scholarly journals A comparative biochemical and ultrastructural study of proteoglycan–collagen interactions in corneal stroma. Functional and metabolic implications

1990 ◽  
Vol 270 (2) ◽  
pp. 491-497 ◽  
Author(s):  
J E Scott ◽  
T R Bosworth
Keyword(s):  
Guinea Pig ◽  
Alcian Blue ◽  
Chondroitin Sulphate ◽  
Corneal Thickness ◽  
Corneal Stroma ◽  
Human Cornea ◽  
Dermatan Sulphate ◽  
Keratan Sulphate ◽  
Cellulose Acetate Membranes ◽  
O2 Supply

1. Corneas of mouse, rat, guinea pig, rabbit, sheep, cat, dog, pig and cow were quantitatively analysed for water, hydroxyproline, nucleic acid, total sulphated polyanion, chondroitin sulphate/dermatan sulphate and keratan sulphate, several samples or pools of tissue from each species being used. Ferret cornea was similarly analysed for water and hydroxyproline on one pool of eight corneas. Pooled frog (38) and ferret (eight) corneas and a single sample of human cornea were qualitatively examined for keratan sulphate and chondroitin sulphate/dermatan sulphate by electrophoresis on cellulose acetate membranes. Nine species (mouse, frog, rat, guinea pig, rabbit, sheep, cat, pig and cow) were examined by light microscopy and six (mouse, frog, rat, guinea pig, rabbit and cow) by electron microscopy, with the use of Alcian Blue or Cupromeronic Blue in critical-electrolyte-concentration (CEC) methods to stain proteoglycans. 2. Water (% of wet weight), hydroxyproline (mg/g dry wt.) and chondroitin sulphate (mg/g of hydroxyproline) contents were approximately constant across the species, except for mouse. 3. Keratan sulphate contents (mg/g of hydroxyproline) increased with corneal thickness, whereas dermatan sulphate contents decreased. The oversulphated domain of keratan sulphate was absent from mouse and frog corneas, increasing as percentage of total keratan sulphate with increasing corneal thickness. Sulphation of dermatan sulphate was essentially complete (i.e. one sulphate group per disaccharide unit). 4. Chondroitin sulphate/dermatan sulphate proteoglycans were present at the d bands of the collagen fibrils of all species examined, orthogonally arrayed, with high frequency, and occasionally at the e bands. Keratan sulphate proteoglycans were present at the a and c bands of all species examined, but with far higher frequency in the thicker corneas, where keratan sulphate contents were high. 5. Alcian Blue CEC staining showed much higher sulphation of keratan sulphate in thick corneas, e.g. that of cow, than in thin corneas, e.g. that of mouse, in keeping with biochemical analyses. 6. It is suggested that the constancy of interfibrillar volumes is regulated via the swelling and osmotic pressure of the interfibrillar polyanions, by adjustment of the extent of sulphation in two independent proteoglycan populations, to achieve an ‘average sulphation’ of the total polyanion similar to that of fully sulphated chondroitin sulphate/dermatan sulphate. 7. The balance of synthesis of the two kinds of proteoglycans may be determined by the O2 supply to the avascular cornea. O2 supply may also determine the conversion of chondroitin sulphate into dermatan sulphate.

scholarly journals Fine-mapping and cell-specific enrichment at corneal resistance factor loci prioritize candidate causal regulatory variants

2020 ◽  
Vol 3 (1) ◽  
Author(s):  
Xinyi Jiang ◽  
Nefeli Dellepiane ◽  
Erola Pairo-Castineira ◽  
Thibaud Boutin ◽  
Yatendra Kumar ◽  
...  
Keyword(s):  
Fine Mapping ◽  
Association Studies ◽  
Corneal Thickness ◽  
Corneal Stroma ◽  
Resistance Factor ◽  
Open Chromatin ◽  
Human Cornea ◽  
Genome Wide Association Studies ◽  
Corneal Resistance Factor ◽  
Matrix Gene

