Sample records for cross-linked hyaluronan gel

  1. Novel enzymatically cross-linked hyaluronan hydrogels support the formation of 3D neuronal networks.

    PubMed

    Broguiere, Nicolas; Isenmann, Luca; Zenobi-Wong, Marcy

    2016-08-01

    Hyaluronan (HA) is an essential component of the central nervous system's extracellular matrix and its high molecular weight (MW) form has anti-inflammatory and anti-fibrotic properties relevant for regenerative medicine. Here, we introduce a new hydrogel based on high MW HA which is cross-linked using the transglutaminase (TG) activity of the activated blood coagulation factor XIII (FXIIIa). These HA-TG gels have significant advantages for neural tissue engineering compared to previous HA gels. Due to their chemical inertness in the absence of FXIIIa, the material can be stored long-term, is stable in solution, and shows no cytotoxicity. The gelation is completely cell-friendly due to the specificity of the enzyme and the gelation rate can be tuned from seconds to hours at physiological pH and independently of stiffness. The gels are injectable, and attach covalently to fibrinogen and fibrin, two common bioactive components in in vitro tissue engineering, as well as proteins present in vivo, allowing the gels to covalently bind to brain or spinal cord defects. These optimal chemical and bioactive properties of HA-TG gels enabled the formation of 3D neuronal cultures of unprecedented performance, showing fast neurite outgrowth, axonal and dendritic speciation, strong synaptic connectivity in 3D networks, and rapidly-occurring and long-lasting coordinated electrical activity. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Evaluation of hyaluronan gel (Gengigel(®) ) as a topical applicant in the treatment of gingivitis.

    PubMed

    Sapna, Nadiger; Vandana, Kharidi Laxman

    2011-08-01

      To clinically and histopathologically evaluate the anti-inflammatory effect of 0.2% hyaluronan gel alone and with mechanical therapy on gingivitis. The argyrophilic nucleolar organizer region staining technique was attempted to routinely determine its diagnostic and prognostic dependability for periodontal lesions.   In each of the 28 gingivitis patients, the four quadrants were subjected to different treatments: scaling, scaling + topical hyaluronan gel, only topical hyaluronan gel, and topical + intrasulcular hyaluronan gel. Clinical parameters were recorded at baseline, and on days 7, 14, and 21. Biopsies were taken from each quadrant, inflammatory infiltrates were graded, and the argyrophilic nucleolar organizer region count was measured before and after treatment.   A significant reduction was seen in clinical parameters, inflammatory infiltrates, and the argyrophilic nucleolar organizer region count within the groups. The effect of topical + intrasulcular gel was equivalent to scaling (P > 0.05). Topical + intrasulcular hyaluronan gel application demonstrated a better reduction than topical hyaluronan gel alone.   Hyaluronan gel is an effective topical agent for treating gingivitis, along with scaling and intrasulcular application. The argyrophilic nucleolar organizer region count can be used as a histopathological indicator in cases of non-responsive gingivitis to assess the severity of gingival inflammation. © 2011 Blackwell Publishing Asia Pty Ltd.

  3. Molecular weight dependency of polyrotaxane-cross-linked polymer gel extensibility.

    PubMed

    Ohmori, Kana; Abu Bin, Imran; Seki, Takahiro; Liu, Chang; Mayumi, Koichi; Ito, Kohzo; Takeoka, Yukikazu

    2016-12-11

    This work investigates the influence of the molecular weight of polyrotaxane (PR) cross-linkers on the extensibility of polymer gels. The polymer gels, which were prepared using PR cross-linkers of three different molecular weights but the same number of cross-linking points per unit volume of gel, have almost the same Young's modulus. By contrast, the extensibility and rupture strength of the polymer gels are substantially increased with increasing molecular weight of the PR cross-linker.

  4. Exploring the kinetics of gelation and final architecture of enzymatically cross-linked chitosan/gelatin gels.

    PubMed

    da Silva, Marcelo A; Bode, Franziska; Grillo, Isabelle; Dreiss, Cécile A

    2015-04-13

    Small-angle neutron scattering (SANS) was used to characterize the nanoscale structure of enzymatically cross-linked chitosan/gelatin hydrogels obtained from two protocols: a pure chemical cross-linking process (C), which uses the natural enzyme microbial transglutaminase, and a physical-co-chemical (PC) hybrid process, where covalent cross-linking is combined with the temperature-triggered gelation of gelatin, occurring through the formation of triple-helices. SANS measurements on the final and evolving networks provide a correlation length (ξ), which reflects the average size of expanding clusters. Their growth in PC gels is restricted by the triple-helices (ξ ∼ 10s of Å), while ξ in pure chemical gels increases with cross-linker concentration (∼100s of Å). In addition, the shear elastic modulus in PC gels is higher than in pure C gels. Our results thus demonstrate that gelatin triple helices provide a template to guide the cross-linking process; overall, this work provides important structural insight to improve the design of biopolymer-based gels.

  5. Robust cross-links in molluscan adhesive gels: Testing for contributions from hydrophobic and electrostatic interactions

    PubMed Central

    Smith, A.M.; Robinson, T. M.; Salt, M. D.; Hamilton, K. S.; Silvia, B. E.; Blasiak, R.

    2009-01-01

    The cross-linking interactions that provide cohesive strength to molluscan adhesive gels were investigated. Metal-based interactions have been shown to play an important role in the glue of the slug Arion subfuscus (Draparnaud), but other types of interactions may also contribute to the glue's strength and their role has not been investigated. This study shows that treatments that normally disrupt hydrophobic or electrostatic interactions have little to no effect on the slug glue. High salt concentrations and non-ionic detergent do not affect the solubility of the proteins in the glue or the ability of the glue proteins to stiffen gels. In contrast, metal chelation markedly disrupts the gel. Experiments with gel filtration chromatography identify a 40 kDa protein that is a central component of the cross-links in the glue. This 40 kDa protein forms robust macromolecular aggregations that are stable even in the presence of high concentrations of salt, non-ionic detergent, urea or metal chelators. Metal chelation during glue secretion, however, may block some of these cross-links. Such robust, non-specific interactions in an aqueous environment are highly unusual for hydrogels and reflect an intriguing cross-linking mechanism. PMID:18952190

  6. Robust cross-links in molluscan adhesive gels: testing for contributions from hydrophobic and electrostatic interactions.

    PubMed

    Smith, A M; Robinson, T M; Salt, M D; Hamilton, K S; Silvia, B E; Blasiak, R

    2009-02-01

    The cross-linking interactions that provide cohesive strength to molluscan adhesive gels were investigated. Metal-based interactions have been shown to play an important role in the glue of the slug Arion subfuscus (Draparnaud), but other types of interactions may also contribute to the glue's strength and their role has not been investigated. This study shows that treatments that normally disrupt hydrophobic or electrostatic interactions have little to no effect on the slug glue. High salt concentrations and non-ionic detergent do not affect the solubility of the proteins in the glue or the ability of the glue proteins to stiffen gels. In contrast, metal chelation markedly disrupts the gel. Experiments with gel filtration chromatography identify a 40 kDa protein that is a central component of the cross-links in the glue. This 40 kDa protein forms robust macromolecular aggregations that are stable even in the presence of high concentrations of salt, non-ionic detergent, urea or metal chelators. Metal chelation during glue secretion, however, may block some of these cross-links. Such robust, non-specific interactions in an aqueous environment are highly unusual for hydrogels and reflect an intriguing cross-linking mechanism.

  7. Comparative study on protein cross-linking and gel enhancing effect of microbial transglutaminase on surimi from different fish.

    PubMed

    Chanarat, Sochaya; Benjakul, Soottawat; H-Kittikun, Aran

    2012-03-15

    Microbial transglutaminase (MTGase) has been used to increase the gel strength of surimi. Nevertheless, its effectiveness varies with fish species. The aim of this study was to elucidate the effect of MTGase at different levels on protein cross-linking and gel property of surimi from threadfin bream, Indian mackerel and sardine in the presence and absence of endogenous transglutaminase. Breaking force of all surimi gels increased as MTGase levels (0-0.6 U g⁻¹) increased except for threadfin bream surimi gel, where the breaking force decreased at 0.6 U g⁻¹ (P < 0.05). In the presence of EDTA, the gel strengthening effect was lower, suggesting the combined effect of endogenous transglutaminase with MTGase. With the addition of MTGase, the gel with the highest increase in breaking force showed highest decrease in myosin heavy chain. When cross-linking activity of MTGase on natural actomyosin (NAM) was determined, the highest decreasing rate in ε-amino group content with the concomitant increased formation of cross-linked proteins was found in NAM from threadfin bream. The reactivity of muscle proteins toward MTGase-induced cross-linking was in agreement with surimi gel strengthening. The composition and properties of muscle proteins of varying fish species more likely determined protein cross-linking induced by MTGase, thereby affecting their gel properties.

  8. Collagen telopeptides (cross-linking sites) play a role in collagen gel lattice contraction

    NASA Technical Reports Server (NTRS)

    Woodley, D. T.; Yamauchi, M.; Wynn, K. C.; Mechanic, G.; Briggaman, R. A.

    1991-01-01

    Solubilized interstitial collagens will form a fibrillar, gel-like lattice when brought to physiologic conditions. In the presence of human dermal fibroblasts the collagen lattice will contract. The rate of contraction can be determined by computer-assisted planemetry. The mechanisms involved in contraction are as yet unknown. Using this system it was found that the rate of contraction was markedly decreased when collagen lacking telopeptides was substituted for native collagen. Histidinohydroxylysinonorleucine (HHL) is a major stable trifunctional collagen cross-link in mature skin that involves a carboxyl terminal, telopeptide site 16c, the sixteenth amino acid residue from the carboxy terminal of the telopeptide region of alpha 1 (I) in type I collagen. Little, if any, HHL was present in native, purified, reconstituted, soluble collagen fibrils from 1% acetic acid-extracted 2-year-old bovine skin. In contrast, HHL cross-links were present (0.22 moles of cross-link per mole of collagen) in lattices of the same collagen contracted by fibroblasts. However, rat tail tendon does not contain HHL cross-links, and collagen lattices made of rat tail tendon collagen are capable of contraction. This suggests that telopeptide sites, and not mature HHL cross-links per se, are essential for fibroblasts to contract collagen lattices. Beta-aminopropionitrile fumarate (BAPN), a potent lathyrogen that perturbs collagen cross-linking by inhibition of lysyl oxidase, also inhibited the rate of lattice cell contraction in lattices composed of native collagen. However, the concentrations of BAPN that were necessary to inhibit the contraction of collagen lattices also inhibited fibroblast growth suggestive of cellular toxicity. In accordance with other studies, we found no inhibition of the rate of lattice contraction when fibronectin-depleted serum was used. Electron microscopy of contracted gels revealed typical collagen fibers with a characteristic axial periodicity. The data

  9. Formation and properties of hyaluronan/nano Ag and hyaluronan-lecithin/nano Ag films.

    PubMed

    Khachatryan, Gohar; Khachatryan, Karen; Grzyb, Jacek; Fiedorowicz, Maciej

    2016-10-20

    A facile and environmentally friendly method of the preparation of silver nanoparticles embedded in hyaluronan (Hyal/Ag) and hyaluronan-lecithin (Hyal-L/Ag) matrix was developed. Thin, elastic foils were prepared from gels by an in situ synthesis of Ag in an aqueous solution of sodium hyaluronate (Hyal), using aq. d-(+)-xylose solution as a reducing agent. The gels were applied to a clean, smooth, defatted Teflon surface and left for drying in the air. The dry foils were stored in a closed container. UV-vis spectroscopy, transmission electron microscopy (TEM) and Fourier transform infrared (FTIR) spectra confirmed formation of about 10nm ball-shaped Ag nanoparticles situated within the polysaccharide template. Thermal properties of the composites were characterized involving differential scanning calorimetry (DSC) and thermogravimetric (TGA) analyses, whereas molecular weights of polysaccharide chains of the matrix were estimated with the size exclusion chromatography coupled with multiangle laser light scattering and refractometric detectors (HPSEC-MALLS-RI). An increase in the molecular weight of the hyaluronate after generation of Ag nanoparticles was observed. The foils showed specific properties. The study confirmed that silver nanoparticles can be successfully prepared with environmentally friendly method, using hyaluronan as a stabilizing template. Hyaluronan and hyaluronan-lecithin matrices provide nanocrystals uniform in size and shape. The composites demonstrated a bacteriostatic activity. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Ionic liquids in a poly ethylene oxide cross-linked gel polymer as an electrolyte for electrical double layer capacitor

    NASA Astrophysics Data System (ADS)

    Chaudoy, V.; Tran Van, F.; Deschamps, M.; Ghamouss, F.

    2017-02-01

    In the present work, we developed a gel polymer electrolyte via the incorporation of a room temperature ionic liquid into a cross-linked polymer matrix. The cross-linked gel electrolyte was prepared using a free radical polymerization of methacrylate and dimethacrylate oligomers dissolved in 1-propyl-1-methylpyrrolidinium bis(fluorosulfonyl)imide. Combining the advantages of the ionic liquids and of conventional polymers, the cross-linked gel polymer electrolyte was used both as a separator and as an electrolyte for a leakage-free and non-flammable EDLC supercapacitor. The quasi-all solid-state supercapacitors showed rather good capacitance, power and energy densities by comparison to a liquid electrolyte-based EDLC.

  11. Iodopropyl-branched polysiloxane gel electrolytes with improved ionic conductivity upon cross-linking.

    PubMed

    De Gregorio, G L; Giannuzzi, R; Cipolla, M P; Agosta, R; Grisorio, R; Capodilupo, A; Suranna, G P; Gigli, G; Manca, M

    2014-11-21

    We here report the implementation of poly[(3-N-methylimidazoliumpropyl)methylsiloxane-co-dimethylsiloxane]iodides as suitable polymeric hosts for a novel class of in situ cross-linkable iodine/iodide-based gel-electrolytes for dye-sensitized solar cells. The polymers are first partially quaternized and then subjected to a thermal cross-linking which allows the formation of a 3D polymeric network which is accompanied by a dramatic enhancement of the ionic conductivity.

  12. Temperature-induced sol-gel transition and microgel formation in α-actinin cross-linked actin networks: A rheological study

    NASA Astrophysics Data System (ADS)

    Tempel, M.; Isenberg, G.; Sackmann, E.

    1996-08-01

    We have studied the sol-gel transition, the viscoelastic and the structural properties of networks constituted of semiflexible actin filaments cross-linked by α-actinin. Cross-linking was regulated in a reversible way by varying the temperature through the association-dissociation equilibrium of the actin-α-actinin system. Viscoelastic parameters [shear storage modulus G'(ω), phase shift tan(Φ)(ω), creep compliance J(t)] were measured as a function of temperature and actin-to-cross-linker ratio by a magnetically driven rotating disc rheometer. G'(ω) and tan(Φ)(ω) were studied at a frequency ω corresponding to the elastic plateau regime of the G'(ω) versus ω spectrum of the purely entangled solution. The microstructure of the networks was viewed by negative staining electron microscopy (EM). The phase shift tan(Φ) (or equivalently the viscosity η) diverges and reaches a maximum when approaching the apparent gel point from lower and higher temperatures, and the maximum defines the gel point (temperature Tg). The elastic plateau modulus G'N diverges at temperatures beyond this gel point TTg. The cross-linking transition (corresponding to a sol-gel transition at zero frequency) is interpreted in terms of a percolation model and the divergence of G'N at Tgel is essentially homogeneous above the cross-linking transition (T>Tg), (2) that microscopic segregation takes place at T<=Tg leading to local formation of clusters (a state termed microgel), and (3) that at low actin-α-actinin ratios (rAα<=10) and low temperatures (T<=10 °C) macroscopic segregation into bundles of cross-linked actin filaments and a diluted solution of actin filaments is observed. The three regimes of network structure are represented by an

  13. Cross-linked Composite Gel Polymer Electrolyte using Mesoporous Methacrylate-Functionalized SiO2 Nanoparticles for Lithium-Ion Polymer Batteries

    PubMed Central

    Shin, Won-Kyung; Cho, Jinhyun; Kannan, Aravindaraj G.; Lee, Yoon-Sung; Kim, Dong-Won

    2016-01-01

    Liquid electrolytes composed of lithium salt in a mixture of organic solvents have been widely used for lithium-ion batteries. However, the high flammability of the organic solvents can lead to thermal runaway and explosions if the system is accidentally subjected to a short circuit or experiences local overheating. In this work, a cross-linked composite gel polymer electrolyte was prepared and applied to lithium-ion polymer cells as a safer and more reliable electrolyte. Mesoporous SiO2 nanoparticles containing reactive methacrylate groups as cross-linking sites were synthesized and dispersed into the fibrous polyacrylonitrile membrane. They directly reacted with gel electrolyte precursors containing tri(ethylene glycol) diacrylate, resulting in the formation of a cross-linked composite gel polymer electrolyte with high ionic conductivity and favorable interfacial characteristics. The mesoporous SiO2 particles also served as HF scavengers to reduce the HF content in the electrolyte at high temperature. As a result, the cycling performance of the lithium-ion polymer cells with cross-linked composite gel polymer electrolytes employing methacrylate-functionalized mesoporous SiO2 nanoparticles was remarkably improved at elevated temperatures. PMID:27189842

  14. Incorporation of Pentraxin 3 into Hyaluronan Matrices Is Tightly Regulated and Promotes Matrix Cross-linking

    PubMed Central

    Baranova, Natalia S.; Inforzato, Antonio; Briggs, David C.; Tilakaratna, Viranga; Enghild, Jan J.; Thakar, Dhruv; Milner, Caroline M.; Day, Anthony J.; Richter, Ralf P.

    2014-01-01

    Mammalian oocytes are surrounded by a highly hydrated hyaluronan (HA)-rich extracellular matrix with embedded cumulus cells, forming the cumulus cell·oocyte complex (COC) matrix. The correct assembly, stability, and mechanical properties of this matrix, which are crucial for successful ovulation, transport of the COC to the oviduct, and its fertilization, depend on the interaction between HA and specific HA-organizing proteins. Although the proteins inter-α-inhibitor (IαI), pentraxin 3 (PTX3), and TNF-stimulated gene-6 (TSG-6) have been identified as being critical for COC matrix formation, its supramolecular organization and the molecular mechanism of COC matrix stabilization remain unknown. Here we used films of end-grafted HA as a model system to investigate the molecular interactions involved in the formation and stabilization of HA matrices containing TSG-6, IαI, and PTX3. We found that PTX3 binds neither to HA alone nor to HA films containing TSG-6. This long pentraxin also failed to bind to products of the interaction between IαI, TSG-6, and HA, among which are the covalent heavy chain (HC)·HA and HC·TSG-6 complexes, despite the fact that both IαI and TSG-6 are ligands of PTX3. Interestingly, prior encounter with IαI was required for effective incorporation of PTX3 into TSG-6-loaded HA films. Moreover, we demonstrated that this ternary protein mixture made of IαI, PTX3, and TSG-6 is sufficient to promote formation of a stable (i.e. cross-linked) yet highly hydrated HA matrix. We propose that this mechanism is essential for correct assembly of the COC matrix and may also have general implications in other inflammatory processes that are associated with HA cross-linking. PMID:25190808

  15. An agarose gel electrophoretic method for analysis of hyaluronan molecular weight distribution.

    PubMed

    Lee, H G; Cowman, M K

    1994-06-01

    An electrophoretic method is described for determining the molecular weight distribution of hyaluronan (HA). The method involves separation of HA by electrophoresis on a 0.5% agarose gel, followed by detection of HA using the cationic dye Stains-All (3,3'-dimethyl-9-methyl-4,5,4'5'-dibenzothiacarbocyanine). The recommended sample load is 7 micrograms. Calibration of the method with HA standards of known molecular weight has established a linear relationship between electrophoretic mobility and the logarithm of the weight-average molecular weight over the range of approximately 0.2-6 x 10(6). The separated HA pattern may also be visualized after electrotransfer of HA from the agarose gel to a nylon membrane. The membrane may be stained with the dye alcian blue. Alternatively, specific detection of HA from impure samples can be achieved by probing the nylon membrane with biotin-labeled HA-binding protein and subsequent interaction with a streptavidin-linked gold reagent and silver staining for amplification. The electrophoretic method was used to analyze HA in two different liquid connective tissues. Normal human knee joint synovial fluid showed a narrow HA molecular weight distribution, with a peak at 6-7 x 10(6). Owl monkey vitreous HA also showed a narrow molecular weight distribution, with a peak at 5-6 x 10(6). These results agree well with available published data and indicate the applicability of the method to the analysis of impure HA samples which may be available in limited amounts.

  16. In situ cross-linkable high molecular weight hyaluronan-bisphosphonate conjugate for localized delivery and cell-specific targeting: a hydrogel linked prodrug approach.

    PubMed

    Varghese, Oommen P; Sun, Weilun; Hilborn, Jöns; Ossipov, Dmitri A

    2009-07-01

    We present here a novel synthesis route to functionalize high molecular weight hyaluronan (HMW-HA) with a hydrazide group and a bioactive ligand, namely bisphosphonate (BP). For this purpose, a new symmetrical self-immolative biscarbazate linker has been devised. The hydrazide group was used to form hydrazone cross-linked hydrogel upon treating with previously described aldehyde modified hyaluronan. The 1:1 weight ratio of these two polymers gave hydrogel in less than 30 s. In this communication we present the first in vitro results showing that even though HA can target CD44 positive cancer cells (HCT-116), receptor mediated endocytosis could only occur by cleavage of high molecular weight HA with an ubiquitous enzyme, hyaluronidase (Hase). The cancer cells are known to overexpress CD44 receptors and also increase the hyaluronidase activity in vivo. Thus the pro-drug design, based on drug conjugation to HMW-HA, represents a new drug delivery platform where the drug potency is triggered by Hase mediated degradation of the HA-drug conjugate. We have successfully demonstrated that the cross-linkable HA-BP conjugate first undergoes Hase-mediated scission to the fragments of suitable sizes so as to be internalized by CD44 positive cells. The specificity of this targeting was proven by comparing the results with less CD44 positive HEK-293T cells. The localized delivery of such drugs at the surgical resection site opens up avenues to control tumor recurrence after removal of the tumor. In the form of hydrogel it would prevent systemic exposure of the drug and would allow its controlled release.

  17. Efficacy and safety of newly developed cross-linked dextran gel injection for glans penis augmentation with a novel technique

    PubMed Central

    Yang, Dae Yul; Ko, Kyungtae; Lee, Seong Ho; Moon, Du Geon; Kim, Jong Wook; Lee, Won Ki

    2018-01-01

    There is no safe and effective standard method for glans penis augmentation. Furthermore, there has been scant research on glans penis augmentation due to a poor understanding of glans anatomy, technical difficulty, and a lack of suitable substances for augmentation. Cross-linked dextran gel is a newly developed filler for soft-tissue augmentation. We evaluated the efficacy and safety of using a novel technique to inject cross-linked dextran gel for glans penis augmentation during a 24-week follow-up study. This prospective, single-arm, multicenter study enrolled twenty healthy adult men who underwent glans penis augmentation between June and August 2013. Cross-linked dextran gel was injected into the glans penis using a simple and easy technique. The sizes of the glans penis and individual satisfaction were assessed. Any adverse event was also reported. A total of 18 individuals were analyzed; two of them were lost to follow-up. The mean procedure time and injected volume were about 30 min and 6.6 ± 0.9 ml, respectively. The mean surface areas of the glans at baseline and 24 weeks were 20.0 ± 3.5 cm2 and 33.6 ± 5.4 cm2, respectively, representing a mean increase of 68.7% ± 14.0% (P < 0.001). Sixteen individuals (88.9%) were satisfied with the outcomes, and none were dissatisfied. There were no serious adverse events during the study. Cross-linked dextran gel injection for glans penis augmentation was easy and showed a significant augmentative effect on the glans penis, good durability, and was well tolerated without serious adverse events. Therefore, cross-linked dextran gel injection may be an effective, new technique for glans penis augmentation. PMID:28303807

  18. Rheological Enhancement of Pork Myofibrillar Protein-Lipid Emulsion Composite Gels via Glucose Oxidase Oxidation/Transglutaminase Cross-Linking Pathway.

    PubMed

    Wang, Xu; Xiong, Youling L; Sato, Hiroaki

    2017-09-27

    Porcine myofibrillar protein (MP) was modified with glucose oxidase (GluOx)-iron that produces hydroxyl radicals then subjected to microbial transglutaminase (TGase) cross-linking in 0.6 M NaCl at 4 °C. The resulting aggregation and gel formation of MP were examined. The GluOx-mediated oxidation promoted the formation of both soluble and insoluble protein aggregates via disulfide bonds and occlusions of hydrophobic groups. The subsequent TGase treatment converted protein aggregates into highly cross-linked polymers. MP-lipid emulsion composite gels formed with such polymers exhibited markedly enhanced gelling capacity: up to 4.4-fold increases in gel firmness and 3.5-fold increases in gel elasticity over nontreated protein. Microstructural examination showed small oil droplets dispersed in a densely packed gel matrix when MP was oxidatively modified, and the TGase treatment further contributed to such packing. The enzymatic GluOx oxidation/TGase treatment shows promise to improve the textural properties of emulsified meat products.

  19. A covalently cross-linked gel derived from the epidermis of the pilot whale Globicephala melas.

    PubMed

    Baum, C; Fleischer, L-G; Roessner, D; Meyer, W; Siebers, D

    2002-01-01

    The rheological properties of the stratum corneum of the pilot whale (Globicephala melas) were investigated with emphasis on their significance to the self-cleaning abilities of the skin surface smoothed by a jelly material enriched with various hydrolytic enzymes. The gel formation of the collected fluid was monitored by applying periodic-harmonic oscillating loads using a stress-controlled rheometer. In the mechanical spectrum of the gel, the plateau region of the storage modulus G' (<1200 Pa) and the loss modulus G" (>120 Pa) were independent of frequency (omega = 43.98 to 0.13 rad x s(-1), tau = 15 Pa, T = 20 degrees C), indicating high elastic performance of a covalently cross-linked viscoelastic solid. In addition, multi-angle laser light scattering experiments (MALLS) were performed to analyse the potential time-dependent changes in the weight-average molar mass of the samples. The observed increase showed that the gel formation is based on the assembly of covalently cross-linked aggregates. The viscoelastic properties and the shear resistance of the gel assure that the enzyme-containing jelly material smoothing the skin surface is not removed from the stratum corneum by shear regimes during dolphin jumping. The even skin surface is considered to be most important for the self-cleaning abilities of the dolphin skin against biofouling.

  20. Hyaluronan: from biomimetic to industrial business strategy.

    PubMed

    Murano, Erminio; Perin, Danilo; Khan, Riaz; Bergamin, Massimo

    2011-04-01

    Hyaluronan (hyaluronic acid) is a naturally occurring polysaccharide of a linear repeating disaccharide unit consisting of beta-(1-->4)-linked D-glucopyranuronic acid and beta-(1-->3)-linked 2-acetamido-2-deoxy-D-glucopyranose, which is present in extracellular matrices, the synovial fluid of joints, and scaffolding that comprises cartilage. In its mechanism of synthesis, its size, and its physico-chemical properties, hyaluronan is unique amongst other glycosaminoglycans. The network-forming, viscoelastic and its charge characteristics are important to many biochemical properties of living tissues. It is an important pericellular and cell surface constituent; its interaction with other macromolecules such as proteins, participates in regulating cell behavior during numerous morphogenic, restorative, and pathological processes in the body. The knowledge of HA in diseases such as various forms of cancers, arthritis and osteoporosis has led to new impetus in research and development in the preparation of biomaterials for surgical implants and drug conjugates for targeted delivery. A concise and focused review on hyaluronan is timely. This review will cover the following important aspects of hyaluronan: (i) biological functions and synthesis in nature; (ii) current industrial production and potential biosynthetic processes of hyaluronan; (iii) chemical modifications of hyaluronan leading to products of commercial significance; and (iv) and the global market position and manufacturers of hyaluronan.

  1. Preparation of a collagen/polymer hybrid gel designed for tissue membranes. Part I: controlling the polymer-collagen cross-linking process using an ethanol/water co-solvent.

    PubMed

    Nam, Kwangwoo; Kimura, Tsuyoshi; Funamoto, Seiichi; Kishida, Akio

    2010-02-01

    The drawback with collagen/2-methacryloyloxyethyl phosphorylcholine (MPC) polymer hybrid gels (collagen/phospholipid polymer hybrid gels) prepared in alkaline morpholinoethane sulfonic acid (MES) aqueous solution is that the cross-linking rate between the polymer and the collagen is low. To solve this problem, ethanol has been adopted as the reaction solvent, to prevent 1-ethyl-3-(3-dimethylaminopropyl)-1-carbodiimide hydrochloride (EDC) hydrolysis. Alterations in the ethanol mole concentration changed the cross-linking rate between the MPC polymer and the collagen gel. Prevention of EDC hydrolysis is clearly observed; protonation of carboxyl groups implies that the ratio of ethanol to water should be controlled. The polymer shows signs of penetration into the collagen gel layer, thus forming a totally homogeneous phase gel. This affects the mechanical strength of the collagen gel, making the gel much stiffer and brittle with an increase in the swelling ratio, as compared with that prepared in MES buffer. However, it is possible to obtain a collagen/phospholipid polymer hybrid gel with a high polymer portion and the cross-linking rate can be successfully controlled.

  2. Enzymatic cross-linking of soy proteins within non-fat set yogurt gel.

    PubMed

    Soleymanpuori, Rana; Madadlou, Ashkan; Zeynali, Fariba; Khosrowshahi, Asghar

    2014-08-01

    Soy proteins as the health-promoting ingredients and candidate fat substitutes in dairy products are good substrates for the cross-linking action of the enzyme transglutaminase. Non-fat set yogurt samples were prepared from the milks enriched with soy protein isolate (SPI) and/or treated with the enzyme transglutaminase. The highest titrable acidity was recorded for the yogurt enriched with SPI and treated with the enzyme throughout the cold storage for 21 d. SPI-enrichment of yogurt milk increased the water holding capacity. Although enrichment with SPI did not influence the count of Streptococcus themophilus, increased that of Lactobacillus bulgaricus ∼3 log cycles. The enzymatic treatment of SPI-enriched milk however, suppressed the bacteria growth-promoting influence of SPI due probably to making the soy proteins inaccessible for Lactobacillus. SPI-enrichment and enzymatic treatment of milk decreased the various organic acids content in yoghurt samples; influence of the former was more significant. The cross-linking of milk proteins to soy proteins was confirmed with the gel electrophoresis results.

  3. Microencapsulation of islets within alginate/poly(ethylene glycol) gels cross-linked via Staudinger ligation

    PubMed Central

    Hall, Kristina K.; Gattás-Asfura, Kerim M.; Stabler, Cherie L.

    2010-01-01

    Functionalized alginate and PEG polymers were used to generate covalently linked alginate-PEG (XAlgPEG) microbeads of high stability. The cell-compatible Staudinger ligation scheme was used to chemoselectively cross-link phosphine-terminated poly(ethylene glycol) (PEG) to azide-functionalized alginate, resulting in XAlgPEG hydrogels. XAlgPEG microbeads were formed by co-incubation of the two polymers, followed by ionic cross-linking of the alginate using barium ions. The enhanced stability and gel properties of the resulting XAlgPEG microbeads, as well as the compatibility of these polymers for the encapsulation of islets and beta cells lines, were investigated. Our data show that XAlgPEG microbeads exhibit superior resistance to osmotic swelling compared to traditional barium cross-linked alginate (Ba-Alg) beads, with a 5-fold reduction in observed swelling, as well as resistance to dissolution via chelation solution. Diffusion and porosity studies found XAlgPEG beads to exhibit properties comparable to standard Ba-Alg. Our data found XAlgPEG microbeads to be highly cell compatible with insulinoma cell lines, as well as rat and human pancreatic islets, where the viability and functional assessment of cells within XAlgPEG were comparable to Ba-Alg controls. The remarkable improved stability, as well as demonstrated cellular compatibility, of XAlgPEG hydrogels makes them an appealing option for a wide variety of tissue engineering applications. PMID:20654745

  4. Transformation of metal-organic framework to polymer gel by cross-linking the organic ligands preorganized in metal-organic framework.

    PubMed

    Ishiwata, Takumi; Furukawa, Yuki; Sugikawa, Kouta; Kokado, Kenta; Sada, Kazuki

    2013-04-10

    Until now, seamless fusion of metal-organic frameworks (MOFs) and covalently cross-linked polymer gels (PG) at molecular level has been extremely rare, since these two matters have been regarded as opposite, that is, hard versus soft. In this report, we demonstrate transformation of cubic MOF crystals to PG via inner cross-linking of the organic linkers in the void space of MOF, followed by decomposition of the metal coordination. The obtained PG behaved as a polyelectrolyte gel, indicating the high content of ionic groups inside. Metal ions were well adsorbed in the PG due to its densely packed carboxylate groups. A chimera-type hybrid material consisting of MOF and PG was obtained by partial hydrolysis of resulting cross-linked MOF. The shape of resulting PG network well reflected the crystal structure of MOF employed as a template. Our results will connect the two different network materials that have been ever studied in the two different fields to provide new soft and hard hybrid materials, and the unique copolymerization in the large void space of the MOF will open a new horizon toward "ideal network polymers" never prepared before now.

  5. Ultraviolet-B radiation induced cross-linking improves physical properties of cold- and warm-water fish gelatin gels and films.

    PubMed

    Otoni, Caio G; Avena-Bustillos, Roberto J; Chiou, Bor-Sen; Bilbao-Sainz, Cristina; Bechtel, Peter J; McHugh, Tara H

    2012-09-01

    Cold- and warm-water fish gelatin granules were exposed to ultraviolet-B radiation for doses up to 29.7 J/cm(2). Solutions and films were prepared from the granules. Gel electrophoresis and refractive index were used to examine changes in molecular weight of the samples. Also, the gel strength and rheological properties of the solutions as well as the tensile and water vapor barrier properties of the films were characterized. SDS-PAGE and refractive index results indicated cross-linking of gelatin chains after exposure to radiation. Interestingly, UV-B treated samples displayed higher gel strengths, with cold- and warm-water fish gelatin having gel strength increases from 1.39 to 2.11 N and from 7.15 to 8.34 N, respectively. In addition, both gelatin samples exhibited an increase in viscosity for higher UV doses. For gelatin films, the cold-water fish gelatin samples made from irradiated granules showed greater tensile strength. In comparison, the warm-water gelatin films made from irradiated granules had lower tensile strength, but better water vapor barrier properties. This might be due to the UV induced cross-linking in warm-water gelatin that disrupted helical structures. Journal of Food Science copy; 2012 Institute of Food Technologists® No claim to original US government works.

  6. Polyimide Aerogels with Three-Dimensional Cross-Linked Structure

    NASA Technical Reports Server (NTRS)

    Meador, Mary Ann B. (Inventor)

    2016-01-01

    A method for creating a three dimensional cross-linked polyimide structure includes dissolving a diamine, a dianhydride, and a triamine in a solvent, imidizing a polyamic acid gel by heating the gel, extracting the gel in a second solvent, supercritically drying the gel, and removing the solvent to create a polyimide aerogel.

  7. Hydrogels Prepared from Cross-Linked Nanofibrillated Cellulose

    Treesearch

    Sandeep S. Nair; J.Y. Zhu; Yulin Deng; Arthur J. Ragauskas

    2014-01-01

    Nanocomposite hydrogels were developed by cross-linking nanofibrillated cellulose with poly(methyl vinyl ether-co-maleic acid) and polyethylene glycol. The cross-linked hydrogels showed enhanced water absorption and gel content with the addition of nanocellulose. In addition, the thermal stability, mechanical strength, and modulus increased with an increase in the...

  8. Agarose and Polyacrylamide Gel Electrophoresis Methods for Molecular Mass Analysis of 5–500 kDa Hyaluronan

    PubMed Central

    Bhilocha, Shardul; Amin, Ripal; Pandya, Monika; Yuan, Han; Tank, Mihir; LoBello, Jaclyn; Shytuhina, Anastasia; Wang, Wenlan; Wisniewski, Hans-Georg; de la Motte, Carol; Cowman, Mary K.

    2011-01-01

    Agarose and polyacrylamide gel electrophoresis systems for the molecular mass-dependent separation of hyaluronan (HA) in the size range of approximately 5–500 kDa have been investigated. For agarose-based systems, the suitability of different agarose types, agarose concentrations, and buffers systems were determined. Using chemoenzymatically synthesized HA standards of low polydispersity, the molecular mass range was determined for each gel composition, over which the relationship between HA mobility and logarithm of the molecular mass was linear. Excellent linear calibration was obtained for HA molecular mass as low as approximately 9 kDa in agarose gels. For higher resolution separation, and for extension to molecular masses as low as approximately 5 kDa, gradient polyacrylamide gels were superior. Densitometric scanning of stained gels allowed analysis of the range of molecular masses present in a sample, and calculation of weight-average and number-average values. The methods were validated for polydisperse HA samples with viscosity-average molecular masses of 112, 59, 37, and 22 kDa, at sample loads of 0.5 µg (for polyacrylamide) to 2.5 µg (for agarose). Use of the methods for electrophoretic mobility shift assays was demonstrated for binding of the HA-binding region of aggrecan (recombinant human aggrecan G1-IGD-G2 domains) to a 150 kDa HA standard. PMID:21684248

  9. Novel cross-linked alcohol-insoluble solid (CL-AIS) affinity gel from pea pod for pectinesterase purification.

    PubMed

    Wu, Ming-Chang; Lin, Guan-Hui; Wang, Yuh-Tai; Jiang, Chii-Ming; Chang, Hung-Min

    2005-10-05

    Alcohol-insoluble solids (AIS) from pea pod were cross-linked (CL-AIS) and used as an affinity gel matrix to isolate pectin esterases (PEs) from tendril shoots of chayote (TSC) and jelly fig achenes (JFA), and the results were compared with those isolated by ion-exchange chromatography with a commercial resin. CL-AIS gel matrix in a column displayed poor absorption and purification fold of PE; however, highly methoxylated CL-AIS (HM-CL-AIS), by exposing CL-AIS to methanolic sulfuric acid to increase the degree of esterification (DE) to 92%, facilitated the enzyme purification. The purified TSC PE and JFA PE by the HM-CL-AIS column were proofed as a single band on an SDS-PAGE gel, showing that the HM-CL-AIS column was a good matrix for purification of PE, either with alkaline isoelectric point (pI) (TSC PE) or with acidic pI (JFA PE).

  10. Ionic cross-linked polyether and silica gel mixed matrix membranes for CO 2 separation from flue gas

    DOE PAGES

    Sekizkardes, Ali K.; Zhou, Xu; Nulwala, Hunaid B.; ...

    2017-09-22

    Mixed matrix membranes (MMMs) were prepared by incorporating 10 wt%, 20 wt% and 30 wt% silica gel filler particles into novel ionic cross-linked polyether (IXPE) polymers. Porous silica gel has the advantage of high surface area that can increase the free volume and permeability in a polymer film while also being commercially available and low cost. The MMMs featured high chemical and thermal stability as well as a modest improvement in storage modulus. These features are due to the excellent interfacial interaction between silica gel filler particles and the polymer matrix. Increasing the loading of silica gel particles in MMMsmore » resulted in higher permeability up to 120 Barrer for CO 2, which is about 40% higher than the neat polymer matrix. Finally, most importantly, the MMMs maintained a very high CO 2/N 2 selectivity performance of around 41 for all particle loadings that were tested.« less

  11. Local delivery of hyaluronan as an adjunct to scaling and root planing in the treatment of chronic periodontitis.

    PubMed

    Johannsen, Annsofi; Tellefsen, Monica; Wikesjö, Ulf; Johannsen, Gunnar

    2009-09-01

    The aim of the present study was to evaluate the adjunctive effect of the local application of a hyaluronan gel to scaling and root planing in the treatment of chronic periodontitis. Twelve patients with chronic periodontitis were recruited to participate in a study with a split-mouth design and provided informed consent. Plaque formation and bleeding on probing were evaluated pretreatment (baseline) and at 1, 4, and 12 weeks post-treatment. Probing depths and attachment levels were evaluated at baseline and at 12 weeks. The patients received full-mouth scaling and root planing. A hyaluronan gel was administered subgingivally in the test sites at baseline and after 1 week. Significant differences between test and control were evaluated using the paired t test, repeated-measures analysis of variance (Wilks lambda), and a non-parametric Wilcoxon signed-rank test. A significant reduction in bleeding on probing scores and probing depths was observed in both groups at 12 weeks (P <0.05). Significantly lower bleeding on probing scores were observed in the hyaluronan group compared to control at 12 weeks (P <0.05). Mean probing depth reductions between baseline and 12 weeks were 1.0 +/- 0.3 mm and 0.8 +/- 0.2 mm for the hyaluronan and control groups, respectively. The difference between the groups was statistically significant (P <0.05). The local application of hyaluronan gel in conjunction with scaling and root planing may have a beneficial effect on periodontal health in patients with chronic periodontitis.

  12. Elasticity, biodegradability and cell adhesive properties of chitosan/hyaluronan multilayer films

    NASA Astrophysics Data System (ADS)

    Schneider, Aurore; Richert, Ludovic; Francius, Gregory; Voegel, Jean-Claude; Picart, Catherine

    2007-03-01

    In the bioengineering field, a recent and promising approach to modifying biomaterial surfaces is the layer-by-layer (LbL) technique used to build thin polyelectrolyte multilayer films. In this work, we focused on polyelectrolyte multilayer films made of two polysaccharides, chitosan (CHI) and hyaluronan (HA), and on the control of their physico-chemical and cell adhesive properties by chemical cross-linking. CHI/HA films were cross-linked using a water soluble carbodiimide and observed by confocal laser scanning microscopy (CLSM) with a fluorescently labeled CHI. Film thicknesses were similar for native and cross-linked films. The film nanometer roughness was measured by atomic force microscopy and was found to be higher for cross-linked films. Cross-linking the films also leads to a drastic change in film stiffness. The elastic modulus of the films (Young's modulus) as measured by AFM nano-indentation was about tenfold increased for cross-linked films as compared to native ones. From a biological point of view, cross-liked films are more resistant to enzymatic degradation by hyaluronidase. Furthermore, the increase in film stiffness has a favorable effect on the adhesion and spreading of chondrosarcoma cells. Thus, the CHI/HA cross-linked films could be used for various applications due to their adhesive properties and to their mechanical properties (including stability in enzymatic media).

  13. Hyaluronan in vaginal secretions: association with recurrent vulvovaginal candidiasis.

    PubMed

    Lev-Sagie, Ahinoam; Nyirjesy, Paul; Tarangelo, Nicholas; Bongiovanni, Ann Marie; Bayer, Cynthia; Linhares, Iara M; Giraldo, Paulo C; Ledger, William J; Witkin, Steven S

    2009-08-01

    We evaluated whether vaginal concentrations of hyaluronan were altered in women with recurrent vulvovaginal candidiasis (RVVC). Lavage samples from 17 women with acute RVVC, 27 women who were receiving a maintenance antifungal regimen, and 24 control women were tested for hyaluronan and interleukin (IL)-6, IL-12, and IL-23 by enzyme-linked immunosorbent assay. Median vaginal hyaluronan concentrations were 33.8 ng/mL (range, 21.6-66.3 ng/mL) in women with acute RVVC, 15.0 ng/mL (range, 11.2-50.6 ng/mL) in women who were receiving maintenance therapy, and 4.2 ng/mL (range, 3.6-12.0 ng/mL) in control subjects (P hyaluronan concentration was 27.4 ng/mL (range, 15.4-37.7 ng/mL) when Candida was detected by microscopy and 9.5 ng/mL (range, 7.7-14.6 ng/mL) in microscopy-negative cases (P = .0354). Elevated hyaluronan levels were associated with itching plus burning (40.7 ng/mL) or itching plus discharge (42.1 ng/mL), as opposed to itching only (6.2 ng/mL; P = .0152). Hyaluronan and IL-6 levels were correlated (P = .0009). Hyaluronan release is a component of the host response to a candidal infection and may contribute to symptoms.

  14. Porous Cross-Linked Polyimide-Urea Networks

    NASA Technical Reports Server (NTRS)

    Meador, Mary Ann B. (Inventor); Nguyen, Baochau N. (Inventor)

    2015-01-01

    Porous cross-linked polyimide-urea networks are provided. The networks comprise a subunit comprising two anhydride end-capped polyamic acid oligomers in direct connection via a urea linkage. The oligomers (a) each comprise a repeating unit of a dianhydride and a diamine and a terminal anhydride group and (b) are formulated with 2 to 15 of the repeating units. The subunit was formed by reaction of the diamine and a diisocyanate to form a diamine-urea linkage-diamine group, followed by reaction of the diamine-urea linkage-diamine group with the dianhydride and the diamine to form the subunit. The subunit has been cross-linked via a cross-linking agent, comprising three or more amine groups, at a balanced stoichiometry of the amine groups to the terminal anhydride groups. The subunit has been chemically imidized to yield the porous cross-linked polyimide-urea network. Also provided are wet gels, aerogels, and thin films comprising the networks, and methods of making the networks.

  15. The properties of water in swollen cross-linked polystyrene sulfo acids

    NASA Astrophysics Data System (ADS)

    Gagarin, A. N.; Tokmachev, M. G.; Kovaleva, S. S.; Ferapontov, N. B.

    2008-11-01

    The properties of water in polystyrene sulfo acid gels with various cross-linking degrees were studied by optical volumetry and dynamic desorption porosimetry. The isotherms of water desorption obtained by dynamic desorption porosimetry coincided with isopiestic isotherms, which allowed this method to be recommended for the determination of the amount of water in polymer gels. Joint optical volumetry and dynamic desorption porosimetry studies showed that the interphase boundary in the cross-liked hydrophilic polymer-water system did not coincide with the visible gel boundary, because gels were two-phase systems, which contained water of two types, “free” and “bound.” The influence of the degree of polymer cross-linking on the amounts and properties of water of the two types was studied. It was shown that constants of water distribution in the polymer could be calculated from the dynamic desorption porosimetry data.

  16. Preparation and characterization of electrospun in-situ cross-linked gelatin-graphite oxide nanofibers.

    PubMed

    Zhan, Jianchao; Morsi, Yosry; Ei-Hamshary, Hany; Al-Deyab, Salem S; Mo, Xiumei

    2016-01-01

    Electrospun gelatin(Gel) nanofibers scaffold has such defects as poor mechanical property and quick degradation due to high solubility. In this study, the in situ cross-linked electrospinning technique was used for the production of gelatin nanofibers. Deionized water was chosen as the spinning solvent and graphite oxide (GO) was chosen as the enhancer. The morphological structure, porosity, thermal property, moisture absorption, and moisture retention performance, hydrolysis resistance, mechanical property, and biocompatibility of the produced nanofibers were investigated. Compared with in situ cross-linked gelatin nanofibers scaffold, in situ cross-linked Gel-GO nanofibers scaffold has the following features: (1) the hydrophilicity, moisture absorption, and moisture retention performance slightly reduce, while the hydrolysis resistance is improved; (2) the breaking strength, breaking elongation, and Young's modulus are significantly improved; (3) the porosity slightly reduces while the biocompatibility considerably increases. The in situ cross-linked Gel-GO nanofibers scaffold is likely to be applied in such fields as drug delivery and scaffold for skin tissue engineering.

  17. Cross-linking by protein oxidation in the rapidly setting gel-based glues of slugs.

    PubMed

    Bradshaw, Andrew; Salt, Michael; Bell, Ashley; Zeitler, Matt; Litra, Noelle; Smith, Andrew M

    2011-05-15

    The terrestrial slug Arion subfuscus secretes a glue that is a dilute gel with remarkable adhesive and cohesive strength. The function of this glue depends on metals, raising the possibility that metal-catalyzed oxidation plays a role. The extent and time course of protein oxidation was measured by immunoblotting to detect the resulting carbonyl groups. Several proteins, particularly one with a relative molecular mass (M(r)) of 165 x 10³, were heavily oxidized. Of the proteins known to distinguish the glue from non-adhesive mucus, only specific size variants were oxidized. The oxidation appears to occur within the first few seconds of secretion. Although carbonyls were detected by 2,4-dinitrophenylhydrazine (DNPH) in denatured proteins, they were not easily detected in the native state. The presence of reversible cross-links derived from carbonyls was tested for by treatment with sodium borohydride, which would reduce uncross-linked carbonyls to alcohols, but stabilize imine bonds formed by carbonyls and thus lead to less soluble complexes. Consistent with imine bond formation, sodium borohydride led to a 20-35% decrease in the amount of soluble protein with a M(r) of 40-165 (x 10³) without changing the carbonyl content per protein. In contrast, the nucleophile hydroxylamine, which would competitively disrupt imine bonds, increased protein solubility in the glue. Finally, the primary amine groups on a protein with a M(r) of 15 x 10³ were not accessible to acid anhydrides. The results suggest that cross-links between aldehydes and primary amines contribute to the cohesive strength of the glue.

  18. Cross-linking by protein oxidation in the rapidly setting gel-based glues of slugs

    PubMed Central

    Bradshaw, Andrew; Salt, Michael; Bell, Ashley; Zeitler, Matt; Litra, Noelle; Smith, Andrew M.

    2011-01-01

    SUMMARY The terrestrial slug Arion subfuscus secretes a glue that is a dilute gel with remarkable adhesive and cohesive strength. The function of this glue depends on metals, raising the possibility that metal-catalyzed oxidation plays a role. The extent and time course of protein oxidation was measured by immunoblotting to detect the resulting carbonyl groups. Several proteins, particularly one with a relative molecular mass (Mr) of 165×103, were heavily oxidized. Of the proteins known to distinguish the glue from non-adhesive mucus, only specific size variants were oxidized. The oxidation appears to occur within the first few seconds of secretion. Although carbonyls were detected by 2,4-dinitrophenylhydrazine (DNPH) in denatured proteins, they were not easily detected in the native state. The presence of reversible cross-links derived from carbonyls was tested for by treatment with sodium borohydride, which would reduce uncross-linked carbonyls to alcohols, but stabilize imine bonds formed by carbonyls and thus lead to less soluble complexes. Consistent with imine bond formation, sodium borohydride led to a 20–35% decrease in the amount of soluble protein with a Mr of 40–165 (×103) without changing the carbonyl content per protein. In contrast, the nucleophile hydroxylamine, which would competitively disrupt imine bonds, increased protein solubility in the glue. Finally, the primary amine groups on a protein with a Mr of 15×103 were not accessible to acid anhydrides. The results suggest that cross-links between aldehydes and primary amines contribute to the cohesive strength of the glue. PMID:21525316

  19. Influence of dialkyne structure on the properties of new click-gels based on hyaluronic acid.

    PubMed

    Testa, Gabriella; Di Meo, Chiara; Nardecchia, Stefania; Capitani, Donatella; Mannina, Luisa; Lamanna, Raffaele; Barbetta, Andrea; Dentini, Mariella

    2009-08-13

    Hydrogels have been widely used in tissue engineering as a support for tissue formation and/or to deliver drug locally. A novel procedure for the in situ rapid chemical gelation of aqueous solutions of hyaluronan (HA) was employed. HA was functionalised with an arm bearing a terminal azido group (HAAA). When HAAA was mixed with a series of dialkyne reagents of different length, a 1,3-dipolar cycloaddition ("click-chemistry") reaction took place in the presence of catalytic amount of Cu(I) resulting in fast gelation at room temperature. The resulting gels were characterised in terms of degree of cross-linking by (1)H HR-MAS NMR. The kinetic of gelation and the determination of elastic moduli as well as the degree of swelling and the controlled release of a model drug, were studied as a function of chemical nature of the dialkyne group, catalyst concentration, HAAA concentration and temperature. All these variables allowed the swelling ratio and the extent of release of a drug, doxorubicin, entrapped within the gel, to be modulated. In all cases the kinetic of release reached the stationary state within 150 h. The height of the plateau was dependent on the overall (chemical and topological) degree of cross-linking.

  20. Tuning chemical and physical cross-links in silk electrogels for morphological analysis and mechanical reinforcement.

    PubMed

    Lin, Yinan; Xia, Xiaoxia; Shang, Ke; Elia, Roberto; Huang, Wenwen; Cebe, Peggy; Leisk, Gary; Omenetto, Fiorenzo; Kaplan, David L

    2013-08-12

    Electrochemically controlled, reversible assembly of biopolymers into hydrogel structures is a promising technique for on-demand cell or drug encapsulation and release systems. An electrochemically sol-gel transition has been demonstrated in regenerated Bombyx mori silk fibroin, offering a controllable way to generate biocompatible and reversible adhesives and other biomedical materials. Despite the involvement of an electrochemically triggered electrophoretic migration of the silk molecules, the mechanism of the reversible electrogelation remains unclear. It is, however, known that the freshly prepared silk electrogels (e-gels) adopt a predominantly random coil conformation, indicating a lack of cross-linking as well as thermal, mechanical, and morphological stabilities. In the present work, the tuning of covalent and physical β-sheet cross-links in silk hydrogels was studied for programming the structural properties. Scanning electron microscopy (SEM) revealed delicate morphology, including locally aligned fibrillar structures, in silk e-gels, preserved by combining glutaraldehyde-cross-linking and ethanol dehydration. Fourier transform infrared (FTIR) spectroscopic analysis of either electrogelled, vortex-induced or spontaneously formed silk hydrogels showed that the secondary structure of silk e-gels was tunable between non-β-sheet-dominated and β-sheet-dominated states. Dynamic oscillatory rheology confirmed the mechanical reinforcement of silk e-gels provided by controlled chemical and physical cross-links. The selective incorporation of either chemical or physical or both cross-links into the electrochemically responsive, originally unstructured silk e-gel should help in the design for electrochemically responsive protein polymers.

  1. Synthesis and characterization of a novel hyaluronic acid hydrogel.

    PubMed

    Zhao, X

    2006-01-01

    Hyaluronic acid (hyaluronan, HA) has many medical applications as a biomaterial. To enhance its biostability, a novel hydrogel of cross-linked hyaluronic acid was prepared using a double cross-linking process, which involves building cross-linkages between hydroxyl group pairs and carboxyl group pairs. The present study explored a number of cross-linking processes in order to obtain different degrees of cross-linking, which were evaluated by the measurement of water absorption capacity as an index of the gel network density. To gain a better understanding of the stability of the gel, the chemical structure and particularly the rheological behaviour of the cross-linked HA, which included the influences of factors, such as degree of cross-linking, HA concentration and gel particle size, were investigated. The in vitro biostability against hyaluronidase and free radical degradation was tested to show that the cross-linked hydrogel had improved resistance to in vitro hyaluronidase and free radical degradation.

  2. Method of Cross-Linking Aerogels Using a One-Pot Reaction Scheme

    NASA Technical Reports Server (NTRS)

    Meador, Ann B.; Capadona, Lynn A.

    2008-01-01

    A document discusses a new, simplified method for cross-linking silica and other oxide aerogels, with a polymeric material to increase strength of such materials without adversely affecting porosity or low density. This innovation introduces the polymer precursor into the sol before gelation either as an agent, which co-reacts with the oxide gel, or as soluble polymer precursors, which do not interact with the oxide gel in any way. Subsequent exposure to heat, light, catalyst or other method of promoting polymerization causes cross-linking without any additional infiltration steps.

  3. Viscoelastic measurements after vocal fold scarring in rabbits--short-term results after hyaluronan injection.

    PubMed

    Hertegård, S; Dahlqvist, A; Goodyer, E

    2006-07-01

    The scarring model resulted in significant damage and elevated viscoelasticity of the lamina propria. Hyaluronan preparations may alter viscoelasticity in scarred rabbit vocal folds. Vocal fold scarring results in stiffness of the lamina propria and severe voice problems. The aims of this study were to examine the degree of scarring achieved in the experiment and to measure the viscoelastic properties after injection of hyaluronan in rabbit vocal folds. Twenty-two vocal folds from 15 New Zealand rabbits were scarred, 8 vocal folds were controls. After 8 weeks 12 of the scarred vocal folds received injections with 2 types of cross-linked hyaluronan products and 10 scarred folds were injected with saline. After 11 more weeks the animals were sacrificed. After dissection, 15 vocal folds were frozen for viscoelastic measurements, whereas 14 vocal folds were prepared and stained. Measurements were made of the lamina propria thickness. Viscoelasticity was measured on intact vocal folds with a linear skin rheometer (LSR) adapted to laryngeal measurements. Measurements on the digitized slides showed a thickened lamina propria in the scarred samples as compared with the normal vocal folds (p<0.05). The viscoelastic analysis showed a tendency to stiffening of the scarred vocal folds as compared with the normal controls (p=0.05). There was large variation in stiffness between the two injected hyaluronan products.

  4. Rheological Properties of Cross-Linked Hyaluronan–Gelatin Hydrogels for Tissue Engineering

    PubMed Central

    Vanderhooft, Janssen L.; Alcoutlabi, Mataz; Magda, Jules J.; Prestwich, Glenn D.

    2009-01-01

    Hydrogels that mimic the natural extracellular matrix (ECM) are used in three-dimensional cell culture, cell therapy, and tissue engineering. A semi-synthetic ECM based on cross-linked hyaluronana offers experimental control of both composition and gel stiffness. The mechanical properties of the ECM in part determine the ultimate cell phenotype. We now describe a rheological study of synthetic ECM hydrogels with storage shear moduli that span three orders of magnitude, from 11 to 3 500 Pa, a range important for engineering of soft tissues. The concentration of the chemically modified HA and the cross-linking density were the main determinants of gel stiffness. Increase in the ratio of thiol-modified gelatin reduced gel stiffness by diluting the effective concentration of the HA component. PMID:18839402

  5. Enzymatically Cross-linked Alginic-Hyaluronic acid Composite Hydrogels As Cell Delivery Vehicles

    PubMed Central

    Ganesh, Nitya; Hanna, Craig; Nair, Shantikumar V.; Nair, Lakshmi S.

