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T N Wight

Publications and source records attributed to T N Wight.

At least 109 records · Page 6Linked to original sources
PubMed1986

Modulation of sulfated proteoglycan synthesis by bovine aortic endothelial cells during migration.

The rates of 35S-sulfate incorporation into proteoglycan were compared in multi-scratch wounded and confluent cultures of bovine aortic endothelial cells to determine whether proteoglycan synthesis is altered as cells are stimulated to migrate and proliferate. Incorporation was found to be stimulated in a time-dependent manner, reaching maximal levels 44-50 h after wounding, as cells migrated into wounded areas of the culture dish. Quantitative autoradiography of 35S-sulfate-labeled single-scratch wounded cultures demonstrated a 2-4-fold increase in the number of silver grains over migrating cells near the wound edge when compared to cells remote from the wound edge. Furthermore, when cell proliferation was blocked by inhibition of DNA synthesis, the increase in 35S-sulfate incorporation into proteoglycan after wounding was unaffected. These data indicate that cell division is not required for the modulation of proteoglycan synthesis to occur after wounding. Characterization of the newly synthesized proteoglycan by ion-exchange and molecular sieve chromatography demonstrated that heparan sulfate proteoglycan constitutes approximately 80% of the labeled proteoglycan in postconfluent cultures, while after wounding, chondroitin sulfate proteoglycan and/or dermatan sulfate proteoglycan (CS/DSPG) increases to as much as 60% of the total labeled proteoglycan. These results suggest that CS/DSPG synthesis is stimulated concomitant with the stimulation of endothelial cell migration after wounding.

Animals↗
PubMed1986

The melanoma proteoglycan: restricted expression on microspikes, a specific microdomain of the cell surface.

A cell surface chondroitin sulfate proteoglycan associated with human melanomas and defined by mAb's F24.47 and 48.7 has been characterized biochemically and localized by indirect immunogold electron microscopy. These antibodies recognize distinct epitopes on the intact proteoglycan. In addition, mAb 48.7 also recognizes an epitope on a 250,000-D glycoprotein and is therefore similar to antibody 9.2.27 (described by Bumol, T.F., and R.A. Reisfeld, 1982, Proc. Natl. Acad. Sci. USA., 79:1245-1249). Furthermore, it was shown that the glycosaminoglycan chains released by alkaline borohydride treatment of the proteoglycan recognized by mAb 48.7 had a size of approximately 60,000 D. Since the intact proteoglycan was estimated to be 420,000 D, there are probably three chondroitin sulfate chains attached to the 250,000-D core glycoprotein. Furthermore, an oligosaccharide fraction containing 42% of the 3H activity (glucosamine as precursor) was isolated. Immunolocalization studies using whole-mount electron microscopy revealed that the chondroitin sulfate proteoglycan was present almost exclusively on microspikes, a microdomain of the melanoma cell surface. These processes were present as 1-2-micron structures on the upper cell surface and as longer (up to 20 micron) structures at the cell periphery. Peripheral microspikes were involved in the initial interactions between adjacent cells and formed complex footpads that made contact with the substratum. Immunogold-labeled cells were also thin sectioned and the specific localization of the chondroitin sulfate proteoglycan antigen was quantitated. The data confirmed the results of whole-mount microscopy and demonstrated a statistically significant association of the antigen with the microspike processes as compared with other areas of the cell surface. By using two different mAb's (48.7 and F24.47) that recognize epitopes on either the core glycoprotein or the intact proteoglycan, respectively, we have demonstrated that both molecules have the same restricted distribution at the cell surface. The specific localization of the antigen to microspikes at the cell surface suggests it may play a role in cell-cell contact and cell-substratum adhesion, which could be important in the metastatic process.

Aggrecans↗
PubMed1986

Undersulfated chondroitin sulfate in cartilage from a miniature poodle with spondyloepiphyseal dysplasia.

