Decorin and a large dermatan sulfate proteoglycan in bovine striated muscle

Proteoglycans were extracted and isolated from adult bovine muscle tissue by dissociative extraction followed by density gradient centrifugation, gel chromatography and ion-exchange chromatography. Two proteoglycans were characterized; one of large molecular size (PG-L) and one of small molecular size (PG-S). The recovery of PG-L and PG-S was 33% and 67% respectively. By cellulose acetate electrophoresis before and after treatment with chondroitinase AC and ABC both samples were shown to carry predominantly dermatan sulfate chains. The large proteoglycan was recognized with an antibody against a large dermatan sulfate proteoglycan from bovine sclera, whereas the small was recognized by an antibody against decorin from bovine sclera. Chondroitinase ABC treatment of PG-S followed by SDS-PAGe showed a core protein with a molecular weight of 45 kDa, which also reacted with the decorin antibody. Amino-acid analysis of both PG-L and PG-S revealed an amino-acid composition closely similar, although not identical, to the large dermatan sulfate proteoglycan from bovine sclera and decorin respectively. Immunohistochemical analyses of muscle tissue sections showed that decorin and the large dermatan sulfate proteoglycan are present in the perimysium layers of muscle tissue, although with a somewhat different pattern of distribution. Decorin was, in addition, found in the endomysium.     

Inventory of human skin fibroblast proteoglycans. Identification of multiple heparan and chondroitin/dermatan sulphate proteoglycans.

Heparan sulphate and chondroitin/dermatan sulphate proteoglycans of human skin fibroblasts were isolated and separated after metabolic labelling for 48 h with 35SO4(2-) and/or [3H]leucine. The proteoglycans were obtained from the culture medium, from a detergent extract of the cells and from the remaining ''matrix'', and purified by using density-gradient centrifugation, gel and ion-exchange chromatography. The core proteins of the various proteoglycans were identified by electrophoresis in SDS after enzymic removal of the glycosaminoglycan side chains. Skin fibroblasts produce a number of heparan sulphate proteoglycans, with core proteins of apparent molecular masses 350, 250, 130, 90, 70, 45 and possibly 35 kDa. The major proteoglycan is that with the largest core, and it is principally located in the matrix. A novel proteoglycan with a 250 kDa core is almost entirely secreted or shed into the culture medium. Two exclusively cell-associated proteoglycans with 90 kDa core proteins, one with heparan sulphate and another novel one with chondroitin/dermatan sulphate, were also identified. The heparan sulphate proteoglycan with the 70 kDa core was found both in the cell layer and in the medium. In a previous study [Fransson, Carlstedt, Cöster & Malmström (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 5657-5661] it was suggested that skin fibroblasts produce a proteoglycan form of the transferrin receptor. However, the core protein of the major heparan sulphate proteoglycan now purified does not resemble this receptor, nor does it bind transferrin. The principal secreted proteoglycans are the previously described large chondroitin sulphate proteoglycan (PG-L) and the small dermatan sulphate proteoglycans (PG-S1 and PG-S2).     

Characterisation of chondroitin/dermatan sulphate proteoglycans synthesised by rat mammary myoepithelial and fibroblastic cell lines.

Chondroitin sulphate proteoglycans were isolated from the culture medium of rat mammary gland fibroblast (Rama 27) and myoepithelial (Rama 401) cell lines which had been labelled with [35S]sulphate. Chromatography on Sepharose CL-4B indicated that the Rama 401 proteoglycan was larger than the Rama 27 proteoglycan (Kav values 0.47 and 0.56, respectively). Treatment of the proteoglycans with alkaline NaBH4 yielded chondroitin sulphate chains with average M(r) values of 37,000 (Rama 401) and 21,000 (Rama 27). Structural analysis of the glycosaminoglycan chains indicated that both were co-polymers of chondroitin and dermatan sulphate although there were differences in the amounts and distribution of the disaccharide repeating units. The M(r) values of the core proteins, determined by immunoblotting, were about 43,000 and 46,000 (Rama 27) and 44,500 (Rama 401). Using an antibody to chondroitin sulphate proteoglycan in immunofluorescence experiments, the proteoglycan was demonstrated on the surface of both cell lines. Rama 27 cells additionally possessed an extensive fibrous extracellular matrix which also stained with the antibody. Staining of sections of lactating mammary gland suggested that the proteoglycan was present in the basement membrane as well as the stromal connective tissue. The presence of chondroitin sulphate proteoglycan in the basement membrane was confirmed by ultrastructural immunolocalisation.     

