Chemical Characterization and Quantification of Proteoglycans in Human Post-Burn Hypertrophic and Mature Scars

1. Samples of normal skin from four patients, post-burn hypertrophic scar from five patients and post-burn mature scar from six patients were analysed for hydroxyproline, water and uronic acid and extracted with guanidinium chloride to yield the proteoglycan pool. A large chondroitin sulphate proteoglycan and biglycan were purified from one hypertrophic scar biopsy and decorin from a normal skin biopsy, by ion-exchange chromatography, gel-filtration and hydrophobic interaction chromatography. These purified proteoglycans were used in an inhibition ELISA assay to estimate the quantities of each in the tissue samples. 2. Samples of post-burn hypertrophic scar had on average 30% less hydroxyproline, 12% more water and 2.4 times as much uronic acid as normal skin. These differences were all statistically significant, whereas the small differences between mature scars and normal skin were not. The content of decorin in hypertrophic scars was only 25% of that in normal skin whereas the large chondroitin sulphate proteoglycan and biglycan were each about 6-fold higher. The mature scars had slightly elevated levels of large chondroitin sulphate proteoglycan and biglycan and a reduced content of decorin compared with normal skin but these differences were not statistically significant. 3. The results suggest that aberrant proteoglycan metabolism is a significant factor contributing to the altered physical properties of hypertrophic scars and that maturation of post-burn scars is dependent on a return of the relative proportions and concentrations of proteoglycans to those characteristic of normal dermis.

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Structural studies on sialylated and sulphated O-glycosidic mannose-linked oligosaccharides in the chondroitin sulphate proteoglycan of brain

Biochemical Journal ◽

10.1042/bj2450229 ◽

1987 ◽

Vol 245 (1) ◽

pp. 229-234 ◽

Cited By ~ 44

Author(s):

T Krusius ◽

V N Reinhold ◽

R K Margolis ◽

R U Margolis

Keyword(s):

Ion Exchange ◽

Chondroitin Sulphate ◽

Gel Filtration ◽

Molecular Size ◽

Ion Exchange Chromatography ◽

Exchange Chromatography ◽

Keratan Sulphate ◽

Size Fractions ◽

Sulphate Proteoglycan ◽

Chondroitin Sulphate Proteoglycan

We have previously described the structures of neutral and sialylated O-glycosidic mannose-linked tetrasaccharides and keratan sulphate polysaccharide chains in the chondroitin sulphate proteoglycan of brain. The present paper provides information on a series of related sialylated and/or sulphated tri- to penta-saccharides released by alkaline-borohydride treatment of the proteoglycan glycopeptides. The oligosaccharides were fractionated by ion-exchange chromatography and gel filtration, and their structural properties were studied by methylation analysis and fast-atom-bombardment mass spectrometry. Five fractions containing [35S]sulphate-labelled oligosaccharides were obtained by ion-exchange chromatography, each of which was eluted from Sephadex G-50 as two well-separated peaks. The apparent Mr values of both the large- and small-molecular-size fractions increased with increasing acidity (and sulphate labelling) of the oligosaccharides. The larger-molecular-size fractions contained short mannose-linked keratan sulphate chains of Mr 3000-4500, together with some asparagine-linked oligosaccharides. The smaller tri- to penta-saccharides, of Mr 800-1400, appear to have a common GlcNac(beta 1-3)Manol core, and to contain one to two residues of sialic acid and/or sulphate.

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Proteoglycans synthesized by an osteoblast-like cell line (UMR 106-01)

Biochemical Journal ◽

10.1042/bj2770199 ◽

1991 ◽

Vol 277 (1) ◽

pp. 199-206 ◽

Cited By ~ 27

Author(s):

D J McQuillan ◽

D M Findlay ◽

A M Hocking ◽

M Yanagishita ◽

R J Midura ◽

...