In order to determine if either the proteoglycans or collagen in the cartilagenous epiphyses of a Miniature Poodle with spondyloepiphyseal dysplasia were abnormal, the cartilage was dissociatively extracted in 4 M guanidine HCl in the presence of protease inhibitors and subjected to isopycnic cesium chloride dissociative density gradient ultracentrifugation. Dissociative extraction solubilized 97% of the uronic acid and 88% of the protein. Uronic acid distributed anomalously in the density gradient in that about 1/3 was recovered in each of the D1 (1.58 g/ml), D2 (1.49 g/ml) and D3 (1.44 g/ml) fractions. Proteoglycans in the D1, D2 and D3 fractions also eluted from Sepharose CL-2B columns in a manner indicative of monomers of a smaller apparent hydrodynamic size than those from normal canine growth plate or articular cartilage. D1, D2 and D3 monomers subjected to the sodium borohydride reaction followed by chromatography on a Sepharose CL-6B column yielded glycosaminoglycan chain molecular weights of 10,200 (D1), 7600 (D2) and 6200 (D3). High pressure liquid chromatography on a Whatman Partisil 10PAC column of the chondroitinase AC II digests of D1, D2 and D3 fractions revealed that 60% of the D1, 81% of the D2 and 88% of the D3 unsaturated disaccharides eluted in the delta DiOS-delta DiHA position. Subsequent HPLC of the unsaturated disaccharides on the Hypersil APS column resulted in the recovery of 97% of the nonsulfated unsaturated disaccharides in the delta DiOS position. Associative extraction in 0.5 M guanidine followed by associative gradient ultracentrifugation resulted in the recovery of 27% of the uronic acid in the aA1 and 47% in the aA2 fractions. Two dimensional SDS gel electrophoresis of the CNBr peptides of the collagen isolated by pepsin digestion and 0.9 M NaCl precipitation revealed type II collagen. This study has demonstrated that spondyloepiphyseal dysplasia in a Miniature Poodle is characterized by cartilage containing undersulfated chondroitin sulfate proteoglycan.

Animals↗
PubMed1986

Proteoglycans in human long-term bone marrow cultures: biochemical and ultrastructural analyses.

Proteoglycans within the extracellular matrix of human bone marrow have been implicated in the process of hematopoiesis, but little is known about the structure and composition of these macromolecules in this tissue. Hematopoietically active human long-term bone marrow cultures were incubated with medium containing 35S-sulfate and 3H-glucosamine as labeling precursors. Proteoglycans present in the medium and cell layer were extracted with 4 mol/L guanidine HCI and purified by diethylaminoethyl (DEAE)-Sephacel ion exchange and molecular sieve chromatography. Both culture compartments contain a large chondroitin sulfate proteoglycan (MI, CI) that eluted in the void volume of a Sepharose CL-4B column and contained glycosaminoglycan chains of molecular weight (mol wt) approximately 38,000. A second population of sulfate-labeled material was identified as a broad heterogenous peak (MII, CII) that was included on Sepharose CL-4B at Kav = 0.31. This material when chromatographed on Sepharose CL-6B could be further separated into a void peak (MIIa, CIIa) and an included peak eluting at Kav = 0.39 (MIIb, CIIb). The void peaks (MIIa, CIIa) were susceptible to chondroitinase ABC digestion (99%) but slightly less susceptible to chondroitinase AC digestion (90%). Papain digestion of these peaks revealed them to be proteoglycans with glycosaminoglycan chains of mol wt approximately 38,000. The included peaks on Sepharose CL-6B (MIIb, CIIb) from both medium and cell layer compartments resisted digestion with papain, indicating the presence of glycosaminoglycan chains of mol wt approximately 38,000 either free or attached to a small peptide. Although this material was susceptible to chondroitinase ABC (98%), it was considerably less susceptible to chondrotinase AC (approximately 60%), indicating that it contained dermatan sulfate. A small amount of heparan sulfate proteoglycan was also identified but constituted only approximately 10% of the total sulfated proteoglycan extracted from these cultures. Additionally, approximately 40% of the incorporated 3H-activity radioactivity was present as hyaluronic acid. Electron microscopy revealed a layer of adherent cells covered by a mat containing ruthenium red-positive granules that were connected by thin filaments. The extracellular matrix layer above the adherent cells contained a mixture of hematopoietic cells. Chondroitinase ABC treatment of the cultures completely removed the ruthenium red-positive granules overlying the cells and resulted in a loss of approximately 70% of the 35S-sulfate-labeled material from the cell layer.(ABSTRACT TRUNCATED AT 400 WORDS)

