surface molecules.