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Sample records for acid glycosaminoglycan mucopolysaccharide

  1. Cytochemical localisation of lysosomal enzymes and acidic mucopolysaccharides in the salivary glands of Aplysia depilans (Opisthobranchia).

    Lobo-da-Cunha, A

    2002-04-01

    Three types of secretory cells were reported in the salivary glands of Aplysia depilans: granular cells, vacuolated cells and mucocytes. To improve the characterisation of these cells, cytochemical methods for the detection of lysosomal enzymes and acidic mucopolysaccharides were applied. In granular cells, acid phosphatase and arylsulphatase were present in small lysosomes and in some secretory granules. The secretory granules could have received these enzymes after fusion with the small lysosomes that were frequently found very close to them. These cells were not stained with colloidal iron because they do not contain acidic mucopolysaccharides. In vacuolated cells, acid phosphatase and arylsulphatase were detected in lysosomes but not in the secretory vacuoles. Colloidal iron staining revealed the presence of acidic mucopolysaccharides in the vacuoles and in the Golgi apparatus of these cells. In mucocytes, lysosomes were very rare, but the secretion of these cells was very rich in acidic mucopolysaccharides. The filamentous network within the secretory vesicles was completely covered with iron particles, but practically no particles were observed over the granular masses attached to the membrane of the vesicles. Iron particles were also found in the trans-face cisternae of the U-shaped Golgi stacks, but were not seen in the cis-face cisternae or in the rough endoplasmic reticulum. PMID:12117284

  2. Immunomodulatory Effect of Stichopus japonicus Acid Mucopolysaccharide on Experimental Hepatocellular Carcinoma in Rats

    Xiao-Jun Ji

    2013-06-01

    Full Text Available Stichopus japonicus acid mucopolysaccharide (SJAMP is an important biologically active compound that can be extracted from the body wall of the sea cucumber. The present study investigated the anti-tumor and immunomodulatory effects of SJAMP in an experimental hepatocellular carcinoma (HCC model in rats. Three doses of SJAMP (17.5 mg/kg, 35 mg/kg, and 70 mg/kg administered 5 days/week via oral gavage were given to rats with diethylnitrosamine (DEN-induced HCC. SJAMP treatment significantly inhibited DEN-induced HCC by reducing both the number and mean volume of nodules, decreasing serum a-fetoprotein (AFP levels and proliferating cell nuclear antigen (PCNA expression in liver, and increasing p21 expression. Furthermore, SJAMP decreased the serum levels of ALT, AST, GGT and TNF-α and increased serum IL-2. SJAMP administration also improved indices of spleen and thymus function and improved both macrophage phagocytosis and NK cell-mediated tumoricidal activity. Moreover, CD3+ and CD4+ T lymphocyte levels recovered significantly and the CD4+/CD8+ T cell ratio normalized in a dose-dependent manner. In conclusion, SJAMP effectively inhibited the growth of HCC through the stimulation of immune organs and tissue proliferation, leading to the enhancement of cellular immunity pathways in rats.

  3. Antiviral effects of Stichopus japonicus acid mucopolysaccharide on hepatitis B virus transgenic mice

    Xin, Yongning; Li, Wei; Lu, Linlin; Zhou, Li; Victor, David W.; Xuan, Shiying

    2016-08-01

    Hepatitis B virus (HBV) is a significant global pathogen and efficient cure for HBV patients is still a challenging goal. We previously reported that acidic mucopolysaccharide from stichopus japonicus selenka (SJAMP) could inhibit HBsAg and HBeAg expression in vitro. However, the potential anti-HBV effects of SJAMP in vivo have not yet been explored. In this study, we show that SJAMP exhibits potent anti-HBV activity in HBV transgenic mice in a dose-dependent manner. Specifically, sixty HBV transgenic male BALB/c mice were randomly selected to receive the treatment of PBS, low dose SJAMP (30 mg kg-1), middle dose SJAMP (40 mg kg-1), high dose SJAMP (50 mg kg-1) and IFN (45 IU kg-1) for 30 d. SJAMP treatment suppressed serum HBV-DNA, and liver HBsAg and HBcAg levels in HBV-transgenic mice. The present study highlights the potential application of SJAMP in HBV therapy.

  4. Tunicamycin inhibits biosynthesis of acid mucopolysaccharides in cultures of chick embryo fibroblasts

    The time course of the incorporation of 14C-glucosamine into extracellular mucopolysaccharides (MPS) and that of H235SO4 into intracellular and extracellular MPS were experimentally studied. The monolayer cultures of chick embrio fibroblasts in test tubes were prepared with modified Eagle's minimal essential medium supplemented with 5% calf serum. After confluence, the medium was replaced with the prewarmed fresh one containing 0.4 μCi/ml glucosamine-1-14C or 2 μCi/ml H235SO4. Tunicamycin (TM) was added at the same time as the radioactive compounds. At the appropriate time intervals of incubation at 370C, cultured cells were sampled, and the medium was withdrawn. The cell sheets were rinsed twice with 1 ml each of could distilled water. Acid MPS were precipitated with cetylpyridinium chloride. The precipitate was collected by centrifugation. Insoluble matter was collected on glass fiber filter. After drying, the radioactivity retained on the filter was counted with a scintillation counter. This fraction is of extracellular MPS. In the case of H235SO4 incorporation, the step of NaOH treatment was omitted. This fraction is of intracellular MPS. As a result, slight difference was observed in the degree of inhibition by 1.0 μg/ml TM between the two fractions. TM also inhibited the incorporation of H235SO4. The degree of inhibition of H235SO4 incorporation into intracellular MPS was similar to that of 14C-glucosamine. The inhibition of MPS biosynthesis by TM suggests the possibility of participation of lipid-linked intermediate in the biosynthesis of MPS. (Iwakiri, K.)

  5. Acid glycosaminoglycans in plasma. I. Determination.

    Friman, C; Juvani, M

    1977-01-01

    Plasma glycosaminoglycans (GAG) were isolated from 10 ml of plasma by a modification of the method of Calatroni et al. (3). DE-52 anion-exchange cellulose was used in the isolation of the fraction operationally defined as free GAG. Chondroitin sulphate and heparin added to plasma were quantitatively recovered in this fraction. After proteolysis with papain the fraction operationally defined as bound GAG was isolated using anion-exchange resin AG 1 X 2. GAG were measured as hexuronate with the m-hydroxydiphenyl method of Blumenkrantz and Asboe-Hansen (7) which was superior to various modifications of the carbazole/borate carbazole procedures. In 15 healthy females and in 15 healthy males the concentrations of the free GAG (mean +/- S.D., expressed as microgram per 10 ml of plasma) were: 12.2 +/- 2.8 and 16.8 +/- 3.8 (P less than 0.001); of the bound GAG 40.4 +/- 7.7, and 40.2 +/- 11.6; and of the total GAG 52.7 +/- 9.0 and 57.0 +/- 10.4, respectively. With the isolation procedures used, plasma GAG were obtained in sufficient quantity for their electrophoretic characterization. Assay of plasma GAG can be performed with satisfactory accuracy and precision within two days by the present method. In clinical chemistry its application to the study of proteoglycan and GAG metabolism in various diseases may prove valuable. PMID:897589

  6. Mucopolysaccharides from psyllium involved in wound healing.

    Westerhof, W; Das, P K; Middelkoop, E; Verschoor, J; Storey, L; Regnier, C

    2001-01-01

    Mucopolysaccharides derived from the husk of psyllium (Plantago ovata) have properties beneficial for wound cleansing and wound healing. Recent studies indicate that these mucopolysaccharides also limit scar formation. Our in vitro and in vivo studies aimed to investigate the mechanisms involved, e.g., fluid absorption, bacterial adherence and in vitro stimulatory effects on macrophages, which are pivotal in wound healing. The mucopolysaccharides contained in a sachet (Askina Cavity) or in a hydrocolloid mixture (Askina Hydro) were found to have a gradual and sustained absorbency over a period of 7 days, amounting to 4-6 times their weight in water. The swelling index was 9 mm after 312 h. Adherence of wound bacteria to the mucopolysaccharides started after 2 h and was more pronounced after 3 h. Semiquantitative measurements of bacterial adherence used centrifugation and subsequent optical density determinations of supernatant. These confirmed the strong adherence potential of psyllium particles. Lactic acid dehydrogenase staining of pretreated cultured human skin explants did not reveal toxicity of the mucopolysaccharides derived from psyllium husk. Langerhans' cell migration from the epidermis was negligible and interleukin-1 beta expression in the explants was not significant, supporting the very low allergenic potential of psyllium. The characteristics of mucopolysaccharide granulate derived from psyllium husk in Askina Cavity and Askina Hydro related to fluid absorption, bacterial adherence, biocompatibility, stimulation of macrophages, irritancy response and allergenicity showed an optimal profile, supporting the good clinical performance of wound healing products containing psyllium husk. PMID:11951574

  7. Emergent Molecular Recognition through Self-Assembly: Unexpected Selectivity for Hyaluronic Acid among Glycosaminoglycans.

    Noguchi, Takao; Roy, Bappaditya; Yoshihara, Daisuke; Sakamoto, Junji; Yamamoto, Tatsuhiro; Shinkai, Seiji

    2016-05-01

    Oligophenylenevinylene (OPV)-based fluorescent (FL) chemosensors exhibiting linear FL responses toward polyanions were designed. Their application to FL sensing of glycosaminoglycans (heparin: HEP, chondroitin 4-sulfate: ChS, and hyaluronic acid: HA) revealed that the charge density encoded as the unit structure directs the mode of OPV self-assembly: H-type aggregate for HEP with 16-times FL increase and J-type aggregate for HA with 93-times FL increase, thus unexpectedly achieving the preferential selectivity for HA in contrast to the conventional HEP selective systems. We have found that the integral magnitude of three factors consisting of binding mechanism, self-assembly, and FL response can amplify the structural information on the target input into the characteristic FL output. This emergent property has been used for a novel molecular recognition system that realizes unconventional FL sensing of HA, potentially applicable to the clinical diagnosis of cancer-related diseases. PMID:27060601

  8. Glycosaminoglycans in the Human Cornea: Age-Related Changes

    Pacella, Elena; Pacella, Fernanda; De Paolis, Giulio; Parisella, Francesca Romana; Turchetti, Paolo; Anello, Giulia; Cavallotti, Carlo

    2015-01-01

    AIM To investigate possible age-related changes in glycosaminoglycans (GAGs) in the human cornea. The substances today called GAGs were previously referred to as mucopolysaccharides. METHODS Samples of human cornea were taken from 12 younger (age 21 ± 1.2) and 12 older (age 72 ± 1.6) male subjects. Samples were weighed, homogenized, and used for biochemical and molecular analyses. All the quantitative results were statistically analyzed. RESULTS The human cornea appears to undergo age-related changes, as evidenced by our biochemical and molecular results. The total GAG and hyaluronic acid counts were significantly higher in the younger subjects than in the older subjects. The sulfated heavy GAGs, such as chondroitin, dermatan, keratan, and heparan sulfate, were lower in the younger subjects than in the older subjects. DISCUSSION GAGs of the human cornea undergo numerous age-related changes. Their quantity is significantly altered in the elderly in comparison with younger subjects. GAGs play an important role in age-related diseases of the human cornea. PMID:25674020

  9. 一种新型粘多糖结构与性能的检测%The structure and performance testing of a new kind of mucopolysaccharide

    丛涛; 徐永斌; 赵晨希; 张淑荣; 刘春巧; 张鹏

    2011-01-01

    粘多糖是由糖醛酸和氨基己糖交替连接成的高分子物质,理化性质独特,应用范围广泛.通过对突变株兽疫链球菌Streptococcus zooepidemicus BU 100进行发酵,可产一种新型粘多糖(下文用粘多糖A代替).利用咔唑法、Elson-Morgan法、考马斯亮蓝法、红外光谱以及13C核磁共振谱测定粘多糖A的结构,结果显示粘多糖A中糖醛酸和氨基糖的摩尔比例接近1∶1,蛋白含量符合标准(<0.1%);粘多糖A图谱中出现的结构特征峰大部分与透明质酸相同.对粘多糖A的实用性能进行检测,并用透明质酸做对比,结果表明透明质酸在两种湿度下的吸湿性均要好于粘多糖A,但粘多糖A的保湿性要好于透明质酸.粘多糖A总体的抗氧化性好于透明质酸,并且粘多糖A耐透明质酸酶.粘多糖A可作为保湿剂、润滑剂、抗氧化剂等被更加有效地应用在医疗和化妆品等领域.%Mucopolysaccharide was made of uronic acid and hexosamine. Due to the unique physical and chemical properties, mucopolysaccharides have been used in various fields. After collecting and enrichment culture of active nasal mucosa scraped from newly slaughtered cattle, we got a kind of streptococcus zooepidemicus which was named BU100. With its culture it can secrete a high yield new Mucopolysaccharide A. As there were uronic acid and hexosamine in mucopolysaccharide, the methods of carbazole and Elson-Morgan were used to detect mucopolysaccharide A. In addition, the major impurity (protein) in mucopolysaccharide was detected by the method of Coomassie brilliant blue. According to these methods, it was found that the ratio of uronic acid and hexosamine in mucopolysaccharide A was nearly 1:1 which was the same as that in Hyaluronic Acid (HA), and the quantity of pro-tein meets the standards (<0.1%) requirements. Furthermore with the IR, NMR, the functional group of mucopolysaccharide A was determined and compared to HA. It therefore indicates that

  10. Suppression of glycosaminoglycan synthesis by articular cartilage, but not of hyaluronic acid synthesis by synovium, after exposure to radiation

    Hugenberg, S.T.; Myers, S.L.; Brandt, K.D.

    1989-04-01

    We recently found that injection of 2 mCi of yttrium 90 (90Y; approximately 23,000 rads) into normal canine knees stimulated glycosaminoglycan (GAG) synthesis by femoral condylar cartilage. The present investigation was conducted to determine whether radiation affects cartilage metabolism directly. Rates of GAG synthesis and degradation in normal canine articular cartilage were studied following irradiation. Cultured synovium from the same knees was treated similarly, to determine the effects of irradiation on hyaluronic acid synthesis. Twenty-four hours after exposure to 1,000 rads, 10,000 rads, or 50,000 rads, 35S-GAG synthesis by the cartilage was 93%, 69%, and 37%, respectively, of that in control, nonirradiated cartilage. The effect was not rapidly reversible: 120 hours after exposure to 50,000 rads, GAG synthesis remained at only 28% of the control level. Autoradiography showed marked suppression of 35S uptake by chondrocytes after irradiation. Cartilage GAG degradation was also increased following irradiation: 4 hours and 8 hours after exposure to 50,000 rads, the cartilage GAG concentration was only 66% and 54%, respectively, of that at time 0, while corresponding values for control, nonirradiated cartilage were 90% and 87%. In contrast to its effects on cartilage GAG metabolism, radiation at these levels had no effect on synovial hyaluronic acid synthesis.

  11. Suppression of glycosaminoglycan synthesis by articular cartilage, but not of hyaluronic acid synthesis by synovium, after exposure to radiation

    We recently found that injection of 2 mCi of yttrium 90 (90Y; approximately 23,000 rads) into normal canine knees stimulated glycosaminoglycan (GAG) synthesis by femoral condylar cartilage. The present investigation was conducted to determine whether radiation affects cartilage metabolism directly. Rates of GAG synthesis and degradation in normal canine articular cartilage were studied following irradiation. Cultured synovium from the same knees was treated similarly, to determine the effects of irradiation on hyaluronic acid synthesis. Twenty-four hours after exposure to 1,000 rads, 10,000 rads, or 50,000 rads, 35S-GAG synthesis by the cartilage was 93%, 69%, and 37%, respectively, of that in control, nonirradiated cartilage. The effect was not rapidly reversible: 120 hours after exposure to 50,000 rads, GAG synthesis remained at only 28% of the control level. Autoradiography showed marked suppression of 35S uptake by chondrocytes after irradiation. Cartilage GAG degradation was also increased following irradiation: 4 hours and 8 hours after exposure to 50,000 rads, the cartilage GAG concentration was only 66% and 54%, respectively, of that at time 0, while corresponding values for control, nonirradiated cartilage were 90% and 87%. In contrast to its effects on cartilage GAG metabolism, radiation at these levels had no effect on synovial hyaluronic acid synthesis

  12. Synthetic Approaches to L-Iduronic Acid and L-Idose: Key Building Blocks for the Preparation of Glycosaminoglycan Oligosaccharides.

    Mohamed, Shifaza; Ferro, Vito

    2015-01-01

    L-Iduronic acid (IdoA) is an important monosaccharide component of glycosaminoglycans (GAGs) such as heparin, heparan sulfate and dermatan sulfate. GAGs are complex, highly sulfated polysaccharides that mediate a multitude of physiological and pathological processes via their interactions with a range of diverse proteins. The main challenge in the synthesis of GAG oligosaccharides is the efficient gram-scale preparation of IdoA building blocks since neither IdoA nor L-idose is commercially available or readily accessible from natural sources. In this review, the different synthetic approaches for the preparation of IdoA and its derivatives, including L-idose, are presented and discussed. Derivatives of the latter are often used in GAG synthesis and are elaborated to IdoA via selective oxidation at C-6 after incorporation into a GAG chain. Particular focus will be given to the preparation of IdoA synthons most commonly used for GAG oligosaccharide synthesis, and on the progress made since the last systematic review in this area. PMID:26613814

  13. Interstitial cystitis/bladder pain syndrome and glycosaminoglycans replacement therapy.

    Cervigni, Mauro

    2015-12-01

    Interstitial cystitis/bladder pain syndrome (IC/BPS) is a debilitating chronic disease characterized by discomfort or recurrent abdominal and pelvic pains in the absence of urinary tract infections. Its symptomatology includes discomfort, increased bladder pressure, sensitivity and intense pain in the bladder and pelvic areas, increased voiding frequency and urgency, or a combination of these symptoms. For these reasons, this pathology has a very negative impact on quality of life. The etiology of IC/BPS is still not well understood and different hypotheses have been formulated, including autoimmune processes, allergic reactions, chronic bacterial infections, exposure to toxins or dietary elements, and psychosomatic factors. The finding of an effective and specific therapy for IC/BPS remains a challenge for the scientific community because of the lack of a consensus regarding the causes and the inherent difficulties in the diagnosis. The last recent hypothesis is that IC/BPS could be pathophysiologically related to a disruption of the bladder mucosa surface layer with consequent loss of glycosaminoglycans (GAGs). This class of mucopolysaccharides has hydrorepellent properties and their alteration expose the urothelium to many urinary toxic agents. It has been hypothesized that when these substances penetrate the bladder wall a chain is triggered in the submucosa. In order to improve the integrity and function of the bladder lining, GAG layer replenishment therapy is widely accepted as therapy for patients with IC/BPS who have poor or inadequate response to conventional therapy. Currently, Chondroitin sulfate (CS), heparin, hyaluronic acid (HA), and pentosan polysulphate (PPS), and combinations of two GAGs (CS and HA) are the available substances with different effectiveness rates in patients with IC/BPS. There are four different commercially available products for GAG replenishment including CS, heparin, HA and PPS. Each product has different concentrations and

  14. Proteoglycans and their heterogeneous glycosaminoglycans at the atomic scale.

    Sattelle, Benedict M; Shakeri, Javad; Cliff, Matthew J; Almond, Andrew

    2015-03-01

    Proteoglycan spatiotemporal organization underpins extracellular matrix biology, but atomic scale glimpses of this microarchitecture are obscured by glycosaminoglycan size and complexity. To overcome this, multimicrosecond aqueous simulations of chondroitin and dermatan sulfates were abstracted into a prior coarse-grained model, which was extended to heterogeneous glycosaminoglycans and small leucine-rich proteoglycans. Exploration of relationships between sequence and shape led to hypotheses that proteoglycan size is dependent on glycosaminoglycan unit composition but independent of sequence permutation. Uronic acid conformational equilibria were modulated by adjacent hexosamine sulfonation and iduronic acid increased glycosaminoglycan chain volume and rigidity, while glucuronic acid imparted chain plasticity. Consequently, block copolymeric glycosaminoglycans contained microarchitectures capable of multivalent binding to growth factors and collagen, with potential for interactional synergy at greater chain number. The described atomic scale views of proteoglycans and heterogeneous glycosaminoglycans provide structural routes to understanding their fundamental signaling and mechanical biological roles and development of new biomaterials. PMID:25645947

  15. Glycosaminoglycans from earthworms (Eisenia andrei)

    Im, A-Rang; Park, Youmie; Sim, Joon-Soo; Zhang, Zhenqing; Liu, Zhenling; Linhardt, Robert J.; Kim, Yeong Shik

    2009-01-01

    The whole tissue of the earthworm (Eisenia andrei) was lyophilized and extracted to purify glycosaminoglycans. Fractions, eluting from an anion-exchange column at 1.0 M and 2.0 M NaCl, showed the presence of acidic polysaccharides on agarose gel electrophoresis. Monosaccharide compositional analysis showed that galactose and glucose were most abundant monosaccharides in both fractions. Depolymerization of the polysaccharide mixture with glycosaminoglycandegrading enzymes confirmed the presenc...

  16. Differentiating chondroitin sulfate glycosaminoglycans using collision-induced dissociation; uronic acid cross-ring diagnostic fragments in a single stage of tandem mass spectrometry.

    Kailemia, Muchena J; Patel, Anish B; Johnson, Dane T; Li, Lingyun; Linhardt, Robert J; Amster, I Jonathan

    2015-01-01

    The stereochemistry of the hexuronic acid residues of the structure of glycosaminoglycans (GAGs) is a key feature that affects their interactions with proteins and other biological functions. Electron based tandem mass spectrometry methods, in particular electron detachment dissociation (EDD), have been able to distinguish glucuronic acid (GlcA) from iduronic acid (IdoA) residues in some heparan sulfate tetrasaccharides by producing epimer-specific fragments. Similarly, the relative abundance of glycosidic fragment ions produced by collision-induced dissociation (CID) or EDD has been shown to correlate with the type of hexuronic acid present in chondroitin sulfate GAGs. The present work examines the effect of charge state and degree of sodium cationization on the CID fragmentation products that can be used to distinguish GlcA and IdoA containing chondroitin sulfate A and dermatan sulfate chains. The cross-ring fragments (2,4)A(n) and (0,2)X(n) formed within the hexuronic acid residues are highly preferential for chains containing GlcA, distinguishing it from IdoA. The diagnostic capability of the fragments requires the selection of a molecular ion and fragment ions with specific ionization characteristics, namely charge state and number of ionizable protons. The ions with the appropriate characteristics display diagnostic properties for all the chondroitin sulfate and dermatan sulfate chains (degree of polymerization of 4-10) studied. PMID:26307707

  17. A method for measuring disease-specific iduronic acid from the non-reducing end of glycosaminoglycan in mucopolysaccharidosis type II mice.

    Shimada, Yohta; Wakabayashi, Taichi; Akiyama, Kazumasa; Hoshina, Hiroo; Higuchi, Takashi; Kobayashi, Hiroshi; Eto, Yoshikatsu; Ida, Hiroyuki; Ohashi, Toya

    2016-02-01

    Mucopolysaccharidosis type II (MPS II) is an X-linked lysosomal storage disorder arising from deficiency of iduronate-2-sulfatase (IDS), which results in progressive accumulation of glycosaminoglycans (GAGs) in multiple tissues. Accumulated GAGs are generally measured as the amount of total GAGs. However, we recently demonstrated that GAG accumulation in the brain of MPS II model mice cannot be reliably detected by conventional dye-binding assay measuring total GAGs. Here we developed a novel quantitative method for measurement of disease-specific GAGs based on the analysis of 2-sulfoiduronic acid levels derived from the non-reducing terminal end of the polysaccharides by using recombinant human IDS (rhIDS) and recombinant human iduronidase (rhIDUA). This method was evaluated on GAGs obtained from the liver and brain of MPS II mice. The GAGs were purified from tissue homogenates and then digested with rhIDS and rhIDUA to generate a desulfated iduronic acid from their non-reducing terminal end. HPLC analysis revealed that the generated iduronic acid levels were markedly increased in the liver and cerebrum of the MPS II mice, whereas the uronic acid was not detected in wild-type mice. These results indicate that this assay clearly detects the disease-specific GAGs in tissues from MPS II mice. PMID:26051019

  18. Autoradiographic study of mucopolysaccharide synthesis in rat ovary

    The study was carried out in an effort to determine the cell type responsible for mucopolysaccharide synthesis in the ovary of mammalia. Sexually mature female rats were used. The animals were injected two times with D-(6-3H) glucosamine hydrochloride ans sacrified at different intervals thereafter to prepare explants from their ovaries. As early as the very first hour after the application of radioactive glucosamine, efficient labelling of the granulosa cells, zona pellucida and, to a lesser extent, the oocytes of the medium and large follicles was established. Labelling of follicular fluid was established one day later. At these earlier terms of the investigation, the granulosa cells, surrounding zona pellicida and antrum folliculi, were better labelled, (7th and 42th day) the autoradiographic reaction over all mentioned structures rapidly decreased, being preserved for the longest above zona pellucida and liquor folliculi. At the first hour of the investigation the silver grains appeared to be situated mainly close to the oocyte membrane, and later onto the granulosa cells. The results show that to all probablity, both the granulosa cells and the oocyte take part in the synthesis of mucopolysaccharides, which are part of the composition of zona pellucida and liquor folliculi. (A.B.)

  19. Glycosaminoglycans and infection.

    Aquino, Rafael S; Park, Pyong Woo

    2016-01-01

    Glycosaminoglycans (GAGs) are complex linear polysaccharides expressed in intracellular compartments, at the cell surface, and in the extracellular environment where they interact with various molecules to regulate many cellular processes implicated in health and disease. Subversion of GAGs is a pathogenic strategy shared by a wide variety of microbial pathogens, including viruses, bacteria, parasites, and fungi. Pathogens use GAGs at virtually every major portals of entry to promote their attachment and invasion of host cells, movement from one cell to another, and to protect themselves from immune attack. Pathogens co-opt fundamental activities of GAGs to accomplish these tasks. This ingenious strategy to subvert essential activities of GAGs likely prevented host organisms from deleting or inactivating these mechanisms during their evolution. The goal of this review is to provide a mechanistic overview of our current understanding of how microbes subvert GAGs at major steps of pathogenesis, using select GAG-pathogen interactions as representative examples. PMID:27100505

  20. Expression of Glycosaminoglycan Epitopes During Zebrafish Skeletogenesis

    Hayes, Anthony J.; Mitchell, Ruth E; Bashford, Andrew; Reynolds, Scott; Caterson, Bruce; Hammond, Chrissy L

    2013-01-01

    Background: The zebrafish is an important developmental model. Surprisingly, there are few studies that describe the glycosaminoglycan composition of its extracellular matrix during skeletogenesis. Glycosaminoglycans on proteoglycans contribute to the material properties of musculo skeletal connective tissues, and are important in regulating signalling events during morphogenesis. Sulfation motifs within the chain structure of glycosaminoglycans on cell-associated and extracellular matrix pro...

  1. Tissue structure-specific distribution of glycosaminoglycans in the human penis.

    Goulas, A; Papakonstantinou, E; Karakiulakis, G; Mirtsou-Fidani, V; Kalinderis, A; Hatzichristou, D G

    2000-09-01

    The aim of this work was to isolate and characterise the glycosaminoglycans present in the different tissue structures of the human penis in view of their potentially significant role in the physiology of erection. Penile tissue samples were obtained from patients who underwent penectomy and were subsequently dissected into individual tissue structures. Total glycosaminoglycans were isolated and purified from tunica albuginea, corpora cavernosa and corpus spongiosum, following tissue mincing, ultrasonication, lipid extraction, extensive digestion with pronase and DNase, treatment with alkali-borohydride and ethanol precipitation. Isolated glycosaminoglycans were separated by cellulose acetate electrophoresis and fractionated by anion exchange chromatography on DEAE Sephacel columns. Different glycosaminoglycan fractions were identified using glycosaminoglycan-degrading enzymes of known specificity. Gradient polyacrylamide gel electrophoresis was used to determine the average molecular mass of the glycosaminoglycans. The corpus cavernosum and the corpus spongiosum extracts contained almost twice the amount of glycosaminoglycan-associated uronic acids as compared to the tunical extracts (1.47+/-0.09, and 1.49+/-0.15 as opposed to 0.75+/-0.15 microg/mg dry defatted tissue, respectively; S.E.M., n=5). With the exception of hyaluronic acid, the relative amount of individual glycosaminoglycan types varied significantly among extracts of different origin. Heparan sulphate was more abundant in cavernosal, dermatan sulphate in tunical, and chondroitin-6-sulphate in corpus spongiosum extracts. No structure-specific differences were detected with respect to the molecular mass distribution of each glycosaminoglycan type. Our study shows that the different structures of the human penis produce distinct profiles of glycosaminoglycans, which are well suited to the individual functional characteristics of these structures. PMID:11084377

  2. Acid glycosaminoglycan (aGAG) excretion is increased in children with autism spectrum disorder, and it can be controlled by diet.

    Endreffy, Ildikó; Bjørklund, Geir; Dicső, Ferenc; Urbina, Mauricio A; Endreffy, Emőke

    2016-04-01

    Autism research continues to receive considerable attention as the options for successful management are limited. The understanding of the autism spectrum disorder (ASD) etiology has now progressed to encompass genetic, epigenetic, neurological, hormonal, and environmental factors that affect outcomes for patients with ASD. Glycosaminoglycans (GAGs) are a family of linear, sulfated polysaccharides that are associated with central nervous system (CNS) development, maintenance, and disorders. Proteoglycans (PG) regulate diverse functions in the central nervous system. Heparan sulfate (HS) and chondroitin sulfate (CS) are two major GAGs present in the PGs of the CNS. As neuroscience advances, biochemical treatments to correct brain chemistry become better defined. Nutrient therapy can be very potent and has minimal to no side effects, since no molecules foreign to the body are needed. Given GAGs are involved in several neurological functions, and that its level can be somewhat modulated by the diet, the present study aimed to evaluate the role of GAGs levels in ASD symptoms. Both tGAG and its different fractions were evaluated in the urine of ASD and healthy control childrens. As levels differed between groups, a second trial was conduted evaluating if diet could reduce tGAG levels and if this in turn decrease ASD symptoms. The present study found that tGAG concentration was significantly higher in the urine of children with ASD compared to healthy control children and this was also evident in all GAG fractions. Within groups (controls and ASD), no gender differences in GAG excretion were found. The use of a 90 days elimination diet (casein-free, special carbohydrates, multivitamin/mineral supplement), had major effects in reducing urinary tGAG excretion in children with ASD. PMID:26464064

  3. Regulation of sulfated glycosaminoglycan production by prostaglandin E2 in cultured lung fibroblasts

    Karlinsky, J.B.; Goldstein, R.H. (Boston Univ. School of Medicine, MA (USA))

    1989-08-01

    Prostaglandin E2 (PGE2) has been shown to increase the synthesis of hyaluronic acid in cultured fibroblasts by increasing the activity of hyaluronate synthetase, a group of plasma membrane-bound synthetic enzymes. We examined whether PGE2 also increased the activity of those enzyme systems involved in the synthesis of sulfated glycosaminoglycan in the human embryonic lung fibroblast. Exposure of cells to PGE2 resulted in dose-dependent increases in glucosamine incorporation into all sulfated glycosaminoglycan subtypes. PGE2 at 10(-7) mol/L increased total glycosaminoglycan per dish to 21.6 +/- 3.1 micrograms versus 12.0 +/- 2.5 micrograms in control untreated cultures. Stimulation of endogenous PGE2 production by bradykinin had a similar effect on glycosaminoglycan synthesis. To examine whether PGE2 affected sulfated glycosaminoglycan protein core production, cells were labeled with tritiated glucosamine in the presence of cycloheximide. Under these conditions, incorporation of radiolabel into all glycosaminoglycan subtypes was reduced. However, when exogenous sulfated glycosaminoglycan chain initiator (p-nitrophenyl beta-D-xyloside) was added, incorporation of tritiated glucosamine into sulfated glycosaminoglycan increased but not to levels found in control cultures. Application of PGE2 to cultures treated with cycloheximide alone, or to cultures treated with cycloheximide plus xyloside, increased tritiated glucosamine incorporation into chondroitin, dermatan sulfate, and to a lesser extent into heparan sulfate. We conclude that PGE2 stimulates synthesis of all sulfated glycosaminoglycan even in the absence of new protein core production, probably by increasing activities of sulfated glycosaminoglycan synthetase enzymes. PGE2 stimulation of heparan sulfate synthesis is partially dependent on the availability of heparan sulfate-specific protein core.

  4. Regulation of sulfated glycosaminoglycan production by prostaglandin E2 in cultured lung fibroblasts

    Prostaglandin E2 (PGE2) has been shown to increase the synthesis of hyaluronic acid in cultured fibroblasts by increasing the activity of hyaluronate synthetase, a group of plasma membrane-bound synthetic enzymes. We examined whether PGE2 also increased the activity of those enzyme systems involved in the synthesis of sulfated glycosaminoglycan in the human embryonic lung fibroblast. Exposure of cells to PGE2 resulted in dose-dependent increases in glucosamine incorporation into all sulfated glycosaminoglycan subtypes. PGE2 at 10(-7) mol/L increased total glycosaminoglycan per dish to 21.6 +/- 3.1 micrograms versus 12.0 +/- 2.5 micrograms in control untreated cultures. Stimulation of endogenous PGE2 production by bradykinin had a similar effect on glycosaminoglycan synthesis. To examine whether PGE2 affected sulfated glycosaminoglycan protein core production, cells were labeled with tritiated glucosamine in the presence of cycloheximide. Under these conditions, incorporation of radiolabel into all glycosaminoglycan subtypes was reduced. However, when exogenous sulfated glycosaminoglycan chain initiator (p-nitrophenyl beta-D-xyloside) was added, incorporation of tritiated glucosamine into sulfated glycosaminoglycan increased but not to levels found in control cultures. Application of PGE2 to cultures treated with cycloheximide alone, or to cultures treated with cycloheximide plus xyloside, increased tritiated glucosamine incorporation into chondroitin, dermatan sulfate, and to a lesser extent into heparan sulfate. We conclude that PGE2 stimulates synthesis of all sulfated glycosaminoglycan even in the absence of new protein core production, probably by increasing activities of sulfated glycosaminoglycan synthetase enzymes. PGE2 stimulation of heparan sulfate synthesis is partially dependent on the availability of heparan sulfate-specific protein core

  5. The Effects of Mucopolysaccharide Polysulphate on Hydration and Elasticity of Human Skin

    Rungsima Wanitphakdeedecha; Sasima Eimpunth; Woraphong Manuskiatti

    2011-01-01

    Background. Mucopolysaccharide polysulphate (MPS) has been used in medicine as an anti-inflammatory and antithrombotic agent for over 50 years. Its chemical structure permits considerable hydrogen bonding with adjacent water molecules, which effectively leads to hydration of the surrounding tissue. In addition, it stimulates endogenous hyaluronate synthesis, resulting in an increase in water-binding capacity and viscoelasticity of the skin. Objective. To study the efficacy of 0.1% MPS on hydr...

  6. New targets for glycosaminoglycans and glycosaminoglycans as novel targets.

    Gesslbauer, Bernd; Theuer, Martina; Schweiger, Daniela; Adage, Tiziana; Kungl, Andreas J

    2013-02-01

    Biological functions of a variety of proteins are mediated via their interaction with glycosaminoglycans (GAGs). The structural diversity within the wide GAG landscape provides individual interaction sites for a multitude of proteins involved in several pathophysiological processes. This 'GAG angle' of such proteins as well as their specific GAG ligands give rise to novel therapeutic concepts for drug development. Current glycomic technologies to elucidate the glycan structure-function relationships, methods to investigate the selectivity and specificity of glycan-protein interactions and existing therapeutic approaches to interfere with GAG-protein interactions are discussed. PMID:23414361

  7. GLYCOSAMINOGLYCANS AND PROTEOGLYCANS IN PALMAR FASCIA OF PATIENTS WITH DUPUYTREN

    Nascimento, Priscilla Carneiro Hirai; Kobayashi, Elsa Yoko; LENZI, Luiz Guilherme de Saboya; dos Santos, João Baptista Gomes; Nader, Helena Bonciani; Faloppa, Flávio

    2016-01-01

    Objective : To evaluate and compare the behavior of glycosaminoglycans (GAGs) in Dupuytren disease (DD). Methods : This is an experimental study with 23 patients diagnosed with DD. Tissue collected through fasciectomy with incision type Brunner or McCash were evaluated by electrophoresis for identification of GAGs. The quantification was carried out by immunofluorescence and dosage of proteins for different types of glycosaminoglycans. The results were expressed in percentage and statistically evaluated. Results : A significant increase was observed through eletrophoresis in GAGs, as compared to the control (p<0.05). Immunofluorescence of hyaluronic acid was reduced (23 times) when compared to the control (p<0.0001). Conclusion : An increase of sulfated GAGs in Dupuytren's disease, mainly dermatan sulfate, was evident from our results, as well as a pronounced decrease of hyaluronic acid in the palmar aponeurosis from the same patients. Level of Evidence III, Case-Control Study. PMID:26981045

  8. Identification and synthesis of a recognition signal for the attachment of glycosaminoglycans to proteins

    Comparison of the amino acid sequences of three different proteoglycan core proteins reveals a 12-amino acid sequence that is about 50% homologous among these proteoglycans. In each of the proteoglycans, this sequence surrounds the serine-glycine dipeptide in which the serine is known or presumed to be substituted with a chondroitin/dermatan sulfate glycosaminoglycan chain. Peptides containing this sequence from two proteoglycans were examined for their ability to serve as acceptors for xylosyltransferase, the enzyme that begins the assembly of glycosaminoglycan chains. Those peptides corresponding to amino acid sequences known to contain glycosaminoglycan-substituted serine residues in the protein were efficient xylosyltransferase acceptors, whereas peptides from sequences with no glycosaminoglycan-substituted serine residues were not. Amino acid substitutions at four critical sites in the acceptor peptides showed that single substitutions could completely abolish acceptor activity or greatly reduce it. The results suggest that the proteoglycan recognition consensus sequence for the attachment of glycosaminoglycans to core proteins consists of acid amino acids closely followed by the tetrapeptide Ser-Gly-Xaa-Gly, where Xaa is any amino acid. The signal appears to be contained in the primary sequence information. In this regard it resembles a number of other signals for protein processing and intracellular routing

  9. Identification and synthesis of a recognition signal for the attachment of glycosaminoglycans to proteins

    Bourdon, M.A.; Krusius, T.; Campbell, S.; Schwartz, N.B.; Ruoslahti, E.

    1987-05-01

    Comparison of the amino acid sequences of three different proteoglycan core proteins reveals a 12-amino acid sequence that is about 50% homologous among these proteoglycans. In each of the proteoglycans, this sequence surrounds the serine-glycine dipeptide in which the serine is known or presumed to be substituted with a chondroitin/dermatan sulfate glycosaminoglycan chain. Peptides containing this sequence from two proteoglycans were examined for their ability to serve as acceptors for xylosyltransferase, the enzyme that begins the assembly of glycosaminoglycan chains. Those peptides corresponding to amino acid sequences known to contain glycosaminoglycan-substituted serine residues in the protein were efficient xylosyltransferase acceptors, whereas peptides from sequences with no glycosaminoglycan-substituted serine residues were not. Amino acid substitutions at four critical sites in the acceptor peptides showed that single substitutions could completely abolish acceptor activity or greatly reduce it. The results suggest that the proteoglycan recognition consensus sequence for the attachment of glycosaminoglycans to core proteins consists of acid amino acids closely followed by the tetrapeptide Ser-Gly-Xaa-Gly, where Xaa is any amino acid. The signal appears to be contained in the primary sequence information. In this regard it resembles a number of other signals for protein processing and intracellular routing.

