Localization Of Glycosaminoglycans Research Articles

Glycosaminoglycans exhibit distinct interactions and signaling with BMP2 according to their nature and localization

The role of glycosaminoglycans (GAGs) in modulating bone morphogenetic protein (BMP) signaling represents a recent and underexplored area. Conflicting reports suggest a dual effect: some indicate a positive influence, while others demonstrate a negative impact. This duality suggests that the localization of GAGs (either at the cell surface or within the extracellular matrix) or the specific type of GAG may dictate their signaling role. The precise sulfation patterns of heparan sulfate (HS) responsible for BMP2 binding remain elusive. BMP2 exhibits a preference for binding to HS over other GAGs. Using well-characterized biomaterials mimicking the extracellular matrix, our research reveals that HS promotes BMP2 signaling in the extracellular space, contrary to chondroitin sulfate (CS), which enhances BMP2 bioactivity at the cell surface. Further observations indicate that a central IdoA (2S)-GlcNS (6S) tri-sulfated motif within HS hexasaccharides enhances binding. Nevertheless, BMP2 exhibits a degree of adaptability to various HS sulfation types and sequences. Molecular dynamic simulations attribute this adaptability to the BMP2 N-terminal end flexibility. Our findings illustrate the complex interplay between GAGs and BMP signaling, highlighting the importance of localization and specific sulfation patterns. This understanding has implications for the development of biomaterials with tailored properties for therapeutic applications targeting BMP signaling pathways.

Distributional characteristics of sulfated glycosaminoglycans in normal human nasal mucosa and nasal polyp

ConclusionThe results of this study suggest that an altered expression pattern of sulfated glycosaminoglycans (GAGs) in nasal polyp may not function as a charge restriction barrier for vascular permeability, contributing to the leakage of protein and fluid.ObjectiveSulfated GAGs are found on the vascular endothelial surface and in the extracellular matrix in various tissues and organs, suggesting that these materials constitute a negatively charged screen restricting the movement of circulating plasma molecules. This study was designed to elucidate the distributional characteristics of sulfated GAGs in normal human nasal mucosa and nasal polyp in order to understand their roles in the formation of nasal polyp.Material and methodsThe expression and localization of sulfated GAGs in normal human nasal mucosa and nasal polyp were detected light microscopically with the histochemical method using poly-L-lysine-conjugated colloidal gold followed by silver enhancement.ResultsSulfated GAGs in normal human nasal mucosa were distributed in the epithelial layer, vascular endothelial surface, submucosal gland and extracellular matrix. In nasal polyp, intense staining was also seen in the glandular structure and epithelial layer. However, the vascular endothelium and extracellular matrix exhibited either a weak reaction or no reaction.

Ultracytochemistry of glycosaminoglycans in the canine knee synovium

The accurate localization and nature of glycosaminoglycans (GAGs) in the canine knee synovium were studied by ultracytochemical methods that involved high or low iron diamine-thiocarbohydrazide-silver proteinate (physical development) staining in combination with enzyme digestion control procedures. The results obtained indicated that heparan sulfates and hyaluronan were present mainly in the plasma membrane of the B (fibroblast-like) cells. In contrast, the plasma membrane of the A (macrophage-like) cells showed negative reactions after the histochemical examination. Dermatan sulfates, chondroitin sulfates (A and/or C) and hyaluronan were localized in the extracellular matrix of the synovial intima, whereby dermatan sulfates were confined to the fibrous component, whereas chondroitin sulfates and hyaluronan were found in the interfibrous matrix. Heparan sulfate was the only notable GAG molecular species localized in the basement membrane of the capillary wall. It is obvious that differences in the quality and localization of glycosaminoglycans in the canine synovial tissue are of specific interest in understanding normal functions as well as pathological alterations of the knee synovium in mammals.

Localization of glycosaminoglycans (GAGs) in pleomorphic adenoma (PA) of salivary glands: An immunohistochemical and histochemical evaluation: Zhao M, Takuta T, Ogawa I, et al. J Oral Pathol Med 27:272, 1998

Localization of glycosaminoglycans (GAGs) in pleomorphic adenoma (PA) of salivary glands: An immunohistochemical and histochemical evaluation: Zhao M, Takuta T, Ogawa I, et al. J Oral Pathol Med 27:272, 1998

Isolation, characterization and localization of glycosaminoglycans in growing antlers of wapiti ( Cervus elaphus)