AbstractCorneal resistance factor (CRF) is altered during corneal diseases progression. Genome-wide-association studies (GWAS) indicated potential CRF and disease genetics overlap. Here, we characterise 135 CRF loci following GWAS in 76029 UK Biobank participants. Enrichment of extra-cellular matrix gene-sets, genetic correlation with corneal thickness (70% (SE = 5%)), reported keratoconus risk variants at 13 loci, all support relevance to corneal stroma biology. Fine-mapping identifies a subset of 55 highly likely causal variants, 91% of which are non-coding. Genomic features enrichments, using all associated variants, also indicate prominent regulatory causal role. We newly established open chromatin landscapes in two widely-used human cornea immortalised cell lines using ATAC-seq. Variants associated with CRF were significantly enriched in regulatory regions from the corneal stroma-derived cell line and enrichment increases to over 5 fold for variants prioritised by fine-mapping-including at GAS7, SMAD3 and COL6A1 loci. Our analysis generates many hypotheses for future functional validation of aetiological mechanisms.

scholarly journals The presence of a cartilage-like proteoglycan in the adult human meniscus

1981 ◽  
Vol 197 (1) ◽  
pp. 77-83 ◽  
Author(s):  
P J Roughley ◽  
D McNicol ◽  
V Santer ◽  
J Buckwalter
Keyword(s):  
Hyaluronic Acid ◽  
Chondroitin Sulphate ◽  
Gradient Centrifugation ◽  
Large Subunit ◽  
Dermatan Sulphate ◽  
Keratan Sulphate ◽  
Adult Human ◽  
Cscl Density Gradient ◽  
Cartilage Proteoglycans ◽  
Aggregate Structures

Proteoglycans were extracted from the adult human meniscus under dissociative conditions and purified by CsCl-density-gradient centrifugation. The preparations of highest density contained proteoglycan that possessed the ability to interact with hyaluronic acid, was of large subunit size and was composed of chondroitin sulphate, keratan sulphate and sialic acid-containing oligosaccharides. This ‘cartilage-like’ proteoglycan also exhibited subunit and aggregate structures analogous to those of hyaline-cartilage proteoglycans when examined by electron microscopy. However, the composition of this proteoglycan was more comparable with proteoglycans from immature cartilage than from age-matched cartilage. The preparations from lower density, which were enriched in dermatan sulphate, contained smaller proteoglycan that was not able to interact with hyaluronic acid. This non-aggregating proteoglycan may be structurally distinct from the ‘cartilage-like’ proteoglycan, which does not contain dermatan sulphate.

scholarly journals The nature of the non-ultrafilterable glycosaminoglycans of normal human urine

1971 ◽  
Vol 122 (3) ◽  
pp. 373-384 ◽  
Author(s):  
E. Wessler
Keyword(s):  
Chondroitin Sulphate ◽  
Heparan Sulphate ◽  
Dermatan Sulphate ◽  
Keratan Sulphate ◽  
Molar Ratios ◽  
Sodium Barbital ◽  
Normal Human ◽  
Normal Men ◽  
Barbital Buffer ◽  
Acidic Glycosaminoglycans

1. The non-ultrafilterable acidic glycosaminoglycans from pooled urine of normal men, aged about 20, were isolated and characterized. The isolation procedure included digestion with sialidase and pronase, and fractionation by stepwise elution from an ECTEOLA-cellulose column. The glycosaminoglycans in each fraction were separated from each other by preparative electrophoresis in sodium barbital buffer and in barium acetate. 2. Approximate relative amounts of the different glycosaminoglycans were: chondroitin sulphate 60%, chondroitin 2%, hyaluronic acid 4%, dermatan sulphate 1%, heparan sulphate 15% and keratan sulphate 18%. Chondroitin sulphate–dermatan sulphate hybrids seemed to occur in trace amounts. 3. Chondroitin sulphate, heparan sulphate and keratan sulphate were heterogeneous with respect to degree of sulphation. Two distinct groups of chondroitin sulphate fractions were found, with sulphate/hexosamine molar ratios of about 0.5 and 1 respectively. The sulphate/hexosamine molar ratios in the heparan sulphate fractions varied from 0.5 to 0.9; the N-sulphate/hexosamine ratio was about 0.5 in all fractions. The sulphate/hexosamine molar ratios in the keratan sulphate fractions varied from 0.2 to 0.7.

scholarly journals Urinary excretion of sulphated N-acetylhexosamines in patients with various mucopolysaccharidoses