  10. Proteinase activity regulation by glycosaminoglycans

    Tersariol I.L.S.

    2002-01-01

    Full Text Available There are few reports concerning the biological role and the mechanisms of interaction between proteinases and carbohydrates other than those involved in clotting. It has been shown that the interplay of enzymes and glycosaminoglycans is able to modulate the activity of different proteases and also to affect their structures. From the large number of proteases belonging to the well-known protease families and also the variety of carbohydrates described as widely distributed, only few events have been analyzed more deeply. The term "family" is used to describe a group of proteases in which every member shows an evolutionary relationship to at least one other protease. This relationship may be evident throughout the entire sequence, or at least in that part of the sequence responsible for catalytic activity. The majority of proteases belong to the serine, cysteine, aspartic or metalloprotease families. By considering the existing limited proteolysis process, in addition to the initial idea that the proteinases participate only in digestive processes, it is possible to conclude that the function of the enzymes is strictly limited to the cleavage of intended substrates since the destruction of functional proteins would result in normal tissue damage. In addition, the location as well as the eventual regulation of protease activity promoted by glycosaminoglycans can play an essential role in the development of several physiopathological conditions.

  11. Hydrolyzable Tannins (Chebulagic Acid and Punicalagin) Target Viral Glycoprotein-Glycosaminoglycan Interactions To Inhibit Herpes Simplex Virus 1 Entry and Cell-to-Cell Spread▿

    Lin, Liang-Tzung; Chen, Ting-Ying; Chung, Chueh-Yao; Noyce, Ryan S.; Grindley, T. Bruce; McCormick, Craig; Lin, Ta-Chen; Wang, Guey-Horng; Lin, Chun-Ching; Richardson, Christopher D.

    2011-01-01

    Herpes simplex virus 1 (HSV-1) is a common human pathogen that causes lifelong latent infection of sensory neurons. Non-nucleoside inhibitors that can limit HSV-1 recurrence are particularly useful in treating immunocompromised individuals or cases of emerging acyclovir-resistant strains of herpesvirus. We report that chebulagic acid (CHLA) and punicalagin (PUG), two hydrolyzable tannins isolated from the dried fruits of Terminalia chebula Retz. (Combretaceae), inhibit HSV-1 entry at noncytot...

  12. Validation of Urinary Glycosaminoglycans in Iranian patients with Mucopolysaccharidase type I: The effect of urine sedimentation characteristics

    Mohammad ABDI

    2014-12-01

    physiological processes. Physiological reviews. 1991 Apr;71(2:481-539.Ghaderi S. The biochemistry base of mucopolysaccharidoses and approach to. Genetics in the 3rd millennium. [Educational]. 2006;4(1:711-22.Mizumoto S, Ikegawa S, Sugahara K. Human genetic disorders caused by mutations in genes encoding biosynthetic enzymes for sulfated glycosaminoglycans. The Journal of biological chemistry. 2013 Apr 19;288(16:10953-61.Salbach J, Rachner TD, Rauner M, Hempel U, Anderegg U, Franz S, et al. Regenerative potential of glycosaminoglycans for skin and bone. Journal of molecular medicine (Berlin, Germany. 2012 Jun;90(6:625-35.Coppa GV, Catassi C, Gabrielli O, Giorgi PL, Dall’Amico R, Naia S, et al. Clinical application of a new simple method for the identification of mucopolysaccharidoses. Helvetica paediatrica acta. 1987 Jun;42(5-6:419-23.Fuller M, Meikle PJ, Hopwood JJ. Glycosaminoglycan degradation fragments in mucopolysaccharidosis I. Glycobiology. 2004 May;14(5:443-50.Fuller M, Rozaklis T, Ramsay SL, Hopwood JJ, Meikle PJ. Disease-specific markers for the mucopolysaccharidoses. Pediatric research. 2004 Nov;56(5:733-8.Blau N, Duran M, Gibson K. Laboratory Guide to the Methods in Biochemical Genetics. First edition ed: Springer-Verlag Berlin Heidelberg; 2008. pp287-324.Dorfman A, Matalon R. The Hurler and Hunter syndromes. The American journal of medicine. 1969 Nov;47(5:691-707.Fratantoni JC, Hall CW, Neufeld EF. Hurler and Hunter syndromes: mutual correction of the defect in cultured fibroblasts. Science (New York, NY. 1968 Nov 1;162(3853:570-2.Fratantoni JC, Hall CW, Neufeld EF. The defect in Hurler and Hunter syndromes. II. Deficiency of specific factors involved in mucopolysaccharide degradation. Proceedings of the National Academy of Sciences of the United States of America. 1969 Sep;64(1:360-6.Fratantoni JC, Neufeld EF, Uhlendorf BW, Jacobson CB. Intrauterine diagnosis of the hurler and hunter syndromes. The New England journal of medicine. 1969 Mar 27

  13. Dietary carotenoid pigment supplementation influences hepatic lipid and mucopolysaccharide levels in rainbow trout (Oncorhynchus mykiss).

    Page, G I; Russell, P M; Davies, S J

    2005-12-01

    We assessed the effects of dietary carotenoid pigment supplementation on liver histochemistry in the rainbow trout. One hundred and eight rainbow trout (mean mass 266+/-10 g) were assigned to each of three replicate tanks for each of three dietary treatments; astaxanthin, canthaxanthin, or control at a target dietary inclusion of 100 mg/kg, by top-coating a pigment-free commercially extruded basal diet (Trouw Aquaculture, U.K.). Fish were fed for 3 weeks at a ration of 1.2% body mass/day, in a recirculating freshwater system maintained at 16 degrees C. Frozen liver sections were stained for total lipids, unsaturated lipids, glycogen, mucopolysaccharides, glycogen phosphorylase and aspartate aminotransferase. Relative amounts were measured quantitatively by image analysis. Carotenoid treatment significantly (P<0.05) altered the total lipid profile and hepatic mucopolysaccharide contents of livers of rainbow trout. Results are discussed in relation to the catabolic potential of the liver in carotenoid pigment metabolism. PMID:16209931

  14. Determinants of Glycosaminoglycan (GAG Structure

    Kristian Prydz

    2015-08-01

    Full Text Available Proteoglycans (PGs are glycosylated proteins of biological importance at cell surfaces, in the extracellular matrix, and in the circulation. PGs are produced and modified by glycosaminoglycan (GAG chains in the secretory pathway of animal cells. The most common GAG attachment site is a serine residue followed by a glycine (-ser-gly-, from which a linker tetrasaccharide extends and may continue as a heparan sulfate, a heparin, a chondroitin sulfate, or a dermatan sulfate GAG chain. Which type of GAG chain becomes attached to the linker tetrasaccharide is influenced by the structure of the protein core, modifications occurring to the linker tetrasaccharide itself, and the biochemical environment of the Golgi apparatus, where GAG polymerization and modification by sulfation and epimerization take place. The same cell type may produce different GAG chains that vary, depending on the extent of epimerization and sulfation. However, it is not known to what extent these differences are caused by compartmental segregation of protein cores en route through the secretory pathway or by differential recruitment of modifying enzymes during synthesis of different PGs. The topic of this review is how different aspects of protein structure, cellular biochemistry, and compartmentalization may influence GAG synthesis.

  15. Determinants of Glycosaminoglycan (GAG) Structure.

    Prydz, Kristian

    2015-01-01

    Proteoglycans (PGs) are glycosylated proteins of biological importance at cell surfaces, in the extracellular matrix, and in the circulation. PGs are produced and modified by glycosaminoglycan (GAG) chains in the secretory pathway of animal cells. The most common GAG attachment site is a serine residue followed by a glycine (-ser-gly-), from which a linker tetrasaccharide extends and may continue as a heparan sulfate, a heparin, a chondroitin sulfate, or a dermatan sulfate GAG chain. Which type of GAG chain becomes attached to the linker tetrasaccharide is influenced by the structure of the protein core, modifications occurring to the linker tetrasaccharide itself, and the biochemical environment of the Golgi apparatus, where GAG polymerization and modification by sulfation and epimerization take place. The same cell type may produce different GAG chains that vary, depending on the extent of epimerization and sulfation. However, it is not known to what extent these differences are caused by compartmental segregation of protein cores en route through the secretory pathway or by differential recruitment of modifying enzymes during synthesis of different PGs. The topic of this review is how different aspects of protein structure, cellular biochemistry, and compartmentalization may influence GAG synthesis. PMID:26308067

  16. Purification and partial characterization of glycosaminoglycans and proteoglycans from cultured rabbit smooth muscle cells

    Sabatino, R.D.

    1985-01-01

    Glycosaminoglycans synthesized by cultured rabbit smooth muscle cells were isolated after incorporation of (/sup 3/H)-glucosamine into glycosaminoglycans in the presence or absence of 10% fetal bovine serum. Glycosaminoglycans were quantitated by two-dimensional electrophoresis after proteolytic digestion of the cell layers and media. The results show that the presence of serum has no effect on the chondroitin sulfate, heparan sulfate and dermatan sulfate content of the cell layers. The incorporation of (/sup 3/H)-glucosamine into hyaluronic acid of the cell layers was three times higher in the presence of serum. In the medium , the quantity of hyaluronic was two times higher in the presence of serum while the other glycosaminoglycans remained unchanged. The incorporation of (/sup 3/H)-glucosamine into hyaluronic acid was unaffected by the presence of serum. Specific proteoglycans were isolated from medium after with (/sup 35/S)-sulfate and (/sup 3/H)-serine by isopycnic ultracentrifugation and chromatography on Sepharose CL-4B and DEAE-cellulose. Preparations contained a chondroitin sulfate proteoglycan, a condroitin sulfate-dermatan sulfate proteoglycan and a heparan sulfate proteoglycan. Glycosaminoglycans and proteoglycans synthesized by rabbit aorta smooth muscle cells are similar to those from human aorta.

  17. Glycosaminoglycan distribution in the rat uterine cervix during the estrous cycle

    Jairo Jose Matozinho Cubas

    2010-01-01

    Full Text Available OBJECTIVE: To analyze the amount of glycosaminoglycans in the uterine cervix during each phase of the rat estrous cycle. DESIGN: Based on vaginal smears, forty female, regularly cycling rats were divided into four groups (n = 10 for each group: GI - proestrous, GII - estrous, GIII - metaestrous and GIV - diestrous. Animals were sacrificed at each phase of the cycle, and the cervix was immediately removed and submitted to biochemical extraction and determination of sulfated glycosaminoglycans and hyaluronic acid. The results were analyzed by ANOVA followed by the Bonferroni post-hoc test. RESULTS: The uterine cervix had the highest amount of total sulfated glycosaminoglycans and dermatan sulfate during the estrous phase (8.90 ± 0.55 mg/g of cetonic extract, p<0.001; and 8.86 ± 0.57 mg/g of cetonic extract, p<0.001. In addition, there was more heparan sulfate at the cervix during the proestrous phase (0.185 ± 0.03 mg/g of cetonic extract than during any other phase (p<0.001. There were no significant changes in the concentration of hyaluronic acid in the uterine cervix during the estrous cycle. CONCLUSION: Our data suggest that the amount of total sulfated glycosaminoglycans may be influenced by hormonal fluctuations related to the estrous cycle, with dermatan sulfate and heparan sulfate being the glycosaminoglycans most sensitive to hormonal change.

  18. Purification and partial characterization of glycosaminoglycans and proteoglycans from cultured rabbit smooth muscle cells

    Glycosaminoglycans synthesized by cultured rabbit smooth muscle cells were isolated after incorporation of [3H]-glucosamine into glycosaminoglycans in the presence or absence of 10% fetal bovine serum. Glycosaminoglycans were quantitated by two-dimensional electrophoresis after proteolytic digestion of the cell layers and media. The results show that the presence of serum has no effect on the chondroitin sulfate, heparan sulfate and dermatan sulfate content of the cell layers. The incorporation of [3H]-glucosamine into hyaluronic acid of the cell layers was three times higher in the presence of serum. In the medium , the quantity of hyaluronic was two times higher in the presence of serum while the other glycosaminoglycans remained unchanged. The incorporation of [3H]-glucosamine into hyaluronic acid was unaffected by the presence of serum. Specific proteoglycans were isolated from medium after with [35S]-sulfate and [3H]-serine by isopycnic ultracentrifugation and chromatography on Sepharose CL-4B and DEAE-cellulose. Preparations contained a chondroitin sulfate proteoglycan, a condroitin sulfate-dermatan sulfate proteoglycan and a heparan sulfate proteoglycan. Glycosaminoglycans and proteoglycans synthesized by rabbit aorta smooth muscle cells are similar to those from human aorta

  19. Structure and anticoagulant properties of sulfated glycosaminoglycans from primitive Chordates

    PAVÃO MAURO S. G.

    2002-01-01

    Full Text Available Dermatan sulfates and heparin, similar to the mammalian glycosaminoglycans, but with differences in the degree and position of sulfation were previously isolated from the body of the ascidian Styela plicata and Ascidia nigra. These differences produce profound effects on their anticoagulant properties. S. plicata dermatan sulfate composed by 2-O-sulfatedalpha-L-iduronic acid and 4-O-sulfated N-acetyl-beta-D-galactosamine residues is a potent anticoagulant due to a high heparin cofactor II activity. Surprisingly, it has a lower potency to prevent thrombus formation on an experimental model and a lower bleeding effect in rats than the mammalian dermatan sulfate. In contrast, A. nigra dermatan sulfate, also enriched in 2-O-sulfated alpha-L-iduronic acid, but in this case sulfated at O-6 of the N-acetyl-beta-D-galactosamine units, has no in vitro or in vivo anticoagulant activity, does not prevent thrombus formation but shows a bleeding effect similar to the mammalian glycosaminoglycan. Ascidian heparin, composed by 2-O-sulfated alpha-L-iduronic acid, N- and 6-O-sulfated glucosamine (75% and alpha-L-iduronic acid, N- and 6-O-sulfated glucosamine (25% disaccharide units has an anticoagulant activity 10 times lower than the mammalian heparin, is about 20 times less potent in the inhibition of thrombin by antithrombin, but has the same heparin cofactor II activity as mammalian heparin.

  20. Urinary glycosaminoglycans excretion and the effect of dimethyl sulfoxide in an experimental model of non-bacterial cystitis

    Roberto Soler; Homero Bruschini; Jose C. Truzzi; Joao R. Martins; Niels O. Camara; Maria T. Alves; Katia R. Leite; Nader, Helena B.; Miguel Srougi; Valdemar Ortiz

    2008-01-01

    PURPOSE: We reproduced a non-bacterial experimental model to assess bladder inflammation and urinary glycosaminoglycans (GAG) excretion and examined the effect of dimethyl sulfoxide (DMSO). MATERIALS AND METHODS: Female rats were instilled with either protamine sulfate (PS groups) or sterile saline (control groups). At different days after the procedure, 24 h urine and bladder samples were obtained. Urinary levels of hyaluronic acid (HA) and sulfated glycosaminoglycans (S-GAG) were determined...

  1. Glycosaminoglycan-lipoprotein complexes from aortas of hypercholesterolemic rabbits. Part 1. Isolation and characterization.

    Mawhinney, T P; Augustyn, J M; Fritz, K E

    1978-10-01

    Glycosaminoglycan-lipoprotein complexes were isolated from rabbit aortas exhibiting nearly confluent cholesterol-induced foam cell lesions by extraction with 0.15 M NaCl. Purification and characterization was achieved by gel chromatography, non-ionic differential flotation and by cellulose polyacetate electrophoresis. Analysis showed that these complexes consisted of very low density lipoproteins, heparan sulfate, chondroitin sulfate-C and hyaluronic acid. The demonstration that rabbit intimal foam cell lesions contain extractable glycosaminoglycan-lipoprotein complexes makes this animal model an excellent tool for further studies on the role of these complexes in the atherogenic process. PMID:215171

  2. Plasmodium vivax adherence to placental glycosaminoglycans.

    Kesinee Chotivanich

    Full Text Available BACKGROUND: Plasmodium vivax infections seldom kill directly but do cause indirect mortality by reducing birth weight and causing abortion. Cytoadherence and sequestration in the microvasculature are central to the pathogenesis of severe Plasmodium falciparum malaria, but the contribution of cytoadherence to pathology in other human malarias is less clear. METHODOLOGY: The adherence properties of P. vivax infected red blood cells (PvIRBC were evaluated under static and flow conditions. PRINCIPAL FINDINGS: P. vivax isolates from 33 patients were studied. None adhered to immobilized CD36, ICAM-1, or thrombospondin, putative ligands for P. falciparum vascular cytoadherence, or umbilical vein endothelial cells, but all adhered to immobilized chondroitin sulphate A (CSA and hyaluronic acid (HA, the receptors for adhesion of P. falciparum in the placenta. PvIRBC also adhered to fresh placental cells (N = 5. Pre-incubation with chondroitinase prevented PvIRBC adherence to CSA, and reduced binding to HA, whereas preincubation with hyaluronidase prevented adherence to HA, but did not reduce binding to CSA significantly. Pre-incubation of PvIRBC with soluble CSA and HA reduced binding to the immobilized receptors and prevented placental binding. PvIRBC adhesion was prevented by pre-incubation with trypsin, inhibited by heparin, and reduced by EGTA. Under laminar flow conditions the mean (SD shear stress reducing maximum attachment by 50% was 0.06 (0.02 Pa but, having adhered, the PvIRBC could then resist detachment by stresses up to 5 Pa. At 37 °C adherence began approximately 16 hours after red cell invasion with maximal adherence at 30 hours. At 39 °C adherence began earlier and peaked at 24 hours. SIGNIFICANCE: Adherence of P. vivax-infected erythrocytes to glycosaminoglycans may contribute to the pathogenesis of vivax malaria and lead to intrauterine growth retardation.

  3. Study on the Fermentation Conditions of a Mucopolysaccharide-producing Bacterium LV-1%一株粘性多糖产生菌LV-1的发酵条件研究

    梁玉丽; 郭继强; 陈晓艺; 刘志文; 李宪臻

    2009-01-01

    [Objective] The fermentation conditions of a mucopolysaccharide-producing bacterium LV-1 which isolated from soil sample were studied.[Method] The polysaccharide-producing bacterium was isolated by serial dilution method, the effects of carbon source, nitrogen source, the initial pH and temperature on producing polysaccharide by it were discussed to confirm the optimum fermentation conditions.[Result] The physicochemical properties showed that the polysaccharide was water-soluble, but insoluble in organic solvents including ethanol, butanol, and chloroform.It was neutral polysaccharide with negative charge and without reducing terminal. The pH of its solution was pH=7.5. There were no protein, fructose, uronic acid, sulphate and starch-like structure included in polysaccharide molecules. The optimum fermentation conditions for polysaccharide production were 3% mannitol as carbon source, 0.25% yeast extract as nitrogen source, culture temperature 28 ℃ and pH=7.5. [Conclusion] The research could provie basis for development and utilization of LV-1 and industrialized production of mucopolysaccharide.

  4. Synthesis of glycosaminoglycans by islets of Langerhans

    Watkins, D.T.; Cooperstein, S.J.

    1988-01-01

    Incorporation of (/sup 35/S)-sulfate into glycosaminoglycans (GAG) of toadfish islets of Langerhans in vitro was examined. (/sup 35/S)-sulfated GAG were synthesized by a component of the microsomal fraction, and subsequently transferred to the secretion granules, mitochondria and nuclei. The predominant type of GAG synthesized was heparan sulfate, but chondroitin 4- and 6-sulfate and dermatan sulfate were also found. 36 references, 3 tables.

  5. Synthesis of glycosaminoglycans by islets of Langerhans

    Incorporation of (35S)-sulfate into glycosaminoglycans (GAG) of toadfish islets of Langerhans in vitro was examined. (35S)-sulfated GAG were synthesized by a component of the microsomal fraction, and subsequently transferred to the secretion granules, mitochondria and nuclei. The predominant type of GAG synthesized was heparan sulfate, but chondroitin 4- and 6-sulfate and dermatan sulfate were also found. 36 references, 3 tables

  6. Accumulation of glycosaminoglycans in radiation-induced muscular fibrosis

    The content and biosynthesis of glycosaminoglycans (GAGs) were studied in pig thigh muscle after acute local γ-irradiation. Seven months following irradiation, the muscular tissue next to the irradiation cone was replaced by severe mutilating fibrosis delimited by an intermediary perifibrotic zone. Results showed a parallel increase of collagen and GAG content in perifibrotic and fibrotic tissues. Sulphated GAGs, heparan sulphate and dermatan sulphate were preferentially accumulated in fibrotic tissue, while the hyaluronic acid content increased only slightly. Synthesis of sulphated GAGs was more elevated in fibrotic tissue than in perifibrotic zone as compared with normal muscle. Seven months after irradiation well-developed fibrotic tissue continued to synthesize and to accumulate extracellular matrix macromolecules. (Author)

  7. Extraction and quantification of sulfated glycosaminoglycan content in five different aquatic species of Malaysia

    Ravi Lokwani; Ramandeep Singh; Gauree Kukreti

    2015-01-01

    Objective: To extract, characterize and quantify glycosaminoglycans (GAGs) from the body of cuttlefish, tennis-ball sea cucumber, shrimp, seabass and fresh water fish Nile tilapia. Methods: The extracted crude powder was evaluated for the content of GAGs. The qualitative analysis of sulfated pattern and other important functional groups related with GAGs were explained in the form of Fourier transform infra-red spectroscopy data. Proteins and nucleic acid in the crude extr...

  8. Glycosaminoglycans that bind cold-insoluble globulin in cell-substratum adhesion sites of murine fibroblasts.

    Laterra, J; Ansbacher, R; Culp, L A

    1980-01-01

    Glycosaminoglycans (GAGs) and glycoprotein-derived glycopeptide from mouse BALB/c3T3 and simian virus 40-transformed 3T3 whole cells or their adhesion sites, which are left bound to the serum-coated tissue culture substratum after detachment of cells mediated by [ethylenebis-(oxyethylenenitrilo]tetraacetic acid (EGTA), were analyzed for specific binding to Sepharose columns derivatized with cold-insoluble globulin (CIg). CIg is the serum-contained form of fibronectin and is required for the a...

  9. The effects of mucopolysaccharide polysulphate on hydration and elasticity of human skin.

    Wanitphakdeedecha, Rungsima; Eimpunth, Sasima; Manuskiatti, Woraphong

    2011-01-01

    Background. Mucopolysaccharide polysulphate (MPS) has been used in medicine as an anti-inflammatory and antithrombotic agent for over 50 years. Its chemical structure permits considerable hydrogen bonding with adjacent water molecules, which effectively leads to hydration of the surrounding tissue. In addition, it stimulates endogenous hyaluronate synthesis, resulting in an increase in water-binding capacity and viscoelasticity of the skin. Objective. To study the efficacy of 0.1% MPS on hydration and elasticity of human skin. Methods. The first part of this study was a randomized double blind placebo-controlled study which included 60 female volunteers aged 30-45 years with dry skin, defined by Corneometer CM 825. The volunteers were treated with either 0.1% MPS or vehicle control. All subjects were asked to apply 1 g of cream to their face twice daily for a total period of 4 weeks. Skin hydration and elasticity were measured at baseline and week 4 with Corneometer CM 825 and cutometer MPA 580, respectively, at forehead and both cheeks. The second part of this study focused on the efficacy of 0.1% MPS on skin hydration after single application. 20 female volunteers aged 30-45 years with dry skin, defined by Corneometer CM 825, were recruited to the study. All subjects were asked to apply 2 g of 0.1% MPS cream on entirely randomly selected forearm. Skin hydration at the middle of both forearms was measured at baseline, immediately after application, and every 1 hour after application for a period of 10 hours. Results. 57 subjects (28 in vehicle control group, 29 in MPS) completed treatment protocol. The baseline skin hydration of both groups was not significantly different (P = 0.47). Hower, there was a statistically significant difference in skin hydration at 4 weeks between MPS and placebo group (P = 0.01). Skin elasticity was significantly improved at week 4 in both groups (vehicle-control, P < 0.01, and MPS, P < 0.01). However, no significant

  10. The Effects of Mucopolysaccharide Polysulphate on Hydration and Elasticity of Human Skin

    Rungsima Wanitphakdeedecha

    2011-01-01

    Full Text Available Background. Mucopolysaccharide polysulphate (MPS has been used in medicine as an anti-inflammatory and antithrombotic agent for over 50 years. Its chemical structure permits considerable hydrogen bonding with adjacent water molecules, which effectively leads to hydration of the surrounding tissue. In addition, it stimulates endogenous hyaluronate synthesis, resulting in an increase in water-binding capacity and viscoelasticity of the skin. Objective. To study the efficacy of 0.1% MPS on hydration and elasticity of human skin. Methods. The first part of this study was a randomized double blind placebo-controlled study which included 60 female volunteers aged 30–45 years with dry skin, defined by Corneometer CM 825. The volunteers were treated with either 0.1% MPS or vehicle control. All subjects were asked to apply 1 g of cream to their face twice daily for a total period of 4 weeks. Skin hydration and elasticity were measured at baseline and week 4 with Corneometer CM 825 and cutometer MPA 580, respectively, at forehead and both cheeks. The second part of this study focused on the efficacy of 0.1% MPS on skin hydration after single application. 20 female volunteers aged 30–45 years with dry skin, defined by Corneometer CM 825, were recruited to the study. All subjects were asked to apply 2 g of 0.1% MPS cream on entirely randomly selected forearm. Skin hydration at the middle of both forearms was measured at baseline, immediately after application, and every 1 hour after application for a period of 10 hours. Results. 57 subjects (28 in vehicle control group, 29 in MPS completed treatment protocol. The baseline skin hydration of both groups was not significantly different (P=0.47. Hower, there was a statistically significant difference in skin hydration at 4 weeks between MPS and placebo group (P=0.01. Skin elasticity was significantly improved at week 4 in both groups (vehicle-control, P<0.01, and MPS, P<0.01. However, no

  11. Development of an enzyme-linked immunosorbent assay (ELISA)-like fluorescence assay to investigate the interactions of glycosaminoglycans to cells

    Boucas, Rodrigo Ippolito [Disciplina de Biologia Molecular, Departamento de Bioquimica, Universidade Federal de Sao Paulo, SP (Brazil); Trindade, Edvaldo S. [Disciplina de Biologia Molecular, Departamento de Bioquimica, Universidade Federal de Sao Paulo, SP (Brazil); Departamento de Biologia Celular, Universidade Federal do Parana, Curitiba, Parana (Brazil); Tersariol, Ivarne L.S. [Disciplina de Biologia Molecular, Departamento de Bioquimica, Universidade Federal de Sao Paulo, SP (Brazil); Centro Interdisciplinar de Investigacao Bioquimica, Universidade de Mogi das Cruzes, Mogi das Cruzes, SP (Brazil); Dietrich, Carl P. [Disciplina de Biologia Molecular, Departamento de Bioquimica, Universidade Federal de Sao Paulo, SP (Brazil); Nader, Helena B. [Disciplina de Biologia Molecular, Departamento de Bioquimica, Universidade Federal de Sao Paulo, SP (Brazil)], E-mail: hbnader.bioq@epm.br

    2008-06-23

    Sulfated glycosaminoglycans were labeled with biotin to study their interaction with cells in culture. Thus, heparin, heparan sulfate, chondroitin 4-sulfate, chondroitin 6-sulfate and dermatan sulfate were labeled using biotin-hydrazide, under different conditions. The structural characteristics of the biotinylated products were determined by chemical (molar ratios of hexosamine, uronic acid, sulfate and biotin) and enzymatic methods (susceptibility to degradation by chondroitinases and heparitinases). The binding of biotinylated glycosaminoglycans was investigated both in endothelial and smooth muscle cells in culture, using a novel time resolved fluorometric method based on interaction of europium-labeled streptavidin with the biotin covalently linked to the compounds. The interactions of glycosaminoglycans were saturable and number of binding sites could be obtained for each individual compound. The apparent dissociation constant varied among the different glycosaminoglycans and between the two cell lines. The interactions of the biotinylated glycosaminoglycans with the cells were also evaluated using confocal microscopy. We propose a convenient and reliable method for the preparation of biotinylated glycosaminoglycans, as well as a sensitive non-competitive fluorescence-based assay for studies of the interactions and binding of these compounds to cells in culture.

  12. Development of an enzyme-linked immunosorbent assay (ELISA)-like fluorescence assay to investigate the interactions of glycosaminoglycans to cells

    Sulfated glycosaminoglycans were labeled with biotin to study their interaction with cells in culture. Thus, heparin, heparan sulfate, chondroitin 4-sulfate, chondroitin 6-sulfate and dermatan sulfate were labeled using biotin-hydrazide, under different conditions. The structural characteristics of the biotinylated products were determined by chemical (molar ratios of hexosamine, uronic acid, sulfate and biotin) and enzymatic methods (susceptibility to degradation by chondroitinases and heparitinases). The binding of biotinylated glycosaminoglycans was investigated both in endothelial and smooth muscle cells in culture, using a novel time resolved fluorometric method based on interaction of europium-labeled streptavidin with the biotin covalently linked to the compounds. The interactions of glycosaminoglycans were saturable and number of binding sites could be obtained for each individual compound. The apparent dissociation constant varied among the different glycosaminoglycans and between the two cell lines. The interactions of the biotinylated glycosaminoglycans with the cells were also evaluated using confocal microscopy. We propose a convenient and reliable method for the preparation of biotinylated glycosaminoglycans, as well as a sensitive non-competitive fluorescence-based assay for studies of the interactions and binding of these compounds to cells in culture

  13. Sulfated glycosaminoglycans in cultured endothelial cells from capillaries and large vessels of human and bovine origin

    The (35S)glycosaminoglycans ((35S)GAG) synthesized by capillary endothelial cells were analyzed and compared to GAG synthesized by endothelial cells cultured from 4 larger vessels. Two separate cultures of endothelial cells were established from bovine fat capillaries and from 4 larger vessels of human origin (umbilical vein) and bovine origin (pulmonary artery, pulmonary vein and aorta). After incubation with 35SO4 for 72 h, the (35S)glycosaminoglycans (GAG) composition of the media, pericellular and cellular fractions of each culture were determined by selective degradation with nitrous acid, chondroitinase ABC and chondroitinase AC. All endothelial cells produced large amounts of (35S)GAG with increased proportions of heparinoids (heparan sulfate and heparin) in the cellular and pericellular fractions. Each culture showed a distinct distribution of (35S)GAG in the media, pericellular and cellular fractions with several specific differences found among the 5 cultures. The differences in GAG content were confirmed in a second group of separate cultures from each of the 5 vessels indicating that, although having several features of GAG metabolism in common, each endothelial cell culture demonstrated a characteristic complement of synthesized, secreted and cell surface-sulfated glycosaminoglycans. (author)

  14. Biosynthesis of glycosaminoglycans: associated disorders and biochemical tests.

    Sasarman, Florin; Maftei, Catalina; Campeau, Philippe M; Brunel-Guitton, Catherine; Mitchell, Grant A; Allard, Pierre

    2016-03-01

    Glycosaminoglycans (GAG) are long, unbranched heteropolymers with repeating disaccharide units that make up the carbohydrate moiety of proteoglycans. Six distinct classes of GAGs are recognized. Their synthesis follows one of three biosynthetic pathways, depending on the type of oligosaccharide linker they contain. Chondroitin sulfate, dermatan sulfate, heparan sulfate, and heparin sulfate contain a common tetrasaccharide linker that is O-linked to specific serine residues in core proteins. Keratan sulfate can contain three different linkers, either N-linked to asparagine or O-linked to serine/threonine residues in core proteins. Finally, hyaluronic acid does not contain a linker and is not covalently attached to a core protein. Most inborn errors of GAG biosynthesis are reported in small numbers of patients. To date, in 20 diseases, convincing evidence for pathogenicity has been presented for mutations in a total of 16 genes encoding glycosyltransferases, sulfotransferases, epimerases or transporters. GAG synthesis defects should be suspected in patients with a combination of characteristic clinical features in more than one connective tissue compartment: bone and cartilage (short long bones with or without scoliosis), ligaments (joint laxity/dislocations), and subepithelial (skin, sclerae). Some produce distinct clinical syndromes. The commonest laboratory tests used for this group of diseases are analysis of GAGs, enzyme assays, and molecular testing. In principle, GAG analysis has potential as a general first-line diagnostic test for GAG biosynthesis disorders. PMID:26689402

  15. Establishment of Glycosaminoglycan Assays for Mucopolysaccharidoses

    Shunji Tomatsu

    2014-08-01

    Full Text Available Mucopolysaccharidoses (MPS are a group of lysosomal storage disorders caused by deficiency of the lysosomal enzymes essential for catabolism of glycosaminoglycans (GAGs. Accumulation of undegraded GAGs results in dysfunction of multiple organs, resulting in distinct clinical manifestations. A range of methods have been developed to measure specific GAGs in various human samples to investigate diagnosis, prognosis, pathogenesis, GAG interaction with other molecules, and monitoring therapeutic efficacy. We established ELISA, liquid chromatography tandem mass spectrometry (LC-MS/MS, and an automated high-throughput mass spectrometry (HT-MS/MS system (RapidFire to identify epitopes (ELISA or disaccharides (MS/MS derived from different GAGs (dermatan sulfate, heparan sulfate, keratan sulfate, and/or chondroitin sulfate. These methods have a high sensitivity and specificity in GAG analysis, applicable to the analysis of blood, urine, tissues, and cells. ELISA is feasible, sensitive, and reproducible with the standard equipment. HT-MS/MS yields higher throughput than conventional LC-MS/MS-based methods while the HT-MS/MS system does not have a chromatographic step and cannot distinguish GAGs with identical molecular weights, leading to a limitation of measurements for some specific GAGs. Here we review the advantages and disadvantages of these methods for measuring GAG levels in biological specimens. We also describe an unexpected secondary elevation of keratan sulfate in patients with MPS that is an indirect consequence of disruption of catabolism of other GAGs.

  16. Glycosaminoglycan interactions in murine gammaherpesvirus-68 infection.

    Laurent Gillet

    Full Text Available Glycosaminoglycans (GAGs commonly participate in herpesvirus entry. They are thought to provide a reversible attachment to cells that promotes subsequent receptor binding. Murine gamma-herpesvirus-68 (MHV-68 infection of fibroblasts and epithelial cells is highly GAG-dependent. This is a function of the viral gp150, in that gp150-deficient mutants are much less GAG-dependent than wild-type. Here we show that the major MHV-68 GAG-binding protein is not gp150 but gp70, a product of ORF4. Surprisingly, ORF4-deficient MHV-68 showed normal cell binding and was more sensitive than wild-type to inhibition by soluble heparin rather than less. Thus, the most obvious viral GAG interaction made little direct contribution to infection. Indeed, a large fraction of the virion gp70 had its GAG-binding domain removed by post-translational cleavage. ORF4 may therefore act mainly to absorb soluble GAGs and prevent them from engaging gp150 prematurely. In contrast to gp70, gp150 bound poorly to GAGs, implying that it provides little in the way of adhesion. We hypothesize that it acts instead as a GAG-sensitive switch that selectively activates MHV-68 entry at cell surfaces.

  17. N-[3H]acetyl-labeling, a convenient method for radiolabeling of glycosaminoglycans

    A method for the introduction of N-[3H]acetyl groups into glycosaminoglycans is described. The procedure is based on [3H]acetylation of N-unsubstituted hexosamine residues by treating the polysaccharides with [3H]acetic anhydride. Preparations of heparin and heparin sulfate were found to contain significant numbers of N-unsubstituted hexosamine residues, as isolates. In contrast, such units could not be detected in chondroitin sulfate, dermatan sulfate, or hyaluronic acid. These polysaccharides were therefore subjected to partial N-deacetylation by reaction with hydrazine in the presence of hydrazine sulfate. After treatment with [3H]acetic anhydride, the specific activities of the resulting labeled polysaccharide preparations ranged between 0.1 X 106 and 0.6 X 106 cpm 3H/μg of uronic acid. The 3H-labeled polysaccharide preparations did not differ significantly from the corresponding unlabeled starting materials with regard to polyanion properties (chromatography on DEAE-cellulose) or polymer chain size (gel chromatography). Further, the radiolabeled polysaccharide derivatives were susceptible to specific enzymatic degradation (chondroitinase ABC and mammalian heparitinase) and retained their ability to interact specifically with certain proteins - for example, [3H]heparin with antithrombin [3H]hyaluronic acid oligosaccharides with chondroitin sulfate proteoglycan. These findings indicate that the labeling procedures did not induce any major structural derangement of the polysaccharide molecules. The method developed should be useful in providing labeled glycosaminoglycans for metabolic and enzymatic experiments as well as for studies on the interacion between glycosaminoglycans and other bilogical macromolecules

  18. N-(/sup 3/H)acetyl-labeling, a convenient method for radiolabeling of glycosaminoglycans

    Hook, M.; Riesenfeld, J.; Lindahl, U.

    1982-01-15

    A method for the introduction of N-(/sup 3/H)acetyl groups into glycosaminoglycans is described. The procedure is based on (/sup 3/H)acetylation of N-unsubstituted hexosamine residues by treating the polysaccharides with (/sup 3/H)acetic anhydride. Preparations of heparin and heparin sulfate were found to contain significant numbers of N-unsubstituted hexosamine residues, as isolates. In contrast, such units could not be detected in chondroitin sulfate, dermatan sulfate, or hyaluronic acid. These polysaccharides were therefore subjected to partial N-deacetylation by reaction with hydrazine in the presence of hydrazine sulfate. After treatment with (/sup 3/H)acetic anhydride, the specific activities of the resulting labeled polysaccharide preparations ranged between 0.1 X 10/sup 6/ and 0.6 X 10/sup 6/ cpm /sup 3/H/..mu..g of uronic acid. The /sup 3/H-labeled polysaccharide preparations did not differ significantly from the corresponding unlabeled starting materials with regard to polyanion properties (chromatography on DEAE-cellulose) or polymer chain size (gel chromatography). Further, the radiolabeled polysaccharide derivatives were susceptible to specific enzymatic degradation (chondroitinase ABC and mammalian heparitinase) and retained their ability to interact specifically with certain proteins - for example, (/sup 3/H)heparin with antithrombin (/sup 3/H)hyaluronic acid oligosaccharides with chondroitin sulfate proteoglycan. These findings indicate that the labeling procedures did not induce any major structural derangement of the polysaccharide molecules. The method developed should be useful in providing labeled glycosaminoglycans for metabolic and enzymatic experiments as well as for studies on the interacion between glycosaminoglycans and other bilogical macromolecules.

  19. Differences between two fractions of glycosaminoglycans of the corneal stroma in their structural relation to collagen.

    Dische, Z; Cremer-Bartels, G.; Kaye, G I

    1985-01-01

    Glycosaminoglycans (GAG) of bovine cornea were sequentially extracted by 0.15 M NaCl and by 1 M CaCl2, pH 8. The amounts of hexosamine (HexN) and hexuronic acid (HexUA), specific hexoses (Hex), and protein were determined in the extracts. The ultrastructure of the corneal stroma after NaCl and NaCl/CaCl2 extraction was also studied. Approximately 70% of the Hex-HexN extractable by 0.15 M NaCl is removed in the first NaCl extract, with the amount decreasing rapidly to the fifth NaCl extract. O...

