Presence Of Keratan Sulfate Research Articles

Morquio disease with CNS involvement: a rare association

Morquio syndrome is a rare type of mucopolysaccharidosis. Our patient presented with uncontrolled seizures and gross skeletal deformity. He was suspected to be suffering from mucopolysaccharidosis based on his disease presentation but the diagnosis could be made as Morquio syndrome on the basis of the presence of keratan sulphate in the urine. DOI: http://dx.doi.org/10.3126/jnps.v34i2.9298 J Nepal Paediatr Soc 2014;34(2):157-159

Regulated expression of keratan sulphate and peanut agglutinin binding sites during organogenesis in the developing chick.

Keratan sulphate proteoglycans are potentially important during development and are possible binding molecules for the lectin, peanut agglutinin, a marker for areas that are inhibitory for axonal growth in early embryos. The present study describes the spatiotemporal distributions of keratan sulphate epitopes and peanut agglutinin binding sites during organogenesis in the developing chick from E5 to hatching. The widespread distributions of these molecules did not often overlap but clearly delimited different carbohydrate compartments demonstrating that peanut agglutinin does not necessarily bind to keratan sulphate proteoglycans. These markers were mostly extracellular but keratan sulphate, in particular, was found within certain specific cells in cartilage, gonad, heart and pancreas, at certain ages. The presence of keratan sulphate in putative germ cells during their migrations and in the gonads may be of particular importance. Their distributions generally evoke modulation of adhesion allowing cell migrations or morphogenetic movements related to epitheliomesenchymal interactions, but may also suggest an involvement in axonal guidance in skin, cartilage, gut and possibly heart. Furthermore, in the kidney, peanut agglutinin binding sites seem to be related to the functional differentiation of the nephrons.

Cloning and chromosomal localization of mouse keratocan, a corneal keratan sulfate proteoglycan.

The corneal stroma consists of a highly structured extracellularmatrix that is maintained by specialized fibroblasts called kerato-cytes. The transparency of the cornea is dependent on the presenceof highly sulfated proteoglycans (PG) that can bind collagen andcan regulate collagen fibril diameter (Rada et al. 1993). The cor-neal proteoglycans that have been identified contain numerousLeu-rich repeats and all belong to the same gene family, the Leu-rich proteoglycans (LRP). These include a chondroitin/dermatansulfate proteoglycan (CS/DSPG), decorin, and three keratan sul-fate proteoglycans (KSPG), lumican, keratocan, and osteoglycin(Funderburgh et al. 1991, 1993; Blochberger et al. 1992; Corpuz etal. 1996). The corneal KSPGs are glycoproteins exclusively duringearly corneal development before corneal transparency (Cornuet etal. 1994) and in macular corneal dystrophy type I, an inheriteddisease that is characterized by corneal opacity and deposits(Klintworth et al. 1977, 1983; Nakazawa et al. 1984). These pre-vious studies show a direct correlation between corneal transpar-ency and the presence of keratan sulfate. The core protein andpoly-lactosamine side chains alone appear to be insufficient extra-cellular matrix components for sustaining a transparent structure inthe cornea.The primary structure of keratocan, based on the deducedamino acid sequence from bovine and chick cDNA sequences,shows N- and C-terminal globular domains containing four andtwo Cys residues forming two and one disulfide bond, respec-tively, and 11 highly conserved Leu-rich repeats located mainlybetween the two globular domains (Corpuz et al. 1996; Dunlevy etal. 1998). The two-dimensional model of chick keratocan, basedon the X-ray crystallography structure of ribonuclease inhibitor, amolecule comprised entirely of Leu-rich repeats, (Kobe et al. 1993,1995), shows the Leu-rich repeats coiled in a spiral with theseregions in close proximity to each other, forming a horseshoestructure (Dunlevy et al. 1998). This model also shows two out ofthree KS chains extended outward from the outer surface of thehorseshoe with the inner surface more likely to be involved incollagen binding.On the basis of mRNA levels, keratocan is most abundant incornea and sclera, but is also found at substantially lower levels inligament, artery, cartilage, skeletal muscle, and skin (Corpuz et al.1996), and keratocan is more specifically expressed in cornea thanis lumican. Consequentially, we are interested in gaining furtherinsights into its tissue-specific regulation of expression, its role indevelopment and maintenance of corneal transparency. In the pre-sent study, we report the cDNA sequence of mouse keratocan andlocalization of the gene to distal Chromosome (Chr) 10 by segre-gation analysis of restriction fragment length variants (RFLVs) inrecombinant inbred (RI) strains of mice.Total RNA was isolated from mouse whole eyes by use of aguanidinium thiocyanate-phenol-chloroform procedure (Chomc-zynski and Sacchi 1987). An mRNA purification kit (PharmaciaBiotech) was used to further purify the RNA, and ∼10 mgofmRNA was used to prepare a cDNA library (Stratagene CustomLibrary Department) in the Uni-Zap XR vector system. The librarywas screened with a high-specific-activity

