Chondroitin Sulfate-dermatan Sulfate Proteoglycan Research Articles

Glycosaminoglycans as Key Molecules in Atherosclerosis: The Role of Versican and Hyaluronan

Cardiovascular disease is the largest cause of death in Western societies and it primarily results from atherosclerosis of large and medium-sized vessels. Atherosclerosis leads to myocardial infarction, when it occurs in the coronary arteries, or stroke, when it occurs in the cerebral arteries. Pathological processes involved in macrovascular disease include the accumulation of lipids which are retained by extracellular matrix (ECM) molecules, especially by the chondroitin sulfate/dermatan sulfate (CS/DS) proteoglycans (CS/DSPGs), such as versican, biglycan and decorin. The sulfation pattern of CS is a key player in protein interactions causing atherosclerosis. Several studies have shown that lipoproteins bind CSPGs via their glycosaminoglycan chains. Galactosaminoglycans, such as CS and DS, bind low density lipoproteins (LDL), affecting the role of these molecules in the arterial wall. In this article, the role of CS and versican in atherosclerosis and hyaluronan in atherogenesis as well as the up to date known mechanisms that provoke this pathological condition are presented and discussed.

Monitoring serum chondroitin sulfate levels in patients submitted to coronary artery bypass surgery

Glycosaminoglycans (GAGs) are functionally important molecules of the arterial wall and play a crucial role in atherogenesis. Chondroitin sulfate/dermatan sulfate proteoglycans (CS/DSPGs) participate in several biological events through their GAG chains, and are also involved in the development of atherosclerosis. The aim of this study was to compare the pre- and post-operative levels of CS in serum of patients after coronary artery bypass graft surgery using a highly sensitive reversed-polarity capillary electrophoresis method and to investigate the correlation of CS with common biochemical lipid markers. It was found that CS values were significantly higher for all patients post-operatively and, furthermore, CS levels were statistically correlated to apolipoprotein A and B levels. Notably, the pre-operational lipid profile of the patient may be indicative of the values of 4-sulfated CS post-operationally. Furthermore, the obtained results highlight the clinical significance of CS levels in serum, since they may provide complementary information for the latent inflammatory state of the patient.

Effects of low molecular weight heparin in obstructed kidneys: decrease of collagen, fibronectin and TGF-β, and increase of chondroitin/dermatan sulfate proteoglycans and macrophage infiltration

Heparin exerts beneficial effects in different experimental models of nephropathy, as observed by the preservation of the structural morphology of the kidney after heparin therapy. Here we investigate molecular and cellular events involved in the protective effects of heparin in the progression of renal disease after unilateral ureteral obstruction. Thirty-six rats were divided into six groups: group C (control) was not subjected to any surgical manipulation; group S (sham) was subjected to surgical manipulation but without ureteral ligation; group UUO was subjected to ureteral obstruction and received no treatment; group UUO + S was subjected to ureteral obstruction and received saline subcutaneously (s.c.) once daily; group UUO + H was subjected to ureteral obstruction and received low molecular weight heparin (LMW-Hep; 4 mg/kg) s.c. once daily; and group C + H was not subjected to any surgical manipulation and received LMW-Hep (4 mg/kg) s.c. once daily. After 14 days, the content of collagen, fibronectin, total glycosaminoglycans (GAGS), chondroitin sulfate/dermatan sulfate proteoglycans (CS/DSPGs), transforming growth factor-beta (TGF-beta) and cellular infiltration were determined in the kidneys by immunohistochemical and biochemical techniques. Collagen, fibronectin, total GAGS, CS/DSPGs, TGF-beta and cellular infiltration increased significantly in group UUO. LMW-Hep treatment reduced collagen, fibronectin and TGF-beta, but induced an increase in the content of total GAGS, CS/DSPGs and macrophage infiltration in group UUO + H when compared with group UUO. LMW-Hep diminishes fibrosis in obstructed kidneys by downregulating the synthesis of collagen, fibronectin and TGF-beta. The mechanisms underlying the overproduction of CS/DSPGs and the increase in cellular infiltration upon LMW-Hep administration remain to be elucidated.

