Showing papers on "Keratan sulfate published in 2005"

Multipotent stem cells in human corneal stroma.

Abstract: Keratocytes of the corneal stroma secrete a specialized extracellular matrix essential for vision. These quiescent cells exhibit limited capacity for self-renewal and after cell division become fibroblastic, secreting nontransparent tissue. This study sought to identify progenitor cells for human keratocytes. Near the corneal limbus, stromal cells expressed ABCG2, a protein present in many adult stem cells. The ABCG2-expressing cell population was isolated as a side population (SP) by cell sorting after exposure to Hoechst 33342 dye. The SP cells exhibited clonal growth and continued to express ABCG2 and also PAX6, product of a homeobox gene not expressed in adult keratocytes. Cloned SP cells cultured in medium with fibroblast growth factor-2 lost ABCG2 and PAX6 expression and upregulated several molecular markers of keratocytes, including keratocan, aldehyde dehydrogenase 3A1, and keratan sulfate. Cloned corneal SP cells under chondrogenic conditions produced matrix staining with toluidine blue and expressed cartilage-specific markers: collagen II, cartilage oligomatrix protein, and aggrecan. Exposure of cloned SP cells to neurogenic culture medium upregulated mRNA and protein for glial fibrillary acidic protein, neurofilament protein, and beta-tubulin II. These results demonstrate the presence of a population of cells in the human corneal stroma expressing stem cell markers and exhibiting multipotent differentiation potential. These appear to be the first human cells identified with keratocyte progenitor potential. Further analysis of these cells will aid elucidation of molecular mechanisms of corneal development, differentiation, and wound healing. These cells may be a resource for bioengineering of corneal stroma and for cell-based therapeutics.

PAX6 expression identifies progenitor cells for corneal keratocytes

Abstract: Keratocytes of the corneal stroma produce a transparent extracellular matrix required for vision. During wound-healing and in vitro, keratocytes proliferate, becoming fibroblastic, and lose biosynthesis of unique corneal matrix components. This study sought identification of cells in the corneal stroma capable of assuming a keratocyte phenotype after extensive proliferation. About 3% of freshly isolated bovine stromal cells exhibited clonal growth. In low-mitogen media, selected clonal cultures displayed dendritic morphology and expressed high levels of keratan sulfate, aldehyde dehydrogenase 3A1, and keratocan, molecular markers of keratocyte phenotype. In protein-free media, both primary keratocytes and selected clonal cells aggregated to form attachment-independent spheroids expressing elevated levels of those marker molecules. The selected clonal cells exhibited normal karyotype and underwent replicative senescence after 65-70 population doublings; however, they continued expression of keratocyte phenotypic markers throughout their replicative life span. The progenitor cells expressed elevated mRNA for several genes characteristic of stem cells and also for genes expressed during ocular development PAX6, Six2, and Six3. PAX6 protein was detected in the cultured progenitor cells and a small number of stromal cells in intact tissue but was absent in cultured keratocytes and fibroblasts. Cytometry demonstrated PAX6 protein in 4% of freshly isolated stromal cells. These results demonstrate the presence of a previously unrecognized population of PAX6-positive cells in adult corneal stroma that maintain the potential to assume a keratocyte phenotype even after extensive replication. The presence of such progenitor cells has implications for corneal biology and for cell-based therapies targeting corneal scarring.

Soluble lumican glycoprotein purified from human amniotic membrane promotes corneal epithelial wound healing.

Abstract: Lumican, a small leucine-rich proteoglycan (SLRP), is one of the major extracellular components in interstitial collagenous matrices of the corneal stroma, aorta, skin, skeletal muscle, lung, kidney, bone, cartilage, and intervertebral discs.1–8 In the cornea, lumican contains keratan sulfate chains. However, it is present as a low or nonsulfated glycoprotein (50 –57 kDa) in noncorneal tissues.1,2,9–11 Its wide distribution implies that it has multiple functions in tissue morphogenesis and maintenance of tissue homeostasis. This is best illustrated by the multiple clinical manifestations observed in Lumican-knockout (lum−/−) mice that exhibit corneal opacity, skin and tendon fragility, delayed wound healing, and low fertility. Indeed, lumican has been shown to play essential roles in corneal transparency by regulating collagen fibrillogenesis8 in wound healing, by modulating epithelial cell migration,12 and in the epithelium–mesenchyme transition of the injured lens.13 These results have led to the speculation that lumican may play an active role in corneal epithelial wound healing. Transplantation of human AM as a temporary or permanent graft has become a popular surgical procedure for ocular surface reconstruction.14,15 Besides the observation that cryopreserved AM reduces inflammation,16–21 scarring,22,23 and neovascularization,24 many clinical studies25–28 have shown that human AM as a graft promotes corneal epithelial wound healing. Furthermore, human AM can help preserve corneal limbal epithelial progenitor cells and maintain the keratocyte phenotype during ex vivo expansion.29–32 Nevertheless, the exact molecular mechanism of the aforementioned effects remains unknown. The present study revealed that human AM is a rich source of soluble lumican glycoprotein and that purified soluble lumican glycoprotein can facilitate proliferation and migration of corneal epithelial cells during wound healing in both wild-type and lum−/− mice.