Bone Marrow↗
PubMed1985

Light microscopic immunolocation of thrombospondin in human tissues.

Affinity-purified antisera against thrombospondin were used to locate the presence of this glycoprotein in frozen sections of several human tissues by immunofluorescence techniques. Immunostaining was observed in the peritubular connective tissue and in basement membrane regions beneath glandular epithelium in skin and lung. Intense immunostaining was observed at the dermal-epidermal junction in skin and in small blood vessels throughout this tissue. Skeletal muscle exhibited positive staining with anti-thrombospondin antisera within interstitial areas. Immunostaining was confined to the luminal portions of large blood vessels such as aorta. In large blood vessels that contained lesions of atherosclerosis, immunostaining was observed throughout the lesion area and was especially prominent surrounding some of the lesion cells. These results indicate that thrombospondin is located within the matrix of a variety of human tissues and supports the suggestion that this glycoprotein is an endogenous component of some extracellular matrices.

Aorta↗
PubMed1985

Proteoglycans in pathological conditions: atherosclerosis.

Proteoglycans accumulate within the innermost layer (intima) of blood vessels during atherosclerosis. This accumulation is marked in some forms of human atherosclerosis and is particularly prominent in vessels that have been experimentally injured and have healed by the process of reendothelialization. The two major cell types of the arterial wall, endothelium and smooth muscle, are the major sources of arterial proteoglycans, and cell cultures have demonstrated that these cells synthesize at least three families of proteoglycans similar to those present in human aorta. Each family differs with regard to molecular size, glycosaminoglycan and oligosaccharide content, and ability to aggregate in the presence of hyaluronic acid. Furthermore, each cell type possesses a distinct pattern of proteoglycan synthesis. Smooth muscle cells synthesize and secrete primarily chondroitin sulfate and dermatan sulfate-containing proteoglycans, whereas endothelial cells synthesize and secrete large amounts of heparan sulfate proteoglycan. Evidence is presented to indicate that the synthesis of proteoglycans is modulated as a function of growth and migratory state of the vascular cells.

Animals↗
PubMed1985

Chondrodysplasia in the Alaskan malamute. Characterization of proteoglycans dissociatively extracted from dwarf growth plates.

Proteoglycan monomers obtained from the dissociative extraction of growth plate cartilages of chondrodysplastic and homozygous nonaffected Alaskan malamute dogs were characterized with regard to hydrodynamic size and glycosaminoglycan composition. Dissociative extraction solubilized 91.7% of the uronic acid and 71.7% of the protein from dwarf growth plates compared to 76.8% of the uronic acid and 50.2% of the protein from normal growth plates. Dissociative density gradient ultracentrifugation of the extracts resulted in the recovery of 84% of the uronic acid from dwarf growth plates and 71% of the uronic acid from normal growth plates in the D1 fraction. High-pressure liquid chromatography of the dwarf D1 monomers revealed a single peak with a retention time of 8.6 minutes while the normal D1 monomers eluted later with a retention time of 8.9 minutes. After reduction of the dwarf D1 monomers, the chondroitin sulfate side chains eluted from Sepharose CL-6B with an approximate molecular weight of 15,000 (Kav of 0.55) while those from the normal eluted with an estimated molecular weight of 9,500 (Kav of 0.64). High-pressure liquid chromatography analysis of the unsaturated disaccharides from the dwarf D1 fractions revealed increased amounts of chondroitin-6-sulfate. Analysis of the fractions for glucosamine and galactosamine revealed that dwarf D1 and D2 fractions were enriched in galactosamine. These findings indicate that the extracellular matrices of dwarf growth plates contain proteoglycan monomers which may be indicative of a less mature extracellular cartilage matrix than the cartilage matrices of age-matched normal dogs.