Transforming growth factor-beta induces selective increase of proteoglycan production and changes in the copolymeric structure of dermatan sulphate in human skin fibroblasts.

Human embryonic skin fibroblasts were pretreated with transforming growth factor-beta (TGF-beta) for 6 h and then labeled with [35S]sulphate and [3H]leucine for 24 h. Radiolabeled proteoglycans from the culture medium and the cell layer were isolated and separated by isopycnic density-gradient centrifugation, followed by gel, ion-exchange and hydrophobic-interaction chromatography. The major proteoglycan species were examined by polyacrylamide gel electrophoresis in sodium dodecyl sulphate before and after enzymatic degradation of the polysaccharide chains. The results showed that TGF-beta increased the production of several different 35S-labelled proteoglycans. A large chondroitin/dermatan sulphate proteoglycan (with core proteins of approximately 400-500 kDa) increased 5-7-fold and a small dermatan sulphate proteoglycan (PG-S1, also termed biglycan, with a core protein of 43 kDa) increased 3-4-fold both in the medium and in the cell layer. Only a small effect was observed on another dermatan sulphate proteoglycan, PG-S2 (also named decorin). These observations are generally in agreement with results of other studies using similar cell types. In addition, we have found that the major heparan sulphate proteoglycan of the cell layer (protein core approximately 350 kDa) was increased by TGF-beta treatment, whereas all the other smaller heparan sulphate proteoglycans with protein cores from 250 kDa to 30 kDa appeared unaffected. To investigate whether TGF-beta also influences the glycosaminoglycan (GAG) chain-synthesizing machinery, we also characterized GAGs derived from proteoglycans synthesized by TGF-beta-treated cells. There was generally no increase in the size of the GAG chains. However, the dermatan sulphate chains on biglycan and decorin from TGF-beta treated cultures contained a larger proportion of D-glucuronosyl residues than those derived from untreated cultures. No effect was noted on the 4- and 6-sulphation of the GAG chains. By the use of p-nitrophenyl beta-D-xyloside (an initiator of GAG synthesis) it could be demonstrated that chain synthesis was also enhanced in TGF-beta-treated cells (approximately twofold). Furthermore, the dermatan sulphate chains synthesized on the xyloside in TGF-beta-treated fibroblasts contained a larger proportion of D-glucuronosyl residues than those of the control. These novel findings indicate that TGF-beta affects proteoglycan synthesis both quantitatively and qualitatively and that it can also change the copolymeric structure of the GAG by affecting the GAG-synthesizing machinery. Altered proteoglycan structure and production may have profound effects on the properties of extracellular matrices, which can affect cell growth and migration as well as organisation of matrix fibres.     

Biochemical and cytochemical characterization of extracellular proteoglycans in the inner circular smooth muscle layer of dog small intestine

Current literature concerning smooth muscle blood vessels has shown versican as the main proteoglycan (PG) component of the matrix. To show whether smooth muscle matrix has the same PG distribution when present in organs, other than the blood vessels, the inner circular smooth muscle layer of the small intestine was obtained by dissection as a highly purified tissue and analyzed by biochemical and cytochemical methods. The smooth muscle layer PGs were extracted from dog small intestine with 4 M guanidine-HCl in the presence of proteinase inhibitors, purified by charge equilibrium, isolated by equilibrium CsCl density gradients, and analyzed in terms of anion exchange, size, and glycosaminoglycan (GAG) distribution. Proteoheparan sulfate itself represented 91.5% of the PGs present in this tissue. The remainder was proteodermatan sulfate. Cytochemical analyses using the cationic dye cuprolinic blue associated with enzymatic treatments with chondroitinases ABC and heparitinase III showed the arrangement and identification of PGs in basal lamina and intramuscular connective tissues. The PGs in the basal lamina were proteoheparan sulfate, and those associated with collagen fibrils in the endomysium and perimysium were rich in dermatan sulfate. In contrast to the blood vessels, inner circular muscle smooth tissue in intestine has, as the main PG, perlecan.     