The role of glucosamine and chondroitin sulfate in treatment for and prevention of osteoarthritis in animals.

Abstract: J disease, in particular osteoarthritis, is an important cause of lameness and debilitation for humans and other animals. Presently, a number of pharmacologic agents are available for treatment for osteoarthritis, including nutraceuticals containing glucosamine and chondroitin sulfate. Although to date clinical trials in veterinary patients are limited, trials conducted in humans have for the most part provided encouraging results. Results of in vitro studies suggest that these compounds may impede the progression of joint degeneration in osteoarthritis. The purpose of this review is to provide veterinary practitioners with up-to-date information regarding the mechanism of action, pharmacokinetics, clinical efficacy, and safety of glucosamine and chondroitin sulfate. Structure of Articular Cartilage and Pathophysiology of Osteoarthritis Chondrocytes, the cellular component of articular cartilage, are responsible for the synthesis and maintenance of the extracellular matrix in which they are embedded. The extracellular matrix is composed of collagen (predominantly type II collagen) and proteoglycans, with a smaller percentage of glycoproteins (Figure 1). Type II collagen arranged in fibrils is responsible for the tensile strength of articular cartilage. Proteoglycans consist of a central protein core to which 1 or more glycosaminoglycan (GAG) side chains are attached. In turn, GAGs are composed of repeating disaccharide units of hexosamine (glucosamine or galactosamine) alternating with another residue of glucuronate, iduronate, or galactose. The largest and most predominant proteoglycan in cartilage is aggrecan. Chondrocytes synthesize aggrecan by covalently attaching GAGs (chondroitin sulfate and keratan sulfate) to a central protein core of proteoglycans in an organized manner. This core protein of the proteoglyThe role of glucosamine and chondroitin sulfate in treatment for and prevention of osteoarthritis in animals

Detection and quantification of sulfated disaccharides from keratan sulfate and chondroitin/dermatan sulfate during chick corneal development by ESI-MS/MS.

Abstract: PURPOSE To identify and quantify changes in keratan sulfate (KS) and chondroitin/dermatan sulfate (CS/DS) sulfated disaccharides in the developing chick cornea using electrospray ionization tandem mass spectrometry (ESI-MS/MS). METHODS Cryostat sections of fresh nonfixed corneas were obtained from White Leghorn embryonic day (E)8 to E20 chicks, and from 4- and 70-week-old chickens. Tissue sections on glass slides were incubated with selected glycosidase enzymes. Digest solutions were analyzed directly by ESI-MS/MS. RESULTS The concentration of KS monosulfated disaccharide (MSD) Gal-beta-1,4-GlcNAc(6S) in E8 cornea equaled that at E20, declined to its lowest level by E10, increased to a second peak by E14, decreased to a second low by E18, peaked again by E20, and remained high in adult corneas. A similar concentration profile was observed for KS disulfated disaccharide (DSD) Gal(6S)-beta-1,4-GlcNAc(6S), and thus also for total sulfated KS disaccharides. The molar percent of DSD was higher than that of MSD from E8 to E18, equivalent at E20, and less than that of MSD in adult corneas. In contrast, total concentration of CS/DS Deltadi-4S plus Deltadi-6S decreases as development progresses and is lowest in adult corneas. Concentration and molar percent of Deltadi-6S is highest at E8, then decreases through development as the concentration and molar percent of Deltadi-4S increases from E8 and exceeds that of Deltadi-6S after E14. CONCLUSIONS New, rapid, direct chemical analysis of extracellular matrix components obtained from sections from embryonic and adult chick corneas reveals heretofore undetected changes in sulfation characteristics of KS and CS/DS disaccharides during corneal development.

