Non-sulphated Glycosaminoglycans Research Articles

Läkning av trumhinneperforation—en komplex process påverkbar av många faktorer

It is known that the healing of a perforated tympanic membrane can be influenced by mechanical procedures as well as drugs. Animal experiments have recently shown that local treatment with steroids delays the healing of a tympanic membrane perforation, whereas hyaluronan (hyaluronic acid) improves the healing rate as well as the quality of the scar. Hyaluronan, a non-sulphated glycosaminoglycan, belongs to the extracellular matrix substances. In the present study another glycosaminoglycan—heparin—was applied on experimental tympanic membrane perforations. This report confirmed that also this extracellular matrix substance improves the healing rate of a tympanic membrane perforation and the scar quality.Spandow O, Hellstrom S. Lakning av trumhinneperforation—en komplex process paverkbar av manga faktorer. Acta Otolaryngol (Stockh) 1992; Suppl 492: 90–93.Spontanlakningen av trumhinneperforationer har studerats i saval kliniska som djurexperimentella material. Det ar kant att denna process kan paverkas bade ...

Antiangiogenic effect of sulphated and nonsulphated glycosaminoglycans and polysaccharides in the chick embryo chorioallantoic membrane

The inhibiting effect of sulphated and nonsulphated glycosaminoglycans and polysaccharides on the normal outgrowth of capillaries was tested in the chick embryo chorioallantoic membrane (CAM) with and without the presence of hydrocortisone. An antiangiogenic response to 50 micrograms of heparin and heparan sulphate (without hydrocortisone present) was observed in 38.8% and 23.1% of the CAMs, respectively, while the antiangiogenic response rate for dermatan sulphate, chondroitin sulphate A or C, hyaluronic acid and keratan sulphate was 15.9-0%. All sulphated homopolysaccharides tested were more effective than the naturally occurring glycosaminoglycans. Nonsulphated dextran and (methyl) cellulose had no antiangiogenic effect, while largely desulphated heparin retained such an effect. Hydrocortisone generally improved the antiangiogenic effect, a 100% response was obtained when it was combined with cellulose sulphate or fucoidan (polyfucose sulphate derived from marine algae), but the antiangiogenic effect of the largely desulphated heparin was unaffected by the presence of hydrocortisone. The results show that different polysulphated polysaccharides also have an antiangiogenic effect, without the addition of corticosteroids. The effect was apparently independent of their degree of sulphation, but the glycosidic structure may be of critical importance.

Endothelial cell stimulation of smooth muscle glycosaminoglycan synthesis can be accounted for by transforming growth factor beta activity

Endothelial cell conditioned medium (ECCM) contains a factor which markedly stimulates smooth muscle cell (SMC) glycosaminoglycan (GAG) synthesis. We report here that the factor responsible is transforming growth factor beta (TGF-β) as assessed by (1) protease and thiol sensitivity, (2) heat and acid enhancement of ECCM activity, and (3) neutralisation of ECCM activity by anti-TGF-β-immunoglobulin. Anti-TGF-β-neutralisation was effective against increases in both sulphated and non-sulphated GAG. Previous studies showed that ECCM from EC of varying densities stimulated individual GAG to varying degrees. ECCM from low density EC preferentially stimulated hyaluronic acid (HA) whereas ECCM from intermediate and high density cultures stimulated increasing amounts of sulphated GAG. Exposure of SMC to varying concentrations of TGF-β produced a similar pattern of response. Very low amounts of TGF-β (< 10–500 pg/10 cells) stimulated a marked and significant increase in HA synthesis. Increase in chondroitin sulphate 4 6 was most marked at TGF-β levels from 500–1000 pg/10 6 cells. At levels above 1000 pg/10 6 cells both HA and sulphated GAG synthesis decreased but still remained elevated above controls. These findings indicate that TGF-β alone can account for the changes in SMC GAG synthesis stimulated by ECCM. It was also found, however, that heat-treated SMC conditioned medium stimulated SMC GAG synthesis, thus SMC may contribute to the control of their own GAG synthesis through autocrine TGF-β activity.