    2013-01-01

    An injectable composite gel was developed from alginic and hyaluronic acid. The ezymatically cross-linked injectable gels were prepared via the oxidative coupling of tyramine modified sodium algiante and sodium hyaluronate in the presence of horse radish peroxidase (HRP) and hydrogen peroxide (H2O2). The composite gels were prepared by mixing equal parts of the two tryaminated polymer solutions in 10U HRP and treating with 1.0% H2O2. The properties of the alginate gels were significanly affected by the addition of hyaluronic acid. The percentage water absorption and storage modulus of the composite gels were found to be lower than the alginate gels. The alginate and composite gels showed lower protein release compared to hyaluronate gels in the absence of hyaluronidase. Even hyaluronate gels showed only approximately 10% protein release after 14 days incubation in phosphate buffer solution. ATDC-5 cells encapsulated in the injectable gels showed high cell viability. The composite gels showed the presence of enlarged spherical cells with significantly higher metabolic activity compared to cells in hyaluronic and alginic acid gels. The results suggest the potential of the composite approach to develop covalently cross-linked hydrogels with tuneable physical, mechanical, and biological properties. PMID:23357799

  6. Antisense inhibition of hyaluronan synthase-2 in human osteosarcoma cells inhibits hyaluronan retention and tumorigenicity

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Nishida, Yoshihiro; Knudson, Warren; Knudson, Cheryl B.

    2005-07-01

    Osteosarcoma is a common malignant bone tumor associated with childhood and adolescence. The results of numerous studies have suggested that hyaluronan plays an important role in regulating the aggressive behavior of various types of cancer cells. However, no studies have addressed hyaluronan with respect to osteosarcomas. In this investigation, the mRNA expression copy number of three mammalian hyaluronan synthases (HAS) was determined using competitive RT-PCR in the osteoblastic osteosarcoma cell line, MG-63. MG-63 are highly malignant osteosarcoma cells with an abundant hyaluronan-rich matrix. The results demonstrated that HAS-2 is the predominant HAS in MG-63. Accumulation of intracellular hyaluronan increased inmore » association with the proliferative phase of these cells. The selective inhibition of HAS-2 mRNA in MG-63 cells by antisense phosphorothioate oligonucleotides resulted in reduced hyaluronan accumulation by these cells. As expected, the reduction in hyaluronan disrupted the assembly of cell-associated matrices. However, of most interest, coincident with the reduction in hyaluronan, there was a substantial decrease in cell proliferation, a decrease in cell motility and a decrease in cell invasiveness. These data suggest that hyaluronan synthesized by HAS-2 in MG-63 plays a crucial role in osteosarcoma cell proliferation, motility, and invasion.« less

  7. Deformation and fracture of cross-linked polymer gels

    NASA Astrophysics Data System (ADS)

    Lin, Wei-Chun

    Because soft materials, particularly polymer gels, are playing a greater role in industrial and biotechnological applications today, the exploration of their mechanical behavior over a range of deformations is becoming more relevant in our daily lives. Understanding these properties is therefore necessary as a means to predict their response for specific applications. To address these concerns, this dissertation presents a set of analytic tools based on flat punch probe indentation tests to predict the response of polymer gels from a mechanical perspective over a large range of stresses and at failure. At small strains, a novel technique is developed to determine the transport properties of gels based on their measured mechanical behavior. Assuming that a polymer gel behaves in a similar manner as a porous structure, the differentiation of solvent flow from viscoelasticity of a gel network is shown to be possible utilizing a flat, circular punch and a flat, rectangular punch under oscillatory conditions. Use of the technique is demonstrated with a poly(N-isopropyl acrylamide) (pNIPAM) hydrogel. Our results indicate that solvent flow is inhibited at temperatures above the critical solution temperature of 35°C. At high stresses and fracture, the flat probe punch indentation geometry is used to understand how the structure and geometry of silicone based gels affect their mechanical properties. A delayed failure response of the gels is observed and the modes of failure are found to be dependent on the geometry of the system. The addition of a sol fraction in these gels was found to toughen the network and play an important role at these large deformations. Potential mechanisms of fracture resistance are discussed, as is the effect of geometric confinement as it relates to large scale deformation and fracture. These results lay the groundwork for understanding the mechanical response of other highly, deformable material systems utilizing this particular geometry.

  8. Repair of osteochondral defects with hyaluronan- and polyester-based scaffolds.

    PubMed

    Solchaga, Luis A; Temenoff, Johnna S; Gao, Jizong; Mikos, Antonios G; Caplan, Arnold I; Goldberg, Victor M

    2005-04-01

    The natural repair of osteochondral defects can be enhanced with biocompatible, biodegradable materials that support the repair process. It is our hypothesis that hyaluronan-based scaffolds are superior to synthetic scaffolds because they provide biological cues. We tested this thesis by comparing two hyaluronan-based scaffolds [auto cross-linked polysaccharide polymer (ACP) and HYAFF-11] to polyester-based scaffolds [poly(DL-lactic-co-glycolic acid) (PLGA) and poly(L-lactic acid) (PLLA)] with similar pore size, porosity and degradation times. Fifty-four rabbits received bilateral osteochondral defects. One defect received a hyaluronan-based scaffold and the contralateral defect received the corresponding polyester-based scaffold. Rabbits were euthanized 4, 12 and 20 weeks after surgery and the condyles dissected and processed for histology. Only ACP-treated defects presented bone at the base of the defect at 4 weeks. At 12 weeks, only defects treated with rapidly dissolving implants (ACP and PLGA) presented bone reconstitution consistently, while bone was present in only one third of those treated with slowly dissolving scaffolds (HYAFF-11 and PLLA). After 20 weeks, the articular surface of PLGA-treated defects presented fibrillation more frequently than in ACP-treated defects. The surface of defects treated with slowly dissolving scaffolds presented more cracks and fissures. The degradation rate of the scaffolds is critical for the repair process. Slowly dissolving scaffolds sustain thicker cartilage at the surface but, it frequently presents cracks and discontinuities. These scaffolds also delay bone formation at the base of the defects. Hyaluronan-based scaffolds appear to allow faster cell infiltration leading to faster tissue formation. The degradation of ACP leads to rapid bone formation while the slow degradation of HYAFF-11 prolongs the presence of cartilage and delays endochondral bone formation.

  9. Hyaluronan (HA) interacting proteins RHAMM and hyaluronidase impact prostate cancer cell behavior and invadopodia formation in 3D HA-based hydrogels.

    PubMed

    Gurski, Lisa A; Xu, Xian; Labrada, Lyana N; Nguyen, Ngoc T; Xiao, Longxi; van Golen, Kenneth L; Jia, Xinqiao; Farach-Carson, Mary C

    2012-01-01

    To study the individual functions of hyaluronan interacting proteins in prostate cancer (PCa) motility through connective tissues, we developed a novel three-dimensional (3D) hyaluronic acid (HA) hydrogel assay that provides a flexible, quantifiable, and physiologically relevant alternative to current methods. Invasion in this system reflects the prevalence of HA in connective tissues and its role in the promotion of cancer cell motility and tissue invasion, making the system ideal to study invasion through bone marrow or other HA-rich connective tissues. The bio-compatible cross-linking process we used allows for direct encapsulation of cancer cells within the gel where they adopt a distinct, cluster-like morphology. Metastatic PCa cells in these hydrogels develop fingerlike structures, "invadopodia", consistent with their invasive properties. The number of invadopodia, as well as cluster size, shape, and convergence, can provide a quantifiable measure of invasive potential. Among candidate hyaluronan interacting proteins that could be responsible for the behavior we observed, we found that culture in the HA hydrogel triggers invasive PCa cells to differentially express and localize receptor for hyaluronan mediated motility (RHAMM)/CD168 which, in the absence of CD44, appears to contribute to PCa motility and invasion by interacting with the HA hydrogel components. PCa cell invasion through the HA hydrogel also was found to depend on the activity of hyaluronidases. Studies shown here reveal that while hyaluronidase activity is necessary for invadopodia and inter-connecting cluster formation, activity alone is not sufficient for acquisition of invasiveness to occur. We therefore suggest that development of invasive behavior in 3D HA-based systems requires development of additional cellular features, such as activation of motility associated pathways that regulate formation of invadopodia. Thus, we report development of a 3D system amenable to dissection of

  10. Fabrication of nonwoven fabrics consisting of gelatin nanofibers cross-linked by glutaraldehyde or N-acetyl-d-glucosamine by aqueous method.

    PubMed

    Furuike, Tetsuya; Chaochai, Thitirat; Okubo, Tsubasa; Mori, Takahiro; Tamura, Hiroshi

    2016-12-01

    Since gelatin (Gel) undergoes a sol-gel transition, a novel dry-spinning procedure for Gel was used. Here, nonwoven fabrics of Gel were electrospun by applying the principles of dry spinning. The diameter of the fibers and the viscosity and flow rate of the solution were directly dependent on the concentration of Gel. Nonwoven fabrics spun with a 25% (w/w) Gel concentration only exhibited a nanoscale fiber diameter. In order to improve the properties of the nonwoven fabrics, they were cross-linked with glutaraldehyde (GTA) vapor after spinning or by the addition of N-acetyl-d-glucosamine (GlcNAc) to the Gel solution prior to spinning followed by heating these fibers. The developed nonwoven fibers were characterized using SEM, rheometry, FTIR, TGA, and mechanical tensile testing. The nonwoven fabrics cross-linked by the GTA vapor exhibited improved mechanical properties compared to those without cross-linking or with GlcNAc cross-linking. The swelling and water uptake ability resulted in no morphological changes in the fibers with GTA cross-linking. The TGA thermogram confirmed no phase change in the composite structure. Further, in vitro cytocompatibility studies using human mesenchymal stem cells showed the compatible nature of the developed nonwoven fibers. Our studies showed that these nonwoven fibers could be useful in medical care. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Assessment of improved buccal permeation and bioavailability of felodipine microemulsion-based cross-linked polycarbophil gel.

    PubMed

    Singh, Mahendra; Kanoujia, Jovita; Parashar, Poonam; Arya, Malti; Tripathi, Chandra B; Sinha, V R; Saraf, Shailendra K; Saraf, Shubhini A

    2018-06-01

    The oral bioavailability of felodipine (FEL) is very low, i.e., about 15%. This could be due to low water solubility and hepatic first-pass effect. The objective of the present study was to develop FEL microemulsion-based gel, to bypass the first pass effect, for buccal delivery. The optimized FEL microemulsion (OPT-MEF) was used to prepare buccoadhesive gels, with varying concentrations of hydroxypropyl methylcellulose (HPMC) E4M and polycarbophil (PCP), and evaluated. The cross-linking of the PCP gelling agent was done by adjusting the pH with a neutralizing agent, triethanolamine (TEA). The formulations, namely drug suspension, OPT-MEF, microemulsion-based buccal gel containing 1% w/v (MEF-E4M1), 2% w/v (MEF-E4M2), and 3% w/v (MEF-E4M3) of HPMC K4M and 1% w/v (MEF-PCP1), 2% w/v (MEF-PCP2), and 3% w/v (MEF-PCP3) of PCP were prepared and optimized on the basis of ex vivo permeation study, mucoadhesion force, and viscosity. The optimized buccal gel (MEF-PCP1) showed significantly higher (p < 0.01) permeation flux (J = 0.44 ± 0.16 mg/cm 2 /h), when compared with the drug suspension (J = 0.17 ± 0.14 mg/cm 2 /h). The permeation enhancement ratio of MEF-PCP1 was found to be 2.59 times higher than that of the aqueous suspension of the drug. The texture profile analysis of MEF-PCP1 was performed which showed spreadability (3.2 mJ), extrudability (151.8 mJ), hardness (13.8 g), and adhesiveness (41.0 g), and results indicated good spreadability and adhesiveness. The rheological study revealed the pseudoplastic flow behavior of MEF-PCP1 buccal gel. The C max value 9.21 ± 2.88 μg/ml of MEF-PCP1 gel was found to be significantly higher (P < 0.01) compared to the same dose administered by oral route (C max value 3.51 ± 1.74 μg/ml). The relative bioavailability (F r ) of the optimized MEF-PCP1 buccal gel was about 397.39% higher than that of oral route. In conclusion, consistent and effective buccal gel containing optimized FEL

  12. Polyimide Aerogels with Three-Dimensional Cross-Linked Structure

    NASA Technical Reports Server (NTRS)

    Panek, John

    2010-01-01

    Polyimide aerogels with three-dimensional cross-linked structure are made using linear oligomeric segments of polyimide, and linked with one of the following into a 3D structure: trifunctional aliphatic or aromatic amines, latent reactive end caps such as nadic anhydride or phenylethynylphenyl amine, and silica or silsesquioxane cage structures decorated with amine. Drying the gels supercritically maintains the solid structure of the gel, creating a polyimide aerogel with improved mechanical properties over linear polyimide aerogels. Lightweight, low-density structures are desired for acoustic and thermal insulation for aerospace structures, habitats, astronaut equipment, and aeronautic applications. Aerogels are a unique material for providing such properties because of their extremely low density and small pore sizes. However, plain silica aerogels are brittle. Reinforcing the aerogel structure with a polymer (X-Aerogel) provides vast improvements in strength while maintaining low density and pore structure. However, degradation of polymers used in cross-linking tends to limit use temperatures to below 150 C. Organic aerogels made from linear polyimide have been demonstrated, but gels shrink substantially during supercritical fluid extraction and may have lower use temperature due to lower glass transition temperatures. The purpose of this innovation is to raise the glass transition temperature of all organic polyimide aerogel by use of tri-, tetra-, or poly-functional units in the structure to create a 3D covalently bonded network. Such cross-linked polyimides typically have higher glass transition temperatures in excess of 300 400 C. In addition, the reinforcement provided by a 3D network should improve mechanical stability, and prevent shrinkage on supercritical fluid extraction. The use of tri-functional aromatic or aliphatic amine groups in the polyimide backbone will provide such a 3D structure.

  13. Gel Point Suppression in RAFT Polymerization of Pure Acrylic Cross-Linker Derived from Soybean Oil.

    PubMed

    Yan, Mengguo; Huang, Yuerui; Lu, Mingjia; Lin, Fang-Yi; Hernández, Nacú B; Cochran, Eric W

    2016-08-08

    Here we report the reversible addition-fragmentation chain transfer (RAFT) polymerization of acrylated epoxidized soybean oil (AESO), a cross-linker molecule, to high conversion (>50%) and molecular weight (>100 kDa) without macrogelation. Surprisingly, gelation is suppressed in this system far beyond the expectations predicated both on Flory-Stockmeyer theory and multiple other studies of RAFT polymerization featuring cross-linking moieties. By varying AESO and initiator concentrations, we show how intra- versus intermolecular cross-linking compete, yielding a trade-off between the degree of intramolecular linkages and conversion at gel point. We measured polymer chain characteristics, including molecular weight, chain dimensions, polydispersity, and intrinsic viscosity, using multidetector gel permeation chromatography and NMR to track polymerization kinetics. We show that not only the time and conversion at macrogelation, but also the chain architecture, is largely affected by these reaction conditions. At maximal AESO concentration, the gel point approaches that predicted by the Flory-Stockmeyer theory, and increases in an exponential fashion as the AESO concentration decreases. In the most dilute solutions, macrogelation cannot be detected throughout the entire reaction. Instead, cyclization/intramolecular cross-linking reactions dominate, leading to microgelation. This work is important, especially in that it demonstrates that thermoplastic rubbers could be produced based on multifunctional renewable feedstocks.

  14. Organization of photosystem I polypeptides examined by chemical cross-linking

    NASA Technical Reports Server (NTRS)

    Armbrust, T. S.; Chitnis, P. R.; Guikema, J. A.; Spooner, B. S. (Principal Investigator)

    1996-01-01

    Photosystem I from the cyanobacterium Synechocystis sp. PCC 6803 was examined using the chemical cross-linkers glutaraldehyde and N-ethyl-1-3-[3-(dimethylamino)propyl]carbodiimide to investigate the organization of the polypeptide subunits. Thylakoid membranes and photosystem I, which was isolated by Triton X-100 fractionation, were treated with cross-linking reagents and were resolved using a Tricine/urea low-molecular-weight resolution gel system. Subunit-specific antibodies and western blotting analysis were used to identify the components of cross-linked species. These analyses identified glutaraldehyde-dependent cross-linking products composed of small amounts of PsaD and PsaC, PsaC and PsaE, and PsaE and PsaF. The novel cross-link between PsaE and PsaF was also observed following treatment with N-ethyl-1-3-[3-(dimethylamino)propyl]carbodiimide. These cross-linking results suggest a structural interaction between PsaE and PsaF and predict a transmembrane topology for PsaF.

  15. Magnetic iron oxide nanoparticles (MIONs) cross-linked natural polymer-based hybrid gel beads: Controlled nano anti-TB drug delivery application.

    PubMed

    Kesavan, Mookkandi Palsamy; Ayyanaar, Srinivasan; Vijayakumar, Vijayaparthasarathi; Dhaveethu Raja, Jeyaraj; Annaraj, Jamespandi; Sakthipandi, Kathiresan; Rajesh, Jegathalaprathaban

    2018-04-01

    The nanosized rifampicin (RIF) has been prepared to increase the solubility in aqueous solution, which leads to remarkable enhancement of its bioavailability and their convenient delivery system studied by newly produced nontoxic, biodegradable magnetic iron oxide nanoparticles (MIONs) cross-linked polyethylene glycol hybrid chitosan (mCS-PEG) gel beads. The functionalization of both nano RIF and mCS-PEG gel beads were studied using various spectroscopic and microscopic techniques. The size of prepared nano RIF was found to be 70.20 ± 3.50 nm. The mechanical stability and swelling ratio of the magnetic gel beads increased by the addition of PEG with a maximum swelling ratio of 38.67 ± 0.29 g/g. Interestingly, this magnetic gel bead has dual responsive assets in the nano drug delivery application (pH and the magnetic field). As we expected, magnetic gel beads show higher nano drug releasing efficacy at acidic medium (pH = 5.0) with maximum efficiency of 71.00 ± 0.87%. This efficacy may also be tuned by altering the external magnetic field and the weight percentage (wt%) of PEG. These results suggest that such a dual responsive magnetic gel beads can be used as a potential system in the nano drug delivery applications. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 1039-1050, 2018. © 2017 Wiley Periodicals, Inc.

  16. Modified gum arabic cross-linked gelatin scaffold for biomedical applications.

    PubMed

    Sarika, P R; Cinthya, Kuriakose; Jayakrishnan, A; Anilkumar, P R; James, Nirmala Rachel

    2014-10-01

    The present work deals with development of modified gum arabic cross-linked gelatin scaffold for cell culture. A new biocompatible scaffold was developed by cross-linking gelatin (Gel) with gum arabic, a polysaccharide. Gum arabic was subjected to periodate oxidation to obtain gum arabic aldehyde (GAA). GAA was reacted with gelatin under appropriate pH to prepare the cross-linked hydrogel. Cross-linking occurred due to Schiff's base reaction between aldehyde groups of oxidized gum arabic and amino groups of gelatin. The scaffold prepared from the hydrogel was characterized by swelling properties, degree of cross-linking, in vitro degradation and scanning electron microscopy (SEM). Cytocompatibility evaluation using L-929 and HepG2 cells confirmed non-cytotoxic and non-adherent nature of the scaffold. These properties are essential for generating multicellular spheroids and hence the scaffold is proposed to be a suitable candidate for spheroid cell culture. Copyright © 2014 Elsevier B.V. All rights reserved.

  17. Preparation and Characterization of Polyurethanes with Cross-Linked Siloxane in the Side Chain by Sol-Gel Reactions

    PubMed Central

    Zhao, Hui; Hao, Tong-Hui; Hu, Guo-Hua; Shi, Dean; Huang, Da; Jiang, Tao; Zhang, Qun-Chao

    2017-01-01

    A series of novel polyurethanes containing cross-linked siloxane in the side chain (SPU) were successfully synthesized through a sol-gel process. The SPU was composed of 0%–20% N-(n-butyl)-3-aminopropyltriethoxysilane (HDI-T) modified hexamethylene diisocynate homopolymer. The effects of HDI-T content on both the structure and properties of SPU were investigated by Fourier transform infrared spectroscopy (FT-IR), X-ray photoelectron spectroscopy (XPS), X-ray diffraction (XRD), scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), differential scanning calorimetry (DSC), thermogravimetric analysis (TGA), mechanical properties tests, gel content test, water contact angle measurement and water absorption test. FT-IR, XPS and XRD results confirmed the successful incorporation of HDI-T onto polyurethanes and the formation of Si–O–Si. The surface roughness and the Si content of SPU enhanced with the increase of HDI-T content. Both crystallization and melting temperature shifted to a lower point after the incorporation of HDI-T. The hydrophobicity, tensile strength, Young’s modulus and pencil hardness overall increased with the increasing of HDI-T content, whereas the thermal stability and the elongation at break of SPU slightly decreased. PMID:28772607

  18. Cutting edge: high molecular weight hyaluronan promotes the suppressive effects of CD4+CD25+ regulatory T cells.

    PubMed

    Bollyky, Paul L; Lord, James D; Masewicz, Susan A; Evanko, Stephen P; Buckner, Jane H; Wight, Thomas N; Nepom, Gerald T

    2007-07-15

    Hyaluronan is a glycosaminoglycan present in the extracellular matrix. When hyaluronan is degraded during infection and injury, low m.w. forms are generated whose interactions influence inflammation and angiogenesis. Intact high m.w. hyaluronan, conversely, conveys anti-inflammatory signals. We demonstrate that high m.w. hyaluronan enhances human CD4(+)CD25(+) regulatory T cell functional suppression of responder cell proliferation, whereas low m.w. hyaluronan does not. High m.w. hyaluronan also up-regulates the transcription factor FOXP3 on CD4(+)CD25(+) regulatory T cells. These effects are only seen with activated CD4(+)CD25(+) regulatory T cells and are associated with the expression of CD44 isomers that more highly bind high m.w. hyaluronan. At higher concentrations, high m.w. hyaluronan also has direct suppressive effects on T cells. We propose that the state of HA in the matrix environment provides contextual cues to CD4(+)CD25(+) regulatory T cells and T cells, thereby providing a link between the innate inflammatory network and the regulation of adaptive immune responses.

  19. Developing and physicochemical evaluation of cross-linked electrospun gelatin-glycerol nanofibrous membranes for medical applications

    NASA Astrophysics Data System (ADS)

    Morsy, Reda; Hosny, Marwa; Reicha, Fikry; Elnimr, Tarek

    2017-05-01

    This study aims to develop optimal cross-linked electrospun gelatin-glycerol (GEL-GLY) nano-fibrous mats suitable for tissue engineering and wound dressing applications. The optimized procedure involves heating the gelatin and gelatin-glycerol solutions up to 90 °C. The electrospinning process was performed, followed by further cross-linking of electrospun films in a container containing glutaraldehyde (GTA) vapor. The results of X-ray diffraction (XRD), Fourier transformed infrared (FTIR), and Differential thermal analysis (DTA) confirmed that heating gelatin solution up to 90 °C in the presence of glycerol affected the cross-linking efficiency and interactions between GTA molecules and gelatin chains. Scanning Electron Microscope (SEM) analysis showed that GEL-GLY nano-fibrous mats with weight ratios less than or equal (12:3 w/w) exhibited a regular morphology with defect free in addition to increasing the degradation time, cross-linking efficiency, and swelling degree of electrospun gelatin/glycerol.

  20. Effects of alginate hydrogel cross-linking density on mechanical and biological behaviors for tissue engineering.

    PubMed

    Jang, Jinah; Seol, Young-Joon; Kim, Hyeon Ji; Kundu, Joydip; Kim, Sung Won; Cho, Dong-Woo

    2014-09-01

    An effective cross-linking of alginate gel was made through reaction with calcium carbonate (CaCO3). We used human chondrocytes as a model cell to study the effects of cross-linking density. Three different pore size ranges of cross-linked alginate hydrogels were fabricated. The morphological, mechanical, and rheological properties of various alginate hydrogels were characterized and responses of biosynthesis of cells encapsulated in each gel to the variation in cross-linking density were investigated. Desired outer shape of structure was maintained when the alginate solution was cross-linked with the applied method. The properties of alginate hydrogel could be tailored through applying various concentrations of CaCO3. The rate of synthesized GAGs and collagens was significantly higher in human chondrocytes encapsulated in the smaller pore structure than that in the larger pore structure. The expression of chondrogenic markers, including collagen type II and aggrecan, was enhanced in the smaller pore structure. It was found that proper structural morphology is a critical factor to enhance the performance and tissue regeneration. Copyright © 2014 Elsevier Ltd. All rights reserved.

  1. N(4)C-ethyl-N(4)C cross-linked DNA: synthesis and characterization of duplexes with interstrand cross-links of different orientations.

    PubMed

    Noronha, Anne M; Noll, David M; Wilds, Christopher J; Miller, Paul S

    2002-01-22

    The preparation and physical properties of short DNA duplexes that contain a N(4)C-ethyl-N(4)C interstrand cross-link are described. Duplexes that contain an interstrand cross-link between mismatched C-C residues and duplexes in which the C residues of a -CG- or -GC- step are linked to give "staggered" interstrand cross-links were prepared using a novel N(4)C-ethyl-N(4)C phosphoramidite reagent. Duplexes with the C-C mismatch cross-link have UV thermal transition temperatures that are 25 degrees C higher than the melting temperatures of control duplexes in which the cross-link is replaced with a G-C base pair. It appears that this cross-link stabilizes adjacent base pairs and does not perturb the structure of the helix, a conclusion that is supported by the CD spectrum of this duplex and by molecular models. An even higher level of stabilization, 49 degrees C, is seen with the duplex that contains a -CG- staggered cross-link. Molecular models suggest that this cross-link may induce propeller twisting in the cross-linked base pairs, and the CD spectrum of this duplex exhibits an unusual negative band at 298 nm, although the remainder of the spectrum is similar to that of B-form DNA. Mismatched C-C or -CG- staggered cross-linked duplexes that have complementary overhanging ends can undergo self-ligation catalyzed by T4 DNA ligase. Analysis of the ligated oligomers by nondenaturing polyacrylamide gel electrophoresis shows that the resulting oligomers migrate in a manner similar to that of a mixture of non-cross-linked control oligomers and suggests that these cross-links do not result in significant bending of the helix. However, the orientation of the staggered cross-link can have a significant effect on the structure and stability of the cross-linked duplex. Thus, the thermal stability of the duplex that contains a -GC- staggered cross-link is 10 degrees C lower than the melting temperature of the control, non-cross-linked duplex. Unlike the -CG- staggered cross-link

  2. Physicochemical properties of collagen solutions cross-linked by glutaraldehyde.

    PubMed

    Tian, Zhenhua; Li, Conghu; Duan, Lian; Li, Guoying

    2014-06-01

    The physicochemical properties of collagen solutions (5 mg/ml) cross-linked by various amounts of glutaraldehyde (GTA) [GTA/collagen (w/w) = 0-0.5] under acidic condition (pH 4.00) were examined. Based on the results of the determination of residual amino group content, sodium dodecyl sulphate-polyacrylamide gel electrophoresis, dynamic rheological measurements, differential scanning calorimetry and atomic force microscopy (AFM), it was proved that the collagen solutions possessed strikingly different physicochemical properties depending on the amount of GTA. At low GTA amounts [GTA/collagen (w/w) ≤ 0.1], the residual amino group contents of the cross-linked collagens decreased largely from 100% to 32.76%, accompanied by an increase in the molecular weight. Additionally, increases of the fiber diameter and the values of G', G″ and η* were measured, while the thermal denaturation temperature (Td) did not change visibly and the fluidity of collagen samples was still retained with increasing the GTA amount. When the ratio of GTA to collagen exceeded 0.1, although the residual amino group content only decreased by ~8.2%, the cross-linked collagen solution [GTA/collagen (w/w) = 0.3] displayed a clear loss of flow and a sudden rise (~2.0 °C) of the Td value compared to the uncross-linked collagen solution, probably illustrating that the collagen solution was converted into a gel with mature network structure-containing nuclei observed in AFM image. It was conjectured that the physicochemical properties of the collagen solutions might be in connection with the cross-linking between collagen molecules from the same aggregate or different aggregates.

  3. Supermacroporous chemically cross-linked poly(aspartic acid) hydrogels.

    PubMed

    Gyarmati, Benjámin; Mészár, E Zsuzsanna; Kiss, Lóránd; Deli, Mária A; László, Krisztina; Szilágyi, András

    2015-08-01

    Chemically cross-linked poly(aspartic acid) (PASP) gels were prepared by a solid-liquid phase separation technique, cryogelation, to achieve a supermacroporous interconnected pore structure. The precursor polymer of PASP, polysuccinimide (PSI) was cross-linked below the freezing point of the solvent and the forming crystals acted as templates for the pores. Dimethyl sulfoxide was chosen as solvent instead of the more commonly used water. Thus larger temperatures could be utilized for the preparation and the drawback of increase in specific volume of water upon freezing could be eliminated. The morphology of the hydrogels was characterized by scanning electron microscopy and interconnectivity of the pores was proven by the small flow resistance of the gels. Compression tests also confirmed the interconnected porous structure and the complete re-swelling and shape recovery of the supermacroporous PASP hydrogels. The prepared hydrogels are of interest for several biomedical applications as scaffolding materials because of their cytocompatibility, controllable morphology and pH-responsive character. Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  4. Liquid droplets of cross-linked actin filaments

    NASA Astrophysics Data System (ADS)

    Weirich, Kimberly; Banerjee, Shiladitya; Dasbiswas, Kinjal; Vaikuntanathan, Suriyanarayan; Gardel, Margaret

    Soft materials constructed from biomolecules self-assemble into a myriad of structures that work in concert to support cell physiology. One critical soft material is the actin cytoskeleton, a viscoelastic gel composed of cross-linked actin filaments. Although actin networks are primarily known for their elastic properties, which are crucial to regulating cell mechanics, the viscous behavior has been theorized to enable shape changes and flows. We experimentally demonstrate a fluid phase of cross-linked actin, where cross-linker condenses dilute short actin filaments into spindle-shaped droplets, or tactoids. Tactoids have shape dynamics consistent with a continuum model of liquid crystal droplets. The cross-linker, which acts as a long range attractive interaction, analogous to molecular cohesion, controls the tactoid shape and dynamics, which reports on the liquid's interfacial tension and viscosity. We investigate how the cross-linker properties and filament length influence the liquid properties. These results demonstrate a novel mechanism to control organization of the actin cytoskeleton and provide insight into design principles for complex, macromolecular liquid phases.

  5. Textural and cargo release attributes of trisodium citrate cross-linked starch hydrogel.

    PubMed

    Abhari, Negar; Madadlou, Ashkan; Dini, Ali; Hosseini Naveh, Ozra

    2017-01-01

    An alkaline starch suspension was charged with citric acid and incubated for different durations (0, 8.5 or 17h). The suspension was then supplemented with caffeine and gelatinized to fabricate hydrogels which were subsequently stored for varying periods (0, 24 or 48h). Charging of the well-dissolved alkaline starch suspension with citric acid decreased at first both the flow index and consistency coefficient (K); however, starch cross-linking over time by the generated trisodium citrate increased the K value. The latter also inhibited gel syneresis and increased its water-holding capacity. Trisodium citrate did not nonetheless influence the gel hardness except for the sample incubated for maximum duration and stored for the longest period. The amount of the caffeine released from hydrogel decreased by citrate cross-linking and was higher at neutral pH than pH 2.0. Fourier-transform infra-red spectroscopy suggested that caffeine was enclosed within the gel network via non-covalent interactions. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Methacrylated gelatin/hyaluronan-based hydrogels for soft tissue engineering

    PubMed Central

    Kessler, Lukas; Gehrke, Sandra; Winnefeld, Marc; Huber, Birgit; Hoch, Eva; Walter, Torsten; Wyrwa, Ralf; Schnabelrauch, Matthias; Schmidt, Malte; Kückelhaus, Maximilian; Lehnhardt, Marcus; Hirsch, Tobias; Jacobsen, Frank

    2017-01-01

    In vitro–generated soft tissue could provide alternate therapies for soft tissue defects. The aim of this study was to evaluate methacrylated gelatin/hyaluronan as scaffolds for soft tissue engineering and their interaction with human adipose–derived stem cells (hASCs). ASCs were incorporated into methacrylated gelatin/hyaluronan hydrogels. The gels were photocrosslinked with a lithium phenyl-2,4,6-trimethylbenzoylphosphinate photoinitiator and analyzed for cell viability and adipogenic differentiation of ASCs over a period of 30 days. Additionally, an angiogenesis assay was performed to assess their angiogenic potential. After 24 h, ASCs showed increased viability on composite hydrogels. These results were consistent over 21 days of culture. By induction of adipogenic differentiation, the mature adipocytes were observed after 7 days of culture, their number significantly increased until day 28 as well as expression of fatty acid binding protein 4 and adiponectin. Our scaffolds are promising as building blocks for adipose tissue engineering and allowed long viability, proliferation, and differentiation of ASCs. PMID:29318000

  7. Cross-Linked Poly(vinylidene fluoride-co-hexafluoropropene) (PVDF-co-HFP) Gel Polymer Electrolyte for Flexible Li-Ion Battery Integrated with Organic Light Emitting Diode (OLED)

    PubMed Central

    Kim, Ilhwan; Kim, Bong Sung; Nam, Seunghoon; Lee, Hoo-Jeong; Chung, Ho Kyoon; Cho, Sung Min; Luu, Thi Hoai Thuong; Hyun, Seungmin; Kang, Chiwon

    2018-01-01

    Here, we fabricate poly(vinylidene fluoride-co-hexafluoropropene) (PVDF-co-HFP) by electrospinning for a gel polymer electrolyte (GPE) for use in flexible Li-ion batteries (LIBs). As a solvent, we use N-methyl-2-pyrrolidone (NMP), which helps produce the cross-linked morphology of PVDF-co-HFP separator, owing to its low volatility. The cross-linked PVDF-co-HFP separator shows an uptake rate higher than that of a commercialized polypropylene (PP) separator. Moreover, the PVDF-co-HFP separator shows an ionic conductivity of 2.3 × 10−3 S/cm at room temperature, comparable with previously reported values. An LIB full-cell assembled with the PVDF-co-HFP-based GPE shows capacities higher than its counterpart with the commercialized PP separator, confirming that the cross-linked PVDF-co-HFP separator provides highly efficient ionic conducting pathways. In addition, we integrate a flexible LIB cell using the PVDF-co-HFP GPE with a flexible organic light emitting diode (OLED), demonstrating a fully flexible unit of LIB and OLED. PMID:29614800

  8. Cross-Linked Poly(vinylidene fluoride-co-hexafluoropropene) (PVDF-co-HFP) Gel Polymer Electrolyte for Flexible Li-Ion Battery Integrated with Organic Light Emitting Diode (OLED).

    PubMed

    Kim, Ilhwan; Kim, Bong Sung; Nam, Seunghoon; Lee, Hoo-Jeong; Chung, Ho Kyoon; Cho, Sung Min; Luu, Thi Hoai Thuong; Hyun, Seungmin; Kang, Chiwon

    2018-04-02

    Here, we fabricate poly(vinylidene fluoride- co -hexafluoropropene) (PVDF- co -HFP) by electrospinning for a gel polymer electrolyte (GPE) for use in flexible Li-ion batteries (LIBs). As a solvent, we use N -methyl-2-pyrrolidone (NMP), which helps produce the cross-linked morphology of PVDF- co -HFP separator, owing to its low volatility. The cross-linked PVDF- co -HFP separator shows an uptake rate higher than that of a commercialized polypropylene (PP) separator. Moreover, the PVDF- co -HFP separator shows an ionic conductivity of 2.3 × 10 -3 S/cm at room temperature, comparable with previously reported values. An LIB full-cell assembled with the PVDF- co -HFP-based GPE shows capacities higher than its counterpart with the commercialized PP separator, confirming that the cross-linked PVDF- co -HFP separator provides highly efficient ionic conducting pathways. In addition, we integrate a flexible LIB cell using the PVDF- co -HFP GPE with a flexible organic light emitting diode (OLED), demonstrating a fully flexible unit of LIB and OLED.

  9. Nondestructive assessment of collagen hydrogel cross-linking using time-resolved autofluorescence imaging

    NASA Astrophysics Data System (ADS)

    Sherlock, Benjamin E.; Harvestine, Jenna N.; Mitra, Debika; Haudenschild, Anne; Hu, Jerry; Athanasiou, Kyriacos A.; Leach, J. Kent; Marcu, Laura

    2018-03-01

    We investigate the use of a fiber-based, multispectral fluorescence lifetime imaging (FLIm) system to nondestructively monitor changes in mechanical properties of collagen hydrogels caused by controlled application of widely used cross-linking agents, glutaraldehyde (GTA) and ribose. Postcross-linking, fluorescence lifetime images are acquired prior to the hydrogels being processed by rheological or tensile testing to directly probe gel mechanical properties. To preserve the sterility of the ribose-treated gels, FLIm is performed inside a biosafety cabinet (BSC). A pairwise correlation analysis is used to quantify the relationship between mean hydrogel fluorescence lifetimes and the storage or Young's moduli of the gels. In the GTA study, we observe strong and specific correlations between fluorescence lifetime and the storage and Young's moduli. Similar correlations are not observed in the ribose study and we postulate a reason for this. Finally, we demonstrate the ability of FLIm to longitudinally monitor dynamic cross-link formation. The strength of the GTA correlations and deployment of our fiber-based FLIm system inside the aseptic environment of a BSC suggests that this technique may be a valuable tool for the tissue engineering community where longitudinal assessment of tissue construct maturation in vitro is highly desirable.

  10. Hyaluronan as an Immune Regulator in Human Diseases

    PubMed Central

    NOBLE, PAUL W.; LIANG, JIURONG; JIANG, DIANHUA

    2010-01-01

    Accumulation and turnover of extracellular matrix components are the hallmarks of tissue injury. Fragmented hyaluronan stimulates the expression of inflammatory genes by a variety of immune cells at the injury site. Hyaluronan binds to a number of cell surface proteins on a variety of cell types. Hyaluronan fragments signal through both Toll-like receptor (TLR) 4 and TLR2 as well as CD44 to stimulate inflammatory genes in inflammatory cells. Hyaluronan is also present on the cell surface of epithelial cells and provides protection against tissue damage by interacting with TLR2 and TLR4 on these parenchymal cells. Hyaluronan and hyaluronan-binding proteins regulate inflammation, tissue injury and repair through regulating inflammatory cell recruitment, release of inflammatory cytokines, and stem cell migration. This review focuses on the role of hyaluronan as an immune regulator in human diseases. PMID:21248167

  11. Using Greener Gels to Explore Rheology

    ERIC Educational Resources Information Center

    Garrett, Brendan; Matharu, Avtar S.; Hurst, Glenn A.

    2017-01-01

    A laboratory experiment was developed to investigate the rheological properties of a green calcium-cross-linked alginate gel as an alternative to the traditional borax-cross-linked poly(vinyl alcohol) gel. As borax is suspected of damaging fertility and the unborn child, a safe, green alternative is necessary. The rheological properties of a…

  12. Transglutaminase catalyzed cross-linking of sodium caseinate improves oxidative stability of flaxseed oil emulsion.

    PubMed

    Ma, Hairan; Forssell, Pirkko; Kylli, Petri; Lampi, Anna-Maija; Buchert, Johanna; Boer, Harry; Partanen, Riitta

    2012-06-20

    Sodium caseinate was modified by transglutaminase catalyzed cross-linking reaction prior to the emulsification process in order to study the effect of cross-linking on the oxidative stability of protein stabilized emulsions. The extent of the cross-linking catalyzed by different dosages of transglutaminase was investigated by following the ammonia production during the reaction and using SDS-PAGE gel. O/W emulsions prepared with the cross-linked and non-cross-linked sodium caseinates were stored for 30 days under the same conditions. Peroxide value measurement, oxygen consumption measurement, and headspace gas chromatography analysis were used to study the oxidative stability of the emulsions. The emulsion made of the cross-linked sodium caseinate showed an improved oxidative stability with reduced formation of fatty acid hydroperoxides and volatiles and a longer period of low rate oxygen consumption. The improving effect of transglutaminase catalyzed cross-linking could be most likely attributed to the enhanced physical stability of the interfacial protein layer against competitive adsorption by oil oxidation products.

  13. Nondestructive assessment of collagen hydrogel cross-linking using time-resolved autofluorescence imaging.

    PubMed

    Sherlock, Benjamin E; Harvestine, Jenna N; Mitra, Debika; Haudenschild, Anne; Hu, Jerry; Athanasiou, Kyriacos A; Leach, J Kent; Marcu, Laura

    2018-03-01

    We investigate the use of a fiber-based, multispectral fluorescence lifetime imaging (FLIm) system to nondestructively monitor changes in mechanical properties of collagen hydrogels caused by controlled application of widely used cross-linking agents, glutaraldehyde (GTA) and ribose. Postcross-linking, fluorescence lifetime images are acquired prior to the hydrogels being processed by rheological or tensile testing to directly probe gel mechanical properties. To preserve the sterility of the ribose-treated gels, FLIm is performed inside a biosafety cabinet (BSC). A pairwise correlation analysis is used to quantify the relationship between mean hydrogel fluorescence lifetimes and the storage or Young's moduli of the gels. In the GTA study, we observe strong and specific correlations between fluorescence lifetime and the storage and Young's moduli. Similar correlations are not observed in the ribose study and we postulate a reason for this. Finally, we demonstrate the ability of FLIm to longitudinally monitor dynamic cross-link formation. The strength of the GTA correlations and deployment of our fiber-based FLIm system inside the aseptic environment of a BSC suggests that this technique may be a valuable tool for the tissue engineering community where longitudinal assessment of tissue construct maturation in vitro is highly desirable. (2018) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE).

  14. [The application of cross-linked hyaluronate hydrogel to trabeculectomy].

    PubMed

    Shoji, Nobuyuki; Shimizu, Kimiya; Takahashi, Katsuya; Nemoto, Rika; Kawai, Hiromi; Tomioka, Toshiya

    2004-05-01

    We investigated the possibility of applying cross-linked hyaluronate hydrogel (HA gel) during trabeculectomy in rabbit eyes. 1. We injected 0.9% salt solution (n = 3), Opegan-HI (n = 3), or HA gel (n = 3) into the subconjunctiva to make a follicle, and investigated its size. 2. After making a limbal-based conjunctival flap and a scleral flap, we injected HA gel or 0.9% salt solution into the subconjunctiva of rabbit eyes and sutured each flap (n = 4 each). Three weeks after the operation, we incised the conjunctiva and investigated the case of peeling away the conjunctival flap or the scleral flap. 3. We performed trabeculectomy with and without subconjunctival HA gel (n = 7 and 6, respectively), and compared the reduction of intraocular pressure between the two groups. The results showed that the formation of the follicle was excellent and it was easy to peel away the adhesion. The reduction of intraocular pressure was statistically significant 4 weeks after the operation in which the HA gel was used. It could be useful to apply HA gel in trabeculectomy to prevent adhesion.

  15. A Refined Model for the TSG-6 Link Module in Complex with Hyaluronan

    PubMed Central

    Higman, Victoria A.; Briggs, David C.; Mahoney, David J.; Blundell, Charles D.; Sattelle, Benedict M.; Dyer, Douglas P.; Green, Dixy E.; DeAngelis, Paul L.; Almond, Andrew; Milner, Caroline M.; Day, Anthony J.

    2014-01-01

    Tumor necrosis factor-stimulated gene-6 (TSG-6) is an inflammation-associated hyaluronan (HA)-binding protein that contributes to remodeling of HA-rich extracellular matrices during inflammatory processes and ovulation. The HA-binding domain of TSG-6 consists solely of a Link module, making it a prototypical member of the superfamily of proteins that interacts with this high molecular weight polysaccharide composed of repeating disaccharides of d-glucuronic acid and N-acetyl-d-glucosamine (GlcNAc). Previously we modeled a complex of the TSG-6 Link module in association with an HA octasaccharide based on the structure of the domain in its HA-bound conformation. Here we have generated a refined model for a HA/Link module complex using novel restraints identified from NMR spectroscopy of the protein in the presence of 10 distinct HA oligosaccharides (from 4- to 8-mers); the model was then tested using unique sugar reagents, i.e. chondroitin/HA hybrid oligomers and an octasaccharide in which a single sugar ring was 13C-labeled. The HA chain was found to make more extensive contacts with the TSG-6 surface than thought previously, such that a d-glucuronic acid ring makes stacking and ionic interactions with a histidine and lysine, respectively. Importantly, this causes the HA to bend around two faces of the Link module (resembling the way that HA binds to CD44), potentially providing a mechanism for how TSG-6 can reorganize HA during inflammation. However, the HA-binding site defined here may not play a role in TSG-6-mediated transfer of heavy chains from inter-α-inhibitor onto HA, a process known to be essential for ovulation. PMID:24403066

  16. Two Players Make a Formidable Combination: In Situ Generated Poly(acrylic anhydride-2-methyl-acrylic acid-2-oxirane-ethyl ester-methyl methacrylate) Cross-Linking Gel Polymer Electrolyte toward 5 V High-Voltage Batteries.

    PubMed

    Ma, Yue; Ma, Jun; Chai, Jingchao; Liu, Zhihong; Ding, Guoliang; Xu, Gaojie; Liu, Haisheng; Chen, Bingbing; Zhou, Xinhong; Cui, Guanglei; Chen, Liquan

    2017-11-29

    Electrochemical performance of high-voltage lithium batteries with high energy density is limited because of the electrolyte instability and the electrode/electrolyte interfacial reactivity. Hence, a cross-linking polymer network of poly(acrylic anhydride-2-methyl-acrylic acid-2-oxirane-ethyl ester-methyl methacrylate) (PAMM)-based electrolyte was introduced via in situ polymerization inspired by "shuangjian hebi", which is a statement in a traditional Chinese Kungfu story similar to the synergetic effect of 1 + 1 > 2. A poly(acrylic anhydride) and poly(methyl methacrylate)-based system is very promising as electrolyte materials for lithium-ion batteries, in which the anhydride and acrylate groups can provide high voltage resistance and fast ionic conductivity, respectively. As a result, the cross-linking PAMM-based electrolyte possesses a significant comprehensive enhancement, including electrochemical stability window exceeding 5 V vs Li + /Li, an ionic conductivity of 6.79 × 10 -4 S cm -1 at room temperature, high mechanical strength (27.5 MPa), good flame resistance, and excellent interface compatibility with Li metal. It is also demonstrated that this gel polymer electrolyte suppresses the negative effect resulting from dissolution of Mn 2+ ions at 25 and 55 °C. Thus, the LiNi 0.5 Mn 1.5 O 4 /Li and LiNi 0.5 Mn 1.5 O 4 /Li 4 Ti 5 O 12 cells using the optimized in situ polymerized cross-linking PAMM-based gel polymer electrolyte deliver stable charging/discharging profiles and excellent rate performance at room temperature and even at 55 °C. These findings suggest that the cross-linking PAMM is an intriguing candidate for 5 V class high-voltage gel polymer electrolyte toward high-energy lithium-on batteries.

  17. γ-PGA and MTGase improve the formation of ε-(γ-glutamyl) lysine cross-links within hairtail (Trichiurus haumela) surimi protein.

    PubMed

    Hu, Yaqin; Shao, Ying; Wu, Chunhua; Yuan, Chunhong; Ishimura, Gakushi; Liu, Wenjuan; Chen, Shiguo

    2018-03-01

    The present study investigated the mechanism of ε-(γ-glutamyl) lysine cross-links within hairtail (Trichiurus haumela) surimi protein via γ-polyglutamic acid (γ-PGA) and MTGase. The results indicated that the addition of MTGase and γ-PGA markedly improved the gelation properties of hairtail surimi protein, including its maximum breaking force and deformation, water holding capacity and gel strength. The maximum improvements were achieved by adding 0.5units MTGase/g meat paste in combination with 0.06% γ-PGA. SDS-PAGE showed that the band intensity of cross-linked proteins increased, whereas that of myosin heavy chain decreased after treatments. Further scanning electron microscopy (SEM) analysis showed the formation of a denser gel matrix, which was caused by much stronger and more inter- and intra-molecular cross-linking of proteins, via MTGase catalysing ε-(γ-glutamyl) lysine cross-links formed between lysine residues in the gel protein and glutamic residues in the hydrolytic γ-PGA. The results provide reliable guidance for the improvement of hairtail surimi protein gelation properties. Copyright © 2017 Elsevier Ltd. All rights reserved.

  18. Pleural tissue hyaluronan produced by postmortem ventilation in rabbits.

    PubMed

    Wang, P M; Lai-Fook, S J

    2000-01-01

    We developed a method that used Alcian blue bound to hyaluronan to measure pleural hyaluronan in rabbits postmortem. Rabbits were killed, then ventilated with 21% O2--5% CO2--74% N2 for 3 h. The pleural liquid was removed by suction and 5 ml Alcian blue stock solution (0.33 mg/ml, 3.3 pH) was injected into each chest cavity. After 10 min, the Alcian blue solution was removed and the unbound Alcian blue solution (supernatant) separated by centrifugation and filtration. The supernatant transmissibility (T) was measured spectrophotometrically at 613 nm. Supernatant Alcian blue concentration (Cab) was obtained from a calibration curve of T versus dilutions of stock solution Cab. Alcian blue bound to pleural tissue hyaluronan was obtained by subtracting supernatant Cab from stock solution Cab. Pleural tissue hyaluronan was obtained from a calibration curve of hyaluronan versus Alcian blue bound to hyaluronan. Compared with control rabbits, pleural tissue hyaluronan (0.21 +/- 0.04 mg/kg) increased twofold, whereas pleural liquid volume decreased by 30% after 3 h of ventilation. Pleural effusions present 3 h postmortem without ventilation did not change pleural tissue hyaluronan from control values. Thus ventilation-induced pleural liquid shear stress, not increased filtration, was the stimulus for the increased hyaluronan produced from pleural mesothelial cells.

  19. Desmosine-Inspired Cross-Linkers for Hyaluronan Hydrogels

    NASA Astrophysics Data System (ADS)

    Hagel, Valentin; Mateescu, Markus; Southan, Alexander; Wegner, Seraphine V.; Nuss, Isabell; Haraszti, Tamás; Kleinhans, Claudia; Schuh, Christian; Spatz, Joachim P.; Kluger, Petra J.; Bach, Monika; Tussetschläger, Stefan; Tovar, Günter E. M.; Laschat, Sabine; Boehm, Heike

    2013-06-01

    We designed bioinspired cross-linkers based on desmosine, the cross-linker in natural elastin, to prepare hydrogels with thiolated hyaluronic acid. These short, rigid cross-linkers are based on pyridinium salts (as in desmosine) and can connect two polymer backbones. Generally, the obtained semi-synthetic hydrogels are form-stable, can withstand repeated stress, have a large linear-elastic range, and show strain stiffening behavior typical for biopolymer networks. In addition, it is possible to introduce a positive charge to the core of the cross-linker without affecting the gelation efficiency, or consequently the network connectivity. However, the mechanical properties strongly depend on the charge of the cross-linker. The properties of the presented hydrogels can thus be tuned in a range important for engineering of soft tissues by controlling the cross-linking density and the charge of the cross-linker.

  20. Influence of cross-linked arabinoxylans on the postprandial blood glucose response in rats.

    PubMed

    Vogel, Barbara; Gallaher, Daniel D; Bunzel, Mirko

    2012-04-18

    Viscous dietary fibers are well established to reduce the blood glucose response to a meal. In this study, arabinoxylans, the most abundant dietary fiber in most cereals, were extracted under alkaline conditions and cross-linked by using laccase. Cross-linking of the arabinoxylans led to gel formation and increased in vitro viscosity almost 100-fold after drying and rehydration. To determine the ability of these cross-linked arabinoxylans to blunt the postprandial blood glucose curve of a meal, arabinoxylans, either native or cross-linked, and either prehydrated or not, were fed to rats as part of a meal, and blood glucose was monitored at intervals after the meal. Cellulose, a nonviscous fiber, served as a control. Cross-linked, but not native, arabinoxylans significantly reduced the area under the blood glucose time curve 5-9% relative to cellulose, indicating that they remained viscous within the gastrointestinal tract, and thus likely provide the health benefits found with other viscous fibers.

  1. Identification of the bombesin receptor on murine and human cells by cross-linking experiments.

    PubMed

    Kris, R M; Hazan, R; Villines, J; Moody, T W; Schlessinger, J

    1987-08-15

    The bombesin receptor present on the surface of murine and human cells was identified using 125I-labeled gastrin-releasing peptide as a probe, the cross-linking agent disuccinimidyl suberate, and sodium dodecyl sulfate gels. A clone of NIH-3T3 cells which possesses approximately 80,000 bombesin receptors/cell with a single binding constant of approximately 1.9 X 10(-9) M was used in these studies. In addition, we used Swiss 3T3 cells and a human glioma cell line which possesses approximately 100,000 and approximately 55,000 bombesin receptors/cell, respectively. Under conditions found optimal for binding, it is demonstrated that 125I-labeled gastrin-releasing peptide can be cross-linked specifically to a glycoprotein of apparent molecular mass of 65,000 daltons on the surface of the NIH-3T3 cells. Similar results were obtained when the cross-linked product was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing or non-reducing conditions. Moreover, the cross-linking reaction is specific and saturable and the 65,000-dalton polypeptide is not observed when the cross-linking experiments were performed with a NIH-3T3 cell line which is devoid of bombesin receptors. Interestingly, glycoproteins with apparent molecular weights of 75,000 were labeled specifically by 125I-labeled gastrin-releasing peptide when similar experiments were performed with Swiss 3T3 cells and with human glioma cell line GM-340. These different molecular weights may indicate differential glycosylation as treatment with the enzyme N-glycanase reduced the apparent molecular weight of the cross-linked polypeptide to 45,000. On the basis of these results it is concluded that the cross-linked polypeptides represent the bombesin receptor or the ligand-binding subunit of a putative larger bombesin receptor expressed on the surface of these cells.

  2. Investigation of Preparation and Mechanisms of a Dispersed Particle Gel Formed from a Polymer Gel at Room Temperature

    PubMed Central

    Zhao, Guang; Dai, Caili; Zhao, Mingwei; You, Qing; Chen, Ang

    2013-01-01

    A dispersed particle gel (DPG) was successfully prepared from a polymer gel at room temperature. The polymer gel system, morphology, viscosity changes, size distribution, and zeta potential of DPG particles were investigated. The results showed that zirconium gel systems with different strengths can be cross-linked within 2.5 h at low temperature. Scanning electron microscopy (SEM), transmission electron microscopy (TEM), and atomic force microscopy (AFM) results showed that the particles were polygonal particles with nano-size distribution. According to the viscosity changes, the whole preparation process can be divided into two major stages: the bulk gel cross-linking reaction period and the DPG particle preparation period. A polymer gel with a 3-dimensional network was formed in the bulk gel cross-linking reaction period whereas shearing force and frictional force were the main driving forces for the preparation of DPG particles, and thus affected the morphology of DPG particles. High shearing force and frictional force reduced the particle size distribution, and then decreased the zeta potential (absolute value). The whole preparation process could be completed within 3 h at room temperature. It could be an efficient and energy-saving technology for preparation of DPG particles. PMID:24324817

  3. To Cross-Link or Not to Cross-Link? Cross-Linking Associated Foreign Body Response of Collagen-Based Devices

    PubMed Central

    Delgado, Luis M.; Bayon, Yves; Pandit, Abhay

    2015-01-01

    Collagen-based devices, in various physical conformations, are extensively used for tissue engineering and regenerative medicine applications. Given that the natural cross-linking pathway of collagen does not occur in vitro, chemical, physical, and biological cross-linking methods have been assessed over the years to control mechanical stability, degradation rate, and immunogenicity of the device upon implantation. Although in vitro data demonstrate that mechanical properties and degradation rate can be accurately controlled as a function of the cross-linking method utilized, preclinical and clinical data indicate that cross-linking methods employed may have adverse effects on host response, especially when potent cross-linking methods are employed. Experimental data suggest that more suitable cross-linking methods should be developed to achieve a balance between stability and functional remodeling. PMID:25517923

  4. Biocompatibility and antifibrotic effect of UV-cross-linked hyaluronate as a release-system for tranilast after trabeculectomy in a rabbit model--a pilot study.