Animals↗
PubMed1984

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.

Animals↗
PubMed1984

Proteoglycans in arterial smooth muscle cell cultures: an ultrastructural histochemical analysis.

The extracellular matrix in cultures of arterial smooth muscle cells has been examined by ultrastructural histochemistry using each of the following cationic dyes: ruthenium red, Alcian blue, acridine orange, and safranin O. All dyes exhibited an affinity for a structural component that was either preserved as a granule with ruthenium red or Alcian blue, or as an extended filament or bottlebrush structure with acridine orange or safranin O. Both granules and filaments were removed when the cultures were pretreated with chondroitinase ABC, an enzyme that degrades the glycosaminoglycan moiety of some proteoglycans. These structural components of the extracellular matrix were not observed when cultures were prepared in the absence of the cationic dyes. Labeling experiments (35S-sulfate) revealed that approximately 40% of the total labeled proteoglycans were lost during routine processing for electron microscopy (i.e., fixation through dehydration). Inclusion of any one of the cationic dyes during fixation reduced the losses to less than 1%. The extended filamentous structure preserved by safranin O and acridine orange resembled the structure of purified proteoglycans prepared from the same cultures and spread on cytochrome c monolayer films. These observations suggest that proteoglycans exist as extended bottlebrush structures within the extracellular matrix, and support the interpretation that the granular deposits observed in the ruthenium red and Alcian blue preparations most likely represent individual proteoglycan monomers that have undergone molecular collapse during processing. In addition, the dyes also exhibited an affinity for chords of fine fibrils that contained small granules and/or filaments. Both the fibrillar material and the associated granular and filamentous structures enmeshed in the fibrils resisted digestion with chondroitinase ABC.

Acridine Orange↗
PubMed1984

Human atherosclerosis. I. Cell constitution and characteristics of advanced lesions of the superficial femoral artery.

This study represents a systematic analysis of the fine-structural characteristics of atherosclerotic lesions of the superficial femoral artery in man together with the growth characteristics in culture of the smooth muscle cells derived from these lesions. Occlusive fibrous atherosclerotic plaques were obtained from 29 male patients at the time of bypass surgery for occlusion of the superficial femoral artery and were studied by light and transmission electron microscopy. The occluded segment of each artery was obtained immediately after removal from the patient and examined with sterile techniques, and representative segments were fixed for light- and electron-microscopic study. Adjacent segments were used for dissection of the lesion away from the underlying media, and smooth muscle cells were cultured from lesion and nonlesion areas and compared in terms of their growth responses to increasing concentrations of a pool of human whole blood serum. The majority of the lesions were fibroproliferative and contained relatively little lipid. The fibrous cap that covered each lesion consisted of a special form of dense connective tissue that contained flat, pancake-shaped smooth muscle cells in a lacunalike space. This space consisted of concentric layers of basement membrane, collagen fibrils, and proteoglycan. The majority of the cells beneath the fibrous cap were smooth muscle cells mixed with small but varying numbers of macrophages. Most of the lesions were occluded by a thrombus, which had undergone organization and recanalization. A small number of the lesions had deep lipid deposits together with foci of degeneration and calcification. The occluded thrombi contained smooth muscle cells and a larger proportion of macrophages than the lesions themselves. The in vitro growth properties of the smooth muscle cells isolated from the lesion and the underlying media suggested that the lesion cells had senesced, compared with the medial smooth muscle cells derived from the same artery.