Proliferation-dependent changes of proteoglycan metabolism in arterial smooth muscle cells

Cultured arterial smooth muscle cells synthesize and secrete two types of sulfated proteoglycans designated as proteoglycan A and proteoglycan B. Proteoglycan A has been characterized as chondroitin sulfate-rich, whereas proteoglycan B was found to be dermatan sulfate-rich [Schmidt, A. & Buddecke, E. (1985) Eur. J. Biochem. 153, 260-273]. During the logarithmic growth phase, arterial smooth muscle cells incorporated about 3 times more [35S]sulfate into the total proteoglycans secreted into the culture medium than did non-dividing cells. When arterial smooth muscle cells stopped proliferating the ratio of [35S]proteoglycan A/B increased. No differences were detected in the respective molecular and chemical characteristics of purified proteoglycans A and B isolated from both proliferating and non-dividing cells. Regardless of the growth phase proteoglycan A had a molecular mass of about 280 kDa and contained 8-9 chondroitin sulfate-rich side chains. Proteoglycan B had a molecular mass of about 180 kDa and contained 6-7 dermatan sulfate-rich side chains. The [35S]methionine-labelled protein cores of proteoglycan A and B had a molecular mass of about 48 kDa, but were distinguishable by their specific reactions to monospecific antibodies. Proliferating cells endocytosed proteoglycan B at a rate up to 100% higher than that of non-dividing cells. In all growth phases proteoglycan A was endocytosed at a 10-fold lower rate than proteoglycan B.     

Purification and characterisation of a minor low-sulphated dermatan sulphate-proteoglycan from ray skin.

A minor low-sulphated dermatan sulphate proteoglycan was isolated from ray skin by extraction with 2% sodium dodecyl sulphate, followed with ion-exchange chromatography, gel chromatography and density gradient centrifugation. The proteoglycan with a relative molecular mass (Mr) ranging from 70 to 120 kDa is composed of about two dermatan sulphate chains (Mr 33 kDa) bound on a protein core of Mr 27 kDa, and oligosaccharides consisting of uronic acids, hexosamines and neutral sugars. The major amino acids of the protein core were glycine (corresponding to about one-fourth of the total amino acids), serine, threonine, glutamic acid/glutamine, leucine and cysteine, together amounting to 56% of the total. The isolated proteoglycan does not interact with hyaluronic acid and does not form self-aggregates. Dermatan sulphate was rich in iduronic acid (62% of total uronic acid) and composed of non-sulphated (44%), and mono-sulphated disaccharides bearing esterified sulphate groups at positions C-4 (53%) or C-6 (3%) of the N-acetyl galactosamine. HPLC analysis of a pure preparation of dermatan sulphate, showed the presence of galactose and glucose possibly as branches on the dermatan sulphate chain.     

Isolation and partial characterization of heparan sulphate proteoglycan from the human glomerular basement membrane.

Heparan sulphate proteoglycan was solubilized from human glomerular basement membranes by guanidine extraction and purified by ion-exchange chromatography and gel filtration. The yield of proteoglycan was approx. 2 mg/g of basement membrane. The glycoconjugate had an apparent molecular mass of 200-400 kDa and consisted of about 75% protein and 25% heparan sulphate. The amino acid composition was characterized by a high content of glycine, proline, alanine and glutamic acid. Hydrolysis with trifluoromethanesulphonic acid yielded core proteins of 160 and 110 kDa (and minor bands of 90 and 60 kDa). Alkaline NaBH4 treatment of the proteoglycan released heparan sulphate chains with an average molecular mass of 18 kDa. HNO2 oxidation of these chains yielded oligosaccharides of about 5 kDa, whereas heparitinase digestion resulted in a more complete degradation. The data suggest a clustering of N-sulphate groups in the peripheral regions of the glycosaminoglycan chains. A polyclonal antiserum raised against the intact proteoglycan showed reactivity against the core protein. It stained all basement membranes in an intense linear fashion in immunohistochemical studies on frozen kidney sections from man and various mammalian species.     