Keyword(s):

Cell Line ◽

Core Protein ◽

Chondroitin Sulphate ◽

Heparan Sulphate ◽

Gel Filtration ◽

Cell Types ◽

Heparan Sulphate Proteoglycan ◽

Sulphate Proteoglycan ◽

Chondroitin Sulphate Proteoglycan ◽

Umr 106

The proteoglycans synthesized by an osteoblast-like cell line of rat origin (UMR 106-01) were defined after biosynthetic labelling with [35S]sulphate and [3H]glucosamine. Newly synthesized labelled proteoglycans were characterized by differential enzymic digestion in combination with analytical gel filtration and SDS/PAGE. UMR 106-01 cells were found to synthesize three major species of proteoglycan: a large chondroitin sulphate proteoglycan of Mr approximately 1 x 10(6), with a core protein of Mr approximately 350,000-400,000; a small chondroitin sulphate-containing species of Mr approximately 120,000 with a core protein of Mr 43,000; and a heparan sulphate proteoglycan of Mr approximately 150,000, with a core protein of Mr approximately 80,000. Over 70% of the newly synthesized intact proteoglycan species are associated with the cell layer of near-confluent cells; however, accessibility to trypsin digestion suggests an extracellular location. Chemical characteristics of the proteoglycans and preliminary mRNA hybridization indicate that the small chondroitin sulphate proteoglycan is probably PG II (decorin). The large chondroitin sulphate proteoglycan is most likely related to a hyaluronate-aggregating species from fibroblasts (versican), and the heparan sulphate proteoglycan bears striking similarities to cell-membrane-intercalated species described for a number of cell types.

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Fibroblasts During Hypertrophic Scar Formation and Resolution

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100072587 ◽

1973 ◽

Vol 31 ◽

pp. 428-429

Author(s):

C. W. Kischer

Keyword(s):

Electron Microscopy ◽

Scanning Electron Microscopy ◽

Cell Proliferation ◽

Fine Structure ◽

Metabolic Activity ◽

Hypertrophic Scar ◽

Normal Skin ◽

Ground Substance ◽

Hypertrophic Scars ◽

Scanning Electron

The morphology of the fibroblasts changes markedly as the healing period from burn wounds progresses, through development of the hypertrophic scar, to resolution of the scar by a self-limiting process of maturation or therapeutic resolution. In addition, hypertrophic scars contain an increased cell proliferation largely made up of fibroblasts. This tremendous population of fibroblasts seems congruous with the abundance of collagen and ground substance. The fine structure of these cells should reflect some aspects of the metabolic activity necessary for production of the scar, and might presage the stage of maturation.A comparison of the fine structure of the fibroblasts from normal skin, different scar types, and granulation tissue has been made by transmission (TEM) and scanning electron microscopy (SEM).

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Chondroitin Sulphate Proteoglycan 4 (NG2/CSPG4) Localization in Low- and High-Grade Gliomas

Cells ◽

10.3390/cells9061538 ◽

2020 ◽

Vol 9 (6) ◽

pp. 1538 ◽

Cited By ~ 1

Author(s):

Marta Mellai ◽

Laura Annovazzi ◽

Ilaria Bisogno ◽

Cristiano Corona ◽

Paola Crociara ◽

...

Keyword(s):