Characterization of chondroitin sulfate and dermatan sulfate proteoglycans of extracellular matrices of human umbilical cord blood vessels and Wharton's jelly

The chondroitin sulfate/dermatan sulfate proteoglycans (CS/DSPGs) of the human umbilical cord vein, arteries and Wharton's jelly matrices were characterized and localized by immunohistochemical analysis. The CS/DSPGs were found to be decorins and biglycans with 43-48 kDa core proteins and are distributed throughout the umbilical cord. A truncated form of decorin having only the approximately 14 kDa NH(2)-terminal portion of the core protein was found exclusively in the vein. The proteoglycans, regardless of their locations, have two types of CS/DS chains, one with approximately 90% CS and approximately 10% DS and the other with approximately 65% CS and approximately 35% DS. The glycosaminoglycan (GAG) chains of the truncated decorin consist of approximately 53% CS and approximately 47% DS. Both decorin and biglycan including the truncated form of decorin could efficiently bind collagen I and fibronectin. The decorin and biglycan with approximately 10% DS and approximately 90% CS were loosely bound in the extracellular matrices, whereas those with approximately 35% DS bound strongly. Together, these data demonstrate that, the GAG chains with 35-47% DS but not those with 10% DS, interact strongly with the matrix. Our data also show that the GAG chain composition is a significant factor in binding of the decorin and biglycan to matrix proteins. The expression of decorin and biglycan with distinctively different CS/DS proportions implies specific biological functions for these PGs in the umbilical cord. The occurrence of the truncated form of decorin exclusively in the umbilical vein suggests a specific functional role.

Distribution of heparan sulfate proteoglycans in embryonic chicken neural retina and isolated inner limiting membrane.

Quantitative distribution of proteoglycans was studied in retinal neural epithelium and its basement membrane (inner limiting membrane). Heparan sulfate proteoglycans (HSPGs) were primarily associated with both inner and outer plexiform (synaptic) layers, and inner limiting membrane (ILM), as determined by autoradiographs of lyase-digested cryosections. Based on distribution of 35S-sulfate-labeled proteoglycans, the isolated ILM contained on average approximately three fourths of its proteoglycans as HSPGs and one fourth as chondroitin sulfate/dermatan sulfate proteoglycans (CS/DSPGs), whereas the remaining retina contained approximately equal amounts of the two proteoglycans (PGs). Immunohistochemical staining indicates that the core proteins of the HSPGs in the ILM are distinct from those of the plexiform layers. The photoreceptor layer, which other studies have shown to contain much of the extracellular CS/DSPGs, was not examined. Enrichment of distinct HSPGs in the ILM and plexiform layers support the conclusion that the HSPGs may be intimately involved in the different developmental events characterizing the two regions: development and extension of ganglion cell axons in the former, synaptogenesis and neuronal function in the latter.

A monoclonal antibody that recognizes hyaluronic acid binding region of aorta proteoglycans.

A chondroitin sulfate-dermatan sulfate proteoglycan was isolated from bovine aorta intima by extraction of the tissue with 4 M guanidine hydrochloride. The proteoglycan was purified by CsCl isopycnic centrifugation followed by gel filtration and ion exchange chromatography. A monoclonal antibody C8F4 was developed to this core protein. The characteristics and specificity of the antibody were studied by an enzyme-linked immunosorbent assay (ELISA) using an alkaline phosphatase conjugated antibody (goat anti-mouse IgG). The antibody binding to the core protein was found specific and optimal at pH 7.0. The antibody recognizes either intact chondroitin sulfate-dermatan sulfate proteoglycan monomer, chondroitinase ABC digested monomer or chemically deglycosylated proteoglycan. Free chondroitin sulfates, keratan sulfate and hyaluronic acid did not compete for the antigenic sites in ELISA. Limited hydrolysis of the core protein by trypsin resulted in three peptides and only the peptide with a molecular weight M(r) = 40,000 was found capable of binding to hyaluronic acid. The antibody C8F4 recognized this hyaluronic acid binding peptide but did not recognize the other two peptides suggesting that the epitope(s) for this antibody is in the hyaluronic acid-binding region of the core protein. The antibody recognized the core proteins from bovine nasal cartilage proteoglycan and human aorta proteoglycan but did not recognize bovine aorta link protein, bovine serum albumin, human serum albumin, human transferrin, collagen Type I and fibronectin. The antibody was found useful to localize proteoglycans in atherosclerotic lesions in human aorta by immunohistochemical techniques.