Aged↗
PubMed1983

Proteoglycans synthesized by smooth muscle cells derived from monkey (Macaca nemestrina) aorta.

Smooth muscle cells derived from monkey aorta were cultured in medium with [35S]sulfate and [3H]glucosamine as labeling precursors. Proteoglycans in the medium and in 4 M guanidine HCl extracts of the cell layer were purified by DEAE-Sephacel and molecular sieve chromatography. Both preparations contained a predominant, large chondroitin sulfate proteoglycan (Kav = 0.30 on Sepharose CL-2B) with glycosaminoglycan chains of Mr approximately 43,000 average containing a ratio of 6-sulfate to 4-sulfate of approximately 2. Approximately 7 and 27% of the 3H label in this proteoglycan were present in N-linked and O-linked oligosaccharides, respectively. Reaggregation experiments indicated that a large proportion of these proteoglycans can form link protein-stabilized aggregates. The medium fraction also contained a smaller dermatan sulfate proteoglycan (Kav = 0.67 on Sepharose CL-2B) with glycosaminoglycan chains of Mr approximately 43,000 containing a ratio of 6-sulfate to 4-sulfate of about 0.5. This proteoglycan contained approximately the same percentage of N-linked oligosaccharides as the chondroitin sulfate proteoglycan, but few or no O-linked oligosaccharides. A smaller dermatan sulfate proteoglycan with a single chain was present only in the cell layer. Additionally, small amounts of heparan sulfate proteoglycans were synthesized by the cells.

Animals↗
PubMed1983

Sulfated proteoglycans synthesized by vascular endothelial cells in culture.

Metabolically labeled proteoglycans were isolated both from the culture medium and from the cell layer of cultured bovine aortic endothelial cells. Proteoglycans were fractionated by sequential gel filtration on Sepharose CL-4B, DEAE-cellulose column chromatography, and CsCl density gradient centrifugation. The culture medium contained two distinct proteoglycans: a large proteoglycan (Kav = 0.07) of low buoyant density containing heparan sulfate side chains (Mr congruent to 36,000) and a smaller proteoglycan (Kav = 0.45) of high density containing chondroitin sulfate chains (Mr congruent to 20,000). The chondroitin sulfate proteoglycan fraction contained a small amount (less than 10%) of dermatan sulfate. A very similar low density heparan sulfate proteoglycan was extracted from the cell layer with 2% sodium dodecyl sulfate in the presence of enzyme inhibitors. In addition, there was a high density proteoglycan of small size (Kav = 0.43) containing heparan sulfate side chains (Mr congruent to 20,000) in the cell layer. Analyses of proteoglycans synthesized by cultured human umbilical vein endothelial cells gave similar results, except that these cells produced more dermatan sulfate and unidentified oversulfated chondroitin sulfate chains. Morphologically atypical endothelial cells contained reduced levels of the large heparan sulfate proteoglycan. Both indirect immunofluorescence and direct immunoelectron microscopy revealed that the basement membrane-like matrix under monolayers of bovine endothelial cells reacted with antibodies against the basement membrane proteoglycan isolated from a basement membrane-producing tumor. By electron microscopy, this material was shown to consist of a fine filamentous meshwork containing discrete 10-20-nm diameter ruthenium red positive granules resembling those present in basement membranes of intact arteries.

Animals↗
PubMed1983

Neovascular responses induced by cultured aortic endothelial cells.