Physical properties of chondroitin sulphate/dermatan sulphate proteoglycans from bovine aorta.

Bovine aortic chondroitin sulphate/dermatan sulphate proteoglycans (PG-25, PG-35 and PG-50) were differentially precipitated with ethanol and analysed by a variety of chemical and physical techniques. The glycosaminoglycan chains of PG-25 and PG-35 contained a mixture of glucuronic acid and iduronic acid, whereas the uronic acid component of PG-50 was primarily glucuronic acid. In addition, various amounts of oligosaccharides containing small amounts of mannose, a galactose/hexosamine ratio of 1:1 and an absence of uronic acid were covalently linked to the core protein of all proteoglycans. The weight-average Mr (Mw) values of the proteoglycans determined by light-scattering in 4 M-guanidinium chloride were 1.3 X 10(6) (PG-25), 0.30 X 10(6) (PG-35) and 0.88 X 10(6) (PG-50). The s0 values of the proteoglycans were distributed between 7 and 8 S, and the reduced viscosities, eta sp./c, of all proteoglycans were dependent on the shear rate and polymer concentration. Electron microscopy of spread molecules revealed that PG-25 contained small structural units that appeared to self-associate into large aggregates, whereas PG-35 and PG-50 appeared mainly as monomers consisting of a core with various numbers of side projections. Hyaluronic acid-proteoglycan complexes occurred only with a small proportion of the molecules present in PG-35, and their formation could be inhibited by oligosaccharides. These results suggest the presence in the aorta of subspecies of chondroitin sulphate and dermatan sulphate proteoglycans, which show large variations in their physicochemical and inter- and intra-molecular association properties.     

Localization of proteoheparan sulfate in rat aorta

Summary This study describes the distribution of heparan sulfate proteoglycan (HSPG) within the rat aorta using immunocytochemical (biotin-avidin-peroxidase) and immunoelectron microscopy (¹²⁵I-autoradiography). Heparan sulfate proteoglycan was isolated from a basement membrane producing mouse EHS sarcoma (Hassell et al. 1980) and used to generate antisera in rabbits. Light microscopic observations revealed intense immunostaining of the intima and media of normal aorta, adventitial vasa vasorum, and aortic intimal fibromuscular thickenings induced by experimental injury (balloon de-endothelialization). Immunoelectron microscopy using ¹²⁵I labeled antibodies to HSPG revealed that proteoheparan sulfate was localized to the amorphous layer of basement membrane below aortic and capillary endothelium. In addition, labeled anti-HSPG could be localized to the external lamina surrounding the smooth muscle cells in the hyperplastic intima. These studies reveal that antibodies prepared against a proteoheparan sulfate isolated from a basement membrane producting EHS sarcoma cross react with basement membrane structures within the aortic wall. Furthermore, these results demonstrate that the basement membranes beneath aortic and capillary endothelium and the external lamina surrounding aortic smooth muscle cells contain a heparan sulfate proteoglycan that is antigenically similar.     

Immunocytochemical localization of heparan sulphate proteoglycan in the rat submandibular gland

Summary Heparan sulphate proteoglycan is the predominant proteoglycan synthesized by the parenchymal cells of the rat submandibular gland. A polyclonal antibody was used to localize this proteoglycan in the adult rat submandibular gland. Localization was accomplished by indirect immunoperoxidase cytochemistry at the light and electron microscopic levels. Heparan sulphate proteoglycan was localized in a continuous, linear pattern in the lamina densa of the basement membrane surrounding all of the epithelial components of the gland as well as the basement membrane of the capillaries and small arterioles in the glandular stroma. In addition, heparan sulphate proteoglycan was seen in vesicles and pits along the acinar cell basal plasmalemma adjacent to the basement membrane and in the endoplasmic reticulum and Golgi apparatus of the acinar cells.     