    PubMed

    Spitzer, Martin S; Sat, Macarena; Schramm, Charlotte; Schnichels, Sven; Schultheiss, Maximilian; Yoeruek, Efdal; Dzhelebov, Dimitar; Szurman, Peter

    2012-06-01

    To analyze the release kinetics and the clinical and histological effects of UV-cross-linked hyaluronic acid as a release-system for the transforming growth factor β-2 antagonist tranilast with anti-phlogistic properties on intraocular pressure after trabeculectomy in an aggressive scarring animal model. Hyaluronate acid was UV-cross linked and loaded with tranilast. The release of tranilast into a buffered salt solution was assessed spectrophotometrically. Glaucoma filtration surgery, similar to that performed in clinical practice, was performed on chinchilla rabbits. The rabbits were divided in 3 groups. (Group A: trabeculectomy alone, group B: trabeculectomy with a cross-linked hyaluronic acid gel preparation and group C: trabeculectomy with cross-linked hyaluronic gel preparation mixed with tranilast). Antifibrotic efficacy was established by clinical response and histologic examination. The cross-linked gels released tranilast for up to 26 h. The release plotted as a function of the square root of time was consistent with a largely diffusion-controlled release system. Both the gel preparation alone and the gel preparation mixed with tranilast were well tolerated in vivo. No adverse effects such as inflammation, corneal toxicity or blurring of the optical media were observed. The intraocular pressure reached preoperative levels within 9 days after surgery in control animals and group B, but remained significantly reduced (p = 0.00016) in the group with tranilast until day 22. The data of this pilot study suggest that the intraoperative application of UV-crossed linked hyaluronic acid used as a slow release system for tranilast may improve the surgical outcome of glaucoma filtration surgery.

  5. Hydration Dynamics of Hyaluronan and Dextran

    PubMed Central

    Hunger, Johannes; Bernecker, Anja; Bakker, Huib J.; Bonn, Mischa; Richter, Ralf P.

    2012-01-01

    Hyaluronan is a polysaccharide, which is ubiquitous in vertebrates and has been reported to be strongly hydrated in a biological environment. We study the hydration of hyaluronan in solution using the rotational dynamics of water as a probe. We measure these dynamics with polarization-resolved femtosecond-infrared and terahertz time-domain spectroscopies. Both experiments reveal that a subensemble of water molecules is slowed down in aqueous solutions of hyaluronan amounting to ∼15 water molecules per disaccharide unit. This quantity is consistent with what would be expected for the first hydration shell. Comparison of these results to the water dynamics in aqueous dextran solution, a structurally similar polysaccharide, yields remarkably similar results. This suggests that the observed interaction with water is a common feature for hydrophilic polysaccharides and is not specific to hyaluronan. PMID:22828349

  6. Strong poly(ethylene oxide) based gel adhesives via oxime cross-linking.

    PubMed

    Ghosh, Smita; Cabral, Jaydee D; Hanton, Lyall R; Moratti, Stephen C

    2016-01-01

    There is a demand for materials to replace or augment the use of sutures and staples in surgical procedures. Currently available commercial surgical adhesives provide either high bond strength with biological toxicity or polymer and protein-based products that are biologically acceptable (though with potential sensitizing potential) but have much reduced bond strength. It is desirable to provide novel biocompatible and biodegradable surgical adhesives/sealants capable of high strength with minimal immune or inflammatory response. In this work, we report the end group derivatization of 8-arm star PEOs with aldehyde and amine end groups. Gels were prepared employing the Schiff-base chemistry between the aldehydes and the amines. Gel setting times, swelling behavior and rheological characterization were carried out for these gels. The mechanical-viscoelastic properties were found to be directly proportional to the crosslinking density of the gels, the 10K PEO gel was stiffer in comparison to the 20K PEO gel. The adhesive properties of these gels were tested using porcine skin and showed excellent adhesion properties. Cytotoxicity studies were carried out for the individual gel components using two different methods: (a) Crystal Violet Staining assay (CVS assay) and (b) impedance and cell index measurement by the xCELLigence system at concentrations >5%. Gels prepared by mixing 20% w/w solutions were also tested for cytotoxicity. The results revealed that the individual gel components as well as the prepared gels and their leachables were non-cytotoxic at these concentrations. This work presents a new type of glue that is aimed at surgery applications using a water soluble star shaped polymer. It show excellent adhesion to skin and is tough and easy to use. We show that it is very biocompatible based on tests on live human cells, and could therefore in principle be used for internal surgery. Comparison with other reported and commercial glues shows that it is stronger

  7. A novel collagen hydrogel cross-linked by gamma-ray irradiation in acidic pH conditions.

    PubMed

    Inoue, Naoki; Bessho, Masahiko; Furuta, Masakazu; Kojima, Takao; Okuda, Shuichi; Hara, Masayuki

    2006-01-01

    We made a new type of collagen gel by gamma-ray irradiation of an acidic solution of type-I collagen, and performed comparative studies on a conventional gel and the new type of gel. The neutral gel, a conventional 0.3% (w/v) collagen gel, was formed at neutral pH and then irradiated by gamma-rays. The acidic gel, a 0.3% (w/v) collagen gel, was formed directly from the acidic solution of collagen by y-ray irradiation. Both types of gel were prepared, swollen in water and then dried for the measurement of specific water content. The neutral gel showed a relatively high specific water content and shrunk moderately, depending on the dose, while the acidic gel showed lower specific water content and shrunk clearly by y-ray irradiation. A three-dimensional tangled network of microfibrils was clearly observed in the neutral gels by scanning electron microscopy, but not in the acidic gels. From these results, we concluded that the acidic gel was quite different from a conventional collagen gel. Sodium dodecylsulfate-polyacrylamide gel electrophoresis showed that the alpha1 subunit and alpha2 subunit of the collagen molecule were cross-linked. The triple-helical structure of collagen was only partially perturbed, but not denatured completely, because the circular dichroism spectrum of the collagen solution irradiated at 1.3 kGy was similar to that of native collagen solution. Amino-acid analysis revealed that tyrosine, phenylalanine and histidine decreased by irradiation in the neutral gel. In the case of the acidic gel, these three amino acids and methionine decreased. We considered that these amino acids were cross-linking points between the collagen subunits during the gamma-ray irradiation.

  8. Induction of hyaluronan cables and monocyte adherence in epidermal keratinocytes.

    PubMed

    Jokela, Tiina A; Lindgren, Antti; Rilla, Kirsi; Maytin, Edward; Hascall, Vincent C; Tammi, Raija H; Tammi, Markku I

    2008-01-01

    Hyaluronan attached to cell surface can form at least two very different structures; a pericellular coat close to plasma membrane and hyaluronan chains coalesced into "cables" that can span several cell lengths. The hyaluronan in cables, induced by many inflammatory agents, can bind leukocytes, whereas that in the pericellular coat does not contribute to leukocyte binding. Therefore, this structural change seems to have a major role in inflammation. In the present study we checked whether cells of squamous epithelium, like epidermal keratinocytes, can form hyaluronan cables and bind leukocytes. In addition, we checked whether hyaluronan synthesis is affected during the induction of cables. Control keratinocytes expressed pericellular hyaluronan as small patches on plasma membrane. But when treated with inflammatory agents or stressful conditions (tunicamycin, interleukin-1beta, tumor necrosis factor-alpha, and high glucose concentration), hyaluronan organization changed into cable-like structures that avidly bound monocytes. Simultaneously, the total amount of secreted hyaluronan was slightly decreased, and the expression levels of hyaluronan synthases (Has1-3) and CD44 were not significantly changed. The results show that epidermal keratinocytes can form cables and bind leukocytes under inflammatory provocation and that these effects are not dependent on stimulation of hyaluronan secretion.

  9. Histidinoalanine, a naturally occurring cross-link derived from phosphoserine and histidine residues in mineral-binding phosphoproteins.

    PubMed

    Marsh, M E

    1986-05-06

    Native mineral-containing phosphoprotein particles were isolated from the Heterodont bivalve Macrocallista nimbosa. The native particles are discrete structures about 40 nm in diameter which migrate as a single band during electrophoresis in agarose gels. Removal of the mineral component with ethylenediaminetetraacetic acid dissociates the native protein into nonidentical subunits. The lower molecular weight subunits, representing 8% of the total protein, were obtained by differential centrifugation. The native protein is characterized by a high content of aspartic acid, phosphoserine, phosphothreonine, histidine, and the bifunctional cross-linking residue histidinoalanine. The low molecular weight subunits have the same amino acid composition except for a reduction in histidinoalanine and a corresponding increase in phosphoserine and histidine residues, demonstrating that the alanine portion of the cross-link is derived from phosphoserine residues. Ion-exchange chromatography and molecular sieve chromatography show that the low molecular weight subunits have a similar charge density but differ in molecular weight, and the relative mobilities of the subunits on agarose gels indicate that they are polymers of a single phosphoprotein molecule. The minimum molecular weight of the monomer is about 140 000 on the basis of the amino acid composition. The high molecular weight subunits are rich in histidinoalanine and too large to be resolved by either molecular sieve chromatography or gel electrophoresis. On the basis of the ultrastructural, electrophoretic, chromatographic, and compositional evidence, native phosphoprotein particles are composed of subunits ionically cross-linked via divalent cations. These subunits are variable molecular weight aggregates of a single phosphoprotein molecule covalently cross-linked via histidinoalanine residues. Evidence for a nonenzymatic cross-linking mechanism is discussed.

  10. Fabrication and characterization of hydrothermal cross-linked chitosan porous scaffolds for cartilage tissue engineering applications.

    PubMed

    Shamekhi, Mohammad Amin; Rabiee, Ahmad; Mirzadeh, Hamid; Mahdavi, Hamid; Mohebbi-Kalhori, Davod; Baghaban Eslaminejad, Mohamadreza

    2017-11-01

    The use of various chemical cross-linking agents for the improvement of scaffolds physical and mechanical properties is a common practical method, which is limited by cytotoxicity effects. Due to exerting contract type forces, chondrocytes are known to implement shrinkage on the tissue engineered constructs, which can be avoided by the scaffold cross-linking. In the this research, chitosan scaffolds are cross-linked with hydrothermal treatment with autoclave sterilization time of 0, 10, 20 and 30min, to avoid the application of the traditional chemical toxic materials. The optimization studies with gel content and crosslink density measurements indicate that for 20min sterilization time, the gel content approaches to ~80%. The scaffolds are fully characterized by the conventional techniques such as SEM, porosity and permeability, XRD, compression, thermal analysis and dynamic mechanical thermal analysis (DMTA). FT-IR studies shows that autoclave inter-chain cross-linking reduces the amine group absorption at 1560cm -1 and increase the absorption of N-acetylated groups at 1629cm -1 . It is anticipated, that this observation evidenced by chitosan scaffold browning upon autoclave cross-linking is an indication of the familiar maillard reaction between amine moieties and carbonyl groups. The biodegradation rate analysis shows that chitosan scaffolds with lower concentrations, possess suitable degradation rate for cartilage tissue engineering applications. In addition, cytotoxicity analysis shows that fabricated scaffolds are biocompatible. The human articular chondrocytes seeding into 3D cross-linked scaffolds shows a higher viability and proliferation in comparison with the uncross-linked samples and 2D controls. Investigation of cell morphology on the scaffolds by SEM, shows a more spherical morphology of chondrocytes on the cross-linked scaffolds for 21days of in vitro culture. Copyright © 2017. Published by Elsevier B.V.

  11. A Study of Cross-linked Regions of Poly(Vinyl Alcohol) Gels by Small-Angle Neutron Scattering

    NASA Astrophysics Data System (ADS)

    Lawrence, Mathias B.; Desa, J. A. E.; Aswal, V. K.

    2011-07-01

    A poly(vinyl alcohol)-borax cross-linked hydrogel has been studied by Small Angle Neutron Scattering as a function of borax concentration in the wave-vector transfer (Q) range of 0.017 Å-1 to 0.36 Å-1. It is found that as the concentration of borax increases, so does the intensity of scattering in this range. Beyond a borax concentration of 2 mg/ml, the increase in cross-linked PVA chains leads to cross-linked units larger than 150 Å as evidenced by a reduction in intensity in the lower Q region.

  12. Caffeine-catalyzed gels.

    PubMed

    DiCiccio, Angela M; Lee, Young-Ah Lucy; Glettig, Dean L; Walton, Elizabeth S E; de la Serna, Eva L; Montgomery, Veronica A; Grant, Tyler M; Langer, Robert; Traverso, Giovanni

    2018-07-01

    Covalently cross-linked gels are utilized in a broad range of biomedical applications though their synthesis often compromises easy implementation. Cross-linking reactions commonly utilize catalysts or conditions that can damage biologics and sensitive compounds, producing materials that require extensive post processing to achieve acceptable biocompatibility. As an alternative, we report a batch synthesis platform to produce covalently cross-linked materials appropriate for direct biomedical application enabled by green chemistry and commonly available food grade ingredients. Using caffeine, a mild base, to catalyze anhydrous carboxylate ring-opening of diglycidyl-ether functionalized monomers with citric acid as a tri-functional crosslinking agent we introduce a novel poly(ester-ether) gel synthesis platform. We demonstrate that biocompatible Caffeine Catalyzed Gels (CCGs) exhibit dynamic physical, chemical, and mechanical properties, which can be tailored in shape, surface texture, solvent response, cargo release, shear and tensile strength, among other potential attributes. The demonstrated versatility, low cost and facile synthesis of these CCGs renders them appropriate for a broad range of customized engineering applications including drug delivery constructs, tissue engineering scaffolds, and medical devices. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

  13. Light-induced cross-linking and post-cross-linking modification of polyglycidol.

    PubMed

    Marquardt, F; Bruns, M; Keul, H; Yagci, Y; Möller, M

    2018-02-08

    The photoinduced radical generation process has received renewed interest due to its economic and ecological appeal. Herein the light-induced cross-linking of functional polyglycidol and its post-cross-linking modification are presented. Linear polyglycidol was first functionalized with a tertiary amine in a two-step reaction. Dimethylaminopropyl functional polyglycidol was cross-linked in a UV-light mediated reaction with camphorquinone as a type II photoinitiator. The cross-linked polyglycidol was further functionalized by quaternization with various organoiodine compounds. Aqueous dispersions of the cross-linked polymers were investigated by means of DLS and zeta potential measurements. Polymer films were evaluated by DSC and XPS.

  14. Triple shape memory effects of cross-linked polyethylene/polypropylene blends with cocontinuous architecture.

    PubMed

    Zhao, Jun; Chen, Min; Wang, Xiaoyan; Zhao, Xiaodong; Wang, Zhenwen; Dang, Zhi-Min; Ma, Lan; Hu, Guo-Hua; Chen, Fenghua

    2013-06-26

    In this paper, the triple shape memory effects (SMEs) observed in chemically cross-linked polyethylene (PE)/polypropylene (PP) blends with cocontinuous architecture are systematically investigated. The cocontinuous window of typical immiscible PE/PP blends is the volume fraction of PE (v(PE)) of ca. 30-70 vol %. This architecture can be stabilized by chemical cross-linking. Different initiators, 2,5-dimethyl-2,5-di(tert-butylperoxy)-hexane (DHBP), dicumylperoxide (DCP) coupled with divinylbenzene (DVB) (DCP-DVB), and their mixture (DHBP/DCP-DVB), are used for the cross-linking. According to the differential scanning calorimetry (DSC) measurements and gel fraction calculations, DHBP produces the best cross-linking and DCP-DVB the worst, and the mixture, DHBP/DCP-DVB, is in between. The chemical cross-linking causes lower melting temperature (Tm) and smaller melting enthalpy (ΔHm). The prepared triple shape memory polymers (SMPs) by cocontinuous immiscible PE/PP blends with v(PE) of 50 vol % show pronounced triple SMEs in the dynamic mechanical thermal analysis (DMTA) and visual observation. This new strategy of chemically cross-linked immiscible blends with cocontinuous architecture can be used to design and prepare new SMPs with triple SMEs.

  15. Sulfur-Containing Organic-Inorganic Hybrid Gel Compositions and Aerogels

    NASA Technical Reports Server (NTRS)

    Evans, Owen R (Inventor); Deshpande, Kiranmayi (Inventor); Dong, Wenting (Inventor)

    2017-01-01

    Methods and materials are described for preparing organic-inorganic hybrid gel compositions where a sulfur-containing cross-linking agent covalently links the organic and inorganic components. The gel compositions are further dried to provide porous gel compositions and aerogels. The mechanical and thermal properties of the dried gel compositions are also disclosed.

  16. Sulfur-Containing Organic-Inorganic Hybrid Gel Compositions and Aerogels

    NASA Technical Reports Server (NTRS)

    Deshpande, Kiranmayi (Inventor); Evans, Owen R. (Inventor); Dong, Wenting (Inventor)

    2015-01-01

    Methods and materials are described for preparing organic-inorganic hybrid gel compositions where a sulfur-containing cross-linking agent covalently links the organic and inorganic components. The gel compositions are further dried to provide porous gel compositions and aerogels. The mechanical and thermal properties of the dried gel compositions are also disclosed.

  17. Bidirectional TSH and IGF-1 receptor cross talk mediates stimulation of hyaluronan secretion by Graves' disease immunoglobins.

    PubMed

    Krieger, Christine C; Neumann, Susanne; Place, Robert F; Marcus-Samuels, Bernice; Gershengorn, Marvin C

    2015-03-01

    There is no pathogenetically linked medical therapy for Graves' ophthalmopathy (GO). Lack of animal models and conflicting in vitro studies have hindered the development of such therapy. Recent reports propose that Graves' Igs bind to and activate thyrotropin receptors (TSHRs) and IGF-1 receptors (IGF-1Rs) on cells in orbital fat, stimulating hyaluronan (HA) secretion, a component of GO. The objective of the study was to investigate potential cross talk between TSHRs and IGF-1Rs in the pathogenesis of GO using a sensitive HA assay. Orbital fibroblasts from GO patients were collected in an academic clinical practice and cultured in a research laboratory. Cells were treated with TSH, IGF-1, and a monoclonal Graves' Ig M22. HA was measured by a modified ELISA. Simultaneous activation by TSH and IGF-1 synergistically increased HA secretion from 320 ± 52 for TSH and 430 ± 65 μg/mL for IGF-1 alone, to 1300 ± 95 μg/mL. IGF-1 shifted the TSH EC50 19-fold to higher potency. The dose response to M22 was biphasic. An IGF-1R antagonist inhibited the higher potency phase but had no effect on the lower potency phase. M22 did not cause IGF-1R autophosphorylation. A TSHR antagonist abolished both phases of M22-stimulated HA secretion. M22 stimulation of HA secretion by GO fibroblasts/preadipocytes involves cross talk between TSHR and IGF-1R. This cross talk relies on TSHR activation rather than direct activation of IGF-1R and leads to synergistic stimulation of HA secretion. These data propose a model for GO pathogenesis that explains previous contradictory results and argues for TSHR as the primary therapeutic target for GO.

  18. Comparative study of thermostability and ester synthesis ability of free and immobilized lipases on cross linked silica gel.

    PubMed

    Kumari, Annapurna; Mahapatra, Paramita; Kumar, Garlapati Vijay; Banerjee, Rintu

    2008-06-01

    A novel support has been utilized for immobilization of lipase, which was prepared by amination of silica with ethanolamine followed by cross linking with glutaraldehyde. Lipases from Rhizopus oryzae 3562 and Enterobacter aerogenes were immobilized on activated silica gel, where they retained 60 and 50% of respective original activity. The thermal stability of the immobilized lipases was significantly improved in comparison to the free forms while the pH stability remained unchanged. E. aerogenes and R. oryzae 3562 lipases retained 75 and 97% of respective initial activity on incubation at 90 degrees C, whereas both the free forms became inactive at this temperature. The conversion yield of isoamyl acetate was found to be higher with the immobilized fungal (90 vs. 21%) and bacterial lipases (64 vs. 18%) than the respective free forms. Immobilized R. oryzae 3562 lipases retained 50% activity for isoamyl acetate synthesis up to ten cycles whereas it was eight cycles for E. aerogenes.

  19. Biocompatible blood pool MRI contrast agents based on hyaluronan

    PubMed Central

    Zhu, Wenlian; Artemov, Dmitri

    2010-01-01

    Biocompatible gadolinium blood pool contrast agents based on a biopolymer, hyaluronan, were investigated for magnetic resonance angiography application. Hyaluronan, a non-sulfated linear glucosaminoglycan composed of 2000–25,000 repeating disaccharide subunits of D-glucuronic acid and N-acetylglucosamine with molecular weight up to 20 MDa, is a major component of the extracellular matrix. Two gadolinium contrast agents based on 16 and 74 kDa hyaluronan were synthesized, both with R1 relaxivity around 5 mM−1 s−1 per gadolinium at 9.4 T at 25°C. These two hyaluronan based agents show significant enhancement of the vasculature for an extended period of time. Initial excretion was primarily through the renal system. Later uptake was observed in the stomach and lower gastrointestinal tract. Macromolecular hyaluronan-based gadolinium agents have a high clinical translation potential as hyaluronan is already approved by FDA for a variety of medical applications. PMID:21504061

  20. Hyaluronan and calcium carbonate hybrid nanoparticles for colorectal cancer chemotherapy

    NASA Astrophysics Data System (ADS)

    Bai, Jinghui; Xu, Jian; Zhao, Jian; Zhang, Rui

    2017-09-01

    A hybrid drug delivery system (DDS) composed of hyaluronan and calcium carbonate (CC) was developed. By taking advantage of the tumor-targeting ability of hyaluronan and the drug-loading property of CC, the well-formed hyaluronan-CC nanoparticles were able to serve as a DDS targeting colorectal cancer with a decent drug loading content, which is beneficial in the chemotherapy of colorectal cancer. In this study, hyaluronan-CC nanoparticles smaller than 100 nm were successfully developed to load the wide-range anti-cancer drug adriamycin (Adr) to construct hyaluronan-CC/Adr nanoparticles. On the other hand, we also found that hyaluronan-CC/Adr nanoparticles can possibly increase the uptake ratio of Adr into HT29 colorectal cancer cells when compared with hyaluronan-free nanoparticles (CC/Adr) via the CD44 receptor-mediated endocytosis via competitive uptake and in vivo imaging assays. Note that both in vitro (CCK-8 assay on HT29 cells) and in vivo (anti-cancer assay on HT-29 tumor-bearing nude mice model) experiments revealed that hyaluronan-CC/Adr nanoparticles exhibited stronger anti-cancer activity than free Adr or CC/Adr nanoparticles with minimized toxic side effects and preferable cancer-suppression potential.

  1. Hyaluronan and Stone Disease

    NASA Astrophysics Data System (ADS)

    Asselman, Marino

    2008-09-01

    Kidney stones cannot be formed as long as crystals are passed in the urine. However, when crystals are retained it becomes possible for them to aggregate and form a stone. Crystals are expected to be formed not earlier than the distal tubules and collecting ducts. Studies both in vitro and in vivo demonstrate that calcium oxalate monohydrate crystals do not adhere to intact distal epithelium, but only when the epithelium is proliferating or regenerating, so that it possesses dedifferentiated cells expressing hyaluronan, osteopontin (OPN) and their mutual receptor CD44 at the apical cell membrane. The polysaccharide hyaluronan is an excellent crystal binding molecule because of its negative ionic charge. We hypothesized that the risk for crystal retention in the human kidney would be increased when tubular cells express hyaluronan at their apical cell membrane. Two different patient categories in which nephrocalcinosis frequently occurs were studied to test this hypothesis (preterm neonates and kidney transplant patients). Hyaluronan (and OPN) expression at the luminal membrane of tubular cells indeed was observed, which preceded subsequent retention of crystals in the distal tubules. Tubular nephrocalcinosis has been reported to be associated with decline of renal function and thus further studies to extend our knowledge of the mechanisms of retention and accumulation of crystals in the kidney are warranted. Ultimately, this may allow the design of new strategies for the prevention and treatment of both nephrocalcinosis and nephrolithiasis in patients.

  2. Tea Derived Galloylated Polyphenols Cross-Link Purified Gastrointestinal Mucins

    PubMed Central

    Georgiades, Pantelis; Pudney, Paul D. A.; Rogers, Sarah; Thornton, David J.; Waigh, Thomas A.

    2014-01-01

    Polyphenols derived from tea are thought to be important for human health. We show using a combination of particle tracking microrheology and small-angle neutron scattering that polyphenols acts as cross-linkers for purified gastrointestinal mucin, derived from the stomach and the duodenum. Both naturally derived purified polyphenols, and green and black tea extracts are shown to act as cross-linkers. The main active cross-linking component is found to be the galloylated forms of catechins. The viscosity, elasticity and relaxation time of the mucin solutions experience an order of magnitude change in value upon addition of the polyphenol cross-linkers. Similarly small-angle neutron scattering experiments demonstrate a sol-gel transition with the addition of polyphenols, with a large increase in the scattering at low angles, which is attributed to the formation of large scale (>10 nm) heterogeneities during gelation. Cross-linking of mucins by polyphenols is thus expected to have an impact on the physicochemical environment of both the stomach and duodenum; polyphenols are expected to modulate the barrier properties of mucus, nutrient absorption through mucus and the viscoelastic microenvironments of intestinal bacteria. PMID:25162539

  3. Factors that influence serum hyaluronan levels in hemodialysis patients.

    PubMed

    de Medina, M; Ashby, M; Diego, J; Pennell, J P; Hill, M; Schiff, E R; Perez, G O

    1999-01-01

    Serum hyaluronan levels are increased in dialysis patients. We evaluated several factors that influence serum hyaluronan levels in 184 patients on chronic hemodialysis (duration 2.3 +/- 2.3 [SD] years). The levels were higher than normal in the whole group and in a subgroup of 133 patients without chronic infection, liver disease, or rheumatoid arthritis (215 +/- 19 and 205 +/- 22 microg/L, respectively). There was a tendency for the levels to be higher in a subgroup of patients with hepatitis c virus (HCV) infection. There was no correlation between hyaluronan levels, alanine aminotransferase (ALT), and duration or dose of dialysis. A weak but highly significant negative correlation between serum albumin levels and serum hyaluronan and ferritin levels was seen. The data suggest that chronic inflammation may explain, at least in part, the increased hyaluronan levels found in chronic dialysis patients.

  4. Hybrid sol-gel optical materials

    DOEpatents

    Zeigler, J.M.

    1993-04-20

    Hybrid sol-gel materials comprise silicate sols cross-linked with linear polysilane, polygermane, or poly(silane-germane). The sol-gel materials are useful as optical identifiers in tagging and verification applications and, in a different aspect, as stable, visible light transparent non-linear optical materials. Methyl or phenyl silicones, polyaryl sulfides, polyaryl ethers, and rubbery polysilanes may be used in addition to the linear polysilane. The linear polymers cross-link with the sol to form a matrix having high optical transparency, resistance to thermooxidative aging, adherence to a variety of substrates, brittleness, and a resistance to cracking during thermal cycling.

  5. Hybrid sol-gel optical materials

    DOEpatents

    Zeigler, John M.

    1993-01-01

    Hybrid sol-gel materials comprise silicate sols cross-linked with linear polysilane, polygermane, or poly(silane-germane). The sol-gel materials are useful as optical identifiers in tagging and verification applications and, in a different aspect, as stable, visible light transparent non-linear optical materials. Methyl or phenyl silicones, polyaryl sulfides, polyaryl ethers, and rubbery polysilanes may be used in addition to the linear polysilane. The linear polymers cross-link with the sol to form a matrix having high optical transparency, resistance to thermooxidative aging, adherence to a variety of substrates, brittleness, and a resistance to cracking during thermal cycling.

  6. Hybrid sol-gel optical materials

    DOEpatents

    Zeigler, John M.

    1992-01-01

    Hybrid sol-gel materials comprise silicate sols cross-linked with linear polysilane, polygermane, or poly(silane-germane). The sol-gel materials are useful as optical identifiers in tagging and verification applications and, in a different aspect, as stable, visible light transparent non-linear optical materials. Methyl or phenyl silicones, polyaryl sulfides, polyaryl ethers, and rubbery polysilanes may be used in addition to the linear polysilane. The linear polymers cross-link with the sol to form a matrix having high optical transparency, resistance to thermooxidative aging, adherence to a variety of substrates, brittleness, and a resistance to cracking during thermal cycling.

  7. Synthesis and properties of reprocessable sulfonated polyimides cross-linked via acid stimulation for use as proton exchange membranes

    NASA Astrophysics Data System (ADS)

    Zhang, Boping; Ni, Jiangpeng; Xiang, Xiongzhi; Wang, Lei; Chen, Yongming

    2017-01-01

    Cross-linked sulfonated polyimides are one of the most promising materials for proton exchange membrane (PEM) applications. However, these cross-linked membranes are difficult to reprocess because they are insoluble. In this study, a series of cross-linkable sulfonated polyimides with flexible pendant alkyl side chains containing trimethoxysilyl groups is successfully synthesized. The cross-linkable polymers are highly soluble in common solvents and can be used to prepare tough and smooth films. Before the cross-linking reaction is complete, the membranes can be reprocessed, and the recovery rate of the prepared films falls within an acceptable range. The cross-linked membranes are obtained rapidly when the cross-linkable membranes are immersed in an acid solution, yielding a cross-linking density of the gel fraction of greater than 90%. The cross-linked membranes exhibit high proton conductivities and tensile strengths under hydrous conditions. Compared with those of pristine membranes, the oxidative and hydrolytic stabilities of the cross-linked membranes are significantly higher. The CSPI-70 membrane shows considerable power density in a direct methanol fuel cell (DMFC) test. All of these results suggest that the prepared cross-linked membranes have great potential for applications in proton exchange membrane fuel cells.

  8. Chromatin structure and methylation of rat rRNA genes studied by formaldehyde fixation and psoralen cross-linking.

    PubMed Central

    Stancheva, I; Lucchini, R; Koller, T; Sogo, J M

    1997-01-01

    By using formaldehyde cross-linking of histones to DNA and gel retardation assays we show that formaldehyde fixation, similar to previously established psoralen photocross-linking, discriminates between nucleosome- packed (inactive) and nucleosome-free (active) fractions of ribosomal RNA genes. By both cross-linking techniques we were able to purify fragments from agarose gels, corresponding to coding, enhancer and promoter sequences of rRNA genes, which were further investigated with respect to DNA methylation. This approach allows us to analyse independently and in detail methylation patterns of active and inactive rRNA gene copies by the combination of Hpa II and Msp I restriction enzymes. We found CpG methylation mainly present in enhancer and promoter regions of inactive rRNA gene copies. The methylation of one single Hpa II site, located in the promoter region, showed particularly strong correlation with the transcriptional activity. PMID:9108154

  9. Photo-Patterned Ion Gel Electrolyte-Gated Thin Film Transistors

    NASA Astrophysics Data System (ADS)

    Choi, Jae-Hong; Gu, Yuanyan; Hong, Kihyun; Frisbie, C. Daniel; Lodge, Timothy P.

    2014-03-01

    We have developed a novel fabrication route to pattern electrolyte thin films in electrolyte-gated transistors (EGTs) using a chemically crosslinkable ABA-triblock copolymer ion gel. In the self-assembly of poly[(styrene-r-vinylbenzylazide)-b-ethylene oxide-b-(styrene-r-vinylbenzylazide)] (SOS-N3) triblock copolymer and the ionic liquid, 1-ethyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide ([EMI][TFSI]), the azide groups of poly(styrene-r-vinylbenzylazide) (PS-N3) end-blocks in the cores can be chemically cross-linked via UV irradiation (λ = 254 nm). Impedance spectroscopy and small-angle X-ray scattering confirmed that ion transport and microstructure of the ion gel are not affected by UV cross-linking. Using this chemical cross-linking strategy, we demonstrate a photo-patterning of ion gels through a patterned mask and the fabricated electrolyte-gated thin film transistors with photo-patterned ion gels as high-capacitance gate insulators exhibited high device performance (low operation voltages and high on/off current ratios).

  10. Computational design of active, self-reinforcing gels.

    PubMed

    Yashin, Victor V; Kuksenok, Olga; Balazs, Anna C

    2010-05-20

    Many living organisms have evolved a protective mechanism that allows them to reversibly alter their stiffness in response to mechanical contact. Using theoretical modeling, we design a mechanoresponsive polymer gel that exhibits a similar self-reinforcing behavior. We focus on cross-linked gels that contain Ru(terpy)(2) units, where both terpyridine ligands are grafted to the chains. The Ru(terpy)(2) complex forms additional, chemoresponsive cross-links that break and re-form in response to a repeated oxidation and reduction of the Ru. In our model, the periodic redox variations of the anchored metal ion are generated by the Belousov-Zhabotinsky (BZ) reaction. Our computer simulations reveal that compression of the BZ gel leads to a stiffening of the sample due to an increase in the cross-link density. These findings provide guidelines for designing biomimetic, active coatings that send out a signal when the system is impacted and use this signaling process to initiate the self-protecting behavior.

  11. Silk Hydrogels of Tunable Structure and Viscoelastic Properties Using Different Chronological Orders of Genipin and Physical Cross-Linking.

    PubMed

    Elliott, Winston H; Bonani, Walter; Maniglio, Devid; Motta, Antonella; Tan, Wei; Migliaresi, Claudio

    2015-06-10

    Catering the hydrogel manufacturing process toward defined viscoelastic properties for intended biomedical use is important to hydrogel scaffolding function and cell differentiation. Silk fibroin hydrogels may undergo "physical" cross-linking through β-sheet crystallization during high pressure carbon dioxide treatment, or covalent "chemical" cross-linking by genipin. We demonstrate here that time-dependent mechanical properties are tunable in silk fibroin hydrogels by altering the chronological order of genipin cross-linking with β-sheet formation. Genipin cross-linking before β-sheet formation affects gelation mechanics through increased molecular weight, affecting gel morphology, and decreasing stiffness response. Alternately, genipin cross-linking after gelation anchored amorphous regions of the protein chain, and increasing stiffness. These differences are highlighted and validated through large amplitude oscillatory strain near physiologic levels, after incorporation of material characterization at molecular and micron length scales.

  12. One-step separation and purification of hydrolysable tannins from Geranium wilfordii Maxim by adsorption chromatography on cross-linked 12% agarose gel.

    PubMed

    Liu, Dan; Ma, Yan; Wang, Ye; Su, Zhiguo; Gu, Ming; Janson, Jan-Christer

    2011-05-01

    The hydrolysable tannins corilagin and geraniin, the major active components of the traditional Chinese medicine Geranium wilfordii Maxim, have been separated and purified from crude extracts in one step by adsorption chromatography on cross-linked 12% agarose gel (Superose 12 10/300 GL). The separation was achieved by gradient elution using mobile phase A composed of 5% ethanol and 5% acetic acid and mobile phase B composed of 30% ethanol and 30% acetic acid. The gradients were composed as follows: 0-240 mL, 0-25% B; 240-480 mL, 25-40% B; after 480 mL, 100% B. The purities of the collected corilagin and geraniin were 92.4 and 87.2%, and the corresponding yields were 88.0 and 76.8%, respectively. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Polyimide aerogels cross-linked through amine functionalized polyoligomeric silsesquioxane.

    PubMed

    Guo, Haiquan; Meador, Mary Ann B; McCorkle, Linda; Quade, Derek J; Guo, Jiao; Hamilton, Bart; Cakmak, Miko; Sprowl, Guilherme

    2011-02-01

    We report the first synthesis of polyimide aerogels cross-linked through a polyhedral oligomeric silsesquioxane, octa(aminophenyl)silsesquioxane (OAPS). Gels formed from polyamic acid solutions of 3,3',4,4'-biphenyltetracarboxylic dianhydride (BPDA), bisaniline-p-xylidene (BAX) and OAPS were chemically imidized and dried using supercritical CO(2) extraction to give aerogels having density around 0.1 g/cm(3). The aerogels are greater than 90 % porous, have high surface areas (230 to 280 m(2)/g) and low thermal conductivity (14 mW/m-K at room temperature). Notably, the polyimide aerogels cross-linked with OAPS have higher modulus than polymer reinforced silica aerogels of similar density and can be fabricated as both monoliths and thin films. Thin films of the aerogel are flexible and foldable making them an ideal insulation for space suits, and inflatable structures for habitats or decelerators for planetary re-entry, as well as more down to earth applications.

  14. Levels of mature cross-links and advanced glycation end product cross-links in human vitreous.

    PubMed

    Matsumoto, Yukihiro; Takahashi, Masaaki; Chikuda, Makoto; Arai, Kiyomi

    2002-01-01

    To determine the levels of pyridinoline and deoxypyridinoline, two mature enzymatic cross-links, and pentosidine, an advanced glycation end product (AGE) cross-link, in the human vitreous, and to investigate the correlations among the cross-links and the effects of aging and diabetes mellitus (DM) on the levels of cross-links. Forty-five vitreous samples were collected from 32 patients (32 eyes) undergoing vitrectomy for diabetic retinopathy (DM group) and from 13 patients (13 eyes) (control group) who were age- and sex-matched patients with idiopathic macular hole or epiretinal membrane with no systemic conditions. The levels of the cross-links were determined using high-performance liquid chromatography after acid hydrolysis and pretreatment with SP-Sephadex. The levels of pentosidine, pyridinoline, and deoxypyridinoline were 27.3 +/- 23.1 (mean +/- SD) pmol/mL (detectable in 45 of 45 specimens), 79.0 +/- 40.2 ng/mL (43 of 45 specimens), and 54.0 +/- 9.5 (32 of 45 specimens) ng/mL, respectively. When the vitreous samples from the DM and the control groups were compared, a significant difference (P <.05) was found in the pentosidine level but not in the levels of pyridinoline or deoxypyridinoline. No significant correlations were found between age and the cross-links. Significant correlations (P <.01) were found among the cross-links. The results indicate that mature cross-link substances exist in the human vitreous. The results also suggest that glycation may occur in the vitreous after mature cross-links form and result in the formation of AGE cross-links. In human vitreous from patients with DM, increased levels of AGE cross-links may stabilize the formation of mature cross-links, but they did not increase the mature cross-links.

  15. Effects of cross-linked high-molecular-weight hyaluronic acid on epidural fibrosis: experimental study.

    PubMed

    Isık, Semra; Taşkapılıoğlu, M Özgür; Atalay, Fatma Oz; Dogan, Seref

    2015-01-01

    Epidural fibrosis is nonphysiological scar formation, usually at the site of neurosurgical access into the spinal canal, in the intimate vicinity of and around the origin of the radicular sheath. The formation of dense fibrous tissue causes lumbar and radicular pain. In addition to radicular symptoms, the formation of scar tissue may cause problems during reoperation. The authors aimed to investigate the effects of cross-linked high-molecular-weight hyaluronic acid (HA), an HA derivative known as HA gel, on the prevention of epidural fibrosis by using histopathological and biochemical parameters. Fifty-six adult female Sprague-Dawley rats were evaluated. The rats were divided into 4 groups. Rats in the sham group (n = 14) underwent laminectomy and discectomy and received no treatment; rats in the control group (n = 14) underwent laminectomy and discectomy and received 0.9% NaCl treatment in the surgical area; rats in the HA group (n = 14) received HA treatment at the surgical area after laminectomy and discectomy; and rats in the HA gel group (n = 14) underwent laminectomy and discectomy in addition to receiving treatment with cross-linked high-molecular-weight HA in the surgical area. All rats were decapitated after 4 weeks, and the specimens were evaluated histopathologically and biochemically. The results were statistically compared using the Mann-Whitney U-test. Compared with the sham and control groups, the HA and HA gel groups showed significantly lower fibroblast cell density and tissue hydroxyproline concentrations (p < 0.05). There was statistically significant lower dural adhesion and foreign-body reaction between the control and HA gel groups (p < 0.05). Granulation tissue and epidural fibrosis were significantly lower in the HA and HA gel groups compared with the sham group (p < 0.05). There were no significant differences in any histopathological parameters or biochemical values between Groups 3 and 4 (p > 0.05). Cross-linked high-molecular-weight HA

  16. Therapeutic Potential of Human Adipose-Derived Stem/Stromal Cell Microspheroids Prepared by Three-Dimensional Culture in Non-Cross-Linked Hyaluronic Acid Gel.

    PubMed

    Mineda, Kazuhide; Feng, Jingwei; Ishimine, Hisako; Takada, Hitomi; Doi, Kentaro; Kuno, Shinichiro; Kinoshita, Kahori; Kanayama, Koji; Kato, Harunosuke; Mashiko, Takanobu; Hashimoto, Ichiro; Nakanishi, Hideki; Kurisaki, Akira; Yoshimura, Kotaro

    2015-12-01

    Three-dimensional culture of mesenchymal stem/stromal cells for spheroid formation is known to enhance their therapeutic potential for regenerative medicine. Spheroids were prepared by culturing human adipose-derived stem/stromal cells (hASCs) in a non-cross-linked hyaluronic acid (HA) gel and compared with dissociated hASCs and hASC spheroids prepared using a nonadherent dish. Preliminary experiments indicated that a 4% HA gel was the most appropriate for forming hASC spheroids with a relatively consistent size (20-50 µm) within 48 hours. Prepared spheroids were positive for pluripotency markers (NANOG, OCT3/4, and SOX-2), and 40% of the cells were SSEA-3-positive, a marker of the multilineage differentiating stress enduring or Muse cell. In contrast with dissociated ASCs, increased secretion of cytokines such as hepatocyte growth factor was detected in ASC spheroids cultured under hypoxia. On microarray ASC spheroids showed upregulation of some pluripotency markers and downregulation of genes related to the mitotic cell cycle. After ischemia-reperfusion injury to the fat pad in SCID mice, local injection of hASC spheroids promoted tissue repair and reduced the final atrophy (1.6%) compared with that of dissociated hASCs (14.3%) or phosphate-buffered saline (20.3%). Part of the administered hASCs differentiated into vascular endothelial cells. ASC spheroids prepared in a HA gel contain undifferentiated cells with therapeutic potential to promote angiogenesis and tissue regeneration after damage. This study shows the therapeutic value of human adipose-derived stem cell spheroids prepared in hyarulonic acid gel. The spheroids have various benefits as an injectable cellular product and show therapeutic potential to the stem cell-depleted conditions such as diabetic chronic skin ulcer. ©AlphaMed Press.

  17. Synthesis and Properties of Cross-Linked Polyamide Aerogels

    NASA Technical Reports Server (NTRS)

    Williams, Jarrod C.; Meador, Mary Ann; McCorkle, Linda

    2015-01-01

    We report the first synthesis of cross-linked polyamide aerogels through step growth polymerization using a combination of diamines, diacid chloride and triacid chloride. Polyamide oligomers endcapped with amines are prepared as stable solutions in N-methylpyrrolidinone from several different diamine precursors and 1,3-benzenedicarbonyl dichloride. Addition of 1,3,5-benzenetricarbonyl trichloride yields gels which form in under five minutes according to the scheme shown. Solvent exchange of the gels into ethanol, followed by drying using supercritical CO2 extraction gives colorless aerogels with densities around 0.1 to 0.2 gcm3. Thicker monolithes of the polyamide aerogels are stiff and strong, while thin films of certain formulations are highly flexible, durable, and even translucent. These materials may have use as insulation for deployable space structures, rovers, habitats or extravehicular activity suits as well as in many terrestrial applications. Strucure property relationships of the aerogels, including surface area, mechanical properties, and thermal conductivity will be discussed.

  18. Chitosan-hyaluronan/nano chondroitin sulfate ternary composite sponges for medical use.

    PubMed

    Anisha, B S; Sankar, Deepthi; Mohandas, Annapoorna; Chennazhi, K P; Nair, Shantikumar V; Jayakumar, R

    2013-02-15

    In this work chitosan-hyaluronan composite sponge incorporated with chondroitin sulfate nanoparticle (nCS) was developed. The fabrication of hydrogel was based on simple ionic cross-linking using EDC, followed by lyophilization to obtain the composite sponge. nCS suspension was characterized using DLS and SEM and showed a size range of 100-150 nm. The composite sponges were characterized using SEM, FT-IR and TG-DTA. Porosity, swelling, biodegradation, blood clotting and platelet activation of the prepared sponges were also evaluated. Nanocomposites showed a porosity of 67% and showed enhanced swelling and blood clotting ability. Cytocompatibility and cell adhesion studies of the sponges were done using human dermal fibroblast (HDF) cells and the nanocomposite sponges showed more than 90% viability. Nanocomposite sponges also showed enhanced proliferation of HDF cells within two days of study. These results indicated that this nanocomposite sponges would be a potential candidate for wound dressing. Copyright © 2012 Elsevier Ltd. All rights reserved.

  19. Tailoring Elastic Properties of Silica Aerogels Cross-Linked with Polystyrene

    NASA Technical Reports Server (NTRS)

    Nguyen, Baochau N.; Meador, Mary Ann B.; Tousley, Marissa E.; Shonkwiler, Brian; McCorkle, Linda; Scheiman, Daniel A.; Palczer, Anna

    2009-01-01

    The effect of incorporating an organic linking group, 1,6-bis(trimethoxysilyl)hexane (BTMSH), into the underlying silica structure of a styrene cross-linked silica aerogel is examined. Vinyltrimethoxysilane (VTMS) is used to provide a reactive site on the silica backbone for styrene polymerization. Replacement of up to 88 mol 1 of the silicon from tetramethoxyorthosilicate with silicon derived from BTMSH and VTMS during the making of silica gels improves the elastic behavior in some formulations of the crosslinked aerogels, as evidenced by measurement of the recovered length after compression of samples to 251 strain. This is especially true for some higher density formulations, which recover nearly 100% of their length after compression to 251 strain twice. The compressive modulus of the more elastic monoliths ranged from 0.2 to 3 MPa. Although some of these monoliths had greatly reduced surface areas, changing the solvent used to produce the gels from methanol to ethanol increased the surface area in one instance from 6 to 220 sq m2/g with little affect on the modulus, elastic recovery, porosity, or density.

  20. Porous Cross-Linked Polyimide Networks

    NASA Technical Reports Server (NTRS)

    Meador, Mary Ann B. (Inventor); Guo, Haiquan (Inventor)

    2015-01-01

    Porous cross-linked polyimide networks are provided. The networks comprise an anhydride end-capped polyamic acid oligomer. The oligomer (i) comprises a repeating unit of a dianhydride and a diamine and terminal anhydride groups, (ii) has an average degree of polymerization of 10 to 50, (iii) has been cross-linked via a cross-linking agent, comprising three or more amine groups, at a balanced stoichiometry of the amine groups to the terminal anhydride groups, and (iv) has been chemically imidized to yield the porous cross-linked polyimide network. Also provided are porous cross-linked polyimide aerogels comprising a cross-linked and imidized anhydride end-capped polyamic acid oligomer, wherein the oligomer comprises a repeating unit of a dianhydride and a diamine, and the aerogel has a density of 0.10 to 0.333 g/cm.sup.3 and a Young's modulus of 1.7 to 102 MPa. Also provided are thin films comprising aerogels, and methods of making porous cross-linked polyimide networks.

  1. Chemically Modified N-Acylated Hyaluronan Fragments Modulate Proinflammatory Cytokine Production by Stimulated Human Macrophages*

    PubMed Central

    Babasola, Oladunni; Rees-Milton, Karen J.; Bebe, Siziwe; Wang, Jiaxi; Anastassiades, Tassos P.

    2014-01-01

    Low molecular mass hyaluronans are known to induce inflammation. To determine the role of the acetyl groups of low molecular mass hyaluronan in stimulating the production of proinflammatory cytokines, partial N-deacetylation was carried out by hydrazinolysis. This resulted in 19.7 ± 3.5% free NH2 functional groups, which were then acylated by reacting with an acyl anhydride, including acetic anhydride. Hydrazinolysis resulted in bond cleavage of the hyaluronan chain causing a reduction of the molecular mass to 30–214 kDa. The total NH2 and N-acetyl moieties in the reacetylated hyaluronan were 0% and 98.7 ± 1.5% respectively, whereas for butyrylated hyaluronan, the total NH2, N-acetyl, and N-butyryl moieties were 0, 82.2 ± 4.6, and 22.7 ± 3.8%, respectively, based on 1H NMR. We studied the effect of these polymers on cytokine production by cultured human macrophages (THP-1 cells). The reacetylated hyaluronan stimulated proinflammatory cytokine production to levels similar to LPS, whereas partially deacetylated hyaluronan had no stimulatory effect, indicating the critical role of the N-acetyl groups in the stimulation of proinflammatory cytokine production. Butyrylated hyaluronan significantly reduced the stimulatory effect on cytokine production by the reacetylated hyaluronan or LPS but had no stimulatory effect of its own. The other partially N-acylated hyaluronan derivatives tested showed smaller stimulatory effects than reacetylated hyaluronan. Antibody and antagonist experiments suggest that the acetylated and partially butyrylated lower molecular mass hyaluronans exert their effects through the TLR-4 receptor system. Selectively N-butyrylated lower molecular mass hyaluronan shows promise as an example of a novel semisynthetic anti-inflammatory molecule. PMID:25053413

  2. Cross-linking of type I collagen with microbial transglutaminase: identification of cross-linking sites.

    PubMed

    Stachel, Ines; Schwarzenbolz, Uwe; Henle, Thomas; Meyer, Michael

    2010-03-08

    Collagen is a popular biomaterial. To deal with its lack of thermal stability and its weak resistance to proteolytic degradation, collagen-based materials are stabilized via different cross-linking procedures. Regarding the potential toxicity of residual cross-linking agents, enzyme-mediated cross-linking would provide an alternative and nontoxic method for collagen stabilization. The results of this study show that type I collagen is a substrate for mTG. However, epsilon-(gamma-glutamyl)lysine cross-links are only incorporated at elevated temperatures when the protein is partially or completely denatured. A maximum number of 5.4 cross-links per collagen monomer were found for heat-denatured collagen. Labeling with the primary amine monodansylcadaverine revealed that at least half of the cross-links are located within the triple helical region of the collagen molecule. Because the triple helix is highly ordered in its native state, this finding might explain why the glutamine residues are inaccessible for mTG under nondenaturing conditions.

  3. Hyaluronan Oligosaccharides for the Promotion of Remyelination (Revised)

    DTIC Science & Technology

    2012-10-01

    AD_________________ Award Number: W81XWH-10-1-0967 TITLE: Hyaluronan oligosaccharides for the...REPORT TYPE FINAL (revised) 3. DATES COVERED 30 2010- 2012 4. TITLE AND SUBTITLE Hyaluronan oligosaccharides for the promotion of...differentiation and remyelination; and if particular formulations of HA oligosaccharides could block the activities of these digestion products

  4. Cyclodextrin-cross-linked diaminotriazine-based hydrogen bonding strengthened hydrogels for drug and reverse gene delivery.

    PubMed

    Hu, Xiufeng; Wang, Ning; Liu, Lu; Liu, Wenguang

    2013-01-01

    A hydrogen bonding strengthened hydrogel was prepared by radical copolymerization of poly(ethylene glycol) methacrylated β-cyclodextrin (PEG-β-CD) and 2-vinyl-4,6-diamino-1,3,5-triazine (VDT) monomer. PEG-β-CD served not only as a cross-linker, but also as a built-in solubilizing agent of the hydrophobic drug in the gel. Increasing VDT content resulted in a notable enhancement in the mechanical strengths of hydrogels whose equilibrium water contents could be modulated from 75% to 85% by varying the ratio of PEG-β-CD cross-linker. It was shown that copolymerizing more PEG-β-CDs could load higher amount of ibuprofen (IBU) in the gels and contribute to a slower release rate of IBU. Plasmid DNA could be anchored onto the surface of hydrogels due to the hydrogen bonding between the base pairs and diaminotriazine, thereby mediating efficient reverse gene transfection of luciferase gene in COS-7 cells cultured on the gel surface. The cytocompatible PEG-β-CD-cross-linked PVDT hydrogels with multifunction of drug and gene delivery hold a potential as tissue engineering scaffold.

  5. Cross-Linked Hydrogel for Pharmaceutical Applications: A Review

    PubMed Central

    2017-01-01

    Hydrogels are promising biomaterials because of their important qualities such as biocompatibility, biodegradability, hydrophilicity and non-toxicity. These qualities make hydrogels suitable for application in medical and pharmaceutical field. Recently, a tremendous growth of hydrogel application is seen, especially as gel and patch form, in transdermal drug delivery. This review mainly focuses on the types of hydrogels based on cross-linking and; secondly to describe the possible synthesis methods to design hydrogels for different pharmaceutical applications. The synthesis and chemistry of these hydrogels are discussed using specific pharmaceutical examples. The structure and water content in a typical hydrogel have also been discussed. PMID:29399542

  6. CD44-mediated hyaluronan binding marks proliferating hematopoietic progenitor cells and promotes bone marrow engraftment

    PubMed Central

    Lee-Sayer, Sally S. M.; Dougan, Meghan N.; Cooper, Jesse; Sanderson, Leslie; Dosanjh, Manisha; Maxwell, Christopher A.

    2018-01-01

    CD44 is a widely expressed cell adhesion molecule that binds to the extracellular matrix component, hyaluronan. However, this interaction is not constitutive in most immune cells at steady state, as the ability of CD44 to engage hyaluronan is highly regulated. While activated T cells and macrophages gain the ability to bind hyaluronan by CD44, the status in other immune cells is less studied. Here we found a percentage of murine eosinophils, natural killer and natural killer T cells were capable of interacting with hyaluronan at steady state. To further investigate the consequences of hyaluronan binding by CD44 in the hematopoietic system, point mutations of CD44 that either cannot bind hyaluronan (LOF-CD44) or have an increased affinity for hyaluronan (GOF-CD44) were expressed in CD44-deficient bone marrow. Competitive bone marrow reconstitution of irradiated mice revealed an early preference for GOF-CD44 over WT-CD44 expressing cells, and for WT-CD44 over LOF-CD44 expressing cells, in the hematopoietic progenitor cell compartment. The advantage of the hyaluronan-binding cells was observed in the hematopoietic stem and progenitor populations, and was maintained throughout the immune system. Hematopoietic stem cells bound minimal hyaluronan at steady state, and this was increased when the cells were induced to proliferate whereas multipotent progenitors had an increased ability to bind hyaluronan at steady state. In vitro, the addition of hyaluronan promoted their proliferation. Thus, proliferating hematopoietic progenitors bind hyaluronan, and hyaluronan binding cells have a striking competitive advantage in bone marrow engraftment. PMID:29684048

  7. Cytocompatible in situ forming chitosan/hyaluronan hydrogels via a metal-free click chemistry for soft tissue engineering.

    PubMed

    Fan, Ming; Ma, Ye; Mao, Jiahui; Zhang, Ziwei; Tan, Huaping

    2015-07-01

    Injectable hydrogels are important cell scaffolding materials for tissue engineering and regenerative medicine. Here, we report a new class of biocompatible and biodegradable polysaccharide hydrogels derived from chitosan and hyaluronan via a metal-free click chemistry, without the addition of copper catalyst. For the metal-free click reaction, chitosan and hyaluronan were modified with oxanorbornadiene (OB) and 11-azido-3,6,9-trioxaundecan-1-amine (AA), respectively. The gelation is attributed to the triazole ring formation between OB and azido groups of polysaccharide derivatives. The molecular structures were verified by FT-IR spectroscopy and elemental analysis, giving substitution degrees of 58% and 47% for chitosan-OB and hyaluronan-AA, respectively. The in vitro gelation, morphologies, equilibrium swelling, compressive modulus and degradation of the composite hydrogels were examined. The potential of the metal-free hydrogel as a cell scaffold was demonstrated by encapsulation of human adipose-derived stem cells (ASCs) within the gel matrix in vitro. Cell culture showed that this metal-free hydrogel could support survival and proliferation of ASCs. A preliminary in vivo study demonstrated the usefulness of the hydrogel as an injectable scaffold for adipose tissue engineering. These characteristics provide a potential opportunity to use the metal-free click chemistry in preparation of biocompatible hydrogels for soft tissue engineering applications. Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  8. Role of hyaluronan chain length in buffering interstitial flow across synovium in rabbits

    PubMed Central

    Coleman, P J; Scott, D; Mason, R M; Levick, J R

    2000-01-01

    Synovial fluid drains out of joints through an interstitial pathway. Hyaluronan, the major polysaccharide of synovial fluid, attenuates this fluid drainage; it creates a graded opposition to outflow that increases with pressure (outflow ‘buffering’). This has been attributed to size-related molecular reflection at the interstitium-fluid interface. Chain length is reduced in inflammatory arthritis. We therefore investigated the dependence of outflow buffering on hyaluronan chain length.Hyaluronan molecules of mean molecular mass ≈2200, 530, 300 and 90 kDa and concentration 3.6 mg ml−1 were infused into the knees of anaesthetized rabbits, with Ringer solution as control in the contralateral joint. Trans-synovial drainage rate was recorded at known joint pressures. Pressure was raised in steps every 30–60 min (range 2–24 cmH2O).With hyaluronan-90 and hyaluronan-300 the fluid drainage rate was reduced relative to Ringer solution (P < 0.001, ANOVA) but increased steeply with pressure. The opposition to outflow, defined as the pressure required to drive unit outflow, did not increase with pressure, i.e. there was no outflow buffering.With hyaluronan-530 and hyaluronan-2000 the fluid drainage rate became relatively insensitive to pressure, causing a near plateau of flow. Opposition to outflow increased markedly with pressure, by up to 3.3 times over the explored pressures.Hyaluronan concentration in the joint cavity increased over the drainage period, indicating partial reflection of hyaluronan by synovial interstitium. Reflected fractions were 0.12, 0.33, 0.25 and 0.79 for hyaluronan-90, -300, -530 and -2200, respectively.Thus the flow-buffering effect of hyaluronan depended on chain length, and shortening the chains reduced the degree of molecular reflection. The latter should reduce the concentration polarization at the tissue interface, and hence the local osmotic pressure opposing fluid drainage. In rheumatoid arthritis the reduced chain length will

  9. Radiation cross-linked collagen/dextran dermal scaffolds: effects of dextran on cross-linking and degradation.

    PubMed

    Zhang, Yaqing; Zhang, Xiangmei; Xu, Ling; Wei, Shicheng; Zhai, Maolin

    2015-01-01

    Ionizing radiation effectively cross-links collagen into network with enhanced anti-degradability and biocompatibility, while radiation-cross-linked collagen scaffold lacks flexibility, satisfactory surface appearance, and performs poor in cell penetration and ingrowth. To make the radiation-cross-linked collagen scaffold to serve as an ideal artificial dermis, dextran was incorporated into collagen. Scaffolds with the collagen/dextran (Col/Dex) ratios of 10/0, 7/3, and 5/5 were fabricated via (60)Co γ-irradiation cross-linking, followed by lyophilization. The morphology, microstructure, physicochemical, and biological properties were investigated. Compared with pure collagen, scaffolds with dextran demonstrated more porous appearance, enhanced hydrophilicity while the cross-linking density was lower with the consequence of larger pore size, higher water uptake, as well as reduced stiffness. Accelerated degradation was observed when dextran was incorporated in both the in vitro and in vivo assays, which led to earlier integration with cell and host tissue. The effect of dextran on degradation was ascribed to the decreased cross-linking density, looser microstructure, more porous and hydrophilic surface. Considering the better appearance, softness, moderate degradation rate due to controllable cross-linking degree and good biocompatibility as well, radiation-cross-linked collagen/dextran scaffolds are expected to serve as promising artificial dermal substitutes.