Neovascularization was studied in the chorioallantoic membrane of the chick embryo after implantation of bovine aortic endothelial and smooth muscle cells, Swiss and BALB/c 3T3 cells and human diploid fibroblasts cultured separately on microcarrier beads. Quantitative analysis of neovascularization indicated a 3 1/2-fold increase in the number of blood vessels responding to endothelial cells while smooth muscle cells induced a twofold increase when compared to the response of beads without cells. Skin fibroblasts and Swiss 3T3 cells did not elicit a comparable response. The marked angiogenic response induced by endothelial cells was characterized by a 137% increase in total vessel length and a 35% increase in average vessel area when compared to controls. Two of the properties required for an angiogenesis factor--stimulation of cellular migration and proliferation--can also be demonstrated using endothelial cell-conditioned medium in cell culture systems. Medium from cultured bovine aortic endothelium stimulates DNA synthesis, proliferation, and migration of smooth muscle cells. In addition, conditioned media from both endothelial cells and smooth muscle cells produced an angiogenic response in the chorioallantoic membrane assay, which was comparable to that produced by intact cells growing on microcarrier beads. Similar responses were not evident with medium conditioned by other cell types. These results indicate the potential importance of endothelial cells and endothelial cell products in regulating blood vessel growth.

Allantois↗
PubMed1983

Proteoglycans in primate arteries. III. Characterization of the proteoglycans synthesized by arterial smooth muscle cells in culture.

Near confluent monolayers of arterial smooth muscle cells derived from Macaca nemestrina were labeled with Na2[35S]O4 and the newly synthesized proteoglycans present in the culture medium and cell layer were extracted with either 4 M guanidine HCl (dissociative solvent) or 0.5 M guanidine HCl (associative solvent) in the presence of protease inhibitors. The proteoglycans in both compartments were further purified by cesium chloride density gradient ultracentrifugation. Two size classes of proteoglycans were observed in the medium as determined by chromatography on Sepharose CL-2B. The large population (Kav = 0.31) contained predominantly chondroitin sulfate chains with Mr = approximately 40,000. The smaller population (Kav = 0.61) contained dermatan sulfate chains of similar Mr (approximately 40,000). When tested for their ability to aggregate, only proteoglycans in the large-sized population were able to aggregate. A chondroitin sulfate containing proteoglycan with identical properties was isolated from the cell layer. In addition, the cell layer contained a dermatan sulfate component which eluted later on Sepharose CL-2B (Kav = 0.78) than the dermatan sulfate proteoglycan present in the medium. Electron microscopy of the purified proteoglycans revealed a bottlebrush structure containing a central core averaging 140 nm in length with an average of 8 to 10 side projections. The length of the side projections varied but averaged between 70 and 75 nm. Similar bottlebrush structures were observed in the intercellular matrix of the smooth muscle cell cultures after staining with Safranin 0. This culture system provides a model to investigate parameters involved in the regulation of synthesis and degradation of arterial proteoglycans.

Animals↗
PubMed1983

Comparative rates of decline in the primary cloning efficiencies of smooth muscle cells from the aging thoracic aorta of two murine species of contrasting maximum life span potentials.

Primary cloning assays of thoracic aortic smooth muscle cells from an F1 hybrid strain of Mus musculus demonstrated linear regressions of replicative potentials as functions of donor age (6-30 months), with regression coefficients, in two independent cohorts, of -1.69 +/- 0.27 (SE) and -1.88 +/- 0.19 (SE) clones per milligram wet weight of intima-media per month and correlation coefficients of -0.83 and -0.92 (P less than 0.001). Secondary cloning (dilute plating from first passages) also demonstrated a high negative correlation (r = -0.90) between donor age and cloning efficiency, thus implicating intrinsic differences in cell populations. Comparable primary cloning assays on the aortas of aging cohorts of Peromyscus leucopus, a murine species with a maximum life span potential approximately twice that of Mus musculus, yielded about twice the number of clonable smooth muscle cells per unit weight; the rate of decline with age was slightly but significantly greater (P less than 0.01). Electron-microscopic studies revealed cellular alterations confined to the first subintimal layer of aortas from mice (Mus musculus) 18 months and older.

Age Factors↗
PubMed1983

Effect of endothelium on glycosaminoglycan accumulation in injured rabbit aorta.