Glycosaminoglycans were isolated from the four sections (tip, upper, middle and base) of the main beam of growing antlers of wapiti ( Cervus elaphus) by papain digestion and DEAE-Sephacel chromatography. Chondroitin sulfate was the major glycosaminoglycan in each section of antler accounting for, on average, 88% of the total uronic acid. The yield of chondroitin sulfate liberated from the tissue was approximately 6-fold greater in the cartilaginous (tip and upper) sections than in the bony (middle and base) sections. This was consistent with the higher intensity of glycosaminoglycan staining with either Alcian Blue or Safranin-O. The majority (average 88%) of chondroitin sulfate was precipitated with 40 and 50% ethanol. The average molecular size of chondroitin sulfate determined by gel chromatography on Sephacryl S-300 tended to be greater in the 40% ethanol than in the 50% ethanol fraction. In either fraction, the molecular size of chondroitin sulfate was smaller in cartilaginous tissues than in osseous tissues of growing antler. In addition to chondroitin sulfate, the antler contained small amounts of hyaluronic acid, dermatan sulfate and keratan sulfate. The immunohistochemical study showed wide distribution of chondroitin sulfate, decorin, and keratan sulfate throughout the antler. On the other hand, keratan sulfate was more prominent in the cartilaginous sections than in the bony sections where the anti-keratan sulfate monoclonal antibody staining was seen in the osteoid tissue only.

Age-related changes in the localization of glycosaminoglycans in condylar cartilage of the mandible in rats.

There is little information available regarding the morphological and biomolecular characteristics of mandibular condylar cartilage. The purpose of this study was to determine the age-related changes in the morphology and immunolocalization of glycosaminoglycans (GAGs) in mandibular condyles. The mandibular condylar cartilages from 4-, 8-, 16-, 32-, and 64-week-old Wistar male rats were examined to verify the localization of chondroitin-4-sulfate (Ch-4S), chondroitin-6-sulfate (Ch-6S) and keratan sulfate (KS) using an indirect immunofluorescent technique with three monoclonal antibodies for glycosaminoglycans, 2-B-6, 3-B-3 and 5-D-4, respectively. Morphologically, the condylar cartilage was a growth cartilage during growing periods, began to differentiate into articular cartilage from the central area of 16-week-old condyles, and became mature articular cartilage at 32 weeks of age. A regional difference was found in the morphological features and distribution of GAGs between the anterior, central, postero-superior and posterior areas of the condyles at each age. The immunohistochemical localizations of these three glycosaminoglycans showed age-related, morphology-dependent changes, from growth cartilage to articular cartilage-like cartilage. Immunoreactions for all of the antibodies decreased progressively with age in the interterritorial matrix, while the pericellular and territorial matrix in the condylar cartilage of the mandible maintained relatively higher immunoreactivity. In conclusion, age-related and regional differences in the localization of glycosaminoglycans Ch-4S, Ch-6S, and KS were found in the mandibular condyles in rats, and these changes are believed to be related to functional and developmental requirements.

Identification of Glycosaminoglycans in Age-Related Macular Deposits

We investigated the presence and localization of glycosaminoglycans in basal laminar deposit and drusen in age-related maculopathy. Conventional histological staining techniques and monoclonal antibodies specific for several glycosaminoglycans were used on paraffin-embedded human maculae. Furthermore, macular homogenates were analyzed with two-dimensional electrophoresis. Quantitative analysis of glycosaminoglycans was done spectrophotometrically using dimethylmethylene blue. Immunohistochemically, all basal laminar deposit stained positive for chondroitin 4-sulfate and focally positive for heparan sulfate proteoglycan. Drusen were not stained with any of the monoclonal antibodies. With two-dimensional electrophoresis, it was demonstrated that macular extracts with and without age-related maculopathy contained chondroitin sulfate. Heparan sulfate was only expressed in maculae with age-related maculopathy. The total amount of glycosaminoglycans was significantly higher in maculae with basal laminar deposit than in maculae without basal laminar deposit (P = .001). There were significant differences in the amount and composition of glycosaminoglycans between maculae with and without age-related maculopathy.

Localization of glycosaminoglycans in periodontal ligament during physiological and experimental tooth movement.