Analysis of keratan sulfate oligosaccharides by electrospray ionization tandem mass spectrometry.

Abstract: Keratan sulfate (KS) is a glycosaminoglycan consisting of repeating disaccharide units composed of alternating residues of d-galactose and N-acetyl-d-glucosamine linked β-(1−4) and β-(1−3), respectively. In this study, electrospray ionization tandem mass spectrometry (ESI-MS/MS) was employed to identify keratan sulfate oligosaccharides. Two nonsulfated disaccharide isomers and two monosulfated disaccharide isomers were distinguished through MS/MS. In MS1 spectra of multiply sulfated KS oligosaccharides, the charge state of the most abundant molecular ion equals the number of sulfates. Subsequent MS2 and MS3 spectra of mono-, di-, tri-, and tetrasulfated KS oligosaccharides and sialylated tetrasaccharides reveal diagnostic ions that can be used as fingerprint maps to identify unknown KS oligosaccharides. Based on the pattern of fragment ions, the compositions of an oligosaccharide mixture from shark cartilage KS and of two enzyme digests of bovine corneal KS were determined directly, without prior isolatio...

Atypical composition and ultrastructure of proteoglycans in the mouse corneal stroma.

Abstract: PURPOSE: Recently, gene-targeted strains of mice with null mutations for specific proteoglycans (PGs) have been used for investigations of the functional role of these molecules. In the present study, the corneal stroma of the mouse was examined to provide some baseline PG morphologies in this species. METHODS: Monoclonal antibodies to specific glycosaminoglycan (GAG) chain sulfation patterns were used to characterize PG composition in corneal extracts by SDS-PAGE and Western blot analysis and to identify their tissue distribution by immunofluorescence microscopy. PGs were also visualized by transmission electron microscopy after contrast enhancement with cationic dye fixation. RESULTS: Western blot analysis of pooled corneal extracts and immunofluorescence of tissue sections identified 4-sulfated, but not 6-sulfated, chondroitin sulfate/dermatan sulfate (CS/DS). Keratan sulfate (KS) was present only as a low-sulfated moiety. Electron microscopic histochemistry disclosed a complex array of corneal PGs present as (1) fine filaments radiating from collagen fibrils, and (2) elongate, straplike structures, running either along the fibril axis or weaving across the primary fibril orientation. These large structures were digested by chondroitinase ABC, but not by keratanase. CONCLUSIONS: KS in the mouse is predominantly undersulfated and generates an immunostaining pattern that differs from that observed in corneas of other mammalian species thus far investigated. The mouse cornea resembles other mammalian corneas in the presence of filamentous arrays of small, collagen-associated stromal PGs visualized by cationic dye staining. However, large dye-positive structures with a CS/DS component are also present and appear to be unique to the cornea of this species.

An x-ray diffraction study of corneal structure in mimecan-deficient mice.

Abstract: PURPOSE: Keratan sulfate proteoglycans (KSPGs) in the corneal stroma are believed to influence collagen fibrillar arrangement. This study was performed to investigate the fibrillar architecture of the corneal stroma in mice homozygous for a null mutation in the corneal KSPG, mimecan. METHODS: Wild-type (n = 9) and mimecan-deficient (n = 10) mouse corneas were investigated by low-angle synchrotron x-ray diffraction to establish the average collagen fibrillar spacing, average collagen fibril diameter, and level of fibrillar organization in the stromal array. RESULTS: The mean collagen fibril diameter in the corneas of mimecan-null mice, as an average throughout the whole thickness of the tissue, was not appreciably different from normal (35.6 +/- 1.1 nm vs. 35.9 +/- 1.0 nm). Average center-to-center collagen fibrillar spacing in the mutant corneas measured 52.6 +/- 2.6 nm, similar to the 53.3 +/- 4.0 nm found in wild-type mice. The degree of local order in the collagen fibrillar array, as indicated by the height-width (H:W) ratio of the background-subtracted interfibrillar x-ray reflection, was also not significantly changed in mimecan-null corneas (23.4 +/- 5.6), when compared with the corneas of wild-types (28.2 +/- 4.8). CONCLUSIONS: On average, throughout the whole depth of the corneal stroma, collagen fibrils in mimecan-null mice, unlike collagen fibrils in lumican-null mice and keratocan-null mice, are of a normal diameter and are normally spaced and arranged. This indicates that, compared with lumican and keratocan, mimecan has a lesser role in the control of stromal architecture in mouse cornea.