  20. Mucopolysaccharide in healing process of gastric ulcer with special reference to 35SO4 autoradiographic findings and amount of hexosamine within mucous membrane

    This study presents protective mechanism of the mucous membrane with our main emphasis on mucopolysaccharides within the mucous membrane at varying stages of gastric ulcer. If we look at the findings based on 35SO4 autoradiography, an increase in uptake of 35SO4 by the epithelial cells of the active stage (A2) and a peak is exhibited during the initial half of the healing stage (H1). If the amount of hexosamine within the mucous membrane is determined, a movement similar to the 35SO4 autoradiographic findings is exhibited with a peak occurring from the A2 to the H1 stage. In summary, it is conceivable that based on the mucopolysaccharide metabolism during the varying stages of gastric ulcer, formation of a mucous barrier is quite rapidly initiated after ulceration and the importance of a protective factor in healing process of gastric ulcer is suggested. (author)

  1. Novel branch patterns and anticoagulant activity of glycosaminoglycan from sea cucumber Apostichopus japonicus.

    Yang, Jie; Wang, Yuanhong; Jiang, Tingfu; Lv, Zhihua

    2015-01-01

    A novel glucosidic pattern of fucose branches was found in the glycosaminoglycan from the sea cucumber Apostichopus japonicus in China. The methylation of desulfated/carboxyl-reduced polysaccharides and analysis of unsaturated disaccharides generated from the enzymolysis of the defucosed polysaccharides demonstrated that the branch is formed by one fucopyranosyl residue, 46.5% of which is linked through the O-3 position of β-D-glucuronic acid, while 8.7% and 43.9% are linked through the O-6 and O-4 positions of the N-acetylgalactosamine moiety. The β-D-glucuronic acid, N-acetyl-β-D-galactosamine, α-L-fucose and sulfate ester with the molecular ratio of 0.97:1.00:1.13:3.85 composed the backbone → 4)GlcUAβ(1 → 3)GalNAcβ(1 → and sulfated fucose branches. The sulfation patterns of fucose branches and the linkage pattern of the backbone structure were determined by 1/2 dimension NMR. The most abundant branch species were 2,4-di-O-sulfated and 3,4-di-O-sulfated fucose, but 4-mono-O-sulfated residue was also present. The structure of presently obtained glycosaminoglycan is different from that previously obtained from Stichopus japonicus (Kariya et al., Carbohyd. Res. 297 (1997) 273-279), which suggests that the structures of glycosaminoglycans from the same species of different regions somehow differ. The anticoagulant assay indicated that the polysaccharide possessed a high anticoagulant activity and the sulfated fucose branches were essential to the activity. PMID:25453278

  2. Synthesis of new glycosaminoglycans-like families by regioselective oxidation followed by sulphation of glucoglucuronan from Rhizobium sp. T1.

    Redouan, Elboutachfaiti; Emmanuel, Petit; Philippe, Michaud; Bernard, Courtois; Josiane, Courtois; Cedric, Delattre

    2012-08-01

    Glycosaminoglycan-like polysaccharides were prepared from Rhizobium sp. T1 polysaccharide using the TEMPO (nitroxyl radical 2,2,6,6-tetramethylpiperidine-1-oxyl radical) mediated oxidation. The structure of this new polyglucuronic acid sodium salt was analyzed by (13)C NMR spectra and HPAEC-PAD chromatography. Therefore, new polysaccharide containing only glucuronic acid monomers in both β-(1,3) and β-(1,4) linkage was obtained by the complete TEMPO-mediated oxidation of C6 primary hydroxyl groups of glucose of glucoglucuronan from Rhizobium sp. T1. Sulphation of this β-(1,3),β-(1,4)-polyglucuronic acid sodium salt was carried out using SO3/DMF reagent. These results suggested a new synthetic route using both TEMPO-mediated oxidation and sulphation of polysaccharides from Rhizobium sp. in developing glycosaminoglycans mimic to enhance the profitability of its low-cost production and processing industries. This novel carbohydrate derivative might find use as cheaper surrogates of glycosaminoglycans in the cosmetics and pharmaceutical fields. PMID:24750940

  3. Glycosaminoglycan sulphation affects the seeded misfolding of a mutant prion protein.

    Victoria A Lawson

    Full Text Available BACKGROUND: The accumulation of protease resistant conformers of the prion protein (PrP(res is a key pathological feature of prion diseases. Polyanions, including RNA and glycosaminoglycans have been identified as factors that contribute to the propagation, transmission and pathogenesis of prion disease. Recent studies have suggested that the contribution of these cofactors to prion propagation may be species specific. METHODOLOGY/PRINCIPAL FINDING: In this study a cell-free assay was used to investigate the molecular basis of polyanion stimulated PrP(res formation using brain tissue or cell line derived murine PrP. Enzymatic depletion of endogenous nucleic acids or heparan sulphate (HS from the PrP(C substrate was found to specifically prevent PrP(res formation seeded by mouse derived PrP(Sc. Modification of the negative charge afforded by the sulphation of glycosaminoglycans increased the ability of a familial PrP mutant to act as a substrate for PrP(res formation, while having no effect on PrP(res formed by wildtype PrP. This difference may be due to the observed differences in the binding of wild type and mutant PrP for glycosaminoglycans. CONCLUSIONS/SIGNIFICANCE: Cofactor requirements for PrP(res formation are host species and prion strain specific and affected by disease associated mutations of the prion protein. This may explain both species and strain dependent propagation characteristics and provide insights into the underlying mechanisms of familial prion disease. It further highlights the challenge of designing effective therapeutics against a disease which effects a range of mammalian species, caused by range of aetiologies and prion strains.

  4. Glycosaminoglycan-protein interactions and human complement factor H

    Blaum, Bärbel

    2010-01-01

    Glycosaminoglycans (GAGs) are linear polysaccharides expressed ubiquitously on animal cell surfaces and within extracellular matrices. GAGs usually occur as parts of proteoglycans and often accomplish their biological functions through their interactions with proteins. GAG oligosaccharides for this work were produced via enzymatic digest of heparin, followed by gel filtration and ion exchange chromatography. Two tetrasaccharide species obtained from this digest were characteris...

  5. Impacts of temperature increase and acidification on thickness of the surface mucopolysaccharide layer of the Caribbean coral Diploria spp.

    Pratte, Zoe A.; Richardson, Laurie L.

    2014-06-01

    Coral mechanisms of resilience and resistance to stressors such as increasing sea surface temperature and ocean acidification must first be understood in order to facilitate the survival of coral reefs as we know them. One such mechanism is production of the protective surface mucopolysaccharide layer (SML). In this study, we investigated changes in the thickness of the SML in response to increasing temperature and acidification for the three Caribbean scleractinian coral species of the genus Diploria, which have been shown to exhibit differential resilience to disease and bleaching. Among the three species, Diploria strigosa is known to have a higher susceptibility to disease, Diploria labyrinthiformis is known to bleach more quickly, and Diploria clivosa is relatively unstudied. When temperature was increased from 25 to 31 °C over a 1- or 6-week period, the overall thickness of the SML decreased from 33 to 55 % for all three species. Average SML thickness at 25 °C for all three species ranged from 106 to 156 μm, while average thickness at 31 °C ranged from 64 to 86 μm. SML thickness was significantly different among species at 25 °C, but not at 31 °C. D. labyrinthiformis demonstrated lower fragment mortality due to thermal stress when compared to the other Diploria species. Acidification from pH 8.2 to 7.7 over 5 weeks had no effect on SML thickness for any species. The observed decrease in SML thickness in response to increased temperature might be attributed to a decrease in the production of mucus or an increase in the viscosity of the SML. These findings may help to explain the increased prevalence of coral disease during the warmer months, since increased temperature compromises an important aspect of coral innate immunity, as well as differences in disease and bleaching susceptibilities between Diploria species.

  6. Study of Glycosaminoglycans of Extracellular Matrix (ECM in Pulp of Developing Tooth

    Kermany T

    2000-05-01

    Full Text Available Mesenchymal- epithelial interactions during embryogenesis have been shown to be important in the fetal development of many organs. Identification of molecules that modulate these interactions is key to our understanding of the pathological conditions. The major groups of extracellular matrix (ECM molecules characterized are glycosaminoglycans that candidate for morphogenesis and differentiation of ceils and tissues. In this study the molecules of ECM were considered in tooth development, pregnant female mice of balb-c were stained (vaginal plug=0 day and embryos (E12-E19 and newborns (PN1-PN9 were collected. Tissues were fixed, processed embedded and sectioned. Sections were stained with the following methods: Alcian Blue (pH=l, PAS-Alcian Blue (pH=2.5, Aician Blue(pH=5.8 prepared with for MgCL2 concentrations (CEC1- CEC4 and toluidin Blue. Non- parametric statistical test (Kruskall- Wallis showed significant difference between groups from the point of hyaluronic acid, chondroitin sulfate, carboxylated and sulfated glycosaminoglycan in pulp. It seems that the synthesis and secretion of components of ECM is important in morphogenic events and followed by a spatiotemporal pattern and developmentally regulated.

  7. Compositional analysis and structural elucidation of glycosaminoglycans in chicken eggs

    Liu, Zhangguo; Zhang, Fuming; Li, Lingyun; LI, GUOYUN; He, Wenqing; Linhardt, Robert J.

    2014-01-01

    Glycosaminoglycans (GAGs) have numerous applications in the fields of pharmaceuticals, cosmetics, nutraceuticals, and foods. GAGs are also critically important in the developmental biology of all multicellular animals. GAGs were isolated from chicken egg components including yolk, thick egg white, thin egg white, membrane, calcified shell matrix supernatant, and shell matrix deposit. Disaccharide compositional analysis was performed using ultra high-performance liquid chromatography-mass spec...

  8. A computational approach for deciphering the organization of glycosaminoglycans.

    Jean L Spencer

    Full Text Available BACKGROUND: Increasing evidence has revealed important roles for complex glycans as mediators of normal and pathological processes. Glycosaminoglycans are a class of glycans that bind and regulate the function of a wide array of proteins at the cell-extracellular matrix interface. The specific sequence and chemical organization of these polymers likely define function; however, identification of the structure-function relationships of glycosaminoglycans has been met with challenges associated with the unique level of complexity and the nontemplate-driven biosynthesis of these biopolymers. METHODOLOGY/PRINCIPAL FINDINGS: To address these challenges, we have devised a computational approach to predict fine structure and patterns of domain organization of the specific glycosaminoglycan, heparan sulfate (HS. Using chemical composition data obtained after complete and partial digestion of mixtures of HS chains with specific degradative enzymes, the computational analysis produces populations of theoretical HS chains with structures that meet both biosynthesis and enzyme degradation rules. The model performs these operations through a modular format consisting of input/output sections and three routines called chainmaker, chainbreaker, and chainsorter. We applied this methodology to analyze HS preparations isolated from pulmonary fibroblasts and epithelial cells. Significant differences in the general organization of these two HS preparations were observed, with HS from epithelial cells having a greater frequency of highly sulfated domains. Epithelial HS also showed a higher density of specific HS domains that have been associated with inhibition of neutrophil elastase. Experimental analysis of elastase inhibition was consistent with the model predictions and demonstrated that HS from epithelial cells had greater inhibitory activity than HS from fibroblasts. CONCLUSIONS/SIGNIFICANCE: This model establishes the conceptual framework for a new class of

  9. Effects of glycosaminoglycan from scallop skirt on foam cell

    Fu-shengSUN; SaiLIU

    2004-01-01

    AIM: To study effects of glycosaminoglycan from scallop skirt (SS-GAG) on NO production, antioxidative enzyme activity,and formation of macrophage-derived and smooth muscle cell-derived foam cell; to study the effects of SS-GAG on VEGF expression, intracellular Ca2~ level, and cytokines secretion of macrophage-derived foam cell. METHODS: Foam-like cells were generated by incubating the U937 cells or porcine artery smooth

  10. Effect of endothelium on glycosaminoglycan accumulation in injured rabbit aorta.

    Wight, T N; Curwen, K. D.; Litrenta, M. M.; Alonso, D. R.; Minick, C. R.

    1983-01-01

    Previous studies have indicated that reendothelialized regions of injured rabbit aortas are more susceptible to diet-induced atherosclerosis than persistently deendothelialized regions or uninjured aortas. However, the mechanism responsible for this selective lipid deposition is not understood. One possibility is that these regions differ with respect to the quantity and type of glycosaminoglycan-containing proteoglycans which are known to interact with lipoproteins. To determine whether thes...

  11. Marine Non-Glycosaminoglycan Sulfated Glycans as Potential Pharmaceuticals

    Vitor H. Pomin

    2015-01-01

    Sulfated fucans (SFs) and sulfated galactans (SGs) are currently the marine non-glycosaminoglycan (GAG) sulfated glycans most studied in glycomics. These compounds exhibit therapeutic effects in several pathophysiological systems such as blood coagulation, thrombosis, neovascularization, cancer, inflammation, and microbial infections. As analogs of the largely employed GAGs and due to some limitations of the GAG-based therapies, SFs and SGs comprise new carbohydrate-based therapeutics availab...

  12. Synthesis of Fluorophore-Tagged Xylosides That Prime Glycosaminoglycan Chains

    Tran, Vy M.; Kuberan, Balagurunathan

    2014-01-01

    Biosynthesis and functions of glycosaminoglycan (GAG) chains are complex and remain elusive. To better understand the factors that regulate the biosynthesis and functions, fluorophore-tagged xylosides carrying two different linkages between fluorophore and xylose residue were synthesized and evaluated for their ability to prime GAG chains such as heparan sulfate (HS), chondroitin sulfate (CS), and dermatan sulfate (DS) in various cell lines. These in vitro studies resulted in the identificati...

  13. Chapter 19 Analysis of Glycosaminoglycans in Stem Cell Glycomics

    Li, Boyangzi; Liu, Haiying; Zhang, Zhenqing; Stansfield, Hope E.; Dordick, Jonathan S.; Linhardt, Robert J.

    2011-01-01

    Glycosaminoglycans (GAGs) play a critical role in the binding and activation of growth factors in cell signal transduction required for biological development. A glycomics approach can be used to examine GAG content, composition, and structure in stem cells in order to characterize their general differentiation. Specifically, this method may be used to evaluate chondrogenic differentiations by profiling for the GAG content of the differentiated cells. Here, embryonic-like teratocarcinoma cell...

  14. Molecular engineering of glycosaminoglycan chemistry for biomolecule delivery

    Miller, Tobias; Goude, Melissa C.; McDevitt, Todd C.; Temenoff, Johnna S.

    2013-01-01

    Glycosaminoglycans (GAGs) are linear, negatively charged polysaccharid es that interact with a variety of positively-charged growth factors. In this review article, the effects of engineering GAG chemistry for molecular delivery applications in regenerative medicine are presented. Three major areas of focus at the structure-function-property interface are discussed: 1) macromolecular properties of GAGs, 2) effects of chemical modifications on protein binding, and 3) degradation mechanisms of ...

  15. Age-dependent alterations of decorin glycosaminoglycans in human skin

    Yong Li; Ying Liu; Wei Xia; Dan Lei; Voorhees, John J.; Fisher, Gary J.

    2013-01-01

    Proteoglycans, a family of glycosaminoglycan (GAG) conjugated proteins, are important constituents of human skin connective tissue (dermis) and are essential for maintaining mechanical strength of the skin. Age-related alterations of dermal proteoglycans have not been fully elucidated. We quantified transcripts of 20 known interstitial proteoglycans in human skin and found that decorin was the most highly expressed. Decorin was predominantly produced by dermal fibroblasts. Decorin was localiz...

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

    Nishida,Keiichiro

    1995-12-01

    Full Text Available 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.

  17. Specificity and affinity of binding of herpes simplex virus type 2 glycoprotein B to glycosaminoglycans.

    Williams, R K; Straus, S E

    1997-01-01

    Herpes simplex virus type 2 (HSV-2) interacts with cell surface glycosaminoglycans during virus attachment. Glycoprotein B of HSV-2 can potentially mediate the interaction between the virion and cell surface glycosaminoglycans. To determine the specificity, kinetics, and affinity of these interactions, we used plasmon resonance-based biosensor technology to measure HSV-2 glycoprotein binding to glycosaminoglycans in real time. The recombinant soluble ectodomain of HSV-2 gB (gB2) but not the s...

  18. Human milk glycosaminoglycans: the state of the art and future perspectives

    Coppa Giovanni Valentino

    2013-01-01

    Full Text Available Abstract Recently, a complete characterization and detailed evaluation of the glycosaminoglycans of human milk were performed. The total glycosaminoglycans content in milk from healthy mothers having delivered term or preterm newborns showed a constant pattern which was essentially composed of two main polysaccharides: chondroitin sulfate (60-70% and heparin (30-40%. Moreover, considerable variations of glycosaminoglycans concentration were found during the first month of lactation, the highest values being present in colostrum compared to mature milk. Metabolism and potential biological functions of human milk glycosaminoglycans are hypothesized and future studies are encouraged.

  19. Mapping by monoclonal antibody detection of glycosaminoglycans in connective tissues

    Couchman, J R; Caterson, B; Christner, J E;

    1984-01-01

    glycosaminoglycan (GAG), particularly with respect to self-association and interactions with other extracellular matrix components. Interactions with specific molecules from different connective tissue types, such as the collagens and their associated glycoproteins, could be favoured by particular charge...... organizations on the GAG molecule endowed by the sulphate groups. So far, it has not been possible to identify and map chondroitins of differing sulphation in tissues, but we have now raised three monoclonal antibodies which specifically recognize unsulphated, 4-sulphated and 6-sulphated chondroitin and...

  20. Nonradioactive glycosyltransferase and sulfotransferase assay to study glycosaminoglycan biosynthesis.

    Ethen, Cheryl M; Machacek, Miranda; Prather, Brittany; Tatge, Timothy; Yu, Haixiao; Wu, Zhengliang L

    2015-01-01

    Glycosaminoglycans (GAGs) are linear polysaccharides with repeating disaccharide units. GAGs include heparin, heparan sulfate, chondroitin sulfate, dermatan sulfate, keratan sulfate, and hyaluronan. All GAGs, except for hyaluronan, are usually sulfated. GAGs are polymerized by mono- or dual-specific glycosyltransferases and sulfated by various sulfotransferases. To further our understanding of GAG chain length regulation and synthesis of specific sulfation motifs on GAG chains, it is imperative to understand the kinetics of GAG synthetic enzymes. Here, nonradioactive colorimetric enzymatic assays are described for these glycosyltransferases and sulfotransferases. In both cases, the leaving nucleotides or nucleosides are hydrolyzed using specific phosphatases, and the released phosphate is subsequently detected using malachite reagents. PMID:25325970

  1. Localization of glycosaminoglycan substitution sites on domain V of mouse perlecan

    Tapanadechopone, P; Hassell, J R; Rigatti, B;

    1999-01-01

    Perlecan, the predominant basement membrane proteoglycan, has previously been shown to contain glycosaminoglycans attached at serine residues, numbers 65, 71, and 76, in domain I. However, the C-terminal domains IV and V of this molecule may also be substituted with glycosaminoglycan chains, but ...

  2. Inhibition of Chemokine-Glycosaminoglycan Interactions in Donor Tissue Reduces Mouse Allograft Vasculopathy and Transplant Rejection

    Dai, Erbin; Liu, Li-Ying; Wang, Hao; McIvor, Dana; Sun, Yun ming; Macaulay, Colin; King, Elaine; Munuswamy-Ramanujam, Ganesh; Bartee, Mee Yong; Williams, Jennifer; Davids, Jennifer; Charo, Israel; McFadden, Grant; Esko, Jeffrey D.; Lucas, Alexandra R.

    2010-01-01

    Background Binding of chemokines to glycosaminoglycans (GAGs) is classically described as initiating inflammatory cell migration and creating tissue chemokine gradients that direct local leukocyte chemotaxis into damaged or transplanted tissues. While chemokine-receptor binding has been extensively studied during allograft transplantation, effects of glycosaminoglycan (GAG) interactions with chemokines on transplant longevity are less well known. Here we examine the impact of interrupting che...

  3. Effect of glycosaminoglycan polysulphate on the human intervertebral discs based on stature measurements.

    Videman, T; Leskinen, T; Stalhammar, H; Riihimaki, H; Takala, E P; Viikari-Juntura, E; Forsskahl, B

    1994-05-01

    Glycosaminoglycan polysulphate has been shown to protect against loss of glycosaminoglycans in experimental osteoarthritis and has been used in humans to achieve the same result. It was assumed that if glycosaminoglycan polysulphate generally affects human connective tissues its use would increase or maintain intervertebral disc height by preventing water loss. Stature was measured during a resting-loading-resting period of 1.5 h. Four baseline measurements were taken of six subjects during a 4-week period. Thereafter each subject was given glycosaminoglycan polysulphate for 3 weeks and stature was measured once a week until a stable condition was reached; no less than three measurements were obtained. No effect of glycosaminoglycan polysulphate on the stature could, however, be seen in the mean values for all six subjects. PMID:23916184

  4. The effects of UVA irradiation on the metabolism of type I collagen and glycosaminoglycans by human dermal fibroblasts in three dimensional culture

    We studied the production of type I collagen and glycosaminoglycans by human dermal fibroblasts derived from young and old individuals after UVA irradiation. In these experiments, we introduced a new three-dimensional culture system supplemented with L-ascorbic acid 2-phosphate. In fibroblasts from old individuals (n=3), the amount of collagens in the cell layer was significantly decreased and matrix metallproteinase-1 (MMP-1) activity in the supernatant was significantly elevated after UVA irradiation, while the amounts of glycosaminoglycans in the cell layer and stromelysin-1 (MMP-3) activity showed no significant changes. In contrast, there were no significant changes after UVA irradiation of fibroblasts from young individuals (n=3). These results suggest that the donor age of dermal fibroblasts may be crucial to investigating the metabolism of type I collagen after UVA irradiation. (author)

  5. Human Genetic Disorders and Knockout Mice Deficient in Glycosaminoglycan

    Shuji Mizumoto

    2014-01-01

    Full Text Available Glycosaminoglycans (GAGs are constructed through the stepwise addition of respective monosaccharides by various glycosyltransferases and maturated by epimerases and sulfotransferases. The structural diversity of GAG polysaccharides, including their sulfation patterns and sequential arrangements, is essential for a wide range of biological activities such as cell signaling, cell proliferation, tissue morphogenesis, and interactions with various growth factors. Studies using knockout mice of enzymes responsible for the biosynthesis of the GAG side chains of proteoglycans have revealed their physiological functions. Furthermore, mutations in the human genes encoding glycosyltransferases, sulfotransferases, and related enzymes responsible for the biosynthesis of GAGs cause a number of genetic disorders including chondrodysplasia, spondyloepiphyseal dysplasia, and Ehlers-Danlos syndromes. This review focused on the increasing number of glycobiological studies on knockout mice and genetic diseases caused by disturbances in the biosynthetic enzymes for GAGs.

  6. Targeting Human Cancer by a Glycosaminoglycan Binding Malaria Protein

    Salanti, Ali; Clausen, Thomas M.; Agerbæk, Mette Ø.;

    2015-01-01

    Plasmodium falciparum engineer infected erythrocytes to present the malarial protein, VAR2CSA, which binds a distinct type chondroitin sulfate (CS) exclusively expressed in the placenta. Here, we show that the same CS modification is present on a high proportion of malignant cells and that it can...... be specifically targeted by recombinant VAR2CSA (rVAR2). In tumors, placental-like CS chains are linked to a limited repertoire of cancer-associated proteoglycans including CD44 and CSPG4. The rVAR2 protein localizes to tumors in vivo and rVAR2 fused to diphtheria toxin or conjugated to hemiasterlin compounds...... strongly inhibits in vivo tumor cell growth and metastasis. Our data demonstrate how an evolutionarily refined parasite-derived protein can be exploited to target a common, but complex, malignancy-associated glycosaminoglycan modification....

  7. [14C] acetylation of a glycosaminoglycan sulphate: Sulodexide

    The synthesis of [14C] labelled Sulodexide is reported. Sulodexide is a sulphated polysaccharide of the class of glycosaminoglycan, containing a heparin-like fraction (70%), dermatan sulphate (20%) and other minor fractions. The heparin-like fraction, suitably isolated from other components, was partially and selectively N-desulphated, thus making few percent unit -NH2 groups available for the labelling with [14C]-acetic anhydride (specific activity 0.15 μCi/mg). Due to the small extent of modification, the sulphate to carboxylate group ratio remained practically unchanged on the heparin-like fraction. Sulodexide was reconstituted adding to the labelled fraction the suitable amount of the other components; the chemical and biological properties of the final labelled Sulodexide were indistinguishable from those of the starting material. (author)

  8. Glycosaminoglycan, computed tomography and gallium-67 scanning in malignant pleural mesothelioma

    Malignant pleural mesothelioma is an unusual disease, often difficult to diagnose. This paper describes the results of quantitative studies on glycosaminoglycan (GAG) in tumor tissues and the findings of chest computed tomography (CT) and gallium-67 scanning in 5 malignant pleural mesotheliomas. The total amount of GAG in tumor tissue was 2.3 to 17.0 times as high as that in adenocarcinoma of the lung. The amount of hyaluronic acid was 3.5 to 170 times higher than that in adenocarcinoma. Also, the amount of chondroitin sulfate increased 2.6 to 10.0 times, but there were no changes in dermatan sulfate and heparan sulfate contents in this neoplasm when compared with adenocarcinoma. The present study suggests that a marked increase of the total amount of GAG and elevation of either hyaluronic acid and chondroitin sulfate content or both is a characteristic abnormality in this neoplasm. In most cases, CT scan of the chest showed pleural effusion, irregular pleural tumorous thickening surrounding the whole lung surface and extension into the fissure. In 3 cases, tumorous lesions extending into the chest wall at the site of pleural biopsy could be visualized on CT. In the terminal stage, the thoracic cavity was occupied by tumor tissues. Gallium-67 scanning showed a diffusely increased radionuclide accumulation over the involved hemithorax with or without particular intensity in the periphery. Conversely, identification of these characteristic findings of CT and gallium-67 scanning indicates the possibility of malignant pleural mesothelioma. (author)

  9. Glycosaminoglycan remodeling during diabetes and the role of dietary factors in their modulation.

    Gowd, Vemana; Gurukar, Abhignan; Chilkunda, Nandini D

    2016-02-25

    Glycosaminoglycans (GAGs) play a significant role in various aspects of cell physiology. These are complex polymeric molecules characterized by disaccharides comprising of uronic acid and amino sugar. Compounded to the heterogeneity, these are variously sulfated and epimerized depending on the class of GAG. Among the various classes of GAG, namely, chondroitin/dermatan sulfate, heparin/heparan sulfate, keratan sulfate and hyaluronic acid (HA), only HA is non-sulfated. GAGs are known to undergo remodeling in various tissues during various pathophysiological conditions, diabetes mellitus being one among them. These changes will likely affect their structure thereby impinging on their functionality. Till date, diabetes has been shown to affect GAGs in organs such as kidney, liver, aorta, skin, erythrocytes, etc. to name a few, with deleterious consequences. One of the mainstays in the treatment of diabetes is though dietary means. Various dietary factors are known to play a significant role in regulating glucose homeostasis. Furthermore, in recent years, there has been a keen interest to decipher the role of dietary factors on GAG metabolism. This review focuses on the remodeling of GAGs in various organs during diabetes and their modulation by dietary factors. While effect of diabetes on GAG metabolism has been worked out quite a bit, studies on the role of dietary factors in their modulation has been few and far between. We have tried our best to give the latest reports available on this subject. PMID:26962410

  10. A study on the relationship between radiologic classification and glycosaminoglycan analysis of cystic fluids in oral region

    Park, In Woo; You, Dong Soo [Dept. of Oral and Maxillofacial Radiology, College of Dentistry, Seoul National University, Seoul (Korea, Republic of)

    1993-08-15

    This study was designed to evaluate the correlationship between radiologic classifications of cysts in oral region and glycosaminoglycan analysis of cystic fluids using cellulose acetate electrophoresis. The materials for this study consisted of 37 cases-8 periapical cysts, 10 dentigerous cysts, 10 primordial cysts, 2 residual cyst, 3 incisive canal cysts, 2 post-operative maxillary cysts, 1 mucocele on maxillary sinus, and 1 unicystic ameloblastoma-diagnosed as cystic lesions radiologically. The obtained results were as follows: 1. At the stepwise discriminant analysis, four variables-low mobility material, hiparin, hyaluronic acid, and dermatan sulfate- were used to define diagnostic model for the odontogenic cyst. The model produced a seventeenths of 100% and a specificity of 85%. 2. The intensities of heparin and chondroitin-4-sulfate were greater in dentigerous cyst than periapical cyst (p<0.05). 3. It showed no statistically significant difference in glycosaminoglycan of the cystic fluids between dentigerous cyst and primordial cyst (p<0.05). 4. On the fluids of the cysts originated from maxillary sinus, there were especially high intensities of heparin and dermatan sulfate, and low intensity of chondroitin-4-sulfate. 5. On the fluids of unicystic ameloblastoma, there were high intensity of dermatan sulfate and low intensity of chondroitin-4-sulfate.

  11. A study on the relationship between radiologic classification and glycosaminoglycan analysis of cystic fluids in oral region

    This study was designed to evaluate the correlationship between radiologic classifications of cysts in oral region and glycosaminoglycan analysis of cystic fluids using cellulose acetate electrophoresis. The materials for this study consisted of 37 cases-8 periapical cysts, 10 dentigerous cysts, 10 primordial cysts, 2 residual cyst, 3 incisive canal cysts, 2 post-operative maxillary cysts, 1 mucocele on maxillary sinus, and 1 unicystic ameloblastoma-diagnosed as cystic lesions radiologically. The obtained results were as follows: 1. At the stepwise discriminant analysis, four variables-low mobility material, hiparin, hyaluronic acid, and dermatan sulfate- were used to define diagnostic model for the odontogenic cyst. The model produced a seventeenths of 100% and a specificity of 85%. 2. The intensities of heparin and chondroitin-4-sulfate were greater in dentigerous cyst than periapical cyst (p<0.05). 3. It showed no statistically significant difference in glycosaminoglycan of the cystic fluids between dentigerous cyst and primordial cyst (p<0.05). 4. On the fluids of the cysts originated from maxillary sinus, there were especially high intensities of heparin and dermatan sulfate, and low intensity of chondroitin-4-sulfate. 5. On the fluids of unicystic ameloblastoma, there were high intensity of dermatan sulfate and low intensity of chondroitin-4-sulfate.

  12. Purification of the lysosomal sialic acid transporter. Functional characteristics of a monocarboxylate transporter

    A.C. Havelaar (Adrie); G.M.S. Mancini (Grazia); C.E.M.T. Beerens (Cecile); R.M. Souren; F.W. Verheijen (Frans)

    1998-01-01

    textabstractSialic acid and glucuronic acid are monocarboxylated monosaccharides, which are normally present in sugar side chains of glycoproteins, glycolipids, and glycosaminoglycans. After degradation of these compounds in lysosomes, the free monosaccharides are relea

  13. Treponema pallidum does not synthesise in vitro a capsule containing glycosaminoglycans or proteoglycans.

    Strugnell, R. A.; Handley, C J; Lowther, D.A.; Faine, S; Graves, S R

    1984-01-01

    Treponema pallidum was investigated for its ability to synthesise glycosaminoglycans or proteoglycans in vitro. Isolated viable T pallidum organisms were incubated with radiolabelled precursors of glycosaminoglycans, sodium 35S-sulphate and 3H-glucosamine (tritiated glucosamine). T pallidum failed to incorporate sodium 35S-sulphate but did incorporate 3H-glucosamine into a macromolecule which may be associated with the surface of the treponeme. This macromolecule was resistant to degradation ...

  14. The structural stability of the endothelial glycocalyx after enzymatic removal of glycosaminoglycans.

    Ye Zeng

    Full Text Available RATIONALE: It is widely believed that glycosaminoglycans (GAGs and bound plasma proteins form an interconnected gel-like structure on the surface of endothelial cells (the endothelial glycocalyx layer-EGL that is stabilized by the interaction of its components. However, the structural organization of GAGs and proteins and the contribution of individual components to the stability of the EGL are largely unknown. OBJECTIVE: To evaluate the hypothesis that the interconnected gel-like glycocalyx would collapse when individual GAG components were almost completely removed by a specific enzyme. METHODS AND RESULTS: Using confocal microscopy, we observed that the coverage and thickness of heparan sulfate (HS, chondroitin sulfate (CS, hyaluronic acid (HA, and adsorbed albumin were similar, and that the thicknesses of individual GAGs were spatially nonuniform. The individual GAGs were degraded by specific enzymes in a dose-dependent manner, and decreased much more in coverage than in thickness. Removal of HS or HA did not result in cleavage or collapse of any of the remaining components. Simultaneous removal of CS and HA by chondroitinase did not affect HS, but did reduce adsorbed albumin, although the effect was not large. CONCLUSION: All GAGs and adsorbed proteins are well inter-mixed within the structure of the EGL, but the GAG components do not interact with one another. The GAG components do provide binding sites for albumin. Our results provide a new view of the organization of the endothelial glycocalyx layer and provide the first demonstration of the interaction between individual GAG components.

  15. The identification of proteoglycans and glycosaminoglycans in archaeological human bones and teeth.

    Yvette M Coulson-Thomas

    Full Text Available Bone tissue is mineralized dense connective tissue consisting mainly of a mineral component (hydroxyapatite and an organic matrix comprised of collagens, non-collagenous proteins and proteoglycans (PGs. Extracellular matrix proteins and PGs bind tightly to hydroxyapatite which would protect these molecules from the destructive effects of temperature and chemical agents after death. DNA and proteins have been successfully extracted from archaeological skeletons from which valuable information has been obtained; however, to date neither PGs nor glycosaminoglycan (GAG chains have been studied in archaeological skeletons. PGs and GAGs play a major role in bone morphogenesis, homeostasis and degenerative bone disease. The ability to isolate and characterize PG and GAG content from archaeological skeletons would unveil valuable paleontological information. We therefore optimized methods for the extraction of both PGs and GAGs from archaeological human skeletons. PGs and GAGs were successfully extracted from both archaeological human bones and teeth, and characterized by their electrophoretic mobility in agarose gel, degradation by specific enzymes and HPLC. The GAG populations isolated were chondroitin sulfate (CS and hyaluronic acid (HA. In addition, a CSPG was detected. The localization of CS, HA, three small leucine rich PGs (biglycan, decorin and fibromodulin and glypican was analyzed in archaeological human bone slices. Staining patterns were different for juvenile and adult bones, whilst adolescent bones had a similar staining pattern to adult bones. The finding that significant quantities of PGs and GAGs persist in archaeological bones and teeth opens novel venues for the field of Paleontology.

  16. NMR structural determination of unique invertebrate glycosaminoglycans endowed with medical properties.

    Pomin, Vitor H

    2015-09-01

    Glycosaminoglycans (GAGs) are sulfated polysaccharides of complex structure endowed with numerous biomedical functions. Although ubiquitously distributed in vertebrates, GAGs can also occur in certain terrestrial or marine invertebrates. Solution nuclear magnetic resonance (NMR) spectroscopy has been the analytical technique mostly employed in structural characterization of GAGs from any source. This review aims at illustrating the application of NMR in structural determination of few representative invertebrate GAG examples of unique structures and endowed with therapeutic actions. They are the holothurian fucosylated chondroitin sulfate, the acharan sulfate isolated from the snail Achatina fulica, the dermatan sulfates with distinct sulfation patterns extracted from ascidian species, the sulfated glucuronic acid-containing heparan sulfate isolated from the gastropode Nodipecten nodosum, and the hybrid heparin/heparan sulfate molecule obtained from the shrimp Litopenaeus vannamei. These invertebrate GAGs exhibit distinct structures when compared to those extracted from mammalian GAGs. The distinct structures of the invertebrate GAGs lead also to different mechanisms of actions as compared to the mammalian GAG standards. Invertebrate GAGs comprise promising therapeutic candidates in fights against diseases. Solution NMR has been playing a pivotal role in this carbohydrate-based drug research, discovery and development. PMID:26083200

  17. "GAG-ing with the neuron": The role of glycosaminoglycan patterning in the central nervous system.

    Smith, Patrice D; Coulson-Thomas, Vivien J; Foscarin, Simona; Kwok, Jessica C F; Fawcett, James W

    2015-12-01

    Proteoglycans (PGs) are a diverse family of proteins that consist of one or more glycosaminoglycan (GAG) chains, covalently linked to a core protein. PGs are major components of the extracellular matrix (ECM) and play critical roles in development, normal function and damage-response of the central nervous system (CNS). GAGs are classified based on their disaccharide subunits, into the following major groups: chondroitin sulfate (CS), heparan sulfate (HS), heparin (HEP), dermatan sulfate (DS), keratan sulfate (KS) and hyaluronic acid (HA). All except HA are modified by sulfation, giving GAG chains specific charged structures and binding properties. While significant neuroscience research has focused on the role of one PG family member, chondroitin sulfate proteoglycan (CSPG), there is ample evidence in support of a role for the other PGs in regulating CNS function in normal and pathological conditions. This review discusses the role of all the identified PG family members (CS, HS, HEP, DS, KS and HA) in normal CNS function and in the context of pathology. Understanding the pleiotropic roles of these molecules in the CNS may open the door to novel therapeutic strategies for a number of neurological conditions. PMID:26277685

  18. Metabolic fate of milk glycosaminoglycans in breastfed and formula fed newborns.

    Maccari, Francesca; Mantovani, Veronica; Gabrielli, Orazio; Carlucci, Antonio; Zampini, Lucia; Galeazzi, Tiziana; Galeotti, Fabio; Coppa, Giovanni V; Volpi, Nicola

    2016-04-01

    In this study, the content, structure and residual percentages of glycosaminoglycans (GAGs) in the feces of seven breastfed newborns after ingesting a known amount of milk were studied. A comparison was made with five newborns fed with formula milk. Characterization of GAGs from milk and feces samples was performed according to previous methodology. Compared to the ingested GAGs present in milk, residual feces GAGs of breastfed newborns were 99 % of human milk GAGs are utilized as opposed to ~96 % of formula milk. Hyaluronic acid utilization was found to be fairly similar contrary to chondroitin sulfate/dermatan sulfate and heparan sulfate, which were found to be ~10-18 times lower in formula milk fed children. Our new results further demonstrate that the elevated content of human milk GAGs passes undigested through the entire digestive system of newborns, possibly protecting the infant from infections. In the distal gastrointestinal tract, these complex macromolecules are catabolized by a cohort of bacterial enzymes and constituent monosaccharides/oligosaccharides utilized for further metabolic purposes potentially useful for bacteria metabolism or internalized by intestinal cells. Thanks to their elevated structural heterogeneity, milk GAGs are used differently depending on their distinct primary structure. Finally, a different utilization and availability was observed for human milk GAGs compared to formula milk due to their various composition and structural heterogeneity. PMID:26873820

  19. Evaluation of the metabolism of glycosaminoglycans in patients with interstitial cystis

    Marcos Lucon

    2014-01-01

    Full Text Available Introduction: Painful bladder syndrome/interstitial cystitis (PBS/IC pathogenesis is not fully known, but evidence shows that glycosaminoglycans (GAG of bladder urothelium can participate in its genesis. The loss of these compounds facilitates the contact of urine compounds with deeper portions of bladder wall triggering an inflammatory process. We investigated GAG in urine and tissue of PBS/IC and pure stress urinary incontinence (SUI patients to better understand its metabolism. Materials and Methods: Tissue and urine of 11 patients with PBS/IC according to NIDDK criteria were compared to 11 SUI patients. Tissue samples were analyzed by histological, immunohistochemistry and immunofluorescence methods. Statistical analysis were performed using t Student test and Anova, considering significant when p < 0.05. Results: PBS/IC patients had lower concentration of GAG in urine when compared to SUI (respectively 0.45 ± 0.11 x 0.62 ± 0.13 mg/mg creatinine, p < 0.05. However, there was no reduction of the content of GAG in the urothelium of both groups. Immunofluorescence showed that PBS/IC patients had a stronger staining of TGF-beta, decorin (a proteoglycan of chondroitin/dermatan sulfate, fibronectin and hyaluronic acid. Conclusion: the results suggest that GAG may be related to the ongoing process of inflammation and remodeling of the dysfunctional urothelium that is present in the PBS/IC.