Cell Transformation ◽

Chondroitin Sulphate ◽

Molecular Genetic ◽

Malignant Gliomas ◽

Malignant Cell ◽

Wild Type ◽

Sulphate Proteoglycan ◽

Chondroitin Sulphate Proteoglycan ◽

Proteoglycan 4

Background: Neuron glial antigen 2 or chondroitin sulphate proteoglycan 4 (NG2/CSPG4) is expressed by immature precursors/progenitor cells and is possibly involved in malignant cell transformation. The aim of this study was to investigate its role on the progression and survival of sixty-one adult gliomas and nine glioblastoma (GB)-derived cell lines. Methods: NG2/CSPG4 protein expression was assessed by immunohistochemistry and immunofluorescence. Genetic and epigenetic alterations were detected by molecular genetic techniques. Results: NG2/CSPG4 was frequently expressed in IDH-mutant/1p19q-codel oligodendrogliomas (59.1%) and IDH-wild type GBs (40%) and rarely expressed in IDH-mutant or IDH-wild type astrocytomas (14.3%). Besides tumor cells, NG2/CSPG4 immunoreactivity was found in the cytoplasm and/or cell membranes of reactive astrocytes and vascular pericytes/endothelial cells. In GB-derived neurospheres, it was variably detected according to the number of passages of the in vitro culture. In GB-derived adherent cells, a diffuse positivity was found in most cells. NG2/CSPG4 expression was significantly associated with EGFR gene amplification (p = 0.0005) and poor prognosis (p = 0.016) in astrocytic tumors. Conclusion: The immunoreactivity of NG2/CSPG4 provides information on the timing of the neoplastic transformation and could have prognostic and therapeutic relevance as a promising tumor-associated antigen for antibody-based immunotherapy in patients with malignant gliomas.

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Morphological studies on endocytosis of chondroitin sulphate proteoglycan by rat liver endothelial cells

Cell and Tissue Research ◽

10.1007/bf00221737 ◽

1988 ◽

Vol 253 (1) ◽

Cited By ~ 18

Author(s):

B�rd Smedsr�d ◽

Marie Malmgren ◽

Jan Ericsson ◽

TorvardC. Laurent

Keyword(s):

Endothelial Cells ◽

Rat Liver ◽

Chondroitin Sulphate ◽

Sulphate Proteoglycan ◽

Morphological Studies ◽

Chondroitin Sulphate Proteoglycan ◽

Liver Endothelial Cells

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Cell surface chondroitin sulphate proteoglycan 4 (CSPG4) binds to the basement membrane heparan sulphate proteoglycan, perlecan, and is involved in cell adhesion

The Journal of Biochemistry ◽

10.1093/jb/mvy008 ◽

2018 ◽

Vol 163 (5) ◽

pp. 399-412 ◽

Cited By ~ 5

Author(s):

Fengying Tang ◽

Megan S Lord ◽

William B Stallcup ◽

John M Whitelock

Keyword(s):

Cell Adhesion ◽

Basement Membrane ◽

Cell Surface ◽

Chondroitin Sulphate ◽

Heparan Sulphate ◽

Heparan Sulphate Proteoglycan ◽

Sulphate Proteoglycan ◽

Chondroitin Sulphate Proteoglycan ◽

Proteoglycan 4

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Chondroitin sulphate proteoglycan in the substratum adhesion sites of Balb/c 3T3 cells. Fractionation on various ion-exchange and affinity columns

Biochemical Journal ◽

10.1042/bj2350469 ◽

1986 ◽

Vol 235 (2) ◽

pp. 469-479 ◽

Cited By ~ 10

Author(s):

B C Wightman ◽

E A Weltman ◽

L A Culp

Keyword(s):