Previous studies have indicated that reendothelialized regions of injured rabbit aortas are more susceptible to diet-induced atherosclerosis than persistently deendothelialized regions or uninjured aortas. However, the mechanism responsible for this selective lipid deposition is not understood. One possibility is that these regions differ with respect to the quantity and type of glycosaminoglycan-containing proteoglycans which are known to interact with lipoproteins. To determine whether these regions differed with respect to their glycosaminoglycan composition, the authors divided 53 rabbits into four groups. Groups IA and IB were fed a regular diet beginning 5 weeks prior to aortic deendothelialization; Groups IIA and IIB were fed the same diet supplemented with 0.5% cholesterol. The rabbits were continued on these diets following aortic deendothelialization with a balloon catheter. Those in Groups IA and IIA were sacrificed either at 2-5 weeks or 6-8 weeks following deendothelialization; proteoglycans were assessed morphometrically following staining with alcian blue. Groups IB and IIB were sacrificed at 10 weeks following injury; glycosaminoglycans were extracted from deendothelialized and reendothelialized aortas, separated by electrophoresis, and quantitated by scanning densitometry. Morphometric analysis of stained aortic sections revealed significantly increased quantities of alcianophilic material in the neointima of reendothelialized aortas as compared with deendothelialized aortas in both diet groups. Chemical analysis revealed significantly more of each glycosaminoglycan in reendothelialized aortas when compared with deendothelialized or uninjured aortas. The major glycosaminoglycans present in all regions were heparan sulfate and chondroitin sulfate; and although absolute quantities of these particular glycosaminoglycans increased in the reendothelialized region, their relative percentages remained the same for each area analyzed. Cholesterol feeding did not appear to influence glycosaminoglycan concentration and composition in reendothelialized and deendothelialized regions when compared with normal diets, but cholesterol feeding alone did increase aortic glycosaminoglycans in uninjured aortas. The results suggest that the presence of endothelium influences the quantity and type of glycosaminoglycans accumulating in the neointima, and that the differences in proteoglycans in the reendothelialized artery may account at least in part for the propensity of this area to accumulate lipid and evolve as atherosclerosis.

Animals↗
PubMed1982

Isolation and characterization of proteoglycans synthesized by human colon and colon carcinoma.

The biosynthesis of proteoglycans in short term organ culture of human colon and colon carcinoma was studied. Proteoglycans, labeled with [35S]sulfate and [3H]serine, were extracted with either 4 M or 0.5 M guanidine HCl in the presence of protease inhibitors and sequentially purified by associative and dissociative CsCl density gradient ultracentrifugation. Normal colon synthesized two polydisperse classes of proteoglycans: a large heparan sulfate-containing monomer, with a Kav of 0.48 on Sepharose CL-2B and a small dermatan sulfate-containing monomer with a Kav of 0.65. A portion (25%) of the proteoglycans was found as aggregate when chromatographed under associative conditions, and the larger monomers interacted with hyaluronic acid to an extent greater than the smaller proteoglycans. Following papain or alkali treatment, the free glycosaminoglycan side chains of both monomers eluted as a single broad peak (Kav = 0.5) from Sepharose CL-6B, with an estimated Mr of 20 X 10(3). In contrast, colon carcinoma synthesized only one proteoglycan monomer, which aggregated to a limited extent (12%). This proteoglycan population, with a Kav of 0.7 on CL-2B, contained chondroitin sulfate as the major glycosaminoglycan (greater than 81%), with small amounts of dermatan sulfate. The glycosaminoglycans had an estimated Mr of 9 X 10(3), and the disaccharides released by chondroitinase ABC consisted of 32% 4-sulfate and 68% 6-sulfate. Electron microscopy of mixed proteoglycancytochrome c monolayers from the associative fractions of normal and neoplastic colon revealed aggregated complexes which were similar in over structure, although smaller than the proteoglycan aggregates from cartilage.

Amino Acids↗