Developmental expression and cellular origin of the laminin alpha2, alpha4, and alpha5 chains in the intestine.

Laminins are extracellular matrix glycoproteins that are involved in various cellular functions, including adhesion, proliferation, and differentiation. In this study, we examine the expression patterns and the cellular origins of the laminin alpha2, alpha4, and alpha5 chains in the developing mouse intestine and in in vitro mouse/chick or chick/mouse interspecies hybrid intestines. In situ hybridization and Northern blot analysis revealed that mRNA levels for all three laminin alpha chains are highest in the fetal intestine undergoing intense morphogenetic movements. Laminin alpha4 mRNA and polypeptide are associated with mesenchyme-derived cell populations such as endothelium and smooth muscle. In contrast, laminin alpha2 and alpha5 chains participate in the structural organization of the subepithelial basement membrane and, in the mature intestine, show a complementary pattern of expression. All three laminin alpha chains occur in the smooth muscle basement membrane, with a differential expression of laminin alpha5 chain in the circular and longitudinal smooth muscle layers. The cellular origin of laminin alpha2 and alpha5 chains found in the subepithelial cell basement membrane was studied by immunocytochemical analysis of mouse/chick or chick/mouse interspecies hybrid intestines at various stages of development using mouse-specific antibodies. Laminin alpha2 was found to be deposited into the basement membrane exclusively by mesenchymal cells, while the laminin alpha5 chain was deposited by both epithelial and mesenchymal cells in an apparently developmentally regulated pattern. We conclude that (1) multiple laminin alpha chains are expressed in the intestine, implying specific roles for individual laminin isoforms during intestinal development, and (2) reciprocal epithelial/mesenchymal interactions are essential for the formation of a structured subepithelial basement membrane.     

Localization of proteoheparan sulfate in rat aorta

This study describes the distribution of heparan sulfate proteoglycan ( HSPG ) within the rat aorta using immunocytochemical (biotin-avidin-peroxidase) and immuno-electron microscopy (125I-autoradiography). Heparan sulfate proteoglycan was isolated from a basement membrane producing mouse EHS sarcoma ( Hassell et al. 1980) and used to generate antisera in rabbits. Light microscopic observations revealed intense immunostaining of the intima and media of normal aorta, adventitial vasa vasorum, and aortic intimal fibromuscular thickenings induced by experimental injury (balloon de-endothelialization). Immunoelectron microscopy using 125I labeled antibodies to HSPG revealed that proteoheparan sulfate was localized to the amorphous layer of basement membrane below aortic and capillary endothelium. In addition, labeled anti- HSPG could be localized to the external lamina surrounding the smooth muscle cells in the hyperplastic intima. These studies reveal that antibodies prepared against a proteoheparan sulfate isolated from a basement membrane producing EHS sarcoma cross react with basement membrane structures within the aortic wall. Furthermore, these results demonstrate that the basement membranes beneath aortic and capillary endothelium and the external lamina surrounding aortic smooth muscle cells contain a heparan sulfate proteoglycan that is antigenically similar.     

Four classes of cell-associated proteoglycans in suspension cultures of articular-cartilage chondrocytes.

The characteristics of cell-associated proteoglycans were studied and compared with those from the medium in suspension cultures of calf articular-cartilage chondrocytes. By including hyaluronic acid or proteoglycan in the medium during [35S]sulphate labelling the proportion of cell-surface-associated proteoglycans could be decreased from 34% to about 15% of all incorporated label. A pulse-chase experiment indicated that this decrease was probably due to blocking of the reassociation with the cells of proteoglycans exported to the medium. Three peaks of [35S]sulphate-labelled proteoglycans from cell extracts and two from the medium were isolated by gel chromatography on Sephacryl S-500. These were characterized by agarose/polyacrylamide-gel electrophoresis, by SDS/polyacrylamide-gel electrophoresis of core proteins, by glycosaminoglycan composition and chain size as well as by distribution of glycosaminoglycans in proteolytic fragments. The results showed that associated with the cells were (a) large proteoglycans, typical for cartilage, apparently bound to hyaluronic acid at the cell surface, (b) an intermediate-size proteoglycan with chondroitin sulphate side chains (this proteoglycan, which had a large core protein, was only found associated with the cells and is apparently not related to the large proteoglycans), (c) a small proteoglycan with dermatan sulphate side chains with a low degree of epimerization, and (d) a somewhat smaller proteoglycan containing heparan sulphate side chains. The medium contained a large aggregating proteoglycan of similar nature to the large cell-associated proteoglycan and small proteoglycans with dermatan sulphate side chains with a higher degree of epimerization than those of the cells, i.e. containing some 20% iduronic acid.     