  10. A cross-linking study of the Ca2+, Mg2+-activated adenosine triphosphatase of Escherichia coli.

    PubMed

    Bragg, P D; Hou, C

    1980-05-01

    The solubilized Ca2+,Mg2+-activated adenosine triphosphatase of Escherichia coli is composed of five subunits designated alpha, beta, gamma, delta and epsilon in order of decreasing molecular weight. The subunit structure of the enzyme has been investigated by the use of the cleavable cross-linking agents dithiobis(succinimidyl propionate), methyl-4-mercaptobutyrimidate, dimethyl-3,3'-dithiobispropionimidate, disuccinimidyl tartarate, and cupric 1,10-phenanthrolinate. The products of cross-linking were analyzed by two different two-dimensional gel electrophoresis systems. The following cross-linked subunit dimers were observed: alpha 2, beta 2, alpha beta, alpha delta, beta gamma, beta delta, beta epsilon and gamma epsilon. These results, together with other published data, are discussed in relation to a model of the arrangement of the subunits in the ATPase molecule.

  11. Cation-induced folding of alginate-bearing bilayer gels: an unusual example of spontaneous folding along the long axis.

    PubMed

    Athas, Jasmin C; Nguyen, Catherine P; Kummar, Shailaa; Raghavan, Srinivasa R

    2018-04-04

    The spontaneous folding of flat gel films into tubes is an interesting example of self-assembly. Typically, a rectangular film folds along its short axis when forming a tube; folding along the long axis has been seen only in rare instances when the film is constrained. Here, we report a case where the same free-swelling gel film folds along either its long or short axis depending on the concentration of a solute. Our gels are sandwiches (bilayers) of two layers: a passive layer of cross-linked N,N'-dimethylyacrylamide (DMAA) and an active layer of cross-linked DMAA that also contains chains of the biopolymer alginate. Multivalent cations like Ca2+ and Cu2+ induce these bilayer gels to fold into tubes. The folding occurs instantly when a flat film of the gel is introduced into a solution of these cations. The likely cause for folding is that the active layer stiffens and shrinks (because the alginate chains in it get cross-linked by the cations) whereas the passive layer is unaffected. The resulting mismatch in swelling degree between the two layers creates internal stresses that drive folding. Cations that are incapable of cross-linking alginate, such as Na+ and Mg2+, do not induce gel folding. Moreover, the striking aspect is the direction of folding. When the Ca2+ concentration is high (100 mM or higher), the gels fold along their long axis, whereas when the Ca2+ concentration is low (40 to 80 mM), the gels fold along their short axis. We hypothesize that the folding axis is dictated by the inhomogeneous nature of alginate-cation cross-linking, i.e., that the edges get cross-linked before the faces of the gel. At high Ca2+ concentration, the stiffer edges constrain the folding; in turn, the gel folds such that the longer edges are deformed less, which explains the folding along the long axis. At low Ca2+ concentration, the edges and the faces of the gel are more similar in their degree of cross-linking; therefore, the gel folds along its short axis. An analogy

  12. Genetically encoded releasable photo-cross-linking strategies for studying protein-protein interactions in living cells.

    PubMed

    Yang, Yi; Song, Haiping; He, Dan; Zhang, Shuai; Dai, Shizhong; Xie, Xiao; Lin, Shixian; Hao, Ziyang; Zheng, Huangtao; Chen, Peng R

    2017-10-01

    Although protein-protein interactions (PPIs) have crucial roles in virtually all cellular processes, the identification of more transient interactions in their biological context remains challenging. Conventional photo-cross-linking strategies can be used to identify transient interactions, but these approaches often suffer from high background due to the cross-linked bait proteins. To solve the problem, we have developed membrane-permeable releasable photo-cross-linkers that allow for prey-bait separation after protein complex isolation and can be installed in proteins of interest (POIs) as unnatural amino acids. Here we describe the procedures for using two releasable photo-cross-linkers, DiZSeK and DiZHSeC, in both living Escherichia coli and mammalian cells. A cleavage after protein photo-cross-linking (CAPP ) strategy based on the photo-cross-linker DiZSeK is described, in which the prey protein pool is released from a POI after affinity purification. Prey proteins are analyzed using mass spectrometry or 2D gel electrophoresis for global comparison of interactomes from different experimental conditions. An in situ cleavage and mass spectrometry (MS)-label transfer after protein photo-cross-linking (IMAPP) strategy based on the photo-cross-linker DiZHSeC is also described. This strategy can be used for the identification of cross-linking sites to allow detailed characterization of PPI interfaces. The procedures for photo-cross-linker incorporation, photo-cross-linking of interaction partners and affinity purification of cross-linked complexes are similar for the two photo-cross-linkers. The final section of the protocol describes prey-bait separation (for CAPP) and MS-label transfer and identification (for IMAPP). After plasmid construction, the CAPP and IMAPP strategies can be completed within 6 and 7 d, respectively.

  13. Modulation of hyaluronan synthase activity in cellular membrane fractions.

    PubMed

    Vigetti, Davide; Genasetti, Anna; Karousou, Evgenia; Viola, Manuela; Clerici, Moira; Bartolini, Barbara; Moretto, Paola; De Luca, Giancarlo; Hascall, Vincent C; Passi, Alberto

    2009-10-30

    Hyaluronan (HA), the only non-sulfated glycosaminoglycan, is involved in morphogenesis, wound healing, inflammation, angiogenesis, and cancer. In mammals, HA is synthesized by three homologous HA synthases, HAS1, HAS2, and HAS3, that polymerize the HA chain using UDP-glucuronic acid and UDP-N-acetylglucosamine as precursors. Since the amount of HA is critical in several pathophysiological conditions, we developed a non-radioactive assay for measuring the activity of HA synthases (HASs) in eukaryotic cells and addressed the question of HAS activity during intracellular protein trafficking. We prepared three cellular fractions: plasma membrane, cytosol (containing membrane proteins mainly from the endoplasmic reticulum and Golgi), and nuclei. After incubation with UDP-sugar precursors, newly synthesized HA was quantified by polyacrylamide gel electrophoresis of fluorophore-labeled saccharides and high performance liquid chromatography. This new method measured HAS activity not only in the plasma membrane fraction but also in the cytosolic membranes. This new technique was used to evaluate the effects of 4-methylumbeliferone, phorbol 12-myristate 13-acetate, interleukin 1beta, platelet-derived growth factor BB, and tunicamycin on HAS activities. We found that HAS activity can be modulated by post-translational modification, such as phosphorylation and N-glycosylation. Interestingly, we detected a significant increase in HAS activity in the cytosolic membrane fraction after tunicamycin treatment. Since this compound is known to induce HA cable structures, this result links HAS activity alteration with the capability of the cell to promote HA cable formation.

  14. Flexible all-solid-state supercapacitors based on graphene/carbon black nanoparticle film electrodes and cross-linked poly(vinyl alcohol)-H2SO4 porous gel electrolytes

    NASA Astrophysics Data System (ADS)

    Fei, Haojie; Yang, Chongyang; Bao, Hua; Wang, Gengchao

    2014-11-01

    Flexible all-solid-state supercapacitors (SCs) are fabricated using graphene/carbon black nanoparticle (GCB) film electrodes and cross-linked poly(vinyl alcohol)-H2SO4 porous gel electrolytes (gPVAP-H2SO4). The GCB composite films, with carbon black (CB) nanoparticles uniformly distributed in the graphene nanosheets, greatly improve the active surface areas and ion transportation of pristine graphene film. The porous structure of as-prepared gPVAP-H2SO4 membrane improves the equilibrium swelling ratio in electrolyte and provides interconnected ion transport channels. The chemical crosslinking solves the fluidity problem of PVA-H2SO4 gel electrolyte at high temperature. As-fabricated GCB//gPVAP(20)-H2SO4//GCB flexible SC displays an increased specific capacitance (144.5 F g-1 at 0.5 A g-1) and a higher specific capacitance retention (67.9% from 0.2 to 4 A g-1). More importantly, the flexible SC possesses good electrochemical performance at high temperature (capacitance retention of 78.3% after 1000 cycles at 70 °C).

  15. Rheological characterization of cataplasm bases composed of cross-linked partially neutralized polyacrylate hydrogel.

    PubMed

    Wang, Jian; Zhang, Hongqin; An, Dianyun; Yu, Jian; Li, Wei; Shen, Teng; Wang, Jianxin

    2014-10-01

    Viscoelasticity is a useful parameter for characterizing the intrinsic properties of the cross-linked polyacrylate hydrogel used in cataplasm bases. The aim of this study was to investigate the effects of various formulation parameters on the rheological characteristics of polyacrylate hydrogel. The hydrogel layers were formed using a partially neutralized polyacrylate (Viscomate(™)), which contained acrylic acid and sodium acrylate in different copolymerization ratios, as the cross-linked gel framework. Dihydroxyaluminum aminoacetate (DAAA), which produces aluminum ions, was used as the cross-linking agent. Rheological analyses were performed using a "stress amplitude sweep" and a "frequency sweep". The results showed that greater amounts of acrylic acid in the structure of Viscomate as well as higher concentrations of DAAA and Viscomate led to an increase in the elastic modulus (G'). However, greater amounts of acrylic acid in the structure of Viscomate and higher concentrations of DAAA had an opposite on the viscous modulus (G″); this might be owing to higher steric hindrance. The results of this study can serve as guidelines for the optimization of formulations for cataplasms.

  16. A rapid increase in macrophage-derived versican and hyaluronan in infectious lung disease.

    PubMed

    Chang, Mary Y; Tanino, Yoshinori; Vidova, Veronika; Kinsella, Michael G; Chan, Christina K; Johnson, Pamela Y; Wight, Thomas N; Frevert, Charles W

    2014-02-01

    The goals of this study were to characterize the changes in chondroitin sulfate proteoglycans and hyaluronan in lungs in acute response to gram-negative bacterial infection and to identify cellular components responsible for these changes. Mice were treated with intratracheal (IT) live Escherichia coli, E. coli lipopolysaccharide (LPS), or PBS. Both E. coli and LPS caused rapid selective increases in mRNA expression of versican and hyaluronan synthase (Has) isoforms 1 and 2 associated with increased immunohistochemical and histochemical staining for versican and hyaluronan in the lungs. Versican was associated with a subset of alveolar macrophages. To examine whether macrophages contribute to versican and hyaluronan accumulation, in vitro studies with primary cultures of bone marrow-derived and alveolar macrophages were performed. Unstimulated macrophages expressed very low levels of versican and hyaluronan synthase mRNA, with no detectible versican protein or hyaluronan product. Stimulation with LPS caused rapid increases in versican mRNA and protein, a rapid increase in Has1 mRNA, and concomitant inhibition of hyaluronidases 1 and 2, the major hyaluronan degrading enzymes. Hyaluronan could be detected following chloroquine pre-treatment, indicating rapid turnover and degradation of hyaluronan by macrophages. In addition, the effects of LPS, the M1 macrophage classical activation agonist, were compared to those of IL-4/IL-13 or IL-10, the M2a and M2c alternative activation agonists, respectively. Versican and Has1 increased only in response to M1 activation. Finally, the up-regulation of versican and Has1 in the whole lungs of wild-type mice following IT LPS was completely abrogated in TLR-4(-/-) mice. These findings suggest that versican and hyaluronan synthesis may play an important role in the innate immune response to gram-negative lung infection. Copyright © 2014 International Society of Matrix Biology. Published by Elsevier B.V. All rights reserved.

  17. Wood Sawdust/Natural Rubber Ecocomposites Cross-Linked by Electron Beam Irradiation

    PubMed Central

    Manaila, Elena; Stelescu, Maria Daniela; Craciun, Gabriela; Ighigeanu, Daniel

    2016-01-01

    The obtaining and characterization of some polymeric eco-composites based on wood sawdust and natural rubber is presented. The natural rubber was cross-linked using the electron beam irradiation. The irradiation doses were of 75, 150, 300 and 600 kGy and the concentrations of wood sawdust were of 10 and 20 phr, respectively. As a result of wood sawdust adding, the physical and mechanical properties such as hardness, modulus at 100% elongation and tensile strength, showed significant improvements. The presence of wood sawdust fibers has a reinforcing effect on natural rubber, similar or better than of mineral fillers. An increase in the irradiation dose leads to the increasing of cross-link density, which is reflected in the improvement of hardness, modulus at 100% elongation and tensile strength of blends. The cross-linking rates, appreciated using the Flory-Rehner equation, have increased with the amount of wood sawdust in blends and with the irradiation dose. Even if the gel fraction values have varied irregularly with the amount of wood sawdust and irradiation dose it was over 90% for all blends, except for the samples without wood sawdust irradiated with 75 kGy. The water uptake increased with increasing of fiber content and decreased with the irradiation dose. PMID:28773626

  18. Wood Sawdust/Natural Rubber Ecocomposites Cross-Linked by Electron Beam Irradiation.

    PubMed

    Manaila, Elena; Stelescu, Maria Daniela; Craciun, Gabriela; Ighigeanu, Daniel

    2016-06-23

    The obtaining and characterization of some polymeric eco-composites based on wood sawdust and natural rubber is presented. The natural rubber was cross-linked using the electron beam irradiation. The irradiation doses were of 75, 150, 300 and 600 kGy and the concentrations of wood sawdust were of 10 and 20 phr, respectively. As a result of wood sawdust adding, the physical and mechanical properties such as hardness, modulus at 100% elongation and tensile strength, showed significant improvements. The presence of wood sawdust fibers has a reinforcing effect on natural rubber, similar or better than of mineral fillers. An increase in the irradiation dose leads to the increasing of cross-link density, which is reflected in the improvement of hardness, modulus at 100% elongation and tensile strength of blends. The cross-linking rates, appreciated using the Flory-Rehner equation, have increased with the amount of wood sawdust in blends and with the irradiation dose. Even if the gel fraction values have varied irregularly with the amount of wood sawdust and irradiation dose it was over 90% for all blends, except for the samples without wood sawdust irradiated with 75 kGy. The water uptake increased with increasing of fiber content and decreased with the irradiation dose.

  19. Di-isodityrosine is the intermolecular cross-link of isodityrosine-rich extensin analogs cross-linked in vitro.

    PubMed

    Held, Michael A; Tan, Li; Kamyab, Abdolreza; Hare, Michael; Shpak, Elena; Kieliszewski, Marcia J

    2004-12-31

    Extensins are cell wall hydroxyproline-rich glycoproteins that form covalent networks putatively involving tyrosyl and lysyl residues in cross-links catalyzed by one or more extensin peroxidases. The precise cross-links remain to be chemically identified both as network components in muro and as enzymic products generated in vitro with native extensin monomers as substrates. However, some extensin monomers contain variations within their putative cross-linking motifs that complicate cross-link identification. Other simpler extensins are recalcitrant to isolation including the ubiquitous P3-type extensin whose major repetitive motif, Hyp)(4)-Ser-Hyp-Ser-(Hyp)(4)-Tyr-Tyr-Tyr-Lys, is of particular interest, not least because its Tyr-Tyr-Tyr intramolecular isodityrosine cross-link motifs are also putative candidates for further intermolecular cross-linking to form di-isodityrosine. Therefore, we designed a set of extensin analogs encoding tandem repeats of the P3 motif, including Tyr --> Phe and Lys --> Leu variations. Expression of these P3 analogs in Nicotiana tabacum cells yielded glycoproteins with virtually all Pro residues hydroxylated and subsequently arabinosylated and with likely galactosylated Ser residues. This was consistent with earlier analyses of P3 glycopeptides isolated from cell wall digests and the predictions of the Hyp contiguity hypothesis. The tyrosine-rich P3 analogs also contained isodityrosine, formed in vivo. Significantly, these isodityrosine-containing analogs were further cross-linked in vitro by an extensin peroxidase to form the tetra-tyrosine intermolecular cross-link amino acid di-isodityrosine. This is the first identification of an inter-molecular cross-link amino acid in an extensin module and corroborates earlier suggestions that di-isodityrosine represents one mechanism for cross-linking extensins in muro.

  20. Viscoelasticity of rabbit vocal folds after injection augmentation.

    PubMed

    Dahlqvist, Ake; Gärskog, Ola; Laurent, Claude; Hertegård, Stellan; Ambrosio, Luigi; Borzacchiello, Assunta

    2004-01-01

    Vocal fold function is related to the viscoelasticity of the vocal fold tissue. Augmentation substances used for injection treatment of voice insufficiency may alter the viscoelastic properties of vocal folds and their vibratory capacity. The objective was to compare the mechanical properties (viscoelasticity) of various injectable substances and the viscoelasticity of rabbit vocal folds, 6 months after injection with one of these substances. Animal model. Cross-linked collagen (Zyplast), double cross-linked hyaluronan (hylan B gel), dextranomers in hyaluronan (DHIA), and polytetrafluoroethylene (Teflon) were injected into rabbit vocal folds. Six months after the injection, the animals were killed and the right- and left-side vocal folds were removed. Dynamic viscosity of the injected substances and the vocal folds was measured with a Bohlin parallel-plate rheometer during small-amplitude oscillation. All injected vocal folds showed a decreasing dynamic viscosity with increasing frequency. Hylan B gel and DiHA showed the lowest dynamic viscosity values, and vocal folds injected with these substances also showed the lowest dynamic viscosity (similar to noninjected control samples). Teflon (and vocal folds injected with Teflon) showed the highest dynamic viscosity values, followed by the collagen samples. Substances with low viscoelasticity alter the mechanical properties of the vocal fold to a lesser degree than substances with a high viscoelasticity. The data indicated that hylan B gel and DiHA render the most natural viscoelastic properties to the vocal folds. These substances seem to be appropriate for preserving or restoring the vibratory capacity of the vocal folds when glottal insufficiency is treated with augmentative injections.

  1. On-line separation and characterization of hyaluronan oligosaccharides derived from radical depolymerization

    PubMed Central

    Zhao, Xue; Yang, Bo; Li, Lingyun; Zhang, Fuming; Linhardt, Robert J.

    2013-01-01

    Hydroxyl radicals are widely implicated in the oxidation of carbohydrates in biological and industrial processes and are often responsible for their structural modification resulting in functional damage. In this study, the radical depolymerization of the polysaccharide hyaluronan was studied in a reaction with hydroxyl radicals generated by Fenton Chemistry. A simple method for isolation and identification of the resulting non-sulfated oligosaccharide products of oxidative depolymerization was established. Hyaluronan oligosaccharides were analyzed using ion-pairing reversed phase high performance liquid chromotography coupled with tandem electrospray mass spectrometry. The sequence of saturated hyaluronan oligosaccharides having even- and odd-numbers of saccharide units, afforded through oxidative depolymerization, were identified. This study represents a simple, effective ‘fingerprinting’ protocol for detecting the damage done to hyaluronan by oxidative radicals. This study should help reveal the potential biological outcome of reactive-oxygen radical-mediated depolymerization of hyaluronan. PMID:23768593

  2. Hypotheses on the evolution of hyaluronan: A highly ironic acid

    PubMed Central

    Csoka, Antonei B; Stern, Robert

    2013-01-01

    Hyaluronan is a high-molecular-weight glycosaminoglycan (GAG) prominent in the extracellular matrix. Emerging relatively late in evolution, it may have evolved to evade immune recognition. Chondroitin is a more ancient GAG and a possible hyaluronan precursor. Epimerization of a 4-hydroxyl in N-acetylgalactosamine in chondroitin to N-acetylglucosamine of hyaluronan is the only structural difference other than chain length between these two polymers. The axial 4-hydroxyl group extends out perpendicular from the equatorial plane of N-acetylgalactosamine in chondroitin. We suspect that this hydroxyl is a prime target for immune recognition. Conversion of a thumbs-up hydroxyl group into a thumbs-down position in the plane of the sugar endows hyaluronan with the ability to avoid immune recognition. Chitin is another potential precursor to hyaluronan. But regardless whether of chondroitin or of chitin origin, an ancient chondroitinase enzyme sequence seems to have been commandeered to catalyze the cleavage of the new hyaluronan substrate. The evolution of six hyaluronidase-like sequences in the human genome from a single chondroitinase as found in Caenorhabditis elegans can now be traced. Confirming our previous predictions, two duplication events occurred, with three hyaluronidase-like sequences occurring in the genome of Ciona intestinalis (sea squirt), the earliest known chordate. This was probably followed by en masse duplication, with six such genes present in the genome of zebra fish onwards. These events occurred, however, much earlier than predicted. It is also apparent on an evolutionary time scale that in several species, this gene family is continuing to evolve. PMID:23315448

  3. Enzymatically cross-linked injectable alginate-g-pyrrole hydrogels for neovascularization.

    PubMed

    Devolder, Ross; Antoniadou, Eleni; Kong, Hyunjoon

    2013-11-28

    Microparticles capable of releasing protein drugs are often incorporated into injectable hydrogels to minimize their displacement at an implantation site, reduce initial drug burst, and further control drug release rates over a broader range. However, there is still a need to develop methods for releasing drug molecules over extended periods of time, in order to sustain the bioactivity of drug molecules at an implantation site. In this study, we hypothesized that a hydrogel formed through the cross-linking of pyrrole units linked to a hydrophilic polymer would release protein drugs in a more sustained manner, because of an enhanced association between cross-linked pyrrole groups and the drug molecules. To examine this hypothesis, we prepared hydrogels of alginate substituted with pyrrole groups, alginate-g-pyrrole, through a horse-radish peroxidase (HRP)-activated cross-linking of the pyrrole groups. The hydrogels were encapsulated with poly(lactic-co-glycolic acid) (PLGA) microparticles loaded with vascular endothelial growth factor (VEGF). The resulting hydrogel system released VEGF in a more sustained manner than Ca(2+) alginate or Ca(2+) alginate-g-pyrrole gel systems. Finally, implantations of the VEGF-releasing HRP-activated alginate-g-pyrrole hydrogel system on chicken chorioallantoic membranes resulted in the formation of blood vessels in higher densities and with larger diameters, compared to other control conditions. Overall, the drug releasing system developed in this study will be broadly useful for regulating release rates of a wide array of protein drugs, and further enhance the quality of protein drug-based therapies. © 2013 Elsevier B.V. All rights reserved.

  4. Quantification and characterization of enzymatically produced hyaluronan with fluorophore-assisted carbohydrate electrophoresis.

    PubMed

    Kooy, Floor K; Ma, Muyuan; Beeftink, Hendrik H; Eggink, Gerrit; Tramper, Johannes; Boeriu, Carmen G

    2009-01-15

    Hyaluronan (HA) is a polysaccharide with high-potential medical applications, depending on the chain length and the chain length distribution. Special interest goes to homogeneous HA oligosaccharides, which can be enzymatically produced using Pasteurella multocida hyaluronan synthase (PmHAS). We have developed a sensitive, simple, and fast method, based on fluorophore-assisted carbohydrate electrophoresis (FACE), for characterization and quantification of polymerization products. A chromatographic pure fluorescent template was synthesized from HA tetrasaccharide (HA4) and 2-aminobenzoic acid. HA4-fluor and HA4 were used as template for PmHAS-mediated polymerization of nucleotide sugars. All products, fluorescent and nonfluorescent, were analyzed with gel electrophoresis and quantified using lane densitometry. Comparison of HA4- and HA4-fluor-derived polymers showed that the fluorophore did not negatively influence the PmHAS-mediated polymerization. Only even-numbered oligosaccharide products were observed using HA4-fluor or HA4 as template. The fluorophore intensity was linearly related to its concentration, and the limit of detection was determined to be 7.4pmol per product band. With this assay, we can now differentiate oligosaccharides of size range DP2 (degree of polymerization 2) to approximately DP400, monitor the progress of polymerization reactions, and measure subtle differences in polymerization rate. Quantifying polymerization products enables us to study the influence of experimental conditions on HA synthesis.

  5. In vitro effects of platelet-rich gel supernatants on histology and chondrocyte apoptosis scores, hyaluronan release and gene expression of equine cartilage explants challenged with lipopolysaccharide.

    PubMed

    Carmona, Jorge U; Ríos, Diana L; López, Catalina; Álvarez, María E; Pérez, Jorge E; Bohórquez, Mabel E

    2016-07-01

    Platelet-rich plasma (PRP) preparations are a common treatment in equine osteoarthritis (OA). However, there are controversies regarding the ideal concentration of platelets and leukocytes in these biological substances necessary to induce an adequate anti-inflammatory and anabolic response in articular cartilage. The aims were to study the influence of leukocyte- and platelet-rich gel (L-PRG) and pure platelet-rich gel (P-PRG) supernatants on the histological changes of cartilage, the degree of chondrocyte apoptosis, the production of hyaluronan (HA) and the gene expression of nuclear factor kappa beta (NFkβ), matrix metalloproteinase 13 (MMP-13), a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS-4), collagen type I alpha 1 (COL1A1), collagen type II alpha 1 (COL2A1) and cartilage oligomeric matrix protein (COMP) in normal cartilage explants (CEs) challenged with lipopolysaccharide (LPS). Overall, 25 % L-PRG supernatant (followed in order of importance by, 50 % P-PRG, 25 % P-PRG and 50 % L-PRG) represented the substance with the most important anti-inflammatory and anabolic effect. 25 % P-PRG supernatant presented important anabolic effects, but it induced a more severe chondrocyte apoptosis than the other evaluated substances. 25 % L-PRG supernatant presented the best therapeutic profile. Our results demonstrate that the biological variability of PRP preparations makes their application rather challenging. Additional in vivo research is necessary to know the effect of PRP preparations at different concentrations.

  6. Inhibition of Ovarian Cancer Chemoresistance and Metastasis with Antagonists of Hyaluronan-CD44-CD147 Interactions

    DTIC Science & Technology

    2013-07-01

    oligosaccharides , which have previously been shown to antagonize hyaluronan-receptor interactions, sensitize cisplatin-resistant human ovarian...carcinoma cells to cisplatin treatment in a mouse xenograft model, as well as in cell culture. Hyaluronan oligosaccharide -decorated nanoparticles... oligosaccharides antagonize hyaluronan-receptor signaling by disrupting constitutive hyaluronan polymer- receptor interaction, thus leading to

  7. Superabsorbing gel for actinide, lanthanide, and fission product decontamination

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Kaminski, Michael D.; Mertz, Carol J.

    The present invention provides an aqueous gel composition for removing actinide ions, lanthanide ions, fission product ions, or a combination thereof from a porous surface contaminated therewith. The composition comprises a polymer mixture comprising a gel forming cross-linked polymer and a linear polymer. The linear polymer is present at a concentration that is less than the concentration of the cross-linked polymer. The polymer mixture is at least about 95% hydrated with an aqueous solution comprising about 0.1 to about 3 percent by weight (wt %) of a multi-dentate organic acid chelating agent, and about 0.02 to about 0.6 molar (M)more » carbonate salt, to form a gel. When applied to a porous surface contaminated with actinide ions, lanthanide ions, and/or other fission product ions, the aqueous gel absorbs contaminating ions from the surface.« less

  8. TMEM2: A missing link in hyaluronan catabolism identified?

    PubMed

    Yamaguchi, Yu; Yamamoto, Hayato; Tobisawa, Yuki; Irie, Fumitoshi

    2018-03-27

    Hyaluronan (HA) is a glycosaminoglycan (GAG) composed of repeating disaccharide units of glucuronic acid and N-acetylglucosamine. HA is an extremely long, unbranched polymer, which often exceeds 10 6  Da and sometimes reaches 10 7  Da. A feature that epitomizes HA is its rapid turnover; one-third of the total body HA is turned over daily. The current model of HA catabolism postulates that high-molecular weight HA in the extracellular space is first cleaved into smaller fragments by a hyaluronidase(s) that resides at the cell surface, followed by internalization of fragments and their degradation into monosaccharides in lysosomes. Over the last decade, considerable research has shown that the HYAL family of hyaluronidases plays significant roles in HA catabolism. Nonetheless, the identity of a hyaluronidase responsible for the initial step of HA cleavage on the cell surface remains elusive, as biochemical and enzymological properties of HYAL proteins are not entirely consistent with those expected of cell surface hyaluronidases. Recent identification of transmembrane 2 (TMEM2) as a cell surface protein that possesses potent hyaluronidase activity suggests that it may be the "missing" cell surface hyaluronidase, and that novel models of HA catabolism should include this protein. Copyright © 2017 International Society of Matrix Biology. Published by Elsevier B.V. All rights reserved.

  9. The Influence of Chitosan Cross-linking on the Properties of Alginate Microparticles with Metformin Hydrochloride-In Vitro and In Vivo Evaluation.

    PubMed

    Szekalska, Marta; Sosnowska, Katarzyna; Zakrzeska, Agnieszka; Kasacka, Irena; Lewandowska, Alicja; Winnicka, Katarzyna

    2017-01-22

    Sodium alginate is a polymer with unique ability to gel with different cross-linking agents in result of ionic and electrostatic interactions. Chitosan cross-linked alginate provides improvement of swelling and mucoadhesive properties and might be used to design sustained release dosage forms. Therefore, the aim of this research was to develop and evaluate possibility of preparing chitosan cross-linked alginate microparticles containing metformin hydrochloride by the spray-drying method. In addition, influence of cross-linking agent on the properties of microparticles was evaluated. Formulation of microparticles prepared by the spray drying of 2% alginate solution cross-linked by 0.1% chitosan was characterized by good mucoadhesive properties, high drug loading and prolonged metformin hydrochloride release. It was shown that designed microparticles reduced rat glucose blood level, delayed absorption of metformin hydrochloride and provided stable plasma drug concentration. Additionally, histopathological studies of pancreas, liver and kidneys indicated that all prepared microparticles improved degenerative changes in organs of diabetic rats. Moreover, no toxicity effect and no changes in rats behavior after oral administration of chitosan cross-linked alginate microparticles were noted.

  10. Provisional matrix: A role for versican and hyaluronan.

    PubMed

    Wight, Thomas N

    2017-07-01

    Hyaluronan and versican are extracellular matrix (ECM) components that are enriched in the provisional matrices that form during the early stages of development and disease. These two molecules interact to create pericellular "coats" and "open space" that facilitate cell sorting, proliferation, migration, and survival. Such complexes also impact the recruitment of leukocytes during development and in the early stages of disease. Once thought to be inert components of the ECM that help hold cells together, it is now quite clear that they play important roles in controlling cell phenotype, shaping tissue response to injury and maintaining tissue homeostasis. Conversion of hyaluronan-/versican-enriched provisional matrix to collagen-rich matrix is a "hallmark" of tissue fibrosis. Targeting the hyaluronan and versican content of provisional matrices in a variety of diseases including, cardiovascular disease and cancer, is becoming an attractive strategy for intervention. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Preliminary evaluation of a gel tube agglutination major cross-match method in dogs.

    PubMed

    Villarnovo, Dania; Burton, Shelley A; Horney, Barbara S; MacKenzie, Allan L; Vanderstichel, Raphaël

    2016-09-01

    A major cross-match gel tube test is available for use in dogs yet has not been clinically evaluated. This study compared cross-match results obtained using the gel tube and the standard tube methods for canine samples. Study 1 included 107 canine sample donor-recipient pairings cross-match tested with the RapidVet-H method gel tube test and compared results with the standard tube method. Additionally, 120 pairings using pooled sera containing anti-canine erythrocyte antibody at various concentrations were tested with leftover blood from a hospital population to assess sensitivity and specificity of the gel tube method in comparison with the standard method. The gel tube method had a good relative specificity of 96.1% in detecting lack of agglutination (compatibility) compared to the standard tube method. Agreement between the 2 methods was moderate. Nine of 107 pairings showed agglutination/incompatibility on either test, too few to allow reliable calculation of relative sensitivity. Fifty percent of the gel tube method results were difficult to interpret due to sample spreading in the reaction and/or negative control tubes. The RapidVet-H method agreed with the standard cross-match method on compatible samples, but detected incompatibility in some sample pairs that were compatible with the standard method. Evaluation using larger numbers of incompatible pairings is needed to assess diagnostic utility. The gel tube method results were difficult to categorize due to sample spreading. Weak agglutination reactions or other factors such as centrifuge model may be responsible. © 2016 American Society for Veterinary Clinical Pathology.

  12. Cross-Link Guided Molecular Modeling with ROSETTA

    PubMed Central

    Leitner, Alexander; Rosenberger, George; Aebersold, Ruedi; Malmström, Lars

    2013-01-01

    Chemical cross-links identified by mass spectrometry generate distance restraints that reveal low-resolution structural information on proteins and protein complexes. The technology to reliably generate such data has become mature and robust enough to shift the focus to the question of how these distance restraints can be best integrated into molecular modeling calculations. Here, we introduce three workflows for incorporating distance restraints generated by chemical cross-linking and mass spectrometry into ROSETTA protocols for comparative and de novo modeling and protein-protein docking. We demonstrate that the cross-link validation and visualization software Xwalk facilitates successful cross-link data integration. Besides the protocols we introduce XLdb, a database of chemical cross-links from 14 different publications with 506 intra-protein and 62 inter-protein cross-links, where each cross-link can be mapped on an experimental structure from the Protein Data Bank. Finally, we demonstrate on a protein-protein docking reference data set the impact of virtual cross-links on protein docking calculations and show that an inter-protein cross-link can reduce on average the RMSD of a docking prediction by 5.0 Å. The methods and results presented here provide guidelines for the effective integration of chemical cross-link data in molecular modeling calculations and should advance the structural analysis of particularly large and transient protein complexes via hybrid structural biology methods. PMID:24069194

  13. Analysis of glycation induced protein cross-linking inhibitory effects of some antidiabetic plants and spices.

    PubMed

    Perera, Handunge Kumudu Irani; Handuwalage, Charith Sandaruwan

    2015-06-09

    Protein cross-linking which occurs towards the latter part of protein glycation is implicated in the development of chronic diabetic complications. Glycation induced protein cross-linking inhibitory effects of nine antidiabetic plants and three spices were evaluated in this study using a novel, simple, electrophoresis based method. Methanol extracts of thirteen plants including nine antidiabetic plants and three spices were used. Lysozyme and fructose were incubated at 37 °C in the presence or absence of different concentrations of plant extracts up to 31 days. Standard glycation inhibitor aminoguanidine and other appropriate controls were included. A recently established sodium dodecyl polyacrylamide gel electrophoresis (SDS-PAGE) method was used to detect the products of protein cross-linking in the incubation mixtures. High molecular weight protein products representing the dimer, trimer and tetramer of lysozyme were detected in the presence of fructose. Among the nine antidiabetic plants, seven showed glycation induced protein cross-linking inhibitory effects namely Ficus racemosa (FR) stem bark, Gymnema sylvestre (GS) leaves, Musa paradisiaca (MP) yam, Phyllanthus debilis (PD) whole plant, Phyllanthus emblica (PE) fruit, Pterocarpus marsupium (PM) latex and Tinospora cordifolia (TC) leaves. Inhibition observed with Coccinia grandis (CG) leaves and Strychnos potatorum (SP) seeds were much low. Leaves of Gymnema lactiferum (GL), the plant without known antidiabetic effects showed the lowest inhibition. All three spices namely Coriandrum sativum (CS) seeds, Cinnamomum zeylanicum (CZ) bark and Syzygium aromaticum (SA) flower buds showed cross-link inhibitory effects with higher effects in CS and SA. PD, PE, PM, CS and SA showed almost complete inhibition on the formation of cross-linking with 25 μg/ml extracts. Methanol extracts of PD, PE, PM, CS and SA have shown promising inhibitory effects on glycation induced protein cross-linking.

  14. Regulation of Hyaluronan (HA) Metabolism Mediated by HYBID (Hyaluronan-binding Protein Involved in HA Depolymerization, KIAA1199) and HA Synthases in Growth Factor-stimulated Fibroblasts.

    PubMed

    Nagaoka, Aya; Yoshida, Hiroyuki; Nakamura, Sachiko; Morikawa, Tomohiko; Kawabata, Keigo; Kobayashi, Masaki; Sakai, Shingo; Takahashi, Yoshito; Okada, Yasunori; Inoue, Shintaro

    2015-12-25

    Regulation of hyaluronan (HA) synthesis and degradation is essential to maintenance of extracellular matrix homeostasis. We recently reported that HYBID (HYaluronan-Binding protein Involved in hyaluronan Depolymerization), also called KIAA1199, plays a key role in HA depolymerization in skin and arthritic synovial fibroblasts. However, regulation of HA metabolism mediated by HYBID and HA synthases (HASs) under stimulation with growth factors remains obscure. Here we report that TGF-β1, basic FGF, EGF, and PDGF-BB commonly enhance total amount of HA in skin fibroblasts through up-regulation of HAS expression, but molecular size of newly produced HA is dependent on HYBID expression levels. Stimulation of HAS1/2 expression and suppression of HYBID expression by TGF-β1 were abrogated by blockade of the MAPK and/or Smad signaling and the PI3K-Akt signaling, respectively. In normal human skin, expression of the TGF-β1 receptors correlated positively with HAS2 expression and inversely with HYBID expression. On the other hand, TGF-β1 up-regulated HAS1/2 expression but exerted only a slight suppressive effect on HYBID expression in synovial fibroblasts from the patients with osteoarthritis or rheumatoid arthritis, resulting in the production of lower molecular weight HA compared with normal skin and synovial fibroblasts. These data demonstrate that although TGF-β1, basic FGF, EGF, and PDGF-BB enhance HA production in skin fibroblasts, TGF-β1 most efficiently contributes to production of high molecular weight HA by HAS up-regulation and HYBID down-regulation and suggests that inefficient down-regulation of HYBID by TGF-β1 in arthritic synovial fibroblasts may be linked to accumulation of depolymerized HA in synovial fluids in arthritis patients. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  15. Controlled Immobilization Strategies to Probe Short Hyaluronan-Protein Interactions

    NASA Astrophysics Data System (ADS)

    Minsky, Burcu Baykal; Antoni, Christiane H.; Boehm, Heike

    2016-02-01

    Well-controlled grafting of small hyaluronan oligosaccharides (sHA) enables novel approaches to investigate biological processes such as angiogenesis, immune reactions and cancer metastasis. We develop two strategies for covalent attachment of sHA, a fast high-density adsorption and a two-layer system that allows tuning the density and mode of immobilization. We monitored the sHA adlayer formation and subsequent macromolecular interactions by label-free quartz crystal microbalance with dissipation (QCM-D). The modified surfaces are inert to unspecific protein adsorption, and yet retain the specific binding capacity of sHA. Thus they are an ideal tool to study the interactions of hyaluronan-binding proteins and short hyaluronan molecules as demonstrated by the specific recognition of LYVE-1 and aggrecan. Both hyaladherins recognize sHA and the binding is independent to the presence of the reducing end.

  16. Controlling adsorption of albumin with hyaluronan on silica surfaces and sulfonated latex particles.

    PubMed

    Berts, Ida; Fragneto, Giovanna; Porcar, Lionel; Hellsing, Maja S; Rennie, Adrian R

    2017-10-15

    Polysaccharides are known to modify binding of proteins at interfaces and this paper describes studies of these interactions and how they are modified by pH. Specifically, the adsorption of human serum albumin on to polystyrene latex and to silica is described, focusing on how this is affected by hyaluronan. Experiments were designed to test how such binding might be modified under relevant physiological conditions. Changes in adsorption of albumin alone and the co-adsorption of albumin and hyaluronan are driven by electrostatic interactions. Multilayer binding is found to be regulated by the pH of the solution and the molecular mass and concentration of hyaluronan. Highest adsorption was observed at pH below 4.8 and for low molecular mass hyaluronan (≤150kDa) at concentrations above 2mgml -1 . On silica with grafted hyaluronan, albumin absorption is reversed by changes in solvent pH due to their strong electrostatic attraction. Albumin physisorbed on silica surfaces is also rinsed away with dilute hyaluronan solution at pH 4.8. The results demonstrate that the protein adsorption can be controlled both by changes of pH and by interaction with other biological macromolecules. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. Quantitative evaluation of protein conformation in pharmaceuticals using cross-linking reactions coupled with LC-MS/MS analysis.

    PubMed

    Yamaguchi, Hideto; Hirakura, Yutaka; Shirai, Hiroki; Mimura, Hisashi; Toyo'oka, Toshimasa

    2011-06-01

    The need for a simple and high-throughput method for identifying the tertiary structure of protein pharmaceuticals has increased. In this study, a simple method for mapping the protein fold is proposed for use as a complementary quality test. This method is based on cross-linking a protein using a [bis(sulfosuccinimidyl)suberate (BS(3))], followed by peptide mapping by LC-MS. Consensus interferon (CIFN) was used as the model protein. The tryptic map obtained via liquid chromatography tandem mass spectroscopy (LC-MS/MS) and the mass mapping obtained via matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy were used to identify cross-linked peptides. While LC-MS/MS analyses found that BS(3) formed cross-links in the loop region of the protein, which was regarded as the biologically active site, sodium dodecyl-sulfate polyacrylamide gel electrophoresis demonstrated that cross-linking occurred within a protein molecule, but not between protein molecules. The occurrence of cross-links at the active site depends greatly on the conformation of the protein, which is determined by the denaturing conditions. Quantitative evaluation of the tertiary structure of CIFN was thus possible by monitoring the amounts of cross-linked peptides generated. Assuming that background information is available at the development stage, this method may be applicable to process development as a complementary test for quality control. Copyright © 2011 Elsevier B.V. All rights reserved.

  18. Effect of depletion of interstitial hyaluronan on hydraulic conductance in rabbit knee synovium

    PubMed Central

    Coleman, P J; Scott, D; Abiona, A; Ashhurst, D E; Mason, R M; Levick, J R

    1998-01-01

    The hydraulic resistance of the synovial lining to fluid outflow from a joint cavity () is important for the retention of intra-articular lubricant. The resistance has been attributed in part to extracellular glycosaminoglycans, including hyaluronan and chondroitin sulphates. Increased permeability in joints infused with testicular hyaluronidase, which digests both chondroitin sulphates and hyaluronan, supports this view. In this study the importance of interstitial hyaluronan per se was assessed using leech and Streptomyces hyaluronidases, which degrade only hyaluronan. Ringer solution was infused into the knee joint cavity of anaesthetized rabbits for 30 min, with or without hyaluronidase, after which intra-articular pressure (Pj) was raised and the relation between pressure and outflow determined. Treatment with Streptomyces, leech or testicular hyaluronidases increased the fluid escape rates by similar factors, namely 4- to 6-fold. After Streptomyces hyaluronidase treatment the slope d/dPj, which at low pressures represents synovial hydraulic conductance, increased from a control of 0.90 ± 0.20 μl min−1 cmH2O−1 (mean ± s.e.m., n = 6) to 4.52 ± 0.70 μl min−1 cmH2O−1. The slope d/dPj increased to a similar level after testicular hyaluronidase, namely to 4.14 ± 1.06 μl min−1 cmH2O−1 (control, 0.54 ± 0.24 μl min−1 cmH2O−1). Streptomyces and leech hyaluronidases were as effective as testicular hyaluronidase (no statistically significant differences) despite differences in substrate specificity. It was shown using histochemical and immunohistochemical techniques that hyaluronan was removed from the synovium by leech, Streptomyces and testicular hyaluronidases. The binding of antibodies 2-B-6 and 3-B-3 showed that the core proteins of the chondroitin sulphate proteoglycans remained intact after treatment with hyaluronidases, and the binding of 5-D-4 showed that keratan sulphate was unaffected. An azocasein digestion assay confirmed that the

  19. Determining equilibrium osmolarity in poly(ethylene glycol)/chondrotin sulfate gels mimicking articular cartilage.

    PubMed

    Sircar, S; Aisenbrey, E; Bryant, S J; Bortz, D M

    2015-01-07

    We present an experimentally guided, multi-phase, multi-species polyelectrolyte gel model to make qualitative predictions on the equilibrium electro-chemical properties of articular cartilage. The mixture theory consists of two different types of polymers: poly(ethylene gylcol) (PEG), chondrotin sulfate (ChS), water (acting as solvent) and several different ions: H(+), Na(+), Cl(-). The polymer chains have covalent cross-links whose effect on the swelling kinetics is modeled via Doi rubber elasticity theory. Numerical studies on equilibrium polymer volume fraction and net osmolarity (difference in the solute concentration across the gel) show a complex interplay between ionic bath concentrations, pH, cross-link fraction and the average charge per monomer. Generally speaking, swelling is aided due to a higher average charge per monomer (or a higher particle fraction of ChS, the charged component of the polymer), low solute concentration in the bath, a high pH or a low cross-link fraction. A peculiar case arises at higher values of cross-link fraction, where it is observed that increasing the average charge per monomer leads to gel deswelling. Copyright © 2014 Elsevier Ltd. All rights reserved.

  20. A RHAMM Mimetic Peptide Blocks Hyaluronan Signaling and Reduces Inflammation and Fibrogenesis in Excisional Skin Wounds

    PubMed Central

    Tolg, Cornelia; Hamilton, Sara R.; Zalinska, Ewa; McCulloch, Lori; Amin, Ripal; Akentieva, Natalia; Winnik, Francoise; Savani, Rashmin; Bagli, Darius J.; Luyt, Len G.; Cowman, Mary K.; McCarthy, Jim B.; Turley, Eva A.

    2013-01-01

    Hyaluronan is activated by fragmentation and controls inflammation and fibroplasia during wound repair and diseases (eg, cancer). Hyaluronan-binding peptides were identified that modify fibrogenesis during skin wound repair. Peptides were selected from 7- to 15mer phage display libraries by panning with hyaluronan-Sepharose beads and assayed for their ability to block fibroblast migration in response to hyaluronan oligosaccharides (10 kDa). A 15mer peptide (P15-1), with homology to receptor for hyaluronan mediated motility (RHAMM) hyaluronan binding sequences, was the most effective inhibitor. P15-1 bound to 10-kDa hyaluronan with an affinity of Kd = 10−7 and appeared to specifically mimic RHAMM since it significantly reduced binding of hyaluronan oligosaccharides to recombinant RHAMM but not to recombinant CD44 or TLR2,4, and altered wound repair in wild-type but not RHAMM−/− mice. One topical application of P15-1 to full-thickness excisional rat wounds significantly reduced wound macrophage number, fibroblast number, and blood vessel density compared to scrambled, negative control peptides. Wound collagen 1, transforming growth factor β-1, and α-smooth muscle actin were reduced, whereas tenascin C was increased, suggesting that P15-1 promoted a form of scarless healing. Signaling/microarray analyses showed that P15-1 blocks RHAMM-regulated focal adhesion kinase pathways in fibroblasts. These results identify a new class of reagents that attenuate proinflammatory, fibrotic repair by blocking hyaluronan oligosaccharide signaling. PMID:22889846

  1. Hyaluronan based materials with catanionic sugar-derived surfactants as drug delivery systems.

    PubMed

    Roig, F; Blanzat, M; Solans, C; Esquena, J; García-Celma, M J

    2018-04-01

    In the present work novel drug delivery systems consisting in highly porous Hyaluronan foams for the administration of a non-steroidal anti-inflammatory drug (NSAID), ketoprofen, have been obtained. A sugar-derived surfactant associated with ketoprofen was prepared and incorporated into the porous hyaluronan materials. The association between a lactose derived surfactant, Lhyd 12 , and ketoprofen was obtained by acid-base reaction and its physicochemical properties were studied. Tensiometric and dynamic light scattering (DLS) determinations showed the formation of catanionic surfactant aggregates, Lhyd 12 /ketoprofen, in aqueous solution. Furthermore, the catanionic surfactants allowed greater solubilisation of ketoprofen. Hyaluronan porous materials were developed using butanediol diglycidyl ether as crosslinking agent. The profile release of Lhyd 12 /ketoprofen from hyaluronan based materials shows differences as a function of the aggregation state of catanionic surfactant. Copyright © 2018 Elsevier B.V. All rights reserved.

  2. 4-Methylumbelliferone Diminishes Catabolically Activated Articular Chondrocytes and Cartilage Explants via a Mechanism Independent of Hyaluronan Inhibition*

    PubMed Central

    Ishizuka, Shinya; Askew, Emily B.; Ishizuka, Naoko; Knudson, Cheryl B.; Knudson, Warren

    2016-01-01

    Depletion of the cartilage proteoglycan aggrecan is one of the earliest events that occurs in association with osteoarthritis. This loss is often accompanied by a coordinate loss in another glycosaminoglycan, hyaluronan. Chondrocytes experimentally depleted of cell-associated hyaluronan respond by switching to a pro-catabolic metabolism that includes enhanced production of endogenous inflammatory mediators and increased synthesis of matrix metalloproteinases. Hyaluronan turnover is also increased. Together, such a response provides for possible establishment of a self-perpetuating spiral of events that maintains or prolongs the pro-catabolic state. Chondrocytes or cartilage can also be activated by treatment with pro-inflammatory cytokines and mediators such as IL-1β, TNFα, LPS, fibronectin fragments, and hyaluronan oligosaccharides. To determine the mechanism of chondrocyte activation due to hyaluronan loss, a depletion method was required that did not include degrading the hyaluronan. In recent years, several laboratories have used the coumarin derivative, 4-methylumbelliferone, as a potent inhibitor of hyaluronan biosynthesis, due in part to its ability to sequester intracellular UDP-glucuronic acid and inhibition of hyaluronan synthase transcription. However, contrary to our expectation, although 4-methylumbelliferone was indeed an inhibitor of hyaluronan biosynthesis, this depletion did not give rise to an activation of chondrocytes or cartilage. Rather, 4-methylumbelliferone directly and selectively blocked gene products associated with the pro-catabolic metabolic state of chondrocytes and did so through a mechanism preceding and independent of hyaluronan inhibition. These data suggest that 4-methylumbelliferone has additional useful applications to block pro-inflammatory cell activation events but complicates how it is used for defining functions related to hyaluronan. PMID:27129266

  3. Adiponectin promotes hyaluronan synthesis along with increases in hyaluronan synthase 2 transcripts through an AMP-activated protein kinase/peroxisome proliferator-activated receptor-{alpha}-dependent pathway in human dermal fibroblasts

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Yamane, Takumi; Kobayashi-Hattori, Kazuo; Oishi, Yuichi, E-mail: y3oishi@nodai.ac.jp

    2011-11-18

    Highlights: Black-Right-Pointing-Pointer Adiponectin promotes hyaluronan synthesis along with an increase in HAS2 transcripts. Black-Right-Pointing-Pointer Adiponectin also increases the phosphorylation of AMPK. Black-Right-Pointing-Pointer A pharmacological activator of AMPK increases mRNA levels of PPAR{alpha} and HAS2. Black-Right-Pointing-Pointer Adiponectin-induced HAS2 mRNA expression is blocked by a PPAR{alpha} antagonist. Black-Right-Pointing-Pointer Adiponectin promotes hyaluronan synthesis via an AMPK/PPAR{alpha}-dependent pathway. -- Abstract: Although adipocytokines affect the functions of skin, little information is available on the effect of adiponectin on the skin. In this study, we investigated the effect of adiponectin on hyaluronan synthesis and its regulatory mechanisms in human dermal fibroblasts. Adiponectin promoted hyaluronan synthesis alongmore » with an increase in the mRNA levels of hyaluronan synthase 2 (HAS2), which plays a primary role in hyaluronan synthesis. Adiponectin also increased the phosphorylation of AMP-activated protein kinase (AMPK). A pharmacological activator of AMPK, 5-aminoimidazole-4-carboxamide-1{beta}-ribofuranoside (AICAR), increased mRNA levels of peroxisome proliferator-activated receptor-{alpha} (PPAR{alpha}), which enhances the expression of HAS2 mRNA. In addition, AICAR increased the mRNA levels of HAS2. Adiponectin-induced HAS2 mRNA expression was blocked by GW6471, a PPAR{alpha} antagonist, in a concentration-dependent manner. These results show that adiponectin promotes hyaluronan synthesis along with increases in HAS2 transcripts through an AMPK/PPAR{alpha}-dependent pathway in human dermal fibroblasts. Thus, our study suggests that adiponectin may be beneficial for retaining moisture in the skin, anti-inflammatory activity, and the treatment of a variety of cutaneous diseases.« less

  4. Self-Assembly of Polysaccharides Gives Rise to Distinct Mechanical Signatures in Marine Gels

    PubMed Central

    Pletikapić, G.; Lannon, H.; Murvai, Ü.; Kellermayer, M.S.Z.; Svetličić, V.; Brujic, J.

    2014-01-01

    Marine-gel biopolymers were recently visualized at the molecular level using atomic force microscopy (AFM) to reveal fine fibril-forming networks with low to high degrees of cross-linking. In this work, we use force spectroscopy to quantify the intra- and intermolecular forces within the marine-gel network. Combining force measurements, AFM imaging, and the known chemical composition of marine gels allows us to identify the microscopic origins of distinct mechanical responses. At the single-fibril level, we uncover force-extension curves that resemble those of individual polysaccharide fibrils. They exhibit entropic elasticity followed by extensions associated with chair-to-boat transitions specific to the type of polysaccharide at high forces. Surprisingly, a low degree of cross-linking leads to sawtooth patterns that we attribute to the unraveling of polysaccharide entanglements. At a high degree of cross-linking, we observe force plateaus that arise from unzipping, as well as unwinding, of helical bundles. Finally, the complex 3D network structure gives rise to force staircases of increasing height that correspond to the hierarchical peeling of fibrils away from the junction zones. In addition, we show that these diverse mechanical responses also arise in reconstituted polysaccharide gels, which highlights their dominant role in the mechanical architecture of marine gels. PMID:25028877

  5. Hyaluronan in aged collagen matrix increases prostate epithelial cell proliferation

    PubMed Central

    Damodarasamy, Mamatha; Vernon, Robert B.; Chan, Christina K.; Plymate, Stephen R.; Wight, Thomas N.

    2015-01-01

    The extracellular matrix (ECM) of the prostate, which is comprised primarily of collagen, becomes increasingly disorganized with age, a property that may influence the development of hyperplasia and cancer. Collageous ECM extracted from the tails of aged mice exhibits many characteristics of collagen in aged tissues, including the prostate. When polymerized into a 3-dimensional (3D) gel, these collagen extracts can serve as models for the study of specific cell-ECM interactions. In the present study, we examined the behaviors of human prostatic epithelial cell lines representing normal prostate epithelial cells (PEC), benign prostatic hyperplasia (BPH-1), and adenocarcinoma (LNCaP) cultured in contact with 3D gels made from collagen extracts of young and aged mice. We found that proliferation of PEC, BPH-1, and LNCaP cells were all increased by culture on aged collagen gels relative to young collagen gels. In examining age-associated differences in the composition of the collagen extracts, we found that aged and young collagen had a similar amount of several collagen-associated ECM components, but aged collagen had a much greater content of the glycosaminoglycan hyaluronan (HA) than young collagen. The addition of HA (of similar size and concentration to that found in aged collagen extracts) to cells placed in young collagen elicited significantly increased proliferation in BPH-1 cells, but not in PEC or LNCaP cells, relative to controls not exposed to HA. Of note, histochemical analyses of human prostatic tissues showed significantly higher expression of HA in BPH and prostate cancer stroma relative to stroma of normal prostate. Collectively, these results suggest that changes in ECM involving increased levels of HA contribute to the growth of prostatic epithelium with aging. PMID:25124870

  6. Priming Adipose-Derived Mesenchymal Stem Cells with Hyaluronan Alters Growth Kinetics and Increases Attachment to Articular Cartilage

    PubMed Central

    Succar, Peter; Medynskyj, Michael; Breen, Edmond J.; Batterham, Tony; Molloy, Mark P.; Herbert, Benjamin R.

    2016-01-01

    Background. Biological therapeutics such as adipose-derived mesenchymal stem cell (MSC) therapy are gaining acceptance for knee-osteoarthritis (OA) treatment. Reports of OA-patients show reductions in cartilage defects and regeneration of hyaline-like-cartilage with MSC-therapy. Suspending MSCs in hyaluronan commonly occurs in animals and humans, usually without supporting data. Objective. To elucidate the effects of different concentrations of hyaluronan on MSC growth kinetics. Methods. Using a range of hyaluronan concentrations, we measured MSC adherence and proliferation on culture plastic surfaces and a novel cartilage-adhesion assay. We employed time-course and dispersion imaging to assess MSC binding to cartilage. Cytokine profiling was also conducted on the MSC-secretome. Results. Hyaluronan had dose-dependent effects on growth kinetics of MSCs at concentrations of entanglement point (1 mg/mL). At higher concentrations, viscosity effects outweighed benefits of additional hyaluronan. The cartilage-adhesion assay highlighted for the first time that hyaluronan-primed MSCs increased cell attachment to cartilage whilst the presence of hyaluronan did not. Our time-course suggested patients undergoing MSC-therapy for OA could benefit from joint-immobilisation for up to 8 hours. Hyaluronan also greatly affected dispersion of MSCs on cartilage. Conclusion. Our results should be considered in future trials with MSC-therapy using hyaluronan as a vehicle, for the treatment of OA. PMID:26981136

  7. A RHAMM mimetic peptide blocks hyaluronan signaling and reduces inflammation and fibrogenesis in excisional skin wounds.