Localization of chondroitin sulphates in periodontal ligaments (PDL) of rat molar roots during physiological and experimental tooth movement were analysed immunohistochemically with the use of monoclonal antibodies, 3B3 and 2B6, specific to chondroitin 6-sulphate (CH-6S) and chondroitin 4-sulphate/dermatan sulfate (CH-4S/DS), respectively. The maxillary first molars of experimental animals were forced to move laterally with a 10 g weight by U-shaped wires for 3 and 7 d. In control animals, 3B3 epitope was seen in the PDL near to the bone surface facing the distal half of roots, which corresponded to the compressive side during physiological tooth movement. Immunoreactivity for 2B6 was weak or negative in the PDL. Both epitopes were present at osteoid, precementum, lacunae and canaliculli of osteocytes and cementocytes. In 3-d-treated animals, the early stage of hyalinization characterized with visible cells and fibres was observed in the PDL at the buccal side of the mesial root, which showed intense immunoreactivity for 3B3. Further 3B3 positive area seen in control animals changed its position from the distal to the buccal side of the PDL. Immunoreactivity for 2B6 did not change in the PDL of 3-d-treated animals. In 7-d-treated animals, the typical hyalinization characterized with no visible cells and fibres was seen in the PDL at the buccal sides of both mesial and disto-buccal roots, where both epitopes were present at the peripheral part of the tissue. Observation of serial sections suggested that the 3B3-positive area was present at the peripheral part of the 2B6-positive area. The present results suggest that the expression of CH-6S is related to the compressive force in non-hyalinized and hyalinized PDL, whereas that of CH-4S/DS is not influenced by the mechanical stress.

The Localization of Glycosaminoglycans in the Glenoicd Labrum

The Localization of Glycosaminoglycans in the Glenoicd Labrum

Localization of the glycosaminoglycans in the synovial tissues from osteoarthritic knees.

Localization of the glycosaminoglycans (GAG) was examined in the synovial membranes of patients with osteoarthritis under light microscopy using a fine cationic colloidal iron staining method combined with enzymatic digestion. Our staining method was very useful for demonstrating the difference in the localization of GAG in regions of the inflammatory site in the osteoarthritic synovial membrane. Hyaluronic acid was mainly located in connective tissues in the surface intercellular and perivascular spaces, chondroitin sulfate A/C in the highly fibrous part of and connective tissue around blood vessels, dermatan sulfate (chondroitin sulfate B) in the subsurface interstitium and vascular endothelial cells and heparan sulfate in part of vascular endothelial cells. No keratan sulfate was detected. GAG is reported to have an important role in cell movement, adherence and aggregation in the inflammatory sites. These findings should be useful for understanding the role of GAG in physiological and pathologic processes of secondary synovitis.

Glycosaminoglycans in porcine lung: an ultrastructural study using cupromeronic blue.

Glycosaminoglycans (GAGs) are essential components of the extracellular matrix contributing to the mechanical properties of connective tissues as well as to cell recognition and growth regulation. The ultrastructural localization of GAGs in porcine lung was studied by means of the dye Cupromeronic Blue in the presence of 0.3 M MgCl2 according to Scott's critical electrolyte concentration technique. GAGs were observed in locations described as follows. Pleura: Dermatan sulphate (DS) and chondroitin sulphate (CS) attached in the region of the d-band of collagen fibrils, interconnecting the fibrils; heparan sulphate (HS) at the surface of elastic fibers and in the basement membrane of the mesothelium and blood vessels. Bronchial cartilage: Abundant amounts of GAGs were observed in three zones: pericellular, in the intercellular matrix and at the perichondrial collagen. By enzyme digestion a superficial cartilage layer with predominantly CS could be distinguished from a deep zone with CS and keratan sulphate. The structure of the large aggregating cartilage proteoglycan was confirmed in situ. Airway epithelium: HS at the whole surface of cilia and microvilli and in the basement membrane of the epithelial cells. Alveolar wall: CS/DS at collagen fibrils, HS at the surface of elastic fibers and in the basement membranes of epithelium and endothelium.

Isolation, characterization, and localization of glycosaminoglycans in rabbit bone marrow