Extracellular Matrix ◽

Affinity Chromatography ◽

Chondroitin Sulphate ◽

Heparan Sulphate ◽

Heparan Sulphate Proteoglycan ◽

3T3 Cells ◽

Platelet Factor ◽

Sulphate Proteoglycan ◽

Adhesion Sites ◽

Chondroitin Sulphate Proteoglycan

Proteoglycans on the cell surface play critical roles in the adhesion of fibroblasts to a fibronectin-containing extracellular matrix, including the model mouse cell line Balb/c 3T3. In order to evaluate the biochemistry of these processes, long-term [35S]sulphate-labelled proteoglycans were extracted quantitatively from the adhesion sites of 3T3 cells, after their EGTA-mediated detachment from the substratum, by using an extractant containing 1% octyl glucoside, 1 M-NaCl and 0.5 M-guanidinium chloride (GdnHCl) in buffer with many proteinase inhibitors. Greater than 90% of the material was identified as a large chondroitin sulphate proteoglycan (Kav. = 0.4 on a Sepharose CL2B column), and the remainder was identified as a smaller heparan sulphate proteoglycan; only small amounts of free chains of glycosaminoglycan were observed in these sites. These extracts were fractionated on DEAE-Sepharose columns under two different sets of elution conditions: with acetate buffer (termed DEAE-I) or with acetate buffer supplemented with 8 M-urea (termed DEAE-II). Under DEAE-I conditions about one-half of the material was eluted as a single peak and the remainder required 4 M-GdnHCl in order to recover it from the column; in contrast, greater than 90% of the material was eluted as a single peak from DEAE-II columns. Comparison of the elution of [35S]sulphate-labelled proteoglycan with that of 3H-labelled proteins from these two columns, as well as mixing experiments, indicated that the GdnHCl-sensitive proteoglycans were trapped at the top of columns, partially as a consequence of their association with proteins in these adhesion-site extracts. Affinity chromatography of these proteoglycans on columns of either immobilized platelet factor 4 or immobilized plasma fibronectin revealed that most of the chondroitin sulphate proteoglycan and the heparan sulphate proteoglycan bound to platelet factor 4 but that only the heparan sulphate proteoglycan bound to fibronectin, providing a ready means of separating the two proteoglycan classes. Affinity chromatography on octyl-Sepharose columns to test for hydrophobic domains in their core proteins demonstrated that a high proportion of the heparan sulphate proteoglycan but none of the chondroitin sulphate proteoglycan bound to the hydrophobic matrix. These results are discussed in light of the possible functional importance of the chondroitin sulphate proteoglycan in the detachment of cells from extracellular matrix and in light of previous affinity fractionations of proteoglycans from the substratum-adhesion sites of simian-virus-40-transformed 3T3 cells.

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A novel keratan sulphate proteoglycan from a human embryonal carcinoma cell line

Biochemical Journal ◽

10.1042/bj2860959 ◽

1992 ◽

Vol 286 (3) ◽

pp. 959-966 ◽

Cited By ~ 22

Author(s):

S Cooper ◽

M F Pera ◽

W Bennett ◽

J T Finch

Keyword(s):

Embryonal Carcinoma ◽

Broad Band ◽

Gel Filtration ◽

Exchange Chromatography ◽

Carcinoma Cells ◽

Embryonal Carcinoma Cell ◽

Embryonal Carcinoma Cells ◽

Keratan Sulphate ◽

Major Band ◽

Sulphate Proteoglycan

We describe here the purification and partial characterization of a 200 kDa keratan sulphate proteoglycan found in the pericellular matrix of human embryonal carcinoma cells. Previously we have shown that this molecule is recognized by a monoclonal antibody (GCTM-2). The antigen was isolated using ion-exchange chromatography and gel filtration, purification being monitored by e.l.i.s.a. using GCTM-2. Metabolic labelling of GCT 27 C-4 embryonal carcinoma cells with sodium [35S]sulphate resulted in the incorporation of [35S]sulphate into the purified molecule. Throughout the purification procedure, the peaks of 35S radioactivity were coincident with the peaks of immunoreactivity, and this label was released both by digestion with keratanase and chondroitinase, confirming the proteoglycan nature of the antigen. The intact molecule ran as a single broad band of 200 kDa, which has been identified by silver staining and immunoblotting following gel electrophoresis. Amino acid analysis of the purified antigen indicated a high content of serine, glycine and aspartic acid/asparagine residues. Visualization by rotary-shadowing electron microscopy suggests that the purified material forms large aggregates, even under denaturing conditions. Deglycosylation of this preparation with trifluoromethanesulphonic acid yielded a major band of 55 kDa and a minor band of 48 kDa. The biochemical nature of the molecule described here, along with tissue distribution studies using GCTM-2, indicates that the antigen is not related to previously described keratan sulphate proteoglycans.

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