Comparison of proteoglycan and collagen in articular cartilage of horses with naturally developing osteochondrosis and healing osteochondral fragments of experimentally induced fractures

Abstract: Objective—To compare articular cartilage from horses with naturally developing osteochondrosis (OC) with normal articular cartilage and healing cartilage obtained from horses with experimentally induced osteochondral fractures. Sample Population—109 specimens of articular cartilage from 78 horses. Procedure—Morphologic characteristics, proteoglycan (PG), and type II collagen were analyzed in articular cartilage of OC specimens (group 1), matched healing cartilage obtained 40 days after experimentally induced osteochondral fractures (group 2), and matched normal cartilage from the same sites (group 3). Results—79 specimens of OC cartilage were obtained from horses. Ex vivo PG synthesis was significantly greater in the femoral cartilage, compared with synthesis in the tibial cartilage, and significantly greater for groups 1 and 2, compared with group 3. For groups 1 and 2, femoral fragments had significantly greater PG content, compared with PG content in tibial fragments. Keratan sulfate content was significantly less in group 3, compared with groups 1 and 2. Cartilage from the OC specimens had loss of structural architecture. The OC tissue bed stained positive for chondroitin sulfate and type II collagen, but the fracture bed did not. Conclusions and Clinical Relevance—Our analyses could not distinguish articular cartilage from horses with OC and a healing fracture. Both resembled an anabolic, reparative process. Immunohistochemical analysis suggested a chondromyxoid tissue in the OC bed that was morphologically similar to fibrous tissue but phenotypically resembled hyaline cartilage. Thus, tissue in the OC bed may be degenerative cartilage, whereas tissue in the fracture bed may be reparative fibrous callus. (Am J Vet Res 2005;66:1881–1890)

Chemistry, Biochemistry, and Pharmaceutical Potentials of Glycosaminoglycans and Related Saccharides

Abstract: A review discusses various aspects of glycosaminoglycans (GAGs) and related saccharides, including their biol. activity, chem., biochem. and pharmaceutical potentials.

Inhibition of antithrombin by hyaluronic acid may be involved in the pathogenesis of rheumatoid arthritis

Abstract: Thrombin is a key factor in the stimulation of fibrin deposition, angiogenesis, proinflammatory processes, and proliferation of fibroblast-like cells. Abnormalities in these processes are primary features of rheumatoid arthritis (RA) in synovial tissues. Tissue destruction in joints causes the accumulation of large quantities of free hyaluronic acid (HA) in RA synovial fluid. The present study was conducted to investigate the effects of HA and several other glycosaminoglycans on antithrombin, a plasma inhibitor of thrombin. Various glycosaminoglycans, including HA, chondroitin sulfate, keratan sulfate, heparin, and heparan, were incubated with human antithrombin III in vitro. The residual activity of antithrombin was determined using a thrombin-specific chromogenic assay. HA concentrations ranging from 250 to 1000 μg/ml significantly blocked the ability of antithrombin to inhibit thrombin in the presence of Ca2+ or Fe3+, and chondroitin A, B and C also reduced this ability under the same conditions but to a lesser extent. Our study suggests that the high concentration of free HA in RA synovium may block antithrombin locally, thereby deregulating thrombin activity to drive the pathogenic process of RA under physiological conditions. The study also helps to explain why RA occurs and develops in joint tissue, because the inflamed RA synovium is uniquely rich in free HA along with extracellular matrix degeneration. Our findings are consistent with those of others regarding increased coagulation activity in RA synovium.