Cross-reactivity of anti-sulfatide antibody with sulfated glycosaminoglycans and DNA in sera from patients with autoimmune hepatitis

Abstract The cross-reactivity of anti-sulfatide antibody in sera from patients with autoimmune hepatitis was studied. The antibody activity, determined by enzyme-linked immunosorbent assay (ELISA), was reduced in the presence of heparin, heparan sulfate, fucoidan, and dextran sulfate, but not in the presence of keratan sulfate, dermatan sulfate and chondroitin sulfate. The reactivity with sulfatide of serum IgG was bound to a heparin-Sepharose column and a double-stranded DNA-cellulose column, and recovered in the fractions eluted with 1.5 M NaCl. Incubation of patients' sera with heparin and calf thymus DNA reduced the reactivity with sulfatide in 8 and 9, respectively, of 11 patients examined. These findings suggested that anti-sulfatide aantibody was cross-reactive with heparan sulfate, especially heparin, and calf thymus DNA, and that this antibody recognizes certain structures containing repetitive, negatively charged groups as functional epitopes.

Immunohistochemical localization of articular cartilage proteoglycan and link protein in situ using monoclonal antibodies and lectin-binding methods.

Lectins have specificity for certain carbohydrate structures in macromolecules. Lectins are, therefore, useful histochemical tools for demonstrating the composition and localization of components of connective tissue matrices, such as articular cartilage. In order to assess the significance of observed lectin-binding patterns, experiments were performed in which monoclonal antibodies against chondroitin sulphate- and keratan sulphate-containing proteoglycans and link proteins were applied to sections of bovine articular cartilage after enzymatic digestion with chondroitinase ABC and keratanase. The following conclusions were made: (1) Binding of peanut agglutinin (PNA) in the interterritorial matrix predominantly indicates the presence of keratan sulphate, but may also detect O-linked oligosaccharides of proteoglycans. (2) In normal cartilage wheat germ agglutinin (WGA) binds nearly exclusively to keratan sulphate. In cartilage degraded with chondroitinase ABC and keratanase this lectin may also detect carbohydrates in link protein due to enhanced accessibility. Binding of WGA to O-linked oligosaccharides may eventually occur. (3) In enzymatically digested cartilage matrix, staining with soybean agglutinin (SBA) may be due to link protein, but not to chondroitin sulphate, because specific breakdown of the glycosaminoglycan chain is required for binding of SBA. (4) Ulex europaeus agglutinin I (UEA I) binding sites are only detectable in digested cartilage matrix.

Nature and distribution of mineral-binding, keratan sulfate-containing glycoconjugates in rat and rabbit bone.