  20. A Complex Dance: The Importance of Glycosaminoglycans and Zinc in the Aggregation of Human Prolactin.

    Christensen, Line Friis Bakmann; Malmos, Kirsten Gade; Christiansen, Gunna; Otzen, Daniel Erik

    2016-07-01

    The zinc binding hormone pituitary human prolactin (hPRL) is stored in secretory granules of specialized cells in an aggregated form. Glycosaminoglycans (GAGs) are anionic polysaccharides commonly associated with secretory granules, indicating their involvement in granule formation. Here we, for the first time, study the impact of GAGs in combination with Zn(2+) on the reversible hPRL aggregation across the pH range of 7.4-5.5. Zn(2+) alone causes hPRL aggregation at pH 7.4, while aggregation between pH 7.4 and 5.5 requires both Zn(2+) and GAGs. GAGs alone cause hPRL aggregation below pH 5.5. Comprehensive thermal stability investigations show that hPRL is particularly destabilized toward thermal denaturation at pH 5.5 and that GAGs increasingly destabilize hPRL at decreasing pH values. We propose that Zn(2+) causes hPRL aggregation through low-affinity Zn(2+) binding sites on hPRL with GAGs facilitating Zn(2+) binding by neutralizing repulsive positive charges of hPRL in the acidic environments of the TGN and mature secretory granules. In a manner independent of the aggregation-causing agent(s), the different hPRL aggregates show very similar secondary structure and amorphous morphology. We speculate that this may be a recognizable sorting signal in the formation of hPRL granular vesicles. PMID:27305175

  1. Extraction and quantification of sulfated glycosaminoglycan content in five different aquatic species of Malaysia

    Ravi Lokwani; Ramandeep Singh; Gauree Kukreti

    2015-01-01

    Objective: To extract, characterize and quantify glycosaminoglycans (GAGs) from the body of cuttlefish, tennis-ball sea cucumber, shrimp, seabass and fresh water fish Nile tilapia. Methods: The extracted crude powder was evaluated for the content of GAGs. The qualitative analysis of sulfated pattern and other important functional groups related with GAGs were explained in the form of Fourier transform infra-red spectroscopy data. Proteins and nucleic acid in the crude extract were determined by the ultraviolet spectrophotometer, while the quantification of total sulfated GAGs and estimation of N-sulfated and O-sulfated GAGs in the crude mixture were performed by using Blyscan kit. Results: The sulfated pattern and other important functional groups related with GAGs were intercepted in Fourier transform infrared analysis. Blyscan quantification method reported that a rare variety of sea cucumber (tennis-ball sea cucumber) emerged as a rich source of GAGs with high values of both N-sulfated and O-sulfated GAGs in comparison to its other counterparts. Conclusions: Findings in this study point out the potential of tennis-ball sea cucumber, a rare variety of sea cucumber to act as an alternative source for GAG extraction for commercial purpose.

  2. Extraction and quantification of sulfated glycosaminoglycan content in five different aquatic species of Malaysia

    Ravi Lokwani

    2015-09-01

    Full Text Available Objective: To extract, characterize and quantify glycosaminoglycans (GAGs from the body of cuttlefish, tennis-ball sea cucumber, shrimp, seabass and fresh water fish Nile tilapia. Methods: The extracted crude powder was evaluated for the content of GAGs. The qualitative analysis of sulfated pattern and other important functional groups related with GAGs were explained in the form of Fourier transform infra-red spectroscopy data. Proteins and nucleic acid in the crude extract were determined by the ultraviolet spectrophotometer, while the quantification of total sulfated GAGs and estimation of N-sulfated and O-sulfated GAGs in the crude mixture were performed by using Blyscan kit. Results: The sulfated pattern and other important functional groups related with GAGs were intercepted in Fourier transform infrared analysis. Blyscan quantification method reported that a rare variety of sea cucumber (tennis-ball sea cucumber emerged as a rich source of GAGs with high values of both N-sulfated and O-sulfated GAGs in comparison to its other counterparts. Conclusions: Findings in this study point out the potential of tennis-ball sea cucumber, a rare variety of sea cucumber to act as an alternative source for GAG extraction for commercial purpose.

  3. Compositional analysis and structural elucidation of glycosaminoglycans in chicken eggs.

    Liu, Zhangguo; Zhang, Fuming; Li, Lingyun; Li, Guoyun; He, Wenqing; Linhardt, Robert J

    2014-11-01

    Glycosaminoglycans (GAGs) have numerous applications in the fields of pharmaceuticals, cosmetics, nutraceuticals, and foods. GAGs are also critically important in the developmental biology of all multicellular animals. GAGs were isolated from chicken egg components including yolk, thick egg white, thin egg white, membrane, calcified shell matrix supernatant, and shell matrix deposit. Disaccharide compositional analysis was performed using ultra high-performance liquid chromatography-mass spectrometry. The results of these analyses showed that all four families of GAGs were detected in all egg components. Keratan sulfate was found in egg whites (thick and thin) and shell matrix (calcified shell matrix supernatant and deposit) with high level. Chondroitin sulfates were much more plentiful in both shell matrix components and membrane. Hyaluronan was plentiful in both shell matrix components and membrane, but was only present in a trace of quantities in the yolk. Heparan sulfate was plentiful in the shell matrix deposit but was present in a trace of quantities in the egg content components (yolk, thick and thin egg whites). Most of the chondroitin and heparan sulfate disaccharides were present in the GAGs found in chicken eggs with the exception of chondroitin and heparan sulfate 2,6-disulfated disaccharides. Both CS and HS in the shell matrix deposit contained the most diverse chondroitin and heparan sulfate disaccharide compositions. Eggs might provide a potential new source of GAGs. PMID:25218438

  4. Roles of Proteoglycans and Glycosaminoglycans in Wound Healing and Fibrosis

    Shibnath Ghatak

    2015-01-01

    Full Text Available A wound is a type of injury that damages living tissues. In this review, we will be referring mainly to healing responses in the organs including skin and the lungs. Fibrosis is a process of dysregulated extracellular matrix (ECM production that leads to a dense and functionally abnormal connective tissue compartment (dermis. In tissues such as the skin, the repair of the dermis after wounding requires not only the fibroblasts that produce the ECM molecules, but also the overlying epithelial layer (keratinocytes, the endothelial cells, and smooth muscle cells of the blood vessel and white blood cells such as neutrophils and macrophages, which together orchestrate the cytokine-mediated signaling and paracrine interactions that are required to regulate the proper extent and timing of the repair process. This review will focus on the importance of extracellular molecules in the microenvironment, primarily the proteoglycans and glycosaminoglycan hyaluronan, and their roles in wound healing. First, we will briefly summarize the physiological, cellular, and biochemical elements of wound healing, including the importance of cytokine cross-talk between cell types. Second, we will discuss the role of proteoglycans and hyaluronan in regulating these processes. Finally, approaches that utilize these concepts as potential therapies for fibrosis are discussed.

  5. The Effect of Glycosaminoglycans (GAGs on Amyloid Aggregation and Toxicity

    Clara Iannuzzi

    2015-02-01

    Full Text Available Amyloidosis is a protein folding disorder in which normally soluble proteins are deposited extracellularly as insoluble fibrils, impairing tissue structure and function. Charged polyelectrolytes such as glycosaminoglycans (GAGs are frequently found associated with the proteinaceous deposits in tissues of patients affected by amyloid diseases. Experimental evidence indicate that they can play an active role in favoring amyloid fibril formation and stabilization. Binding of GAGs to amyloid fibrils occurs mainly through electrostatic interactions involving the negative polyelectrolyte charges and positively charged side chains residues of aggregating protein. Similarly to catalyst for reactions, GAGs favor aggregation, nucleation and amyloid fibril formation functioning as a structural templates for the self-assembly of highly cytotoxic oligomeric precursors, rich in β-sheets, into harmless amyloid fibrils. Moreover, the GAGs amyloid promoting activity can be facilitated through specific interactions via consensus binding sites between amyloid polypeptide and GAGs molecules. We review the effect of GAGs on amyloid deposition as well as proteins not strictly related to diseases. In addition, we consider the potential of the GAGs therapy in amyloidosis.

  6. Glycosaminoglycans and Glycomimetics in the Central Nervous System

    Dáire Rowlands

    2015-02-01

    Full Text Available With recent advances in the construction of synthetic glycans, selective targeting of the extracellular matrix (ECM as a potential treatment for a wide range of diseases has become increasingly popular. The use of compounds that mimic the structure or bioactive function of carbohydrate structures has been termed glycomimetics. These compounds are mostly synthetic glycans or glycan-binding constructs which manipulate cellular interactions. Glycosaminoglycans (GAGs are major components of the ECM and exist as a diverse array of differentially sulphated disaccharide units. In the central nervous system (CNS, they are expressed by both neurons and glia and are crucial for brain development and brain homeostasis. The inherent diversity of GAGs make them an essential biological tool for regulating a complex range of cellular processes such as plasticity, cell interactions and inflammation. They are also involved in the pathologies of various neurological disorders, such as glial scar formation and psychiatric illnesses. It is this diversity of functions and potential for selective interventions which makes GAGs a tempting target. In this review, we shall describe the molecular make-up of GAGs and their incorporation into the ECM of the CNS. We shall highlight the different glycomimetic strategies that are currently being used in the nervous system. Finally, we shall discuss some possible targets in neurological disorders that may be addressed using glycomimetics.

  7. Glycosaminoglycan analogs as a novel anti-inflammatory strategy

    Severin, India C.; Soares, Adriano; Hantson, Jennifer; Teixeira, Mauro; Sachs, Daniela; Valognes, Delphine; Scheer, Alexander; Schwarz, Matthias K.; Wells, Timothy N. C.; Proudfoot, Amanda E. I.; Shaw, Jeffrey

    2012-01-01

    Heparin, a glycosaminoglycan (GAG), has both anti-inflammatory and anti-coagulant properties. The clinical use of heparin against inflammation, however, has been limited by concerns about increased bleeding. While the anti-coagulant activity of heparin is well understood, its anti-inflammatory properties are less so. Heparin is known to bind to certain cytokines, including chemokines, small proteins which mediate inflammation through their control of leukocyte migration and activation. Molecules which can interrupt the chemokine-GAG interaction without inhibiting coagulation could therefore, represent a new class of anti-inflammatory agents. In the present study, two approaches were undertaken, both focusing on the heparin-chemokine relationship. In the first, a structure based strategy was used: after an initial screening of potential small molecule binders using protein NMR on a target chemokine, binding molecules were optimized through structure-based design. In the second approach, commercially available short oligosaccharides were polysulfated. In vitro, these molecules prevented chemokine-GAG binding and chemokine receptor activation without disrupting coagulation. However, in vivo, these compounds caused variable results in a murine peritoneal recruitment assay, with a general increase of cell recruitment. In more disease specific models, such as antigen-induced arthritis and delayed-type hypersensitivity, an overall decrease in inflammation was noted, suggesting that the primary anti-inflammatory effect may also involve factors beyond the chemokine system. PMID:23087686

  8. Furosemide modifies heart hypertrophy and glycosaminoglycan myocardium content in a rat model of neurogenic hypertension.

    Pourzitaki, Chryssa; Tsaousi, Georgia; Manthou, Maria Eleni; Karakiulakis, Georgios; Kouvelas, Dimitrios; Papakonstantinou, Eleni

    2016-08-01

    Hypertension is a major risk factor for atherogenesis and heart hypertrophy, both of which are associated with specific morphological and functional changes of the myocardium. Glycosaminoglycans (GAGs) are complex molecules involved both in tissue morphology and function. In the present study, we investigated the effects of neurogenic hypertension and subsequent antihypertensive treatment with furosemide, on heart hypertrophy and the content of GAGs in the myocardium. Neurogenic hypertension was achieved in male Wistar rats by bilateral aortic denervation (bAD). At days 2, 7 and 15 after surgery, animals were sacrificed and the hearts were dissected away, weighted, and homogenized. Total GAGs were assessed by measuring the uronic acid content colorimetrically and individual GAGs were isolated and characterized by enzymatic treatment, with GAG-degrading enzymes, using electrophoresis on polyacrylamide gradient gels and cellulose acetate membranes. In bAD-animals blood pressure, blood pressure lability, heart rate and heart weight were significantly increased 15 days postoperatively. These effects were prevented by treatment with furosemide. Major GAGs identified in the heart were chondroitin sulphates, heparin (H), heparan sulphate (HS) and hyaluronic acid. The content of uronic and the relative content of H and HS in the heart in bAD animals significantly decreased from day 2 to day 15 postoperatively. Furosemide prevented the bAD induced decrease in GAG content. Considering that H and HS are potent inhibitors of cardiomyocyte hypertrophy, our results indicate that heart hypertrophy induced by neurogenic hypertension may be associated with decreases in the relative content of heparin and heparan sulphate in the heart. PMID:27221775

  9. Macrophage polarization alters the expression and sulfation pattern of glycosaminoglycans.

    Martinez, Pierre; Denys, Agnès; Delos, Maxime; Sikora, Anne-Sophie; Carpentier, Mathieu; Julien, Sylvain; Pestel, Joël; Allain, Fabrice

    2015-05-01

    Macrophages are major cells of inflammatory process and take part in a large number of physiological and pathological processes. According to tissue environment, they can polarize into pro-inflammatory (M1) or alternative (M2) cells. Although many evidences have hinted to a potential role of cell-surface glycosaminoglycans (GAGs) in the functions of macrophages, the effect of M1 or M2 polarization on the biosynthesis of these polysaccharides has not been investigated so far. GAGs are composed of repeat sulfated disaccharide units. Heparan (HS) and chondroitin/dermatan sulfates (CS/DS) are the major GAGs expressed at the cell membrane. They are involved in numerous biological processes, which rely on their ability to selectively interact with a large panel of proteins. More than 20 genes encoding sulfotransferases have been implicated in HS and CS/DS biosynthesis, and the functional repertoire of HS and CS/DS has been related to the expression of these isoenzymes. In this study, we analyzed the expression of sulfotransferases as a response to macrophage polarization. We found that M1 and M2 activation drastically modified the profiles of expression of numerous HS and CS/DS sulfotransferases. This was accompanied by the expression of GAGs with distinct structural features. We then demonstrated that GAGs of M2 macrophages were efficient to present fibroblast growth factor-2 in an assay of tumor cell proliferation, thus indicating that changes in GAG structure may contribute to the functions of polarized macrophages. Altogether, our findings suggest a regulatory mechanism in which fine modifications in GAG biosynthesis may participate to the plasticity of macrophage functions. PMID:25504800

  10. Glycosaminoglycan synthesis in amiodarone-induced pulmonary fibrosis

    Glycosaminoglycans (GAG) have previously been demonstrated to be synthesized in greater than normal amounts following a single intratracheal insufflation of bleomycin in hamsters. This suggests that GAG may play a role in the propagation of pulmonary fibrotic reactions. To further test this hypothesis, GAG synthesis was studied in a new hamster model of interstitial lung injury, induced by the cardiac drug, aminodarone. Animals received a single intratracheal instillation of 1.25 mg aminodarone. At 4, 9, and 21 days post-insufflation, the animals were sacrificed, their lungs removed, and 1 mm fragments placed in explant culture for 6 hours at 370C in the presence of 35S-sulfate. The labeled GAG were isolated and measured for 35S incorporation. The author then isolated the hexosamine portions of the respective GAGs, Heparan Sulfate (HEP S), Chondroitin-6-Sulfate (Ch-6-S) and Chondroitin-4-Sulfate and Dermatan Sulfate (CH-4-S and DS) using the enzyme ABC and paper chromatography. They also studied the GAG content and distribution in hamster lung fibroblasts incorporated with 35S for 48 hours and subjected to either 0, 0.01 mg, 0.1 mg, or 1 mg of aminodarone. GAG synthesis is increased at an early stage following the induction of lung injury by aminodarone and remains elevated for a 3 week period. The change in GAG distribution boards elevated CH-4-S and DS may be characteristic of interstitial diseases in general. The GAGs that are synthesized by fibroblasts may be responsible for the increased CH-4-S and DS synthesis

  11. Glycosaminoglycan synthesis in amiodarone-induced pulmonary fibrosis

    Farinas, E.M.

    1986-01-01

    Glycosaminoglycans (GAG) have previously been demonstrated to be synthesized in greater than normal amounts following a single intratracheal insufflation of bleomycin in hamsters. This suggests that GAG may play a role in the propagation of pulmonary fibrotic reactions. To further test this hypothesis, GAG synthesis was studied in a new hamster model of interstitial lung injury, induced by the cardiac drug, aminodarone. Animals received a single intratracheal instillation of 1.25 mg aminodarone. At 4, 9, and 21 days post-insufflation, the animals were sacrificed, their lungs removed, and 1 mm fragments placed in explant culture for 6 hours at 37/sup 0/C in the presence of /sup 35/S-sulfate. The labeled GAG were isolated and measured for /sup 35/S incorporation. The author then isolated the hexosamine portions of the respective GAGs, Heparan Sulfate (HEP S), Chondroitin-6-Sulfate (Ch-6-S) and Chondroitin-4-Sulfate and Dermatan Sulfate (CH-4-S and DS) using the enzyme ABC and paper chromatography. They also studied the GAG content and distribution in hamster lung fibroblasts incorporated with /sup 35/S for 48 hours and subjected to either 0, 0.01 mg, 0.1 mg, or 1 mg of aminodarone. GAG synthesis is increased at an early stage following the induction of lung injury by aminodarone and remains elevated for a 3 week period. The change in GAG distribution boards elevated CH-4-S and DS may be characteristic of interstitial diseases in general. The GAGs that are synthesized by fibroblasts may be responsible for the increased CH-4-S and DS synthesis.

  12. Urinary glycosaminoglycans excretion and the effect of dimethyl sulfoxide in an experimental model of non-bacterial cystitis

    Roberto Soler

    2008-08-01

    Full Text Available PURPOSE: We reproduced a non-bacterial experimental model to assess bladder inflammation and urinary glycosaminoglycans (GAG excretion and examined the effect of dimethyl sulfoxide (DMSO. MATERIALS AND METHODS: Female rats were instilled with either protamine sulfate (PS groups or sterile saline (control groups. At different days after the procedure, 24 h urine and bladder samples were obtained. Urinary levels of hyaluronic acid (HA and sulfated glycosaminoglycans (S-GAG were determined. Also to evaluate the effect of DMSO animals were instilled with either 50% DMSO or saline 6 hours after PS instillation. To evaluate the effect of DMSO in healthy bladders, rats were instilled with 50% DMSO and controls with saline. RESULTS: In the PS groups, bladder inflammation was observed, with polymorphonuclear cells during the first days and lymphomononuclear in the last days. HA and S-GAG had 2 peaks of urinary excretion, at the 1st and 7th day after PS injection. DMSO significantly reduced bladder inflammation. In contrast, in healthy bladders, DMSO produced mild inflammation and an increase in urinary HA levels after 1 and 7 days and an increase of S-GAG level in 7 days. Animals instilled with PS and treated with DMSO had significantly reduced levels of urinary HA only at the 1st day. Urinary S-GAG/Cr levels were similar in all groups. CONCLUSIONS: Increased urinary levels of GAG were associated with bladder inflammation in a PS-induced cystitis model. DMSO significantly reduced the inflammatory process after urothelial injury. Conversely, this drug provoked mild inflammation in normal mucosa. DMSO treatment was shown to influence urinary HA excretion.

  13. Effects of endothelial removal and regeneration on smooth muscle glycosaminoglycan synthesis and growth in rat carotid artery in organ culture

    Segments of rat carotid artery were maintained in serum-free and serum-supplemented media with endothelium both present and substantially removed by air drying. At intervals of 3, 7, and 14 days the synthesis of glycosaminoglycan across the vessel walls was determined by autoradiographic detection of incorporated [3H]glucosamine. In control carotids the typical pattern of incorporation was 40% of label in the intima, consisting of endothelium and subendothelial matrix, 23, 13, and 15% in the three medial layers (M1, M2, M3, respectively), and 9% in the adventitia. During the first week in culture the proportion, and often the amount, of label in M1 increased significantly. Following air drying labeling decreased markedly in M1 but often increased in M2 and M3. By 14 days residual endothelial cells had regenerated, and the pattern of incorporation in the medial layers beneath this new endothelium was the same as for the controls with a high level of labeling in M1. In areas free of endothelium incorporation in M1 remained at a low level. Digestion with chondroitinase ABC and Streptomyces hyaluronidase showed that the changes in M1-labeling levels were due to changes in the amounts of both hyaluronic acid and sulfated glycosaminoglycan, whereas pulse and continuous labeling studies showed that the different labeling levels for the various layers and conditions were due to different rates of synthesis and not degradation. Carotids were also labeled with [3H]thymidine. Control and regenerating endothelia were active in serum- free and serum-supplemented media and had similar mitotic indices. Indices for smooth muscle cells in M1, however, were generally very low and were not affected by the presence or absence of endothelium

  14. Effects of endothelial removal and regeneration on smooth muscle glycosaminoglycan synthesis and growth in rat carotid artery in organ culture

    Merrilees, M.J.; Scott, L.J.

    1985-04-01

    Segments of rat carotid artery were maintained in serum-free and serum-supplemented media with endothelium both present and substantially removed by air drying. At intervals of 3, 7, and 14 days the synthesis of glycosaminoglycan across the vessel walls was determined by autoradiographic detection of incorporated (/sup 3/H)glucosamine. In control carotids the typical pattern of incorporation was 40% of label in the intima, consisting of endothelium and subendothelial matrix, 23, 13, and 15% in the three medial layers (M1, M2, M3, respectively), and 9% in the adventitia. During the first week in culture the proportion, and often the amount, of label in M1 increased significantly. Following air drying labeling decreased markedly in M1 but often increased in M2 and M3. By 14 days residual endothelial cells had regenerated, and the pattern of incorporation in the medial layers beneath this new endothelium was the same as for the controls with a high level of labeling in M1. In areas free of endothelium incorporation in M1 remained at a low level. Digestion with chondroitinase ABC and Streptomyces hyaluronidase showed that the changes in M1-labeling levels were due to changes in the amounts of both hyaluronic acid and sulfated glycosaminoglycan, whereas pulse and continuous labeling studies showed that the different labeling levels for the various layers and conditions were due to different rates of synthesis and not degradation. Carotids were also labeled with (/sup 3/H)thymidine. Control and regenerating endothelia were active in serum- free and serum-supplemented media and had similar mitotic indices. Indices for smooth muscle cells in M1, however, were generally very low and were not affected by the presence or absence of endothelium.

  15. Effect of transforming growth factor beta on synthesis of glycosaminoglycans by human lung fibroblasts

    Dubaybo, B.A.; Thet, L.A. (Veterans Administration Medical Center, Allen Park, MI (USA))

    1990-09-01

    The processes of lung growth, injury, and repair are characterized by alterations in fibroblast synthesis and interstitial distribution of extracellular matrix components. Transforming growth factor beta (TGF-beta), which is postulated to play a role in modulating lung repair, alters the distribution of several matrix components such as collagen and fibronectin. We studied the effect of TGF-beta on the synthesis and distribution of the various glycosaminoglycans (GAGs) and whether these effects may explain its role in lung repair. Human diploid lung fibroblasts (IMR-90) were exposed to various concentrations of TGF-beta (0-5 nM) for variable periods of time (0-18 h). Newly synthesized GAGs were labeled with either (3H)glucosamine or (35S)sulfate. Individual GAGs were separated by size exclusion chromatography after serial enzymatic and chemical digestions and quantitated using scintillation counting. There was a dose-dependent increase in total GAG synthesis with maximal levels detected after 6 h of exposure. This increase was noted in all individual GAG types measured and was observed in both the cell associated GAGs (cell-matrix fraction) as well as the GAGs released into the medium (medium fraction). In the cell-matrix fraction, TGF-beta increased the proportion of heparan sulfate that was membrane bound as well as the proportion of dermatan sulfate in the intracellular compartment. In the medium fraction, TGF-beta increased the proportion of hyaluronic acid, chondroitin sulfate and dermatan sulfate released. We conclude that the role of TGF-beta in lung growth and repair may be related to increased synthesis of GAGs by human lung fibroblasts as well as alterations in the distribution of individual GAGs.

  16. Possible genetic defects in regulation of glycosaminoglycans in patients with diabetic nephropathy

    Deckert, T.; Horowitz, I.M.; Kofoed-Enevoldsen, A.; Kjellen, L.; Deckert, M.; Lykkelund, C.; Burcharth, F. (Steno Memorial Hospital, Gentofte (Denmark))

    1991-06-01

    The hypothesis of genetic defects in glycosaminoglycan (GAG) regulation among patients with insulin-dependent diabetes mellitus (IDDM) and nephropathy was assessed by studies in tissue cultures of fibroblasts obtained from 7 patients with normal urinary albumin excretion, 11 patients with diabetic nephropathy, and 6 nondiabetic control subjects. The incorporation of (2H) glucosamine and (35S) sulfate into hyaluronic acid (HA), chondroitin sulfate and dermatan sulfate (CS + DS), and heparan sulfate (HS) was measured in cells, matrix, and medium and related to micrograms of tissue protein. Large interindividual variations were seen in all three groups, and the incorporation of (3H) glucosamine into HA, CS + DS, and HS and (35S) sulfate into CS + DS and HS were not significantly different between the three groups. However, the fractional incorporation of (3H)glucosamine into HS was significantly reduced in diabetic patients with nephropathy compared with control subjects. This was the case not only when related to the total amount of GAGs (P = 0.014) but also when related to HA (P = 0.014). No significant difference was seen between control subjects and normoalbuminuric diabetic patients. The degree of N-sulfation of HS was not significantly different between the experimental groups. The results suggest that patients with diabetic nephropathy may suffer from deficiencies of coordinate regulation in the biosynthesis of GAG in fibroblasts, which may lead to a reduced density of HS in the extracellular matrix. If these changes reflect alterations in the biosynthesis of GAG from endothelial, myomedial, and mesangial cells, this observation may be relevant for the pathogenesis of severe diabetic complications.

  17. Effect of transforming growth factor beta on synthesis of glycosaminoglycans by human lung fibroblasts

    The processes of lung growth, injury, and repair are characterized by alterations in fibroblast synthesis and interstitial distribution of extracellular matrix components. Transforming growth factor beta (TGF-beta), which is postulated to play a role in modulating lung repair, alters the distribution of several matrix components such as collagen and fibronectin. We studied the effect of TGF-beta on the synthesis and distribution of the various glycosaminoglycans (GAGs) and whether these effects may explain its role in lung repair. Human diploid lung fibroblasts (IMR-90) were exposed to various concentrations of TGF-beta (0-5 nM) for variable periods of time (0-18 h). Newly synthesized GAGs were labeled with either [3H]glucosamine or [35S]sulfate. Individual GAGs were separated by size exclusion chromatography after serial enzymatic and chemical digestions and quantitated using scintillation counting. There was a dose-dependent increase in total GAG synthesis with maximal levels detected after 6 h of exposure. This increase was noted in all individual GAG types measured and was observed in both the cell associated GAGs (cell-matrix fraction) as well as the GAGs released into the medium (medium fraction). In the cell-matrix fraction, TGF-beta increased the proportion of heparan sulfate that was membrane bound as well as the proportion of dermatan sulfate in the intracellular compartment. In the medium fraction, TGF-beta increased the proportion of hyaluronic acid, chondroitin sulfate and dermatan sulfate released. We conclude that the role of TGF-beta in lung growth and repair may be related to increased synthesis of GAGs by human lung fibroblasts as well as alterations in the distribution of individual GAGs

  18. IL-8 dictates glycosaminoglycan binding and stability of IL-18 in cystic fibrosis.

    Reeves, Emer P

    2010-02-01

    Dysregulation of airway inflammation contributes to lung disease in cystic fibrosis (CF). Inflammation is mediated by inflammatory cytokines, including IL-8, which illustrates an increase in biological half-life and proinflammatory activity when bound to glycosaminoglycans (GAGs). The aim of this project was to compare IL-8 and IL-18 for their relative stability, activity, and interaction with GAGs, including chondroitin sulfate, hyaluronic acid, and heparan sulfate, present in high quantities in the lungs of patients with CF. Bronchoalveolar lavage fluid was collected from patients with CF (n = 28), non-CF controls (n = 14), and patients with chronic obstructive pulmonary disease (n = 12). Increased levels of IL-8 and reduced concentrations of IL-18 were detected in bronchial samples obtained from CF individuals. The low level of IL-18 was not a defect in IL-18 production, as the pro- and mature forms of the molecule were expressed and produced by CF epithelial cells and monocytes. There was, however, a marked competition between IL-8 and IL-18 for binding to GAGs. A pronounced loss of IL-18 binding capacity occurred in the presence of IL-8, which displaced IL-18 from these anionic-matrices, rendering the cytokine susceptible to proteolytic degradation by neutrophil elastase. As a biological consequence of IL-18 degradation, reduced levels of IL-2 were secreted by Jurkat T lymphocytes. In conclusion, a novel mechanism has been identified highlighting the potential of IL-8 to determine the fate of other inflammatory molecules, such as IL-18, within the inflammatory milieu of the CF lung.

  19. Isolation and characterization of the glycosaminoglycan component of rabbit thrombomodulin proteoglycan

    Bourin, M.C.; Lundgren-Akerlund, E.; Lindahl, U. (Swedish Univ. of Agricultural Sciences, Uppsala (Sweden))

    1990-09-15

    Previous studies on rabbit thrombomodulin (TM) revealed that certain anticoagulant activities expressed by TM depend on the presence of an acidic domain tentatively identified as a sulfated galactosaminoglycan. The glycan was released by alkaline beta-elimination, isolated by ion-exchange chromatography, and radiolabeled by partial N-deacetylation (hydrazinolysis) followed by re-N-(3H)acetylation. The labeled product behaved like standard chondroitin sulfate on ion-exchange chromatography, exhibited a Mr of 10-12 x 10(3) on gel chromatography, and was susceptible to degradation by chondroitinase and testicular hyaluronidase. The major labeled degradation products following digestion of the glycosaminoglycan with chondroitinase were identified, depending on the incubation conditions, either as 4/6-mono-O-sulfated, 4,5-unsaturated disaccharides (delta HexA-GalNAc)S and N-acetylgalactosamine 4,6-di-O-sulfate GalcNAc (diS), the latter component accounting for approximately 25% of the total label, or as a major fraction of labeled trisaccharide, with the predominant structure GalNAc(diS)-GlcA-GalNAc(diS). The terminal GalNAc(diS) unit (not substituted at C3) was shown to be more susceptible to N-deacetylation during hydrazinolysis than were the internal GalNAc units (substituted at C3), and thus was more extensively labeled, resulting in over-representation of this unit. It is concluded that rabbit TM is a chondroitin sulfate proteoglycan, which carries a single glycan side chain characterized by an unusual accumulation of sulfate groups at the nonreducing terminus. Metabolically 35S-labeled TM was isolated from cultured rabbit heart endothelial cells and characterized as a chondroitin sulfate proteoglycan which accounted for 1-2% of the total 35S-labeled cell-associated macromolecules.

  20. Possible genetic defects in regulation of glycosaminoglycans in patients with diabetic nephropathy

    The hypothesis of genetic defects in glycosaminoglycan (GAG) regulation among patients with insulin-dependent diabetes mellitus (IDDM) and nephropathy was assessed by studies in tissue cultures of fibroblasts obtained from 7 patients with normal urinary albumin excretion, 11 patients with diabetic nephropathy, and 6 nondiabetic control subjects. The incorporation of [2H] glucosamine and [35S] sulfate into hyaluronic acid (HA), chondroitin sulfate and dermatan sulfate (CS + DS), and heparan sulfate (HS) was measured in cells, matrix, and medium and related to micrograms of tissue protein. Large interindividual variations were seen in all three groups, and the incorporation of [3H] glucosamine into HA, CS + DS, and HS and [35S] sulfate into CS + DS and HS were not significantly different between the three groups. However, the fractional incorporation of [3H]glucosamine into HS was significantly reduced in diabetic patients with nephropathy compared with control subjects. This was the case not only when related to the total amount of GAGs (P = 0.014) but also when related to HA (P = 0.014). No significant difference was seen between control subjects and normoalbuminuric diabetic patients. The degree of N-sulfation of HS was not significantly different between the experimental groups. The results suggest that patients with diabetic nephropathy may suffer from deficiencies of coordinate regulation in the biosynthesis of GAG in fibroblasts, which may lead to a reduced density of HS in the extracellular matrix. If these changes reflect alterations in the biosynthesis of GAG from endothelial, myomedial, and mesangial cells, this observation may be relevant for the pathogenesis of severe diabetic complications

  1. Manganese supplementation enhances the synthesis of glycosaminoglycan in eggshell membrane: a strategy to improve eggshell quality in laying hens.

    Xiao, J F; Zhang, Y N; Wu, S G; Zhang, H J; Yue, H Y; Qi, G H

    2014-02-01

    This study investigated the effect of dietary Mn supplementation on eggshell quality, ultrastructure, glycosaminoglycan (GAG), and uronic acid content, and mRNA and protein expression of Galβ1,3-glucuronosyltransferase (GlcAT-I). A total of 216 layers (Hy-Line Grey) at age of 50 wk were divided into 3 groups. In the first 8 wk of the 12-wk feeding trial, all groups were fed a basal diet that met all layer nutrient requirements except for Mn. In the last 4 wk, each group was fed 1 of 3 diets supplemented with Mn levels at 0, 25, or 100 mg Mn/kg. Dietary Mn deficiency did not affect the egg performance of layers. Dietary Mn supplementation significantly improved the breaking strength, thickness, and fracture toughness of eggshells (P < 0.05). In photographs of eggshell ultrastructure, the size of mammillary cones and cracks in the outer surface were decreased by dietary Mn supplementation. The contents of GAG and uronic acids in eggshell membrane were significantly increased by dietary Mn addition (P < 0.05). This result was further confirmed by increased mRNA expression and protein expression of GlcAT-I when Mn was added to the diet. This study suggests that dietary Mn supplementation can improve eggshell quality by enhancing the GAG and uronic acid synthesis in the eggshell glands, which can affect the ultrastructure of eggshells. PMID:24570460

  2. Down-regulation of UDP-glucose dehydrogenase affects glycosaminoglycans synthesis and motility in HCT-8 colorectal carcinoma cells

    Wang, Tsung-Pao; Pan, Yun-Ru; Fu, Chien-Yu; Chang, Hwan-You, E-mail: hychang@life.nthu.edu.tw

    2010-10-15

    UDP-glucose dehydrogenase (UGDH) catalyzes oxidation of UDP-glucose to yield UDP-glucuronic acid, a precursor of hyaluronic acid (HA) and other glycosaminoglycans (GAGs) in extracellular matrix. Although association of extracellular matrix with cell proliferation and migration has been well documented, the importance of UGDH in these behaviors is not clear. Using UGDH-specific small interference RNA to treat HCT-8 colorectal carcinoma cells, a decrease in both mRNA and protein levels of UGDH, as well as the cellular UDP-glucuronic acid and GAG production was observed. Treatment of HCT-8 cells with either UGDH-specific siRNA or HA synthesis inhibitor 4-methylumbelliferone effectively delayed cell aggregation into multicellular spheroids and impaired cell motility in both three-dimensional collagen gel and transwell migration assays. The reduction in cell aggregation and migration rates could be restored by addition of exogenous HA. These results indicate that UGDH can regulate cell motility through the production of GAG. The enzyme may be a potential target for therapeutic intervention of colorectal cancers.

  3. Down-regulation of UDP-glucose dehydrogenase affects glycosaminoglycans synthesis and motility in HCT-8 colorectal carcinoma cells

    UDP-glucose dehydrogenase (UGDH) catalyzes oxidation of UDP-glucose to yield UDP-glucuronic acid, a precursor of hyaluronic acid (HA) and other glycosaminoglycans (GAGs) in extracellular matrix. Although association of extracellular matrix with cell proliferation and migration has been well documented, the importance of UGDH in these behaviors is not clear. Using UGDH-specific small interference RNA to treat HCT-8 colorectal carcinoma cells, a decrease in both mRNA and protein levels of UGDH, as well as the cellular UDP-glucuronic acid and GAG production was observed. Treatment of HCT-8 cells with either UGDH-specific siRNA or HA synthesis inhibitor 4-methylumbelliferone effectively delayed cell aggregation into multicellular spheroids and impaired cell motility in both three-dimensional collagen gel and transwell migration assays. The reduction in cell aggregation and migration rates could be restored by addition of exogenous HA. These results indicate that UGDH can regulate cell motility through the production of GAG. The enzyme may be a potential target for therapeutic intervention of colorectal cancers.

  4. Reduced supportive capacity of bone marrow stroma upon chemotherapy is mediated via changes in glycosaminoglycan profile

    Zweegman, Sonja; Kessler, Floortje L.; Kerkhoven, Ron M.; Heimerikx, Mike; Celie, Johanna W. A. M.; Janssen, Jeroen J. W. M.; Huijgens, Peter C.; Drager, Angelika M.; Van den Born, Jacob

    2007-01-01

    High dose chemotherapy and radiation have been found to impair the hematopoiesis-supportive capacity of bone marrow stroma. We now provide evidence for an important role of chemotherapy-induced alterations in stromal glycosaminoglycans (GAGs) in reduction of the supportive properties of stromal fibr

  5. Changes in the extracellular matrix and glycosaminoglycan synthesis during the initiation of regeneration in adult newt forelimbs

    Mescher, A.L.; Munaim, S.I.

    1986-04-01

    The extracellular matrix (ECM) of the distal tissues in a newt limb stump is completely reorganized in the 2-3-week period following amputation. In view of numerous in vitro studies showing that extracellular material influences cellular migration and proliferation, it is likely that the changes in the limb's ECM are important activities in the process leading to regeneration of such limbs. Using biochemical, autoradiographic, and histochemical techniques we studied temporal and spatial differences in the synthesis of glycosaminoglycans (GAGs) during the early, nerve-dependent phase of limb regeneration. Hyaluronic acid synthesis began with the onset of tissue dedifferentiation, became maximal within 1 weeks, and continued throughout the period of active cell proliferation. Chondroitin sulfate synthesis began somewhat later, increased steadily, and reached very high levels during chondrogenesis. During the first 10 days after amputation, distributions of sulfated and nonsulfated GAGs were both uniform throughout dedifferentiating tissues, except for a heavier localization near the bone. Since nerves are necessary to promote the regenerative process, we examined the neural influence on synthesis and accumulation of extracellular GAGs. Denervation decreased GAG production in all parts of the limb stump by approximately 50%. Newt dorsal root ganglia and brain-derived fibroblast growth factor each produced twofold stimulation of GAG synthesis in cultured 7-day regenerates. The latter effect was primarily on synthesis of hyaluronic acid. The results indicate that the trophic action of nerves on amphibian limb regeneration includes a positive influence on synthesis and extracellular accumulation of GAGs.