1985 ◽  
Vol 229 (3) ◽  
pp. 579-586 ◽  
Author(s):  
J J Hopwood ◽  
H Elliott
Keyword(s):  
Human Urine ◽  
Chondroitin Sulphate ◽  
Mucopolysaccharidosis Type ◽  
Type Ii ◽  
Dermatan Sulphate ◽  
Keratan Sulphate ◽  
Normal Individuals ◽  
Deficient Patient ◽  
Mucolipidosis Type ◽  
Normal Controls

Sulphated N-acetylhexosamines have been isolated from human urine and tentatively identified as N-acetylglucosamine 6-sulphate (GlcNAc6S), N-acetylgalactosamine 6-sulphate (GalNAc6S), N-acetylgalactosamine 4-sulphate (GalNAc4S) and N-acetylgalactosamine 4,6-disulphate (GalNAc4,6diS). Urine from mucopolysaccharidosis-Type-IIID, -IVA and -VI patients compared with that from normal individuals contains elevated levels of GlcNAc6S (380-fold), GalNAc6S (180-fold) and GalNAc4S (420-fold) respectively. Urine from mucopolysaccharidosis-Type-VI patients also contain more than 600 times the normal level of GalNAc4,6diS. Urine from a mucolipidosis-Type-II and a multiple-sulphatase-deficient patient, and, in general, all mucopolysaccharidosis patients studied, contain at least 5-10-fold elevations of sulphated N-acetylhexosamines over the levels detected in urine from normal controls and a alpha-mannosidosis patient. Urine from patients with clinically mild phenotypes contains less sulphated N-acetylhexosamines than isolated from urine of clinically severe mucopolysaccharidosis patients. The source of the four sulphated N-acetylhexosamines is not known. However, incubation of a series of oligosaccharide substrates, derived from keratan sulphate and chondroitin 6-sulphate and containing non-reducing-end beta-linked 6-sulphated N-acetylhexosamine residues, with homogenates of cultured human skin fibroblasts has indirectly been shown to release GlcNA6S and GalNAc6S respectively. Release of GalNAc4S could not be demonstrated in similar incubations of oligosaccharide substrates derived from chondroitin 4-sulphate and containing non-reducing-end beta-linked GalNAc4S residues. We propose that some, if not all, of the sulphated N-acetylhexosamine present in human urine is derived from the action of beta-N-acetylhexosaminidase on sulphated GlcNAc or GalNAc residues at the non-reducing end of keratan sulphate, dermatan sulphate or chondroitin sulphate.

scholarly journals Fractionation of proteoglycans from bovine corneal stroma

1975 ◽  
Vol 145 (3) ◽  
pp. 491-500 ◽  
Author(s):  
I Axelsson ◽  
D Heinegård
Keyword(s):  
Chondroitin Sulphate ◽  
Corneal Stroma ◽  
Molecular Size ◽  
Equal Weight ◽  
Keratan Sulphate ◽  
Before And After ◽  
Weight One ◽  
Tenfold Excess ◽  
Covalently Linked ◽  
The Individual

Proteoglycans were extracted from bovine corneal stroma with 4M-guanidinum chloride, purified by DEAE-dellulose chromatography (Antonopoulos et al., 1974) and fractionated by precipitation with ethanol into three fractions of approximately equal weight. One of these fractions consisted of a proteoglycan that contained keratan sulphate as the only glycosaminoglycan. In the othertwo fractions proteoglycans that contained chondroitin sulphate, dermatan sulphate and keratan sulphate were present. Proteoglycans which had a more than tenfold excess of galactosaminoglycans over keratan sulphate could be obtianed by further subfractionation. The gel-chromatographic patterns of the glucosaminoglycans before and after digestion with chondroitinase AC differed for the fractions. The individual chondroitin sulphate chains seemed to be larger in cornea than in cartilage. Oligosaccharides, possibly covalently linked to the protein core of the proteoglycans, could be isolated from all fractions. The corneal proteoglycans were shown to have higher protein contents and to be of smaller molecular size than cartilage proteoglycans.