The glycosaminoglycans that exist in rabbit bone marrow were analyzed chemically, and their in situ localization was studied immunohistochemically. Femoral bone marrow of 3-month-old rabbits was defatted with organic solvents. Glycosaminoglycans were prepared from the defatted tissue after its digestion with pronase, treatment with mild alkali, and then digestion with DNase-I. The tissue contained glycosaminoglycans equivalent to 195 mg of hexosamine per femur, which accounted for 27.3% of the total hexosamine in the tissue. Studies with hyaluronidase from Streptomyces hyalurolyticus and chondroitinase ABC showed that the glycosaminoglycans were composed of hyaluronic acid (16% of the total glycosaminoglycan) and chondroitin 6-sulfate (79%). The chondroitin 6-sulfate was separated on Bio-Gel A-0.5m gel into two molecular species with mol wt of greater than 12,000 (Kd greater than 0.2) and approximately 8,000 (Kd = 0.47). Bone marrow digested with chondroitinase ABC and then treated with three monoclonal antibodies 4/8/9-A-2, 5/6/3-B-3, and 5/6/1-B-5, which were specific for unsaturated 4-sulfated, 6-sulfated, and nonsulfated disaccharide structures, respectively, at the nonreducing end of chondroitin sulfate chains, reacted with only 5/6/3-B-3. This result indicated that the chondroitin sulfate, isomer in the bone marrow is chondroitin 6- sulfate, consistent with the biochemical results. The chondroitin 6- sulfate was localized mainly in the extracellular compartment and was considered to be involved in construction of the hemopoietic microenvironment in the bone marrow.

Isolation, Characterization, and Localization of Glycosaminoglycans in Rabbit Bone Marrow

The glycosaminoglycans that exist in rabbit bone marrow were analyzed chemically, and their in situ localization was studied immunohistochemically. Femoral bone marrow of 3-month-old rabbits was defatted with organic solvents. Glycosaminoglycans were prepared from the defatted tissue after its digestion with pronase, treatment with mild alkali, and then digestion with DNase-I. The tissue contained glycosaminoglycans equivalent to 195 mg of hexosamine per femur, which accounted for 27.3% of the total hexosamine in the tissue. Studies with hyaluronidase from Streptomyces hyalurolyticus and chondroitinase ABC showed that the glycosaminoglycans were composed of hyaluronic acid (16% of the total glycosaminoglycan) and chondroitin 6-sulfate (79%). The chondroitin 6-sulfate was separated on Bio-Gel A-0.5m gel into two molecular species with mol wt of greater than 12,000 (Kd greater than 0.2) and approximately 8,000 (Kd = 0.47). Bone marrow digested with chondroitinase ABC and then treated with three monoclonal antibodies 4/8/9-A-2, 5/6/3-B-3, and 5/6/1-B-5, which were specific for unsaturated 4-sulfated, 6-sulfated, and nonsulfated disaccharide structures, respectively, at the nonreducing end of chondroitin sulfate chains, reacted with only 5/6/3-B-3. This result indicated that the chondroitin sulfate, isomer in the bone marrow is chondroitin 6-sulfate, consistent with the biochemical results. The chondroitin 6-sulfate was localized mainly in the extracellular compartment and was considered to be involved in construction of the hemopoietic microenvironment in the bone marrow.

A Method for Histochemical Localization of Glycosaminoglycans in Semi-thin Tissue Sections Embedded in GMA, Quetol 523 and Methyl Methacrylate

A Method for Histochemical Localization of Glycosaminoglycans in Semi-thin Tissue Sections Embedded in GMA, Quetol 523 and Methyl Methacrylate

The synthesis and localization of glycosaminoglycans in striated muscle differentiating in cell culture.

AbstractDay 3 mass muscle cultures incorporated 35SO4= and 3H· glucosamine into specific glycosaminoglycans that were identified by a variety of methods (enzyme susceptibility, nitrous acid degradation, electrophoresis, thin layer chromatography). More than 75% of the newly synthesized glycosaminoglycans appeared in the medium. In the medium, the glycosaminoglycans consisted of primarily hyaluronic acid (∼35%) and chondroitin sulfate (∼40%) with smaller amounts of chondroitin (∼15%) and heparan sulfate (∼10%). Among the glycosaminoglycans that remained in the cell layer, chondroitin sulfate (∼45%) and heparan sulfate (20‐40%) predominated and only a small amount of chondroitin (∼15%) and hyaluronic acid (∼5%) were present. In both the medium and cell layer, greater than 80% of the chondroitin sulfate was chondroitin‐6‐sulfate. Although mass cultures contain some non‐muscle cell types which undoubtedly contributed to total glycosaminoglycan synthesis, susceptibility of the autoradiographic grains associated with cells in muscle clones to testicular hyaluronidase digestion suggests that at least some glycosaminoglycans (chondroitin sulfate, in particular) were synthesized by muscle cells.Transmission electron microscopy revealed the presence of ruthenium red staining globular units along the surfaces of myotubes. Destruction of the integrity of surface associated ruthenium red stained material by both Streptomyces hyaluronidase and testicular hyaluronidase suggests that both hyaluronic acid and chondroitin sulfate are present on muscle cell surfaces.