Characterization of the non-collagenous proteins in avian cortical and medullary bone

Abstract: Northern blotting, RT-PCR, and Western blotting techniques were used to characterize the matrix constituents of avian cortical and medullary bone. Extracts of bone tissue were found to contain multiple isoforms of bone sialoprotein (BSP), osteopontin (OPN), osteonectin (ON), osteocalcin (OC), and dentin matrix protein-1 (DMP-1). Only single transcripts were observed with Northern blotting; therefore it was concluded that the isoforms were due to differences in post-translational modifications. Since medullary bone is rich in keratan sulfate (KS), RT-PCR was used to investigate the expression of known keratan sulfate-containing proteoglycans (KSPGs). Although this tissue was found to express lumican and osteoglycin/mimecan, there was little evidence to suggest that these proteoglycans were a major source of the keratan sulfate glycosaminoglycans. Treatment of medullary bone extracts with keratanase resulted in the appearance of a BSP immunoactive band of approximately 59 kDa. However, it was not possible to isolate and identify the intact keratan sulfate proteoglycan.

Expression of a chondroitin sulfate proteoglycan, versican (PG-M), during development of rat cornea.

Abstract: Purpose: To understand the role of chondroitin sulfate proteoglycans during the development of rat cornea, expression of chondroitin sulfate and versican (PG-M) was studied. Methods: Chondroitin sulfate and keratan sulfate in rat cornea were analyzed by immunohistochemical techniques. Reverse transcription polymerase chain reaction (RT-PCR) for chondroitin sulfate proteoglycans was performed. Versican expression was studied by RT-PCR, immunohistochemical, and dot blot analyses. Expression of hyaluronan was evaluated histochemically using biotinylated hyaluronan binding protein. Results: Chondroitin sulfate was abundant in rat cornea at postnatal day 1 (P1) and became undetectable at P14. RT-PCR analysis showed that versican mRNA was highly expressed at P1 but was little expressed at P42. mRNAs for other chondroitin sulfate proteoglycans including biglycan, aggrecan, and decorin did not change much between P1 and P42. Expression for all versican splicing isoforms (V0–V3) was detectable from P1 through P14 ...

Ultrastructural immunolocalization of [alpha]-elastin and keratan sulfate proteoglycan in normal and scoliotic lumbar disc

Abstract: Study Design. Comparative ultrastructural study of intervertebral discs from normal subjects and patients with scoliosis. Objective. To identify ultrastructural relations among keratan sulfate (KS) proteoglycan, [alpha]-elastin, collagen fibers, and elastic fibers in normal and scoliotic discs. Summary of Background. KS proteoglycans, elastic fibers, and collagen fibers play important mechanical roles in the intervertebral disc, but the distributions of KS proteoglycans and elastin in this tissue have received little attention. Methods. Tissues were fixed in 4% paraformaldehyde. Monoclonal antibody 5-D-4 (which recognizes a KS epitope on aggrecan, fibromodulin, and lumican) and polyclonal anti-[alpha]-elastin were visualized with a 10-nm immunogold-conjugated secondary antibody. Results. In a normal disc, a regular pattern of KS labeling occurred around collagen fibers, in the cell cytoplasm, and in the rough endoplasmic reticulum; the nucleus pulposus was more densely labeled for KS than was the anulus fibrosus. In scoliotic disc anulus fibrosus, KS labeling was weak throughout the matrix and pericellularly but abundant in lysosomes and on electron-dense material in degenerate cells. Degenerate collagen fibers in scoliotic tissue bore less KS than did normal fibers. KS labeling of the microfibrillar region of elastic fibers was strong in normal disc but weak in scoliotic disc. Elastin labeling of elastic fibers was weaker in scoliotic than in normal tissue. Conclusion. KS proteoglycans and elastic fibers are closely associated with the lamellar organization of the collagen fibers in a normal disc. In scoliosis, impaired regulation of collagen fibrillogenesis by lumican or fibromodulin may result in disruption of the lamellar structure.

Ultrastructural and immunohistochemical studies of medullary bone calcification, with special reference to sulphated glycosaminoglycans

Abstract: Histochemical, immunohistochemical and electron energy-loss spectroscopic studies were performed to examine the relationship between sulphated glycosaminoglycans and medullary bone calcification using oestrogen-injected male Japanese quail. Sulphated glycosaminoglycans, detected by high iron diamine (HID) or HID-thiocarbohydrazide-silver protein (HID-TCH-SP) methods, were distributed throughout the matrix of medullary bone, some periphery and extending tips of the trabeculae stained weakly, and the globular structures at osteoid areas were exclusively positive for HID-TCH-SP stain. Immunohistochemistry identified keratan sulphate located in the globular structures at osteoid areas and calcified matrix, but chondroitin-4 sulphate and chondroitin-6 sulphate were not detected in the matrix. Using electron spectroscopic imaging, sulphur was determined to be localized in the globular structures. These results demonstrate that medullary bone matrix accumulates keratan sulphate in the globular structures, which are the foci for calcification, and eventually in the calcified areas. This suggests that keratan sulphate containing sulphur is maintained in the calcified matrix. These results indicate a unique process of calcification exists in medullary bone.