Proteoglycan-collagen interactions in intervertebral disc. A chondroitin sulphate proteoglycan associates with collagen fibrils in rabbit annulus fibrosus at the d-e bands

1986 ◽  
Vol 6 (10) ◽  
pp. 879-888 ◽  
Author(s):  
J. E. Scott ◽  
M. Haigh
Keyword(s):  
Electron Microscopy ◽  
Intervertebral Disc ◽  
Annulus Fibrosus ◽  
Electrolyte Concentration ◽  
Chondroitin Sulphate ◽  
Collagen Fibrils ◽  
Dermatan Sulphate ◽  
Keratan Sulphate ◽  
Sulphate Proteoglycan ◽  
Chondroitin Sulphate Proteoglycan

Rabbit annulus fibrosus and nucleus pulposus were analysed for hydroxyproline, chondroitin sulphate, keratan sulphate and dermatan sulphate. Tissue proteoglycans were stained for electron microscopy with Cupromeronic blue, used in the critical electrolyte concentration mode, with and without prior digestion by chondroitinase AC or ABC, hyaluronidase or keratanase. Collagen bands, a—e were demonstrated with UO2++. A chondroitin sulphate proteoglycan was found orthogonally associated with loosely packed collagen fibrils in annulus fibrosus at the d and e bands. The close metabolic and structural analogies with the dermatan sulphate proteoglycans previously shown to be located at collagen d-e bands in tendon, skin, etc. (Scott and Haigh (1985) Biosci. Rep.5:71–81), are discussed. Tightly packed annulus collagen fibrils were surrounded by axially oriented proteoglycan filaments, mostly without specific locations.

scholarly journals Identification of specific binding sites for keratan sulphate proteoglycans and chondroitin-dermatan sulphate proteoglycans on collagen fibrils in cornea by the use of cupromeronic blue in ‘critical-electrolyte-concentration’ techniques

1988 ◽  
Vol 253 (2) ◽  
pp. 607-610 ◽  
Author(s):  
J E Scott ◽  
M Haigh
Keyword(s):  
Collagen Fibril ◽  
Electrolyte Concentration ◽  
Specific Binding ◽  
Corneal Stroma ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Dermatan Sulphate ◽  
Keratan Sulphate ◽  
Specific Binding Sites ◽  
The One

Proteoglycans (PGs) in bovine corneal stroma were stained with Cupromeronic Blue in ‘critical-electrolyte-concentration’ (CEC) methods for electron microscopy, and were located vis-à-vis collagen fibril a-e banding patterns. Keratanase and chondroitin ABC lyase digestion showed that a + c-band- and d + e-band-associated PGs were keratan sulphate-rich and chondroitin (dermatan) sulphate-rich respectively. The CEC pattern proved that the keratan sulphate PGs at the a and c bands differed. Comparison of their CECs with their behaviour on anion-exchange chromatography confirmed previous (indirect) attempts at identification [Scott & Haigh (1985) Biosci. Rep. 5, 765-774]. Similar arguments were applied to the dermatan sulphate PGs at the d and e bands. These results strongly support the one-PG-one-binding-site hypothesis [e.g. Scott (1988) Biochem. J. 252, 313-323]. Remarkable inter-species variations in the keratan sulphate PG patterns contrast with the relatively constant picture of dermatan sulphate PG-collagen fibril interactions.

scholarly journals Hurler's, Hunter's and Morquio's syndromes. A biochemical study in the light of current views of the underlying defects

1971 ◽  
Vol 123 (5) ◽  
pp. 883-894 ◽  
Author(s):  
M. F. Dean ◽  
Helen Muir ◽  
R. J. F. Ewins
Keyword(s):  
Molecular Weight ◽  
Chondroitin Sulphate ◽  
Biochemical Study ◽  
Heparan Sulphate ◽  
Total Output ◽  
Dermatan Sulphate ◽  
Keratan Sulphate ◽  
Hurler’S Syndrome ◽  
Morquio's Syndrome ◽  
Hunter's Syndrome