Stimulation of human arterial smooth muscle cell chondroitin sulfate proteoglycan synthesis by transforming growth factor-beta

Summary Human platelet-derived transforming growth factor-beta (TGF-beta) is a cell-type specific promotor of proteoglycan synthesis in human adult arterial cells. Cultured human adult arterial smooth muscle cells synthesized chondroitin sulfate, dermatan sulfate, and heparan sulfate proteoglycans, and the percent composition of these three proteoglycan subclasses varied to some extent from cell strain to cell strain. However, TGF-beta consistently stimulated the synthesis of chondroitin sulfate proteoglycan. Both chondroitin 4- and chondroitin 6-sulfate were stimulated by TGF-beta to the same extent. TGF-beta had no stimulatory effect on either class of [³⁵S]sulfate-labeled proteoglycans which appeared in an approximately 1:1 and 2:1 ratio of heparan sulfate to dermatan sulfate of the medium and cell layers, respectively, of arterial endothelial cells. Human adult arterial endothelial cells synthesized little or no chondroitin sulfate proteoglycan. Pulse-chase labeling revealed that the appearance of smooth muscle cell proteoglycans into the medium over a 36-h period equaled the disappearance of labeled proteoglycans from the cell layer, independent of TGF-beta. Inhibitors of RNA synthesis blocked TGF-beta-stimulated proteoglycan synthesis in the smooth muscle cells. The incorporation of [³⁵S]methionine into chondroitin sulfate proteoglycan core proteins was stimulated by TGF-beta. Taken together, the results presented indicate that TGF-beta stimulates chondroitin sulfate proteoglycan synthesis in human adult arterial smooth muscle cells by promoting the core protein synthesis. Supported in part by grants from the Public Health Service, U.S. Department of Health and Human Services, Washington, DC (CA 37589 and HL 33842), RJR Nabisco, Inc., and Chang Gung Biomedical Research Foundation (CMRP 291).     

The accumulation of basement membrane components during the onset of chondrogenesis and myogenesis in the chick wing bud

The study describes the distribution of several basement membrane molecules in the embryonic chick wing bud from stages 23 to 26, during the onset of myogenesis and chondrogenesis, and then later at stage 28. Laminin is localized as early as stage 23, prior to the onset of myogenesis, in regions corresponding to the position of the future dorsal and ventral myogenic areas. Other matrix components, including fibronectin, do not differentially accumulate in these same regions. Fibronectin, basement membrane heparan sulphate proteoglycan and type IV collagen are more widespread in their distribution than laminin, and are even present between mesenchymal cells. These results suggest a role for laminin in the initial differentiation of the muscle masses and emphasize that components of basement membrane can also be associated with mesenchymal cells.     

Mapping of proteoglycans in human arterial tissue

The major families of proteoglycans in human arterial tissue have been localized and characterized by electron microscopy. After staining with the polycationic dye cuprolinic blue in the presence of a critical electrolyte concentration, three differently sized populations of proteoglycan-cuprolinic blue precipitates are found. The precipitates are distinguished of the basis of their morphology, topographical distribution and susceptibility to specific glycosaminoglycan-degrading enzymes. Each type of proteoglycan is preferentially associated with one connective tissue component: (a) a dermatan sulfate proteoglycan interacts with collagenous fibers, (b) a heparan sulfate proteoglycan is associated with elastic fibers and with the exterior surface of the basement membrane-like layer surrounding smooth muscle cells, and (c) a chondroitin sulfate proteoglycan forms aggregates with hyaluronate in the soluble matrix. Information about the pattern of proteoglycans in normal human arterial tissue should constitute a useful basis for evaluating perturbations in proteoglycan distribution in arteriosclerotic plaques.     