An autoradiographic study using [3H]glucosamine of gonadotrophin regulation of proteoglycan and glycoprotein synthesis in developing mouse follicles.

Autoradiography of serial sections of ovaries of immature mice was used, after an injection of [3H]glucosamine, to follow the migration of newly synthesized macromolecules into preantral follicles during the period of treatment with PMSG and hCG. [3H]Glucosamine was injected at the same time as the PMSG or 2-h before the time of autopsy. PMSG stimulated a modest uptake of [3H]glucosamine into the zona pellucida of preantral follicles. However, the in-vivo synthesis of labelled macromolecules increased substantially during the period of hCG stimulation, especially in those mice in which the label was injected at the same time as the PMSG. After both short and longer term exposure to [3H]glucosamine, the maximum uptake of label in preantral follicles occurred 4-8 h after the injection of hCG, indicating that hCG rather than PMSG probably exerts the greatest control over the uptake and incorporation of [3H]glucosamine into the zona pellucida and oocyte of preantral follicles. It is suggested that [3H]glucosamine is largely incorporated into non-sulphated glycosaminoglycans.

Histochemical localization of skin glycosaminoglycans during feather development in the chick embryo.

The distribution of glycosaminoglycans (GAGs) was studied in embryonic chick skin, using alcian blue staining with critical electrolyte concentration and glycanase treatment, immunofluorescence and transmission electron microscopy. Light microscopy revealed an uneven distribution of sulphated and non-sulphated GAGs at all stages of feather development. Along the dermal-epidermal junction and throughout the depth of the dermis, staining was stronger inside the feathers than in the interplumar skin. With increasing MgCl2 concentration, the decrease in stain intensity along the dermal-epidermal junction was stronger in interplumar skin than inside feather structures, indicating that sulphated GAGs are more abundant within feathers than in interplumar skin. The same differential sensitivity to electrolyte concentration was noted in the dermis, except at the feather placode stage, when labelling inside the dermal condensation was virtually wiped out at 0.6 M MgCl2 and higher concentrations, whereas it persisted in the surrounding dermis up to 0.8 M MgCl2, indicating that the dermal condensation contains a larger amount of hyaluronate than non-feather-forming dermis. Enzyme treatment of sections with Streptomyces hyaluronidase as compared with those treated with chondroitinase ABC corroborated these findings. Immunofluorescent detection of heparan sulphate proteoglycan revealed the presence of the antigen along the dermal-epidermal junction at all stages of feather development, with peaks of brightness in discrete spots of feather structures. Electron microscopy revealed the presence of ruthenium red and tannic acid positive material in the dermal-epidermal junctional zone and inside the dermis. The density of marked granules was somewhat higher in intraplumar than in interplumar regions. These observations demonstrate that certain sulphated and non-sulphated GAGs are distributed in a microheterogeneous manner, which appears to be related to the morphogenetic events of feather development. They are discussed in view of the possible role these components might play in dermal-epidermal interactions. They strengthen the notion, already gained from previous studies on the localization of interstitial collagens and fibronectin, that extracellular matrix components play an important structural and informative role in organogenesis.

Glycosaminoglycans in human gingival crevicular fluid as indicators of active periodontal disease

The glycosaminoglycans (GAG) in gingival crevicular fluid (GCF) were investigated by cellulose-acetate electrophoresis of samples from individual sites of defined clinical conditions variously affecting the tissues of the periodontium. The non-sulphated GAG, hyaluronic acid, was present in all samples and was the only major band from sites of chronic gingivitis. An additional sulphated GAG band identified by enzymic digestions as chondroitin-4-sulphate, was detected in GCF from sites of untreated-advanced periodontitis. Initial samples from sites of early periodontitis and juvenile periodontitis yielded a similar additional band which was not detected, however, in samples collected after either surgery to eliminate deep pockets or daily subgingival irrigation with a chlorhexidine solution. Sulphated GAG was also present in fluid from the control situations, i.e. of teeth either undergoing orthodontic movement or showing evidence of trauma from occlusion, and from healing tooth-extraction wounds. Thus the presence of such a component in GCF correlates with those clinical conditions in which degradative changes are occurring in the deeper-periodontal tissues. The electrophoretic profile of GAG in a sample of GCF may be a sensitive laboratory method of indicating active phases of destructive periodontal disease at individual sites.