  6. Changes in the extracellular matrix and glycosaminoglycan synthesis during the initiation of regeneration in adult newt forelimbs

    The extracellular matrix (ECM) of the distal tissues in a newt limb stump is completely reorganized in the 2-3-week period following amputation. In view of numerous in vitro studies showing that extracellular material influences cellular migration and proliferation, it is likely that the changes in the limb's ECM are important activities in the process leading to regeneration of such limbs. Using biochemical, autoradiographic, and histochemical techniques we studied temporal and spatial differences in the synthesis of glycosaminoglycans (GAGs) during the early, nerve-dependent phase of limb regeneration. Hyaluronic acid synthesis began with the onset of tissue dedifferentiation, became maximal within 1 weeks, and continued throughout the period of active cell proliferation. Chondroitin sulfate synthesis began somewhat later, increased steadily, and reached very high levels during chondrogenesis. During the first 10 days after amputation, distributions of sulfated and nonsulfated GAGs were both uniform throughout dedifferentiating tissues, except for a heavier localization near the bone. Since nerves are necessary to promote the regenerative process, we examined the neural influence on synthesis and accumulation of extracellular GAGs. Denervation decreased GAG production in all parts of the limb stump by approximately 50%. Newt dorsal root ganglia and brain-derived fibroblast growth factor each produced twofold stimulation of GAG synthesis in cultured 7-day regenerates. The latter effect was primarily on synthesis of hyaluronic acid. The results indicate that the trophic action of nerves on amphibian limb regeneration includes a positive influence on synthesis and extracellular accumulation of GAGs

  7. Anticoagulant properties and cytotoxic effect against HCT116 human colon cell line of sulfated glycosaminoglycans isolated from the Norway lobster (Nephrops norvegicus) shell.

    Sayari, Nadhem; Balti, Rafik; Ben Mansour, Mohamed; Ben Amor, Ikram; Graiet, Imen; Gargouri, Jalel; Bougatef, Ali

    2016-05-01

    Sulfated glycosaminoglycans (SGNL) were extracted for the first time from Norway lobster (Nephrops norvegicus) shell. The monosaccharide composition analysed by GC/MS revealed the presence of galacturonic acid, glucuronic acid, N-acetylgalactosamine and N-acetylglucosamine. The analysis of SGNL with acetate cellulose electrophoresis in Zn-acetate revealed the presence of heparan sulfate (HS) and dermatan sulfate (DS). SGNL were evaluated for their anticoagulant activities using activated partial thromboplastin time (aPTT), thrombin time (TT) and prothrombine time (PT) tests. After 21h incubation, HCT116 cell proliferation was inhibited (psulfate content. SGNL demonstrated promising antiproliferative and anticoagulant potential, which may be used as a novel, effective and promising antithrombotic agent. PMID:27133072

  8. A Modified Glycosaminoglycan, GM-0111, Inhibits Molecular Signaling Involved in Periodontitis

    Savage, Justin R.; Pulsipher, Abigail; Rao, Narayanam V.; Kennedy, Thomas P.; Prestwich, Glenn D.; Ryan, Maria E.; Lee, Won Yong

    2016-01-01

    Background Periodontitis is characterized by microbial infection, inflammation, tissue breakdown, and accelerated loss of alveolar bone matrix. Treatment targeting these multiple stages of the disease provides ways to treat or prevent periodontitis. Certain glycosaminoglycans (GAGs) block multiple inflammatory mediators as well as suppress bacterial growth, suggesting that these GAGs may be exploited as a therapeutic for periodontitis. Methods We investigated the effects of a synthetic GAG, GM-0111, on various molecular events associated with periodontitis: growth of Porphyromonas gingivalis (P. gingivalis) and Aggregatibacter actinomycetemcomitans (A. actinomycetemcomitans) pathogenic bacteria associated with periodontitis; activation of pro-inflammatory signaling through TLR2 and TLR4 in mouse macrophage RAW 264.7 cells and heterologously expressed HEK 293 cells; osteoclast formation and bone matrix resorption in cultured mouse pre-osteoclasts. Results (1) GM-0111 suppressed the growth of P. gingivalis and A. actinomycetemcomitans even at 1% (w/v) solution. The antibacterial effects of GM-0111 were stronger than hyaluronic acid (HA) or xylitol in P. gingivalis at all concentrations and comparable to xylitol in A. actinomycetemcomitans at ≥2% (w/v) solution. We also observed that GM-0111 suppressed biofilm formation of P. gingivalis and these effects were much stronger than HA. (2) GM-0111 inhibited TLR-mediated pro-inflammatory cellular signaling both in macrophage and HEK 293 cells with higher selectivity for TLR2 than TLR4 (IC50 of 1–10 ng/mL vs. > 100 μg/mL, respectively). (3) GM-0111 blocked RANKL-induced osteoclast formation (as low as 300 ng/mL) and bone matrix resorption. While GM-0111 showed high affinity binding to RANKL, it did not interfere with RANKL/RANK/NF-κB signaling, suggesting that GM-0111 inhibits osteoclast formation by a RANKL-RANK-independent mechanism. Conclusions We report that GM-0111 inhibits multiple molecular events involved in

  9. A Modified Glycosaminoglycan, GM-0111, Inhibits Molecular Signaling Involved in Periodontitis.

    Justin R Savage

    Full Text Available Periodontitis is characterized by microbial infection, inflammation, tissue breakdown, and accelerated loss of alveolar bone matrix. Treatment targeting these multiple stages of the disease provides ways to treat or prevent periodontitis. Certain glycosaminoglycans (GAGs block multiple inflammatory mediators as well as suppress bacterial growth, suggesting that these GAGs may be exploited as a therapeutic for periodontitis.We investigated the effects of a synthetic GAG, GM-0111, on various molecular events associated with periodontitis: growth of Porphyromonas gingivalis (P. gingivalis and Aggregatibacter actinomycetemcomitans (A. actinomycetemcomitans pathogenic bacteria associated with periodontitis; activation of pro-inflammatory signaling through TLR2 and TLR4 in mouse macrophage RAW 264.7 cells and heterologously expressed HEK 293 cells; osteoclast formation and bone matrix resorption in cultured mouse pre-osteoclasts.(1 GM-0111 suppressed the growth of P. gingivalis and A. actinomycetemcomitans even at 1% (w/v solution. The antibacterial effects of GM-0111 were stronger than hyaluronic acid (HA or xylitol in P. gingivalis at all concentrations and comparable to xylitol in A. actinomycetemcomitans at ≥2% (w/v solution. We also observed that GM-0111 suppressed biofilm formation of P. gingivalis and these effects were much stronger than HA. (2 GM-0111 inhibited TLR-mediated pro-inflammatory cellular signaling both in macrophage and HEK 293 cells with higher selectivity for TLR2 than TLR4 (IC50 of 1-10 ng/mL vs. > 100 μg/mL, respectively. (3 GM-0111 blocked RANKL-induced osteoclast formation (as low as 300 ng/mL and bone matrix resorption. While GM-0111 showed high affinity binding to RANKL, it did not interfere with RANKL/RANK/NF-κB signaling, suggesting that GM-0111 inhibits osteoclast formation by a RANKL-RANK-independent mechanism.We report that GM-0111 inhibits multiple molecular events involved in periodontitis, spanning from the

  10. Small molecule inhibitors of protein interaction with glycosaminoglycans (SMIGs), a novel class of bioactive agents with anti-inflammatory properties

    Harris, N.; Kogan, F. Y.; Ilková, G.; Juhás, Štefan; Lahmy, O.; Gregor, Y. I.; Koppel, J.; Zhuk, R.; Gregor, P.

    2014-01-01

    Roč. 1840, č. 1 (2014), s. 245-254. ISSN 0304-4165 Institutional support: RVO:67985904 Keywords : heparan sulfate * heparin binding protein * glycosaminoglycan Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.381, year: 2014

  11. Otolaryngological findings in mucopolysaccharidosis

    Cingi, Cemal; MULUK, Nuray Bayar; Hancı, Deniz; Şahin, Ethem; Acar, Mustafa

    2014-01-01

    In this review paper, we reported otolaryngological problems in patients with mucopolysaccharidoses (MPSs). Mucopolysaccharidoses are a group of lysosomal storage diseases, each of which is produced by an inherited deficiency of an enzyme involved in the degradation of acid mucopolysaccharides, now called glycosaminoglycans (GAGs). The mucopolysaccharidoses consist of a group of 7 metabolic disorders, known as mucopolysaccharidoses types I-VII. In all groups, there are clinical and otolaryng...

  12. Validation of Urinary Glycosaminoglycans in Iranian patients with Mucopolysaccharidase type I: The effect of urine sedimentation characteristics

    Abdi, Mohammad; Hakhamaneshi, Mohammad Said; Alaei, Mohammad Reza; AZADI, Namam-Ali; Vakili, Rahim; Zamanfar, Daniel; Taghikhani, Mohammad; KHATAMI, Shohreh

    2014-01-01

    How to Cite This Article:Abdi M, Khatami Sh, Hakhamaneshi MS, Alaei MR, Azadi NA, Zamanfar D, Taghikhani M.Validation of Urinary Glycosaminiglycans in Iranian Patients with Mucopolysaccharidose Type I: The Effect of Urine Sedimentation Characteristics. Iran J Child Neurol. 2014; 8(4):39-45. AbstractObjectiveThe first line-screening test for mucopolysaccharidosis is based on measurement of urinary glycosaminoglycans. The most reliable test for measurement of urine glycosaminoglycans is the 1,9...

  13. Binding and Clustering of Glycosaminoglycans: A Common Property of Mono- and Multivalent Cell-Penetrating Compounds

    Ziegler, André; Seelig, Joachim

    2007-01-01

    Recent observations in cell culture provide evidence that negatively charged glycosaminoglycans (GAGs) at the surface of biological cells bind cationic cell-penetrating compounds (CPCs) and cluster during CPC binding, thereby contributing to their endocytotic uptake. The GAG binding and clustering occur in the low-micromolar concentration range and suggest a tight interaction between GAGs and CPCs, although the relation between binding affinity and specificity of this interaction remains to b...

  14. Glycosaminoglycan-Mediated Selective Changes in the Aggregation States, Zeta Potentials, and Intrinsic Stability of Liposomes

    Nyren-Erickson, Erin K; Haldar, Manas K.; Totzauer, Jessica R.; Ceglowski, Riley; Patel, Dilipkumar S.; Daniel L. Friesner; Srivastava, D. K.; Mallik, Sanku

    2012-01-01

    Though the aggregation of glycosaminoglycans (GAGs) in the presence of liposomes and divalent cations has been previously reported, the effect of different GAG species, as well as minor changes in GAG composition on the aggregates formed is yet unknown. If minor changes in GAG composition produce observable changes in liposome aggregate diameter or zeta potential, such a phenomenon may be used to detect potentially dangerous over-sulfated contaminants in heparin. We studied the mechanism of t...

  15. Glycosaminoglycan-mediated selective changes in the aggregation states, zeta potentials, and intrinsic stability of liposomes.

    Nyren-Erickson, Erin K; Haldar, Manas K; Totzauer, Jessica R; Ceglowski, Riley; Patel, Dilipkumar S; Friesner, Daniel L; Srivastava, D K; Mallik, Sanku

    2012-11-20

    Though the aggregation of glycosaminoglycans (GAGs) in the presence of liposomes and divalent cations has been previously reported, the effects of different GAG species and minor changes in GAG composition on the aggregates that are formed are yet unknown. If minor changes in GAG composition produce observable changes in the liposome aggregate diameter or zeta potential, such a phenomenon may be used to detect potentially dangerous oversulfated contaminants in heparin. We studied the mechanism of the interactions between heparin and its oversulfated glycosaminoglycan contaminants with liposomes. Herein, we demonstrate that Mg(2+) acts to shield the incoming glycosaminoglycans from the negatively charged phosphate groups of the phospholipids and that changes in the aggregate diameter and zeta potential are a function of the glycosaminoglycan species and concentration as well as the liposome bilayer composition. These observations are supported by TEM studies. We have shown that the organizational states of the liposome bilayers are influenced by the presence of GAG and excess Mg(2+), resulting in a stabilizing effect that increases the T(m) value of DSPC liposomes; the magnitude of this effect is also dependent on the GAG species and concentration present. There is an inverse relationship between the percent change in aggregate diameter and the percent change in aggregate zeta potential as a function of GAG concentration in solution. Finally, we demonstrate that the diameter and zeta potential changes in POPC liposome aggregates in the presence of different oversulfated heparin contaminants at low concentrations allow for an accurate detection of oversulfated chondroitin sulfate at concentrations of as low as 1 mol %. PMID:23102026

  16. Glycosaminoglycan Binding Facilitates Entry of a Bacterial Pathogen into Central Nervous Systems

    Chang, Yung-Chi; Wang, Zhipeng; Flax, Lindsay A.; Xu, Ding; Esko, Jeffrey D.; Nizet, Victor; Baron, Miriam J.

    2011-01-01

    Certain microbes invade brain microvascular endothelial cells (BMECs) to breach the blood-brain barrier (BBB) and establish central nervous system (CNS) infection. Here we use the leading meningitis pathogen group B Streptococcus (GBS) together with insect and mammalian infection models to probe a potential role of glycosaminoglycan (GAG) interactions in the pathogenesis of CNS entry. Site-directed mutagenesis of a GAG-binding domain of the surface GBS alpha C protein impeded GBS penetration ...

  17. Lysyl oxidase activity and elastin/glycosaminoglycan interactions in growing chick and rat aortas

    1987-01-01

    Hydrophobic tropoelastin molecules aggregate in vitro in physiological conditions and form fibers very similar to natural ones (Bressan, G. M., I. Pasquali Ronchetti, C. Fornieri, F. Mattioli, I. Castellani, and D. Volpin, 1986, J. Ultrastruct. Molec. Struct. Res., 94:209-216). Similar hydrophobic interactions might be operative in in vivo fibrogenesis. Data are presented suggesting that matrix glycosaminoglycans (GAGs) prevent spontaneous tropoelastin aggregation in vivo, at least up to the ...

  18. A Modified Glycosaminoglycan, GM-0111, Inhibits Molecular Signaling Involved in Periodontitis

    Savage, Justin R.; Pulsipher, Abigail; Rao, Narayanam V.; Thomas P Kennedy; Glenn D Prestwich; Ryan, Maria E.; Lee, Won Yong

    2016-01-01

    Background Periodontitis is characterized by microbial infection, inflammation, tissue breakdown, and accelerated loss of alveolar bone matrix. Treatment targeting these multiple stages of the disease provides ways to treat or prevent periodontitis. Certain glycosaminoglycans (GAGs) block multiple inflammatory mediators as well as suppress bacterial growth, suggesting that these GAGs may be exploited as a therapeutic for periodontitis. Methods We investigated the effects of a synthetic GAG, G...

  19. High-performance liquid chromatography-mass spectrometry for mapping and sequencing glycosaminoglycan-derived oligosaccharides

    Volpi, Nicola; Linhardt, Robert J.

    2010-01-01

    Glycosaminoglycans (GAGs) have proven to be very difficult to analyze and characterize because of their high negative charge density, polydispersity and sequence heterogeneity. As the specificity of the interactions between GAGs and proteins results from the structure of these polysaccharides, an understanding of GAG structure is essential for developing a structure–activity relationship. Electrospray ionization (ESI) mass spectrometry (MS) is particularly promising for the analysis of oligos...

  20. Structural Requirements for Glycosaminoglycan Recognition by the Lyme Disease Spirochete, Borrelia burgdorferi†

    Leong, John M.; Robbins, Douglas; Rosenfeld, Louis; Lahiri, Biswajit; Parveen, Nikhat

    1998-01-01

    Borrelia burgdorferi, the Lyme disease agent, binds glycosaminoglycans (GAGs) such as heparin, heparan sulfate, and dermatan sulfate. Heparin or heparan sulfate fractions separated by size or charge were tested for their ability to inhibit attachment of B. burgdorferi to Vero cells. GAG chains of increasing length and/or charge showed increasing inhibitory potency, and detectable heparin inhibition of bacterial binding required a minimum of 16 residues. The ability of a given heparin fraction...

  1. Pattern-Based Sensing of Sulfated Glycosaminoglycans with a Dynamic Mixture of Iron Complexes

    Müller-Graff, Peter-Korbinian; Szelke, Helga; Severin, Kay; Krämer, Roland

    2010-01-01

    A dynamic mixture of iron complexes was used as a colorimetric sensor for sulfated glycosaminoglycans. The sensing ensemble was prepared by mixing [FeCl2(H2O)4] with dipicolylamine, the functionalized bipyridyl ligand N-(6-aminohexyl)-4'-methyl-2,2'-bipyridine-4-carboxamide, and the dye Evans Blue. Upon addition of the analytes, characteristic changes in the UV-Vis spectrum of the solutions were observed. The spectral changes allowed indentifying different sulfated glucosaminoglycans (unfr...

  2. Human Genetic Disorders Caused by Mutations in Genes Encoding Biosynthetic Enzymes for Sulfated Glycosaminoglycans*

    Mizumoto, Shuji; Ikegawa, Shiro; Sugahara, Kazuyuki

    2013-01-01

    A number of genetic disorders are caused by mutations in the genes encoding glycosyltransferases and sulfotransferases, enzymes responsible for the synthesis of sulfated glycosaminoglycan (GAG) side chains of proteoglycans, including chondroitin sulfate, dermatan sulfate, and heparan sulfate. The phenotypes of these genetic disorders reflect disturbances in crucial biological functions of GAGs in human. Recent studies have revealed that mutations in genes encoding chondroitin sulfate and derm...

  3. Effects of Immobilized Glycosaminoglycans on the Proliferation and Differentiation of Mesenchymal Stem Cells

    Uygun, Basak E.; Stojsih, Sarah E.; Matthew, Howard W.T.

    2009-01-01

    Mesenchymal stem cells (MSCs) are adult stem cells with potential for multilineage differentiation. They represent an attractive cell source alternative to embryonic stem cells for therapeutic applications. Optimal utilization of MSCs for tissue engineering requires improved biomaterials that can enhance their growth and direct differentiation. The biological activity of glycosaminoglycans (GAGs) has been previously exploited for use in tissue engineering applications. In this study, MSC prol...

  4. Strain Variation in Glycosaminoglycan Recognition Influences Cell-Type-Specific Binding by Lyme Disease Spirochetes

    Parveen, Nikhat; Robbins, Douglas; Leong, John M.

    1999-01-01

    Lyme disease, a chronic multisystemic disorder that can affect the skin, heart, joints, and nervous system is caused by Borrelia burgdorferi sensu lato. Lyme disease spirochetes were previously shown to bind glycosaminoglycans (GAGs). In the current study, the GAG-binding properties of eight Lyme disease strains were determined. Binding by two high-passage HB19 derivatives to Vero cells could not be inhibited by enzymatic removal of GAGs or by the addition of exogenous GAG. The other six stra...

  5. Linear polyalkylamines as fingerprinting agents in capillary electrophoresis of low molecular weight heparins and glycosaminoglycans

    King, J. Timothy; Desai, Umesh R.

    2011-01-01

    Glycosaminoglycan (GAG) analysis represents a challenging frontier despite the advent of many high resolution technologies because of their unparalleled structural complexity. We previously developed a resolving agent aided capillary electrophoretic approach for fingerprinting low molecular weight heparins (LMWHs) to profile their microscopic differences and assess batch-to-batch variability. In this work, we study the application of this approach for fingerprinting other GAGs and analyze the...

  6. Exogenous glycosaminoglycans coat damaged bladder surfaces in experimentally damaged mouse bladder

    Hurst Robert E; Coffman Jean; Kyker Kimberly D

    2005-01-01

    Abstract Background Interstital cystitis is often treated with exogenous glycosaminoglycans such as heparin, chondroitin sulphate (Uracyst), hyaluronate (Cystistat) or the semi-synthetic pentosan polysulphate (Elmiron). The mechanism of action is presumed to be due to a coating of the bladder surface to replace the normally present chondroitin sulphate and heparan sulphate lost as a result of the disease. This study used fluorescent labelled chondroitin sulphate to track the distribution of g...

  7. Study of Glycosaminoglycans of Extracellular Matrix (ECM) in Pulp of Developing Tooth

    Kermany T; AR. Varasteh; Nicravesh MR; oradi M

    2000-01-01

    Mesenchymal- epithelial interactions during embryogenesis have been shown to be important in the fetal development of many organs. Identification of molecules that modulate these interactions is key to our understanding of the pathological conditions. The major groups of extracellular matrix (ECM) molecules characterized are glycosaminoglycans that candidate for morphogenesis and differentiation of ceils and tissues. In this study the molecules of ECM were considered in tooth development, pre...

  8. Exopolysaccharides produced by marine bacteria and their applications as glycosaminoglycan-like molecules

    Delbarre-Ladrat, Christine; Sinquin, Corinne; Lebellenger, Lou; Zykwinska, Agata; Colliec-Jouault, Sylvia

    2014-01-01

    Although polysaccharides are ubiquitous and the most abundant renewable bio-components, their studies, covered by the glycochemistry and glycobiology fields, remain a challenge due to their high molecular diversity and complexity. Polysaccharides are industrially used in food products; human therapeutics fall into a more recent research field and pharmaceutical industry is looking for more and more molecules with enhanced activities. Glycosaminoglycans (GAGs) found in animal tissues play a cr...

  9. Glycosaminoglycans in extracts of cardiac amyloid fibrils from familial amyloid cardiomyopathy of Danish origin related to variant transthyretin Met 111.

    Magnus, J H; Stenstad, T; Kolset, S O; Husby, G

    1991-07-01

    We have previously demonstrated an association between secondary AA type amyloid fibrils and glycosaminoglycans (GAGs) in human liver. The present study was aimed at investigating whether a similar association could be demonstrated in isolated cardiac amyloid fibrils from a unique Danish family with amyloid cardiomyopathy related to variant transthyretin (TTR) with a single amino acid substitution of a methionin for leucine at position 111 (TTR Met 111). Using gel filtration and ion exchange chromatography, significant amounts of GAGs were detected in close association with purified myocardial amyloid fibrils, whereas only trace amounts of polysaccharides were present in the corresponding normal preparation. The GAGs were identified as 50% chondroitin sulfate, 33% heparin/heparan sulfate, and 17% hyaluronan. With the methods used the amyloid associated GAGs appeared as high molecular weight free polysaccharide chains, and not as part of intact proteoglycans (PGs) in the fibril extracts. We conclude that the association between purified amyloid fibrils and GAGs may be a general feature of amyloid deposits. Also, we suggest that the proportion of different GAGs in the amyloid deposits may depend both on the organ or tissues affected and the type of proteins making up the fibrils. PMID:2068532

  10. Stimulation of sulfated glycosaminoglycan synthesis by the tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu2+.

    Wegrowski, Y; Maquart, F X; Borel, J P

    1992-01-01

    Glycyl-L-histidyl-L-lysine-copper (II) complex (GHK-Cu) is a naturally occurring tripeptide with potential healing properties. We studied the effect of GHK-Cu on the synthesis of glycosaminoglycans (GAGs) by normal human fibroblasts in culture. Cells were incubated with 3H glucosamine and 35S sulfate and the radioactivity of isolated GAGs was determined. GHK-Cu induced a dose-dependent increase of the synthesis of total GAGs secreted into the culture medium and those associated with the cell layer. The effect of GHK-Cu was biphasic with a maximal stimulation at 10(-9) to 10(-8) M. At higher concentrations, the rate of synthesis returned progressively to that of control cultures. Electrophoretic analysis of the different GAG populations showed that GHK-Cu preferentially stimulated the synthesis of extracellular dermatan sulfate and cell layer associated heparan sulfate. No influence of GHK-Cu on the synthesis of hyaluronic acid was observed. GHK-Cu stimulation of GAG synthesis may be one of the phenomenons implicated in the wound healing properties of the peptide. PMID:1522753

  11. Glycosaminoglycans affect the interaction of human plasma kallikrein with plasminogen, factor XII and inhibitors

    Gozzo A.J.

    2003-01-01

    Full Text Available Human plasma kallikrein, a serine proteinase, plays a key role in intrinsic blood clotting, in the kallikrein-kinin system, and in fibrinolysis. The proteolytic enzymes involved in these processes are usually controlled by specific inhibitors and may be influenced by several factors including glycosaminoglycans, as recently demonstrated by our group. The aim of the present study was to investigate the effect of glycosaminoglycans (30 to 250 µg/ml on kallikrein activity on plasminogen and factor XII and on the inhibition of kallikrein by the plasma proteins C1-inhibitor and antithrombin. Almost all available glycosaminoglycans (heparin, heparan sulfate, bovine and tuna dermatan sulfate, chondroitin 4- and 6-sulfates reduced (1.2 to 3.0 times the catalytic efficiency of kallikrein (in a nanomolar range on the hydrolysis of plasminogen (0.3 to 1.8 µM and increased (1.9 to 7.7 times the enzyme efficiency in factor XII (0.1 to 10 µM activation. On the other hand, heparin, heparan sulfate, and bovine and tuna dermatan sulfate improved (1.2 to 3.4 times kallikrein inhibition by antithrombin (1.4 µM, while chondroitin 4- and 6-sulfates reduced it (1.3 times. Heparin and heparan sulfate increased (1.4 times the enzyme inhibition by the C1-inhibitor (150 nM.

  12. Exogenous glycosaminoglycans coat damaged bladder surfaces in experimentally damaged mouse bladder

    Hurst Robert E

    2005-03-01

    Full Text Available Abstract Background Interstital cystitis is often treated with exogenous glycosaminoglycans such as heparin, chondroitin sulphate (Uracyst, hyaluronate (Cystistat or the semi-synthetic pentosan polysulphate (Elmiron. The mechanism of action is presumed to be due to a coating of the bladder surface to replace the normally present chondroitin sulphate and heparan sulphate lost as a result of the disease. This study used fluorescent labelled chondroitin sulphate to track the distribution of glycosaminoglycans administered intravesically to mouse bladder that had been damaged on the surface. Methods The surfaces of mouse bladders were damaged by 3 mechanisms – trypsin, 10 mM HCl, and protamine sulphate. Texas Red-labeled chondroitin sulphate was instilled into the bladders of animals with damaged bladders and controls instilled only with saline. Bladders were harvested, frozen, and sectioned for examination by fluorescence. Results The normal mouse bladder bound a very thin layer of the labelled chondroitin sulphate on the luminal surface. Trypsin- and HCl-damaged bladders bound the labelled chondroitin sulphate extensively on the surface with little penetration into the bladder muscle. Protamine produced less overt damage, and much less labelling was seen, presumably due to loss of the label as it complexed with the protamine intercalated into the bladder surface. Conclusion Glycosaminoglycan administered intravesically does bind to damaged bladder. Given that the changes seen following bladder damage resemble those seen naturally in interstitial cystitis, the mechanisms proposed for the action of these agents is consistent with a coating of damaged bladder.

  13. Interactions of bovine viral diarrhoea virus glycoprotein E(rns) with cell surface glycosaminoglycans.

    Iqbal, M; Flick-Smith, H; McCauley, J W

    2000-02-01

    Recombinant E(rns) glycoprotein of bovine viral diarrhoea virus (BVDV) has been tagged with a marker epitope or linked to an immunoglobulin Fc tail and expressed in insect and mammalian cell lines. The product was shown to be functional, both having ribonuclease activity and binding to a variety of cells that were permissive and non-permissive for replication of BVDV. Addition of soluble E(rns) to the medium blocked replication of BVDV in permissive cells. Binding of epitope-tagged E(rns) to permissive calf testes (CTe) cells was abolished and virus infection was reduced when cells were treated with heparinases I or III. E(rns) failed to bind to mutant Chinese hamster ovary (CHO) cells that lacked glycosaminoglycans (pgsA-745 cells) or heparan sulphate (pgsD-677 cells) but bound to normal CHO cells. E(rns) also bound to heparin immobilized on agarose and could be eluted by heparin and by a high concentration of salt. Flow cytometric analysis of E(rns) binding to CTe cell cultures showed that glycosaminoglycans such as heparin, fucoidan and dermatan sulphate all inhibit binding but dextran sulphate, keratan sulphate, chondroitin sulphate and mannan fail to inhibit binding. The low molecular mass polysulphonated inhibitor suramin also inhibited binding to CTe cells but poly-L-lysine did not. Furthermore, suramin, the suramin analogue CPD14, fucoidan and pentosan polysulphate inhibited the infectivity of virus. It is proposed that binding of E(rns) to cells is through an interaction with glycosaminoglycans and that BVDV may bind to cells initially through this interaction. PMID:10644844

  14. Anti-inflammatory Effect of Isaria sinclairii Glycosaminoglycan in an Adjuvant-treated Arthritis Rat Model

    Ahn, Mi Young; Jee, Sang Duck; Hwang, Jae Sam; Yun, Eun Young; Ahn, Kwang Seok; Kim, Yeong Shik

    2013-01-01

    The anti-inflammatory effects of glycosaminoglycan (GAG) derived from Isaria sinclairii (IS) and of IS extracts were investigated in a complete Freund’s adjuvant (CFA)-treated chronic arthritis rat model. Groups of rats were treated orally with 30 mg/kg one of the following: [1] saline control, extracts of [2] water-IS, [3] methanol-IS, [4] butanol-IS, [5] ethyl acetate-IS, or [6] Indomethacin® as the positive control for a period of two weeks. The anti-paw edema effects of the individual ext...

  15. 多磺酸粘多糖治疗面部激素依赖性皮炎临床观察%Clinical observation of Mucopolysaccharide Polysulphate Cream in the treatment of facial corticosteroid addictive dermatitis

    陈香梅

    2015-01-01

    Objective: To evaluate the therapy efficacy and safety of Mucopolysacde Polysulphate Cream and Hydro-cortisone on facial corticosteroid addictive dermatitis. Me thods:98 patients with mild to moderate facial corticosteroid addictive dermatitis were selected and randomly divided into two groups: one was treated with Mucopolysaccharide Polysulphate Cream (the treatment group), the other one was treated with Hydrocortisone cream (the control group).The cream was applied 2 times per day for 4 weeks continuously, the efficacy and adverse events were evaluated after drug withdrawal. At the 4 weeks after stopping drugs, the recovery of the primary lesions was observed and recorded as well as the recurrence rate. Results:83 patients completed the treatment and the follow-up, including 45 cases in Mu-copolysaccharide Polysulphate group and 38 cases in Hydrocortisone group;The effective rate in the treatment group was 71.11%, and 76.32%in the control group, there was no significant difference between two groups (P>0. 05). There was significant difference about the incidence rate of adverse events between two groups(P0.05);不良反应发生率治疗组为28.89%,对照组为18.42%,两组差异有显著性(P<0.05);在随后第4周的随访中,复发率分别是治疗组为11.11%,对照组为42.11%,两组差异有显著性(P<0.05)。结论:与丁酸氢化可的松比较,多磺酸粘多糖对轻中度面部激素依赖性皮炎的疗效无明显差异,但是其可减少面部激素依赖性皮炎的复发率。

  16. The Immunosuppressant FTY720 (Fingolimod enhances Glycosaminoglycan depletion in articular cartilage

    Stradner Martin H

    2011-12-01

    Full Text Available Abstract Background FTY720 (Fingolimod is a novel immunosuppressive drug investigated in clinical trials for organ transplantation and multiple sclerosis. It acts as a functional sphingosine-1-phosphate (S1P receptor antagonist, thereby inhibiting the egress of lymphocytes from secondary lymphoid organs. As S1P is able to prevent IL-1beta induced cartilage degradation, we examined the direct impact of FTY720 on cytokine induced cartilage destruction. Methods Bovine chondrocytes were treated with the bioactive phosphorylated form of FTY720 (FTY720-P in combination with IL-1beta or TNF-alpha. Expression of MMP-1,-3.-13, iNOS and ADAMTS-4,-5 and COX-2 was evaluated using quantitative real-time PCR and western blot. Glycosaminoglycan depletion from cartilage explants was determined using a 1,9-dimethylene blue assay and safranin O staining. Results FTY720-P significantly reduced IL-1beta and TNF-alpha induced expression of iNOS. In contrast FTY720-P increased MMP-3 and ADAMTS-5 mRNA expression. Furthermore depletion of glycosaminoglycan from cartilage explants by IL-1beta and TNF-alpha was significantly enhanced by FTY720-P in an MMP-3 dependent manner. Conclusions Our results suggest that FTY720 may enhance cartilage degradation in pro-inflammatory environment.

  17. Nanocoating of titanium implant surfaces with organic molecules. Polysaccharides including glycosaminoglycans.

    Gurzawska, Katarzyna; Svava, Rikke; Jørgensen, Niklas Rye; Gotfredsen, Klaus

    2012-12-01

    Long-term stability of titanium implants are dependent on a variety of factors. Nanocoating with organic molecules is one of the method used to improve osseointegration. Nanoscale modification of titanium implants affects surface properties, such as hydrophilicity, biochemical bonding capacity and roughness. This influences cell behaviour on the surface such as adhesion, proliferation and differentiation of cells as well as the mineralization of the extracellular matrix at the implant surfaces. The aim of the present systematic review was to describe organic molecules used for surface nanocoating with focus on polysaccharides including glycosaminoglycans, and how these molecules change surface properties, cell reactions and affect on osseointegartion. The included in vitro studies demonstrated increased cell adhesion, proliferation and mineralization of a number of the tested polysaccharide nanocoatings. The included in vivo studies, showed improvement of bone interface reactions measured as increased Bone-to-Implant Contact length and Bone Mineral Density adjacent to the polysaccharide coated surfaces. Based on existing literature, surface modification with polysaccharide and glycosaminoglycans appears to be an effective way to stimulate bone regeneration on bone-implant interface. PMID:23030010

  18. Glycosaminoglycan-bound and free inter-alpha-trypsin inhibitor components of follicular fluid.

    Odum, L; Jessen, T E; Andersen, C Y

    2001-11-01

    The proteinase inhibitor inter-alpha trypsin inhibitor (ITI) is a blood-derived protein necessary for normal female fertility. Absence of ITI leads to ovulation of naked oocytes that cannot fertilise. ITI consists of two heavy chains (ITI-HC) and bikunin linked by a chrondroitin sulphate. By binding to hyaluronate, ITI-HC stabilises the extracellular matrix, but ITI-HC also binds to proteoglycans in follicular fluid. In vivo concentrations of ITI components in preovulatory follicular fluid, free as well as bound to hyaluronate or proteoglycan, are unknown. In order to quantify these components, 58 follicular fluids and 13 blood samples were collected in connection with in vitro fertilisation and embryo transfer treatment of 13 women. Quantitation of glycosaminoglycan-bound ITI-HC was performed after separation from free ITI in agarose gel. ITI components were determined by immunoelectrophoresis and hyaluronate by an ELISA method. The follicular fluid concentration of ITI was on average 70% of that in plasma and the concentration of hyaluronate remained low despite follicular production, suggesting that the production of hyaluronate is the rate-limiting step in the formation of the extracellular matrix of the oocyte-cumulus complex. In follicular fluid, the concentration of free ITI-HC was higher than that of glycosaminoglycan-bound ITI-HC. Addition of exogeneous hyaluronate doubled the amount of hyaluronate-bound ITI-HC, further supporting the notion that ITI in follicular fluid is not rate-limiting for cumulus expansion in vivo. PMID:11771893

  19. An oral nutraceutical containing antioxidants, minerals and glycosaminoglycans improves skin roughness and fine wrinkles.

    Udompataikul, M; Sripiroj, P; Palungwachira, P

    2009-12-01

    Various nutraceuticals (dietary supplements) are claimed to have cutaneous antiageing properties, however, there are a limited number of research studies supporting these claims. The objective of this research was to study the effectiveness of an oral nutraceutical containing antioxidants, minerals and glycosaminoglycans on cutaneous ageing. In this double-blind, placebo-controlled trial, 60 women aged 35-60 years were randomized to receive oral dietary supplement (n = 30) or placebo (n = 30), once daily for 12 weeks. The depth of skin roughness and fine wrinkles were measured using surface evaluation of skin parameters for living skin (Visioscan) at baseline, and at the 4, 8 and 12 weeks of treatment. Surface evaluation using a replica film (Visiometer) at baseline and at the 12th week of treatment was also carried out. Statistical differences in objective skin improvement were assessed by the independent t-test. The volunteers' satisfaction was tested using the chi-squared test. The baseline depth of skin roughness and fine wrinkles in the treatment group and the placebo group were 100.5 and 100 mum, respectively. At the end of the study, the depth of skin roughness and fine wrinkles in the treatment group showed a 21.2% improvement, whereas improvement in the control group was 1.7%. This difference was statistically significant (P minerals and glycosaminoglycans improved skin roughness and fine wrinkles but did not affect skin colour change in female volunteers. PMID:19570098

  20. Benzylidene Acetal Protecting Group as Carboxylic Acid Surrogate: Synthesis of Functionalized Uronic Acids and Sugar Amino Acids.

    Banerjee, Amit; Senthilkumar, Soundararasu; Baskaran, Sundarababu

    2016-01-18

    Direct oxidation of the 4,6-O-benzylidene acetal protecting group to C-6 carboxylic acid has been developed that provides an easy access to a wide range of biologically important and synthetically challenging uronic acid and sugar amino acid derivatives in good yields. The RuCl3 -NaIO4 -mediated oxidative cleavage method eliminates protection and deprotection steps and the reaction takes place under mild conditions. The dual role of the benzylidene acetal, as a protecting group and source of carboxylic acid, was exploited in the efficient synthesis of six-carbon sialic acid analogues and disaccharides bearing uronic acids, including glycosaminoglycan analogues. PMID:26572799

  1. The influence of the type of sulphate bond and degree of sulphation of glycosaminoglycans on their interaction with lysosomal enzymes.

    Avila, J L

    1978-01-01

    Significant differences occur between the interaction of several sulphated glycosaminoglycans with a particular lysosomal protein, leading to inhibition in the case of lysosomal enzymes. The order of strength of inhibition at pH4 was: heparin greater than chondroitin 4-sulphate = chondroitin 6-sulphate greater than dermatan sulphate. PMID:656058

  2. LL-37 complexation with glycosaminoglycans in cystic fibrosis lungs inhibits antimicrobial activity, which can be restored by hypertonic saline.

    Bergsson, Gudmundur

    2009-07-01

    There is an abundance of antimicrobial peptides in cystic fibrosis (CF) lungs. Despite this, individuals with CF are susceptible to microbial colonization and infection. In this study, we investigated the antimicrobial response within the CF lung, focusing on the human cathelicidin LL-37. We demonstrate the presence of the LL-37 precursor, human cathelicidin precursor protein designated 18-kDa cationic antimicrobial protein, in the CF lung along with evidence that it is processed to active LL-37 by proteinase-3. We demonstrate that despite supranormal levels of LL-37, the lung fluid from CF patients exhibits no demonstrable antimicrobial activity. Furthermore Pseudomonas killing by physiological concentrations of exogenous LL-37 is inhibited by CF bronchoalveolar lavage (BAL) fluid due to proteolytic degradation of LL-37 by neutrophil elastase and cathepsin D. The endogenous LL-37 in CF BAL fluid is protected from this proteolysis by interactions with glycosaminoglycans, but while this protects LL-37 from proteolysis it results in inactivation of LL-37 antimicrobial activity. By digesting glycosaminoglycans in CF BAL fluid, endogenous LL-37 is liberated and the antimicrobial properties of CF BAL fluid restored. High sodium concentrations also liberate LL-37 in CF BAL fluid in vitro. This is also seen in vivo in CF sputum where LL-37 is complexed to glycosaminoglycans but is liberated following nebulized hypertonic saline resulting in increased antimicrobial effect. These data suggest glycosaminoglycan-LL-37 complexes to be potential therapeutic targets. Factors that disrupt glycosaminoglycan-LL-37 aggregates promote the antimicrobial effects of LL-37 with the caveat that concomitant administration of antiproteases may be needed to protect the now liberated LL-37 from proteolytic cleavage.