The role of proteoglycans in idiopathic carpal tunnel syndrome.

Abstract: Decompression of the carpal canal is the most common hand surgery performed in the United States. Hand surgeons perform 460,000 carpal tunnel releases (CTR) each year, which cost the medical industry in excess of two billion dollars per year. The focus of this investigation was to identify the changes, which occur in the flexor tenosynovium of patients undergoing CTR at the connective tissue level. The connective tissues determine the amount and arrangement of macromolecules (fibers, proteoglycans, and glycoproteins) in the extracellular matrix. The proteoglycans are soluble macromolecules that have both structural and metabolic roles. Glycoproteins help to form the interstitial space, basement membrane and function as cell surface receptors. The mechanical function of the proteoglycans includes stabilization of the collagen fibers as well as function in the hydration of the tissues. It has been previous shown that changes in the oxygen concentration at the tissue level can alter the proteoglycans profile of the tissue. During periods of hypoxia, such as those obtained during repetitive motion CTS; the glycolytic pathway acts as the energy source for the tissue. Productions of chondroitin sulfates are a process consumes NAD and would be potentially toxic to the cells under anaerobic conditions. Production of keratan sulfate is NAD sparing product, and may act as a survival pathway for cells under adverse conditions. The disruption in the proteoglycan balance will allow for alterations in the ECM and changes in hydration status of the tissues may have serious implication in CTS because the carpal canal is anatomically very narrow and increases in volume within the canal can result in further compression of the nerve. Flexor tenosyioum was obtained from patients undergoing CTR and compared with control tissue for dermatan, keratan and chondroitin sulfate. The results show a greater density of keratan reactivity in CTS tissues identified by immunostaining. In addition to changes in proteoglycan content there was also an increase in new vessel formation in the CTS tissues. The data obtained suggests that the shifts in the proteoglycan ratios may render the tissues less able to withstand the compressive forces and therefore allow for more force to be placed on the median nerve within the carpal canal.

Immunohistochemical characterization of noncollagenous matrix molecules on the alveolar bone surface at the initial principal fiber attachment in rat molars.

Abstract: This study was designed to immunodetect proteoglycans (PGs) and the noncollagenous glycoproteins, bone sialoprotein (BSP) and osteopontin (OPN) on developing alveolar bone surface in rat molars by the indirect immunoperoxidase method, and to discuss the roles of these molecules at the initial principal fiber (PF) attachment. To characterize PGs, antibodies against five species of glycosaminoglycans (GAGs), chondroitin-4-sulfate (C4S), chondroitin-6-sulfate (C6S), unsulfated chondroitin (C0S), dermatan sulfate (DS), and keratan sulfate (KS) were used. Maxillary alveolar bone facing the distal root of the second molar was examined in 20- and 25-day-old male Wistar rats. Routine histological staining was also used. A hematoxylin-stained, fibril-poor layer always appeared on the alveolar bone surface just prior to the initial PF organization. This layer was strongly immunoreactive for C4S, C0S, OPN, and BSP, and weakly for C6S, but not for DS and KS. Then the initial PFs were attached to this layer. When new bone containing Sharpey's fibers covered this layer, it remained as a hematoxylin-stained, fibril-poor layer between Sharpey's fiber-containing and -lacking bone. The layer was consistently immunoreactive for OPN and BSP but had no immunoreactivity for GAGs. The results suggest that the accumulation of C4S-, C0S-, and C6S-carrying PGs, and of BSP and OPN is a primary event at the initial PF attachment, and is involved in the adhesion of PFs and mineralization of the initial attachment layer. The BSP and OPN act to maintain the interface integrity between Sharpey's fiber-containing and Sharpey's fiber-lacking alveolar bone after the PF attachment is established.