Cell-free translation of mRNA encoding an arterial smooth muscle cell proteoglycan core protein

The size and immunological reactivity of the primary gene products of a small non-aggregating dermatan sulfate proteoglycan from bovine and monkey arterial smooth muscle cells were examined after cell-free translation of mRNA. Antisera against the dermatan sulfate proteoglycans from bovine articular cartilage, DSPG II [Rosenberg et al. J. Biol. Chem. 260, 6304 (1985)] and human skin fibroblasts [Glossl et al. J. Biol. Chem. 259, 14144 (1984)] were used to show that the unmodified smooth muscle precursor core protein was immunologically related to both the cartilage and fibroblast core proteins. The size of the precursor core proteins within each species was identical regardless of the tissue source. Comparison of the precursor core proteins synthesized by primate and bovine cells revealed that the bovine core proteins were approximately 1500 Da larger than the primate core proteins as determined by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. A similar size difference was observed when the mature core proteins of monkey smooth muscle cells and bovine articular chondrocytes were compared after removal of the glycosaminoglycan chains. These results indicate that arterial smooth muscle cells synthesize a dermatan sulfate proteoglycan whose core protein is similar to, if not the same as, the cartilage and fibroblast dermatan sulfate proteoglycan core proteins. These core proteins may be encoded by the same gene that has diverged in size during speciation.     

Biosynthesis and properties of a further member of the small chondroitin/dermatan sulfate proteoglycan family

Human osteosarcoma cells express a 78-kDa proteoglycan core protein to which an asparagine-bound oligosaccharide, O-glycosidically linked oligosaccharides and probably only a single chondroitin 6-sulfate chain of 29-kDa are bound. Prior to O-glycosylation, the N-glycosylated core protein exhibits a mass of 83 kDa. Upon digestion of the secreted proteoglycan with chondroitin ABC lyase a mature core protein with an apparent molecular mass of 106 kDa is obtained. Smaller amounts of core proteins of 101 and 115 kDa can be detected occasionally. The glycosaminoglycan composition and the relative molecular mass of the glycosaminoglycan chain distinguish this proteoglycan, tentatively named proteoglycan 100 (PG-100), from biglycan (small proteoglycan I) and decorin (small proteoglycan II) which are also expressed by osteosarcoma cells. An antiserum against PG-100 shows partial cross-reactivity with decorin, but in contrast to the latter proteoglycan it does not bind to type I collagen fibrils. PG-100 is not a unique product of osteosarcoma cells. It has also been found in the secretions of human skin fibroblasts.     

Differential deposition of basement membrane components during formation of the caudal neural tube in the mouse embryo

The distribution of basement membrane and extracellular matrix components laminin, fibronectin, type IV collagen and heparan sulphate proteoglycan was examined during posterior neuropore closure and secondary neurulation in the mouse embryo. During posterior neuropore closure, these components were densely deposited in basement membranes of neuroepithelium, blood vessels, gut and notochord; although deposition was sparse in the midline of the regressing primitive streak. During secondary neurulation, mesenchymal cells formed an initial aggregate near the dorsal surface, which canalized and merged with the anterior neuroepithelium. With aggregation, fibronectin and heparan sulphate proteoglycan were first detected at the base of a 3- to 4-layer zone of radially organized cells. With formation of a lumen within the aggregate, laminin and type IV collagen were also deposited in the forming basement membrane. During both posterior neuropore closure and secondary neurulation, fibronectin and heparan sulphate proteoglycan were associated with the most caudal portion of the neuroepithelium, the region where newly formed epithelium merges with the consolidated neuroepithelium. In regions of neural crest migration, the deposition of basement membrane components was altered, lacking laminin and type IV collagen, with increased deposition of fibronectin and heparan sulphate proteoglycan.