    PubMed

    Tolg, Cornelia; Hamilton, Sara R; Zalinska, Ewa; McCulloch, Lori; Amin, Ripal; Akentieva, Natalia; Winnik, Francoise; Savani, Rashmin; Bagli, Darius J; Luyt, Len G; Cowman, Mary K; McCarthy, Jim B; Turley, Eva A

    2012-10-01

    Hyaluronan is activated by fragmentation and controls inflammation and fibroplasia during wound repair and diseases (eg, cancer). Hyaluronan-binding peptides were identified that modify fibrogenesis during skin wound repair. Peptides were selected from 7- to 15mer phage display libraries by panning with hyaluronan-Sepharose beads and assayed for their ability to block fibroblast migration in response to hyaluronan oligosaccharides (10 kDa). A 15mer peptide (P15-1), with homology to receptor for hyaluronan mediated motility (RHAMM) hyaluronan binding sequences, was the most effective inhibitor. P15-1 bound to 10-kDa hyaluronan with an affinity of K(d) = 10(-7) and appeared to specifically mimic RHAMM since it significantly reduced binding of hyaluronan oligosaccharides to recombinant RHAMM but not to recombinant CD44 or TLR2,4, and altered wound repair in wild-type but not RHAMM(-/-) mice. One topical application of P15-1 to full-thickness excisional rat wounds significantly reduced wound macrophage number, fibroblast number, and blood vessel density compared to scrambled, negative control peptides. Wound collagen 1, transforming growth factor β-1, and α-smooth muscle actin were reduced, whereas tenascin C was increased, suggesting that P15-1 promoted a form of scarless healing. Signaling/microarray analyses showed that P15-1 blocks RHAMM-regulated focal adhesion kinase pathways in fibroblasts. These results identify a new class of reagents that attenuate proinflammatory, fibrotic repair by blocking hyaluronan oligosaccharide signaling. Copyright © 2012 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  8. On the Formation and Properties of Interstrand DNA-DNA Cross-links Forged by Reaction of an Abasic Site With the Opposing Guanine Residue of 5′-CAp Sequences in Duplex DNA

    PubMed Central

    Johnson, Kevin M.; Price, Nathan E.; Wang, Jin; Fekry, Mostafa I.; Dutta, Sanjay; Seiner, Derrick R.; Wang, Yinsheng; Gates, Kent S.

    2014-01-01

    We recently reported that the aldehyde residue of an abasic (Ap) site in duplex DNA can generate an interstrand cross-link via reaction with a guanine residue on the opposing strand. This finding is intriguing because the highly deleterious nature of interstrand cross-links suggests that even small amounts of Ap-derived cross-links could make a significant contribution to the biological consequences stemming from the generation of Ap sites in cellular DNA. Incubation of 21-bp duplexes containing a central 5′-CAp sequence under conditions of reductive amination (NaCNBH3, pH 5.2) generated much higher yields of cross-linked DNA than reported previously. At pH 7, in the absence of reducing agents, these Ap-containing duplexes also produced cross-linked duplexes that were readily detected on denaturing polyacrylamide gels. Cross-link formation was not highly sensitive to reaction conditions and, once formed, the cross-link was stable to a variety of work-up conditions. Results of multiple experiments including MALDI-TOF mass spectrometry, gel mobility, methoxyamine capping of the Ap aldehyde, inosine-for-guanine replacement, hydroxyl radical footprinting, and LCMS/MS were consistent with a cross-linking mechanism involving reversible reaction of the Ap aldehyde residue with the N2-amino group of the opposing guanine residue in 5′-CAp sequences to generate hemiaminal, imine, or cyclic hemiaminal cross-links (7-10) that were irreversibly converted under conditions of reductive amination (NaCNBH3/pH 5.2) to a stable amine linkage. Further support for the importance of the exocyclic N2-amino group in this reaction was provided by an experiment showing that installation of a 2-aminopurine-thymine base pair at the cross-linking site produced high yields (15-30%) of a cross-linked duplex at neutral pH, in the absence of NaCNBH3. PMID:23215239

  9. The effects of hyaluronan and its fragments on lipid models exposed to UV irradiation.

    PubMed

    Trommer, Hagen; Wartewig, Siegfried; Böttcher, Rolf; Pöppl, Andreas; Hoentsch, Joachim; Ozegowski, Jörg H; Neubert, Reinhard H H

    2003-03-26

    The effects of hyaluronan and its degradation products on irradiation-induced lipid peroxidation were investigated. Liposomal skin lipid models with increasing complexity were used. Hyaluronan and its fragments were able to reduce the amount of lipid peroxidation secondary products quantified by the thiobarbituric acid (TBA) assay. The qualitative changes were studied by mass spectrometry. To elucidate the nature of free radical involvement electron paramagnetic resonance (EPR) studies were carried out. The influence of hyaluronan and its fragments on the concentration of hydroxyl radicals generated by the Fenton system was examined using the spin trapping technique. Moreover, the mucopolysaccharide's ability to react with stable radicals was checked. The quantification assay of 2,2-diphenyl-1-picrylhydrazyl hydrate (DPPH) showed no concentration changes of the stable radical caused by hyaluronan. Hyaluronan was found to exhibit prooxidative effects in the Fenton assay in a concentration dependent manner. A transition metal chelation was proposed as a mechanism of this behavior. Considering human skin and its constant exposure to UV light and oxygen and an increased pool of iron in irradiated skin the administration of hyaluronan or its fragments in cosmetic formulations or sunscreens could be helpful for the protection of the human skin. Copyright 2003 Elsevier Science B.V.

  10. Viscoelastic properties of rabbit vocal folds after augmentation.

    PubMed

    Hertegård, Stellan; Dahlqvist, Ake; Laurent, Claude; Borzacchiello, Assunta; Ambrosio, Luigi

    2003-03-01

    Vocal fold function is closely related to tissue viscoelasticity. Augmentation substances may alter the viscoelastic properties of vocal fold tissues and hence their vibratory capacity. We sought to investigate the viscoelastic properties of rabbit vocal folds in vitro after injections of various augmentation substances. Polytetrafluoroethylene (Teflon), cross-linked collagen (Zyplast), and cross-linked hyaluronan, hylan b gel (Hylaform) were injected into the lamina propria and the thyroarytenoid muscle of rabbit vocal folds. Dynamic viscosity of the injected vocal fold as a function of frequency was measured with a Bohlin parallel-plate rheometer during small-amplitude oscillation. All injected vocal folds showed a decreasing dynamic viscosity with increasing frequency. Vocal fold samples injected with hylan b gel showed the lowest dynamic viscosity, quite close to noninjected control samples. Vocal folds injected with polytetrafluoroethylene showed the highest dynamic viscosity followed by the collagen samples. The data indicated that hylan b gel in short-term renders the most natural viscoelastic properties to the vocal fold among the substances tested. This is of importance to restore/preserve the vibratory capacity of the vocal folds when glottal insufficiency is treated with injections.

  11. Structure-Property Relationships in Porous 3-D Nanostructures as a Function of Preparation Conditions: Isocyanate Cross-Linked Silica Aerogels

    NASA Technical Reports Server (NTRS)

    Meador, Mary Ann B.; Capadona, Lynn A.; McCorkle, Linda; Padadopoulos, Demetrios S.; Leventis, Nicholas

    2007-01-01

    Sol-gel derived silica aerogels are attractive candidates for many unique thermal, optical, catalytic, and chemical applications because of their low density and high mesoporosity. However, their inherent fragility has restricted use of aerogel monoliths to applications where they are not subject to any load. We have previously reported cross-linking the mesoporous silica structure of aerogels with di-isocyanates, styrenes or epoxies reacting with amine decorated silica surfaces. These approaches have been shown to significantly increase the strength of aerogels with only a small effect on density or porosity. Though density is a prime predictor of properties such as strength and thermal conductivity for aerogels, it is becoming clear from previous studies that varying the silica backbone and size of the polymer cross-link independently can give rise to combinations of properties which cannot be predicted from density alone. Herein, we examine the effects of four processing parameters for producing this type of polymer cross-linked aerogel on properties of the resulting monoliths. We focus on the results of C-13 CP-MAS NMR which gives insight to the size and structure of polymer cross-link present in the monoliths, and relates the size of the cross-links to microstructure, mechanical properties and other characteristics of the materials obtained.

  12. Structure-Property Relationships in Porous 3-D Nanostructures as a Function of Preparation Conditions: Isocyanate Cross-Linked Silica Aerogels

    NASA Technical Reports Server (NTRS)

    Meador, Mary Ann B.; Capadona, Lynn A.; McCorkle, Linda; Papadopoulos, Demetrios S.; Leventis, Nicholas

    2007-01-01

    Sol-gel derived silica aerogels are attractive candidates for many unique thermal, optical, catalytic, and chemical applications because of their low density and high mesoporosity. However, their inherent fragility has restricted use of aerogel monoliths to applications where they are not subject to any load. We have previously reported cross-linking the mesoporous silica structure of aerogels with di-isocyanates, styrenes or epoxies reacting with amine decorated silica surfaces. These approaches have been shown to significantly increase the strength of aerogels with only a small effect on density or porosity. Though density is a prime predictor of properties such as strength and thermal conductivity for aerogels, it is becoming clear from previous studies that varying the silica backbone and size of the polymer cross-link independently can give rise to combinations of properties which cannot be predicted from density alone. Herein, we examine the effects of four processing parameters for producing this type of polymer cross-linked aerogel on properties of the resulting monoliths. We focus on the results of 13C CP-MAS NMR which gives insight to the size and structure of polymer cross-link present in the monoliths, and relates the size of the cross-links to microstructure, mechanical properties and other characteristics of the materials obtained.

  13. Non-volatile copolymer compositions for fabricating gel element microarrays

    PubMed Central

    Golova, Julia B.; Chernov, Boris K.; Perov, Alexander N.; Reynolds, Jennifer; Linger, Yvonne L.; Kukhtin, Alexander; Chandler, Darrell P.

    2011-01-01

    By modifying polymer compositions and cross-linking reagents, we have developed a simple yet effective manufacturing strategy for copolymerized three-dimensional gel element arrays. A new gel-forming monomer (2-(hydroxyethyl) methacrylamide; HEMAA) was used that possesses low volatility and improves the stability of copolymerized gel element arrays to on-chip thermal cycling procedures relative to previously used monomers. Probe immobilization efficiency within the new polymer was 55%, equivalent to that obtained with acrylamide (AA) and methacrylamide (MA) monomers. Non-specific binding of single stranded targets was equivalent for all monomers. Increasing cross-linker chain length improved hybridization kinetics and end-point signal intensities relative to N,N-methylenebisacrylamide (Bis). The new copolymer formulation was successfully applied to a model orthopox array. Because HEMAA greatly simplifies gel element array manufacture, we expect it (in combination with new cross-linkers described herein) to find widespread application in microarray science. PMID:22033291

  14. New Aptes Cross-linked Polymers from Poly(ethylene oxide)s and Cyanuric Chloride for Lithium Batteries

    NASA Technical Reports Server (NTRS)

    Tigelaar, Dean M.; Meador, Mary Ann B.; Kinder, James D.; Bennett, William R.

    2005-01-01

    A new series of polymer electrolytes for use as membranes for lithium batteries are described. Electrolytes were made by polymerization between cyanuric chloride and diamino-terminated poly(ethylene oxide)s, followed by cross-linking via a sol-gel process. Thermal analysis and lithium conductivity of freestanding polymer films were studied. The effects of several variables on conductivity were investigated, such as length of backbone PEO chain, length of branching PEO chain, extent of branching, extent of cross-linking, salt content, and salt counterion. Polymer films with the highest percentage of PEO were found to be the most conductive, with a maximum lithium conductivity of 3.9 x 10(exp -5) S/cm at 25 C. Addition of plasticizer to the dry polymers increased conductivity by an order of magnitude.

  15. Role of Hyaluronan in Schwannoma Growth

    DTIC Science & Technology

    2008-06-01

    schwannomatosis . Although schwannomas occur due to mutations in the neurofibromatosis 2 gene, which encodes the merlin tumor suppressor protein, recent...pre-disposition syndromes neurofibromatosis 2 (NF2) and schwannomatosis . We and others found that the glycosaminoglycan hyaluronan (HA) is present

  16. Multifunctional Nanostructured Conductive Polymer Gels: Synthesis, Properties, and Applications

    DOE PAGES

    Zhao, Fei; Shi, Ye; Pan, Lijia; ...

    2017-06-26

    Conductive polymers have attracted significant interest over the past few decades because they synergize the advantageous features of conventional polymeric materials and organic conductors. With rationally designed nanostructures, conductive polymers can further exhibit exceptional mechanical, electrical, and optical properties because of their confined dimensions at the nanoscale level. Among various nanostructured conductive polymers, conductive polymer gels (CPGs) with synthetically tunable hierarchical 3D network structures show great potential for a wide range of applications, such as bioelectronics, and energy storage/conversion devices owing to their structural features. CPGs retain the properties of nanosized conductive polymers during the assembly of the nanobuilding blocksmore » into a monolithic macroscopic structure while generating structure-derived features from the highly cross-linked network. In this Account, we review our recent progress on the synthesis, properties, and novel applications of dopant cross-linked CPGs. We first describe the synthetic strategies, in which molecules with multiple functional groups are adopted as cross-linkers to cross-link conductive polymer chains into a 3D molecular network. These cross-linking molecules also act as dopants to improve the electrical conductivity of the gel network. The microstructure and physical/chemical properties of CPGs can be tuned by controlling the synthetic conditions such as species of monomers and cross-linkers, reaction temperature, and solvents. By incorporating other functional polymers or particles into the CPG matrix, hybrid gels have been synthesized with tailored structures. These hybrid gel materials retain the functionalities from each component, as well as enable synergic effects to improve mechanical and electrical properties of CPGs. We then introduce the unique structure-derived properties of the CPGs. The network facilitates both electronic and ionic transport owing to the

  17. Multifunctional Nanostructured Conductive Polymer Gels: Synthesis, Properties, and Applications

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Zhao, Fei; Shi, Ye; Pan, Lijia

    Conductive polymers have attracted significant interest over the past few decades because they synergize the advantageous features of conventional polymeric materials and organic conductors. With rationally designed nanostructures, conductive polymers can further exhibit exceptional mechanical, electrical, and optical properties because of their confined dimensions at the nanoscale level. Among various nanostructured conductive polymers, conductive polymer gels (CPGs) with synthetically tunable hierarchical 3D network structures show great potential for a wide range of applications, such as bioelectronics, and energy storage/conversion devices owing to their structural features. CPGs retain the properties of nanosized conductive polymers during the assembly of the nanobuilding blocksmore » into a monolithic macroscopic structure while generating structure-derived features from the highly cross-linked network. In this Account, we review our recent progress on the synthesis, properties, and novel applications of dopant cross-linked CPGs. We first describe the synthetic strategies, in which molecules with multiple functional groups are adopted as cross-linkers to cross-link conductive polymer chains into a 3D molecular network. These cross-linking molecules also act as dopants to improve the electrical conductivity of the gel network. The microstructure and physical/chemical properties of CPGs can be tuned by controlling the synthetic conditions such as species of monomers and cross-linkers, reaction temperature, and solvents. By incorporating other functional polymers or particles into the CPG matrix, hybrid gels have been synthesized with tailored structures. These hybrid gel materials retain the functionalities from each component, as well as enable synergic effects to improve mechanical and electrical properties of CPGs. We then introduce the unique structure-derived properties of the CPGs. The network facilitates both electronic and ionic transport owing to the

  18. Multifunctional Nanostructured Conductive Polymer Gels: Synthesis, Properties, and Applications.

    PubMed

    Zhao, Fei; Shi, Ye; Pan, Lijia; Yu, Guihua

    2017-07-18

    Conductive polymers have attracted significant interest over the past few decades because they synergize the advantageous features of conventional polymeric materials and organic conductors. With rationally designed nanostructures, conductive polymers can further exhibit exceptional mechanical, electrical, and optical properties because of their confined dimensions at the nanoscale level. Among various nanostructured conductive polymers, conductive polymer gels (CPGs) with synthetically tunable hierarchical 3D network structures show great potential for a wide range of applications, such as bioelectronics, and energy storage/conversion devices owing to their structural features. CPGs retain the properties of nanosized conductive polymers during the assembly of the nanobuilding blocks into a monolithic macroscopic structure while generating structure-derived features from the highly cross-linked network. In this Account, we review our recent progress on the synthesis, properties, and novel applications of dopant cross-linked CPGs. We first describe the synthetic strategies, in which molecules with multiple functional groups are adopted as cross-linkers to cross-link conductive polymer chains into a 3D molecular network. These cross-linking molecules also act as dopants to improve the electrical conductivity of the gel network. The microstructure and physical/chemical properties of CPGs can be tuned by controlling the synthetic conditions such as species of monomers and cross-linkers, reaction temperature, and solvents. By incorporating other functional polymers or particles into the CPG matrix, hybrid gels have been synthesized with tailored structures. These hybrid gel materials retain the functionalities from each component, as well as enable synergic effects to improve mechanical and electrical properties of CPGs. We then introduce the unique structure-derived properties of the CPGs. The network facilitates both electronic and ionic transport owing to the

  19. Development of casein microgels from cross-linking of casein micelles by genipin.

    PubMed

    Silva, Naaman F Nogueira; Saint-Jalmes, Arnaud; de Carvalho, Antônio F; Gaucheron, Frédéric

    2014-09-02

    Casein micelles are porous colloidal particles, constituted of casein molecules, water, and minerals. The vulnerability of the supramolecular structure of casein micelles face to changes in the environmental conditions restrains their applications in other domains besides food. Thus, redesigning casein micelles is a challenge to create new functionalities for these biosourced particles. The objective of this work was to create stable casein microgels from casein micelles using a natural cross-linker, named genipin. Suspensions of purified casein micelles (25 g L(-1)) were mixed with genipin solutions to have final concentrations of 5, 10, and 20 mM genipin. Covalently linked casein microgels were formed via cross-linking of lysyl and arginyl residues of casein molecules. The reacted products exhibited blue color. The cross-linking reaction induced gradual changes on the colloidal properties of the particles. The casein microgels were smaller and more negatively charged and presented smoother surfaces than casein micelles. These results were explained based on the cross-linking of free NH2 present in an external layer of κ-casein. Light scattering and rheological measurements showed that the reaction between genipin and casein molecules was intramicellar, as one single population of particles was observed and the values of viscosity (and, consequently, the volume fraction of the particles) were reduced. Contrary to the casein micelles, the casein microgels were resistant to the presence of dissociating agents, e.g., citrate (calcium chelating) and urea, but swelled as a consequence of internal electrostatic repulsion and the disruption of hydrophobic interactions between protein chains. The casein microgels did not dissociate at the air-solution interface and formed solid-like interfaces rather than a viscoelastic gel. The potential use of casein microgels as adaptable nanocarriers is proposed in the article.

  20. Self-assembly of polysaccharides gives rise to distinct mechanical signatures in marine gels.

    PubMed

    Pletikapić, G; Lannon, H; Murvai, Ü; Kellermayer, M S Z; Svetličić, V; Brujic, J

    2014-07-15

    Marine-gel biopolymers were recently visualized at the molecular level using atomic force microscopy (AFM) to reveal fine fibril-forming networks with low to high degrees of cross-linking. In this work, we use force spectroscopy to quantify the intra- and intermolecular forces within the marine-gel network. Combining force measurements, AFM imaging, and the known chemical composition of marine gels allows us to identify the microscopic origins of distinct mechanical responses. At the single-fibril level, we uncover force-extension curves that resemble those of individual polysaccharide fibrils. They exhibit entropic elasticity followed by extensions associated with chair-to-boat transitions specific to the type of polysaccharide at high forces. Surprisingly, a low degree of cross-linking leads to sawtooth patterns that we attribute to the unraveling of polysaccharide entanglements. At a high degree of cross-linking, we observe force plateaus that arise from unzipping, as well as unwinding, of helical bundles. Finally, the complex 3D network structure gives rise to force staircases of increasing height that correspond to the hierarchical peeling of fibrils away from the junction zones. In addition, we show that these diverse mechanical responses also arise in reconstituted polysaccharide gels, which highlights their dominant role in the mechanical architecture of marine gels. Copyright © 2014 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  1. Cross-linking Chemistry of Squid Beak*

    PubMed Central

    Miserez, Ali; Rubin, Daniel; Waite, J. Herbert

    2010-01-01

    In stark contrast to most aggressive predators, Dosidicus gigas (jumbo squids) do not use minerals in their powerful mouthparts known as beaks. Their beaks instead consist of a highly sclerotized chitinous composite with incremental hydration from the tip to the base. We previously reported l-3,4-dihydroxyphenylalanine (dopa)-histidine (dopa-His) as an important covalent cross-link providing mechanical strengthening to the beak material. Here, we present a more complete characterization of the sclerotization chemistry and describe additional cross-links from D. gigas beak. All cross-links presented in this report share common building blocks, a family of di-, tri-, and tetra-histidine-catecholic adducts, that were separated by affinity chromatography and high performance liquid chromatography (HPLC) and identified by tandem mass spectroscopy and proton nuclear magnetic resonance (1H NMR). The data provide additional insights into the unusually high cross-link density found in mature beaks. Furthermore, we propose both a low molecular weight catechol, and peptidyl-dopa, to be sclerotization agents of squid beak. This appears to represent a new strategy for forming hard tissue in animals. The interplay between covalent cross-linking and dehydration on the graded properties of the beaks is discussed. PMID:20870720

  2. Cross-linking chemistry of squid beak.

    PubMed

    Miserez, Ali; Rubin, Daniel; Waite, J Herbert

    2010-12-03

    In stark contrast to most aggressive predators, Dosidicus gigas (jumbo squids) do not use minerals in their powerful mouthparts known as beaks. Their beaks instead consist of a highly sclerotized chitinous composite with incremental hydration from the tip to the base. We previously reported l-3,4-dihydroxyphenylalanine (dopa)-histidine (dopa-His) as an important covalent cross-link providing mechanical strengthening to the beak material. Here, we present a more complete characterization of the sclerotization chemistry and describe additional cross-links from D. gigas beak. All cross-links presented in this report share common building blocks, a family of di-, tri-, and tetra-histidine-catecholic adducts, that were separated by affinity chromatography and high performance liquid chromatography (HPLC) and identified by tandem mass spectroscopy and proton nuclear magnetic resonance ((1)H NMR). The data provide additional insights into the unusually high cross-link density found in mature beaks. Furthermore, we propose both a low molecular weight catechol, and peptidyl-dopa, to be sclerotization agents of squid beak. This appears to represent a new strategy for forming hard tissue in animals. The interplay between covalent cross-linking and dehydration on the graded properties of the beaks is discussed.

  3. Hyaluronan Protects Bovine Articular Chondrocytes against Cell Death Induced by Bupivacaine under Supraphysiologic Temperatures

    PubMed Central

    Liu, Sen; Zhang, Qing-Song; Hester, William; O’Brien, Michael J.; Savoie, Felix H.; You, Zongbing

    2013-01-01

    Background Bupivacaine and supraphysiologic temperature can independently reduce cell viability of articular chondrocytes. In combination these two deleterious factors could further impair cell viability. Hypothesis Hyaluronan may protect chondrocytes from death induced by bupivacaine at supraphysiologic temperatures. Study Design Controlled laboratory study. Methods Bovine articular chondrocytes were treated with hyaluronan at physiologic (37°C) and supraphysiologic temperatures (45°C and 50°C) for one hour, and then exposed to bupivacaine for one hour at room temperature. Cell viability was assessed at three time points: immediately after treatment, six hours later, and twenty-four hours later using flow cytometry and fluorescence microscopy. The effects of hyaluronan on the levels of sulfated glycosaminoglycan in the chondrocytes were determined using Alcian blue staining. Results (1) Bupivacaine alone did not induce noticeable chondrocyte death at 37°C; (2) bupivacaine and temperature synergistically increased chondrocyte death, that is, when the chondrocytes were conditioned to 45°C and 50°C, 0.25% and 0.5% bupivacaine increased the cell death rate by 131% to 383% in comparison to the phosphate-buffered saline control group; and, (3) addition of hyaluronan reduced chondrocyte death rates to approximately 14% and 25% at 45°C and 50°C, respectively. Hyaluronan’s protective effects were still observed at six and twenty-four hours after bupivacaine treatment at 45°C. However, at 50°C, hyaluronan delayed but did not prevent the cell death caused by bupivacaine. One-hour treatment with hyaluronan significantly increased sulfated glycosaminoglycan levels in the chondrocytes. Conclusions Bupivacaine and supraphysiologic temperature synergistically increase chondrocyte death and hyaluronan may protect articular chondrocytes from death caused by bupivacaine. Clinical Relevance This study provides a rationale to perform pre-clinical and clinical studies to

  4. Heat shrinkable behavior, physico-mechanical and structure properties of electron beam cross-linked blends of high-density polyethylene with acrylonitrile-butadiene rubber

    NASA Astrophysics Data System (ADS)

    Reinholds, Ingars; Kalkis, Valdis; Merijs-Meri, Remo; Zicans, Janis; Grigalovica, Agnese

    2016-03-01

    In this study, heat-shrinkable composites of electron beam irradiated high-density polyethylene (HDPE) composites with acrylonitrile-butadiene rubber (NBR) were investigated. HDPE/NBR blends at a ratio of components 100/0, 90/10, 80/20, 50/50 and 20/80 wt% were prepared using a two-roll mill. The compression molded films were irradiated high-energy (5 MeV) accelerated electrons up to irradiation absorbed doses of 100-300 kGy. The effect of electron beam induced cross-linking was evaluated by the changes of mechanical properties, gel content and by the differences of thermal properties, detected by differential scanning calorimetry. The thermo-shrinkage forces were determined as the kinetics of thermorelaxation and the residual shrinkage stresses of previously oriented (stretched up to 100% at above melting temperature of HDPE and followed by cooling to room temperature) specimens of irradiated HDPE/NBR blends under isometric heating-cooling mode. The compatibility between the both components was enhanced due to the formation of cross-linked sites at amorphous interphase. The results showed increase of mechanical stiffness of composites with increase of irradiation dose. The values of gel fraction compared to thermorelaxation stresses increased with the growth of irradiation dose level, as a result of formation cross-linked sites in amorphous PP/NBR interphase.

  5. Skin bacterial flora as a potential risk factor predisposing to late bacterial infection after cross-linked hyaluronic acid gel augmentation.

    PubMed

    Netsvyetayeva, Irina; Marusza, Wojciech; Olszanski, Romuald; Szyller, Kamila; Krolak-Ulinska, Aneta; Swoboda-Kopec, Ewa; Sierdzinski, Janusz; Szymonski, Zachary; Mlynarczyk, Grazyna

    2018-01-01

    Cross-linked hyaluronic acid (HA) gel is widely used in esthetic medicine. Late bacterial infection (LBI) is a rare, but severe complication after HA augmentation. The aim of this study was to determine whether patients who underwent the HA injection procedure and developed LBI had qualitatively different bacterial flora on the skin compared to patients who underwent the procedure without any complications. The study group comprised 10 previously healthy women with recently diagnosed, untreated LBI after HA augmentation. The control group comprised 17 healthy women who had a similar amount of HA injected with no complications. To assess the difference between the two groups, their skin flora was cultured from nasal swabs, both before and after antibiotic treatment in the study group. A significant increase in the incidence of Staphylococcus epidermidis was detected in the control group ( P =0.000) compared to the study group. The study group showed a significantly higher incidence of Staphylococcus aureus ( P =0.005), Klebsiella pneumoniae ( P =0.006), Klebsiella oxytoca ( P =0.048), and Staphylococcus haemolyticus ( P =0.048) compared to the control group. The bacterial flora on the skin differed in patients with LBI from the control group. The control group's bacterial skin flora was dominated by S. epidermidis . Patients with LBI had a bacterial skin flora dominated by potentially pathogenic bacteria.

  6. Nano and micro mechanical properties of uncross-linked and cross-linked chitosan films

    PubMed Central

    Aryaei, Ashkan; Jayatissa, Ahalapitiya H.; Jayasuriya, A. Champa

    2016-01-01

    The aim of this study is to determine the nano and micro mechanical properties for uncross-linked and cross-linked chitosan films. Specifically, we looked at nanoindentation hardness, microhardness, and elastic modulus. It is important to study the nano and microscale mechanical properties of chitosan since chitosan has been widely used for biomedical applications. Using the solvent-cast method, the chitosan films were prepared at room temperature on the cleaned glass plates. The chitosan solution was prepared by dissolving chitosan in acetic acid 1% (v/v). Tripolyphosphate (TPP) was used to create the cross-links between amine groups in chitosan and phosphate groups in TPP. In this study, atomic force microscopy was used to measure the nanoindentation hardness and surface topography of the uncross-linked and cross-linked chitosan films. Elastic modulus was then calculated from the nanoindentation results. The effective elastic modulus was determined by microhardness with some modifications to previous theories. The microhardness of the chitosan films were measured using Vicker’s hardness meter under three different loads. Our results show that the microhardness and elastic modulus for cross-linked chitosan films are higher than the uncross-linked films. However, the cross-linked chitosan films show increased brittleness when compared to uncross-linked films. By increasing the load magnitude, the microhardness increases for both uncross-linked and cross-linked chitosan films. PMID:22100082

  7. Oral administration of hyaluronan prevents skin dryness and epidermal thickening in ultraviolet irradiated hairless mice.

    PubMed

    Kawada, Chinatsu; Kimura, Mamoru; Masuda, Yasunobu; Nomura, Yoshihiro

    2015-12-01

    Hyaluronan is a component of the extracellular matrix that plays a role in water retention in tissues. In this study, we orally administered hyaluronans of varying molecular weights (300k and less than 10k) repeatedly to hairless mice exposed to ultraviolet (UV) irradiation and examined their effects on the skin of these mice. UV irradiation induces a marked increase in the epidermal thickness of the dorsal skin and a marked decrease in the skin moisture content; however, orally administered hyaluronan, particularly that with a molecular weight of less than 10k, markedly reversed the increase and decrease in the epidermal thickness and skin moisture content, respectively. Furthermore, on analyzing the mice skin, orally administered hyaluronan with a molecular weight of less than 10k increased the levels of the HAS2 gene expression in the skin. Based on these findings, it is assumed that orally administered hyaluronans, with molecular weight of 300k and less than 10k, reversed UV irradiation-induced skin disturbance. In particular, it was considered that the increase in the skin moisture content by orally administered hyaluronan, with a molecular weight of less than 10k, was related to the effect on skin cells. Copyright © 2015 Elsevier B.V. All rights reserved.

  8. Recent advances in corneal collagen cross-linking

    PubMed Central

    Sachdev, Gitansha Shreyas; Sachdev, Mahipal

    2017-01-01

    Corneal collagen cross-linking has become the preferred modality of treatment for corneal ectasia since its inception in late 1990s. Numerous studies have demonstrated the safety and efficacy of the conventional protocol. Our understanding of the cross-linking process is ever evolving, with its wide implications in the form of accelerated and pulsed protocols. Newer advancements in technology include various riboflavin formulations and the ability to deliver higher fluence protocols with customised irradiation patterns. A greater degree of customisation is likely the path forward, which will aim at achieving refractive improvements along with disease stability. The use of cross-linking for myopic correction is another avenue under exploration. Combination of half fluence cross-linking with refractive correction for high errors to prevent post LASIK regression is gaining interest. This review aims to highlight the various advancements in the cross-linking technology and its clinical applications. PMID:28905820

  9. Hyaluronan synthase 3 (HAS3) overexpression downregulates MV3 melanoma cell proliferation, migration and adhesion

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Takabe, Piia, E-mail: piia.takabe@uef.fi; Bart, Geneviève; Ropponen, Antti

    2015-09-10

    Malignant skin melanoma is one of the most deadly human cancers. Extracellular matrix (ECM) influences the growth of malignant tumors by modulating tumor cells adhesion and migration. Hyaluronan is an essential component of the ECM, and its amount is altered in many tumors, suggesting an important role for hyaluronan in tumorigenesis. Nonetheless its role in melanomagenesis is not understood. In this study we produced a MV3 melanoma cell line with inducible expression of the hyaluronan synthase 3 (HAS3) and studied its effect on the behavior of the melanoma cells. HAS3 overexpression expanded the cell surface hyaluronan coat and decreased melanomamore » cell adhesion, migration and proliferation by cell cycle arrest at G1/G0. Melanoma cell migration was restored by removal of cell surface hyaluronan by Streptomyces hyaluronidase and by receptor blocking with hyaluronan oligosaccharides, while the effect on cell proliferation was receptor independent. Overexpression of HAS3 decreased ERK1/2 phosphorylation suggesting that inhibition of MAP-kinase signaling was responsible for these suppressive effects on the malignant phenotype of MV3 melanoma cells. - Highlights: • Inducible HAS3-MV3 melanoma cell line was generated using Lentiviral transduction. • HAS3 overexpression inhibits MV3 cell migration via hyaluronan–receptor interaction. • HAS3 overexpression decreases MV3 melanoma cell proliferation and adhesion. • ERK1/2 phosphorylation is downregulated by 50% in HAS3 overexpressing cells. • The results suggest that hyaluronan has anti-cancer like effects in melanoma.« less

  10. Hyaluronan hydrogels with a low degree of modification as scaffolds for cartilage engineering.

    PubMed

    La Gatta, Annalisa; Ricci, Giulia; Stellavato, Antonietta; Cammarota, Marcella; Filosa, Rosanna; Papa, Agata; D'Agostino, Antonella; Portaccio, Marianna; Delfino, Ines; De Rosa, Mario; Schiraldi, Chiara

    2017-10-01

    In the field of cartilage engineering, continuing efforts have focused on fabricating scaffolds that favor maintenance of the chondrocytic phenotype and matrix formation, in addition to providing a permeable, hydrated, microporous structure and mechanical support. The potential of hyaluronan-based hydrogels has been well established, but the ideal matrix remains to be developed. This study describes the development of hyaluronan sponges-based scaffolds obtained by lysine methyl-ester crosslinking. The reaction conditions are optimized with minimal chemical modifications to obtain materials that closely resemble elements in physiological cellular environments. Three hydrogels with different amounts of crosslinkers were produced that show morphological, water-uptake, mechanical, and stability properties comparable or superior to those of currently available hyaluronan-scaffolds, but with significantly fewer hyaluronan modifications. Primary human chondrocytes cultured with the most promising hydrogel were viable and maintained lineage identity for 3 weeks. They also secreted cartilage-specific matrix proteins. These scaffolds represent promising candidates for cartilage engineering. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Determination of the parameters controlling swelling of chemically cross-linked pH-sensitive poly(N-vinylimidazole) hydrogels.

    PubMed

    Molina, M Jesús; Gómez-Antón, M Rosa; Piérola, Inés F

    2007-10-25

    The number of variables controlling the behavior of ionic gels is large and very often some of them are unknown. The aim of this work is to interpret quantitatively the swelling behavior of pH sensitive gels, with the minimum number of simplifying assumptions. With this purpose, the equilibrium degree of swelling (S) and protonation (alpha) of chemically cross-linked poly(N-vinylimidazole) (PVI) immersed in aqueous salt solutions were measured as a function of the ionic strength (mu), in the whole range of pH. In acid solutions with pH in the range 0 to 4, imidazole moieties become protonated, and PVI behaves as a polyelectrolyte gel: S decreases upon increasing mu both for NaCl and for CaCl(2), with HCl as protonating acid. In aqueous solutions with larger pH, between 4 and 12, the hydrogel is practically neutral, and S increases as mu rises, showing a salting-in effect. From the quantitative analysis of these results, the following facts emerged. Protonation induces chain stiffness (as measured by the non-Gaussian factor) and worsening of the solvent quality of the aqueous media (as measured by the polymer-solvent interaction parameter). For alpha below 33%, swelling seems to be governed by the excess of mobile counterions inside the gel with respect to the bath, with a minor but still significantly negative contribution of the osmotic swelling pressure due to polymer-solvent mixing. Above 33% protonation, it is necessary to consider Manning counterion condensation to get parameters with physical meaning. The crossover between polyelectrolyte and salting-in effects corresponds to alpha and mu values with the same ionic and mixing contributions to the osmotic swelling pressure. The formation of ionic nonpermanent cross-links, with H(2)SO(4) as the protonating acid, was discarded.

  12. Sol-Gel Manufactured Energetic Materials

    DOEpatents

    Simpson, Randall L.; Lee, Ronald S.; Tillotson, Thomas M.; Hrubesh, Lawrence W.; Swansiger, Rosalind W.; Fox, Glenn A.

    2005-05-17

    Sol-gel chemistry is used for the preparation of energetic materials (explosives, propellants and pyrotechnics) with improved homogeneity, and/or which can be cast to near-net shape, and/or made into precision molding powders. The sol-gel method is a synthetic chemical process where reactive monomers are mixed into a solution, polymerization occurs leading to a highly cross-linked three dimensional solid network resulting in a gel. The energetic materials can be incorporated during the formation of the solution or during the gel stage of the process. The composition, pore, and primary particle sizes, gel time, surface areas, and density may be tailored and controlled by the solution chemistry. The gel is then dried using supercritical extraction to produce a highly porous low density aerogel or by controlled slow evaporation to produce a xerogel. Applying stress during the extraction phase can result in high density materials. Thus, the sol-gel method can be used for precision detonator explosive manufacturing as well as producing precision explosives, propellants, and pyrotechnics, along with high power composite energetic materials.

  13. Sol-gel manufactured energetic materials

    DOEpatents

    Simpson, Randall L.; Lee, Ronald S.; Tillotson, Thomas M.; Hrubesh, Lawrence W.; Swansiger, Rosalind W.; Fox, Glenn A.

    2003-12-23

    Sol-gel chemistry is used for the preparation of energetic materials (explosives, propellants and pyrotechnics) with improved homogeneity, and/or which can be cast to near-net shape, and/or made into precision molding powders. The sol-gel method is a synthetic chemical process where reactive monomers are mixed into a solution, polymerization occurs leading to a highly cross-linked three dimensional solid network resulting in a gel. The energetic materials can be incorporated during the formation of the solution or during the gel stage of the process. The composition, pore, and primary particle sizes, gel time, surface areas, and density may be tailored and controlled by the solution chemistry. The gel is then dried using supercritical extraction to produce a highly porous low density aerogel or by controlled slow evaporation to produce a xerogel. Applying stress during the extraction phase can result in high density materials. Thus, the sol-gel method can be used for precision detonator explosive manufacturing as well as producing precision explosives, propellants, and pyrotechnics, along with high power composite energetic materials.

  14. Nano- and microsized cubic gel particles from cyclodextrin metal-organic frameworks.

    PubMed

    Furukawa, Yuki; Ishiwata, Takumi; Sugikawa, Kouta; Kokado, Kenta; Sada, Kazuki

    2012-10-15

    Sweet cube o' mine: Bottom-up control of gel particles has been regarded as a great challenge. By employing internal cross-linking of cyclodextrin metal-organic frameworks, cubic sugar gels were formed with sharp edges that reflect the shape of the crystals. This enabled the fabrication of shape- and size-controlled polymer gels from porous crystals (see picture). Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Absence of skin rash in Goodpasture's syndrome: the hyaluronan effect.

    PubMed

    Stern, A; Stern, R

    2014-12-01

    Goodpasture's syndrome is a rare disease that involves rapidly progressive kidney failure as well as hemorrhagic lung disease. It is a form of autoimmune disorder with unusual features; marked male preponderance, in contrast with other autoimmune disease in which females are at far greater risk. The autoantibodies are directed again the carboxy-extension non-collagenous (NC1) portion of one of the basement membrane-specific collagen IV alpha 3 chains. Basal laminas throughout the body share this structure, including those in kidney, lung and skin. But curiously, skin is rarely involved in Goodpasture's syndrome. Hyaluronan is a large extracellular matrix carbohydrate polymer. Half of total body hyaluronan occurs in skin. High molecular weight hyaluronan, a potent immunosuppressive polymer, might be functioning as an immune shield for skin in Goodpasture's syndrome, and be the basis for the anomaly. A summary of this putative effect is described, including possible molecular mechanisms involved, and suggestions for testing this hypothesis. Copyright © 2014 Elsevier Ltd. All rights reserved.

  16. Nitric oxide-induced interstrand cross-links in DNA.

    PubMed

    Caulfield, Jennifer L; Wishnok, John S; Tannenbaum, Steven R

    2003-05-01

    The DNA damaging effects of nitrous acid have been extensively studied, and the formation of interstrand cross-links have been observed. The potential for this cross-linking to occur through a common nitrosating intermediate derived from nitric oxide is investigated here. Using a HPLC laser-induced fluorescence (LIF) system, the amount of interstrand cross-link formed on nitric oxide treatment of the 5'-fluorescein-labeled oligomer ATATCGATCGATAT was determined. This self-complimentary sequence contains two 5'-CG sequences, which is the preferred site for nitrous acid-induced cross-linking. Nitric oxide was delivered to an 0.5 mM oligomer solution at 15 nmol/mL/min to give a final nitrite concentration of 652 microM. The resulting concentration of the deamination product, xanthine, in this sample was found to be 211 +/- 39 nM, using GC/MS, and the amount of interstrand cross-link was determined to be 13 +/- 2.5 nM. Therefore, upon nitric oxide treatment, the cross-link is found at approximately 6% of the amount of the deamination product. Using this system, detection of the cross-link is also possible for significantly lower doses of nitric oxide, as demonstrated by treatment of the same oligomer with NO at a rate of 18 nmol/mL/min resulting in a final nitrite concentration of 126 microM. The concentration of interstrand cross-link was determined to be 3.6 +/- 0.1 nM in this sample. Therefore, using the same dose rate, when the total nitric oxide concentration delivered drops by a factor of approximately 5, the concentration of cross-link drops by a factor of about 4-indicating a qausi-linear response. It may now be possible to predict the number of cross-links in a small genome based on the number of CpG sequences and the yield of xanthine derived from nitrosative deamination.

  17. Characterization of rat leydig cell gonadotropin receptor structure by affinity cross-linking

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Zhang, Q.Y.; Hwang, J.; Menon, K.M.J.

    1986-05-01

    The gonadotropin receptor from rat leydig cell has been characterized with respect to binding kinetics and physiological regulation. The present study was intended to examine the structure of the receptor. Leydig cell suspension was prepared by either collagenase digestion or by mechanical disruption of the testis. The cells were incubated with /sup 125/I-hCG and the unreacted hCG was removed by centrifugation. The /sup 125/I-hCG was then covalently linked to the cell surface receptor using cleavable (dithiobis (succinimidyl propionate)) and non-cleavable (disuccinimidyl suberate) cross-linking reagents. The extracted cross-linked membrane proteins were resolved on SDS-polyacrylamide gels under reducing and non-reducing conditions andmore » subjected to autoradiographic analysis. Under non-reducing conditions, two labeled species with M/sub r/ = 87,000 and 120,000 were detected. However, only one labeled band was detected under reducing conditions with M/sub r/ = 64,000. The binding of /sup 125/I-hCG to the receptor was inhibited by hCG and LH, but not by a number of peptides and proteins. The data suggest that hCG receptor in leydig cell is an oligomeric complex consisting of four subunits, ..cap alpha cap alpha beta gamma... The ..beta.. and ..gamma.. subunits are each linked to an ..cap alpha.. subunit through disulfide linkage and the hormone binds to each ..cap alpha.. subunit. The two dimers formed (..cap alpha beta cap alpha gamma..) are associated by noncovalent interactions.« less

  18. Identification of mammalian proteins cross-linked to DNA by ionizing radiation.

    PubMed

    Barker, Sharon; Weinfeld, Michael; Zheng, Jing; Li, Liang; Murray, David

    2005-10-07

    Ionizing radiation (IR) is an important environmental risk factor for various cancers and also a major therapeutic agent for cancer treatment. Exposure of mammalian cells to IR induces several types of damage to DNA, including double- and single-strand breaks, base and sugar damage, as well as DNA-DNA and DNA-protein cross-links (DPCs). Little is known regarding the biological consequences of DPCs. Identifying the proteins that become cross-linked to DNA by IR would be an important first step in this regard. We have therefore undertaken a proteomics study to isolate and identify proteins involved in IR-induced DPCs. DPCs were induced in AA8 Chinese hamster ovary or GM00637 human fibroblast cells using 0-4 gray of gamma-rays under either aerated or hypoxic conditions. DPCs were isolated using a recently developed method, and proteins were identified by mass spectrometry. We identified 29 proteins as being cross-linked to DNA by IR under aerated and/or hypoxic conditions. The identified proteins include structural proteins, actin-associated proteins, transcription regulators, RNA-splicing components, stress-response proteins, cell cycle regulatory proteins, and GDP/GTP-binding proteins. The involvement of several proteins (actin, histone H2B, and others) in DPCs was confirmed by using Western blot analysis. The dose responsiveness of DPC induction was examined by staining one-dimensional SDS-polyacrylamide gels with SYPRO Tangerine followed by analysis using fluorescence imaging. Quantitation of the fluorescence signal indicated no significant difference in total yields of IR-induced DPCs generated under aerated or hypoxic conditions, although differences were observed for several individual protein bands.

  19. 21 CFR 177.1211 - Cross-linked polyacrylate copolymers.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... polyacrylate copolymers consist of: (1) The grafted copolymer of cross-linked sodium polyacrylate identified as... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Cross-linked polyacrylate copolymers. 177.1211... Basic Components of Single and Repeated Use Food Contact Surfaces § 177.1211 Cross-linked polyacrylate...

  20. 21 CFR 177.1211 - Cross-linked polyacrylate copolymers.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... polyacrylate copolymers consist of: (1) The grafted copolymer of cross-linked sodium polyacrylate identified as... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Cross-linked polyacrylate copolymers. 177.1211... Basic Components of Single and Repeated Use Food Contact Surfaces § 177.1211 Cross-linked polyacrylate...

  1. Study of interactions between hyaluronan and cationic surfactants by means of calorimetry, turbidimetry, potentiometry and conductometry.

    PubMed

    Krouská, J; Pekař, M; Klučáková, M; Šarac, B; Bešter-Rogač, M

    2017-02-10

    The thermodynamics of the micelle formation of the cationic surfactants tetradecyltrimethylammonium bromide (TTAB) and cetyltrimethylammonium bromide (CTAB) with and without the addition of hyaluronan of two molecular weights was studied in aqueous solution by titration calorimetry. Macroscopic phase separation, which was detected by calorimetry and also by conductometry, occurs when charges on the surfactant and hyaluronan are balanced. In contrast, turbidimetry and potentiometry showed hyaluronan-surfactant interactions at very low surfactant concentrations. The observed differences between systems prepared with CTAB and TTAB indicate that besides the electrostatic interactions, which probably predominate, hydrophobic effects also play a significant role in hyaluronan interactions with cationic surfactants. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Towards aging mechanisms of cross-linked polyethylene (XLPE) cable insulation materials in nuclear power plants

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Liu, Shuaishuai; Fifield, Leonard S.; Bowler, Nicola

    Cross-linked polyethylene (XLPE) cable insulation material undergoes simultaneous, accelerated thermal and gamma-radiation aging to simulate the long-term aging environment within nuclear power plants (NPPs). A variety of materials characterization tests, including scanning electron microscopy, thermo-gravimetric analysis, differential scanning calorimetry, oxidation induction time, gel-fraction and dielectric properties measurement, are conducted on pristine and differently aged XLPE samples. A preliminary model of one possible aging mechanism of XLPE cable insulation material under gamma radiation at elevated temperature of 115 °C is suggested.

  3. Nanoscale supramolecular ordering in gel-surfactant complexes: Sodium alkyl sulfates in poly(diallyldimethylammonium chloride)

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Sokolov, E.L.; Yeh, F.; Khokhlov, A.

    1996-12-25

    Studies of slightly cross-linked polycationic gels interacting with anionic surfactants have been performed by using random copolymers of poly(diallyldimethylammonium chloride) (PDADMACl) and polyacrylamide (PAAm) with varying content of PDADMACl and degree of cross-linking. Gel samples which had been fully swollen in water were placed in aqueous solutions of sodium alkyl sulfates (octyl(SOS), decyl-(SDCS), dodecyl (SDS), tetradecyl (STS), and hexyl (SHS) sulfates). The degree of the sample volume contraction depends on the PDADMACl content. The collapsed gel-surfactant complexes were studied using synchrotron small-angle X-ray scattering. All studied samples containing PDADMACl exhibited pronounced supramolecular nanostructures. The gel-SDCS complex exhibited a cubic structuremore » with a periodicity (7.75 nm) of approximately 4 times the surfactant molecular length, while the gel-SDS, gel-STS, and gel-SHS complexes showed hexagonal supramolecular ordering with a periodicity of approximately 2 times the surfactant molecular length. The d spacing of the longest periodicity in the complexes was dependent on the PDADMACl content and the surfactant. The d spacing generally increased with decreasing PDADMACl (charge) content and increasing number of carbon atoms in the surfactant alkyl chain. 20 refs., 11 figs., 5 tabs.« less

  4. The chain of cross-contamination: link-by-link.

    PubMed

    Schwartz, Jeanne

    2002-01-01

    Conscious efforts must be made to break the chain of cross-contamination--link-by-link. Pay attention to detail Avoid being careless Avoid touching objects while wearing soiled gloves Frequently wash hands (15-second hand washing) to remove pathogens--before and after gloving, and before handling food or drink Wear all PPE, and change accordingly.

  5. Probing vocal fold fibroblast response to hyaluronan in 3D contexts.

    PubMed

    Munoz-Pinto, Dany J; Jimenez-Vergara, Andrea Carolina; Gelves, L Marcela; McMahon, Rebecca E; Guiza-Arguello, Viviana; Hahn, Mariah S

    2009-11-01

    A number of treatments are being investigated for vocal fold (VF) scar, including designer implants. The aim of the present study was to validate a 3D model system for probing the effects of various bioactive moieties on VF fibroblast (VFF) behavior toward rational implant design. We selected poly(ethylene glycol) diacrylate (PEGDA) hydrogels as our base-scaffold due to their broadly tunable material properties. However, since cells encapsulated in PEGDA hydrogels are generally forced to take on rounded/stellate morphologies, validation of PEGDA gels as a 3D VFF model system required that the present work directly parallel previous studies involving more permissive scaffolds. We therefore chose to focus on hyaluronan (HA), a polysaccharide that has been a particular focus of the VF community. Toward this end, porcine VFFs were encapsulated in PEGDA hydrogels containing consistent levels of high Mw HA (HA(HMW)), intermediate Mw HA (HA(IMW)), or the control polysaccharide, alginate, and cultured for 7 and 21 days. HA(HMW) promoted sustained increases in active ERK1/2 relative to HA(IMW). Furthermore, VFFs in HA(IMW) gels displayed a more myofibroblast-like phenotype, higher elastin production, and greater protein kinase C (PkC) levels at day 21 than VFFs in HA(HMW) and alginate gels. The present results are in agreement with a previous 3D study of VFF responses to HA(IMW) relative to alginate in collagen-based scaffolds permissive of cell elongation, indicating that PEGDA hydrogels may serve as an effective 3D model system for probing at least certain aspects of VFF behavior. Copyright 2009 Wiley Periodicals, Inc.

  6. Formation and rupture of Ca(2+) induced pectin biopolymer gels.

    PubMed

    Basak, Rajib; Bandyopadhyay, Ranjini

    2014-10-07

    When calcium salts are added to an aqueous solution of polysaccharide pectin, ionic cross-links form between pectin chains, giving rise to a gel network in dilute solution. In this work, dynamic light scattering (DLS) is employed to study the microscopic dynamics of the fractal aggregates (flocs) that constitute the gels, while rheological measurements are carried out to study the process of gel rupture. As the calcium salt concentration is increased, DLS experiments reveal that the polydispersity of the flocs increase simultaneously with the characteristic relaxation times of the gel network. Above a critical salt concentration, the flocs become interlinked to form a reaction-limited fractal gel network. Rheological studies demonstrate that the limits of the linear rheological response and the critical stresses required to rupture these networks both decrease with the increase in salt concentration. These features indicate that the ion-mediated pectin gels studied here lie in a 'strong link' regime that is characterised by inter-floc links that are stronger than intra-floc links. A scaling analysis of the experimental data presented here demonstrates that the elasticities of the individual fractal flocs exhibit power-law dependences on the added salt concentration. We conclude that when both pectin and salt concentrations are increased, the number of fractal flocs of pectin increases simultaneously with the density of crosslinks, giving rise to very large values of the bulk elastic modulus.

  7. Manufacture of Regularly Shaped Sol-Gel Pellets

    NASA Technical Reports Server (NTRS)

    Leventis, Nicholas; Johnston, James C.; Kinder, James D.

    2006-01-01

    An extrusion batch process for manufacturing regularly shaped sol-gel pellets has been devised as an improved alternative to a spray process that yields irregularly shaped pellets. The aspect ratio of regularly shaped pellets can be controlled more easily, while regularly shaped pellets pack more efficiently. In the extrusion process, a wet gel is pushed out of a mold and chopped repetitively into short, cylindrical pieces as it emerges from the mold. The pieces are collected and can be either (1) dried at ambient pressure to xerogel, (2) solvent exchanged and dried under ambient pressure to ambigels, or (3) supercritically dried to aerogel. Advantageously, the extruded pellets can be dropped directly in a cross-linking bath, where they develop a conformal polymer coating around the skeletal framework of the wet gel via reaction with the cross linker. These pellets can be dried to mechanically robust X-Aerogel.

  8. Apparatus for the production of gel beads containing a biocatalyst

    DOEpatents

    Scott, C.D.; Scott, T.C.; Davison, B.H.

    1998-03-19

    An apparatus is described for the large-scale and continuous production of gel beads containing a biocatalyst. The apparatus is a columnar system based on the chemical cross-linking of hydrocolloidal gels that contain and immobilize a biocatalyst, the biocatalyst being a microorganism or an enzyme. Hydrocolloidal gels, such as alginate, carrageenan, and a mixture of bone gelatin and modified alginate, provide immobilization matrices that can be used to entrap and retain the biocatalyst while allowing effective contact with substrates and release of products. Such immobilized biocatalysts are generally formulated into small spheres or beads that have high concentrations of the biocatalyst within the gel matrix. The columnar system includes a gel dispersion nozzle submerged in a heated non-interacting liquid, typically an organic liquid, that is immiscible with water to allow efficient formation of spherical gel droplets, the non-interacting liquid having a specific gravity that is less than water so that the gel droplets will fall through the liquid by the force of gravity. The heated non-interacting liquid is in direct contact with a chilled upflowing non-interacting liquid that will provide sufficient residence time for the gel droplets as they fall through the liquid so that they will be cooled below the gelling temperature and form solid spheres. The upflowing non-interacting liquid is in direct contact with an upflowing temperature-controlled aqueous solution containing the necessary chemicals for cross-linking or fixing of the gel beads to add the necessary stability. The flow rates of the two liquid streams can be varied to control the proper residence time in each liquid section to accommodate the production of gel beads of differing settling velocities. A valve is provided for continuous removal of the stabilized gel beads from the bottom of the column. 1 fig.

  9. Apparatus for the production of gel beads containing a biocatalyst

    DOEpatents

    Scott, Charles D.; Scott, Timothy C.; Davison, Brian H.

    1998-01-01

    An apparatus for the large-scale and continuous production of gel beads containing a biocatalyst. The apparatus is a columnar system based on the chemical cross-linking of hydrocolloidal gels that contain and immobilize a biocatalyst, the biocatalyst being a microorganism or an enzyme. Hydrocolloidal gels, such as alginate, carrageenan, and a mixture of bone gelatin and modified alginate, provide immobilization matrices that can be used to entrap and retain the biocatalyst while allowing effective contact with substrates and release of products. Such immobilized biocatalysts are generally formulated into small spheres or beads that have high concentrations of the biocatalyst within the gel matrix. The columnar system includes a gel dispersion nozzle submerged in a heated non-interacting liquid, typically an organic liquid, that is immiscible with water to allow efficient formation of spherical gel droplets, the non-interacting liquid having a specific gravity that is less than water so that the gel droplets will fall through the liquid by the force of gravity. The heated non-interacting liquid is in direct contact with a chilled upflowing non-interacting liquid that will provide sufficient residence time for the gel droplets as they fall through the liquid so that they will be cooled below the gelling temperature and form solid spheres. The upflowing non-interacting liquid is in direct contact with an upflowing temperature-controlled aqueous solution containing the necessary chemicals for cross-linking or fixing of the gel beads to add the necessary stability. The flow rates of the two liquid streams can be varied to control the proper residence time in each liquid section to accommodate the production of gel beads of differing settling velocities. A valve is provided for continuous removal of the stabilized gel beads from the bottom of the column.