Glycosaminoglycans were isolated from the urine of three patients with Hurler's, Hunter's and Morquio's syndromes and also from the liver and spleen of the case of Hurler's syndrome by a procedure avoiding further degradation. A method of determining the proportions of dermatan sulphate, heparan sulphate and chondroitin sulphate in each preparation is described. The relative proportions of these glycosaminoglycans in the urine and organs of the case of Hurler's syndrome were very similar. Glycosaminoglycans from the organs were of much lower molecular weight than normal, consisting of single chains of molecular weight about 5000 together with multiples of up to four such chains attached to peptide moieties. The linkage region normally attaching glycosaminoglycan chains to protein in whole protein–polysaccharides of connective tissue was degraded progressively towards serine. The total output and relative proportions of abnormal glycosaminoglycans in the urine were compared in two brothers with Hunter's syndrome examined on two occasions 4 years apart. At comparable ages they excreted about the same amount, and the relative proportions of each glycosaminoglycan remained essentially constant. The composition and chromatographic behaviour of the glycosaminoglycan in the urine from the case of Morquio's syndrome indicated that it consisted of material containing about one-third keratan sulphate and two-thirds chondroitin sulphate as part of the same molecule, as in proteoglycans of cartilage. The total output of glycosaminoglycans, although higher than normal, was considerably less than in other types of Mucopolysaccharidoses.

Healing of Circular Defects in the Rabbit Medial Meniscus Can Occur Spontaneously and Is not Improved by Intra-Articular Hyaluronic Acid

1996 ◽  
Vol 09 (02) ◽  
pp. 60-5 ◽  
Author(s):  
N. Hope ◽  
P. Ghosh ◽  
S. Collier
Keyword(s):  
Hyaluronic Acid ◽  
Medial Meniscus ◽  
Chondroitin Sulphate ◽  
Rabbit Model ◽  
Type Ii Collagen ◽  
Type Ii ◽  
Keratan Sulphate ◽  
Meniscal Healing ◽  
Meniscus Healing ◽  
Made In

SummaryThe aim of this study was to determine the effects of intra-articular hyaluronic acid on meniscal healing. Circular defects, 1.0 mm in diameter, were made in the anterior third of the medial meniscus in rabbits. In one joint, 0.4 ml hyaluronic acid (Healon®) was instilled, and in the contralateral (control) joint, 0.4 ml Ringer’s saline. Four rabbits were killed after four, eight and 12 weeks and the menisci examined histologically. By eight weeks most of the lesions had healed by filling with hyaline-like cartilage. Healing was not improved by hyaluronic acid treatment. The repair tissue stained strongly with alcian blue, and the presence of type II collagen, keratan sulphate, and chondroitin sulphate was demonstrated by immunohistochemical localisation. In contrast to the circular defects, longitudinal incisions made in the medial menisci of a further six rabbits did not show any healing after 12 weeks, indicating that the shape of the lesion largely determined the potential for healing.The effect of hyaluronic acid on meniscal healing was tested in a rabbit model. With one millimeter circular lesions in the medial meniscus, healing by filling with hyalinelike cartilage was not significantly affected by the application of hyaluronic acid intra-articularly at the time of surgery, compared to saline controls, as assessed histologically four, eight and 12 weeks after the operation.

scholarly journals Attempted isolation of a heparin proteoglycan from bovine liver capsule

1970 ◽  
Vol 116 (1) ◽  
pp. 27-34 ◽  
Author(s):  
U. Lindahl
Keyword(s):  
Potassium Chloride ◽  
Chondroitin Sulphate ◽  
Cetylpyridinium Chloride ◽  
Bovine Liver ◽  
Gel Chromatography ◽  
Dermatan Sulphate ◽  
Liver Capsule ◽  
Neutral Sugars ◽  
Degraded Material ◽  
Heparin Preparation

(1) Polysaccharides were isolated from bovine liver capsule by extraction with 2m-potassium chloride followed by precipitation from 0.8m-potassium chloride with cetylpyridinium chloride. Chondroitin sulphate was eliminated by digestion with hyaluronidase. The yield of heparin was approx. 40% of that obtained after extraction of the papain-digested tissue. (2) The macromolecular properties of the hyaluronidase-digested polysaccharide were studied by gel chromatography on Sephadex G-200 of the intact, as well as of the alkali-degraded, material. The results suggested the presence of single heparin chains in addition to a dermatan sulphate proteoglycan. (3) A purified heparin preparation was analysed for amino acids and neutral sugars. Xylose, galactose and serine were found in amounts corresponding to 0.1, 0.2, and 0.4 residue/polysaccharide chain (mol.wt. 7400), respectively. It is suggested that the isolated material had been degraded by a polysaccharidase with endo-enzyme properties.

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