  3. The involvement of glycosaminoglycans in airway disease associated with cystic fibrosis.

    Reeves, Emer P

    2012-02-01

    Individuals with cystic fibrosis (CF) present with severe airway destruction and extensive bronchiectasis. It has been assumed that these structural airway changes have occurred secondary to infection and inflammation, but recent studies suggest that glycosaminoglycan (GAG) remodelling may be an important independent parallel process. Evidence is accumulating that not only the concentration, but also sulphation of GAGs is markedly increased in CF bronchial cells and tissues. Increased expression of GAGs and, in particular, heparan sulphate, has been linked to a sustained inflammatory response and neutrophil recruitment to the CF airways. This present review discusses the biological role of GAGs in the lung, as well as their involvement in CF respiratory disease, and their potential as therapeutic targets.

  4. A Dilemma in the Glycosaminoglycan-Based Therapy: Synthetic or Naturally Unique Molecules?

    Pomin, Vitor H

    2015-11-01

    Glycosaminoglycans (GAGs) are widely explored in the biomedical market as functional ingredients in pharmaceutical or nutraceutical preparations. This extensive application of GAGs is justified by their multiple activities across several systems including, but not limited to, coagulation, thrombosis, inflammation, cancer, angiogenesis, cell differentiation, tissue repair, and microbial infections. Therapeutic GAGs are commonly extracted from mammalian tissues. Although functional in diverse systems, mammalian GAGs present serious downsides in therapy such as contamination risk from the mammalian tissues. In order to overcome some of the downsides, two new GAG sources have been appearing as alternatives to the mammalian-derived molecules. They are the synthetic GAGs and those extracted from nonmammalian origins such as invertebrate animals. This report overviews the general aspects of each GAG alternative and compares critically their pros and cons attributes in light of the prospects for the future of GAG-based therapy. PMID:26152885

  5. Glycosaminoglycan binding facilitates entry of a bacterial pathogen into central nervous systems.

    Chang, Yung-Chi; Wang, Zhipeng; Flax, Lindsay A; Xu, Ding; Esko, Jeffrey D; Nizet, Victor; Baron, Miriam J

    2011-06-01

    Certain microbes invade brain microvascular endothelial cells (BMECs) to breach the blood-brain barrier (BBB) and establish central nervous system (CNS) infection. Here we use the leading meningitis pathogen group B Streptococcus (GBS) together with insect and mammalian infection models to probe a potential role of glycosaminoglycan (GAG) interactions in the pathogenesis of CNS entry. Site-directed mutagenesis of a GAG-binding domain of the surface GBS alpha C protein impeded GBS penetration of the Drosophila BBB in vivo and diminished GBS adherence to and invasion of human BMECs in vitro. Conversely, genetic impairment of GAG expression in flies or mice reduced GBS dissemination into the brain. These complementary approaches identify a role for bacterial-GAG interactions in the pathogenesis of CNS infection. Our results also highlight how the simpler yet genetically conserved Drosophila GAG pathways can provide a model organism to screen candidate molecules that can interrupt pathogen-GAG interactions for future therapeutic applications. PMID:21731486

  6. Glycosaminoglycan binding facilitates entry of a bacterial pathogen into central nervous systems.

    Yung-Chi Chang

    2011-06-01

    Full Text Available Certain microbes invade brain microvascular endothelial cells (BMECs to breach the blood-brain barrier (BBB and establish central nervous system (CNS infection. Here we use the leading meningitis pathogen group B Streptococcus (GBS together with insect and mammalian infection models to probe a potential role of glycosaminoglycan (GAG interactions in the pathogenesis of CNS entry. Site-directed mutagenesis of a GAG-binding domain of the surface GBS alpha C protein impeded GBS penetration of the Drosophila BBB in vivo and diminished GBS adherence to and invasion of human BMECs in vitro. Conversely, genetic impairment of GAG expression in flies or mice reduced GBS dissemination into the brain. These complementary approaches identify a role for bacterial-GAG interactions in the pathogenesis of CNS infection. Our results also highlight how the simpler yet genetically conserved Drosophila GAG pathways can provide a model organism to screen candidate molecules that can interrupt pathogen-GAG interactions for future therapeutic applications.

  7. Glycosaminoglycans in the accessory sex glands, testes and seminal plasma of alpaca and ram.

    Kershaw-Young, Claire M; Evans, G; Maxwell, W M C

    2012-01-01

    The viscous nature of alpaca semen limits its use in cryopreservation and other assisted reproductive technologies. The cause and source of this viscosity is unknown although it has been postulated, but never proven, that glycosaminoglycans (GAGs) secreted by the bulbourethral gland are responsible. The present study investigated the concentration and composition of GAGs in alpaca seminal plasma, testes, bulbourethral gland and prostate gland and compared them to those in the ram to determine the relationship between seminal plasma GAGs and viscosity and to identify the source of seminal plasma GAGs. Alpaca seminal plasma contained more GAGs than ram (Pviscosity (P=0.05, R(2)=0.2635). The alpaca bulbourethral gland contained most GAGs compared with prostate or testis (Pviscosity in alpacas, and that the seminal plasma GAGs originate from the bulbourethral gland. PMID:22281083

  8. Nanocoating of titanium implant surfaces with organic molecules. Polysaccharides including glycosaminoglycans

    Gurzawska, Katarzyna; Svava, Rikke; Jørgensen, Niklas Rye;

    2012-01-01

    Long-term stability of titanium implants are dependent on a variety of factors. Nanocoating with organic molecules is one of the method used to improve osseointegration. Nanoscale modification of titanium implants affects surface properties, such as hydrophilicity, biochemical bonding capacity and...... roughness. This influences cell behaviour on the surface such as adhesion, proliferation and differentiation of cells as well as the mineralization of the extracellular matrix at the implant surfaces. The aim of the present systematic review was to describe organic molecules used for surface nanocoating...... nanocoatings. The included in vivo studies, showed improvement of bone interface reactions measured as increased Bone-to-Implant Contact length and Bone Mineral Density adjacent to the polysaccharide coated surfaces. Based on existing literature, surface modification with polysaccharide and glycosaminoglycans...

  9. Effects of glycosaminoglycan polysulphate on the organisation of collagen fibres in experimentally induced tendonitis in horses.

    Moraes, J R E; Facco, G G; Moraes, F R; Engracia Filho, J R; Miyazato, L G; Beretta, D C

    2009-08-15

    An inflammatory process was induced by intratendinous injection of bacterial collagenase into the superficial digital flexor tendon (SDFT) of the left thoracic limb of 10 horses. One week later, the tendons in five of the horses (group 1) were treated with glycosaminoglycan polysulphate (GAGPS), and the tendons of the other five (group 2) were treated with saline solution. The horses were euthanased 150 days after the collagenase injections, and samples of the SDFTs were frozen at -14 degrees C, sectioned at 5 to 7 mum longitudinally and transversely, and stained by the picrosirius red method. Morphometric analysis was used to quantify the organised and disorganised bundles of collagen in the samples from groups 1 and 2. Significantly more organised bundles of collagen were observed in the tendons treated with GAGPS. PMID:19684346

  10. Treatment of renal calcium stone disease with the synthetic glycosaminoglycan pentosan polysulphate.

    Fellström, B; Backman, U; Danielson, B; Wikström, B

    1994-01-01

    Glycosaminoglycans (GAGs) are potent inhibitors of calcium oxalate growth and aggregation. The synthetic GAG pentosan polysulphate (PPS) was used in the treatment of patients with renal calcium stone disease. Altogether, 121 patients were included in an open trial over a 3-year-period. The average stone episode rate and the stone operation rate were no different during treatment and in the pretreatment period. Altogether 48% of the patients were entirely stone-free during follow-up, whereas 29/56 patients who continued to form stones reported smaller stones that were more easily passed. It is concluded that there may be a role for PPS in the treatment of recurrent renal calcium stone disease, but a controlled study may be needed. PMID:7516780

  11. Removal of glycosaminoglycans from bovine granulosa cells contributes to increased binding of hydrogen-3 heparin

    Ax, R.L.; Stodd, C.M.; Boehm, S.K.; Bellin, M.E.

    1986-02-01

    Granulosa cells from small or large bovine follicles were pretreated with enzymes that hydrolyze various glycosaminoglycans, and binding of (/sup 3/H)-heparin to the granulosa was measured. Binding of (/sup 3/H) heparin increased significantly after enzymatic pretreatments with chondroitinase ABC and fungal hyaluronidase, and similar results were obtained with granulosa from small and large follicles. No changes in binding of (/sup 3/H) heparin were detected after hydrolyses with chondroitinase AC and heparinase in either follicle size. Heparitinase, which hydrolyzes heparan sulfate, led to a significant 50% increase in binding of (/sup 3/H) heparin to granulosa from large follicles but was without effect in small follicles. These results suggest that the lower binding of (/sup 3/H) heparin, which has been reported with follicular enlargement, may be due to heparan sulfate occupying or obstructing binding sites for heparin on granulosa from large follicles.

  12. Tensile force transmission in human patellar tendon fascicles is not mediated by glycosaminoglycans

    Svensson, René B; Hassenkam, Tue; Hansen, Philip;

    2011-01-01

    suggested that the proteoglycan-associated glycosaminoglycans (GAGs) found on the surface of the collagen fibrils may be an important transmitter of load, but existing results are ambiguous and have not investigated human tendons. We have used a small-scale mechanical testing system to measure the...... change the tendon modulus, relative energy dissipation, peak stress, or peak strain. The effect of deformation rate was not modulated by the treatment either, indicating no effect on viscosity. These results suggest that GAGs cannot be considered mediators of tensile force transmission in the human...... patellar tendon, and as such, force transmission must either take place through other matrix components or the fibrils must be mechanically continuous at least to the tested length of 7 mm....

  13. Influence of Hyaluronic Acid in Periodontal Tissue Regeneration

    Radojkova-Nikolovska, Vera; Popovska, Mirjana; Minovska, Ana; Belazelkovska, Zlatanka

    2013-01-01

    Hyaluronic acid is a high molecular weight polysaccharide - glycosaminoglycan, which plays a vital role in the functioning of extracellular matrices, including those of mineralized and non-mineralized periodontal tissues. Hyaluronic acid is also important because of its numerous actions in the mechanisms associated with inflammation and the wound healing process. Hyaluronic acid has been identified in all periodontal tissues in varying quantities, being more prominent in ...

  14. Diabetes-Impaired Wound Healing Is Improved by Matrix Therapy With Heparan Sulfate Glycosaminoglycan Mimetic OTR4120 in Rats

    Tong, Miao; Tuk, Bastiaan; Shang, Peng; Hekking, Ineke M.; Esther M G Fijneman; Guijt, Marnix; Hovius, Steven E. R.; Johan W van Neck

    2012-01-01

    Wound healing in diabetes is frequently impaired, and its treatment remains a challenge. We tested a therapeutic strategy of potentiating intrinsic tissue regeneration by restoring the wound cellular environment using a heparan sulfate glycosaminoglycan mimetic, OTR4120. The effect of OTR4120 on healing of diabetic ulcers was investigated. Experimental diabetes was induced by intraperitoneal injection of streptozotocin. Seven weeks after induction of diabetes, rats were ulcerated by clamping ...

  15. Peptide p5 binds both heparinase-sensitive glycosaminoglycans and fibrils in patient-derived AL amyloid extracts

    Highlights: •Polybasic peptide p5 binds human light chain amyloid extracts. •The binding of p5 with amyloid involves both glycosaminoglycans and fibrils. •Heparinase treatment led to a correlation between p5 binding and fibril content. •p5 binding to AL amyloid requires electrostatic interactions. -- Abstract: In previously published work, we have described heparin-binding synthetic peptides that preferentially recognize amyloid deposits in a mouse model of reactive systemic (AA) amyloidosis and can be imaged by using positron and single photon emission tomographic imaging. We wanted to extend these findings to the most common form of visceral amyloidosis, namely light chain (AL); however, there are no robust experimental animal models of AL amyloidosis. To further define the binding of the lead peptide, p5, to AL amyloid, we characterized the reactivity in vitro of p5 with in situ and patient-derived AL amyloid extracts which contain both hypersulfated heparan sulfate proteoglycans as well as amyloid fibrils. Histochemical staining demonstrated that the peptide specifically localized with tissue-associated AL amyloid deposits. Although we anticipated that p5 would undergo electrostatic interactions with the amyloid-associated glycosaminoglycans expressing heparin-like side chains, no significant correlation between peptide binding and glycosaminoglycan content within amyloid extracts was observed. In contrast, following heparinase I treatment, although overall binding was reduced, a positive correlation between peptide binding and amyloid fibril content became evident. This interaction was further confirmed using synthetic light chain fibrils that contain no carbohydrates. These data suggest that p5 can bind to both the sulfated glycosaminoglycans and protein fibril components of AL amyloid. Understanding these complex electrostatic interactions will aid in the optimization of synthetic peptides for use as amyloid imaging agents and potentially as

  16. Increased physical activity severely induces osteoarthritic changes in knee joints with papain induced sulfate-glycosaminoglycan depleted cartilage

    Siebelt, M.; Groen, H.C.; Koelewijn, S. J.; de Blois, E.; Sandker, M.; Waarsing, J. H.; Müller, C.(Dr. Remeis-Sternwarte and ECAP, Universität Erlangen-Nürnberg, Sternwartstr. 7, 96049 , Bamberg, Germany); van Osch, G. J. V. M.; de Jong, M.; Weinans, H.H.

    2014-01-01

    Introduction Articular cartilage needs sulfated-glycosaminoglycans (sGAGs) to withstand high pressures while mechanically loaded. Chondrocyte sGAG synthesis is regulated by exposure to compressive forces. Moderate physical exercise is known to improve cartilage sGAG content and might protect against osteoarthritis (OA). This study investigated whether rat knee joints with sGAG depleted articular cartilage through papain injections might benefit from moderate exercise, or whether this increase...

  17. The Structure of Chondroitin B Lyase Complexed with Glycosaminoglycan Oligosaccharides Unravels a Calcium-dependent Catalytic Machinery*

    Michel, Gurvan; Pojasek, Kevin; Li, Yunge; Sulea, Traian; Linhardt, Robert J.; Raman, Rahul; Prabhakar, Vikas; Sasisekharan, Ram; Cygler, Miroslaw

    2004-01-01

    Chondroitinase B from Pedobacter heparinus is the only known enzyme strictly specific for dermatan sulfate and is a widely used enzymatic tool for the structural characterization of glycosaminoglycans. This β-helical polysaccharide lyase belongs to family PL-6 and cleaves the β(1,4) linkage of dermatan sulfate in a random manner, yielding 4,5-unsaturated dermatan sulfate disaccharides as the product. The previously reported structure of its complex with a dermatan sulfate disaccharide product...

  18. Positive effect of oral supplementation with glycosaminoglycans and antioxidants on the regeneration of osteochondral defects in the knee joint

    Handl, M.; Amler, Evžen; Bräun, K.; Holzheu, J.; Imhoff, A.B.; Lytvynets, Andrej; Filová, Eva; Kotyk, Arnošt; Martínek, V.

    2007-01-01

    Roč. 56, č. 2 (2007), s. 243-249. ISSN 0862-8408 R&D Projects: GA AV ČR(CZ) 1ET400110403; GA AV ČR(CZ) IAA500390702 Institutional research plan: CEZ:AV0Z50390512; CEZ:AV0Z50110509 Keywords : Cartilage * Glycosaminoglycans * Oral supplementatio Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.505, year: 2007

  19. Inhibition of Chemokine-Glycosaminoglycan Interactions in Donor Tissue Reduces Mouse Allograft Vasculopathy and Transplant Rejection

    Dai, Erbin; Liu, Li-Ying; Wang, Hao; McIvor, Dana; Sun, Yun ming; Macaulay, Colin; King, Elaine; Munuswamy-Ramanujam, Ganesh; Bartee, Mee Yong; Williams, Jennifer; Davids, Jennifer; Charo, Israel; McFadden, Grant; Esko, Jeffrey D.; Lucas, Alexandra R.

    2010-01-01

    Background Binding of chemokines to glycosaminoglycans (GAGs) is classically described as initiating inflammatory cell migration and creating tissue chemokine gradients that direct local leukocyte chemotaxis into damaged or transplanted tissues. While chemokine-receptor binding has been extensively studied during allograft transplantation, effects of glycosaminoglycan (GAG) interactions with chemokines on transplant longevity are less well known. Here we examine the impact of interrupting chemokine-GAG interactions and chemokine-receptor interactions, both locally and systemically, on vascular disease in allografts. Methodology/Principal Findings Analysis of GAG or CC chemokine receptor 2 (CCR2) deficiency were coupled with the infusion of viral chemokine modulating proteins (CMPs) in mouse aortic allograft transplants (n = 239 mice). Inflammatory cell invasion and neointimal hyperplasia were significantly reduced in N-deacetylase-N-sulfotransferase-1 (Ndst1f/fTekCre+) heparan sulfate (GAG)-deficient (Ndst1−/−, p<0.044) and CCR2-deficient (Ccr2−/−, p<0.04) donor transplants. Donor tissue GAG or CCR2 deficiency markedly reduced inflammation and vasculopathy, whereas recipient deficiencies did not. Treatment with three CMPs was also investigated; Poxviral M-T1 blocks CC chemokine receptor binding, M-T7 blocks C, CC, and CXC GAG binding, and herpesviral M3 binds receptor and GAG binding for all classes. M-T7 reduced intimal hyperplasia in wild type (WT) (Ccr2+/+, p≤0.003 and Ccr2−/−, p≤0.027) aortic allografts, but not in Ndst1−/− aortic allografts (p = 0.933). M-T1 and M3 inhibited WT (Ccr2+/+ and Ndst1+/+, p≤0.006) allograft vasculopathy, but did not block vasculopathy in Ccr2−/− (p = 0.61). M-T7 treatment alone, even without immunosuppressive drugs, also significantly prolonged survival of renal allograft transplants (p≤0.001). Conclusions/Significance Interruption of chemokine-GAG interactions, even in the absence of

  20. Comparison of Engineered Peptide-Glycosaminoglycan Microfibrous Hybrid Scaffolds for Potential Applications in Cartilage Tissue Regeneration

    Steven M. Romanelli

    2015-07-01

    Full Text Available Advances in tissue engineering have enabled the ability to design and fabricate biomaterials at the nanoscale that can actively mimic the natural cellular environment of host tissue. Of all tissues, cartilage remains difficult to regenerate due to its avascular nature. Herein we have developed two new hybrid polypeptide-glycosaminoglycan microfibrous scaffold constructs and compared their abilities to stimulate cell adhesion, proliferation, sulfated proteoglycan synthesis and soluble collagen synthesis when seeded with chondrocytes. Both constructs were designed utilizing self-assembled Fmoc-protected valyl cetylamide nanofibrous templates. The peptide components of the constructs were varied. For Construct I a short segment of dentin sialophosphoprotein followed by Type I collagen were attached to the templates using the layer-by-layer approach. For Construct II, a short peptide segment derived from the integrin subunit of Type II collagen binding protein expressed by chondrocytes was attached to the templates followed by Type II collagen. To both constructs, we then attached the natural polymer N-acetyl glucosamine, chitosan. Subsequently, the glycosaminoglycan chondroitin sulfate was then attached as the final layer. The scaffolds were characterized by Fourier transform infrared spectroscopy (FT-IR, differential scanning calorimetry (DSC, atomic force microscopy and scanning electron microscopy. In vitro culture studies were carried out in the presence of chondrocyte cells for both scaffolds and growth morphology was determined through optical microscopy and scanning electron microscopy taken at different magnifications at various days of culture. Cell proliferation studies indicated that while both constructs were biocompatible and supported the growth and adhesion of chondrocytes, Construct II stimulated cell adhesion at higher rates and resulted in the formation of three dimensional cell-scaffold matrices within 24 h. Proteoglycan

  1. A biomimetic porous hydrogel of gelatin and glycosaminoglycans cross-linked with transglutaminase and its application in the culture of hepatocytes

    The development of blended gelatin and glycosaminoglycan (GAG) scaffolds can potentially be used in many soft tissue engineering applications since these scaffolds mimic the structure and biological function of native extracellular matrix (ECM). In this study, we were able to obtain a gelatin–GAG scaffold by using a concentrated emulsion templating technique known as high internal phase emulsion (HIPE), in which a prevailing in volume organic phase is dispersed in the form of discrete droplets inside an aqueous solution of three biopolymers represented by gelatin, hyaluronic acid (HA) and chondroitin sulfate (CS) in the presence of a suitable surfactant. In order to preserve the bioactive potential of the biopolymers employed, the cross-linking procedure involved the use of transglutaminase (MTGase) that catalyzes the formation of covalent N-ε-(γ-glutamyl) lysine amide bonds. Since neither HA nor CS possess the necessary primary amino groups toward which MTGase is active, they were functionalized with the dipeptide glycine-lysine (GK). In this way the introduction of foreign cross-linking bridging units with an unpredictable biocompatibility was avoided. These enzymatic cross-linked gelatin–GAG scaffolds were tested in the culture of primary rat and C3A hepatocytes. Results underlined the good performance of this novel support in maintaining and promoting hepatocyte functions in vitro. (paper)

  2. Selectivity in glycosaminoglycan binding dictates the distribution and diffusion of fibroblast growth factors in the pericellular matrix.

    Sun, Changye; Marcello, Marco; Li, Yong; Mason, David; Lévy, Raphaël; Fernig, David G

    2016-03-01

    The range of biological outcomes generated by many signalling proteins in development and homeostasis is increased by their interactions with glycosaminoglycans, particularly heparan sulfate (HS). This interaction controls the localization and movement of these signalling proteins, but whether such control depends on the specificity of the interactions is not known. We used five fibroblast growth factors with an N-terminal HaloTag (Halo-FGFs) for fluorescent labelling, with well-characterized and distinct HS-binding properties, and measured their binding and diffusion in pericellular matrix of fixed rat mammary 27 fibroblasts. Halo-FGF1, Halo-FGF2 and Halo-FGF6 bound to HS, whereas Halo-FGF10 also interacted with chondroitin sulfate/dermatan sulfate, and FGF20 did not bind detectably. The distribution of bound FGFs in the pericellular matrix was not homogeneous, and for FGF10 exhibited striking clusters. Fluorescence recovery after photobleaching showed that FGF2 and FGF6 diffused faster, whereas FGF1 diffused more slowly, and FGF10 was immobile. The results demonstrate that the specificity of the interactions of proteins with glycosaminoglycans controls their binding and diffusion. Moreover, cells regulate the spatial distribution of different protein-binding sites in glycosaminoglycans independently of each other, implying that the extracellular matrix has long-range structure. PMID:27009190

  3. INCREASE OF GLYCOSAMINOGLYCANS AND METALLOPROTEINASES 2 AND 9 IN LIVER EXTRACELLULAR MATRIX ON EARLY STAGES OF EXTRAHEPATIC CHOLESTASIS

    Pedro Luiz Rodrigues GUEDES

    2014-12-01

    Full Text Available Context Cholestasis produces hepatocellular injury, leukocyte infiltration, ductular cells proliferation and fibrosis of liver parenchyma by extracellular matrix replacement. Objective Analyze bile duct ligation effect upon glycosaminoglycans content and matrix metalloproteinase (MMPs activities. Methods Animals (6-8 weeks; n = 40 were euthanized 2, 7 or 14 days after bile duct ligation or Sham-surgery. Disease evolution was analyzed by body and liver weight, seric direct bilirubin, globulins, gamma glutamyl transpeptidase (GGT, alkaline phosphatase (Alk-P, alanine and aspartate aminotransferases (ALT and AST, tissue myeloperoxidase and MMP-9, pro MMP-2 and MMP-2 activities, histopathology and glycosaminoglycans content. Results Cholestasis caused cellular damage with elevation of globulins, GGT, Alk-P, ALT, AST. There was neutrophil infiltration observed by the increasing of myeloperoxidase activity on 7 (P = 0.0064 and 14 (P = 0.0002 groups which leads to the magnification of tissue injuries. Bile duct ligation increased pro-MMP-2 (P = 0.0667, MMP-2 (P = 0.0003 and MMP-9 (P<0.0001 activities on 14 days indicating matrix remodeling and establishment of inflammatory process. Bile duct ligation animals showed an increasing on dermatan sulfate and/or heparan sulfate content reflecting extracellular matrix production and growing mitosis due to parenchyma depletion. Conclusions Cholestasis led to many changes on rats’ liver parenchyma, as so as on its extracellular matrix, with major alterations on MMPs activities and glycosaminoglycans content.

  4. Xyloside primed glycosaminoglycans alter hair bundle micromechanical coupling and synaptic transmission: Pharmacokinetics

    Holman, Holly A.; Tran, Vy M.; Nguyen, Lynn Y.; Arungundram, Sailaja; Kalita, Mausam; Kuberan, Balagurunathan; Rabbitt, Richard D.

    2015-12-01

    Glycosaminoglycans (GAGs) are ubiquitous in the inner ear, and disorders altering their structure or production often result in debilitating hearing and balance deficits. The specific mechanisms responsible for loss of hair-cell function are not well understood. We recently reported that introduction of a novel BODIPY conjugated xyloside (BX) into the endolymph primes fluorescent GAGs in vivo [6, 15]. Confocal and two-photon fluorescence imaging revealed rapid turnover and assembly of a glycocalyx enveloping the kinocilia and extending into the cupula, a structure that presumably serves as a mechanical link between the hair bundle and the cupula. Extracellular fluorescence was also observed around the basolateral surface of hair cells and surrounding afferent nerve projections into the crista. Single unit afferent recordings during mechanical hair bundle stimulation revealed temporary interruption of synaptic transmission following BX administration followed by recovery, demonstrating an essential role for GAGs in function of the hair cell synapse. In the present work we present a pharmacokinetic model to quantify the time course of BX primed GAG production and turnover in the ear.

  5. Cell-Penetrating Ability of Peptide Hormones: Key Role of Glycosaminoglycans Clustering

    Armelle Tchoumi Neree

    2015-11-01

    Full Text Available Over the last two decades, the potential usage of cell-penetrating peptides (CPPs for the intracellular delivery of various molecules has prompted the identification of novel peptidic identities. However, cytotoxic effects and unpredicted immunological responses have often limited the use of various CPP sequences in the clinic. To overcome these issues, the usage of endogenous peptides appears as an appropriate alternative approach. The hormone pituitary adenylate-cyclase-activating polypeptide (PACAP38 has been recently identified as a novel and very efficient CPP. This 38-residue polycationic peptide is a member of the secretin/glucagon/growth hormone-releasing hormone (GHRH superfamily, with which PACAP38 shares high structural and conformational homologies. In this study, we evaluated the cell-penetrating ability of cationic peptide hormones in the context of the expression of cell surface glycosaminoglycans (GAGs. Our results indicated that among all peptides evaluated, PACAP38 was unique for its potent efficiency of cellular uptake. Interestingly, the abilities of the peptides to reach the intracellular space did not correlate with their binding affinities to sulfated GAGs, but rather to their capacity to clustered heparin in vitro. This study demonstrates that the uptake efficiency of a given cationic CPP does not necessarily correlate with its affinity to sulfated GAGs and that its ability to cluster GAGs should be considered for the identification of novel peptidic sequences with potent cellular penetrating properties.

  6. Influence of the sulfation degree of glycosaminoglycans on their multilayer assembly with poly-l-lysine.

    Teixeira, Raquel; Reis, Rui L; Pashkuleva, Iva

    2016-09-01

    We report on the build-up and the intrinsic properties of polyelectrolyte multilayer films from poly-l-lysine and glycosaminoglycans (GAGs) with different sulfation degree, i.e. different charge. We used three complementary techniques, namely electrokinetic analysis (EKA), quartz-crystal microbalance with dissipation (QCM-D) and surface plasmon resonance (SPR), to characterize the assembly process and to assess the properties of the obtained films. EKA elucidated the contribution of the polymers charged groups to the net surface charge of the films and suggested that the assembly process is not solely driven by electrostatic interactions. The combined analysis of QCM-D and SPR data demonstrated that the mechanical properties of the films are dependent on the polymer charge: sulfated GAGs (heparin and chondroitin sulfate) form elastic films while hyaluronan (no sulfation) assembles into multilayer constructs with viscous behavior. The contribution of the water content to these distinct regimes is also discussed. Finally, we show that rather complete characterization of the film properties is possible by SPR employing the two-wavelength and two-media approach: thickness, adsorbed mass, refractive index, and interaction kinetics of the assembly process can be studied by SPR alone. PMID:27285729

  7. Glycosaminoglycan Monosaccharide Blocks Analysis by Quantum Mechanics, Molecular Dynamics, and Nuclear Magnetic Resonance

    Sergey A. Samsonov

    2014-01-01

    Full Text Available Glycosaminoglycans (GAGs play an important role in many biological processes in the extracellular matrix. In a theoretical approach, structures of monosaccharide building blocks of natural GAGs and their sulfated derivatives were optimized by a B3LYP6311ppdd//B3LYP/6-31+G(d method. The dependence of the observed conformational properties on the applied methodology is described. NMR chemical shifts and proton-proton spin-spin coupling constants were calculated using the GIAO approach and analyzed in terms of the method's accuracy and sensitivity towards the influence of sulfation, O1-methylation, conformations of sugar ring, and ω dihedral angle. The net sulfation of the monosaccharides was found to be correlated with the 1H chemical shifts in the methyl group of the N-acetylated saccharides both theoretically and experimentally. The ω dihedral angle conformation populations of free monosaccharides and monosaccharide blocks within polymeric GAG molecules were calculated by a molecular dynamics approach using the GLYCAM06 force field and compared with the available NMR and quantum mechanical data. Qualitative trends for the impact of sulfation and ring conformation on the chemical shifts and proton-proton spin-spin coupling constants were obtained and discussed in terms of the potential and limitations of the computational methodology used to be complementary to NMR experiments and to assist in experimental data assignment.

  8. Systematic protein interactome analysis of glycosaminoglycans revealed YcbS as a novel bacterial virulence factor.

    Hsiao, Felix Shih-Hsiang; Sutandy, Fx Reymond; Syu, Guan-Da; Chen, Yi-Wen; Lin, Jun-Mu; Chen, Chien-Sheng

    2016-01-01

    Microbial pathogens have evolved several strategies for interacting with host cell components, such as glycosaminoglycans (GAGs). Some microbial proteins involved in host-GAG binding have been described; however, a systematic study on microbial proteome-mammalian GAG interactions has not been conducted. Here, we used Escherichia coli proteome chips to probe four typical mammalian GAGs, heparin, heparan sulphate (HS), chondroitin sulphate B (CSB), and chondroitin sulphate C (CSC), and identified 185 heparin-, 62 HS-, 98 CSB-, and 101 CSC-interacting proteins. Bioinformatics analyses revealed the unique functions of heparin- and HS-specific interacting proteins in glycine, serine, and threonine metabolism. Among all the GAG-interacting proteins, three were outer membrane proteins (MbhA, YcbS, and YmgH). Invasion assays confirmed that mutant E. coli lacking ycbS could not invade the epithelial cells. Introducing plasmid carrying ycbS complemented the invading defects at ycbS lacking E. coli mutant, that can be further improved by overexpressing ycbS. Preblocking epithelial cells with YcbS reduced the percentage of E. coli invasions. Moreover, we observed that whole components of the ycb operon were crucial for invasion. The displacement assay revealed that YcbS binds to the laminin-binding site of heparin and might affect the host extracellular matrix structure by displacing heparin from laminin. PMID:27323865

  9. Nephroprotective action of glycosaminoglycans: why the pharmacological properties of sulodexide might be reconsidered

    Antonio V Gaddi

    2010-07-01

    Full Text Available Antonio V Gaddi1, Arrigo FG Cicero1, Giovanni Gambaro21Atherosclerosis and Metabolic disease Research Unit, Internal Medicine, Aging and Kidney diseases Dept., University of Bologna, Italy; 2Nephrology and Dialysis Unit, Gemelli University Hospital, Sacred Heart Catholic University, Rome, ItalyAbstract: A relatively large body of evidence supports the notion that glomerular capillary wall and mesangial alterations in diabetic nephropathy involve biochemical alterations of glycoproteins in these structures. Evidence in experimental animals rendered diabetic reveals that the administration of heparin and other anionic glycoproteins can effectively prevent the biochemical alterations that promote albuminuria. Moreover, angiotensin II inhibits heparan sulfate synthesis, while heparins modulate angiotensin II signaling in glomerular cells, inhibiting aldosterone synthesis and lowering proteinuria in diabetes patients. Sulodexide, a mixture of heparin and dermatan sulfate, appears to be a promising treatment for diabetic proteinuria partially resistant to renin–angiotensin system blocking agents. Sulodexide prevents heparan sulfate degradation, thus allowing reconstruction of heparan sulfate content and restoration of glomerular basement membrane ionic permselectivity. The antiproteinuric effect appears to be mainly related to the basal proteinuria and consequently to the duration of treatment in a relatively large number of small clinical trials. On the other hand, several sulodexide pharmacodynamic properties could improve the prognosis of chronic kidney disease patients, also independently from its antiproteinuric effect. However, sulodexide development as an antiproteinuric drug needs to be continued, in order to define which kind of patients could better respond to this treatment.Keywords: glycosaminoglycans, sulodexide, albuminuria, proteinuria, diabetic nephropathy

  10. Binding of anti-prion agents to glycosaminoglycans: Evidence from electronic absorption and circular dichroism spectroscopy

    The polyanionic glycosaminoglycans (GAGs) are intimately involved in the pathogenesis of protein conformational disorders such as amyloidosis and prion diseases. Several cationic agents are known to exhibit anti-prion activity but their mechanism of action is poorly understood. In this study, UV absorption and circular dichroism (CD) spectroscopic techniques were used to investigate the interaction between heparin and chondroitin-6-sulfate and anti-prion drugs including acridine, quinoline, and phenothiazine derivatives. UV band hypochromism of (±)-quinacrine, (±)-primaquine, tacrine, quinidine, chlorpromazine, and induced CD spectra of (±)-quinacrine upon addition of GAGs provided evidence for the GAG binding of these compounds. The association constants (∼106-107 M-1) estimated from the UV titration curves show high-affinity drug-heparin interactions. Ionic strength-dependence of the absorption spectra suggested that the interaction between GAGs and the cationic drugs is principally electrostatic in nature. Drug binding differences of heparin and chondroitin-6-sulfate were attributed to their different negative charge density. These results call the attention to the alteration of GAG-prion/GAG-amyloid interactions by which these compounds might exert their anti-prion/anti-amyloidogenic activities

  11. Structural Evidence for the Tetrameric Assembly of Chemokine CCL11 and the Glycosaminoglycan Arixtra™

    Dykstra, Andrew B.; Sweeney, Matt D.; Leary, Julie A.

    2013-01-01

    Understanding chemokine interactions with glycosaminoglycans (GAG) is critical as these interactions have been linked to a number of inflammatory medical conditions, such as arthritis and asthma. To better characterize in vivo protein function, comprehensive knowledge of multimeric species, formed by chemokines under native conditions, is necessary. Herein is the first report of a tetrameric assembly of the human chemokine CCL11, which was shown bound to the GAG Arixtra™. Isothermal titration calorimetry data indicated that CCL11 interacts with Arixtra, and ion mobility mass spectrometry (IM-MS) was used to identify ions corresponding to the CCL11 tetrameric species bound to Arixtra. Collisional cross sections (CCS) of the CCL11 tetramer-Arixtra noncovalent complex were compared to theoretical CCS values calculated using a preliminary structure of the complex deduced using X-ray crystallography. Experimental CCS values were in agreement with theoretical values, strengthening the IM-MS evidence for the formation of the noncovalent complex. Tandem mass spectrometry data of the complex indicated that the tetramer-GAG complex dissociates into a monomer and a trimer-GAG species, suggesting that two CC-like dimers are bridged by Arixtra. As development of chemokine inhibitors is of utmost importance to treatment of medical inflammatory conditions, these results provide vital insights into chemokine-GAG interactions. PMID:24970196

  12. Exopolysaccharides produced by marine bacteria and their applications as glycosaminoglycan-like molecules.

    Delbarre-Ladrat, Christine; Sinquin, Corinne; Lebellenger, Lou; Zykwinska, Agata; Colliec-Jouault, Sylvia

    2014-10-01

    Although polysaccharides are ubiquitous and the most abundant renewable bio-components, their studies, covered by the glycochemistry and glycobiology fields, remain a challenge due to their high molecular diversity and complexity. Polysaccharides are industrially used in food products; human therapeutics fall into a more recent research field and pharmaceutical industry is looking for more and more molecules with enhanced activities. Glycosaminoglycans (GAGs) found in animal tissues play a critical role in cellular physiological and pathological processes as they bind many cellular components. Therefore, they present a great potential for the design and preparation of therapeutic drugs. On the other hand, microorganisms producing exopolysaccharides (EPS) are renewable resources meeting well the actual industrial demand. In particular, the diversity of marine microorganisms is still largely unexplored offering great opportunities to discover high value products such as new molecules and biocatalysts. EPS-producing bacteria from the marine environment will be reviewed with a focus on marine-derived EPS from bacteria isolated from deep-sea hydrothermal vents. Information on chemical and structural features, putative pathways of biosynthesis, novel strategies for chemical and enzymatic modifications and potentialities in the biomedical field will be provided. An integrated approach should be used to increase the basic knowledge on these compounds and their applications; new clean environmentally friendly processes for the production of carbohydrate bio-active compounds should also be proposed for a sustainable industry.

  13. Tensile force transmission in human patellar tendon fascicles is not mediated by glycosaminoglycans

    Svensson, René B; Hassenkam, Tue; Hansen, Philip;

    2011-01-01

    Correct mechanical function of tendons is essential to human physiology and therefore the mechanical properties of tendon have been a subject of research for many decades now. However, one of the most fundamental questions remains unanswered: How is load transmitted through the tendon? It has bee...... patellar tendon, and as such, force transmission must either take place through other matrix components or the fibrils must be mechanically continuous at least to the tested length of 7 mm.......Correct mechanical function of tendons is essential to human physiology and therefore the mechanical properties of tendon have been a subject of research for many decades now. However, one of the most fundamental questions remains unanswered: How is load transmitted through the tendon? It has been...... suggested that the proteoglycan-associated glycosaminoglycans (GAGs) found on the surface of the collagen fibrils may be an important transmitter of load, but existing results are ambiguous and have not investigated human tendons. We have used a small-scale mechanical testing system to measure the...

  14. Glycosaminoglycan and DNA Binding Induced Intra- and Intermolecular Exciton Coupling of the bis-4-Aminoquinoline Surfen.