  10. Enzymatically cross-linked tilapia gelatin hydrogels: physical, chemical, and hybrid networks.

    PubMed

    Bode, Franziska; da Silva, Marcelo Alves; Drake, Alex F; Ross-Murphy, Simon B; Dreiss, Cécile A

    2011-10-10

    This Article investigates different types of networks formed from tilapia fish gelatin (10% w/w) in the presence and absence of the enzymatic cross-linker microbial transglutaminase. The influence of the temperature protocol and cross-linker concentration (0-55 U mTGase/g gelatin) was examined in physical, chemical, and hybrid gels, where physical gels arise from the formation of triple helices that act as junction points when the gels are cooled below the gelation point. A combination of rheology and optical rotation was used to study the evolution of the storage modulus (G') over time and the number of triple helices formed for each type of gel. We attempted to separate the final storage modulus of the gels into its chemical and physical contributions to examine the existence or otherwise of synergism between the two types of networks. Our experiments show that the gel characteristics vary widely with the thermal protocol. The final storage modulus in chemical gels increased with enzyme concentration, possibly due to the preferential formation of closed loops at low cross-linker amount. In chemical-physical gels, where the physical network (helices) was formed consecutively to the covalent one, we found that below a critical enzyme concentration the more extensive the chemical network is (as measured by G'), the weaker the final gel is. The storage modulus attributed to the physical network decreased exponentially as a function of G' from the chemical network, but both networks were found to be purely additive. Helices were not thermally stabilized. The simultaneous formation of physical and chemical networks (physical-co-chemical) resulted in G' values higher than the individual networks formed under the same conditions. Two regimes were distinguished: at low enzyme concentration (10-20 U mTGase/g gelatin), the networks were formed in series, but the storage modulus from the chemical network was higher in the presence of helices (compared to pure chemical gels

  11. Dog erythrocyte antigens 1.1, 1.2, 3, 4, 7, and Dal blood typing and cross-matching by gel column technique.

    PubMed

    Kessler, Rebecca J; Reese, Jessica; Chang, Denise; Seth, Mayank; Hale, Anne S; Giger, Urs

    2010-09-01

    Testing for canine blood types other than dog erythrocyte antigen 1.1 (DEA 1.1) is controversial and complicated by reagent availability and methodology. The objectives of this study were to use available gel column technology to develop an extended blood-typing method using polyclonal reagents for DEA 1.1, 1.2, 3, 4, 7, and Dal and to assess the use of gel columns for cross-matching. Dogs (43-75) were typed for DEA 1.1, 1.2, 3, 4, 7, and Dal. METHODS included tube agglutination (Tube) using polyclonal reagents, a commercially available DEA 1.1 gel column test kit (Standard-Gel) using monoclonal reagent, and multiple gel columns (Extended-Gel) using polyclonal reagents. Blood from 10 recipient and 15 donor dogs was typed as described above and cross-matched using the gel column technique. Of 43 dogs typed for DEA 1.1, 23, 25, and 20 dogs were positive using Standard-Gel, Extended-Gel, and Tube, respectively. Typing for DEA 1.2 was not achievable with Extended-Gel. For 75 dogs typed for DEA 3, 4, and 7, concordance of Extended-Gel with Tube was 94.7%, 100%, and 84%, respectively. Dal, determined only by Extended-Gel, was positive for all dogs. Post-transfusion major cross-matches were incompatible in 10 of 14 pairings, but none were associated with demonstrable blood type incompatibilities. Gel column methodology can be adapted for use with polyclonal reagents for detecting DEA 1.1, 3, 4, 7, and Dal. Agglutination reactions are similar between Extended-Gel and Tube, but are more easily interpreted with Extended-Gel. When using gel columns for cross-matching, incompatible blood cross-matches can be detected following sensitization by transfusion, although in this study incompatibilities associated with any tested DEA or Dal antigens were not found. ©2010 American Society for Veterinary Clinical Pathology.

  12. Donor cross-linking for keratoplasty: a laboratory evaluation.

    PubMed

    Mukherjee, Achyut; Hayes, Sally; Aslanides, Ioannis; Lanchares, Elena; Meek, Keith M

    2015-12-01

    This laboratory-based investigation compares the topographic outcomes of conventional penetrating keratoplasty with that of a novel procedure in which donor corneas are cross-linked prior to keratoplasty. Penetrating keratoplasty procedures with continuous running sutures were carried out in a porcine whole globe model. Sixty eyes were randomly paired as 'donor' and 'host' tissue before being assigned to one of two groups. In the cross-linked group, donor corneas underwent riboflavin/UVA cross-linking prior to being trephined and sutured to untreated hosts. In the conventional keratoplasty group, both host and donor corneas remained untreated prior to keratoplasty. Topographic and corneal wavefront measurements were performed following surgery, and technical aspects of the procedure evaluated. Mean keratometric astigmatism was significantly lower in the cross-linked donor group at 3.67D (SD 1.8 D), vs. 8.43 D (SD 2.4 D) in the conventional keratoplasty group (p < 0.005). Mean wavefront astigmatism was also significantly reduced in the cross-linked donor group 4.71 D (SD 2.1) vs. 8.29D (SD 3.6) in the conventional keratoplasty group (p < 0.005). Mean RMS higher order aberration was significantly lower in the cross-linked donor group at 1.79 um (SD 0.98), vs. 3.05 um (SD 1.9) in the conventional keratoplasty group (P = 0.02). Qualitative analysis revealed less tissue distortion at the graft-host junction in the cross-linked group. Cross-linking of donor corneas prior to keratoplasty reduces intraoperative induced astigmatism and aberrations in an animal model. Further studies are indicated to evaluate the implications of this potential modification of keratoplasty surgery.

  13. Examination of synovial fluid hyaluronan quantity and quality in stifle joints of dogs with osteoarthritis.

    PubMed

    Venable, Rachel O; Stoker, Aaron M; Cook, Cristi R; Cockrell, Mary K; Cook, James L

    2008-12-01

    To determine the quantity (concentration) and quality (molecular weight) of synovial fluid hyaluronan with respect to presence and severity of osteoarthritis in stifle joints of dogs. 21 purpose-bred dogs and 6 clinically affected large-breed dogs (cranial cruciate ligament [CrCL] disease with secondary osteoarthritis). Research dogs underwent arthroscopic surgery in 1 stifle joint to induce osteoarthritis via CrCL transection (CrCLt; n=5 stifle joints), femoral condylar articular cartilage groove creation (GR; 6), or meniscal release (MR; 5); 5 had sham surgery (SH) performed. Contralateral stifle joints (n=21) were used as unoperated control joints. Synovial fluid was obtained from research dogs at time 0 and 12 weeks after surgery and from clinically affected dogs prior to surgery. All dogs were assessed for lameness, radiographic signs of osteoarthritis, and pathologic findings on arthroscopy as well as for quantity and quality of hyaluronan. Clinically affected dogs had significantly greater degrees of pathologic findings, compared with dogs with surgically induced osteoarthritis (ie, those with CrCLt, GR, and MR stifle joints), and with respect to lameness scores, radiographic signs of osteoarthritis, pathologic findings on arthroscopy, and synovial fluid hyaluronan concentration. Synovial fluid from stifle joints of dogs with surgically induced osteoarthritis had hyaluronan bands at 35 kd on western blots that synovial fluid from SH and clinically affected stifle joints did not. Synovial fluid hyaluronan quantity and quality were altered in stifle joints of dogs with osteoarthritis, compared with control stifle joints. A specific hyaluronan protein fragment may be associated with early pathologic changes in affected joints.

  14. Characterization of the Deoxyguanosine–Lysine Cross-Link of Methylglyoxal

    PubMed Central

    2015-01-01

    Methylglyoxal is a mutagenic bis-electrophile that is produced endogenously from carbohydrate precursors. Methylglyoxal has been reported to induce DNA–protein cross-links (DPCs) in vitro and in cultured cells. Previous work suggests that these cross-links are formed between guanine and either lysine or cysteine side chains. However, the chemical nature of the methylglyoxal induced DPC have not been determined. We have examined the reaction of methylglyoxal, deoxyguanosine (dGuo), and Nα-acetyllysine (AcLys) and determined the structure of the cross-link to be the N2-ethyl-1-carboxamide with the lysine side chain amino group (1). The cross-link was identified by mass spectrometry and the structure confirmed by comparison to a synthetic sample. Further, the cross-link between methylglyoxal, dGuo, and a peptide (AcAVAGKAGAR) was also characterized. The mechanism of cross-link formation is likely to involve an Amadori rearrangement. PMID:24801980

  15. Photocontrolled Cargo Release from Dual Cross-Linked Polymer Particles.

    PubMed

    Tan, Shereen; Cui, Jiwei; Fu, Qiang; Nam, Eunhyung; Ladewig, Katharina; Ren, Jing M; Wong, Edgar H H; Caruso, Frank; Blencowe, Anton; Qiao, Greg G

    2016-03-09

    Burst release of a payload from polymeric particles upon photoirradiation was engineered by altering the cross-linking density. This was achieved via a dual cross-linking concept whereby noncovalent cross-linking was provided by cyclodextrin host-guest interactions, and irreversible covalent cross-linking was mediated by continuous assembly of polymers (CAP). The dual cross-linked particles (DCPs) were efficiently infiltrated (∼80-93%) by the biomacromolecule dextran (molecular weight up to 500 kDa) to provide high loadings (70-75%). Upon short exposure (5 s) to UV light, the noncovalent cross-links were disrupted resulting in increased permeability and burst release of the cargo (50 mol % within 1 s) as visualized by time-lapse fluorescence microscopy. As sunlight contains UV light at low intensities, the particles can potentially be incorporated into systems used in agriculture, environmental control, and food packaging, whereby sunlight could control the release of nutrients and antimicrobial agents.

  16. Cross-linked polyvinyl alcohol and method of making same

    NASA Technical Reports Server (NTRS)

    Hsu, L. C.; Sheibley, D. W.; Philipp, W. H. (Inventor)

    1981-01-01

    A film-forming polyvinyl alcohol polymer is mixed with a polyaldehyde-polysaccharide cross-linking agent having at least two monosaccharide units and a plurality of aldehyde groups per molecule, perferably an average of at least one aldehyde group per monosaccharide units. The cross-linking agent, such as a polydialdehyde starch, is used in an amount of about 2.5 to 20% of the theoretical amount required to cross-link all of the available hydroxyl groups of the polyvinyl alcohol polymer. Reaction between the polymer and cross-linking agent is effected in aqueous acidic solution to produce the cross-linked polymer. The polymer product has low electrical resistivity and other properties rendering it suitable for making separators for alkaline batteries.

  17. Structuring of Amide Cross-Linked Non-Bridged and Bridged Alkyl-Based Silsesquioxanes.

    PubMed

    Nunes, S C; de Zea Bermudez, V

    2018-02-06

    The development of sophisticated organized materials exhibiting enhanced properties is a challenging topic of the domain of organic/inorganic hybrid materials. This review, composed of four sections, reports the work we have carried out over the last 10 years on the synthesis of amide cross-linked alkyl/siloxane hybrids by means of sol-gel chemistry and self-directed assembly/self-organization routes relying on weak interactions (hydrophobic interactions and hydrogen bonding). The various as-produced lamellar structures displaying a myriad of morphologies, often closely resembling those found in natural materials, are discussed. The major role played by the synthetic conditions (pH, water content, co-solvent(s) nature/concentration and dopant presence/concentration), the alkyl chains (length and presence of ramification or not) and the number of the amide cross-links present in the precursor, is evidenced. Examples of highly organized hybrids structures incorporating ionic species (alkali and alkaline earth metal salts) and optically-active centers (organic dyes and lanthanide ions) are described. A useful qualitative relationship deduced between the emission quantum yield of the ordered hybrid materials and the degree of order of the hydrogen-bonded network is highlighted. © 2018 The Chemical Society of Japan & Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Maize arabinoxylan gels as protein delivery matrices.

    PubMed

    Berlanga-Reyes, Claudia M; Carvajal-Millán, Elizabeth; Lizardi-Mendoza, Jaime; Rascón-Chu, Agustin; Marquez-Escalante, Jorge A; Martínez-López, Ana Luisa

    2009-04-08

    The laccase induced gelation of maize bran arabinoxylans at 2.5% (w/v) in the presence of insulin or beta-lactoglobulin at 0.1% (w/v) was investigated. Insulin and beta-lacto-globulin did not modify either the gel elasticity (9 Pa) or the cross-links content (0.03 and 0.015 microg di- and triferulic acids/mg arabinoxylan, respectively). The protein release capability of the gel was also investigated. The rate of protein release from gels was dependent on the protein molecular weight. The apparent diffusion coefficient was 0.99 x 10(-7) and 0.79 x 10(-7) cm(2)/s for insulin (5 kDa) and beta-lactoglobulin (18 kDa), respectively. The results suggest that maize bran arabinoxylan gels can be potential candidates for the controlled release of proteins.

  19. Characterization of Hyaluronan-Degrading Enzymes from Yeasts.

    PubMed

    Smirnou, Dzianis; Krčmář, Martin; Kulhánek, Jaromír; Hermannová, Martina; Bobková, Lenka; Franke, Lukáš; Pepeliaev, Stanislav; Velebný, Vladimír

    2015-10-01

    Hyaluronidases (HAases) from yeasts were characterized for the first time. The study elucidated that hyaluronate 4-glycanohydrolase and hyaluronan (HA) lyase can be produced by yeasts. Six yeasts producing HAases were found through express screening of activities. The extracellular HAases from two of the yeast isolates, Pseudozyma aphidis and Cryptococcus laurentii, were characterized among them. P. aphidis HAase hydrolyzed β-1,4 glycosidic bonds of HA, yielding even-numbered oligosaccharides with N-acetyl-D-glucosamine at the reducing end. C. laurentii produced hyaluronan lyase, which cleaved β-1,4 glycosidic bonds of HA in β-elimination reaction, and the products of HA degradation were different-sized even-numbered oligosaccharides. The shortest detected HA oligomer was dimer. The enzymes' pH and temperature optima were pH 3.0 and 37-45 °C (P. aphidis) and pH 6.0 and 37 °C (C. laurentii), respectively. Both HAases showed good thermostability.

  20. Cross-linking and the molecular packing of corneal collagen

    NASA Technical Reports Server (NTRS)

    Yamauchi, M.; Chandler, G. S.; Tanzawa, H.; Katz, E. P.

    1996-01-01

    We have quantitatively characterized, for the first time, the cross-linking in bovine cornea collagen as a function of age. The major iminium reducible cross-links were dehydro-hydroxylysinonorleucine (deH-HLNL) and dehydro-histidinohydroxymerodesmosine (deH-HHMD). The former rapidly diminished after birth; however, the latter persisted in mature animals at a level of 0.3 - 0.4 moles/mole of collagen. A nonreducible cross-link, histidinohydroxylysinonorleucine (HHL), previously found only in skin, was also found to be a major mature cross-link in cornea. The presence of HHL indicates that cornea fibrils have a molecular packing similar to skin collagen. However, like deH-HHMD, the HHL content in corneal fibrils only reaches a maximum value with time about half that of skin. These data suggest that the corneal fibrils are comprised of discrete filaments that are internally stabilized by HHL and deH-HHMD cross-links. This pattern of intermolecular cross-linking would facilitate the special collagen swelling property required for corneal transparency.

  1. Thiolated hyaluronan-based hydrogels crosslinked using oxidized glutathione: an injectable matrix designed for ophthalmic applications.

    PubMed

    Zarembinski, Thomas I; Doty, Nathaniel J; Erickson, Isaac E; Srinivas, Ramya; Wirostko, Barbara M; Tew, William P

    2014-01-01

    Future ophthalmic therapeutics will require the sustained delivery of bioactive proteins and nucleic acid-based macromolecules and/or provide a suitable microenvironment for the localization and sustenance of reparative progenitor cells after transplantation into or onto the eye. Water-rich hydrogels are ideal vehicles for such cargo, but few have all the qualities desired for novel ophthalmic use, namely in situ gelation speed, cytocompatibility, biocompatibility and capacity to functionalize. We describe here the development of an ophthalmic-compatible crosslinking system using oxidized glutathione (GSSG), a physiologically relevant molecule with a history of safe use in humans. When GSSG is used in conjunction with an existing hyaluronate-based, in situ crosslinkable hydrogel platform, gels form in less than 5 min using the thiol-disulfide exchange reaction. This GSSG hydrogel supports the 3-D culture of adipose-derived stem cells in vitro and shows biocompatibility in preliminary intracutaneous and subconjunctival experiments in vivo. In addition, the thiol-disulfide exchange reaction can also be used in conjunction with other thiol-compatible chemistries to covalently link peptides for more complex formulations. These data suggest that this hydrogel could be well suited for local ocular delivery, focusing initially on front of the eye therapies. Subsequent uses of the hydrogel include delivery of back of the eye treatments and eventually into other soft, hyaluronan-rich tissues such as those from the liver and brain. Copyright © 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  2. DNA cross-linking by dehydromonocrotaline lacks apparent base sequence preference.

    PubMed

    Rieben, W Kurt; Coulombe, Roger A

    2004-12-01

    Pyrrolizidine alkaloids (PAs) are ubiquitous plant toxins, many of which, upon oxidation by hepatic mixed-function oxidases, become reactive bifunctional pyrrolic electrophiles that form DNA-DNA and DNA-protein cross-links. The anti-mitotic, toxic, and carcinogenic action of PAs is thought to be caused, at least in part, by these cross-links. We wished to determine whether the activated PA pyrrole dehydromonocrotaline (DHMO) exhibits base sequence preferences when cross-linked to a set of model duplex poly A-T 14-mer oligonucleotides with varying internal and/or end 5'-d(CG), 5'-d(GC), 5'-d(TA), 5'-d(CGCG), or 5'-d(GCGC) sequences. DHMO-DNA cross-links were assessed by electrophoretic mobility shift assay (EMSA) of 32P endlabeled oligonucleotides and by HPLC analysis of cross-linked DNAs enzymatically digested to their constituent deoxynucleosides. The degree of DNA cross-links depended upon the concentration of the pyrrole, but not on the base sequence of the oligonucleotide target. Likewise, HPLC chromatograms of cross-linked and digested DNAs showed no discernible sequence preference for any nucleotide. Added glutathione, tyrosine, cysteine, and aspartic acid, but not phenylalanine, threonine, serine, lysine, or methionine competed with DNA as alternate nucleophiles for cross-linking by DHMO. From these data it appears that DHMO exhibits no strong base preference when forming cross-links with DNA, and that some cellular nucleophiles can inhibit DNA cross-link formation.

  3. Characterization of the effect of high molecular weight hyaluronan on trans-synovial flow in rabbit knees.

    PubMed

    Coleman, P J; Scott, D; Mason, R M; Levick, J R

    1999-01-01

    1. The effect of a rooster comb hyaluronan (3.6-4.0 g l-1) of similar chain length to rabbit synovial fluid hyaluronan, on the trans-synovial escape of fluid from the joint cavity in the steady state ( 8d s) was studied in 29 rabbit knees at controlled intra-articular pressures (Pj). 2. Rooster hyaluronan caused the pressure-flow relation to flatten out as pressure was raised. At 10-20 cmH2O the slope of the quasi-plateau, 0.05 +/- 0.01 microliter min-1 cmH2O-1 (mean +/- s.e.m.), was 1/39th that for Ringer solution (1.94 +/- 0.01 microliter 2O-1 ). 3. Bovine synovial fluid had a similar effect to hyaluronan in Ringer solution. 4. The quasi-plateau was caused by increasing opposition to outflow; the pressure required to drive unit outflow increased 4.4-fold between 5 and 20 cmH2O. The increased opposition to outflow at 20 cmH2O was equivalent to an effective osmotic pressure of 13-17 cmH2O at the interface. Since the infusate's osmotic pressure was only 0.9 cmH2O, this implied concentration polarization to 15-18 g l-1 hyaluronan at the interface. 5. Mechanical perforation of the lining, or enzymatic degradation of the interstitial matrix by chymopapain, abolished the quasi-plateau. Hydrational expansion of the matrix by approximately 2-fold did not. The increased opposition to outflow was reversible by washing out the hyaluronan, or by reducing Pj. It was unaffected by interruption of tissue blood flow or synoviocyte oxidative metabolism. These properties are compatible with a concentration polarization mechanism, i.e. flow-induced concentration of hyaluronan at the synovial interface due to molecular reflection. 6. A concentration polarization theory was developed for a partially reflected solute. Numerical solutions supported the feasibility of this osmotic explanation of the quasi-plateau. Additional mechanisms may also be involved. 7. It is concluded that native-size hyaluronan helps to retain synovial fluid in the joint cavity when pressure is raised and acts

  4. Characterization of the effect of high molecular weight hyaluronan on trans-synovial flow in rabbit knees

    PubMed Central

    Coleman, P J; Scott, D; Mason, R M; Levick, J R

    1999-01-01

    The effect of a rooster comb hyaluronan (3.6–4.0 g l−1) of similar chain length to rabbit synovial fluid hyaluronan, on the trans-synovial escape of fluid from the joint cavity in the steady state (Q̇s) was studied in 29 rabbit knees at controlled intra-articular pressures (Pj).Rooster hyaluronan caused the pressure-flow relation to flatten out as pressure was raised. At 10–20 cmH2O the slope of the quasi-plateau, 0.05 ± 0.01 μl min−1 cmH2O−1 (mean ±s.e.m.), was 1/39th that for Ringer solution (1.94 ± 0.01 μl min−1 cmH2O−1).Bovine synovial fluid had a similar effect to hyaluronan in Ringer solution.The quasi-plateau was caused by increasing opposition to outflow; the pressure required to drive unit outflow increased 4.4-fold between 5 and 20 cmH2O. The increased opposition to outflow at 20 cmH2O was equivalent to an effective osmotic pressure of 13–17 cmH2O at the interface. Since the infusate's osmotic pressure was only 0.9 cmH2O, this implied concentration polarization to 15–18 g l−1 hyaluronan at the interface.Mechanical perforation of the lining, or enzymatic degradation of the interstitial matrix by chymopapain, abolished the quasi-plateau. Hydrational expansion of the matrix by /2-fold did not. The increased opposition to outflow was reversible by washing out the hyaluronan, or by reducing Pj. It was unaffected by interruption of tissue blood flow or synoviocyte oxidative metabolism. These properties are compatible with a concentration polarization mechanism, i.e. flow-induced concentration of hyaluronan at the synovial interface due to molecular reflection.A concentration polarization theory was developed for a partially reflected solute. Numerical solutions supported the feasibility of this osmotic explanation of the quasi-plateau. Additional mechanisms may also be involved.It is concluded that native-size hyaluronan helps to retain synovial fluid in the joint cavity when pressure is raised and acts, at least in part, by exerting

  5. Cross-linked hyaluronic acid in pressure ulcer prevention.

    PubMed

    Beniamino, P; Vadalà, M; Laurino, C

    2016-07-02

    Long-term bedridden patients are at high risk of acquring pressure ulcers (PUs). In this group of patients, prevention is necessary to cut the health costs, improve quality of life and reduce the mortality. Here, we evaluated the effectiveness of a cross-linked hyaluronic acid (HA) as plastic bulking-agent filling and remodelling the deep dermis and subcutaneous space of the skin areas exposed to the risk of necrosis. Our work hypothesis has been to inflate a sub-dermal elastic cushion, filled with a natural ECM component, with the aim to induce a stronger tissue background resistant to the ulcerative process. All the patients had an increased risk of PUs, at the sacral, ileum or heel skin. Patients were being nursed accordingly to the standard orthopaedic ward management with a pressure relieveing air mattress. The standard protocol consisted in body mobilisation every 3 hours, 24 hours a day and accurate cleaning of the skin with liquid soap and water without any towel friction and without adding any cream or lotion for the skin protection. Our filling protocol enclosed: accurate disinfection of the skin to be injected with povidone-iodine solution, followed by a local anaesthesia with 28G 13 mm needle, injecting 1.5 ml of 1% xylocaine. Then slow, deep, subcutaneous injection of cross-linked HA was performed with a 18G long needle, in order to deliver a homogeneous, soft gel layer underneath and around the whitish erythematous skin edges at risk of ulceration. Patients' tolerability of the compound and adverse events were also recorded. There were 15 patients (78-94 years old) who participated in the study. All tolerated the procedure very well and no serious side effects were declared. No skin pressure ulceration was detected in the four weeks follow-up Conclusion: We have demonstrated the safety and tolerability of a cross-linked HA subdermal injection in PUs prevention. The compound stratifies in a soft, elastic, interstitial bulk into the deep dermis, thus

  6. Tissue simulating gel for medical research

    NASA Technical Reports Server (NTRS)

    Companion, John A. (Inventor)

    1989-01-01

    A tissue simulating gel and a method for preparing the tissue simulating gel are disclosed. The tissue simulating gel is prepared by a process using water, gelatin, ethylene gylcol, and a cross-linking agent. In order to closely approximate the characteristics of the type of tissue being simulated, other material has been added to change the electrical, sound conducting, and wave scattering properties of the tissue simulating gel. The result of the entire process is a formulation that will not melt at the elevated temperatures involved in hyperthermia medical research. Furthermore, the tissue simulating gel will not support mold or bacterial growth, is of a sufficient mechanical strength to maintain a desired shape without a supporting shell, and is non-hardening and non-drying. Substances were injected into the tissue simulating gel prior to the setting-up thereof just as they could be injected into actual tissue, and the tissue simulating gel is translucent so as to permit visual inspection of its interior. A polyurethane spray often used for coating circuit boards can be applied to the surface of the tissue simulating gel to give a texture similar to human skin, making the tissue simulating gel easier to handle and contributing to its longevity.

  7. Tissue simulating gel for medical research

    NASA Technical Reports Server (NTRS)

    Companion, John A. (Inventor)

    1991-01-01

    A tissue simulating gel and a method for preparing the tissue simulating gel are disclosed. The tissue simulating gel is prepared by a process using water, gelatin, ethylene glycol, and a cross-linking agent. In order to closely approximate the characteristics of the type of tissue being simulated, other material has been added to change the electrical, sound conducting, and wave scattering properties of the tissue simulating gel. The result of the entire process is a formulation that will not melt at the elevated temperatures involved in hyperthermia medical research. Furthermore, the tissue simulating gel will not support mold or bacterial growth, is of a sufficient mechanical strength to maintain a desired shape without a supporting shell, and is non-hardening and non-drying. Substances have been injected into the tissue simulating gel prior to the setting-up thereof just as they could be injected into actual tissue, and the tissue simulating gel is translucent so as to permit visual inspection of its interior. A polyurethane spray often used for coating circuit boards can be applied to the surface of the tissue simulating gel to give a texture similar to human skin, making the tissue simulating gel easier to handle and contributing to its longevity.

  8. UV laser-induced cross-linking in peptides

    PubMed Central

    Leo, Gabriella; Altucci, Carlo; Bourgoin-Voillard, Sandrine; Gravagnuolo, Alfredo M.; Esposito, Rosario; Marino, Gennaro; Costello, Catherine E.; Velotta, Raffaele; Birolo, Leila

    2013-01-01

    RATIONALE The aim of this study was to demonstrate, and to characterize by high resolution mass spectrometry, that it is possible to preferentially induce covalent cross-links in peptides by using high energy femtosecond UV laser pulses. The cross-link is readily formed only when aromatic amino acids are present in the peptide sequence. METHODS Three peptides, xenopsin, angiotensin I, interleukin, individually or in combination, were exposed to high energy femtosecond UV laser pulses, either alone or in the presence of spin trapping molecules, the reaction products being characterized by high resolution mass spectrometry. RESULTS High resolution mass spectrometry and spin trapping strategies showed that cross-linking occurs readily, proceeds via a radical mechanism, and is the highly dominant reaction, proceeding without causing significant photo-damage in the investigated range of experimental parameters. CONCLUSIONS High energy femtosecond UV laser pulses can be used to induce covalent cross-links between aromatic amino acids in peptides, overcoming photo-oxidation processes, that predominate as the mean laser pulse intensity approaches illumination conditions achievable with conventional UV light sources. PMID:23754800

  9. Multiscale mechanical effects of native collagen cross-linking in tendon.

    PubMed

    Eekhoff, Jeremy D; Fang, Fei; Lake, Spencer P

    2018-06-06

    The hierarchical structure of tendon allows for attenuation of mechanical strain down decreasing length scales. While reorganization of collagen fibers accounts for microscale strain attenuation, cross-linking between collagen molecules contributes to deformation mechanisms at the fibrillar and molecular scales. Divalent and trivalent enzymatic cross-links form during the development of collagen fibrils through the enzymatic activity of lysyl oxidase (LOX). By establishing connections between telopeptidyl and triple-helical domains of adjacent molecules within collagen fibrils, these cross-links stiffen the fibrils by resisting intermolecular sliding. Ultimately, greater enzymatic cross-linking leads to less compliant and stronger tendon as a result of stiffer fibrils. In contrast, nonenzymatic cross-links such as glucosepane and pentosidine are not produced during development but slowly accumulate through glycation of collagen. Therefore, these cross-links are only expected to be present in significant quantities in advanced age, where there has been sufficient time for glycation to occur, and in diabetes, where the presence of more free sugar in the extracellular matrix increases the rate of glycation. Unlike enzymatic cross-links, current evidence suggests that nonenzymatic cross-links are at least partially isolated to the surface of collagen fibers. As a result, glycation has been proposed to primarily impact tendon mechanics by altering molecular interactions at the fiber interface, thereby diminishing sliding between fibers. Thus, increased nonenzymatic cross-linking decreases microscale strain attenuation and the viscous response of tendon. In conclusion, enzymatic and nonenzymatic collagen cross-links have demonstrable and distinct effects on the mechanical properties of tendon across different length scales.

  10. Living additive manufacturing: Transformation of parent gels into diversely functionalized daughter gels made possible by visible light photoredox catalysis

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Chen, Mao; Gu, Yuwei; Singh, Awaneesh

    Light-initiated additive manufacturing techniques typically rely on layer-by-layer addition or continuous extraction of polymers formed via nonliving, free radical polymerization methods that render the final materials “dead” toward further monomer insertion; the polymer chains within the materials cannot be reactivated to induce chain extension. An alternative “living additive manufacturing” strategy would involve the use of photocontrolled living radical polymerization to spatiotemporally insert monomers into dormant “parent” materials to generate more complex and diversely functionalized “daughter” materials. Here, we demonstrate a proof-of-concept study of living additive manufacturing using end-linked polymer gels embedded with trithiocarbonate iniferters that can be activated by photoinducedmore » single-electron transfer from an organic photoredox catalyst in solution. This system enables the synthesis of a wide range of chemically and mechanically differentiated daughter gels from a single type of parent gel via light-controlled modification of the parent’s average composition, strand length, and/or cross-linking density. Furthermore, daughter gels that are softer than their parent, stiffer than their parent, larger but with the same modulus as their parent, thermally responsive, polarity responsive, healable, and weldable are all realized.« less

  11. Living Additive Manufacturing: Transformation of Parent Gels into Diversely Functionalized Daughter Gels Made Possible by Visible Light Photoredox Catalysis

    PubMed Central

    2017-01-01

    Light-initiated additive manufacturing techniques typically rely on layer-by-layer addition or continuous extraction of polymers formed via nonliving, free radical polymerization methods that render the final materials “dead” toward further monomer insertion; the polymer chains within the materials cannot be reactivated to induce chain extension. An alternative “living additive manufacturing” strategy would involve the use of photocontrolled living radical polymerization to spatiotemporally insert monomers into dormant “parent” materials to generate more complex and diversely functionalized “daughter” materials. Here, we demonstrate a proof-of-concept study of living additive manufacturing using end-linked polymer gels embedded with trithiocarbonate iniferters that can be activated by photoinduced single-electron transfer from an organic photoredox catalyst in solution. This system enables the synthesis of a wide range of chemically and mechanically differentiated daughter gels from a single type of parent gel via light-controlled modification of the parent’s average composition, strand length, and/or cross-linking density. Daughter gels that are softer than their parent, stiffer than their parent, larger but with the same modulus as their parent, thermally responsive, polarity responsive, healable, and weldable are all realized. PMID:28280779

  12. Living Additive Manufacturing: Transformation of Parent Gels into Diversely Functionalized Daughter Gels Made Possible by Visible Light Photoredox Catalysis.

    PubMed

    Chen, Mao; Gu, Yuwei; Singh, Awaneesh; Zhong, Mingjiang; Jordan, Alex M; Biswas, Santidan; Korley, LaShanda T J; Balazs, Anna C; Johnson, Jeremiah A

    2017-02-22

    Light-initiated additive manufacturing techniques typically rely on layer-by-layer addition or continuous extraction of polymers formed via nonliving, free radical polymerization methods that render the final materials "dead" toward further monomer insertion; the polymer chains within the materials cannot be reactivated to induce chain extension. An alternative "living additive manufacturing" strategy would involve the use of photocontrolled living radical polymerization to spatiotemporally insert monomers into dormant "parent" materials to generate more complex and diversely functionalized "daughter" materials. Here, we demonstrate a proof-of-concept study of living additive manufacturing using end-linked polymer gels embedded with trithiocarbonate iniferters that can be activated by photoinduced single-electron transfer from an organic photoredox catalyst in solution. This system enables the synthesis of a wide range of chemically and mechanically differentiated daughter gels from a single type of parent gel via light-controlled modification of the parent's average composition, strand length, and/or cross-linking density. Daughter gels that are softer than their parent, stiffer than their parent, larger but with the same modulus as their parent, thermally responsive, polarity responsive, healable, and weldable are all realized.

  13. Living additive manufacturing: Transformation of parent gels into diversely functionalized daughter gels made possible by visible light photoredox catalysis

    DOE PAGES

    Chen, Mao; Gu, Yuwei; Singh, Awaneesh; ...

    2017-01-13

    Light-initiated additive manufacturing techniques typically rely on layer-by-layer addition or continuous extraction of polymers formed via nonliving, free radical polymerization methods that render the final materials “dead” toward further monomer insertion; the polymer chains within the materials cannot be reactivated to induce chain extension. An alternative “living additive manufacturing” strategy would involve the use of photocontrolled living radical polymerization to spatiotemporally insert monomers into dormant “parent” materials to generate more complex and diversely functionalized “daughter” materials. Here, we demonstrate a proof-of-concept study of living additive manufacturing using end-linked polymer gels embedded with trithiocarbonate iniferters that can be activated by photoinducedmore » single-electron transfer from an organic photoredox catalyst in solution. This system enables the synthesis of a wide range of chemically and mechanically differentiated daughter gels from a single type of parent gel via light-controlled modification of the parent’s average composition, strand length, and/or cross-linking density. Furthermore, daughter gels that are softer than their parent, stiffer than their parent, larger but with the same modulus as their parent, thermally responsive, polarity responsive, healable, and weldable are all realized.« less

  14. Wear effects on microscopic morphology and hyaluronan uptake in siloxane-hydrogel contact lenses.

    PubMed

    Tavazzi, Silvia; Tonveronachi, Martina; Fagnola, Matteo; Cozza, Federica; Ferraro, Lorenzo; Borghesi, Alessandro; Ascagni, Miriam; Farris, Stefano

    2015-07-01

    The purpose of this study was a comparison between new and worn siloxane-hydrogel contact lenses in terms of microscopic structure, surface morphology, and loading of hyaluronan. The analyses were performed by scanning electron microscopy, with the support of the freeze-drying technique, and by fluorescence confocal microscopy. Along the depth profile of new lenses, a thin porous top layer was observed, which corresponds to the region of hyaluronan penetration inside well-defined channels. The time evolution was followed from one day to two weeks of daily wear, when a completely different scenario was found. Clear experimental evidence of a buggy surface was observed with several crests and regions of swelling, which could be filled by the hyaluronan solution. The modifications are attributed to the progressive relaxation of the structure of the polymeric network. © 2014 Wiley Periodicals, Inc.

  15. A Structural Approach to Establishing a Platform Chemistry for the Tunable, Bulk Electron Beam Cross-Linking of Shape Memory Polymer Systems

    PubMed Central

    Hearon, Keith; Besset, Celine J.; Lonnecker, Alexander T.; Ware, Taylor; Voit, Walter E.; Wilson, Thomas S.; Wooley, Karen L.; Maitland, Duncan J.

    2014-01-01

    The synthetic design and thermomechanical characterization of shape memory polymers (SMPs) built from a new polyurethane chemistry that enables facile, bulk and tunable cross-linking of low-molecular weight thermoplastics by electron beam irradiation is reported in this study. SMPs exhibit stimuli-induced geometry changes and are being proposed for applications in numerous fields. We have previously reported a polyurethane SMP system that exhibits the complex processing capabilities of thermoplastic polymers and the mechanical robustness and tunability of thermomechanical properties that are often characteristic of thermoset materials. These previously reported polyurethanes suffer practically because the thermoplastic molecular weights needed to achieve target cross-link densities severely limit high-throughput thermoplastic processing and because thermally unstable radiation-sensitizing additives must be used to achieve high enough cross-link densities to enable desired tunable shape memory behavior. In this study, we demonstrate the ability to manipulate cross-link density in low-molecular weight aliphatic thermoplastic polyurethane SMPs (Mw as low as ~1.5 kDa) without radiation-sensitizing additives by incorporating specific structural motifs into the thermoplastic polymer side chains that we hypothesized would significantly enhance susceptibility to e-beam cross-linking. A custom diol monomer was first synthesized and then implemented in the synthesis of neat thermoplastic polyurethane SMPs that were irradiated at doses ranging from 1 to 500 kGy. Dynamic mechanical analysis (DMA) demonstrated rubbery moduli to be tailorable between 0.1 and 55 MPa, and both DMA and sol/gel analysis results provided fundamental insight into our hypothesized mechanism of electron beam cross-linking, which enables controllable bulk cross-linking to be achieved in highly processable, low-molecular weight thermoplastic shape memory polymers without sensitizing additives. PMID

  16. Modeling Protein Excited-state Structures from "Over-length" Chemical Cross-links.

    PubMed

    Ding, Yue-He; Gong, Zhou; Dong, Xu; Liu, Kan; Liu, Zhu; Liu, Chao; He, Si-Min; Dong, Meng-Qiu; Tang, Chun

    2017-01-27

    Chemical cross-linking coupled with mass spectroscopy (CXMS) provides proximity information for the cross-linked residues and is used increasingly for modeling protein structures. However, experimentally identified cross-links are sometimes incompatible with the known structure of a protein, as the distance calculated between the cross-linked residues far exceeds the maximum length of the cross-linker. The discrepancies may persist even after eliminating potentially false cross-links and excluding intermolecular ones. Thus the "over-length" cross-links may arise from alternative excited-state conformation of the protein. Here we present a method and associated software DynaXL for visualizing the ensemble structures of multidomain proteins based on intramolecular cross-links identified by mass spectrometry with high confidence. Representing the cross-linkers and cross-linking reactions explicitly, we show that the protein excited-state structure can be modeled with as few as two over-length cross-links. We demonstrate the generality of our method with three systems: calmodulin, enzyme I, and glutamine-binding protein, and we show that these proteins alternate between different conformations for interacting with other proteins and ligands. Taken together, the over-length chemical cross-links contain valuable information about protein dynamics, and our findings here illustrate the relationship between dynamic domain movement and protein function. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  17. Xilmass: A New Approach toward the Identification of Cross-Linked Peptides.

    PubMed

    Yılmaz, Şule; Drepper, Friedel; Hulstaert, Niels; Černič, Maša; Gevaert, Kris; Economou, Anastassios; Warscheid, Bettina; Martens, Lennart; Vandermarliere, Elien

    2016-10-18

    Chemical cross-linking coupled with mass spectrometry plays an important role in unravelling protein interactions, especially weak and transient ones. Moreover, cross-linking complements several structural determination approaches such as cryo-EM. Although several computational approaches are available for the annotation of spectra obtained from cross-linked peptides, there remains room for improvement. Here, we present Xilmass, a novel algorithm to identify cross-linked peptides that introduces two new concepts: (i) the cross-linked peptides are represented in the search database such that the cross-linking sites are explicitly encoded, and (ii) the scoring function derived from the Andromeda algorithm was adapted to score against a theoretical tandem mass spectrometry (MS/MS) spectrum that contains the peaks from all possible fragment ions of a cross-linked peptide pair. The performance of Xilmass was evaluated against the recently published Kojak and the popular pLink algorithms on a calmodulin-plectin complex data set, as well as three additional, published data sets. The results show that Xilmass typically had the highest number of identified distinct cross-linked sites and also the highest number of predicted cross-linked sites.

  18. Infrared microspectroscopic determination of collagen cross-links in articular cartilage

    NASA Astrophysics Data System (ADS)

    Rieppo, Lassi; Kokkonen, Harri T.; Kulmala, Katariina A. M.; Kovanen, Vuokko; Lammi, Mikko J.; Töyräs, Juha; Saarakkala, Simo

    2017-03-01

    Collagen forms an organized network in articular cartilage to give tensile stiffness to the tissue. Due to its long half-life, collagen is susceptible to cross-links caused by advanced glycation end-products. The current standard method for determination of cross-link concentrations in tissues is the destructive high-performance liquid chromatography (HPLC). The aim of this study was to analyze the cross-link concentrations nondestructively from standard unstained histological articular cartilage sections by using Fourier transform infrared (FTIR) microspectroscopy. Half of the bovine articular cartilage samples (n=27) were treated with threose to increase the collagen cross-linking while the other half (n=27) served as a control group. Partial least squares (PLS) regression with variable selection algorithms was used to predict the cross-link concentrations from the measured average FTIR spectra of the samples, and HPLC was used as the reference method for cross-link concentrations. The correlation coefficients between the PLS regression models and the biochemical reference values were r=0.84 (p<0.001), r=0.87 (p<0.001) and r=0.92 (p<0.001) for hydroxylysyl pyridinoline (HP), lysyl pyridinoline (LP), and pentosidine (Pent) cross-links, respectively. The study demonstrated that FTIR microspectroscopy is a feasible method for investigating cross-link concentrations in articular cartilage.

  19. Apparatus and method for the production of gel beads containing a biocatalyst

    DOEpatents

    Scott, Charles D.; Scott, Timothy C.; Davison, Brian H.

    1998-01-01

    An apparatus and method for the large-scale and continuous production of gel beads containing a biocatalyst. The apparatus is a columnar system based on the chemical cross-linking of hydrocolloidal gels that contain and immobilize a biocatalyst, the biocatalyst being a microorganism or an enzyme. Hydrocolloidal gels, such as alginate, carrageenan, and a mixture of bone gelatin and modified alginate, provide immobilization matrices that can be used to entrap and retain the biocatalyst while allowing effective contact with substrates and release of products. Such immobilized biocatalysts are generally formulated into small spheres or beads that have high concentrations of the biocatalyst within the gel matrix. The columnar system includes a gel dispersion nozzle submerged in a heated non-interacting liquid, typically an organic liquid, that is immiscible with water to allow efficient formation of spherical gel droplets, the non-interacting liquid having a specific gravity that is less than water so that the gel droplets will fall through the liquid by the force of gravity. The heated non-interacting liquid is in direct contact with a chilled upflowing non-interacting liquid that will provide sufficient residence time for the gel droplets as they fall through the liquid so that they will be cooled below the gelling temperature and form solid spheres. The upflowing non-interacting liquid is in direct contact with an upflowing temperature-controlled aqueous solution containing the necessary chemicals for cross-linking or fixing of the gel beads to add the necessary stability. The flow rates of the two liquid streams can be varied to control the proper residence time in each liquid section to accommodate the production of gel beads of differing settling velocities. A valve is provided for continuous removal of the stabilized gel beads from the bottom of the column.

  20. Apparatus and method for the production of gel beads containing a biocatalyst

    DOEpatents

    Scott, C.D.; Scott, T.C.; Davison, B.H.

    1998-01-27

    An apparatus and method are disclosed for the large-scale and continuous production of gel beads containing a biocatalyst. The apparatus is a columnar system based on the chemical cross-linking of hydrocolloidal gels that contain and immobilize a biocatalyst, the biocatalyst being a microorganism or an enzyme. Hydrocolloidal gels, such as alginate, carrageenan, and a mixture of bone gelatin and modified alginate, provide immobilization matrices that can be used to entrap and retain the biocatalyst while allowing effective contact with substrates and release of products. Such immobilized biocatalysts are generally formulated into small spheres or beads that have high concentrations of the biocatalyst within the gel matrix. The columnar system includes a gel dispersion nozzle submerged in a heated non-interacting liquid, typically an organic liquid, that is immiscible with water to allow efficient formation of spherical gel droplets, the non-interacting liquid having a specific gravity that is less than water so that the gel droplets will fall through the liquid by the force of gravity. The heated non-interacting liquid is in direct contact with a chilled upflowing non-interacting liquid that will provide sufficient residence time for the gel droplets as they fall through the liquid so that they will be cooled below the gelling temperature and form solid spheres. The upflowing non-interacting liquid is in direct contact with an upflowing temperature-controlled aqueous solution containing the necessary chemicals for cross-linking or fixing of the gel beads to add the necessary stability. The flow rates of the two liquid streams can be varied to control the proper residence time in each liquid section to accommodate the production of gel beads of differing settling velocities. A valve is provided for continuous removal of the stabilized gel beads from the bottom of the column. 1 fig.

  1. Cross-linked polyvinyl alcohol films as alkaline battery separators

    NASA Technical Reports Server (NTRS)

    Sheibley, D. W.; Manzo, M. A.; Gonzalez-Sanabria, O. D.

    1983-01-01

    Cross-linking methods have been investigated to determine their effect on the performance of polyvinyl alcohol (PVA) films as alkaline battery separators. The following types of cross-linked PVA films are discussed: (1) PVA-dialdehyde blends post-treated with an acid or acid periodate solution (two-step method) and (2) PVA-dialdehyde blends cross-linked during film formation (drying) by using a reagent with both aldehyde and acid functionality (one-step method). Laboratory samples of each cross-linked type of film were prepared and evaluated in standard separator screening tests. Then pilot-plant batches of films were prepared and compared to measure differences due to the cross-linking method. The pilot-plant materials were then tested in nickel oxide-zinc cells to compare the two methods with respect to performance characteristics and cycle life. Cell test results are compared with those from tests with Celgard.

  2. Cross-linked polyvinyl alcohol films as alkaline battery separators

    NASA Technical Reports Server (NTRS)

    Sheibley, D. W.; Manzo, M. A.; Gonzalez-Sanabria, O. D.

    1982-01-01

    Cross-linking methods were investigated to determine their effect on the performance of polyvinyl alcohol (PVA) films as alkaline battery separators. The following types of cross-linked PVA films are discussed: (1) PVA-dialdehyde blends post-treated with an acid or acid periodate solution (two-step method) and (2) PVA-dialdehyde blends cross-linked during film formation (drying) by using a reagent with both aldehyde and acid functionality (one-step method). Laboratory samples of each cross-linked type of film were prepared and evaluated in standard separator screening tests. The pilot-plant batches of films were prepared and compared to measure differences due to the cross-linking method. The pilot-plant materials were then tested in nickel oxide - zinc cells to compare the two methods with respect to performance characteristics and cycle life. Cell test results are compared with those from tests with Celgard.

  3. Hyaluronan synthesis in cultured tobacco cells (BY-2) expressing a chlorovirus enzyme: cytological studies.

    PubMed

    Rakkhumkaew, Numfon; Shibatani, Shigeo; Kawasaki, Takeru; Fujie, Makoto; Yamada, Takashi

    2013-04-01

    Extraction of hyaluronan from animals or microbial fermentation has risks including contamination with pathogens and microbial toxins. In this work, tobacco cultured-cells (BY-2) were successfully transformed with a chloroviral hyaluronan synthase (cvHAS) gene to produce hyaluronan. Cytological studies revealed accumulation of HA on the cells, and also in subcellular fractions (protoplasts, miniplasts, vacuoplasts, and vacuoles). Transgenic BY-2 cells harboring a vSPO-cvHAS construct containing the vacuolar targeting signal of sporamin connected to the N-terminus of cvHAS accumulated significant amounts of HA in vacuoles. These results suggested that cvHAS successfully functions on the vacuolar membrane and synthesizes/transports HA into vacuoles. Efficient synthesis of HA using this system provides a new method for practical production of HA. Copyright © 2012 Wiley Periodicals, Inc.

  4. Hyaluronan, CD44, and Emmprin Regulate Lactate Efflux and Membrane Localization of Monocarboxylate Transporters in Human Breast Carcinoma Cells

    PubMed Central

    Slomiany, Mark G.; Grass, G. Daniel; Robertson, Angela D.; Yang, Xiao Y.; Maria, Bernard L.; Beeson, Craig; Toole, Bryan P.

    2013-01-01

    Interactions of hyaluronan with CD44 in tumor cells play important cooperative roles in various aspects of malignancy and drug resistance. Emmprin (CD147; basigin)is a cell surface glycoprotein of the immunoglobulin superfamily that is highly up-regulated in malignant cancer cells and stimulates hyaluronan production, as well as several downstream signaling pathways. Emmprin also interacts with various monocarboxylate transporters (MCT). Malignant cancer cells use the glycolytic pathway and require MCTs to efflux lactate that results from glycolysis. Glycolysis and lactate secretion contribute to malignant cell behaviors and drug resistance in tumor cells. In the present study, we find that perturbation of endogenous hyaluronan, using small hyaluronan oligosaccharides, rapidly inhibits lactate efflux from breast carcinoma cells; down-regulation of emmprin, using emmprin small interfering RNA, also results in decreased efflux. In addition, we find that CD44 coimmunoprecipitates with MCT1, MCT4, and emmprin and colocalizes with these proteins at the plasma membrane. Moreover, after treatment of the cells with hyaluronan oligosaccharides, CD44, MCT1, and MCT4 become localized intracellularly whereas emmprin remains at the cell membrane. Together, these data indicate that constitutive interactions among hyaluronan, CD44, and emmprin contribute to regulation of MCT localization and function in the plasma membrane of breast carcinoma cells. PMID:19176383

  5. Ionic liquid compatibility in polyethylene oxide/siloxane ion gel membranes

    DOE PAGES

    Kusuma, Victor A.; Macala, Megan K.; Liu, Jian; ...

    2018-10-02

    Ion gel films were prepared by incorporating eight commercially available ionic liquids in two different cross-linked polymer matrices to evaluate their phase miscibility, gas permeability and ionic conductivity for potential applications as gas separation membranes and solid electrolyte materials. The ionic liquids cations were 1-ethyl-3-methylimidazolium, 1-ethyl-3-methylpyridinium, 1-butyl-1-methylpyrrolidinium, tributylmethylphosphonium, and butyltrimethylammonium with a common anion (bis(trifluoromethylsulfonyl)imide). In addition, ionic liquids with 1-ethyl-3-methylimidazolium cation with acetate, dicyanamide and tetrafluoroborate counterions were evaluated. The two polymers were cross-linked poly(ethylene oxide) and cross-linked poly(ethylene oxide)/siloxane copolymer. Differential scanning calorimetry, X-ray diffractometry and visual observations were performed to evaluate the ion gels’ miscibility, thermal stabilitymore » and homogeneity. Ionic liquids with the least basic anion (bis(trifluoromethylsulfonyl)imide) and aromatic cations containing acidic proton (e.g. imidazolium and pyridinium) gave the most stable and miscible ion gels. Phase stability was shown to be a function of both ionic liquid content and temperature, with phase separation observed at elevated temperatures. In conclusion, gas permeability testing with carbon dioxide and nitrogen and ionic conductivity measurements confirmed that these ionic liquids increased the gas permeability and ionic conductivity of the polymers.« less

  6. Ionic liquid compatibility in polyethylene oxide/siloxane ion gel membranes

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Kusuma, Victor A.; Macala, Megan K.; Liu, Jian

    Ion gel films were prepared by incorporating eight commercially available ionic liquids in two different cross-linked polymer matrices to evaluate their phase miscibility, gas permeability and ionic conductivity for potential applications as gas separation membranes and solid electrolyte materials. The ionic liquids cations were 1-ethyl-3-methylimidazolium, 1-ethyl-3-methylpyridinium, 1-butyl-1-methylpyrrolidinium, tributylmethylphosphonium, and butyltrimethylammonium with a common anion (bis(trifluoromethylsulfonyl)imide). In addition, ionic liquids with 1-ethyl-3-methylimidazolium cation with acetate, dicyanamide and tetrafluoroborate counterions were evaluated. The two polymers were cross-linked poly(ethylene oxide) and cross-linked poly(ethylene oxide)/siloxane copolymer. Differential scanning calorimetry, X-ray diffractometry and visual observations were performed to evaluate the ion gels’ miscibility, thermal stabilitymore » and homogeneity. Ionic liquids with the least basic anion (bis(trifluoromethylsulfonyl)imide) and aromatic cations containing acidic proton (e.g. imidazolium and pyridinium) gave the most stable and miscible ion gels. Phase stability was shown to be a function of both ionic liquid content and temperature, with phase separation observed at elevated temperatures. In conclusion, gas permeability testing with carbon dioxide and nitrogen and ionic conductivity measurements confirmed that these ionic liquids increased the gas permeability and ionic conductivity of the polymers.« less

  7. Diffusion and convection in collagen gels: implications for transport in the tumor interstitium.

    PubMed Central

    Ramanujan, Saroja; Pluen, Alain; McKee, Trevor D; Brown, Edward B; Boucher, Yves; Jain, Rakesh K

    2002-01-01

    Diffusion coefficients of tracer molecules in collagen type I gels prepared from 0-4.5% w/v solutions were measured by fluorescence recovery after photobleaching. When adjusted to account for in vivo tortuosity, diffusion coefficients in gels matched previous measurements in four human tumor xenografts with equivalent collagen concentrations. In contrast, hyaluronan solutions hindered diffusion to a lesser extent when prepared at concentrations equivalent to those reported in these tumors. Collagen permeability, determined from flow through gels under hydrostatic pressure, was compared with predictions obtained from application of the Brinkman effective medium model to diffusion data. Permeability predictions matched experimental results at low concentrations, but underestimated measured values at high concentrations. Permeability measurements in gels did not match previous measurements in tumors. Visualization of gels by transmission electron microscopy and light microscopy revealed networks of long collagen fibers at lower concentrations along with shorter fibers at high concentrations. Negligible assembly was detected in collagen solutions pregelation. However, diffusion was similarly hindered in pre and postgelation samples. Comparison of diffusion and convection data in these gels and tumors suggests that collagen may obstruct diffusion more than convection in tumors. These findings have significant implications for drug delivery in tumors and for tissue engineering applications. PMID:12202388

  8. The molecular basis of the solution properties of hyaluronan investigated by confocal fluorescence recovery after photobleaching.

    PubMed Central

    Gribbon, P; Heng, B C; Hardingham, T E

    1999-01-01

    Hyaluronan (HA) is a highly hydrated polyanion, which is a network-forming and space-filling component in the extracellular matrix of animal tissues. Confocal fluorescence recovery after photobleaching (confocal-FRAP) was used to investigate intramolecular hydrogen bonding and electrostatic interactions in hyaluronan solutions. Self and tracer lateral diffusion coefficients within hyaluronan solutions were measured over a wide range of concentrations (c), with varying electrolyte and at neutral and alkaline pH. The free diffusion coefficient of fluoresceinamine-labeled HA of 500 kDa in PBS was 7.9 x 10(-8) cm(2) s(-1) and of 830 kDa HA was 5.6 x 10(-8) cm(2) s(-1). Reductions in self- and tracer-diffusion with c followed a stretched exponential model. Electrolyte-induced polyanion coil contraction and destiffening resulted in a 2.8-fold increase in self-diffusion between 0 and 100 mM NaCl. Disruption of hydrogen bonds by strong alkali (0.5 M NaOH) resulted in further larger increases in self- and tracer-diffusion coefficients, consistent with a more dynamic and permeable network. Concentrated hyaluronan solution properties were attributed to hydrodynamic and entanglement interactions between domains. There was no evidence of chain-chain associations. At physiological electrolyte concentration and pH, the greatest contribution to the intrinsic stiffness of hyaluronan appeared to be due to hydrogen bonds between adjacent saccharides. PMID:10512840

  9. A monolithic column based on covalent cross-linked polymer gels for online extraction and analysis of trace aflatoxins in food sample.

    PubMed

    Wei, Tianfu; Chen, Zhengyi; Li, Gongke; Zhang, Zhuomin

    2018-05-04

    Aflatoxins are highly toxic mycotoxin contamination, which pose serious food safety incidents. It is very important to precisely and rapidly determine trace aflatoxins in food. In this study, we designed porous monolithic column based on covalent cross-linked polymer gels for online extraction and analysis of trace aflatoxins in food samples with complicated matrices coupled with high-performance liquid chromatography-ultraviolet detector (HPLC-UV). The prepared monolithic column showed excellent enrichment performance due to its good permeability, good reproducibility and long life span. The study of adsorption mechanism suggested that the excellent enrichment performance of this monolithic column was attributed to the multiple effect of π-π stacking interaction, hydrophobic effect and steric effect. When the online analytical method was applied for the determine of trace aflatoxins in real food samples, aflatoxins G 1 and aflatoxins B 1 could be actually found in one positive bean sauce sample and quantified to be 32.8 and 26.4 μg/kg, respectively. Aflatoxins G 1 in one bean sample could be also found and quantified to be 25.9 μg/kg. The low detection limits of the developed method were achieved in range of 0.08-0.2 μg/kg. And the recoveries for spiked samples were in range from 76.1 to 113% with RSDs of 1.1-9.6%. The developed method was proved to be a promising method for online enrichment and analysis of trace aflatoxins in complicated food samples. Copyright © 2018 Elsevier B.V. All rights reserved.