Metabolism of low-density lipoprotein-proteoglycan complex by macrophages: further evidence for a receptor pathway

Earlier, we (Vijayagopal, P., et al. (1985) Biochim. Biophys. Acta 837-251) have shown that complexes of plasma low-density lipoproteins (LDL) and arterial chondroitin sulfate-dermatan sulfate proteoglycan aggregate promote LDL degradation and cholesteryl ester accumulation in mouse peritoneal macrophages. Further studies were conducted to determine whether LDL-proteoglycan complex is metabolized by a receptor-mediated process. Native proteoglycan aggregate was isolated from bovine aorta by associative CsCI isopycnic centrifugation. Complex of 125I-labeled LDL and proteoglycan aggregate formed in the presence of 30 mM Ca 2+ was incubated with macrophages, and the binding at 4°C and degradation at 37°C of 125I-labeled LDL in the complex was monitored. Both binding and degradation of the complex were specific and saturable, suggesting that the processes are receptor mediated. The K d for binding was 23μg LDL protein per ml in the complex. Degradation of 125I-labeled LDL-proteoglycan complex was not suppressed by preincubation of macrophages with excess unlabeled complex, suggesting that the receptor for the complex is not subject to down regulation. Both binding and degradation of the complex and the resultant stimulation of cholesteryl ester synthesis were inhibited by limited treatment of cells with low doses of trypsin and pronase, indicating that the binding sites are protein or glycoprotein in nature. Binding was not inhibited by an excess of native LDL and β-VLDL and exhibited only partial competition by excess unlabeled acetyl-LDL; however, polyinosinic acid, fucoidin and dextran sulfate, known inhibitors of acetyl-LDL binding and degradation in macrophages, did not affect LDL-proteoglycan complex binding and degradation. Similarly, excess unlabeled LDL-proteoglycan complex produced only partial inhibition of the binding and degradation of 125I-labeled acetyl-LDL by macrophages, suggesting that the binding sites for acetyl-LDL and LDL-proteoglycan complex are probably not identical. These studies provide evidence for a receptor-mediated pathway for the metabolism of LDL-proteoglycan complex in macrophages.

Proteoglycans synthesized by embryonic chicken retina in culture: Composition and compartmentalization

Characteristics of the chondroitin sulfate/dermatan sulfate proteoglycans (CS/DSPGs) and heparan sulfate proteoglycans (HSPGs) from retinas of 14-day chicken embryos were examined following specific lyase digestion of the HSPG and CS/DSPG glycosaminoglycans, respectively. On the basis of gel exclusion chromatography the prevalent CS/DSPGs in the tissue were above M r 400 × 10 3 with two or three glycosaminoglycan chains of M r 60–70 × 10 3. The HSPGs existed in two distinct populations in the tissue. Those in the dominant population appeared to be in the range of M r 250–300 × 10 3 with 9 to 12 glycosaminoglycan chains of M r 15–25 × 10 3. The other population consisted of free heparan sulfate chains of M r15–25 × 10 3. The HSPGs in the medium tended to be intermediate in size. To examine the distribution of proteoglycans, tissues were sequentially homogenized and extracted in saline and reextracted with 4 m guanidine HCl (GdnHCl) and Triton X-100 (TX), or they were washed in heparin solution and dissociated to single cells with trypsin before sequential extraction in saline and GdnHCl with TX. Through comparison of the results of these two extraction methods, CS/DSPGs were found to be almost entirely within the medium or matrix or loosely associated with the cell surface, and most HSPGs were associated with either the basal lamina or the plasma membrane. The single heparan sulfate glycosaminoglycan chains appeared to be intracellular degradation products. These results support reports that CS/DSPGs may be present in the retina interphotoreceptor matrix and that HSPGs may be present in regions of synaptogenesis, associated with cell membranes.

Immunocytochemical investigation on the distribution of small chondroitin sulfate-dermatan sulfate proteoglycan in the human.