The presence of keratan sulfate (KS) and KS proteoglycans in bone has been demonstrated in birds and rabbits but comparison with other animal species has not been investigated. The nature and distribution of mineral-binding, KS-containing glycoconjugates in rat and rabbit bone were investigated with a monoclonal antibody (MAb 5D4) specific for KS. Mineral-binding proteins were extracted from the mineralized bone with 0.4 M EDTA without guanidine-HCl (E-extract). On Western blot analysis of SDS-polyacrylamide gel electrophoresis, rat E-extract gave a weak 5D4-reactive band, M(r) 66,000-68,000, whereas rabbit E-extract produced two major reactive populations of small and large molecular size; one population consisted of two closely spaced bands at M(r) 61,000-63,000 and 66,000-68,000, and the other population consisted of one band at approximately M(r) 200,000. The identity of KS chains was further established by the sensitivity of these bands to keratanase II (Bacillus sp. Ks 36) and endo-beta-galactosidase. Immunocytochemistry with MAb 5D4 showed that, in rat bone, staining associated with the mineral phase was limited to the walls of osteocytic lacunae and bone canaliculi, whereas the remainder of the mineralized matrix lacked staining. In contrast, in rabbit bone the staining was distributed over the entire portion of the mineralized matrix with focal accumulation of staining in the wall of the lacunocanalicular system. These results indicate that rat bone contains a mineral-binding, KS-containing glycoconjugate with preferential localization in the wall of the lacunocanalicular system, whereas rabbit bone contains at least two or possibly three types of KS-containing glycoconjugates distributed over the entire portion of the mineralized matrix.

Influence of Constituents of Proteoglycans on Type II Collagen Fibrillogenesis

Turbimetry was used to study the influence of glycosaminoglycans of cartilage proteoglycans on type II collagen fibrillogenesis. The monosaccharides, D-glucuronate, N-acetyl-D-galactosamine, N-acetyl-D-glucosamine and D-galactose, all decreased the rate of the fibril formation. D-glucuronate had the strongest effect. The influence of chondroitin sulphate on type II collagen fibrillogenesis depended on the pH of the final solution, the length of the chondroitin sulphate chains and the concentration of chondroitin sulphate. At pH = 7.3 all chondroitin sulphate preparations decreased the rate of fibrillogenesis, while at pH = 6.9 and lower fibrillogenesis was stimulated by chondroitin sulphate (whale/shark), chondroitin 4-sulphate (whale) and chondroitin 6-sulphate (shark). The chain length of these three appeared to be longer than the chain length of chondroitin sulphate (human) and chondroitin sulphate oligosaccharides (whale/shark). At high concentrations (more than 3 mg/ml) fibril formation was less strongly retarded by keratan sulphate (human) than by chondroitin sulphate. At low concentrations a slight stimulation was observed in the presence of keratan sulphate. Glycosaminoglycans did not bind to collagen fibrils. At 0.5 mg/ml chondroitin 4-sulphate had a large solubilizing effect on fibrils compared to chondroitin 6-sulphate. Fibrillogenesis of type II collagen is in many aspects not comparable with fibrillogenesis of type I collagen.

Topographical distribution of sulfated glycosaminoglycans in the surface layers of the human temporomandibular joint. A histochemical study of an autopsy material.

The right temporomandibular joint (TMJ) was removed at autopsy from 18 individuals and the articular surfaces were scored for macroscopic lesions. Soft tissue specimens were cut out of the medial, lateral and posterior parts of the temporal and condylar components and frozen. The sections were stained with toluidine blue (1-N HCl solution) and with alcian blue (CEC method) for sulfated glycosaminoglycans. The sections were examined for inter- and pericellular metachromasia and alcian blue staining. Macroscopic surface lesions were found in 14 of the joints. The highest score of interand pericellular metachromasia was found in the anterior parts of the joints and a negative correlation was found between the score of surface lesions and that of intercellular metachromasia. Pericellular metachromasia was found mainly in the mineralized cartilage and cartilage layers of the articular surface. Although the CEC values of pericellular staining commonly reached a level indicating the presence of keratan sulfate, the majority of the CEC values found corresponded to those of chondroitin-dermatan sulfate. It was concluded that sulfated GAG are localized mainly to the anterior part of the TMJ and that macroscopic TMJ surface lesions are associated with a reduction of sulfated glycosaminoglycans.