  10. Structurally stable gel bead containing entrapped enzyme and method for manufacture thereof

    DOEpatents

    Woodward, Jonathan

    1998-01-01

    A structurally stable gel bead containing an entrapped enzyme and a method for its manufacture. The enzyme is covalently cross-linked to gelatin in the presence of glutaraldehyde prior to the formation of the gel bead, to prevent leakage of the enzyme. Propylene glycol alginate is then added to the mixture. Once the gel beads are formed, they are then soaked in glutaraldehyde, which imparts structural stability to the gel beads. This method can be used with many types of enzymes, such as proteases, carbohydrases, proteases, ligases, isomerases, oxidoreductases, and specialty enzymes. These and other enzymes can be immobilized in the gel beads and utilized in a number of enzymatic processes. Exogenously added ions are not required to maintain the structural stability of these gel beads.

  11. Ionically cross-linked poly(allylamine) as a stimulus-responsive underwater adhesive: ionic strength and pH effects.

    PubMed

    Lawrence, Patrick G; Lapitsky, Yakov

    2015-02-03

    Gel-like coacervates that adhere to both hydrophilic and hydrophobic substrates under water have recently been prepared by ionically cross-linking poly(allylamine) (PAH) with pyrophosphate (PPi) and tripolyphosphate (TPP). Among the many advantages of these underwater adhesives (which include their simple preparation and low cost) is their ability to dissolve on demand when exposed to high or low pH. To further analyze their stimulus-responsive properties, we have investigated the pH and ionic strength effects on the formation, rheology and adhesion of PAH/PPi and PAH/TPP complexes. The ionic cross-linker concentrations needed to form these adhesives decreased with increasing pH and ionic strength (although the complexes ceased to form when the parent solution pH exceeded ca. 8.5; i.e., the effective pKa of PAH). Once formed, their ionic cross-links were most stable (as inferred from their relaxation times) at near-neutral or slightly alkaline pH values (of roughly 6.5-9) and at low ionic strengths. The decrease in ionic cross-link stability within complexes prepared at other pH values and at elevated (150-300 mM) NaCl concentrations diminished both the strength and longevity of adhesion (although, under most conditions tested, the short-term tensile adhesion strengths remained above 10(5) Pa). Additionally, the sensitivity of PAH/PPi and PAH/TPP complexes to ionic strength was demonstrated as a potential route to injectable adhesive design (where spontaneous adhesive formation was triggered via injection of low-viscosity, colloidal PAH/TPP dispersions into phosphate buffered saline). Thus, while the sensitivity of ionically cross-linked PAH networks to pH and ionic strength can weaken their adhesion, it can also impart them with additional functionality, such as minimally invasive, injectable delivery, and ability to form and dissolve their bonds on demand.

  12. Preparation and characterization of a magneto-polymeric nanocomposite: Fe 3O 4 nanoparticles in a grafted, cross-linked and plasticized poly(vinyl chloride) matrix

    NASA Astrophysics Data System (ADS)

    Rodríguez-Fernández, Oliverio S.; Rodríguez-Calzadíaz, C. A.; Yáñez-Flores, Isaura G.; Montemayor, Sagrario M.

    In this work two kind of materials: (1) grafted, cross-linked and plasticized poly(vinyl chloride) (PVC) "plastic films" and (2) magnetic plastic films "magneto-polymeric nanocomposites" were prepared. Precursor solutions or "plastisols" used to obtain the plastic films were obtained by mixing PVC (emulsion grade) as polymeric matrix, di(2-ethylhexyl)phthalate (DOP) as plasticizer, a thermal stabilizer based in Ca/Zn salts, and a cross-linking agent, 3-mercaptopropyltrimethoxysilane (MTMS) or 3-aminopropyltriethoxysilane (ATES), at several concentrations. Flexible films were obtained from the plastisols using static casting. The stress-strain behavior and the gel content (determined by Soxhlet extraction with boiling THF) of the flexible films were measured in order to evaluate the effect of the cross-linking agent and their content on the degree of cross-linking. The magneto-polymeric nanocomposites were obtained by mixing the optimum composition of the plastisols (analyzed previously) with magnetite (Fe 3O 4)-based ferrofluid and DOP. Later, flexible films were obtained by static casting of the plastisol/ferrofluid systems. The magnetic films were characterized by the above-mentioned techniques and X-ray diffraction, vibrating sample magnetometry and thermogravimetrical analysis.

  13. Investigation into response characteristics of the chitosan gel artificial muscle

    NASA Astrophysics Data System (ADS)

    Zhao, Gang; Yang, Junjie; Wang, Yujian; Zhao, Honghao; Fu, Yu; Zhang, Guangli; Yu, Shuqin; Wu, Yuda; Wei, Chengye; Liu, Xuxiong; Wang, Zhijie

    2018-01-01

    Bionic artificial muscle made from chitosan gel is an emerging type of the ionic electro active polymer with advantages of large deformation, low cost and environmental protection etc, which leads to a research focus and wide application in the fields of bionic engineering and intelligence material recently. In this paper, effects and improvement mechanisms of the direct casting and genipin cross-linking processes on response speed properties of the chitosan gel artificial muscle (CGAM) were mainly studied. Based on in-depth analysis of the CGAM response mechanism, a platform was built for testing the response performance of the CGAM, then its equivalent circuit and mathematical models were also established. Furthermore, control experiments were carried out to test and analyze several performances of the CGAM on response speed, electrical conductivity, mechanical properties and microstructure with different control variables. The experimental results illustrated that the CGAM assembled by direct casting enabled its electric actuating membrane and non-metallic electrode membrane tightly attached together with low contact resistance, which dramatically promoted the electrical conductivity of the CGAM resulting in nearly doubled response speed. Besides, different concentrations of genipin were adopted to cross-link the CGAM actuating membranes, and then it was found that the response speed of the uncross-linked CGAM was fast in the initial stage, but as time increased, it declined rapidly with poor steadiness. While there was no obvious decrease over time on the response speed of the CGAM cross-linked with low genipin concentration. Namely, its stability was getting better and better. In addition, the response speed of the CGAM cross-linked with low concentration of genipin was roughly the same as uncross-linked CGAM, which was quicker than that of high concentration. In this work, its internal mechanisms, feasible assembly technique and green modification method were

  14. In vivo oxidation in remelted highly cross-linked retrievals.

    PubMed

    Currier, B H; Van Citters, D W; Currier, J H; Collier, J P

    2010-10-20

    Elimination of free radicals to prevent oxidation has played a major role in the development and product differentiation of the latest generation of highly cross-linked ultra-high molecular weight polyethylene bearing materials. In the current study, we (1) examined oxidation in a series of retrieved remelted highly cross-linked ultra-high molecular weight polyethylene bearings from a number of device manufacturers and (2) compared the retrieval results with findings for shelf-stored control specimens. The hypothesis was that radiation-cross-linked remelted ultra-high molecular weight polyethylene would maintain oxidative stability in vivo comparable with the stability during shelf storage and in published laboratory aging tests. Fifty remelted highly cross-linked ultra-high molecular weight polyethylene acetabular liners and nineteen remelted highly cross-linked ultra-high molecular weight polyethylene tibial inserts were received after retrieval from twenty-one surgeons from across the U.S. Thirty-two of the retrievals had been in vivo for two years or more. Each was measured for oxidation with use of Fourier transform infrared spectroscopy. A control series of remelted highly cross-linked ultra-high molecular weight polyethylene acetabular liners from three manufacturers was analyzed with electron paramagnetic resonance spectroscopy to measure free radical content and with Fourier transform infrared spectroscopy to measure oxidation initially and after eight to nine years of shelf storage in air. The never-implanted, shelf-aged controls had no measurable free-radical content initially or after eight to nine years of shelf storage. The never-implanted controls showed no increase in oxidation during shelf storage. Oxidation measurements showed measurable oxidation in 22% of the retrieved remelted highly cross-linked liners and inserts after an average of two years in vivo. Because never-implanted remelted highly cross-linked ultra-high molecular weight

  15. Shear Stress Regulates Adhesion and Rolling of CD44+ Leukemic and Hematopoietic Progenitor Cells on Hyaluronan

    PubMed Central

    Christophis, Christof; Taubert, Isabel; Meseck, Georg R.; Schubert, Mario; Grunze, Michael; Ho, Anthony D.; Rosenhahn, Axel

    2011-01-01

    Leukemic cells and human hematopoietic progenitor cells expressing CD44 receptors have the ability to attach and roll on hyaluronan. We investigated quantitatively the adhesion behavior of leukemic cell lines and hematopoietic progenitor cells on thin films of the polysaccharides hyaluronan and alginate in a microfluidic system. An applied flow enhances the interaction between CD44-positive cells and hyaluronan if a threshold shear stress of 0.2 dyn/cm2 is exceeded. At shear stress ∼1 dyn/cm2, the cell rolling speed reaches a maximum of 15 μm/s. Leukemic Jurkat and Kasumi-1 cells lacking CD44-expression showed no adhesion or rolling on the polysaccharides whereas the CD44-expressing leukemic cells KG-1a, HL-60, K-562, and hematopoietic progenitor cells attached and rolled on hyaluronan. Interestingly, the observations of flow-induced cell rolling are related to those found in the recruitment of leukocytes to inflammatory sites and the mechanisms of stem-cell homing into the bone marrow. PMID:21806926

  16. Hyaluronan suppresses lidocaine-induced apoptosis of human chondrocytes in vitro by inhibiting the p53-dependent mitochondrial apoptotic pathway.

    PubMed

    Lee, Yoon-Jin; Kim, Soo A; Lee, Sang-Han

    2016-05-01

    Intra-articular injection of local anesthetics (LAs) is a common procedure for therapeutic purposes. However, LAs have been found toxic to articular cartilage, and hyaluronan may attenuate this toxicity. In this study we investigated whether hyaluronan attenuated lidocaine-induced chondrotoxicity, and if so, to elucidate the underlying mechanisms. Human chondrocyte cell line SW1353 and newly isolated murine chondrocytes were incubated in culture medium containing hyaluronan and/or lidocaine for 72 h. Cell viability was evaluated using MTT assay. Cell apoptosis was detected with DAPI staining, caspase 3/7 activity assay and flow cytometry. Cell cycle distributions, ROS levels and mitochondrial membrane potential (ΔΨm) were determined using flow cytometry. The expression of p53 and p53-regulated gene products was measured with Western blotting. Lidocaine (0.005%-0.03%) dose-dependently decreased the viability of SW1353 cells. This local anesthetic (0.015%, 0.025%) induced apoptosis, G2/M phase arrest and loss of ΔΨm, and markedly increased ROS production in SW1353 cells. Hyaluronan (50-800 μg/mL) alone did not affect the cell viability, but co-treatment with hyaluronan (200 μg/mL) significantly attenuated lidocaine-induced apoptosis and other abnormalities in SW1353 cells. Furthermore, co-treatment with lidocaine and hyaluronan significantly decreased the levels of p53 and its transcription targets Bax and p21 in SW1353 cells, although treatment with lidocaine alone did not significantly change these proteins. Similar results were obtained in ex vivo cultured murine chondrocytes. Hyaluronan suppresses lidocaine-induced apoptosis of human chondrocytes in vitro through inhibiting the p53-dependent mitochondrial apoptotic pathway.

  17. The effect of mixed dopants on the stability of Fricke gel dosimeters

    NASA Astrophysics Data System (ADS)

    Penev, K.; Mequanint, K.

    2013-06-01

    Auto-oxidation and fast diffusion in Fricke gels are major drawbacks to wide-spread application of these gels in 3D dosimetry. Aiming to limit both processes, we used mixed dopants: the ferric-specific ligand xylenol orange with a ferrous-specific ligand (1,10-phenanthroline) and/or a bi-functional cross-linking agent (glyoxal). Markedly improved auto-oxidation stability was observed in the xylenol orange and phenanthroline doped gel at the expense of increased background absorbance and faster diffusion. Addition of glyoxal limited the diffusion rate and led to a partial bleaching of the gel. It is conceivable that these two new compositions may find useful practical application.

  18. Cross-Linking Studies of Lysozyme Nucleation

    NASA Technical Reports Server (NTRS)

    Forsythe, Elizabeth; Pusey, Marc

    2000-01-01

    Tetragonal chicken egg white crystals consist of 4(sub 3) helices running in alternating directions, the helix rows having a two fold symmetry with each other. The unit cell consists of one complete tetrameric turn from each of two adjacent helices (an octamer). PBC analysis indicates that the helix intermolecular bonds are the strongest in the crystal, therefore likely formed first. AFM analysis of the (110) surface shows only complete helices, no half steps or bisected helices being found, while AFM line scans to measure the growth step increments show that they are multiples of the 4(sub 3) helix tetramer dimensions. This supports our thesis that the growth units are in fact multiples of the four molecule 4(sub 3) helix unit, the "average" growth unit size for the (110) face being an octamer (two turns about the helix) and the (101) growth unit averaging about the size of a hexamer. In an effort to better understand the species involved in the crystal nucleation and growth process, we have initiated an experimental program to study the species formed in solution compared to what is found in the crystal through covalent cross-linking studies. These experiments use the heterobifunctional cross-linking agent aminoethyl-4-azidonitroanaline (AEANA). An aliphatic amine at one end is covalently attached to the protein by a carbodiimide-mediated reaction, and a photo reactive group at the other can be used to initiate crosslinking. Modifications to the parent structure can be used to alter the distance between the two reactive groups and thus the cross-linking agents "reach". In practice, the cross-linking agent is first coupled to the asp101 side chain through the amine group. Asp101 lies within the active site cleft, and previous work with fluorescent probes had shown that derivatives at this site still crystallize in the tetragonal space group. This was also found to be the case with the AEANA derivative, which gave red tetragonal crystals. The protein now has a

  19. Spectroscopic characterization of collagen cross-links in bone

    NASA Technical Reports Server (NTRS)

    Paschalis, E. P.; Verdelis, K.; Doty, S. B.; Boskey, A. L.; Mendelsohn, R.; Yamauchi, M.

    2001-01-01

    Collagen is the most abundant protein of the organic matrix in mineralizing tissues. One of its most critical properties is its cross-linking pattern. The intermolecular cross-linking provides the fibrillar matrices with mechanical properties such as tensile strength and viscoelasticity. In this study, Fourier transform infrared (FTIR) spectroscopy and FTIR imaging (FTIRI) analyses were performed in a series of biochemically characterized samples including purified collagen cross-linked peptides, demineralized bovine bone collagen from animals of different ages, collagen from vitamin B6-deficient chick homogenized bone and their age- and sex-matched controls, and histologically stained thin sections from normal human iliac crest biopsy specimens. One region of the FTIR spectrum of particular interest (the amide I spectral region) was resolved into its underlying components. Of these components, the relative percent area ratio of two subbands at approximately 1660 cm(-1) and approximately 1690 cm(-1) was related to collagen cross-links that are abundant in mineralized tissues (i.e., pyridinoline [Pyr] and dehydrodihydroxylysinonorleucine [deH-DHLNL]). This study shows that it is feasible to monitor Pyr and DHLNL collagen cross-links spatial distribution in mineralized tissues. The spectroscopic parameter established in this study may be used in FTIRI analyses, thus enabling the calculation of relative Pyr/DHLNL amounts in thin (approximately 5 microm) calcified tissue sections with a spatial resolution of approximately 7 microm.

  20. Intra-molecular cross-linking of acidic residues for protein structure studies.

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Kruppa, Gary Hermann; Young, Malin M.; Novak, Petr

    2005-03-01

    Intra-molecular cross-linking has been suggested as a method of obtaining distance constraints that would be useful in developing structural models of proteins. Recent work published on intra-molecular cross-linking for protein structural studies has employed commercially available primary amine selective reagents that can cross-link lysine residues to other lysine residues or the amino terminus. Previous work using these cross-linkers has shown that for several proteins of known structure, the number of cross-links that can be obtained experimentally may be small compared to what would be expected from the known structure, due to the relative reactivity, distribution, and solvent accessibility of themore » lysines in the protein sequence. To overcome these limitations we have investigated the use of cross-linking reagents that can react with other reactive sidechains in proteins. We used 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) to activate the carboxylic acid containing residues, aspartic acid (D), glutamic acid (E), and the carboxy terminus (O), for cross-linking reactions. Once activated, the DEO sidechains can react to form 'zero-length' cross-links with nearby primary amine containing resides, lysines (K) and the amino terminus (X), via the formation of a new amide bond. We also show that the EDC-activated DEO sidechains can be cross-linked to each other using dihydrazides, two hydrazide moieties connected by an alkyl cross-linker ann of variable length. Using these reagents, we have found three new 'zero-length' cross-links in ubiquitin consistent with its known structure (M1-E16, M1-E18, and K63-E64). Using the dihydrazide cross-linkers, we have identified 2 new cross-links (D21-D32 and E24-D32) unambiguously. Using a library of dihydrazide cross-linkers with varying arm length, we have shown that there is a minimum arm length required for the DEO-DEO cross-links of 5.8 angstroms. These results show that additional structural

  1. Diepoxybutane Interstrand Cross-Links Induce DNA Bending

    PubMed Central

    Millard, Julie T.; McGowan, Erin E.; Bradley, Sharonda Q.

    2011-01-01

    The bifunctional alkylating agent 1,2,3,4-diepoxybutane (DEB) is thought to be a major contributor to the carcinogenicity of 1,3-butadiene, from which it is derived in vivo. DEB forms DNA interstrand cross-links primarily between distal deoxyguanosine residues at the duplex sequence 5’-GNC. In order for the short butanediol tether to span this distance, distortion of the DNA target has been postulated. We determined that the electrophoretic mobility of ligated DNA oligomers containing DEB cross-links was retarded in comparison with control, uncross-linked DNA. Our data are consistent with DNA bending of ~34° per lesion towards the major groove. PMID:21839139

  2. Fabrication of chemically cross-linked porous gelatin matrices.

    PubMed

    Bozzini, Sabrina; Petrini, Paola; Altomare, Lina; Tanzi, Maria Cristina

    2009-01-01

    The aim of this study was to chemically cross-link gelatin, by reacting its free amino groups with an aliphatic diisocyanate. To produce hydrogels with controllable properties, the number of reacting amino groups was carefully determined. Porosity was introduced into the gelatin-based hydrogels through the lyophilization process. Porous and non-porous matrices were characterized with respect to their chemical structure, morphology, water uptake and mechanical properties. The physical, chemical and mechanical properties of the porous matrices are related to the extent of their cross-linking, showing that they can be controlled by varying the reaction parameters. Water uptake values (24 hours) vary between 160% and 200% as the degree of cross-linking increases. The flexibility of the samples also decreases by changing the extent of cross-linking. Young's modulus shows values between 0.188 KPa, for the highest degree, and 0.142 KPa for the lowest degree. The matrices are potential candidates for use as tissue-engineering scaffolds by modulating their physical chemical properties according to the specific application.

  3. Practical application of thermoreversibly Cross-linked rubber products

    NASA Astrophysics Data System (ADS)

    Polgar, L. M.; Picchioni, F.; de Ruiter, E.; van Duin, M.

    2017-07-01

    Currently, rubber products cannot simply be reprocessed after their product life, due to the irreversible cross-linking methods traditionally applied. The purpose of this work is to investigate how thermoreversible cross-linking of rubbers via Diels Alder chemistry can be used for the development of recyclable rubber products. Unfortunately, the applicability of the thermoreversible EPM-g-furan/BM system appears to be limited to room temperature applications, because of the rapid deterioration of the compression set at elevated temperatures compared to irreversibly cross-linked EPM. However, the use of EPM rubber modified with thiophene or cyclopentadiene moieties may extend the temperature application range and results in rubber products with acceptable properties. Finally, rubber products generally comprise fillers such as silica, carbon black or fibers. In this context, the reinforcing effect of short cut aramid fibers on the material properties of the newly developed thermoreversibly cross-linked EPM rubbers was also studied. The material properties of the resulting products were found to be comparable to those of a fiber reinforced, peroxide cured reference sample.

  4. HcRed, a Genetically Encoded Fluorescent Binary Cross-Linking Agent for Cross-Linking of Mitochondrial ATP Synthase in Saccharomyces cerevisiae

    PubMed Central

    Gong, Lan; Ramm, Georg; Devenish, Rodney J.; Prescott, Mark

    2012-01-01

    Genetically encoded fluorescent cross-linking agents represent powerful tools useful both for visualising and modulating protein interactions in living cells. The far-red fluorescent protein HcRed, which is fluorescent only in a dimer form, can be used to promote the homo-dimerisation of target proteins, and thereby yield useful information about biological processes. We have in yeast cells expressed HcRed fused to a subunit of mitochondrial ATP synthase (mtATPase). This resulted in cross-linking of the large multi-subunit mtATPase complex within the inner-membrane of the mitochondrion. Fluorescence microscopy revealed aberrant mitochondrial morphology, and mtATPase complexes isolated from mitochondria were recovered as fluorescent dimers under conditions where complexes from control mitochondria were recovered as monomers. When viewed by electron microscopy normal cristae were absent from mitochondria in cells in which mATPase complexes were cross-linked. mtATPase dimers are believed to be the building blocks that are assembled into supramolecular mtATPase ribbons that promote the formation of mitochondrial cristae. We propose that HcRed cross-links mATPase complexes in the mitochondrial membrane hindering the normal assembly/disassembly of the supramolecular forms of mtATPase. PMID:22496895

  5. On the Reproducibility of Label-Free Quantitative Cross-Linking/Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Müller, Fränze; Fischer, Lutz; Chen, Zhuo Angel; Auchynnikava, Tania; Rappsilber, Juri

    2018-02-01

    Quantitative cross-linking/mass spectrometry (QCLMS) is an emerging approach to study conformational changes of proteins and multi-subunit complexes. Distinguishing protein conformations requires reproducibly identifying and quantifying cross-linked peptides. Here we analyzed the variation between multiple cross-linking reactions using bis[sulfosuccinimidyl] suberate (BS3)-cross-linked human serum albumin (HSA) and evaluated how reproducible cross-linked peptides can be identified and quantified by LC-MS analysis. To make QCLMS accessible to a broader research community, we developed a workflow that integrates the established software tools MaxQuant for spectra preprocessing, Xi for cross-linked peptide identification, and finally Skyline for quantification (MS1 filtering). Out of the 221 unique residue pairs identified in our sample, 124 were subsequently quantified across 10 analyses with coefficient of variation (CV) values of 14% (injection replica) and 32% (reaction replica). Thus our results demonstrate that the reproducibility of QCLMS is in line with the reproducibility of general quantitative proteomics and we establish a robust workflow for MS1-based quantitation of cross-linked peptides.

  6. Structurally stable gel bead containing entrapped enzyme and method for manufacture thereof

    DOEpatents

    Woodward, J.

    1998-12-08

    This research provides a structurally stable gel bead containing an entrapped enzyme and a method for its manufacture. The enzyme is covalently cross-linked to gelatin in the presence of glutaraldehyde prior to the formation of the gel bead, to prevent leakage of the enzyme. Propylene glycol alginate is then added to the mixture. Once the gel beads are formed, they are then soaked in glutaraldehyde, which imparts structural stability to the gel beads. This method can be used with many types of enzymes, such as proteases, carbohydrases, proteases, ligases, isomerases, oxidoreductases, and specialty enzymes. These and other enzymes can be immobilized in the gel beads and utilized in a number of enzymatic processes. Exogenously added ions are not required to maintain the structural stability of these gel beads. 7 figs.

  7. Hyaluronan suppresses lidocaine-induced apoptosis of human chondrocytes in vitro by inhibiting the p53-dependent mitochondrial apoptotic pathway

    PubMed Central

    Lee, Yoon-Jin; Kim, Soo A; Lee, Sang-Han

    2016-01-01

    Aim: Intra-articular injection of local anesthetics (LAs) is a common procedure for therapeutic purposes. However, LAs have been found toxic to articular cartilage, and hyaluronan may attenuate this toxicity. In this study we investigated whether hyaluronan attenuated lidocaine-induced chondrotoxicity, and if so, to elucidate the underlying mechanisms. Methods: Human chondrocyte cell line SW1353 and newly isolated murine chondrocytes were incubated in culture medium containing hyaluronan and/or lidocaine for 72 h. Cell viability was evaluated using MTT assay. Cell apoptosis was detected with DAPI staining, caspase 3/7 activity assay and flow cytometry. Cell cycle distributions, ROS levels and mitochondrial membrane potential (ΔΨm) were determined using flow cytometry. The expression of p53 and p53-regulated gene products was measured with Western blotting. Results: Lidocaine (0.005%−0.03%) dose-dependently decreased the viability of SW1353 cells. This local anesthetic (0.015%, 0.025%) induced apoptosis, G2/M phase arrest and loss of ΔΨm, and markedly increased ROS production in SW1353 cells. Hyaluronan (50−800 μg/mL) alone did not affect the cell viability, but co-treatment with hyaluronan (200 μg/mL) significantly attenuated lidocaine-induced apoptosis and other abnormalities in SW1353 cells. Furthermore, co-treatment with lidocaine and hyaluronan significantly decreased the levels of p53 and its transcription targets Bax and p21 in SW1353 cells, although treatment with lidocaine alone did not significantly change these proteins. Similar results were obtained in ex vivo cultured murine chondrocytes. Conclusion: Hyaluronan suppresses lidocaine-induced apoptosis of human chondrocytes in vitro through inhibiting the p53-dependent mitochondrial apoptotic pathway. PMID:27041463

  8. Synthesis and enzymatic degradation of epichlorohydrin cross-linked pectins.

    PubMed

    Semdé, Rasmané; Moës, André J; Devleeschouwer, Michel J; Amighi, Karim

    2003-02-01

    The water solubility of pectin was successfully decreased by cross-linking with increasing amounts of epichlorohydrin in the reaction media. The initial molar ratios of epichlorohydrin/ galacturonic acid monomer in the reaction mixtures were 0, 0.37, 0.56, 0.74, 1.00, 1.47, and 2.44. The resulting epichlorohydrin cross-linked pectins were thus referred to as C-LP0, C-LP37, C-LP56, C-LP75, C-LP100, C-LP150, and C-LP250, respectively. Methoxylation degrees ranged from 60.5 +/- 0.9% to 68.0 +/- 0.6%, and the effective cross-linking degrees, determined by quantification of the hydroxyl anions consumed during the reaction, were 0, 17.8, 26.0, 38.3, 46.5, 53.5, and 58.7%. respectively. After incubating the different cross-linked pectins (0.5% w/v) in 25 mL of 0.05 M acetate-phosphate buffer (pH 4.5), containing 50 microL of Pectinex Ultra SP-L (pectinolytic enzymes), between 60 and 80% of the pectin osidic bounds were broken in less than 1 hr. Moreover, increasing the cross-linking degree only resulted in a weak slowing on the enzymatic degradation velocity.

  9. Cross-linking of surface Ig receptors on murine B lymphocytes stimulates the expression of nuclear tetradecanoyl phorbol acetate-response element-binding proteins

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Chiles, T.C.; Liu, J.L.; Rothstein, T.L.

    1991-03-15

    Cross-linking of sIg on primary B lymphocytes leads to increased nuclear DNA-binding activity specific for the tetradecanoyl phorbol acetate-response element (TRE), as judged by gel mobility shift assays. Stimulation of B cells to enter S phase of the cell cycle by treatment with the combination of phorbol ester plus calcium ionophore also stimulated nuclear TRE-binding activity within 2 h, with maximal expression at 4 h; however, phorbol ester and calcium ionophore were not as effective in stimulating binding activity when examined separately. Stimulated nuclear expression of TRE-binding activity appears to require protein synthesis. Fos- and Jun/AP-1-related proteins participate directly inmore » the identified nucleoprotein complex, as shown by the ability of c-fos- and c-jun-specific antisera to either alter or completely abolish electrophoretic migration of the complex in native gels. Further, UV photo-cross-linking studies identified two major TRE-binding protein species, whose sizes correspond to TRE-binding proteins derived from HeLa cell nuclear extracts. The results suggest that in primary B cells nuclear TRE-binding activity represents a downstream signaling event that occurs subsequent to changes in protein kinase C activity and intracellular Ca2+ but that can be triggered physiologically through sIg.« less

  10. Highly cross-linked nanoporous polymers

    DOEpatents

    Steckle, Jr., Warren P.; Apen, Paul G.; Mitchell, Michael A.

    1998-01-01

    Condensation polymerization followed by a supercritical extraction step can be used to obtain highly cross-linked nanoporous polymers with high surface area, controlled pore sizes and rigid structural integrity. The invention polymers are useful for applications requiring separation membranes.

  11. Highly cross-linked nanoporous polymers

    DOEpatents

    Steckle, Jr., Warren P.; Apen, Paul G.; Mitchell, Michael A.

    1997-01-01

    Condensation polymerization followed by a supercritical extraction step can be used to obtain highly cross-linked nanoporous polymers with high surface area, controlled pore sizes and rigid structural integrity. The invention polymers are useful for applications requiring separation membranes.

  12. Effect of gel formation on the dissolution behavior of clarithromycin tablets.

    PubMed

    Inukai, Koki; Takiyama, Kei; Noguchi, Shuji; Iwao, Yasunori; Itai, Shigeru

    2017-04-15

    Clarithromycin (CAM) is a macrolide antibiotic that is widely used at clinical sites. We found that release of CAM is suppressed when tablets of CAM were exposed to an external solvent containing carboxylate buffers such as citrate. The suppressed release of CAM can be attributed to the formation of gels on the tablet surfaces, which inhibits penetration of the solvent into the tablet and thus disintegration of the tablets. Delayed disintegration of the tablets was also observed for commercial tablets. This suggests that taking CAM and carboxylates at the same time might be avoided. The crystal structure of CAM citrate reveals that molecular chains of CAM are cross-linked by hydrogen bond between citrate groups in the crystal. The crystal structure indicates that cross-linked CAM chains of the three-dimensional mesh structure might also be formed in high concentration CAM solutions in the presence of carboxylates, resulting in gel formation. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Formaldehyde cross-linking and structural proteomics: Bridging the gap.

    PubMed

    Srinivasa, Savita; Ding, Xuan; Kast, Juergen

    2015-11-01

    Proteins are dynamic entities constantly moving and altering their structures based on their functions and interactions inside and outside the cell. Formaldehyde cross-linking combined with mass spectrometry can accurately capture interactions of these rapidly changing biomolecules while maintaining their physiological surroundings. Even with its numerous established uses in biology and compatibility with mass spectrometry, formaldehyde has not yet been applied in structural proteomics. However, formaldehyde cross-linking is moving toward analyzing tertiary structure, which conventional cross-linkers have already accomplished. The purpose of this review is to describe the potential of formaldehyde cross-linking in structural proteomics by highlighting its applications, characteristics and current status in the field. Copyright © 2015 Elsevier Inc. All rights reserved.

  14. Cross-Linked Nanotube Materials with Variable Stiffness Tethers

    NASA Technical Reports Server (NTRS)

    Frankland, Sarah-Jane V.; Odegard, Gregory M.; Herzog, Matthew N.; Gates, Thomas S.; Fay, Catherine C.

    2004-01-01

    The constitutive properties of a cross-linked single-walled carbon nanotube material are predicted with a multi-scale model. The material is modeled as a transversely isotropic solid using concepts from equivalent-continuum modeling. The elastic constants are determined using molecular dynamics simulation. Some parameters of the molecular force field are determined specifically for the cross-linker from ab initio calculations. A demonstration of how the cross-linked nanotubes may affect the properties of a nanotube/polyimide composite is included using a micromechanical analysis.

  15. 11β-Hydroxylation of Cortexolone (Reichstein Compound S) to Hydrocortisone by Curvularia lunata Entrapped in Photo-Cross-Linked Resin Gels

    PubMed Central

    Sonomoto, Kenji; Hoq, M. Mozammel; Tanaka, Atsuo; Fukui, Saburo

    1983-01-01

    Spores of Curvularia lunata were immobilized by entrapment with photo-cross-linkable resin prepolymers and incubated to form mycelium in potato dextrose broth containing cortexolone (Reichstein compound S) as an inducer of steroid 11β-hydroxylase. In a buffer system containing 2.5% dimethyl sulfoxide, this immobilized mycelium hydroxylated cortexolone to hydrocortisone. The activity of this mycelium was comparable to the activity of free mycelium. Dimethyl sulfoxide did not inhibit hydroxylase activity at the concentration used and was effective in dissolving the product. Of the various photo-cross-linkable resin prepolymers examined, use of ENT-4000, whose main chain was polyethylene glycol 4000 (chain length, approximately 40 nm), resulted in maximum hydroxylation activity of the entrapped mycelium. The chain length of prepolymers affected markedly mycelial growth in the gels and, subsequently, the activity of the entrapped mycelium. The immobilized hydroxylation system was more stable than the system in free mycelium and could be reactivated by incubation of the entrapped mycelium in potato dextrose broth containing cortexolone. The system was tested 50 times during 100 days of operation and was found to carry out the desired transformation with overall yields of 60%. Images PMID:16346194

  16. Highly cross-linked nanoporous polymers

    DOEpatents

    Steckle, W.P. Jr.; Apen, P.G.; Mitchell, M.A.

    1998-01-20

    Condensation polymerization followed by a supercritical extraction step can be used to obtain highly cross-linked nanoporous polymers with high surface area, controlled pore sizes and rigid structural integrity. The invention polymers are useful for applications requiring separation membranes. 1 fig.

  17. Dynamic and static fluctuations in polymer gels studied by neutron spin-echo

    NASA Astrophysics Data System (ADS)

    Kanaya, T.; Takahashi, N.; Nishida, K.; Seto, H.; Nagao, M.; Takeba, Y.

    2006-11-01

    We report neutron spin-echo measurements on three types of poly(vinyl alcohol) (PVA) gels. The first is PVA gel in a mixture of dimethyl sulfoxide (DMSO) and water with volume ratio 60/40, the second is PVA gel in an aqueous borax solution and the third is chemically cross-linked PVA gel. The observed normalized intermediate scattering functions I( Q, t)/ I( Q,0) were very different among them. The I( Q, t)/ I( Q,0) of the first and third gels showed a non-decaying component in addition to a decaying component, but the second one did not have the non-decaying one. This clearly indicates that the fluctuations in the first and third PVA gels consist of static and dynamic fluctuations whereas the second PVA gel does include only the dynamic fluctuations. The dynamic and static fluctuations of the PVA gels were analyzed in terms of a restricted motion in the gel network and the Zimm motion, respectively.

  18. Probing structures of large protein complexes using zero-length cross-linking.

    PubMed

    Rivera-Santiago, Roland F; Sriswasdi, Sira; Harper, Sandra L; Speicher, David W

    2015-11-01

    Structural mass spectrometry (MS) is a field with growing applicability for addressing complex biophysical questions regarding proteins and protein complexes. One of the major structural MS approaches involves the use of chemical cross-linking coupled with MS analysis (CX-MS) to identify proximal sites within macromolecules. Identified cross-linked sites can be used to probe novel protein-protein interactions or the derived distance constraints can be used to verify and refine molecular models. This review focuses on recent advances of "zero-length" cross-linking. Zero-length cross-linking reagents do not add any atoms to the cross-linked species due to the lack of a spacer arm. This provides a major advantage in the form of providing more precise distance constraints as the cross-linkable groups must be within salt bridge distances in order to react. However, identification of cross-linked peptides using these reagents presents unique challenges. We discuss recent efforts by our group to minimize these challenges by using multiple cycles of LC-MS/MS analysis and software specifically developed and optimized for identification of zero-length cross-linked peptides. Representative data utilizing our current protocol are presented and discussed. Copyright © 2015 Elsevier Inc. All rights reserved.

  19. Study on the preparation process of cross-linked porous cassava starch

    NASA Astrophysics Data System (ADS)

    Yin, Xiulian; You, Qinghong; Wan, Miaomiao; Zhang, Xuejuan; Dai, Chunhua

    2017-04-01

    Using cassava starch as raw material, preparation process of porous cross-linked cassava starch was studied. Using TSTP as cross-linking agents, Orthogonal design was applied for the optimization of cross-linked porous starch preparation process. The results showed that the opitmal conditions of cross-linked porous cassava starch were as follows: reaction temperature 45°C, reaction time 20 h, 1% of the amount of the enzyme, the enzyme ratio of 1:5, pH 5.50, substrate concentration of 40%.

  20. Enrichment of Cross-Linked Peptides Using Charge-Based Fractional Diagonal Chromatography (ChaFRADIC).

    PubMed

    Tinnefeld, Verena; Venne, A Saskia; Sickmann, Albert; Zahedi, René P

    2017-02-03

    Chemical cross-linking of proteins is an emerging field with huge potential for the structural investigation of proteins and protein complexes. Owing to the often relatively low yield of cross-linking products, their identification in complex samples benefits from enrichment procedures prior to mass spectrometry analysis. So far, this is mainly accomplished by using biotin moieties in specific cross-linkers or by applying strong cation exchange chromatography (SCX) for a relatively crude enrichment. We present a novel workflow to enrich cross-linked peptides by utilizing charge-based fractional diagonal chromatography (ChaFRADIC). On the basis of two-dimensional diagonal SCX separation, we could increase the number of identified cross-linked peptides for samples of different complexity: pure cross-linked BSA, cross-linked BSA spiked into a simple protein mixture, and cross-linked BSA spiked into a HeLa lysate. We also compared XL-ChaFRADIC with size exclusion chromatography-based enrichment of cross-linked peptides. The XL-ChaFRADIC approach is straightforward, reproducible, and independent of the cross-linking chemistry and cross-linker properties.

  1. Effect of Carboxymethylation on the Rheological Properties of Hyaluronan

    PubMed Central

    Wendling, Rian J.; Christensen, Amanda M.; Quast, Arthur D.; Atzet, Sarah K.; Mann, Brenda K.

    2016-01-01

    Chemical modifications made to hyaluronan to enable covalent crosslinking to form a hydrogel or to attach other molecules may alter the physical properties as well, which have physiological importance. Here we created carboxymethyl hyaluronan (CMHA) with varied degree of modification and investigated the effect on the viscosity of CMHA solutions. Viscosity decreased initially as modification increased, with a minimum viscosity for about 30–40% modification. This was followed by an increase in viscosity around 45–50% modification. The pH of the solution had a variable effect on viscosity, depending on the degree of carboxymethyl modification and buffer. The presence of phosphates in the buffer led to decreased viscosity. We also compared large-scale production lots of CMHA to lab-scale and found that large-scale required extended reaction times to achieve the same degree of modification. Finally, thiolated CMHA was disulfide crosslinked to create hydrogels with increased viscosity and shear-thinning aspects compared to CMHA solutions. PMID:27611817

  2. The role of temperature in forming sol-gel biocomposites containing polydopamine.

    PubMed

    Dyke, Jason Christopher; Hu, Huamin; Lee, Dong Joon; Ko, Ching-Chang; You, Wei

    2014-11-28

    To further improve the physical strength and biomedical applicability of bioceramicsbuilt on hydroxyapatite-gelatin (HAp-Gel) and siloxane sol-gel reactions, we incorporated mussel adhesive inspired polydopamine (PD) into our original composite based on HAp-Gel cross-linked with siloxane. Surprisingly, with the addition of PD, we observed that the processing conditions and temperatures play an important role in the structure and performance of these materials. A systematic study to investigate this temperature dependence behavior discloses that the rate of crosslinking of silane during the sol-gel process is significantly influenced by the temperature, whereas the polymerization of the dopamine only shows minor temperature dependence. With this discovery, we report an innovative thermal process for the design and application of these biocomposites.

  3. The role of temperature in forming sol-gel biocomposites containing polydopamine

    PubMed Central

    Dyke, Jason Christopher; Hu, Huamin; Lee, Dong Joon; Ko, Ching-Chang; You, Wei

    2014-01-01

    To further improve the physical strength and biomedical applicability of bioceramicsbuilt on hydroxyapatite-gelatin (HAp-Gel) and siloxane sol-gel reactions, we incorporated mussel adhesive inspired polydopamine (PD) into our original composite based on HAp-Gel cross-linked with siloxane. Surprisingly, with the addition of PD, we observed that the processing conditions and temperatures play an important role in the structure and performance of these materials. A systematic study to investigate this temperature dependence behavior discloses that the rate of crosslinking of silane during the sol-gel process is significantly influenced by the temperature, whereas the polymerization of the dopamine only shows minor temperature dependence. With this discovery, we report an innovative thermal process for the design and application of these biocomposites. PMID:25485111

  4. Citric-Acid-Derived Photo-cross-Linked Biodegradable Elastomers

    PubMed Central

    Gyawali, Dipendra; Tran, Richard T.; Guleserian, Kristine J.; Tang, Liping; Yang, Jian

    2010-01-01

    Citric-acid-derived thermally cross-linked biodegradable elastomers (CABEs) have recently received significant attention in various biomedical applications, including tissue-engineering orthopedic devices, bioimaging and implant coatings. However, citric-acid-derived photo-cross-linked biodegradable elastomers are rarely reported. Herein, we report a novel photo-cross-linked biodegradable elastomer, referred to as poly(octamethylene maleate citrate) (POMC), which preserves pendant hydroxyl and carboxylic functionalities after cross-linking for the potential conjugation of biologically active molecules. POMC is a low-molecular-mass pre-polymer with a molecular mass average between 701 and 1291 Da. POMC networks are soft and elastic with an initial modulus of 0.07 to 1.3 MPa and an elongation at break between 38 and 382%. FT-IR–ATR results confirmed the successful surface immobilization of type-I collagen onto POMC films, which enhanced in vitro cellular attachment and proliferation. Photo-polymerized POMC films implanted subcutaneously into Sprague–Dawley rats demonstrated minimal in vivo inflammatory responses. The development of POMC enriches the family of citric-acid-derived biodegradable elastomers and expands the available biodegradable polymers for versatile needs in biomedical applications. PMID:20557687

  5. Cross-linked polymeric membranes for carbon dioxide separation

    DOEpatents

    Hong, Tao; Chatterjee, Sabornie; Mahurin, Shannon Mark; Long, Brian Keith; Jiang, De-en; Mays, Jimmy Wayne; Sokolov, Alexei P.; Saito, Tomonori

    2018-01-23

    A membrane useful in gas separation, the membrane comprising a cross-linked polysiloxane structure having a cross-link density of about 0.1.times.10.sup.-5 mol/cm.sup.3 to about 6.times.10.sup.-5 mol/cm.sup.3, where, in particular embodiments, the cross-linked polysiloxane structure has the following general structure: ##STR00001## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5, and R.sup.6 are independently selected from hydrocarbon groups having at least 1 and up to 6 carbon atoms; A.sup.1 and A.sup.2 are independently selected from cyclic hydrocarbon groups; L.sup.1 and L.sup.2 are linking groups or covalent bonds; n is an integer of at least 1; r and s are independently selected from integers of at least 1; and p is an integer of at least 10. The invention also includes methods for making and using the above-described membranes for gas separation.

  6. Cross-Linked Enzyme Aggregates for Applications in Aqueous and Nonaqueous Media.

    PubMed

    Roy, Ipsita; Mukherjee, Joyeeta; Gupta, Munishwar N

    2017-01-01

    Extensive cross-linking of a precipitate of a protein by a cross-linking reagent (glutaraldehyde has been most commonly used) creates an insoluble enzyme preparation called cross-linked enzyme aggregates (CLEAs). CLEAs show high stability and performance in conventional aqueous as well as nonaqueous media. These are also stable at fairly high temperatures. CLEAs with more than one kind of enzyme activity can be prepared, and such CLEAs are called combi-CLEAs or multipurpose CLEAs. Extent of cross-linking often influences their morphology, stability, activity, and enantioselectivity.

  7. A General Method for Targeted Quantitative Cross-Linking Mass Spectrometry.

    PubMed

    Chavez, Juan D; Eng, Jimmy K; Schweppe, Devin K; Cilia, Michelle; Rivera, Keith; Zhong, Xuefei; Wu, Xia; Allen, Terrence; Khurgel, Moshe; Kumar, Akhilesh; Lampropoulos, Athanasios; Larsson, Mårten; Maity, Shuvadeep; Morozov, Yaroslav; Pathmasiri, Wimal; Perez-Neut, Mathew; Pineyro-Ruiz, Coriness; Polina, Elizabeth; Post, Stephanie; Rider, Mark; Tokmina-Roszyk, Dorota; Tyson, Katherine; Vieira Parrine Sant'Ana, Debora; Bruce, James E

    2016-01-01

    Chemical cross-linking mass spectrometry (XL-MS) provides protein structural information by identifying covalently linked proximal amino acid residues on protein surfaces. The information gained by this technique is complementary to other structural biology methods such as x-ray crystallography, NMR and cryo-electron microscopy[1]. The extension of traditional quantitative proteomics methods with chemical cross-linking can provide information on the structural dynamics of protein structures and protein complexes. The identification and quantitation of cross-linked peptides remains challenging for the general community, requiring specialized expertise ultimately limiting more widespread adoption of the technique. We describe a general method for targeted quantitative mass spectrometric analysis of cross-linked peptide pairs. We report the adaptation of the widely used, open source software package Skyline, for the analysis of quantitative XL-MS data as a means for data analysis and sharing of methods. We demonstrate the utility and robustness of the method with a cross-laboratory study and present data that is supported by and validates previously published data on quantified cross-linked peptide pairs. This advance provides an easy to use resource so that any lab with access to a LC-MS system capable of performing targeted quantitative analysis can quickly and accurately measure dynamic changes in protein structure and protein interactions.

  8. Virtual Cross-Linking of the Active Nemorubicin Metabolite PNU-159682 to Double-Stranded DNA.

    PubMed

    Scalabrin, Matteo; Quintieri, Luigi; Palumbo, Manlio; Riccardi Sirtori, Federico; Gatto, Barbara

    2017-02-20

    The DNA alkylating mechanism of PNU-159682 (PNU), a highly potent metabolite of the anthracycline nemorubicin, was investigated by gel-electrophoretic, HPLC-UV, and micro-HPLC/mass spectrometry (MS) measurements. PNU quickly reacted with double-stranded oligonucleotides, but not with single-stranded sequences, to form covalent adducts which were detectable by denaturing polyacrylamide gel electrophoresis (DPAGE). Ion-pair reverse-phase HPLC-UV analysis on CG rich duplex sequences having a 5'-CCCGGG-3' central core showed the formation of two types of adducts with PNU, which were stable and could be characterized by micro-HPLC/MS. The first type contained one alkylated species (and possibly one reversibly bound species), and the second contained two alkylated species per duplex DNA. The covalent adducts were found to produce effective bridging of DNA complementary strands through the formation of virtual cross-links reminiscent of those produced by classical anthracyclines in the presence of formaldehyde. Furthermore, the absence of reactivity of PNU with CG-rich sequence containing a TA core (CGTACG), and the minor reactivity between PNU and CGC sequences (TACGCG·CGCGTA) pointed out the importance of guanine sequence context in modulating DNA alkylation.

  9. An Investigation of Siloxane Cross-linked Hydroxyapatite-Gelatin/Copolymer Composites for Potential Orthopedic Applications†

    PubMed Central

    Dyke, Jason Christopher; Knight, Kelly Jane; Zhou, Huaxing; Chiu, Chi-Kai; Ko, Ching-Chang; You, Wei

    2012-01-01

    Causes of bone deficiency are numerous, but biomimetic alloplastic grafts provide an alternative to repair tissue naturally. Previously, a hydroxyapatite-gelatin modified siloxane (HAp-Gemosil) composite was prepared by cross-linking (N, N′-bis[(3-trimethoxysilyl)propyl]ethylene diamine (enTMOS) around the HAp-Gel nanocomposite particles, to mimic the natural composition and properties of bone. However, the tensile strength remained too low for many orthopedic applications. It was hypothesized that incorporating a polymer chain into the composite could help improve long range interaction. Furthermore, designing this polymer to interact with the enTMOS siloxane cross-linked matrix would provide improved adhesion between the polymer and the ceramic composite, and improve mechanical properties. To this end, copolymers of L-Lactide (LLA), and a novel alkyne derivatized trimethylene carbonate, propargyl carbonate (PC), were synthesized. Incorporation of PC during copolymerization affects properties of copolymers such as molecular weight, Tg, and % PC incorporation. More importantly, PC monomers bear a synthetic handle, allowing copolymers to undergo post-polymerization functionalization with graft monomers to specifically tailor the properties of the final composite. For our investigation, P(LLA-co-PC) copolymers were functionalized by an azido-silane (AS) via copper catalyzed azide-alkyne cycloaddition (CuAAC) through terminal alkyne on PC monomers. The new functionalized polymer, P(LLA-co-PC)(AS) was blended with HAp-Gemosil, with the azido-silane linking the copolymer to the silsesquioxane matrix within the final composite. These HAp-Gemosil/P(LLA-co-PC)(AS) composites were subjected to mechanical and biological testing, and the results were compared with those from the HAp-Gemosil composites. This study revealed that incorporating a cross-linkable polymer served to increase the flexural strength of the composite by 50%, while maintaining the biocompatibility of

  10. Polymerase cross-linking spiral reaction (PCLSR) for detection of African swine fever virus (ASFV) in pigs and wild boars.

    PubMed

    Woźniakowski, Grzegorz; Frączyk, Magdalena; Kowalczyk, Andrzej; Pomorska-Mól, Małgorzata; Niemczuk, Krzysztof; Pejsak, Zygmunt

    2017-02-15

    The study reports the development of a polymerase cross-linking spiral reaction (PCLSR) for the detection of African swine fever virus (ASFV) DNA in blood collected from infected pigs and wild boars. The method uses 3 specifically designed primers. Two outer-spiral primers comprising of 3' sequences complementary to ASFV p72 gene sequence and 5'end sequences complementary to exogenous gene of black widow alpha-latrotoxin as well as additional ASFV specific cross-linking primer. The method is specific exclusively to ASFV DNA without cross-reactions with cDNA of classical swine fever virus (CSFV), porcine reproductive respiratory syndrome (PRRSV) or porcine epidemic diarrhea virus (PEDV). The sensitivity of this technique reached 7.2 × 10 2 copies per μl -1 of plasmid containing p72 gene. The PCLSR was conducted at 65 °C creating cross-linked complex structures. The results of PCLSR were visualized using SYBR Green I dye, gel electrophoresis while the reaction progress was traced using real-time PCR system that resulted in registration of fluorescent curves and melting peaks at 85.3 °C. The developed PCLSR was examined using blood or tissue samples collected from selected 17 ASF cases from infected wild boars and 3 outbreaks in pigs. Further tests have been also conducted using 55 tissue samples from 23 outbreaks and 22 cases. These results showed that PCLSR might be further used for preliminary and cost-effective detection and surveillance of ASFV.

  11. Injectable biocompatible and biodegradable pH-responsive hollow particle gels containing poly(acrylic acid): the effect of copolymer composition on gel properties.

    PubMed

    Halacheva, Silvia S; Adlam, Daman J; Hendow, Eseelle K; Freemont, Tony J; Hoyland, Judith; Saunders, Brian R

    2014-05-12

    The potential of various pH-responsive alkyl (meth)acrylate ester- and (meth)acrylic acid-based copolymers, including poly(methyl methacrylate-co-acrylic acid) (PMMA-AA) and poly(n-butyl acrylate-co-methacrylic acid) (PBA-MAA), to form pH-sensitive biocompatible and biodegradable hollow particle gel scaffolds for use in non-load-bearing soft tissue regeneration have been explored. The optimal copolymer design criteria for preparation of these materials have been established. Physical gels which are both pH- and redox-sensitive were formed only from PMMA-AA copolymers. MMA is the optimal hydrophobic monomer, whereas the use of various COOH-containing monomers, e.g., MAA and AA, will always induce a pH-triggered physical gelation. The PMMA-AA gels were prepared at physiological pH range from concentrated dispersions of swollen, hollow, polymer-based particles cross-linked with either cystamine (CYS) or 3,3'-dithiodipropionic acid dihydrazide (DTP). A linear relationship between particle swelling ratios, gel elasticity, and ductility was observed. The PMMA-AA gels with lower AA contents feature lower swelling ratios, mechanical strengths, and ductilities. Increasing the swelling ratio (e.g., through increasing AA content) decreased the intraparticle elasticity; however, intershell contact and gel elasticity were found to increase. The mechanical properties and performance of the gels were tuneable upon varying the copolymers' compositions and the structure of the cross-linker. Compared to PMMA-AA/CYS, the PMMA-AA/DTP gels were more elastic and ductile. The biodegradability and cytotoxicity of the new hollow particle gels were tested for the first time and related to their composition, mechanical properties, and morphology. The new PMMA-AA/CYS and PMMA-AA/DTP gels have shown good biocompatibility, biodegradability, strength, and interconnected porosity and therefore have good potential as a tissue repair agent.

  12. Molecular Model for HNBR with Tunable Cross-Link Density.

    PubMed

    Molinari, N; Khawaja, M; Sutton, A P; Mostofi, A A

    2016-12-15

    We introduce a chemically inspired, all-atom model of hydrogenated nitrile butadiene rubber (HNBR) and assess its performance by computing the mass density and glass-transition temperature as a function of cross-link density in the structure. Our HNBR structures are created by a procedure that mimics the real process used to produce HNBR, that is, saturation of the carbon-carbon double bonds in NBR, either by hydrogenation or by cross-linking. The atomic interactions are described by the all-atom "Optimized Potentials for Liquid Simulations" (OPLS-AA). In this paper, first, we assess the use of OPLS-AA in our models, especially using NBR bulk properties, and second, we evaluate the validity of the proposed model for HNBR by investigating mass density and glass transition as a function of the tunable cross-link density. Experimental densities are reproduced within 3% for both elastomers, and qualitatively correct trends in the glass-transition temperature as a function of monomer composition and cross-link density are obtained.

  13. Probing actin polymerization by intermolecular cross-linking.

    PubMed

    Millonig, R; Salvo, H; Aebi, U

    1988-03-01

    We have used N,N'-1,4-phenylenebismaleimide, a bifunctional sulfhydryl cross-linking reagent, to probe the oligomeric state of actin during the early stages of its polymerization into filaments. We document that one of the first steps in the polymerization of globular monomeric actin (G-actin) under a wide variety of ionic conditions is the dimerization of a significant fraction of the G-actin monomer pool. As polymerization proceeds, the yield of this initial dimer ("lower" dimer with an apparent molecular mass of 86 kD by SDS-PAGE [LD]) is attenuated, while an actin filament dimer ("upper" dimer with an apparent molecular mass of 115 kD by SDS-PAGE [UD] as characterized [Elzinga, M., and J. J. Phelan. 1984. Proc. Natl. Acad. Sci. USA. 81:6599-6602]) is formed. This shift from LD to UD occurs concomitant with formation of filaments as assayed by N-(1-pyrenyl)iodoacetamide fluorescence enhancement and electron microscopy. Isolated cross-linked LD does not form filaments, while isolated cross-linked UD will assemble into filaments indistinguishable from those polymerized from unmodified G-actin under typical filament-forming conditions. The presence of cross-linked LD does not effect the kinetics of polymerization of actin monomer, whereas cross-linked UD shortens the "lag phase" of the polymerization reaction in a concentration-dependent fashion. Several converging lines of evidence suggest that, although accounting for a significant oligomeric species formed during early polymerization, the LD is incompatible with the helical symmetry defining the mature actin filament; however, it could represent the interfilament dimer found in paracrystalline arrays or filament bundles. Furthermore, the LD is compatible with the unit cell structure and symmetry common to various types of crystalline actin arrays (Aebi, U., W. E. Fowler, G. Isenberg, T. D. Pollard, and P. R. Smith. 1981. J. Cell Biol. 91:340-351) and might represent the major structural state in which a mutant

  14. A Study into the Collision-induced Dissociation (CID) Behavior of Cross-Linked Peptides*

    PubMed Central

    Giese, Sven H.; Fischer, Lutz; Rappsilber, Juri

    2016-01-01

    Cross-linking/mass spectrometry resolves protein–protein interactions or protein folds by help of distance constraints. Cross-linkers with specific properties such as isotope-labeled or collision-induced dissociation (CID)-cleavable cross-linkers are in frequent use to simplify the identification of cross-linked peptides. Here, we analyzed the mass spectrometric behavior of 910 unique cross-linked peptides in high-resolution MS1 and MS2 from published data and validate the observation by a ninefold larger set from currently unpublished data to explore if detailed understanding of their fragmentation behavior would allow computational delivery of information that otherwise would be obtained via isotope labels or CID cleavage of cross-linkers. Isotope-labeled cross-linkers reveal cross-linked and linear fragments in fragmentation spectra. We show that fragment mass and charge alone provide this information, alleviating the need for isotope-labeling for this purpose. Isotope-labeled cross-linkers also indicate cross-linker-containing, albeit not specifically cross-linked, peptides in MS1. We observed that acquisition can be guided to better than twofold enrich cross-linked peptides with minimal losses based on peptide mass and charge alone. By help of CID-cleavable cross-linkers, individual spectra with only linear fragments can be recorded for each peptide in a cross-link. We show that cross-linked fragments of ordinary cross-linked peptides can be linearized computationally and that a simplified subspectrum can be extracted that is enriched in information on one of the two linked peptides. This allows identifying candidates for this peptide in a simplified database search as we propose in a search strategy here. We conclude that the specific behavior of cross-linked peptides in mass spectrometers can be exploited to relax the requirements on cross-linkers. PMID:26719564

  15. Inverted bulk-heterojunction solar cell with cross-linked hole-blocking layer

    PubMed Central

    Udum, Yasemin; Denk, Patrick; Adam, Getachew; Apaydin, Dogukan H.; Nevosad, Andreas; Teichert, Christian; S. White, Matthew.; S. Sariciftci, Niyazi.; Scharber, Markus C.