    Zsila, Ferenc

    2015-09-01

    Despite the diverse biological activities of the glycosaminoglycan (GAG) antagonist surfen, the molecular details of its interaction with biomacromolecules remain poorly understood. Therefore, heparin and DNA binding properties of surfen were studied by circular dichroism (CD) and UV absorption spectroscopy methods. High-affinity (Ka  ~ 10(7)  M(-1)) association of surfen to the chiral heparin chain gives rise to a characteristic biphasic CD pattern due to the conformational twist of the aminoquinoline moieties around the central urea bridge. At higher drug loading, intermolecular stacking of surfen molecules alters the induced CD profile and also provokes strong UV hypochromism. In contrast to the right-handed heparin template, binding of surfen to the left-helicity chondroitin sulfate chains produces inverted CD pattern. Large UV hypochromism as well as polyphasic induced ellipticity bands indicate that surfen intercalates between the base pairs of calf-thymus DNA. Extensive CD spectroscopic changes observed at higher drug binding ratios refer to cooperative binding interactions between the intercalated drug molecules. The inherent conformational flexibility of surfen demonstrated here for the first time is important in its binding to distinct macromolecular targets and should be considered for rational drug design of novel GAG antagonists. PMID:26096963

  15. Capillary electrophoresis of heparin and other glycosaminoglycans using a polyamine running electrolyte

    Highlights: ► Ethylenediamine is likely acting as an ion-pairing agent. ► Oversulfated chondroitin sulfate is last peak instead of first peak. ► There is about a factor of five improved detectability with a 12.5 min analysis time. ► Use of a 50 μm ID capillary is possible. - Abstract: This study involves the use of polyamines as potential resolving agents for the capillary electrophoresis (CE) of glycosaminoglycans (GAGs), specifically heparin, dermatan sulfate, chondroitin sulfate, over-sulfated chondroitin sulfate (OSCS), and hyaluronan. All of the compounds can be separated from each other with the exception of chondroitin sulfate and hyaluronan. Using optimization software, the final run conditions are found to be 200 mM ethylenediamine and 45.5 mM phosphate as the electrolyte with −14 V applied across a 50 μm ID × 24.5 cm fused silica capillary at 15 °C. The ion migration order, with OSCS as the last instead of the first peak, is in contrast to previous reports using either a high molarity TRIS or lithium phosphate run buffer with narrower bore capillaries. Total analysis time is 12. 5 min and the relative standard deviation of the heparin migration time is about 2.5% (n = 5). The interaction mechanism between selected polyamines and heparin is explored using conductivity measurements in addition to CE experiments to show that an ion-pairing mechanism is likely.

  16. New roles of glycosaminoglycans in α-synuclein aggregation in a cellular model of Parkinson disease.

    Sonia Lehri-Boufala

    Full Text Available The causes of Parkinson disease (PD remain mysterious, although some evidence supports mitochondrial dysfunctions and α-synuclein accumulation in Lewy bodies as major events. The abnormal accumulation of α-synuclein has been associated with a deficiency in the ubiquitin-proteasome system and the autophagy-lysosomal pathway. Cathepsin D (cathD, the major lysosomal protease responsible of α-synuclein degradation was described to be up-regulated in PD model. As glycosaminoglycans (GAGs regulate cathD activity, and have been recently suggested to participate in PD physiopathology, we investigated their role in α-synuclein accumulation by their intracellular regulation of cathD activity. In a classical neuroblastoma cell model of PD induced by MPP+, the genetic expression of GAGs-biosynthetic enzymes was modified, leading to an increase of GAGs amounts whereas intracellular level of α-synuclein increased. The absence of sulfated GAGs increased intracellular cathD activity and limited α-synuclein accumulation. GAGs effects on cathD further suggested that specific sequences or sulfation patterns could be responsible for this regulation. The present study identifies, for the first time, GAGs as new regulators of the lysosome degradation pathway, regulating cathD activity and affecting two main biological processes, α-synuclein aggregation and apoptosis. Finally, this opens new insights into intracellular GAGs functions and new fields of investigation for glycobiological approaches in PD and neurobiology.

  17. Interleukin-1 induced nitric oxide inhibits sulphation of glycosaminoglycan chains in human articular chondrocytes.

    Hickery, M S; Bayliss, M T

    1998-10-23

    Incubation of human articular cartilage explants with interleukin-1alpha (IL-1alpha) inhibited the rate of [35S]sulphate incorporation into glycosaminoglycan (GAG) chains concomitant with an increase in nitric oxide (NO) production. Measurement of the [35S]sulphate showed that IL-1alpha inhibited the synthesis of both keratan sulphate and chondroitin sulphate (CS) chains to a similar extent. This effect was reversed by the NO synthase inhibitor Nomega-iminoethyl-l-ornithine (l-NIO). Analysis of alkali borohydride cleaved GAG chains showed that IL-1alpha had no effect on their size. Similarly when GAG chains were coupled to xyloside the size of the GAG chains attached to the exogenous acceptor decreased but IL-1alpha had no further effect on hydrodynamic size. IL-1alpha did, however, inhibit [35S]sulphate incorporation into xyloside-linked CS chains. In both experiments l-NIO reversed the inhibitory effect on sulphation. Disaccharide analysis of the [35S]GAG chains showed that IL-1alpha preferentially inhibited sulphation of the 6-sulphated isomer and that l-NIO reversed this effect. Thus, IL-1alpha-induced NO mediates the inhibition of sulphate incorporation and alters the sulphation pattern of newly synthesised GAG chains. PMID:9795242

  18. Structural Evidence for the Tetrameric Assembly of Chemokine CCL11 and the Glycosaminoglycan Arixtra™

    Julie A. Leary

    2013-11-01

    Full Text Available Understanding chemokine interactions with glycosaminoglycans (GAG is critical as these interactions have been linked to a number of inflammatory medical conditions, such as arthritis and asthma. To better characterize in vivo protein function, comprehensive knowledge of multimeric species, formed by chemokines under native conditions, is necessary. Herein is the first report of a tetrameric assembly of the human chemokine CCL11, which was shown bound to the GAG Arixtra™. Isothermal titration calorimetry data indicated that CCL11 interacts with Arixtra, and ion mobility mass spectrometry (IM-MS was used to identify ions corresponding to the CCL11 tetrameric species bound to Arixtra. Collisional cross sections (CCS of the CCL11 tetramer-Arixtra noncovalent complex were compared to theoretical CCS values calculated using a preliminary structure of the complex deduced using X-ray crystallography. Experimental CCS values were in agreement with theoretical values, strengthening the IM-MS evidence for the formation of the noncovalent complex. Tandem mass spectrometry data of the complex indicated that the tetramer-GAG complex dissociates into a monomer and a trimer-GAG species, suggesting that two CC-like dimers are bridged by Arixtra. As development of chemokine inhibitors is of utmost importance to treatment of medical inflammatory conditions, these results provide vital insights into chemokine-GAG interactions.

  19. Enthalpy-driven interactions with sulfated glycosaminoglycans promote cell membrane penetration of arginine peptides.

    Takechi-Haraya, Yuki; Nadai, Ryo; Kimura, Hitoshi; Nishitsuji, Kazuchika; Uchimura, Kenji; Sakai-Kato, Kumiko; Kawakami, Kohsaku; Shigenaga, Akira; Kawakami, Toru; Otaka, Akira; Hojo, Hironobu; Sakashita, Naomi; Saito, Hiroyuki

    2016-06-01

    The first step of cell membrane penetration of arginine peptides is thought to occur via electrostatic interactions between positive charges of arginine residues and negative charges of sulfated glycosaminoglycans (GAGs) on the cell surface. However, the molecular interaction of arginine peptides with GAG still remains unclear. Here, we compared the interactions of several arginine peptides of Tat, R8, and Rev and their analogues with heparin in relation to the cell membrane penetration efficiency. The high-affinity binding of arginine peptides to heparin was shown to be driven by large favorable enthalpy contributions, possibly reflecting multidentate hydrogen bondings of arginine residues with sulfate groups of heparin. Interestingly, the lysine peptides in which all arginine residues are substituted with lysine residues exhibited negligible binding enthalpy despite of their considerable binding to heparin. In CHO-K1 cells, arginine peptides exhibited a great cell-penetrating ability whereas their corresponding lysine peptides did not penetrate into cells. The degree of cell penetration of arginine peptides markedly decreased by the chlorate treatment of cells which prevents the sulfation of GAG chains. Significantly, the cell penetration efficiency of arginine peptides was found to be correlated with the favorable enthalpy of binding to heparin. These results suggest that the enthalpy-driven strong interaction with sulfated GAGs such as heparan sulfate plays a critical role in the efficient cell membrane penetration of arginine peptides. PMID:27003128

  20. Xyloside primed glycosaminoglycans alter hair bundle micromechanical coupling and synaptic transmission: Pharmacokinetics

    Holman, Holly A.; Nguyen, Lynn Y. [Bioengineering, University of Utah, Salt Lake City, Utah (United States); Tran, Vy M.; Arungundram, Sailaja; Kalita, Mausam [Medicinal Chemistry, University of Utah, Salt Lake City, Utah (United States); Kuberan, Balagurunathan [Medicinal Chemistry, University of Utah, Salt Lake City, Utah (United States); Neuroscience Program, University of Utah, Salt Lake City, Utah (United States); Rabbitt, Richard D. [Bioengineering, University of Utah, Salt Lake City, Utah (United States); Neuroscience Program, University of Utah, Salt Lake City, Utah (United States); Otolaryngology, University of Utah, Salt Lake City, Utah (United States); Marine Biological Laboratory, Woods Hole, Massachusetts (United States)

    2015-12-31

    Glycosaminoglycans (GAGs) are ubiquitous in the inner ear, and disorders altering their structure or production often result in debilitating hearing and balance deficits. The specific mechanisms responsible for loss of hair-cell function are not well understood. We recently reported that introduction of a novel BODIPY conjugated xyloside (BX) into the endolymph primes fluorescent GAGs in vivo [6, 15]. Confocal and two-photon fluorescence imaging revealed rapid turnover and assembly of a glycocalyx enveloping the kinocilia and extending into the cupula, a structure that presumably serves as a mechanical link between the hair bundle and the cupula. Extracellular fluorescence was also observed around the basolateral surface of hair cells and surrounding afferent nerve projections into the crista. Single unit afferent recordings during mechanical hair bundle stimulation revealed temporary interruption of synaptic transmission following BX administration followed by recovery, demonstrating an essential role for GAGs in function of the hair cell synapse. In the present work we present a pharmacokinetic model to quantify the time course of BX primed GAG production and turnover in the ear.

  1. Assignment of hexuronic acid stereochemistry in synthetic heparan sulfate tetrasaccharides with 2-O-sulfo uronic acids using electron detachment dissociation

    Agyekum, Isaac; Patel, Anish B.; Zong, Chengli; Boons, Geert Jan; Amster, I. Jonathan

    2015-01-01

    The present work focuses on the assignment of uronic acid stereochemistry in heparan sulfate (HS) oligomers. The structural elucidation of HS glycosaminoglycans is the subject of considerable importance due to the biological and biomedical significance of this class of carbohydrates. They are highly

  2. Capillary electrophoresis of heparin and other glycosaminoglycans using a polyamine running electrolyte

    Loegel, Thomas N.; Trombley, John D.; Taylor, Richard T. [Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056 (United States); Danielson, Neil D., E-mail: danielnd@muohio.edu [Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056 (United States)

    2012-11-13

    Highlights: Black-Right-Pointing-Pointer Ethylenediamine is likely acting as an ion-pairing agent. Black-Right-Pointing-Pointer Oversulfated chondroitin sulfate is last peak instead of first peak. Black-Right-Pointing-Pointer There is about a factor of five improved detectability with a 12.5 min analysis time. Black-Right-Pointing-Pointer Use of a 50 {mu}m ID capillary is possible. - Abstract: This study involves the use of polyamines as potential resolving agents for the capillary electrophoresis (CE) of glycosaminoglycans (GAGs), specifically heparin, dermatan sulfate, chondroitin sulfate, over-sulfated chondroitin sulfate (OSCS), and hyaluronan. All of the compounds can be separated from each other with the exception of chondroitin sulfate and hyaluronan. Using optimization software, the final run conditions are found to be 200 mM ethylenediamine and 45.5 mM phosphate as the electrolyte with -14 V applied across a 50 {mu}m ID Multiplication-Sign 24.5 cm fused silica capillary at 15 Degree-Sign C. The ion migration order, with OSCS as the last instead of the first peak, is in contrast to previous reports using either a high molarity TRIS or lithium phosphate run buffer with narrower bore capillaries. Total analysis time is 12. 5 min and the relative standard deviation of the heparin migration time is about 2.5% (n = 5). The interaction mechanism between selected polyamines and heparin is explored using conductivity measurements in addition to CE experiments to show that an ion-pairing mechanism is likely.

  3. Structural basis for oligomerization and glycosaminoglycan binding of CCL5 and CCL3.

    Liang, Wenguang G; Triandafillou, Catherine G; Huang, Teng-Yi; Zulueta, Medel Manuel L; Banerjee, Shiladitya; Dinner, Aaron R; Hung, Shang-Cheng; Tang, Wei-Jen

    2016-05-01

    CC chemokine ligand 5 (CCL5) and CCL3 are critical for immune surveillance and inflammation. Consequently, they are linked to the pathogenesis of many inflammatory conditions and are therapeutic targets. Oligomerization and glycosaminoglycan (GAG) binding of CCL5 and CCL3 are vital for the functions of these chemokines. Our structural and biophysical analyses of human CCL5 reveal that CCL5 oligomerization is a polymerization process in which CCL5 forms rod-shaped, double-helical oligomers. This CCL5 structure explains mutational data and offers a unified mechanism for CCL3, CCL4, and CCL5 assembly into high-molecular-weight, polydisperse oligomers. A conserved, positively charged BBXB motif is key for the binding of CC chemokines to GAG. However, this motif is partially buried when CCL3, CCL4, and CCL5 are oligomerized; thus, the mechanism by which GAG binds these chemokine oligomers has been elusive. Our structures of GAG-bound CCL5 and CCL3 oligomers reveal that these chemokine oligomers have distinct GAG-binding mechanisms. The CCL5 oligomer uses another positively charged and fully exposed motif, KKWVR, in GAG binding. However, residues from two partially buried BBXB motifs along with other residues combine to form a GAG-binding groove in the CCL3 oligomer. The N termini of CC chemokines are shown to be involved in receptor binding and oligomerization. We also report an alternative CCL3 oligomer structure that reveals how conformational changes in CCL3 N termini profoundly alter its surface properties and dimer-dimer interactions to affect GAG binding and oligomerization. Such complexity in oligomerization and GAG binding enables intricate, physiologically relevant regulation of CC chemokine functions. PMID:27091995

  4. Altered interaction and distribution of glycosaminoglycans and growth factors in mucopolysaccharidosis type I bone disease.

    Kingma, Sandra D K; Wagemans, Tom; IJlst, Lodewijk; Bronckers, Antonius L J J; van Kuppevelt, Toin H; Everts, Vincent; Wijburg, Frits A; van Vlies, Naomi

    2016-07-01

    The mucopolysaccharidoses (MPSs) comprise a group of lysosomal storage disorders characterized by deficient degradation and subsequent accumulation of glycosaminoglycans (GAGs). Progressive bone and joint disease are a major cause of morbidity, and current therapeutic strategies have limited effect on these symptoms. By elucidating pathophysiological mechanisms underlying bone disease, new therapeutic targets may be identified. Longitudinal growth is regulated by interaction between GAGs and growth factors. Because GAGs accumulate in the MPSs, we hypothesized that altered interaction between growth factors and GAGs contribute to the pathogenesis of MPS bone disease. In this study, binding between GAGs from MPS I chondrocytes and fibroblast growth factor 2 (FGF2) was not significantly different from binding of FGF2 to GAGs from control chondrocytes. FGF2 signaling, however, was increased in MPS I chondrocytes after incubation with FGF2, as compared to control chondrocytes. Using bone cultures, we demonstrated decreased growth of WT mouse bones after incubation with FGF2, but no effect on MPS I bone growth. However, MPS I bones showed decreased growth in the presence of GAGs from MPS I chondrocytes. Finally, we demonstrate altered GAG distribution in MPS I chondrocytes, and altered GAG, FGF2 and Indian hedgehog distribution in growth plates from MPS I mice. In summary, our results suggest that altered interaction and distribution of growth factors and accumulated GAGs may contribute to the pathogenesis of MPS bone disease. In the future, targeting growth factor regulation or the interaction between in growth factors and GAGs might be a promising therapeutic strategy for MPS bone disease. PMID:27105565

  5. N-acetylneuraminlactose sulfate in milk and its role in the synthesis of glycosaminoglycans during development

    Mammals in early life are incapable of synthesizing inorganic sulfate, an important constituent of connective tissue glycosaminoglycans (GAGs). Studies have shown that [35S]Na2SO4 injected into lactating rats is secreted into the milk as [35S] N-acetylneuraminlactose sulfate (NLS); this compound accounts for ≥ 95% of the radioactivity found in milk. They have studied the metabolic fate of the sulfate moiety of NLS in the newborn rat. Lactating rat dams were injected with [35S] Na2SO4 at various times postpartum and pups were allowed to suckle for 48 hrs before sacrifice. Pups of varying ages (3 d. - 19 d.) were sacrificed and the GAGs were prepared from 9 different organs and tissues. Pups of all ages showed appreciable radiosulfate incorporated into all of the tissues and organs examined. In young pups (3 and 5 d old) a high of 75% of the total [35S] sulfate in the homogenate was found in the GAGs of bone, and a low of 27% in the GAGs of liver. Analysis of the composition of the GAGs by enzymatic degradation showed that bone GAGs consisted entirely of chondroitin 4/6 sulfate (C4/6S) while liver GAGs contained 50% C4/6S, 10% dermatan sulfate (DS), and 40% chondroitinase resistant material (probably heparan sulfate). These data show that sulfate in the form of NLS is incorporated in substantial amounts into the sulfated GAGs of the developing rat and support the hypothesis that NLS is an important source of nutrient sulfate during rat development

  6. N-acetylneuraminlactose sulfate in milk and its role in the synthesis of glycosaminoglycans during development

    Kieras, F.J.; Kastin, S.; Rerecich, M.; Sturman, J.A.

    1986-05-01

    Mammals in early life are incapable of synthesizing inorganic sulfate, an important constituent of connective tissue glycosaminoglycans (GAGs). Studies have shown that (/sup 35/S)Na/sub 2/SO/sub 4/ injected into lactating rats is secreted into the milk as (/sup 35/S) N-acetylneuraminlactose sulfate (NLS); this compound accounts for greater than or equal to 95% of the radioactivity found in milk. They have studied the metabolic fate of the sulfate moiety of NLS in the newborn rat. Lactating rat dams were injected with (/sup 35/S) Na/sub 2/SO/sub 4/ at various times postpartum and pups were allowed to suckle for 48 hrs before sacrifice. Pups of varying ages (3 d. - 19 d.) were sacrificed and the GAGs were prepared from 9 different organs and tissues. Pups of all ages showed appreciable radiosulfate incorporated into all of the tissues and organs examined. In young pups (3 and 5 d old) a high of 75% of the total (/sup 35/S) sulfate in the homogenate was found in the GAGs of bone, and a low of 27% in the GAGs of liver. Analysis of the composition of the GAGs by enzymatic degradation showed that bone GAGs consisted entirely of chondroitin 4/6 sulfate (C4/6S) while liver GAGs contained 50% C4/6S, 10% dermatan sulfate (DS), and 40% chondroitinase resistant material (probably heparan sulfate). These data show that sulfate in the form of NLS is incorporated in substantial amounts into the sulfated GAGs of the developing rat and support the hypothesis that NLS is an important source of nutrient sulfate during rat development.

  7. Apoptosis induced by granzyme B-glycosaminoglycan complexes: implications for granule-mediated apoptosis in vivo.

    Galvin, J P; Spaeny-Dekking, L H; Wang, B; Seth, P; Hack, C E; Froelich, C J

    1999-05-01

    Lymphocyte granule-mediated apoptosis occurs by perforin-mediated intracellular delivery of granule-associated serine proteases (granzymes). A granule-associated proteoglycan, namely serglycin, that contains chondroitin 4-sulfate (CS) glycosaminoglycans is present in the granules of cytotoxic cells. Serglycin acts as scaffold for packaging the positively charged granzymes and probably chaperones the proteases secreted extracellularly. To learn how the interaction of granzyme B (GrB) with serglycin might influence the apoptotic potential of this proteases, we have evaluated a model system where desalted CS is combined with isolated human granzyme. CS-GrB complexes were very stable, remaining undissociated in salt concentrations upwards to 500 mM (pH 7.4). On the basis of a capture enzyme immunoassay that accurately detects GrB, equivalent amounts of active free and CS-GrB, delivered by perforin or adenovirus, efficiently induced apoptosis in Jurkat cells and produced a similar time-dependent increase in caspase-3-like activity. CS-GrB processed isolated caspases-3 and -7 less efficiently than free granzyme. However, when added to cytosolic extracts, rates of processing were nearly equivalent for the two forms, suggesting cationic GrB may nonspecifically bind cytosolic proteins, leading to reduce proteolytic activity. Finally, GrB was found to be exocytosed from lymphocyte-activated killer cells as a neutral, high macromolecular weight complex, which possessed apoptotic activity. Collectively, the results indicate that neutral, high m.w. GrB has the capacity to induce cell death and will be useful to study the mechanism of cytotoxic cell-mediated apoptosis in vitro. PMID:10228010

  8. Enterovirus 71 Uses Cell Surface Heparan Sulfate Glycosaminoglycan as an Attachment Receptor

    Tan, Chee Wah; Poh, Chit Laa; Sam, I-Ching

    2013-01-01

    Enterovirus 71 (EV-71) infections are usually associated with mild hand, foot, and mouth disease in young children but have been reported to cause severe neurological complications with high mortality rates. To date, four EV-71 receptors have been identified, but inhibition of these receptors by antagonists did not completely abolish EV-71 infection, implying that there is an as yet undiscovered receptor(s). Since EV-71 has a wide range of tissue tropisms, we hypothesize that EV-71 infections may be facilitated by using receptors that are widely expressed in all cell types, such as heparan sulfate. In this study, heparin, polysulfated dextran sulfate, and suramin were found to significantly prevent EV-71 infection. Heparin inhibited infection by all the EV-71 strains tested, including those with a single-passage history. Neutralization of the cell surface anionic charge by polycationic poly-d-lysine and blockage of heparan sulfate by an anti-heparan sulfate peptide also inhibited EV-71 infection. Interference with heparan sulfate biosynthesis either by sodium chlorate treatment or through transient knockdown of N-deacetylase/N-sulfotransferase-1 and exostosin-1 expression reduced EV-71 infection in RD cells. Enzymatic removal of cell surface heparan sulfate by heparinase I/II/III inhibited EV-71 infection. Furthermore, the level of EV-71 attachment to CHO cell lines that are variably deficient in cell surface glycosaminoglycans was significantly lower than that to wild-type CHO cells. Direct binding of EV-71 particles to heparin-Sepharose columns under physiological salt conditions was demonstrated. We conclude that EV-71 infection requires initial binding to heparan sulfate as an attachment receptor. PMID:23097443

  9. Dexamethasone regulation of glycosaminoglycan synthesis in cultured human skin fibroblasts. Similar effects of glucocorticoid and thyroid hormones.

    Smith, T. J.

    1984-01-01

    The effects of dexamethasone on glycosaminoglycan accumulation were examined in confluent human skin fibroblasts in vitro. The glucocorticoid consistently inhibited the incorporation of either [3H]acetate or [3H]glucosamine into hyaluronate when added to culture medium 72 h before harvest. This effect was half-maximal at approximately 1 nM and maximal at 5-10 nM. Inhibition occurred within 5 h of hormone addition and was near maximal by 25 h. 11 alpha-hydrocortisone (10 nM), deoxycorticostero...

  10. Efficacy of Annona squamosa L in the Synthesis of Glycosaminoglycans and Collagen during Wound Repair in Streptozotocin Induced Diabetic Rats

    Thangavel Ponrasu; Lonchin Suguna

    2014-01-01

    The aim of this work was to find out the effects of Annona squamosa on the formation of glycosaminoglycans and collagen during wound healing in normal and diabetic rats. Diabetes induced rats were segregated into 4 groups, each containing six animals. Groups I and III served as the normal and diabetic control while groups II and IV served as normal and diabetic treated. The animals were treated with 200  μ L of Annona squamosa extract topically. The granulation tissues formed were removed on ...

  11. Extraction and Biochemical Characterization of Sulphated Glycosaminoglycans from Chicken Keel Cartilage

    Humaira Majeed Khan1, Muhammad Ashraf2, Abu Saeed Hashmi3, Mansur-ud-Din Ahmad4 and Aftab Ahmad Anjum5

    2013-11-01

    Full Text Available The present study was conducted to explore the potential and cheaper source of major and abundantly found sulphated glycosaminoglycans (GAGs in chicken keel cartilage. Chicken is comparatively readily accessible to all the communities of Pakistan and its cartilages are the rich source of sulphated GAGs. The GAGs were extracted from prewashed and ground keel cartilages (n=3 of chicken using 3 M MgCl2, dialyzed, digested with papain, precipitated with three volumes of ethanol, and finally lyophilized to dry powder. The dry products were used for proximate analysis (carbohydrates 65.49±0.10, crude protein 12.82±0.26, ash 11.12±.56, moisture 9.88±0.32 and fat 0.69±0.14%. Dimethylmethylene blue binding (DMMB assay was performed to determine the quantity of total GAGs in each group of product and protein contents were estimated by Bradford method. Identification of extracted samples of GAGs was performed with FTIR spectrometer using KBr disc and purity of the samples was determined by SDS-PAGE. Quantity of total GAGs in extracted samples was 70.77±2.27% and estimated amount of protein was 4.64±0.29%. FTIR spectra of standard and samples of CS showed identical and characteristic peaks in finger print region. Finger print region revealed the presence of C-O-S, S=O, -COO, -C-C, R-SO2–R, -CONH2 and R-SO2-NH2 molecules. SDS-PAGE analysis revealed the presence of 77.8 and 50.5 kDa proteins in all extracted samples of GAGs. It can be concluded that chicken keel cartilage is the potential and cheap source of GAGs. Analysis by SDS-PAGE revealed that most of the non-collagen protein can be removed by three volumes of solvent extraction and FTIR is an advance technique for identification of GAGs in mid infrared region (400-4000 cm-1.

  12. Glycosaminoglycan-functionalized poly-lactide-co-glycolide nanoparticles: synthesis, characterization, cytocompatibility, and cellular uptake

    Lamichhane SP

    2015-01-01

    Full Text Available Surya P Lamichhane,1 Neha Arya,1,2 Nirdesh Ojha,3 Esther Kohler,1 V Prasad Shastri1,2,41Institute for Macromolecular Chemistry, University of Freiburg, Freiburg, 2Helmholtz Virtual Institute on “Multifunctional Biomaterials for Medicine”, 3Laboratory for Process Technology, Department of Microsystems Engineering, University of Freiburg, Freiburg, 4Centre for Biological Signaling Studies (BIOSS, University of Freiburg, Freiburg, GermanyAbstract: The efficient delivery of chemotherapeutics to the tumor via nanoparticle (NP-based delivery systems remains a significant challenge. This is compounded by the fact that the tumor is highly dynamic and complex environment composed of a plurality of cell types and extracellular matrix. Since glycosaminoglycan (GAG production is altered in many diseases (or pathologies, NPs bearing GAG moieties on the surface may confer some unique advantages in interrogating the tumor microenvironment. In order to explore this premise, in the study reported here poly-lactide-co-glycolide (PLGA NPs in the range of 100–150 nm bearing various proteoglycans were synthesized by a single-step nanoprecipitation and characterized. The surface functionalization of the NPs with GAG moieties was verified using zeta potential measurements and X-ray photoelectron spectroscopy. To establish these GAG-bearing NPs as carriers of therapeutics, cellular toxicity assays were undertaken in lung epithelial adenocarcinoma (A549 cells, human pulmonary microvascular endothelial cells (HPMEC, and renal proximal tubular epithelial cells. In general NPs were well tolerated over a wide concentration range (100–600 µg/mL by all cell types and were taken up to appreciable extents without any adverse cell response in A549 cells and HPMEC. Further, GAG-functionalized PLGA NPs were taken up to different extents in A459 cells and HPMEC. In both cell systems, the uptake of heparin-modified NPs was diminished by 50%–65% in comparison to that of

  13. Fact versus artifact: Avoiding erroneous estimates of sulfated glycosaminoglycan content using the dimethylmethylene blue colorimetric assay for tissue-engineered constructs

    CH Zheng

    2015-04-01

    Full Text Available The 1,9-dimethylmethylene blue (DMMB assay is widely used to quantify sulfated glycosaminoglycan (sGAG contents of engineered tissues, culture media, tissue samples and bodily fluids, but the assay is subject to interference from polyanions such as hyaluronic acid (HA, DNA and RNA. We examined whether specific combinations of dye pH and absorbance wavelength could minimize non-sGAG artifacts without compromising DMMB assay sensitivity. HA and DNA solutions generated substantial signal at pH 3 but not at pH 1.5. Reducing dye pH did not significantly alter sGAG measurements for normal cartilage and meniscus tissues, but eliminated anomalously high apparent sGAG contents for enzymatically isolated chondrocytes, adipose-derived stem cell (ADSC-agarose constructs and ADSC pellets. In a cartilage tissue-engineering case study, pH 3 dye indicated high apparent sGAG readings throughout culture in both basal and chondrogenic media, with a marked decline between day 14 and 21 for chondrogenic constructs. The pH 1.5 dye, however, indicated minimal sGAG accumulation in basal medium and stable sGAG content throughout culture in chondrogenic medium. As it is often difficult to know a priori whether all groups in a study will have sGAG contents high enough to overwhelm artifacts, we recommend modifying the standard DMMB assay to reduce the risk of spurious findings in tissue engineering and clinical research. Specifically, we recommend shifting to a pH 1.5 DMMB dye and basing quantification on the absorbance difference between 525 nm (µ peak and 595 nm (β peak to compensate for the moderate loss of sensitivity associated with reducing the dye pH.

  14. Oxidative and nitrative stress and pro-inflammatory cytokines in Mucopolysaccharidosis type II patients: effect of long-term enzyme replacement therapy and relation with glycosaminoglycan accumulation.

    Jacques, Carlos Eduardo Diaz; Donida, Bruna; Mescka, Caroline P; Rodrigues, Daiane G B; Marchetti, Desirèe P; Bitencourt, Fernanda H; Burin, Maira G; de Souza, Carolina F M; Giugliani, Roberto; Vargas, Carmen Regla

    2016-09-01

    Mucopolysaccharidosis type II (MPS II) is a lysosomal storage disease caused by a deficient activity of iduronate-2-sulfatase, leading to abnormal accumulation of glycosaminoglycans (GAG). The main treatment for MPS II is enzyme replacement therapy (ERT). Previous studies described potential benefits of six months of ERT against oxidative stress in patients. Thus, the aim of this study was to investigate oxidative, nitrative and inflammatory biomarkers in MPS II patients submitted to long term ERT. It were analyzed urine and blood samples from patients on ERT (mean time: 5.2years) and healthy controls. Patients presented increased levels of lipid peroxidation, assessed by urinary 15-F2t-isoprostane and plasmatic thiobarbituric acid-reactive substances. Concerning to protein damage, urinary di-tyrosine (di-Tyr) was increased in patients; however, sulfhydryl and carbonyl groups in plasma were not altered. It were also verified increased levels of urinary nitrate+nitrite and plasmatic nitric oxide (NO) in MPS II patients. Pro-inflammatory cytokines IL-1β and TNF-α were increased in treated patients. GAG levels were correlated to di-Tyr and nitrate+nitrite. Furthermore, IL-1β was positively correlated with TNF-α and NO. Contrastingly, we did not observed alterations in erythrocyte superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase activities, in reduced glutathione content and in the plasmatic antioxidant capacity. Although some parameters were still altered in MPS II patients, these results may suggest a protective role of long-term ERT against oxidative stress, especially upon oxidative damage to protein and enzymatic and non-enzymatic defenses. Moreover, the redox imbalance observed in treated patients seems to be GAG- and pro-inflammatory cytokine-related. PMID:27251652

  15. Unusual Glycosaminoglycans from a Deep Sea Hydrothermal Bacterium Improve Fibrillar Collagen Structuring and Fibroblast Activities in Engineered Connective Tissues

    Jean Guezennec

    2013-04-01

    Full Text Available Biopolymers produced by marine organisms can offer useful tools for regenerative medicine. Particularly, HE800 exopolysaccharide (HE800 EPS secreted by a deep-sea hydrothermal bacterium displays an interesting glycosaminoglycan-like feature resembling hyaluronan. Previous studies demonstrated its effectiveness to enhance in vivo bone regeneration and to support osteoblastic cell metabolism in culture. Thus, in order to assess the usefulness of this high-molecular weight polymer in tissue engineering and tissue repair, in vitro reconstructed connective tissues containing HE800 EPS were performed. We showed that this polysaccharide promotes both collagen structuring and extracellular matrix settle by dermal fibroblasts. Furthermore, from the native HE800 EPS, a low-molecular weight sulfated derivative (HE800 DROS displaying chemical analogy with heparan-sulfate, was designed. Thus, it was demonstrated that HE800 DROS mimics some properties of heparan-sulfate, such as promotion of fibroblast proliferation and inhibition of matrix metalloproteinase (MMP secretion. Therefore, we suggest that the HE800EPS family can be considered as an innovative biotechnological source of glycosaminoglycan-like compounds useful to design biomaterials and drugs for tissue engineering and repair.

  16. Hyaluronic acid changes of the extracellular matrix in colon carcinoma

    Arab MR.; Allahyari A.; Sargolzaie Aval F.; Rafighdoost H; Karimi M

    2007-01-01

    Background: The extracellular matrix is a complex three-dimensional network of proteins and glycosaminoglycans, which have important roles in cellular physiology and cell-cell and cell-extracellular matrix interactions. Any changes in the extracellular matrix of tumors may be implicated in cellular transformation and metastasis. The aim of the present study was to identify changes in the hyaluronic acid of the stroma of colonic carcinoma.Methods: Paraffin blocks of 30 patients with colon carc...

  17. Hyaluronic acid: A promising mediator for periodontal regeneration

    Bansal Jyoti; Kedige Suresh; Anand Samir

    2010-01-01

    Hyaluronic acid (HA) is a natural-non sulphated high molecular weight glycosaminoglycan that forms a critical component of the extracellular matrix and contributes significantly to tissue hydrodynamics, cell migration and proliferation. The use of HA in the treatment of inflammatory process is established in medical areas such as orthopedics, dermatology and ophthalmology. In the field of dentistry, hyaluronate has shown anti-inflammatory, antiedematous and anti-bacterial effects for the trea...

  18. Chitosan Scaffolds Containing Hyaluronic Acid for Cartilage Tissue Engineering

    Correia, Clara R.; Moreira Teixeira, Liliana S.; Moroni, Lorenzo; Reis, Rui L.; Blitterswijk, van, C.A.; Karperien, Marcel; Mano, João F.

    2011-01-01

    Scaffolds derived from natural polysaccharides are very promising in tissue engineering applications and regenerative medicine, as they resemble glycosaminoglycans in the extracellular matrix (ECM). In this study, we have prepared freeze-dried composite scaffolds of chitosan (CHT) and hyaluronic acid (HA) in different weight ratios containing either no HA (control) or 1%, 5%, or 10% of HA. We hypothesized that HA could enhance structural and biological properties of CHT scaffolds. To test thi...

  19. The TSG-6 and IαI Interaction Promotes a Transesterification Cleaving the Protein-Glycosaminoglycan-Protein (PGP) Cross-link

    Sanggaard, Kristian Wejse; Karring, Henrik; Valnickova, Zuzana;

    2005-01-01

    , were formed. The formation of these complexes was prevented by the addition of hyaluronan. The cross-links stabilizing these complexes displaying properties similar to the protein-glycosaminoglycan-protein (PGP) cross-link. The TSG-6-containing SDS-stable complexes were composed of HC1·TSG-6 or HC2·TSG...

  20. Effects of retinal growth factor and of the increase of the number of subcultures on sulfated glycosaminoglycans of bovine lens epithelial cells

    Sulfated glycosaminoglycans of cultured bovine lens epithelial cells grown in the presence and in the absence of a retinal growth factor were investigated comparatively. The newly formed [35S] sulfate-labeled glycosaminoglycans were analysed in the extra-, peri- and intracellular compartments of early (4-5th) and late (17-18h) subcultures. The following results were obtained: (1) Cultured lens epithelial cells grown in the presence or in the absence of the growth factor synthesize chondroitin 4- and 6-sulfates and dermatan sulfate, with heparan sulfate as the main component, the pericellular compartments were particularly rich in heparan sulfate; (2) The distribution pattern of the glycosaminoglycans changes during successive subcultures; the proportion of heparan sulfate increases in the pericellular compartment, the dermatan sulfate to chondroitin sulfate ratio increases in all three compartments; (3) In contrast to the drastic decrease in the fibronectin levels in the presence of growth factor in the early subcultures, only minor differences were found between the glycosaminoglycan patterns for the treated and non-treated cells. ( orig.)

  1. Evaluation of early healing events around mesenchymal stem cell-seeded collagen-glycosaminoglycan scaffold. An experimental study in Wistar rats.

    Alhag, Mohamed

    2011-03-01

    Tissue engineering using cell-seeded biodegradable scaffolds offers a new bone regenerative approach that might circumvent many of the limitations of current therapeutic modalities. The aim of this experiment was to study the early healing events around mesenchymal stem cell-seeded collagen-glycosaminoglycan scaffolds.

  2. Identification of Chemically Sulfated/desulfated Glycosaminoglycans in Contaminated Heparins and Development of a Simple Assay for the Detection of Most Contaminants in Heparin

    Pan, Jing; Qian, Yi; Zhou, Xiaodong; Pazandak, Andrew; Frazier, Sarah B.; Weiser, Peter; Lu, Hong; Zhang, Lijuan

    2010-01-01

    Contaminated heparin was linked to at least 149 deaths and hundreds of adverse reactions. Published report indicates that heparin contaminants were a natural impurity, dermatan sulfate, and a contaminant, oversulfated chondroitin sulfate (OSCS). OSCS was assumed to derive from animal cartilage. By analyzing 26 contaminated heparin lots from different sources, our data indicate that the heparin contaminants were chemically sulfated or chemically sulfated/desulfated glycosaminoglycans (GAGs) co...

  3. Collagen and Glycosaminoglycan Profiles in the Canine Cervix during Different Stages of the Estrous Cycle and in Open- and Closed-Cervix Pyometra

    LINHARATTANARUKSA, Pichanun; SRISUWATANASAGUL, Sayamon; PONGLOWHAPAN, Suppawiwat; Khalid, Muhammad; Chatdarong, Kaywalee

    2013-01-01

    ABSTRACT The extracellular matrix of the cervix that comprises collagen, elastin, proteoglycans and glycosaminoglycans (GAGs) is thought to have an essential role in cervical relaxation. This study investigated the proportion of collagen and smooth muscle as well as the GAGs in cervices obtained from healthy bitches at different stages of the estrous cycle and bitches with open- and closed-cervix pyometra. Cervices were collected after ovariohysterectomy. The proportion of collagen to smooth ...