Extracellular matrix production by mouse 3T3-F442A cells during adipose differentiation in culture

When cultured 3T3-F442A cells undergo adipose differentiation, they produce extracellular matrix (ECM) that is not present in undifferentiated cells. This ECM stains strongly with ruthenium red, tannic acid and with Alcian blue at both pH 1 and 2.5, showing histochemical characteristics similar to sulphated and non-sulphated glycosaminoglycans. Under the electron microscope, ECM was observed bound to the cell surface and in the intercellular space; it was composed of fibrils of several thicknesses with attached granules and fibrous long-spacing forms of collagen. In addition, adipocytes were observed as rounded cells interconnected with the ECM fibrils, thus giving rise to fat cell clusters similar to the adipocyte lobules found in adipose tissue. Since fat cell clusters in culture emerge by clonal expansion of one adipose precursor cell, we suggest that this ECM can keep daughter adipocytes interconnected during differentiation. ECM production by adipocytes might have some significance for the formation of fat cell lobules in vivo.

Synthesis of glycosaminoglycans and other macromolecules by embryo calf epiphyseal cells in culture.

Monolayer cell cultures from the distal femoral epiphyses of embryo calves were studied following the first subculture, which was carried out after confluence in primary culture. Light microscopic examination revealed scattered deposits of metachromatic-staining material; on electron microscopy fine fibrils considered to be developmental collagen were seen. After several days in culture lacuna-like patterns of cells were seen. The nature of the cell secretions were studied by radioactive precursors, which were fractionated on Sephadex G200 and by ion exchange chromatography. Enzyme digestion with bacterial and testicular hyaluronidase and chondroitinase-AC and-ABC revealed that the cells synthesized 70% sulphated, and 30% non-sulphated glycosaminoglycans. Of the sulphated glycosaminoglycans 70% was chondroitin-4-sulphate, 20% chondroitin-6-sulphate, and the remainder probably keratansulphate. Studies were labelled amino acid precursors suggested that the cells synthesized a high-molecular weight protein containing hydroxyproline, as well as some non-collagenous protein, shown by tryptophan incorporation

Molecular size and xylose content of urinary glycosaminoglycans

1. 1. The molecular weights of the non-ultrafilterable glycosaminoglycans isolated from normal human urine were determined by gel chromatography, and their neutral-sugar contents were analysed by gas-liquid chromatography. 2. 2. The molecular weights ( M W ) of the polysaccharides were as follows: chondroitin sulphate fractions, from 8000 to 13000; dermatan sulphate, 16000; heparan sulphate fractions, from 8000 to approx. 30000; the non-sulphated glycosaminoglycans (hyaluronic acid and chondroitin), 8000. The ratio of M W to M n did not exceed 1.5 in any of the fractions. The calculated Stokes' radii of the chondroitin sulphate fractions were 24–36 Å, whereas the Strokes' radii of the largest heparan sulphate fractions were about 60 Å. 3. 3. The chondroitin sulphates, the dermatan sulphate, and the heparan sulphates contained slightly less than one residue of xylose per chain, independently of size. This finding suggests that no significant endopolysaccharidase activity had been exerted on the excreted glycosaminoglycans. 4. 4. The non-sulphated urinary glycosaminoglycans, which consisted of hyaluronic acid ( 2 3 ) and chondroitin ( 1 3 ), were found to contain about one xylose residue and about two residues of galactose per chain.