Role of Heparan Sulfate in Cancer

Abstract: Publisher Summary Glycosaminoglycans (GAGs) include heparin (HP), heparan sulfate (HS), dermatan sulfate (DS), chondroitin sulfate (CS), keratan sulfate (KS), and hyaluronic acid (HA). These are the polymers of a disaccharide repeat unit, comprising a uronic acid and a hexosamine. Chondroitin and DS arise from a single homogeneous precursor, chondroitin, with the disaccharide repeat unit. KS is the only GAG that does not contain uronic acid; therefore, it is not cleavable by the eliminative enzymatic cleavage. Almost all glucosamines and some of galactose units are 6-sulfated in KS. Proteoglycans are heavily glycosylated proteins, generally consisting of a core protein with one or more covalently linked GAG chains that constitute most of the proteoglycan. The storage function of HS-like GAGs (HSGAGs) is exemplified by the HP found in the granules of mast cells, where these highly sulfated polysaccharides bind to specific granule proteases and store them in an inactive form.

Remodeling of Heparan Sulfate Sulfation by Extracellular Endosulfatases

Abstract: Publisher Summary This chapter reviews current knowledge of the 6-O-HS endosulfatase (Sulf) enzymes, including their unique structural and enzymatic properties as novel extracellular heparan endosulfatases, their regulatory functions in extracellular HS-dependent Wnt and fibroblast growth factors (FGF) signaling in stem cell progenitors, and their roles as well as therapeutic applications in the control of tumor growth and angiogenesis. Conservation of the hydrophilic domain (HD) sequence in association with catalytic and C-terminal domains has enabled identification of Sulf proteins in vertebrates and invertebrates. Sulf enzymes are functionally distinct from other sulfatases, including GlcNR6Sase, in their restricted expression in stem cells, their localization on the cell surface, and their substrate specificity and activity as heparan sulfate (HS)-specific 6-O-endosulfatases. The cell surface localization of Sulf on expressing cells suggests its cell autonomous function. Sulfs were predicted to act on GlcN 6-O-sulfate residues in HS based on their sequence homology with lysosomal GlcNR6Sase, an exoenzyme that removes 6-O-sulfate groups from the nonreducing-terminal GlcN residue of HS, heparin (HP), and keratan sulfate chains.

Glycosaminoglycans of the vertebral body growth plate in patients with idiopathic scoliosis.

Abstract: The concentrations of keratan sulfates and unmodified keratan sulfates increased in the vertebral body growth plate in patients with idiopathic scoliosis. Sulfation and acetylation of total glycosaminoglycans decreased by 50 and 30%, respectively. These changes reflect the decrease in biological activity of molecules that modulate function of the growth plate.

Spatial distribution of keratan sulfate in the rabbit cornea following photorefractive keratectomy.

Abstract: PURPOSE: To examine the keratan sulfate content of the stroma and to assess its correlation with the healing process (epithelialization and keratocyte density) after photorefractive keratectorny (PRK). METHODS: Using an Aesculap Meditec MEL 70 G-scan excimer laser, -6.0 diopters (6.0-mm diameter, 82 μm photoablation depth), PRK was carried out on the right eye of 32 New Zealand pigmented rabbits. After enucleation (at days 1, 4, 7, 14, and 28 and months 2, 3, and 7; subl-groups of 4 animals), fluorescent immunohistochemistry was performed on sections from the central cornea using monoclonal mouse anti-keratan-sulfate antibody, immunohistochemistry with proliferative cell nuclear antigen antibody, and hematoxylin-eosin histology. The left, untreated eyes served as controls. Cellular morphology and spatial distribution of keratan sulfate were recorded, stromal thickness measured, and keratocyte density calculated. RESULTS: Keratan sulfate was found on the surface of migrating epithelial cells in the early stage (from days 1 to 7). In the stroma, three phases were noted. Phase 1) Day 1 to 14, intense granular fluorescence appeared in the anterior stroma with hypocellularity. Phase 2) Month 1 to 2, newly synthesized lamellar keratan sulfate restored the repopulating anterior stroma. Endothelial cells became keratan sulfate positive, while in the posterior stroma, lamellar-form keratan sulfate increased from week 1 and peaked at months 1 (100% increase). Phase 3) Month 2 to 7, remodeling and depositior of keratan sulfate was noted, which was produced in phase 2. CONCLUSIONS: Keratan sulfate was found in the epithelium, stroma, and endothelium. By controlling the interlamellar spacing, keratan sulfate plays a role in post-operative edema, remodeling of the corneal stroma, and simultaneous regulation of inflammation after PRK.