  4. Clinical cases of joint disease in horse. Total glycosaminoglycans sulphate and keratansulphate in synovial fluid as markers of degenerative cartilage processes

    Total glycosaminoglycans sulphate (GAGs) and keratan sulphate (KS) were measured in synovial fluid (SF) obtained from 28 horses with different joint diseases (degenerative joint disease (DJD), osteochondrosis (OCD), positivity to Flex Test (FT)) and 15 horses without any clinical sign of lameness. All groups of animals with joint disease showed levels of total GAGs significantly higher (P0.001) than normal. On the contrary, only DJD affected joints showed a significantly (P0.01) higher level of KS

  5. Preventive effect of glycosaminoglycans from Amussium pleuronectus (Linne) on biomolecules, lactate dehydrogenase-isoenzyme and electrocardiographic patterns in isoproterenol-induced myocardial infarction in Wistar rats

    Ramachandran Saravanan; Annaian Shanmugam; Devaraj Rajkumar

    2012-01-01

    Objectives: This study was aimed to assess the cardioprotective role of low molecular weight glycosaminoglycans (LMW-GAG) in isoproterenol (ISO)-induced myocardial infarction (MI) in male Wistar rats. Effect of LMW-GAG on biomolecules, lactate dehydrogenase (LDH)-isoenzyme, and electrocardiographic (ECG)-patterns was studied as evidence of cardioprotection. Materials and Methods: Male Wistar rats (140 ± 10 g) were divided into four groups; untreated control (group I), LMW-GAG treated (300 ...

  6. Preparation and characterization of molecular weight fractions of glycosaminoglycan from sea cucumber Thelenata ananas using free radical depolymerization.

    Wu, Mingyi; Xu, Shimin; Zhao, Jinhua; Kang, Hui; Ding, Hui

    2010-03-30

    A glycosaminoglycan from sea cucumber Thelenata anana (THG) was isolated as a polymer of molecular weight of around 70 kDa. Its low molecular weight derivatives were first prepared by free radical depolymerization with hydrogen peroxide in the presence of copper(II) ion. The parameters of the process were investigated by a high-performance gel permeation chromatography. Analyses of chemical composition and molecular weight distribution indicated that the fragmentation of the main-chain of THG occurred randomly, obeyed pseudo first-order kinetics, and produced species with rather narrow and unimodal distribution of molar mass. The characterization of different molecular weight fractions was investigated by using viscometry and atomic force microscopy (AFM). Analysis of molecular weight and intrinsic viscosity in terms of the known theories for unperturbed wormlike cylinder yielded 1201+/-110 nm(-1), 15.3+/-1.5 nm, and 1.5+/-0.3 nm for molar mass per unit contour length M(L), persistence length q, and diameter d, respectively. The M(L) and d values were approximately consistent with those observed by AFM. The present data suggest that THG may dissolve in 0.1M aqueous NaCl as single-stranded helical chains. PMID:20117762

  7. The effect of glycosaminoglycan enzymes and proteases on the viscosity of alpaca seminal plasma and sperm function.

    Kershaw-Young, C M; Stuart, C; Evans, G; Maxwell, W M C

    2013-05-01

    In order to advance the development of cryopreservation and other assisted reproductive technologies in camelids it is necessary to eliminate the viscous component of the seminal plasma without impairing sperm function. It has been postulated that glycosaminoglycans (GAGs) or proteoglycans are responsible for this viscosity. This study investigated the effect of the GAG enzymes hyaluronidase, chondroitinase ABC and keratanase and the proteases papain and proteinase K on seminal plasma viscosity and sperm function in order to aid identification of the cause of seminal plasma viscosity and propose methods for the reduction of viscosity. Sperm motility, DNA integrity, acrosome integrity and viability were assessed during 2h incubation. All enzymes reduced seminal plasma viscosity compared to control (Pviscosity within 30 min of treatment. Sperm motility and DNA integrity was not affected by enzyme treatment. The proportion of viable, acrosome intact sperm was reduced in all enzyme treated samples except those treated with papain (Pviscosity. Papain treatment of alpaca semen may be a suitable technique for reduction of seminal plasma viscosity prior to sperm cryopreservation. PMID:23537479

  8. Effect of castration on renal glycosaminoglycans and their urinary excretion in male and female rats with chronic renal failure

    Lemos, C.C.S. [Disciplina de Nefrologia, Faculdade de Ciências Médicas, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, RJ (Brazil); Tovar, A.M.F. [Laboratório de Tecido Conjuntivo, Instituto de Bioquímica Médica, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ (Brazil); Guimarães, M.A.M. [Departamento de Patologia e Laboratórios, Faculdade de Ciências Médicas, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, RJ (Brazil); Bregman, R. [Disciplina de Nefrologia, Faculdade de Ciências Médicas, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, RJ (Brazil)

    2013-08-10

    Glycosaminoglycans (GAGs) participate in a variety of processes in the kidney, and evidence suggests that gender-related hormones participate in renal function. The aim of this study was to analyze the relationship of GAGs, gender, and proteinuria in male and female rats with chronic renal failure (CRF). GAGs were analyzed in total kidney tissue and 24-h urine of castrated (c), male (M), and female (F) Wistar control (C) rats (CM, CMc, CF, CFc) and after 30 days of CRF induced by 5/6 nephrectomy (CRFM, CRFMc, CRFF, CRFFc). Total GAG quantification and composition were determined using agarose and polyacrylamide gel electrophoresis, respectively. Renal GAGs were higher in CF compared to CM. CRFM presented an increase in renal GAGs, heparan sulfate (HS), and proteinuria, while castration reduced these parameters. However, CRFF and CRFFc groups showed a decrease in renal GAGs concomitant with an increase in proteinuria. Our results suggest that, in CRFM, sex hormones quantitatively alter GAGs, mainly HS, and possibly the glomerular filtration barrier, leading to proteinuria. The lack of this response in CRFMc, where HS did not increase, corroborates this theory. This pattern was not observed in females. Further studies of CRF are needed to clarify gender-dependent differences in HS synthesis.

  9. Host cell heparan sulfate glycosaminoglycans are ligands for OspF-related proteins of the Lyme disease spirochete.

    Lin, Yi-Pin; Bhowmick, Rudra; Coburn, Jenifer; Leong, John M

    2015-10-01

    Borrelia burgdorferi, the agent of Lyme disease, spreads from the site of the tick bite to tissues such as heart, joints and the nervous tissues. Host glycosaminoglycans, highly modified repeating disaccharides that are present on cell surfaces and in extracellular matrix, are common targets of microbial pathogens during tissue colonization. While several dermatan sulfate-binding B. burgdorferi adhesins have been identified, B. burgdorferi adhesins documented to promote spirochetal binding to heparan sulfate have not yet been identified. OspEF-related proteins (Erps), a large family of plasmid-encoded surface lipoproteins that are produced in the mammalian host, can be divided into the OspF-related, OspEF-leader peptide (Elp) and OspE-related subfamilies. We show here that a member of the OspF-related subfamily, ErpG, binds to heparan sulfate and when produced on the surface of an otherwise non-adherent B. burgdorferi strain, ErpG promotes heparan sulfate-mediated bacterial attachment to the glial but not the endothelial, synovial or respiratory epithelial cells. Six other OspF-related proteins were capable of binding heparan sulfate, whereas representative OspE-related and Elp proteins lacked this activity. These results indicate that OspF-related proteins are heparan sulfate-binding adhesins, at least one of which promotes bacterial attachment to glial cells. PMID:25864455

  10. Glycosaminoglycan Profiling in Patients' Plasma and Urine Predicts the Occurrence of Metastatic Clear Cell Renal Cell Carcinoma.

    Gatto, Francesco; Volpi, Nicola; Nilsson, Helén; Nookaew, Intawat; Maruzzo, Marco; Roma, Anna; Johansson, Martin E; Stierner, Ulrika; Lundstam, Sven; Basso, Umberto; Nielsen, Jens

    2016-05-24

    Metabolic reprogramming is a hallmark of clear cell renal cell carcinoma (ccRCC) progression. Here, we used genome-scale metabolic modeling to elucidate metabolic reprogramming in 481 ccRCC samples and discovered strongly coordinated regulation of glycosaminoglycan (GAG) biosynthesis at the transcript and protein levels. Extracellular GAGs are implicated in metastasis, so we speculated that such regulation might translate into a non-invasive biomarker for metastatic ccRCC (mccRCC). We measured 18 GAG properties in 34 mccRCC samples versus 16 healthy plasma and/or urine samples. The GAG profiles were distinctively altered in mccRCC. We derived three GAG scores that distinguished mccRCC patients with 93.1%-100% accuracy. We validated the score accuracies in an independent cohort (up to 18 mccRCC versus nine healthy) and verified that the scores normalized in eight patients with no evidence of disease. In conclusion, coordinated regulation of GAG biosynthesis occurs in ccRCC, and non-invasive GAG profiling is suitable for mccRCC diagnosis. PMID:27184840

  11. Inhibition of herpes simplex virus infection by lactoferrin is dependent on interference with the virus binding to glycosaminoglycans

    Previous reports have indicated that lactoferrin inhibits herpes simplex virus (HSV) infection during the very early phases of the viral replicative cycle. In the present work we investigated the mechanism of the antiviral activity of lactoferrin in mutant glycosaminoglycan (GAG)-deficient cells. Bovine lactoferrin (BLf) was a strong inhibitor of HSV-1 infection in cells expressing either heparan sulfate (HS) or chondroitin sulfate (CS) or both, but was ineffective or less efficient in GAG-deficient cells or in cells treated with GAG-degrading enzymes. In contrast to wild-type HSV-1, virus mutants devoid of glycoprotein C (gC) were significantly less inhibited by lactoferrin in GAG-expressing cells, indicating that lactoferrin interfered with the binding of viral gC to cell surface HS and/or CS. Finally, we demonstrated that lactoferrin bound directly to both HS and CS isolated from surfaces of the studied cells, as well as to commercial preparations of GAG chains. The results support the hypothesis that the inhibition of HSV-1 infectivity by lactoferrin is dependent on its interaction with cell surface GAG chains of HS and CS

  12. Artificial Extracellular Matrices with Oversulfated Glycosaminoglycan Derivatives Promote the Differentiation of Osteoblast-Precursor Cells and Premature Osteoblasts

    Ute Hempel

    2014-01-01

    Full Text Available Sulfated glycosaminoglycans (GAG are components of the bone marrow stem cell niche and to a minor extent of mature bone tissue with important functions in regulating stem cell lineage commitment and differentiation. We anticipated that artificial extracellular matrices (aECM composed of collagen I and synthetically oversulfated GAG derivatives affect preferentially the differentiation of osteoblast-precursor cells and early osteoblasts. A set of gradually sulfated chondroitin sulfate and hyaluronan derivatives was used for the preparation of aECM. All these matrices were analysed with human bone marrow stromal cells to identify the most potent aECM and to determine the influence of the degree and position of sulfate groups and the kind of disaccharide units on the osteogenic differentiation. Oversulfated GAG derivatives with a sulfate group at the C-6 position of the N-acetylglycosamine revealed the most pronounced proosteogenic effect as determined by tissue nonspecific alkaline phosphatase activity and calcium deposition. A subset of the aECM was further analysed with different primary osteoblasts and cell lines reflecting different maturation stages to test whether the effect of sulfated GAG derivatives depends on the maturation status of the cells. It was shown that the proosteogenic effect of aECM was most prominent in early osteoblasts.

  13. Effect of castration on renal glycosaminoglycans and their urinary excretion in male and female rats with chronic renal failure

    Glycosaminoglycans (GAGs) participate in a variety of processes in the kidney, and evidence suggests that gender-related hormones participate in renal function. The aim of this study was to analyze the relationship of GAGs, gender, and proteinuria in male and female rats with chronic renal failure (CRF). GAGs were analyzed in total kidney tissue and 24-h urine of castrated (c), male (M), and female (F) Wistar control (C) rats (CM, CMc, CF, CFc) and after 30 days of CRF induced by 5/6 nephrectomy (CRFM, CRFMc, CRFF, CRFFc). Total GAG quantification and composition were determined using agarose and polyacrylamide gel electrophoresis, respectively. Renal GAGs were higher in CF compared to CM. CRFM presented an increase in renal GAGs, heparan sulfate (HS), and proteinuria, while castration reduced these parameters. However, CRFF and CRFFc groups showed a decrease in renal GAGs concomitant with an increase in proteinuria. Our results suggest that, in CRFM, sex hormones quantitatively alter GAGs, mainly HS, and possibly the glomerular filtration barrier, leading to proteinuria. The lack of this response in CRFMc, where HS did not increase, corroborates this theory. This pattern was not observed in females. Further studies of CRF are needed to clarify gender-dependent differences in HS synthesis

  14. Identification and tissue-specific distribution of sulfated glycosaminoglycans in the blood-sucking bug Rhodnius prolixus (Linnaeus).

    Costa-Filho, Adilson; Souza, Maisa L S; Martins, Rita C L; dos Santos, André V F; Silva, Gabriela V; Comaru, Michele W; Moreira, Mônica F; Atella, Georgia C; Allodi, Silvana; Nasciutti, Luiz E; Masuda, Hatisaburo; Silva, Luiz-Claudio F

    2004-03-01

    We have previously characterized heparan sulfate (HS) as the major ovarian sulfated glycosaminoglycan (GAG) in females of Rhodnius prolixus, while chondroitin sulfate (CS) was the minor component. Using histochemical procedures we found that GAGs were concentrated in the ovarian tissue but not found inside the oocytes. Here, we extend our initial observations of GAG expression in R. prolixus by characterizing these molecules in other organs: the fat body, intestinal tract, and the reproductive tracts. Only HS and CS were found in the three organs analyzed, however CS was the major GAG species in these tissues. We also determined the compartmental distribution of GAGs in these organs by histochemical analysis using 1,9-dimethylmethylene blue, and evaluated the specific distribution of CS within both male and female reproductive tracts by immunohistochemistry using an anti-CS antibody. We also determined the GAG composition in eggs at days 0 and 6 of embryonic development. Only HS and CS were found in eggs at day 6, while no sulfated GAGs were detected at day 0. Our results demonstrate that HS and CS are the only sulfated GAG species expressed in the fat body and in the intestinal and reproductive tracts of Rhodnius male and female adults. Both sulfated GAGs were also identified in Rhodnius embryos. Altogether, these results show no qualitative differences in the sulfated GAG composition regarding tissue-specific or development-specific distribution. PMID:14871621

  15. Nuclear Magnetic Resonance Insight into the Multiple Glycosaminoglycan Binding Modes of the Link Module from Human TSG-6.

    Park, Younghee; Jowitt, Thomas A; Day, Anthony J; Prestegard, James H

    2016-01-19

    Tumor necrosis factor-stimulated gene-6 (TSG-6) is a hyaluronan (HA)-binding protein that is essential for stabilizing and remodeling the extracellular matrix (ECM) during ovulation and inflammatory disease processes such as arthritis. The Link module, one of the domains of TSG-6, is responsible for binding hyaluronan and other glycosaminoglycans found in the ECM. In this study, we used a well-defined chondroitin sulfate (CS) hexasaccharide (ΔC444S) to determine the structure of the Link module, in solution, in its chondroitin sulfate-bound state. A variety of nuclear magnetic resonance techniques were employed, including chemical shift perturbation, residual dipolar couplings (RDCs), nuclear Overhauser effects, spin relaxation measurements, and paramagnetic relaxation enhancements from a spin-labeled analogue of ΔC444S. The binding site for ΔC444S on the Link module overlapped with that of HA. Surprisingly, ΔC444S binding induced dimerization of the Link module (as confirmed by analytical ultracentrifugation), and a second weak binding site that partially overlapped with a previously identified heparin site was detected. A dimer model was generated using chemical shift perturbations and RDCs as restraints in the docking program HADDOCK. We postulate that the molecular cross-linking enhanced by the multiple binding modes of the Link module might be critical for remodeling the ECM during inflammation/ovulation and might contribute to other functions of TSG-6. PMID:26685054

  16. Evaluation of the experimental parameters which control electron detachment dissociation, and their effect on the fragmentation efficiency of glycosaminoglycan carbohydrates

    Leach, Franklin E., III; Wolff, Jeremy J.; Laremore, Tatiana N.; Linhardt, Robert J.; Amster, I. Jonathan

    2008-10-01

    The efficiency of conversion of precursor ions to observable products for electron detachment dissociation (EDD) was measured as a function of the key experimental parameters to determine their optimal values for the Fourier transform mass spectrometry analysis of anionic glycosaminoglycan carbohydrates. These parameters include electron current, electron energy, dispenser cathode heater current, electron beam duration, charge state of the precursor ion, oligomer length, and precursor ion number accumulated in an external radio frequency multipole trap. Precursor conversion is most efficient at an electron current of 15 [mu]A, and decreases at higher and lower values. The conversion of precursor to product ions increases in efficiency as the electron pulse duration is increased. Together, these data suggest that a radially repulsive electric field is produced between the electron beam and negative ions during EDD which causes the reaction cross-section to decrease at higher values of electron current (>15 [mu]A). Elevating the heater current of the dispenser cathode increases the electron flux, but also causes ion activation, presumably by blackbody infrared irradiation. An electronic circuit is described that allows the electron current produced by the dispenser cathode to be measured during an EDD or electron capture dissociation (ECD) experiment.

  17. In vitro evaluation of antiviral and virucidal activity of a high molecular weight hyaluronic acid

    Blasi Elisabetta; Ardizzoni Andrea; Neglia Rachele G; Bettua Clotilde; Scuri Monica; Cuoghi Alessandro; Cermelli Claudio; Iannitti Tommaso; Palmieri Beniamino

    2011-01-01

    Abstract Background hyaluronic acid (HA), a non-sulphated glycosaminoglycan, is present in synovial fluid, vitreous humour serum and many connective tissues. Pharmaceutical preparations of HA are used in clinical practice for wound healing, joint pain, kerato-conjunctivitis, asthma, mouth care, oesophageal-reflux, and gastritis. Moreover, it is used as a filler to counteract ageing and facial lipoatrophy. Our study aims at investigating the in vitro antiviral activity of a high molecular weig...

  18. Molecular transformations in connective tissue hyaluronic acid

    Free radicals, either induced by the action of ionizing radiations or produced by metal ion induced electron transfer reactions in situ, can initiate a marked reduction in the viscoelasticity of the connective tissue matrix. This paper examines the dominant role of hyaluronic acid in controlling this behavior at molecular level. The results indicate that after a dose of 5Gy (500 rads), the average molecular weight of hyaluronic acid in skin would be reduced by a factor of 4, which would lead to a 60-fold reduction in viscosity of the glycosaminoglycan. Shorter chains so produced would further inhibit hyperentanglement and chain-chain interactions which are responsible for the viscoelasticity of the hyaluronic acid-polymer network. The results are relevant to preservation of skin grafts and to the effects of low-dose radiation in vivo

  19. Biochemical imaging of cervical intervertebral discs with glycosaminoglycan chemical exchange saturation transfer magnetic resonance imaging: feasibility and initial results

    Schleich, Christoph; Mueller-Lutz, Anja; Zimmermann, Lisa; Boos, Johannes; Wittsack, Hans-Joerg; Antoch, Gerald; Miese, Falk [Department of Diagnostic and Interventional Radiology, University Dusseldorf, Medical Faculty, Dusseldorf (Germany); Schmitt, Benjamin [Siemens Ltd. Australia, Healthcare Sector, Macquarie Park, NSW (Australia)

    2016-01-15

    To evaluate glycosaminoglycan chemical exchange saturation transfer (gagCEST) imaging at 3T in the assessment of the GAG content of cervical IVDs in healthy volunteers. Forty-two cervical intervertebral discs of seven healthy volunteers (four females, three males; mean age: 21.4 ± 1.4 years; range: 19-24 years) were examined at a 3T MRI scanner in this prospective study. The MRI protocol comprised standard morphological, sagittal T2 weighted (T2w) images to assess the magnetic resonance imaging (MRI) based grading system for cervical intervertebral disc degeneration (IVD) and biochemical imaging with gagCEST to calculate a region-of-interest analysis of nucleus pulposus (NP) and annulus fibrosus (AF). GagCEST of cervical IVDs was technically successful at 3T with significant higher gagCEST values in NP compared to AF (1.17 % ± 1.03 % vs. 0.79 % ± 1.75 %; p = 0.005). We found topological differences of gagCEST values of the cervical spine with significant higher gagCEST effects in lower IVDs (r = 1; p = 0). We could demonstrate a significant, negative correlation between gagCEST values and cervical disc degeneration of NP (r = -0.360; p = 0.019). Non-degenerated IVDs had significantly higher gagCEST effects compared to degenerated IVDs in NP (1.76 % ± 0.92 % vs. 0.52 % ± 1.17 %; p < 0.001). Biochemical imaging of cervical IVDs is feasible at 3T. GagCEST analysis demonstrated a topological GAG distribution of the cervical spine. The depletion of GAG in the NP with increasing level of morphological degeneration can be assessed using gagCEST imaging. (orig.)

  20. A glycosaminoglycan based, modular tissue scaffold system for rapid assembly of perfusable, high cell density, engineered tissues.

    Ramkumar Tiruvannamalai-Annamalai

    Full Text Available The limited ability to vascularize and perfuse thick, cell-laden tissue constructs has hindered efforts to engineer complex tissues and organs, including liver, heart and kidney. The emerging field of modular tissue engineering aims to address this limitation by fabricating constructs from the bottom up, with the objective of recreating native tissue architecture and promoting extensive vascularization. In this paper, we report the elements of a simple yet efficient method for fabricating vascularized tissue constructs by fusing biodegradable microcapsules with tunable interior environments. Parenchymal cells of various types, (i.e. trophoblasts, vascular smooth muscle cells, hepatocytes were suspended in glycosaminoglycan (GAG solutions (4%/1.5% chondroitin sulfate/carboxymethyl cellulose, or 1.5 wt% hyaluronan and encapsulated by forming chitosan-GAG polyelectrolyte complex membranes around droplets of the cell suspension. The interior capsule environment could be further tuned by blending collagen with or suspending microcarriers in the GAG solution These capsule modules were seeded externally with vascular endothelial cells (VEC, and subsequently fused into tissue constructs possessing VEC-lined, inter-capsule channels. The microcapsules supported high density growth achieving clinically significant cell densities. Fusion of the endothelialized, capsules generated three dimensional constructs with an embedded network of interconnected channels that enabled long-term perfusion culture of the construct. A prototype, engineered liver tissue, formed by fusion of hepatocyte-containing capsules exhibited urea synthesis rates and albumin synthesis rates comparable to standard collagen sandwich hepatocyte cultures. The capsule based, modular approach described here has the potential to allow rapid assembly of tissue constructs with clinically significant cell densities, uniform cell distribution, and endothelialized, perfusable channels.

  1. Interaction of collagen-like peptide models of asymmetric acetylcholinesterase with glycosaminoglycans: spectroscopic studies of conformational changes and stability.

    Doss-Pepe, E; Deprez, P; Inestrosa, N C; Brodsky, B

    2000-12-01

    The effect of heparin on the conformation and stability of triple-helical peptide models of the collagen tail of asymmetric acetylcholinesterase expands our understanding of heparin interactions with proteins and presents an opportunity for clarifying the nature of binding of ligands to collagen triple-helix domains. Within the collagen tail of AChE, there are two consensus sequences for heparin binding of the form BBXB, surrounded by additional basic residues. Circular dichroism studies were used to determine the effect of the addition of increasing concentrations of heparin on triple-helical peptide models for the heparin binding domains, including peptides in which the basic residues within and surrounding the consensus sequence were replaced by alanine residues. The addition of heparin caused an increased triple-helix content with saturation properties for the peptide modeling the C-terminal site, while precipitation, with no increased helix content resulted from heparin addition to the peptide modeling the N-terminal site. The results suggest that the two binding sites with a similar triple-helical conformation have distinctive ways of interacting with heparin, which must relate to small differences in the consensus sequence (GRKGR vs GKRGK) and in the surrounding basic residues. Addition of heparin increased the thermal stability of all peptides containing the consensus sequence. Heparan sulfate produced conformational and stabilization effects similar to those of heparin, while chondroitin sulfate led to a cloudy solution, loss of circular dichroism signal, and a smaller increase in thermal stability. Thus, specificity in both the sequence of the triple helix and the type of glycosaminoglycan is required for this interaction. PMID:11101304

  2. Regulation of RPTPbeta/phosphacan expression and glycosaminoglycan epitopes in injured brain and cytokine-treated glia.

    Dobbertin, Alexandre; Rhodes, Kate E; Garwood, Jeremy; Properzi, Francesca; Heck, Nicolas; Rogers, John H; Fawcett, James W; Faissner, Andreas

    2003-12-01

    Several chondroitin sulfate proteoglycans (CSPGs) are upregulated after CNS injury and are thought to limit axonal regeneration in the adult mammalian CNS. Therefore, we examined the expression of the CSPG, receptor protein tyrosine phosphatase beta (RPTPbeta)/phosphacan, after a knife lesion to the cerebral cortex and after treatment of glial cultures with regulatory factors. The three splice variants of this CSPG gene, the secreted isoform, phosphacan, and the two transmembrane isoforms, the long and short RPTPbeta, were examined. Western blot and immunostaining analysis of injured and uninjured tissue revealed a transient decrease of phosphacan protein levels, but not of short RPTPbeta, in the injured tissue from 1 to 7 days postlesion (dpl). By real time RT-PCR, we show that phosphacan and long RPTPbeta mRNA levels are transiently down-regulated at 2 dpl, unlike those of short RPTPbeta which increased after 4 dpl. In contrast to the core glycoprotein, the phosphacan chondroitin sulfate (CS) glycosaminoglycan epitope DSD-1 was up-regulated after 7 dpl. Phosphacan was expressed by cultivated astrocytes and oligodendrocyte precursors but was more glycanated in oligodendrocyte precursors, which produce more of DSD-1 epitope than astrocytes. Epidermal growth factor/transforming growth factor alpha strongly increased the astrocytic expression of long RPTPbeta and phosphacan and slightly the short RPTPbeta protein levels, while interferon gamma and tumor necrosis factor alpha reduced astrocytic levels of phosphacan, but not of the receptor forms. Examining the effects of phosphacan on axon growth from rat E17 cortical neurons, we found that phosphacan stimulates outgrowth in a largely CS dependent manner, while it blocks the outgrowth-promoting effects of laminin through an interaction that is not affected by removal of the CS chains. These results demonstrate complex injury-induced modifications in phosphacan expression and glycanation that may well influence axonal

  3. Research progress of glycosaminoglycans from marine animals%海洋动物糖胺聚糖的研究进展

    周小双; 王锦旭; 杨贤庆; 魏涯

    2015-01-01

    糖胺聚糖存在于大多数海洋动物的组织中,目前主要利用酶解法进行提取,已确定的糖胺聚糖一共有7种。因为糖胺聚糖具有高生物活性和低毒副作用的特点,所以近年来国内外学者对海洋动物中糖胺聚糖的抗肿瘤、降血糖、增强免疫等活性研究逐渐增多,研究发现海洋动物糖胺聚糖可以抑制 S180肿瘤细胞生长,促进 T、B淋巴细胞增殖,增强 NK细胞活性,清除 O2-·、·OH、DPPH·自由基,降低血清总胆固醇(total cholesterol, TC)、低密度脂蛋白(low density lipoprotein, LDL)和甘油三酯(triglyceride, TG)的含量,升高高密度脂蛋白(high density lipoprotein, HDL)水平,促进胆固醇代谢。这些特性都将使海洋动物糖胺聚糖成为医学及保健品行业重点研究对象,使得一些海洋动物得以高值化利用,促进海洋经济发展。本文主要对海洋动物中糖胺聚糖的研究现状、结构分类、提取方法、分离纯化、分析测定和生物活性进行了概述。%ABSTRACT:Glycosaminoglycans were found in the tissue of most marine animals, which were extracted by enzymatic method. Seven kinds of glycosaminoglycans have been identified. In recent years, due to their high biological activities and low toxicity, experts worldwide were attracted by the anti-tumor, lowering blood glucose, immuno-enhancement effects of glycosaminoglycans from marine animals. Researches indicated that glycosaminoglycans of marine animal can inhibit the growth of S180 tumor cells, promote proliferation of T and B lymphocytes, enhance the activity of NK cells, scavenge free radicalof O2-·,·OH and DPPH·, reduce content of serum total cholesterol, low density lipoprotein and triglyceride , increase the level of high density lipoprotein, promote the metabolism of cholesterol. These features of glycosaminoglycans have attracted attention from medicine and health products industry. Marine animals can be utilized for high value, and

  4. Assessment of lumbar intervertebral disc glycosaminoglycan content by gadolinium-enhanced MRI before and after 21-days of head-down-tilt bedrest.

    Timmo Koy

    Full Text Available During spaceflight, it has been shown that intervertebral discs (IVDs increase in height, causing elongation of the spine up to several centimeters. Astronauts frequently report dull lower back pain that is most likely of discogenic origin and may result from IVD expansion. It is unknown whether disc volume solely increases by water influx, or if the content of glycosaminoglycans also changes in microgravity. Aim of this pilot study was to investigate effects of the spaceflight analog of bedrest on the glycosaminoglycan content of human lumbar IVDs. Five healthy, non-smoking, male human subjects of European descent were immobilized in 6° head-down-tilt bedrest for 21 days. Subjects remained in bed 24 h a day with at least one shoulder on the mattress. Magnetic Resonance Imaging (MRI scans were taken according to the delayed gadolinium-enhanced magnetic resonance imaging (dGEMRIC protocol before and after bedrest. The outcome measures were T1 and ΔT1. Scans were performed before and after administration of the contrast agent Gd-DOTA, and differences between T1-values of both scans (ΔT1 were computed. ΔT1 is the longitudinal relaxation time in the tissue and inversely related to the glycosaminoglycan-content. For data analysis, IVDs L1/2 to L4/5 were semi-automatically segmented. Zones were defined and analyzed separately. Results show a highly significant decrease in ΔT1 (p<0.001 after bedrest in all IVDs, and in all areas of the IVDs. The ΔT1-decrease was most prominent in the nucleus pulposus and in L4/5, and was expressed slightly more in the posterior than anterior IVD. Unexpected negative ΔT1-values were found in Pfirrmann-grade 2-discs after bedrest. Significantly lower T1 before contrast agent application was found after bedrest compared to before bedrest. According to the dGEMRIC-literature, the decrease in ΔT1 may be interpreted as an increase in glycosaminoglycans in healthy IVDs during bedrest. This interpretation seems

  5. Avaliação dos glicosaminoglicanos do tecido periuretral de pacientes com e sem prolapso genital Evaluation of glycosaminoglycans of periurethral tissue in patients with and without pelvic organ prolapse

    Paulo Cezar Feldner Jr

    2008-04-01

    tissue during surgery and assessed by biochemical methods. The GAGs were obtained by proteolysis and precipitated by trichloroacetic acid. The relative concentration of sulfated GAGs was determined by densitometry of toluidine blue stained gel using a spectrophotometer with a 525 nm wavelength. Data were compared using analysis of variance (ANOVA. RESULTS: In the two groups dermatan sulphate (DS was the predominant glycosaminoglycan (85%, followed by chondroitin sulphate (CS and heparan sulphate (HS. Women with pelvic organ prolapse had significantly more total GAGs, DS and HS. Differences in CS were not observed. CONCLUSIONS: This study showed altered biochemical characteristics in the extracellular matrix of periurethral tissue and also accumulation of GAGs, DS and CS, in women with pelvic organ prolapse.

  6. Rotavirus NSP4 is secreted from infected cells as an oligomeric lipoprotein and binds to glycosaminoglycans on the surface of non-infected cells

    Didsbury Alicia

    2011-12-01

    Full Text Available Abstract Background Nonstructural glycoprotein 4 (NSP4 encoded by rotavirus is the only viral protein currently believed to function as an enterotoxin. NSP4 is synthesized as an intracellular transmembrane glycoprotein and as such is essential for virus assembly. Infection of polarized Caco-2 cells with rotavirus also results in the secretion of glycosylated NSP4 apparently in a soluble form despite retention of its transmembrane domain. We have examined the structure, solubility and cell-binding properties of this secreted form of NSP4 to further understand the biochemical basis for its enterotoxic function. We show here that NSP4 is secreted as discrete detergent-sensitive oligomers in a complex with phospholipids and demonstrate that this secreted form of NSP4 can bind to glycosaminoglycans present on the surface of a range of different cell types. Methods NSP4 was purified from the medium of infected cells after ultracentrifugation and ultrafiltration by successive lectin-affinity and ion exchange chromatography. Oligomerisation of NSP4 was examined by density gradient centrifugation and chemical crosslinking and the lipid content was assessed by analytical thin layer chromatography and flame ionization detection. Binding of NSP4 to various cell lines was measured using a flow cytometric-based assay. Results Secreted NSP4 formed oligomers that contained phospholipid but dissociated to a dimeric species in the presence of non-ionic detergent. The purified glycoprotein binds to the surface of various non-infected cells of distinct lineage. Binding of NSP4 to HT-29, a cell line of intestinal origin, is saturable and independent of divalent cations. Complementary biochemical approaches reveal that NSP4 binds to sulfated glycosaminoglycans on the plasma membrane. Conclusion Our study is the first to analyze an authentic (i.e. non-recombinant form of NSP4 that is secreted from virus-infected cells. Despite retention of the transmembrane domain

  7. Assessment of quantitative and qualitative changes of proteoglycans and glycosaminoglycans in normal breast tissue during the follicular and luteal phases of the menstrual cycle.

    Júnior, J A Dos Santos; de Lima, C R; Michelacci, Y M C da Silva; Nazário, A C Pinto

    2015-01-01

    The effect of sex hormones on extracellular matrix compounds, such as proteoglycans (PGs) and glycosaminoglycans (GAGs), in mammary tissue remains poorly understood. The elucidation of extracellular matrix component functions could clarify pathophysiological conditions, such as cyclical mastalgia (breast pain). The authors examined the quantitative and qualitative changes of PGs and GAGs in normal breast tissue during the follicular and luteal phases of the menstrual cycle. Twenty-eight eumenorrheic patients with benign breast nodules were divided into groups: Group A included 15 follicular patients and Group B included 13 luteal phase patients. Breast tissue adjacent to the nodules was biochemically analyzed to evaluate the types and concentrations of PGS and GAGs. The distribution of proteoglycans during the menstrual cycle was analyzed with immunofluorescence. PG concentrations were elevated (p dermatan and heparan sulfate, varied cyclically, the differences were not significant. PMID:26524806

  8. Characterization of osteoprotegerin binding to glycosaminoglycans by surface plasmon resonance: Role in the interactions with receptor activator of nuclear factor κB ligand (RANKL) and RANK

    Osteoprotegerin (OPG) is a decoy receptor for receptor activator of nuclear factor κB ligand (RANKL), a key inducer of osteoclastogenesis via its receptor RANK. We previously showed that RANK, RANKL, and OPG are able to form a tertiary complex and that OPG must be also considered as a direct effector of osteoclast functions. As OPG contains a heparin-binding domain, the present study investigated the interactions between OPG and glycosaminoglycans (GAGs) by surface plasmon resonance and their involvement in the OPG functions. Kinetic data demonstrated that OPG binds to heparin with a high-affinity (K D: 0.28 nM) and that the pre-incubation of OPG with heparin inhibits in a dose-dependent manner the OPG binding to the complex RANK-RANKL. GAGs from different structure/origin (heparan sulfate, dermatan sulfate, and chondroitin sulfate) exert similar activity on OPG binding. The contribution of the sulfation pattern and the size of the oligosaccharide were determined in this inhibitory mechanism. The results demonstrated that sulfation is essential in the OPG-blocking function of GAGs since a totally desulfated heparin loses its capacity to bind and to block OPG binding to RANKL. Moreover, a decasaccharide is the minimal structure that totally inhibits the OPG binding to the complex RANK-RANKL. Western blot analysis performed in 293 cells surexpressing RANKL revealed that the pre-incubation of OPG with these GAGs strongly inhibits the OPG-induced decrease of membrane RANKL half-life. These data support an essential function of the related glycosaminoglycans heparin and heparan sulfate in the activity of the triad RANK-RANKL-OPG

  9. Isolation, characterization and properties of the oversulphated chondroitin sulphate proteoglycan from squid skin with peculiar glycosaminoglycan sulphation pattern.

    Karamanos, N K; Aletras, A J; Tsegenidis, T; Tsiganos, C P; Antonopoulos, C A

    1992-03-01

    Oversulphated chondroitin sulphate proteoglycan from squid skin was isolated from 4 M guanidine hydrochloride extract by ion-exchange chromatography, gel chromatography and density gradient centrifugation. The proteoglycan had Mr 3.5 x 10(5), contained on average six oversulphated chondroitin sulphate chains (Mr 4 x 10(4)) bound on a polypeptide of Mr 2.8 x 10(4), and oligosaccharides consisting of both hexosamines, glucuronic acid, sulphates and fucose as the only neutral monosaccharide. The major amino acids of the proteoglycan protein core are glycine (corresponding to about one third of the total amino acids), aspartic acid/asparagine and serine, together amounting to 50% of the total. The proteoglycan was resistant to the proteolytic enzymes V8 protease, trypsin (treated with diphenylcarbamoyl chloride), alpha-chymotrypsin and pronase, while it was completely degraded by papain and to a large extent by collagenase. Pretreated proteoglycan with chondroitinase AC was degraded by pronase to a large extent and slightly by V8 protease and trypsin. The proteoglycan did not interact with hyaluronic acid and did not form self-aggregates. Oversulphated chondroitin sulphate chains were composed of unusual sulphated disaccharide units which were isolated and characterized by HPLC. In particular, it contained 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid)-D-galactose 4-sulphate (delta di-4S) and disulphated disaccharides (delta di-diS) [90% 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid 2/3-sulphate)-D-galactose 6-sulphate (delta di-diSD) and 10% 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid 2/3-sulphate)-D-galactose 4-sulphate (delta di-diSK)] as the major disaccharides, significant amounts of trisulphated disaccharides (delta di-triS) and small amounts of 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4-enopyranosyluronic acid)-D-galactose 6-sulphate (delta di-6S) and 2-acetamido-2-deoxy-3-O-(alpha-L-threo-4

  10. Molecular cloning of a novel glucuronokinase/putative pyrophosphorylase from zebrafish acting in an UDP-glucuronic acid salvage pathway.

    Roman Gangl

    Full Text Available In animals, the main precursor for glycosaminoglycan and furthermore proteoglycan biosynthesis, like hyaluronic acid, is UDP-glucuronic acid, which is synthesized via the nucleotide sugar oxidation pathway. Mutations in this pathway cause severe developmental defects (deficiency in the initiation of heart valve formation. In plants, UDP-glucuronic acid is synthesized via two independent pathways. Beside the nucleotide sugar oxidation pathway, a second minor route to UDP-glucuronic acid exist termed the myo-inositol oxygenation pathway. Within this myo-inositol is ring cleaved into glucuronic acid, which is subsequently converted to UDP-glucuronic acid by glucuronokinase and UDP-sugar pyrophosphorylase. Here we report on a similar, but bifunctional enzyme from zebrafish (Danio rerio which has glucuronokinase/putative pyrophosphorylase activity. The enzyme can convert glucuronic acid into UDP-glucuronic acid, required for completion of the alternative pathway to UDP-glucuronic acid via myo-inositol and thus establishes a so far unknown second route to UDP-glucuronic acid in animals. Glucuronokinase from zebrafish is a member of the GHMP-kinase superfamily having unique substrate specificity for glucuronic acid with a Km of 31 ± 8 µM and accepting ATP as the only phosphate donor (Km: 59 ± 9 µM. UDP-glucuronic acid pyrophosphorylase from zebrafish has homology to bacterial nucleotidyltransferases and requires UTP as nucleosid diphosphate donor. Genes for bifunctional glucuronokinase and putative UDP-glucuronic acid pyrophosphorylase are conserved among some groups of lower animals, including fishes, frogs, tunicates, and polychaeta, but are absent from mammals. The existence of a second pathway for UDP-glucuronic acid biosynthesis in zebrafish likely explains some previous contradictory finding in jekyll/ugdh zebrafish developmental mutants, which showed residual glycosaminoglycans and proteoglycans in knockout mutants of UDP