Polyclonal antibodies against the core protein of the small chondroitin sulfate-dermatan sulfate proteoglycan from human skin fibroblast secretions were used, after affinity-purification, as a probe to study localization of crossreactive material in several human tissues by indirect immunocytochemistry. In contrast to skin, kidney, and the adventitial layer of aorta, positive staining of brain, liver, cartilage, and intimal and medial layers of aorta required pre-treatment of tissue sections with chondroitin ABC lyase. In all tissues investigated, antigenic material was present in the interstitial space. Filamentous structures were perpendicularly oriented towards basement membranes. In liver, specific staining was seen along the sinusoidal walls. Reticular fibers with or without focal condensations were seen in cerebral cortex and cerebellum. The results suggest a role of small chondroitin sulfate-dermatan sulfate proteoglycan in cell-matrix interactions.

Hemostatic properties and serum lipoprotein binding of a heparan sulfate proteoglycan from bovine aorta

The biologic properties of two major proteoglycans of bovine aorta, heparan sulfate proteoglycan and chondroitin sulfate-dermatan sulfate proteoglycan were compared. The heparan sulfate proteoglycan was isolated either by elastase digestion or by 4.0 M guanidine hydrochloride extraction, of aorta tissue, fractionated by CsCl isopycnic centrifugation and purified by chondrointinase ABC treatment. The first method resulted in considerably greater yield (about 70% of the total heparan sulfate proteoglycan of the tissue) than the second procedure (12% of total). The chondroitin sulfate-dermatan sulfate proteoglycan was obtained by 4.0 M guanidine-HCl extraction of aorta tissue followed by CsCl isopycnic centrifugation. The chemical composition of both heparan sulfate proteoglycan preparations was similar. Unlike the chondroitin sulfate-dermatan sulfate proteoglycan, which eluted in the void volume of Sepharose CL-6B column, the heparan sulfate proteoglycan preparations were each resolved into a high molecular weight fraction ( k av = 0.18 and 0.13) and a low molecular weight fraction ( k av = 0.47 and 0.36). The heparan sulfate proteoglycan preparations exhibited significantly more potent anticoagulant and platelet aggregation inhibitory activities than the chondroitin sulfate-dermatan sulfate proteoglycan. The protein core of the proteoglycan molecules did not seem to be essential for their hemostatic properties. The complex forming ability of the heparan sulfate proteoglycan with serum low density lipoproteins (LDL) was much less than that of chondroitin sulfate-dermatan sulfate proteoglycan in the presence and absence of Ca 2+. Interaction between heparan sulfate proteoglycan and LDL was also much more sensitive to changes in the ionic strength of the medium than that of chondroitin sulfate-dermatan sulfate proteoglycan and the lipoprotein. Since the total sulfate content of both proteoglycans is almost similar, the smaller molecular size and hence the lower overall charge density of the heparan sulfate proteoglycan appears to be partly responsible for its low affinity for LDL. The differences in biologic properties of the two proteoglycans might have implications in the pathophysiology of cardiovascular diseases.

Interaction of serum lipoproteins and a proteoglycan from bovine aorta.

The interactions of a chondroitin sulfate-dermatan sulfate proteoglycan with serum very low (VLDL), low (LDL), and high density (HDL3) lipoproteins were studied with special reference to the nature of the interaction of LDL and the proteoglycan. The proteoglycan formed insoluble complexes with VLDL and LDL, but no complex was formed with HDL3. The proteoglycan (40 micrograms/ml) converted 98% of added LDL (150 micrograms of cholesterol/ml) into insoluble complex at a Ca2+ concentration of 30 mM. Physiologic concentrations of albumin inhibited insoluble complex formation with VLDL and LDL by 61.5 and 40.7%, respectively. The proteoglycan formed soluble complexes with LDL even in the absence of Ca2+. Optimum soluble complex formation occurred when the medium contained 66.6 micrograms of proteoglycan and 166.6 micrograms of LDL cholesterol/ml. Specific modifications of the lysine and/or arginine residues of LDL prevented complex formation with the proteoglycan, thus indicating the requirement of the positive charges of the protein moiety of LDL in the interaction. Removal of the protein core or the glycosaminoglycan chains of the proteoglycan prevented interaction with LDL. Desulfation of the proteoglycan molecule also inhibited complex formation with LDL. Thus, the native state of the arterial proteoglycan molecule confers optimum charge density for specific interaction with serum apoprotein B-containing lipoproteins.