    2014-01-01

    We have developed a hole-blocking layer for bulk-heterojunction solar cells based on cross-linked polyethylenimine (PEI). We tested five different ether-based cross-linkers and found that all of them give comparable solar cell efficiencies. The initial idea that a cross-linked layer is more solvent resistant compared to a pristine PEI layer could not be confirmed. With and without cross-linking, the PEI layer sticks very well to the surface of the indium–tin–oxide electrode and cannot be removed by solvents used to process PEI or common organic semiconductors. The cross-linked PEI hole-blocking layer functions for multiple donor–acceptor blends. We found that using cross-linkers improves the reproducibility of the device fabrication process. PMID:24817837

  16. A Review of Collagen Cross-Linking in Cornea and Sclera

    PubMed Central

    Zhang, Xiao; Tao, Xiang-chen; Zhang, Jian; Li, Zhi-wei; Xu, Yan-yun; Wang, Yu-meng; Zhang, Chun-xiao; Mu, Guo-ying

    2015-01-01

    Riboflavin/UVA cross-linking is a technique introduced in the past decades for the treatment of keratoconus, keratectasia, and infectious keratitis. Its efficacy and safety have been investigated with clinical and laboratory studies since its first clinical application by Wollensak for the treatment of keratoconus. Although its complications are encountered during clinical practice, such as infection inducing risk, minimal invasion merits a further investigation on its future application in clinical practice. Recently, collagen cross-linking in sclera shows a promising prospect. In present study, we summarized the representative studies describing the clinical and laboratory application of collagen cross-linking published in past decades and provided our opinion on the positive and negative results of cross-linking in the treatment of ophthalmic disorders. PMID:25922758

  17. Inhibition of Ovarian Cancer Chemoresistance and Metastasis with Antagonists of Hyaluronan-CD44-CD147 Interactions

    DTIC Science & Technology

    2015-09-01

    malignant and drug- resistant properties. This most likely occurs through assembly and/or stabilization of plasma membrane signaling complexes ...interactions of hyaluronan polymer with CD44 are necessary for stabilizing CD44-CD147 signaling complexes , and that small, monovalent, hyaluronan...transporter complexes (Ghatak et al., 2005; Grass et al., 2012; Grass et al., 2013; Qin et al., 2011; Slomiany et al., 2009a; Slomiany et al., 2009b

  18. Atomic force microscopy evaluation of aqueous interfaces of immobilized hyaluronan.

    PubMed

    Morra, Marco; Cassinelli, Clara; Pavesio, Alessandra; Renier, Davide

    2003-03-15

    Hyaluronan (HA) was immobilized on aminated glass surfaces in three different ways: by simple ionic interaction and by covalent linking at low density and at full density. In agreement with previous reports, in vitro experiments show that the outcome of fibroblast adhesion tests is markedly affected by the details of the coupling procedure, suggesting that different interfacial forces are operating at the aqueous/HA interface in the three cases investigated. The interfacial properties of the HA-coated surfaces were probed by force-distance curves obtained with the atomic force microscope (AFM). This approach readily shows significant differences among the tested samples, which are directly related to the coupling strategy and to results of cell adhesion tests. In particular, the range of interaction between the tip and the surface is much lower when HA is covalently linked than when it is ionically coupled, suggesting a more compact surface structure in the former case. Increasing HA surface density minimizes the interaction force between the surface and the AFM tip, likely reflecting more complete shielding by the HA chains of the underlying substrate. In summary, these measurements clearly show the different nature of the aqueous interfaces tested, and underline the role of this analytical approach in the development and control of finely tuned biomaterial surfaces.

  19. Hierarchical Sol-Gel Transition Induced by Thermosensitive Self-Assembly of an ABC Triblock Polymer in an Ionic Liquid

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Kitazawa, Yuzo; Ueki, Takeshi; McIntosh, Lucas D.

    2016-04-29

    Here we investigate a hierarchical morphology change and accompanying sol–gel transition using a doubly thermosensitive ABC-triblock copolymer in an ionic liquid (IL). The triblock copolymer contains two different lower critical solution temperature (LCST) thermosensitive polymers, poly(benzyl methacrylate) (PBnMA) and poly(2-phenylethyl methacrylate) (PPhEtMA), as the end blocks and poly(methyl methacrylate) (PMMA) as the middle block (PBnMA-b-PMMA-b-PPhEtMA: BMP). BMP undergoes a hierarchical phase transition corresponding to the self-assembly of each of the thermosensitive blocks in the IL, and a sol–gel transition was observed in concentrated, above 10 wt %, polymer solutions. The gelation behavior was affected by polymer concentration, and at 20more » wt %, the BMP/IL composite showed a phase transition, with increasing temperature, from solution through a jammed micelle suspension to a physically cross-linked gel. For each phase was formed reversibly and rapidly over the corresponding temperature range. Finally, the jammed micelle and cross-linked gel states were characterized using viscoelastic measurements and small-angle X-ray scattering (SAXS).« less

  20. Synthesis, Characterization, and Antibacterial Activity of Cross-Linked Chitosan-Glutaraldehyde

    PubMed Central

    Li, Bin; Shan, Chang-Lin; Zhou, Qing; Fang, Yuan; Wang, Yang-Li; Xu, Fei; Han, Li-Rong; Ibrahim, Muhammad; Guo, Long-Biao; Xie, Guan-Lin; Sun, Guo-Chang

    2013-01-01

    This present study deals with synthesis, characterization and antibacterial activity of cross-linked chitosan-glutaraldehyde. Results from this study indicated that cross-linked chitosan-glutaraldehyde markedly inhibited the growth of antibiotic-resistant Burkholderia cepacia complex regardless of bacterial species and incubation time while bacterial growth was unaffected by solid chitosan. Furthermore, high temperature treated cross-linked chitosan-glutaraldehyde showed strong antibacterial activity against the selected strain 0901 although the inhibitory effects varied with different temperatures. In addition, physical-chemical and structural characterization revealed that the cross-linking of chitosan with glutaraldehyde resulted in a rougher surface morphology, a characteristic Fourier transform infrared (FTIR) band at 1559 cm−1, a specific X-ray diffraction peak centered at 2θ = 15°, a lower contents of carbon, hydrogen and nitrogen, and a higher stability of glucose units compared to chitosan based on scanning electron microscopic observation, FTIR spectra, X-ray diffraction pattern, as well as elemental and thermo gravimetric analysis. Overall, this study indicated that cross-linked chitosan-glutaraldehyde is promising to be developed as a new antibacterial drug. PMID:23670533

  1. Ice templated and cross-linked xylan/nanocrystalline cellulose hydrogels

    Treesearch

    Tobias Köhnke; Thomas Elder; Hans Theliander; Arthur J. Ragauskas

    2014-01-01

    Structured xylan-based hydrogels, reinforced with cellulose nanocrystals (CNCs), have successfully been prepared from water suspensions by cross-linking during freeze-casting. In order to induce cross-linking during the solidification/sublimation operation, xylan was first oxidized using sodium periodate to introduce dialdehydes. The oxidized xylan was then mixed with...

  2. UV cross-linking of donor corneas confers resistance to keratolysis.

    PubMed

    Arafat, Samer N; Robert, Marie-Claude; Shukla, Anita N; Dohlman, Claes H; Chodosh, James; Ciolino, Joseph B

    2014-09-01

    The aim of this study was to develop a modified ex vivo corneal cross-linking method that increases stromal resistance to enzymatic degradation for use as a carrier for the Boston keratoprosthesis. Ex vivo cross-linking of human corneas was performed using Barron artificial anterior chambers. The corneas were deepithelialized, pretreated with riboflavin solution (0.1% riboflavin/20% dextran), and irradiated with ultraviolet A (UV-A) light (λ = 370 nm, irradiance = 3 mW/cm) for various durations. The combined effect of UV-A and gamma (γ) irradiation was also assessed using the commercially available γ-irradiated corneal donors. The corneas were then trephined and incubated at 37°C with 0.3% collagenase A solution. The time to dissolution of each cornea was compared across treatments. Deepithelialized corneas (no UV light, no riboflavin) dissolved in 5.8 ± 0.6 hours. Cross-linked corneas demonstrated increased resistance to dissolution, with a time to dissolution of 17.8 ± 2.6 hours (P < 0.0001). The corneal tissues' resistance to collagenase increased with longer UV-A exposure, reaching a plateau at 30 minutes. Cross-linking both the anterior and posterior corneas did not provide added resistance when compared with cross-linking the anterior corneas only (P > 0.05). γ-irradiated corneas dissolved as readily as deepithelialized controls regardless of whether they were further cross-linked (5.6 ± 1.2 hours) or not (6.1 ± 0.6 hours) (P = 0.43). Collagen cross-linking of the deepithelialized anterior corneal surface for 30 minutes conferred optimal resistance to in vitro keratolysis by collagenase A.

  3. Effects of ventilation on hyaluronan and protein concentration in pleural liquid of anesthetized and conscious rabbits.

    PubMed

    Wang, P M; Lai-Fook, S J

    1998-01-01

    The hypothesis of this study is that pleural lubrication is enhanced by hyaluronan acting as a boundary lubricant in pleural liquid and by pleural filtration as reflected in changes in protein concentration with ventilation. Anesthetized rabbits were injected intravenously with Evans blue dye and ventilated with 100% O2 at either of two levels of ventilation for 6 h. Postmortem values of hyaluronan, total protein, and Evans blue-dyed albumin (EBA) concentrations in pleural liquid were greater at the higher ventilation, consistent with increases in boundary lubrication, pleural membrane permeability, and pleural filtration. To determine whether these effects were caused by hyperoxia or anesthesia, conscious rabbits were ventilated with either 3% CO2 or room air in a box for 6, 12, or 24 h. Similar to the anesthetized rabbits, pleural liquid hyaluronan concentration after 24 h was higher in the conscious rabbits with the hypercapnic-induced greater ventilation. By contrast, the time course of total protein and EBA in pleural liquid was similar in both groups of conscious rabbits, indicating no effect of ventilation on pleural permeability. The increase in pleural liquid hyaluronan concentration might be the result of mesothelial cell stimulation by a ventilation-induced increase in pleural liquid shear stress.

  4. Collagen cross-linking of skin in patients with amyotrophic lateral sclerosis

    NASA Technical Reports Server (NTRS)

    Ono, S.; Yamauchi, M.

    1992-01-01

    Collagen cross-links of skin tissue (left upper arm) from 11 patients with amyotrophic lateral sclerosis (ALS) and 9 age-matched control subjects were quantified. It was found that patients with ALS had a significant reduction in the content of an age-related, stable cross-link, histidinohydroxylysinonorleucine, that was negatively correlated with the duration of illness. The contents of sodium borohydride-reducible labile cross-links, dehydro-hydroxylysinonorleucine and dehydro-histidinohydroxymerodesmosine, were significantly increased and were positively associated with the duration of illness (r = 0.703, p less than 0.05 and r = 0.684, p less than 0.05, respectively). The results clearly indicate that during the course of ALS, the cross-linking pathway of skin collagen runs counter to its normal aging, resulting in a "rejuvenation" phenomenon of skin collagen. Thus, cross-linking of skin collagen is affected in ALS.

  5. Thermo-and pH-sensitive hydrogel membranes composed of poly(N-isopropylacrylamide)-hyaluronan for biomedical applications: Influence of hyaluronan incorporation on the membrane properties.

    PubMed

    Kamoun, Elbadawy A; Fahmy, Alaa; Taha, Tarek H; El-Fakharany, Esmail M; Makram, Mohamed; Soliman, Hesham M A; Shehata, Hassan

    2018-01-01

    Interpenetrating hydrogel membranes consisting of pH-sensitive hyaluronan (HA) and thermo-sensitive poly(N-isopropylacrylamide) (PNIPAAM) were synthesized using redox polymerization, followed by N,N-methylenebisacrylamide (BIS) and epichlorohydrin (EPI) were added as chemical crosslinkers. The interaction between membrane compositions has been characterized by FTIR spectroscopy and discussed intensively. The result indicates that HA incorporation in membranes increase the gel fraction, swelling uptake, and the flexibility/elasticity of crosslinked membranes, however it reduced oppositely the mechanical elongation of membranes. PNIPAAm-HA hydrogels responded to both temperature and pH changes and the stimuli-responsiveness was reversible. However, in vitro bioevaluation results revealed that the released ampicillin during the burst release time was sharply influenced and increased with increasing HA contents in membranes; afterwards it became sustainable. Whereas, high HA contents in hydrogels unexpectedly impacted negatively on the cells viability, owing to the viscosity of cell culture media changed. A big resistance was observed against microbial growth of Staphylococcus aureus, Salmonella typhi, and Candida albicans in case of pure PNIPAAm hydrogel membranes without HA or ampicillin. However, HA incorporation or the loaded ampicillin in membranes showed unexpected easily microbial growth. The fast release performance with dual pH-thermo-sensitive hydrogels were suggested as promising materials for quick drug carrier in the biomedical field. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Polypeptide Grafted Hyaluronan: Synthesis and Characterization

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Wang, Xiaojun; Messman, Jamie M; Mays, Jimmy

    2010-01-01

    Poly(L-leucine) grafted hyaluronan (HA-g-PLeu) has been synthesized via a Michael addition reaction between primary amine terminated poly(L-leucine) and acrylate-functionalized HA (TBAHA-acrylate). The precursor hyaluronan was first functionalized with acrylate groups by reaction with acryloyl chloride in the presence of triethylamine in N,N-dimethylformamide. 1H NMR analysis of the resulting product indicated that an increase in the concentration of acryloylchoride with respect to hydroxyl groups on HA has only a moderate effect on functionalization efficiency, f. A precise control of stoichiometry was not achieved, which could be attributed to partial solubility of intermolecular aggregates and the hygroscopic nature of HA. Michael additionmore » at high [PLeu- NH2]/[acrylate]TBAHA ratios gave a molar grafting ratio of only 0.20 with respect to the repeat unit of HA, indicating grafting limitation due to insolubility of the grafted HA-g-PLeu. Soluble HA-g-PLeu graft copolymers were obtained for low grafting ratios (<0.039) with <8.6% by mass of PLeu and were characterized thoroughly using light scattering, 1H NMR, FT-IR, and AFM techniques. Light scattering experiments showed a strong hydrophobic interaction between PLeu chains, resulting in aggregates with segregated nongrafted HA segments. This yields local networks of aggregates, as demonstrated by atomic force microscopy. Circular dichroism spectroscopy showed a -sheet conformation for aggregates of poly(L-leucine).« less

  7. Ara h 2 cross-linking catalyzed by MTGase decreases its allergenicity.

    PubMed

    Wu, Zhihua; Lian, Jun; Zhao, Ruifang; Li, Kun; Li, Xin; Yang, Anshu; Tong, Ping; Chen, Hongbing

    2017-03-22

    Peanuts, whose major allergen is Ara h 2, are included among the eight major food allergens. After reduction using dithiothreitol (DTT), cross-linking of Ara h 2 could be catalyzed by microbial transglutaminase (MTGase), a widely used enzyme in the food industry. In this study, Ara h 2 cross-linking was catalyzed by MTGase after it was reduced by DTT. Using mass spectrometry and PLINK software, five cross-linkers were identified, and five linear allergen epitopes were found to be involved in the reactions. The IgE binding capacity of cross-linked Ara h 2 was found to be significantly lower compared to that of native and reduced Ara h 2. After simulated gastric fluid (SGF) digestion, the digested products of the cross-linked Ara h 2, again, had a significantly lower IgE binding capacity compared to untreated and reduced Ara h 2. Furthermore, reduced and cross-linked Ara h 2 (RC-Ara h 2) induced lower sensitization in mice, indicating its lower allergenicity. Reduction and MTGase-catalyzed cross-linking are effective methods to decrease the allergenicity of Ara h 2. The reactions involved linear allergen epitopes destroying the material basis of the allergenicity, and this might develop a new direction for protein desensitization processes.

  8. Direct evidence for the partial reflection of hyaluronan molecules by the lining of rabbit knee joints during trans-synovial flow

    PubMed Central

    Scott, D; Coleman, P J; Mason, R M; Levick, J R

    1998-01-01

    In synovial joints hydraulic and turnover studies indicate that the synovial lining may partially reflect large macromolecules like hyaluronan, despite discontinuities in the lining cell layer. The reflection hypothesis was tested directly in the present study. Solutions of high molecular weight hyaluronan were infused at controlled pressures into the cavity of rabbit knees under anaesthesia, at concentrations of 0.2 g l−1 (n = 5), 2 g l−1 (n = 5) and 4 g l−1 (n = 6). Time-averaged trans-synovial flows were 9.6, 4.8 and 2.9 μl min−1, respectively. After 5 h infusion the intra-articular fluid was mixed and sampled. Hyaluronan concentration was determined by size-exclusion chromatography. In all sixteen experiments the hyaluronan concentration in the aspirate was greater than that in the infusate (P = 0.0001, Student's paired t test). The increases averaged 2.28 ± 0.04 times at high filtration rates (0.2 g l−1 infusates; mean ± s.e.m.), 1.60 ± 0.09 times at intermediate filtration rates (2 g l−1 infusates) and 1.26 ± 0.08 times at low filtration rates (4 g l−1 infusates). Between 48 and 95% of the hyaluronan in the filtrand was retained in the joint cavity. The greater retention at 2 g l−1, viz.95%, than at 0.2 g l−1, viz.48%, was attributed to interactions between overlapping molecular domains in the more concentrated solution. It is concluded that synovial interstitial matrix can partially reflect hyaluronan molecules, and thus conserve intra-articular lubricant. PMID:9508822

  9. Linking Gestures: Cross-Cultural Variation during Instructional Analogies

    ERIC Educational Resources Information Center

    Richland, Lindsey Engle

    2015-01-01

    Deictic linking gestures, hand and arm motions that physically embody links being communicated between two or more objects in the shared communicative environment, are explored in a cross-cultural sample of mathematics instruction. Linking gestures are specifically examined here when they occur in the context of communicative analogies designed to…

  10. Cross-linked polyelectrolyte for direct methanol fuel cells applications based on a novel sulfonated cross-linker

    NASA Astrophysics Data System (ADS)

    Li, Mingyu; Zhang, Gang; Xu, Shuai; Zhao, Chengji; Han, Miaomiao; Zhang, Liyuan; Jiang, Hao; Liu, Zhongguo; Na, Hui

    2014-06-01

    A novel type of cross-linked proton exchange membrane of lower methanol permeation and high proton conductivity is prepared, based on a newly synthesized sulfonated cross-linker: carboxyl terminated benzimidazole trimer bearing sulfonic acid groups (s-BI). Compared to membranes cross-linked with non-sulfonated cross-linker (BI), SPEEK/s-BI-n membranes show higher IEC values and proton conductivities. Meanwhile, oxidative stability and mechanical property of SPEEK/s-BI-n membranes are obviously improved. Among SPEEK/s-BI-n membranes, SPEEK/s-BI-2 exhibits high proton conductivity, low swelling ratio (0.122 S cm-1 and 15.2% at 60 °C, respectively) and low methanol permeability coefficient. These results imply that the cross-linked membranes prepared with the newly sulfonated cross-linker are promising for the direct methanol fuel cells (DMFCs) application.

  11. J-integral fracture toughness and tearing modulus measurement of radiation cross-linked UHMWPE.

    PubMed

    Gomoll, A; Wanich, T; Bellare, A

    2002-11-01

    Radiation and chemical cross-linking of medical grade ultrahigh molecular weight polyethylene (UHMWPE) has recently been utilized in an effort to improve wear performance of total joint replacement components. However, reductions in mechanical properties with cross-linking are cause for concern regarding the use of cross-linked UHMWPE for high-stress applications such as in total knee replacement prostheses. In this study, the fracture behavior of radiation cross-linked UHMWPE was compared to that of uncross-linked UHMWPE. The Rice and Sorensen model that utilizes mechanical parameters obtained from uniaxial tensile and compact tension tests was used to calculate the steady state J-integral fracture toughness, Jss, for radiation cross-linked UHMWPE. Jss decreased monotonically with increase in radiation dose. UHMWPE exhibited tough, ductile tearing behavior with stable crack growth when it was cross-linked using a gamma radiation dose of 0-50 kGy. However, in cross-linked UHMWPE irradiated to a dose of 100 and 200 kGy, unstable fracture occurred spontaneously upon attaining the initial crack driving force, J1c. This indicates that a high degree of cross-linking is less desirable for high-stress applications in orthopaedic implants. However, a substantial increase in J1c, even at a low degree of cross-linking, suggests that a low degree of cross-linking may be beneficial for resistance to delamination and catastrophic failure, both of which require an initiation step for the fracture to propagate in the material. This mechanical test should, however, be considered along with fatigue tests and joint simulator testing before determination of an appropriate amount of cross-linking for total joint replacement prostheses that experience high stresses.

  12. Actin Cross-link Assembly and Disassembly Mechanics for α-Actinin and Fascin*

    PubMed Central

    Courson, David S.; Rock, Ronald S.

    2010-01-01

    Self-assembly of complex structures is commonplace in biology but often poorly understood. In the case of the actin cytoskeleton, a great deal is known about the components that include higher order structures, such as lamellar meshes, filopodial bundles, and stress fibers. Each of these cytoskeletal structures contains actin filaments and cross-linking proteins, but the role of cross-linking proteins in the initial steps of structure formation has not been clearly elucidated. We employ an optical trapping assay to investigate the behaviors of two actin cross-linking proteins, fascin and α-actinin, during the first steps of structure assembly. Here, we show that these proteins have distinct binding characteristics that cause them to recognize and cross-link filaments that are arranged with specific geometries. α-Actinin is a promiscuous cross-linker, linking filaments over all angles. It retains this flexibility after cross-links are formed, maintaining a connection even when the link is rotated. Conversely, fascin is extremely selective, only cross-linking filaments in a parallel orientation. Surprisingly, bundles formed by either protein are extremely stable, persisting for over 0.5 h in a continuous wash. However, using fluorescence recovery after photobleaching and fluorescence decay experiments, we find that the stable fascin population can be rapidly competed away by free fascin. We present a simple avidity model for this cross-link dissociation behavior. Together, these results place constraints on how cytoskeletal structures assemble, organize, and disassemble in vivo. PMID:20551315

  13. Gait patterns after intraarticular treatment of patients with osteoarthritis of the Knee - Hyaluronan versus triamcinolone: a prospective, randomized, doubleblind, monocentric study

    PubMed Central

    2009-01-01

    Objective Evaluation of gait performance and muscle activity patterns as well as clinical efficacy and safety after single intraarticular injection with hyaluronan compared with triamcinolone in patients with knee osteoarthritis. Materials and Methods This trial evaluated the influence of a single injection of hyaluronan or triamcinolone on gait pattern and muscle activity. For clinical evaluation a visual analogue scale for pain, Lequesne index, and Knee Society Score were used. Quality of life was assessed with the SF-36. Results The complete analysis was performed in 50 of 60 patients. 26 patients were treated with triamcinolone and 24 with hyaluronan. Hyaluronan treatment led to significant improvement of range of motion at hip and knee. Significant improvement could be either demonstrated for the pain scale, Lequesne and Knee Society score in both groups. Quality of life showed greater improvement in the triamcinolone group. Conclusion Single application of high-viscosity hyaluronan shows superior range of motion and pain reduction as well as improvement in clinical results. Even if there was a lack of significant differences compared to triamcinolone, this therapy classified as safe and effective in the short follow up. PMID:19380288

  14. The distribution of renal hyaluronan and the expression of hyaluronan synthases during water deprivation in the Spinifex hopping mouse, Notomys alexis.

    PubMed

    Bartolo, Ray C; Donald, John A

    2007-12-01

    Hyaluronan (HA) is a glycosaminoglycan that is synthesized by a family of enzymes called hyaluronan synthases (HASs), of which there are three isoforms (HAS1, 2 and 3) in mammals. The HASs have different tissue expression patterns and function, indicating that synthesis of HA and formation of the HA matrix may be regulated by various factors. The HA matrix has an important role in renal water handling and the production of a concentrated urine. We investigated the distribution of HA and the expression of HAS1, HAS2 and HAS3 mRNAs in the kidney of the Spinifex hopping mouse, Notomys alexis, a native Australian desert rodent that is reported to produce the most concentrated urine of any mammal. After periods of three, seven and fourteen days of water deprivation, the distribution of renal HA changed considerably, and there was a general down-regulation of HAS mRNA expression. It is proposed that the regulation of HA synthesis by the different HAS isoforms during water deprivation in N. alexis, could be influenced by the molecular mass of the HA chains produced by each isoform, followed by the rate at which the individual HAS produces HA.

  15. Targeting of Prostate Cancer with Hyaluronan-Binding Proteins

    DTIC Science & Technology

    2005-06-01

    16). Briefly, bovine nasal cartilage (Pel-Freez, Rogers, AR) was shredded with a Sure-Form blade (Stanley). extracted overnight with 4 M guanidine... nasal cartilage by affinity chromatography on hyaluronan- E Sepharose. As shown in Fig. 1, Metastatin consisted of two molecular C 40 factions as...biotinylated 490 pg/ml.) was isolated from the medium of rat fibrosarcoma cells grown tachvplesin (Fig. IB). However, this interaction was

  16. Hyaluronan Benzyl Ester as a Scaffold for Tissue Engineering

    PubMed Central

    Vindigni, Vincenzo; Cortivo, Roberta; Iacobellis, Laura; Abatangelo, Giovanni; Zavan, Barbara

    2009-01-01

    Tissue engineering is a multidisciplinary field focused on in vitro reconstruction of mammalian tissues. In order to allow a similar three-dimensional organization of in vitro cultured cells, biocompatible scaffolds are needed. This need has provided immense momentum for research on “smart scaffolds” for use in cell culture. One of the most promising materials for tissue engineering and regenerative medicine is a hyaluronan derivative: a benzyl ester of hyaluronan (HYAFF®). HYAFF® can be processed to obtain several types of devices such as tubes, membranes, non-woven fabrics, gauzes, and sponges. All these scaffolds are highly biocompatible. In the human body they do not elicit any adverse reactions and are resorbed by the host tissues. Human hepatocytes, dermal fibroblasts and keratinocytes, chondrocytes, Schwann cells, bone marrow derived mesenchymal stem cells and adipose tissue derived mesenchymal stem cells have been successfully cultured in these meshes. The same scaffolds, in tube meshes, has been applied for vascular tissue engineering that has emerged as a promising technology for the design of an ideal, responsive, living conduit with properties similar to that of native tissue. PMID:19742179

  17. Mechanical Strength Improvements of Carbon Nanotube Threads through Epoxy Cross-Linking

    PubMed Central

    Yu, Qingyue; Alvarez, Noe T.; Miller, Peter; Malik, Rachit; Haase, Mark R.; Schulz, Mark; Shanov, Vesselin; Zhu, Xinbao

    2016-01-01

    Individual Carbon Nanotubes (CNTs) have a great mechanical strength that needs to be transferred into macroscopic fiber assemblies. One approach to improve the mechanical strength of the CNT assemblies is by creating covalent bonding among their individual CNT building blocks. Chemical cross-linking of multiwall CNTs (MWCNTs) within the fiber has significantly improved the strength of MWCNT thread. Results reported in this work show that the cross-linked thread had a tensile strength six times greater than the strength of its control counterpart, a pristine MWCNT thread (1192 MPa and 194 MPa, respectively). Additionally, electrical conductivity changes were observed, revealing 2123.40 S·cm−1 for cross-linked thread, and 3984.26 S·cm−1 for pristine CNT thread. Characterization suggests that the obtained high tensile strength is due to the cross-linking reaction of amine groups from ethylenediamine plasma-functionalized CNT with the epoxy groups of the cross-linking agent, 4,4-methylenebis(N,N-diglycidylaniline). PMID:28787868

  18. Polymerase cross-linking spiral reaction (PCLSR) for detection of African swine fever virus (ASFV) in pigs and wild boars

    PubMed Central

    Woźniakowski, Grzegorz; Frączyk, Magdalena; Kowalczyk, Andrzej; Pomorska-Mól, Małgorzata; Niemczuk, Krzysztof; Pejsak, Zygmunt

    2017-01-01

    The study reports the development of a polymerase cross-linking spiral reaction (PCLSR) for the detection of African swine fever virus (ASFV) DNA in blood collected from infected pigs and wild boars. The method uses 3 specifically designed primers. Two outer-spiral primers comprising of 3′ sequences complementary to ASFV p72 gene sequence and 5′end sequences complementary to exogenous gene of black widow alpha-latrotoxin as well as additional ASFV specific cross-linking primer. The method is specific exclusively to ASFV DNA without cross-reactions with cDNA of classical swine fever virus (CSFV), porcine reproductive respiratory syndrome (PRRSV) or porcine epidemic diarrhea virus (PEDV). The sensitivity of this technique reached 7.2 × 102 copies per μl−1 of plasmid containing p72 gene. The PCLSR was conducted at 65 °C creating cross-linked complex structures. The results of PCLSR were visualized using SYBR Green I dye, gel electrophoresis while the reaction progress was traced using real-time PCR system that resulted in registration of fluorescent curves and melting peaks at 85.3 °C. The developed PCLSR was examined using blood or tissue samples collected from selected 17 ASF cases from infected wild boars and 3 outbreaks in pigs. Further tests have been also conducted using 55 tissue samples from 23 outbreaks and 22 cases. These results showed that PCLSR might be further used for preliminary and cost-effective detection and surveillance of ASFV. PMID:28198455

  19. Annulus Fibrosus Repair Using High-Density Collagen Gel: An In Vivo Ovine Model.

    PubMed

    Pennicooke, Brenton; Hussain, Ibrahim; Berlin, Connor; Sloan, Stephen R; Borde, Brandon; Moriguchi, Yu; Lang, Gernot; Navarro-Ramirez, Rodrigo; Cheetham, Jonathan; Bonassar, Lawrence J; Härtl, Roger

    2018-02-15

    Ovine in vivo study. To perform lateral approach lumbar surgery in an ovine model to administer an injectable riboflavin cross-linked high-density collagen (HDC) gel and to assess its ability to mitigate intervertebral disc (IVD) degeneration after induced annulus fibrosus (AF) injury. Biological-based injectable gels have shown efficacy in restoring biomechanical, radiographic, and histological parameters in IVD-injured animal models. Riboflavin cross-linked HDC gel has previously demonstrated retention of nucleus pulposus (NP) tissue, reduced loss of disc height, and prevention of terminal cellular degenerative changes in rat-tail spines. However, this biological therapy has never been tested in large animal models. Forty lumbar IVDs were accessed from eight sheep via lateral approach surgery. IVDs were randomly assigned to healthy control, injury and HDC treatment, or negative control with injury and no treatment. IVD injury was carried out using a drill-bit through the AF followed by needle puncture of the NP. Sheep were followed for 16 weeks and underwent qualitative/quantitative magnetic resonance imaging, x-ray, and histological analyses of collagen and proteoglycan content. The lateral approach to the ovine lumbar spine to deliver HDC gel proved to be safe and reproducible. IVDs treated with the HDC gel revealed less degenerative changes at the microscopic level based on AF and NP histology. However, mean Pfirrmann grade, T2 relaxation time, NP voxel size, and disc height index were not significantly different between the two injury groups. Injectable HDC gel can be administered safely via lateral approach surgery in an ovine AF injury model. IVDs treated with HDC gel demonstrated less degeneration at the microscopic level though radiographic changes were slight when comparing treated to untreated IVDs. Future studies will need to elucidate the role of injury technique and time frame for follow-up in correlating histological and radiographical outcomes. N

  20. Quantification of hyaluronan (HA) using a simplified fluorophore-assisted carbohydrate electrophoresis (FACE) procedure.

    PubMed

    Midura, Ronald J; Cali, Valbona; Lauer, Mark E; Calabro, Anthony; Hascall, Vincent C

    2018-01-01

    Hyaluronan (HA) exhibits numerous important roles in physiology and pathologies, and these facts necessitate an ability to accurately and reproducibly measure its quantities in tissues and cell cultures. Our group previously reported a rigorous and analytical procedure to quantify HA (and chondroitin sulfate, CS) using a reductive amination chemistry and separation of the fluorophore-conjugated, unsaturated disaccharides unique to HA and CS on high concentration acrylamide gels. This procedure is known as fluorophore-assisted carbohydrate electrophoresis (FACE) and has been adapted for the detection and quantification of all glycosaminoglycan types. While this previous FACE procedure is relatively straightforward to implement by carbohydrate research investigators, many nonglycoscience laboratories now studying HA biology might have difficulties establishing this prior FACE procedure as a routine assay for HA. To address this need, we have greatly simplified our prior FACE procedure for accurate and reproducible assessment of HA in tissues and cell cultures. This chapter describes in detail this simplified FACE procedure and, because it uses an enzyme that degrades both HA and CS, investigators will also gain additional insight into the quantities of CS in the same samples dedicated for HA analysis. © 2018 Elsevier Inc. All rights reserved.

  1. Cross-Linked Micellar Spherical Nucleic Acids from Thermoresponsive Templates

    PubMed Central

    2017-01-01

    A one-pot synthesis of micellar spherical nucleic acid (SNA) nanostructures using Pluronic F127 as a thermoresponsive template is reported. These novel constructs are synthesized in a chemically straightforward process that involves intercalation of the lipid tails of DNA amphiphiles (CpG motifs for TLR-9 stimulation) into the hydrophobic regions of Pluronic F127 micelles, followed by chemical cross-linking and subsequent removal of non-cross-linked structures. The dense nucleic acid shell of the resulting cross-linked micellar SNA enhances their stability in physiological media and facilitates their rapid cellular internalization, making them effective TLR-9 immunomodulatory agents. These constructs underscore the potential of SNAs in regulating immune response and address the relative lack of stability of noncovalent constructs. PMID:28207251

  2. Recyclable cross-linked anion exchange membrane for alkaline fuel cell application

    NASA Astrophysics Data System (ADS)

    Hou, Jianqiu; Liu, Yazhi; Ge, Qianqian; Yang, Zhengjin; Wu, Liang; Xu, Tongwen

    2018-01-01

    Cross-linking can effectively solve the conductivity-swelling dilemma in anion exchange membranes (AEMs) but will generate solid wastes. To address this, we developed an AEM cross-linked via disulfide bonds, bearing quaternary ammonium groups, which can be easily recycled. The membrane (RC-QPPO) with IEC of 1.78 mmol g-1, when cross-linked, showed enhanced mechanical properties and good hydroxide conductivity (24.6 mS cm-1 at 30 °C). Even at higher IEC value (2.13 mmol g-1), it still has low water uptake, low swelling ratio and delivers a peak power density of 150 mW cm-2 at 65 °C. Exploiting the formation of disulfide bonds from -SH groups, the membrane can be readily cross-linked in alkaline condition and recycled by reversibly breaking disulfide bonds with dithiothreitol (DTT). The recycled membrane solution can be directly utilized to cast a brand-new AEM. By washing away the residual DTT with water and exposure to air, it can be cross-linked again and this process is repeatable. During the recycling and cross-linking processes, the membrane showed a slight IEC decrease of 5% due to functional group degradation. The strategy presented here is promising in enhancing AEM properties and reducing the impact of artificial polymers on the environment.

  3. Freeze-dried eudragit-hyaluronan multicompartment liposomes to improve the intestinal bioavailability of curcumin.

    PubMed

    Catalan-Latorre, Ana; Ravaghi, Maryam; Manca, Maria Letizia; Caddeo, Carla; Marongiu, Francesca; Ennas, Guido; Escribano-Ferrer, Elvira; Peris, José Esteban; Diez-Sales, Octavio; Fadda, Anna Maria; Manconi, Maria

    2016-10-01

    This work aimed at finding an innovative vesicle-type formulation able to improve the bioavailability of curcumin upon oral administration. To this purpose, phospholipid, Eudragit® S100 and hyaluronan sodium salt were combined to obtain eudragit-hyaluronan immobilized vesicles using an easy and environmentally-friendly method. For the first time, the two polymers were combined in a system intended for oral delivery, to enhance curcumin stability when facing the harsh environment of the gastrointestinal tract. Four different formulations were prepared, keeping constant the amount of the phospholipid and varying the eudragit-hyaluronan ratio. The freeze-drying of the samples, performed to increase their stability, led to a reduction of vesicle size and a good homogeneity of the systems, after simple rehydration with water. X-ray diffraction study demonstrated that after the freeze-drying process, curcumin remained successfully incorporated within the vesicles. All the vesicles displayed similar features: size ranging from 220 to 287nm, spherical or oval shape, multilamellar or large unilamellar morphology with a peculiar multicompartment organization involving 1-4 smaller vesicles inside. In vitro studies demonstrated the ability of the combined polymers to protect the vesicles from the harsh conditions of the gastro-intestinal tract (i.e., ionic strength and pH variation), which was confirmed in vivo by the greater deposition of curcumin in the intestinal region, as compared to the free drug in dispersion. This enhanced accumulation of curcumin provided by the eudragit-hyaluronan immobilized vesicles, together with an increase in Caco-2 cell viability exposed to hydrogen peroxide, indicated that vesicles can ensure a local protection against oxidative stress and an increase in its intestinal absorption. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Yield and Failure Behavior Investigated for Cross-Linked Phenolic Resins Using Molecular Dynamics

    NASA Technical Reports Server (NTRS)

    Monk, Joshua D.; Lawson, John W.

    2016-01-01

    Molecular dynamics simulations were conducted to fundamentally evaluate the yield and failure behavior of cross-linked phenolic resins at temperatures below the glass transition. Yield stress was investigated at various temperatures, strain rates, and degrees of cross-linking. The onset of non-linear behavior in the cross-linked phenolic structures was caused by localized irreversible molecular rearrangements through the rotation of methylene linkers followed by the formation or annihilation of neighboring hydrogen bonds. The yield stress results, with respect to temperature and strain rate, could be fit by existing models used to describe yield behavior of amorphous glasses. The degree of cross-linking only indirectly influences the maximum yield stress through its influence on glass transition temperature (Tg), however there is a strong relationship between the degree of cross-linking and the failure mechanism. Low cross-linked samples were able to separate through void formation, whereas the highly cross-linked structures exhibited bond scission.

  5. Effect of molecular weight and concentration of hyaluronan on cell proliferation and osteogenic differentiation in vitro

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Zhao, Ningbo, E-mail: curl-zhao@163.com; Wang, Xin, E-mail: 394041230@qq.com; Qin, Lei, E-mail: qinlei30@126.com

    Hyaluronan (HA), the simplest glycosaminoglycan and a major component of the extracellular matrix, exists in various tissues. It is involved in some critical biological procedures, including cellular signaling, cell adhesion and proliferation, and cell differentiation. The effect of molecular weight (MW) and concentration of HA on cell proliferation and differentiation was controversial. In this study, we investigated the effect of MW and concentration of HA on the proliferation and osteogenic differentiation of rabbit bone marrow-derived stem cells in vitro. Results showed that high MW HA decreased the cell adhesion rate in a concentration-dependant manner. The cell adhesion rate was decreased bymore » increasing MW of HA. Cell proliferation was significantly enhanced by low MW HA (P < 0.05). The factorial analysis indicated that MW and concentration had an interactive effect on the cell adhesion rate and cell proliferation (P < 0.05). High MW HA increased the mRNA expressions of ALP, RUNX-2 and OCN. The higher the MW was, the higher the mRNA expressions were. The factorial analysis indicated that MW and concentration had an interactive effect on ALP mRNA expression (P < 0.05). HA of higher MW and higher concentration promoted bone formation. These findings provide some useful information in understanding the mechanism underlying the effect of MW and concentration of HA on cell proliferation and differentiation. - Highlights: • Effect of hyaluronan on cell proliferation and differentiation is evaluated in vitro. • Hyaluronan of low molecular weight increases cell proliferation. • Hyaluronan of high molecular weight promotes cell osteogenic differentiation. • Molecular weight and concentration of hyaluronan show interactive effect.« less

  6. Lysyl hydroxylase 2 induces a collagen cross-link switch in tumor stroma

    PubMed Central

    Chen, Yulong; Terajima, Masahiko; Yang, Yanan; Sun, Li; Ahn, Young-Ho; Pankova, Daniela; Puperi, Daniel S.; Watanabe, Takeshi; Kim, Min P.; Blackmon, Shanda H.; Rodriguez, Jaime; Liu, Hui; Behrens, Carmen; Wistuba, Ignacio I.; Minelli, Rosalba; Scott, Kenneth L.; Sanchez-Adams, Johannah; Guilak, Farshid; Pati, Debananda; Thilaganathan, Nishan; Burns, Alan R.; Creighton, Chad J.; Martinez, Elisabeth D.; Zal, Tomasz; Grande-Allen, K. Jane; Yamauchi, Mitsuo; Kurie, Jonathan M.

    2015-01-01

    Epithelial tumor metastasis is preceded by an accumulation of collagen cross-links that heighten stromal stiffness and stimulate the invasive properties of tumor cells. However, the biochemical nature of collagen cross-links in cancer is still unclear. Here, we postulated that epithelial tumorigenesis is accompanied by changes in the biochemical type of collagen cross-links. Utilizing resected human lung cancer tissues and a p21CIP1/WAF1-deficient, K-rasG12D-expressing murine metastatic lung cancer model, we showed that, relative to normal lung tissues, tumor stroma contains higher levels of hydroxylysine aldehyde–derived collagen cross-links (HLCCs) and lower levels of lysine aldehyde–derived cross-links (LCCs), which are the predominant types of collagen cross-links in skeletal tissues and soft tissues, respectively. Gain- and loss-of-function studies in tumor cells showed that lysyl hydroxylase 2 (LH2), which hydroxylates telopeptidyl lysine residues on collagen, shifted the tumor stroma toward a high-HLCC, low-LCC state, increased tumor stiffness, and enhanced tumor cell invasion and metastasis. Together, our data indicate that LH2 enhances the metastatic properties of tumor cells and functions as a regulatory switch that controls the relative abundance of biochemically distinct types of collagen cross-links in the tumor stroma. PMID:25664850

  7. Hyper alginate gel microbead formation by molecular diffusion at the hydrogel/droplet interface.

    PubMed

    Hirama, Hirotada; Kambe, Taisuke; Aketagawa, Kyouhei; Ota, Taku; Moriguchi, Hiroyuki; Torii, Toru

    2013-01-15

    We report a simple method for forming monodispersed, uniformly shaped gel microbeads with precisely controlled sizes. The basis of our method is the placement of monodispersed sodium alginate droplets, formed by a microfluidic device, on an agarose slab gel containing a high-osmotic-pressure gelation agent (CaCl(2) aq.): (1) the droplets are cross-linked (gelated) due to the diffusion of the gelation agent from the agarose slab gel to the sodium alginate droplets and (2) the droplets simultaneously shrink to a fraction of their original size (<100 μm in diameter) due to the diffusion of water molecules from the sodium alginate droplets to the agarose slab gel. We verified the mass transfer mechanism between the droplet and the agarose slab gel. This method circumvents the limitations of gel microbead formation, such as the need to prepare microchannels of various sizes, microchannel clogging, and the deformation of the produced gel microbeads.

  8. Macromolecular cross-linked enzyme aggregates (M-CLEAs) of α-amylase.

    PubMed

    Nadar, Shamraja S; Muley, Abhijeet B; Ladole, Mayur R; Joshi, Pranoti U

    2016-03-01

    Macromolecular cross-linked enzyme aggregates (M-CLEAs) of α-amylase were prepared by precipitation and subsequent cross-linking. The non-toxic, biodegradable, biocompatible, renewable polysaccharide based macromolecular cross-linkers viz. agar, chitosan, dextran, and gum arabic were used as a substitute for traditional glutaraldehyde to augment activity recovery toward macromolecular substrate. Macromolecular cross-linkers were prepared by periodate mediated controlled oxidation of polysaccharides. The effects of precipitating agent, concentration and different cross-linkers on activity recovery of α-amylase CLEAs were investigated. α-Amylase aggregated with ammonium sulphate and cross-linked by dextran showed 91% activity recovery, whereas glutaraldehyde CLEAs (G-CLEAs) exhibited 42% activity recovery. M-CLEAs exhibited higher thermal stability in correlation with α-amylase and G-CLEAs. Moreover, dextran and chitosan M-CLEAs showed same affinity for starch hydrolysis as of free α-amylase. The changes in secondary structures revealed the enhancements in structural and conformational rigidity attributed by cross-linkers. Finally, after five consecutive cycles dextran M-CLEAs retained 1.25 times higher initial activity than G-CLEAs. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. (ZrO 2) 0.85( REO 1.5) 0.15 ( RE=Sc, Y) solid solutions prepared via three Pechini-type gel routes: 1—gel formation and calcination behaviors

    NASA Astrophysics Data System (ADS)

    Zhang, Yawen; Li, Ang; Yan, Zhengguang; Xu, Gang; Liao, Chunsheng; Yan, Chunhua

    2003-02-01

    (ZrO2)0.85(REO1.5)0.15 (RE=Sc, Y) nanoparticles with pure cubic fluorite structure have been synthesized by three Pechini-type gel routes, viz. poly(vinyl alcohol) containing-process (route I), poly(ethylene glycol) and formic acid-containing process (route II), and in situ polymerizable complex method (route III). The coordination modes between metal ions and polymers in the gels are shown to be highly correlative with the synthesis route used. The gels prepared by route III have the strongest coordination throughout their network and therefore the best chemical homogeneity. The altered variety of polymer and cross-linking within the gels adopted by these three routes has made the as-synthesized samples show appreciable differences in thermal behavior, powder reactivity and nanoparticle properties.

  10. Photouncaged Sequence-specific Interstrand DNA Cross-Linking with Photolabile 4-oxo-enal-modified Oligonucleotides

    PubMed Central

    Sun, Jingjing; Tang, Xinjing

    2015-01-01

    DNA cross-linking technology is an attractive tool for the detection, regulation, and manipulation of genes. In this study, a series of photolabile 4-oxo-enal-modified oligonucleotides functionalized with photosensitive ο-nitrobenzyl derivatives were rationally designed as a new kind of photocaged cross-linking agents. A comprehensive evaluation of cross-linking reactions for different nucleobases in complementary strands under different conditions suggested that the modified DNA oligonucleotides tended to form interstrand cross-linking to nucleobases with the potential of thymidine > guanosine » cytidine ~ adenosine. Different from previous literature reports that cytidine and adenosine were preferential cross-linked nucleobases with 4-oxo-enal moieties, our study represents the first example of DNA cross-linking for T and G selectivity using 4-oxo-enal moiety. The cross-linked adducts were identified and their cross-linking mechanism was also illustrated. This greatly expands the applications of 4-oxo-enal derivatives in the studies of DNA damage and RNA structure PMID:26020694

  11. Photouncaged Sequence-specific Interstrand DNA Cross-Linking with Photolabile 4-oxo-enal-modified Oligonucleotides.

    PubMed

    Sun, Jingjing; Tang, Xinjing

    2015-05-28

    DNA cross-linking technology is an attractive tool for the detection, regulation, and manipulation of genes. In this study, a series of photolabile 4-oxo-enal-modified oligonucleotides functionalized with photosensitive ο-nitrobenzyl derivatives were rationally designed as a new kind of photocaged cross-linking agents. A comprehensive evaluation of cross-linking reactions for different nucleobases in complementary strands under different conditions suggested that the modified DNA oligonucleotides tended to form interstrand cross-linking to nucleobases with the potential of thymidine > guanosine » cytidine ~ adenosine. Different from previous literature reports that cytidine and adenosine were preferential cross-linked nucleobases with 4-oxo-enal moieties, our study represents the first example of DNA cross-linking for T and G selectivity using 4-oxo-enal moiety. The cross-linked adducts were identified and their cross-linking mechanism was also illustrated. This greatly expands the applications of 4-oxo-enal derivatives in the studies of DNA damage and RNA structure.

  12. In vitro cross-linking of elastin peptides and molecular characterization of the resultant biomaterials.

    PubMed

    Heinz, Andrea; Ruttkies, Christoph K H; Jahreis, Günther; Schräder, Christoph U; Wichapong, Kanin; Sippl, Wolfgang; Keeley, Fred W; Neubert, Reinhard H H; Schmelzer, Christian E H

    2013-04-01

    Elastin is a vital protein and the major component of elastic fibers which provides resilience to many vertebrate tissues. Elastin's structure and function are influenced by extensive cross-linking, however, the cross-linking pattern is still unknown. Small peptides containing reactive allysine residues based on sequences of cross-linking domains of human elastin were incubated in vitro to form cross-links characteristic of mature elastin. The resultant insoluble polymeric biomaterials were studied by scanning electron microscopy. Both, the supernatants of the samples and the insoluble polymers, after digestion with pancreatic elastase or trypsin, were furthermore comprehensively characterized on the molecular level using MALDI-TOF/TOF mass spectrometry. MS(2) data was used to develop the software PolyLinX, which is able to sequence not only linear and bifunctionally cross-linked peptides, but for the first time also tri- and tetrafunctionally cross-linked species. Thus, it was possible to identify intra- and intermolecular cross-links including allysine aldols, dehydrolysinonorleucines and dehydromerodesmosines. The formation of the tetrafunctional cross-link desmosine or isodesmosine was unexpected, however, could be confirmed by tandem mass spectrometry and molecular dynamics simulations. The study demonstrated that it is possible to produce biopolymers containing polyfunctional cross-links characteristic of mature elastin from small elastin peptides. MALDI-TOF/TOF mass spectrometry and the newly developed software PolyLinX proved suitable for sequencing of native cross-links in proteolytic digests of elastin-like biomaterials. The study provides important insight into the formation of native elastin cross-links and represents a considerable step towards the characterization of the complex cross-linking pattern of mature elastin. Copyright © 2013 Elsevier B.V. All rights reserved.

  13. Weak reversible cross links may decrease the strength of aligned fiber bundles.

    PubMed

    Nabavi, S Soran; Hartmann, Markus A

    2016-02-21

    Reversible cross-linking is an effective strategy to specifically tailor the mechanical properties of polymeric materials that can be found in a variety of biological as well as man-made materials. Using a simple model in this paper the influence of weak, reversible cross-links on the mechanical properties of aligned fiber bundles is investigated. Special emphasis in this analysis is put on the strength of the investigated structures. Using Monte Carlo methods two topologies of cross-links exceeding the strength of the covalent backbone are studied. Most surprisingly only two cross-links are sufficient to break the backbone of a multi chain system, resulting in a reduced strength of the material. The found effect crucially depends on the ratio of inter- to intra-chain cross-links and, thus, on the grafting density that determines this ratio.

  14. General protein-protein cross-linking.

    PubMed

    Alegria-Schaffer, Alice

    2014-01-01

    This protocol describes a general protein-to-protein cross-linking procedure using the water-soluble amine-reactive homobifunctional BS(3) (bis[sulfosuccinimidyl] suberate); however, the protocol can be easily adapted using other cross-linkers of similar properties. BS(3) is composed of two sulfo-NHS ester groups and an 11.4 Å linker. Sulfo-NHS ester groups react with primary amines in slightly alkaline conditions (pH 7.2-8.5) and yield stable amide bonds. The reaction releases N-hydroxysuccinimide (see an application of NHS esters on Labeling a protein with fluorophores using NHS ester derivitization). © 2014 Elsevier Inc. All rights reserved.

  15. Structural interactions between retroviral Gag proteins examined by cysteine cross-linking.

    PubMed Central

    Hansen, M S; Barklis, E

    1995-01-01

    We have examined structural interactions between Gag proteins within Moloney murine leukemia virus (M-MuLV) particles by making use of the cysteine-specific cross-linking agents iodine and bis-maleimido hexane. Virion-associated wild-type M-MuLV Pr65Gag proteins in immature particles were intermolecularly cross-linked at cysteines to form Pr65Gag oligomers, from dimers to pentamers or hexamers. Following a systematic approach of cysteine-to-serine mutagenesis, we have shown that cross-linking of Pr65Gag occurred at cysteines of the nucleocapsid (NC) Cys-His motif, suggesting that the Cys-His motifs within virus particles are packed in close proximity. The M-MuLV Pr65Gag protein did not cross-link to the human immunodeficiency virus Pr55Gag protein when the two molecules were coexpressed, indicating either that they did not coassemble or that heterologous Gag proteins were not in close enough proximity to be cross-linked. Using an assembly-competent, protease-minus, cysteine-minus Pr65Gag protein as a template, novel cysteine residues were generated in the M-MuLV capsid domain major homology region (MHR). Cross-linking of proteins containing MHR cysteines showed above-background levels of Gag-Gag dimers but also identified a novel cellular factor, present in virions, that cross-linked to MHR residues. Although the NC cysteine mutation was compatible with M-MuLV particle assembly, deletions of the NC domain were not tolerated. These results suggest that the Cys-His motif is held in close proximity within immature M-MuLV particles by interactions between CA domains and/or non-Cys-His motif domains of the NC. PMID:7815493

  16. Novel magnetic cross-linked lipase aggregates for improving the resolution of (R, S)-2-octanol.

    PubMed

    Liu, Ying; Guo, Chen; Liu, Chun-Zhao

    2015-03-01

    Novel magnetic cross-linked lipase aggregates were fabricated by immobilizing the cross-linked lipase aggregates onto magnetic particles with a high number of -NH2 terminal groups using p-benzoquinone as the cross-linking agent. At the optimal fabrication conditions, 100% of immobilization efficiency and 139% of activity recovery of the magnetic cross-linked lipase aggregates were achieved. The magnetic cross-linked lipase aggregates were able to efficiently resolve (R, S)-2-octanol, and retained 100% activity and 100% enantioselectivity after 10 cycles of reuse, whereas the cross-linked lipase aggregates only retained about 50% activity and 70% enantioselectivity due to insufficient cross-linking. These results provide a great potential for industrial applications of the magnetic cross-linked lipase aggregates. © 2014 Wiley Periodicals, Inc.

  17. Gels prepared from egg yolk and its fractions for tissue engineering.

    PubMed

    Rodil, Andrea; Laca, Amanda; Paredes, Benjamín; Rendueles, Manuel; Meana, Álvaro; Díaz, Mario

    2016-11-01

    New biomaterials prepared from egg yolk and its main fractions (plasma and granules) have been developed for use in tissue engineering. Protein gels obtained via transglutaminase cross-linking were characterized by rheometry, texturometry and scanning electron microscopy. All the gels exhibited suitable physical and mechanical characteristics for use as potential biomaterials in skin regeneration. Specifically, results showed that these materials presented a compact, uniform structure, with granular gel being found to be the most resistant as well as the most elastic material. Accordingly, these gels were subsequently evaluated as scaffolds for murine fibroblast growth. The best results were obtained with granule gels. Not only adhesion and cell growth were detected when using these gels, but also continuous coatings of cells growing on their surface. These findings can be attributed to the higher protein content of this fraction and to the particular structure of its proteins. Thus, granules have proved to be an interesting potential raw material for scaffold development. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1577-1583, 2016. © 2016 American Institute of Chemical Engineers.

  18. The High and Low Molecular Weight Forms of Hyaluronan Have Distinct Effects on CD44 Clustering*

    PubMed Central

    Yang, Cuixia; Cao, Manlin; Liu, Hua; He, Yiqing; Xu, Jing; Du, Yan; Liu, Yiwen; Wang, Wenjuan; Cui, Lian; Hu, Jiajie; Gao, Feng

    2012-01-01

    CD44 is a major cell surface receptor for the glycosaminoglycan hyaluronan (HA). Native high molecular weight hyaluronan (nHA) and oligosaccharides of hyaluronan (oHA) provoke distinct biological effects upon binding to CD44. Despite the importance of such interactions, however, the feature of binding with CD44 at the cell surface and the molecular basis for functional distinction between different sizes of HA is still unclear. In this study we investigated the effects of high and low molecular weight hyaluronan on CD44 clustering. For the first time, we provided direct evidence for a strong relationship between HA size and CD44 clustering in vivo. In CD44-transfected COS-7 cells, we showed that exogenous nHA stimulated CD44 clustering, which was disrupted by oHA. Moreover, naturally expressed CD44 was distributed into clusters due to abundantly expressed nHA in HK-2 cells (human renal proximal tubule cells) and BT549 cells (human breast cancer cell line) without exogenous stimulation. Our results suggest that native HA binding to CD44 selectively induces CD44 clustering, which could be inhibited by oHA. Finally, we demonstrated that HA regulates cell adhesion in a manner specifically dependent on its size. oHA promoted cell adhesion while nHA showed no effects. Our results might elucidate a molecular- and/or cellular-based mechanism for the diverse biological activities of nHA and oHA. PMID:23118219

  19. Ion exchange selectivity for cross-linked polyacrylic acid

    NASA Technical Reports Server (NTRS)

    May, C. E.; Philipp, W. H.

    1983-01-01

    The ion separation factors for 21 common metal ions with cross-linked polyacrylic acid were determined as a function of pH and the percent of the cross-linked polyacrylic acid neutralized. The calcium ion was used as a reference. At a pH of 5 the decreasing order of affinity of the ions for the cross-linked polyacrylic acid was found to be: Hg++, Fe+++, Pb++, Cr+++, Cu++, Cd++, Al+++, Ag+, Zn++, Ni++, Mn++, Co++, Ca++, Sr++, Ba++, Mg++, K+, Rb+, Cs+, Na+, and Li+. Members of a chemical family exhibited similar selectivities. The Hg++ ion appeared to be about a million times more strongly bound than the alkali metal ions. The relative binding of most of the metal ions varied with pH; the very tightly and very weakly bound ions showed the largest variations with pH. The calcium ion-hydrogen ion equilibrium was perturbed very little by the presence of the other ions. The separation factors and selectivity coefficients are discussed in terms of equilibrium and thermodynamic significance.

  20. Photoinduced DNA Interstrand Cross-Link Formation by Naphthalene Boronates via a Carbocation.

    PubMed

    Wang, Yibin; Lin, Zechao; Fan, Heli; Peng, Xiaohua

    2016-07-18

    Most photoinduced DNA cross-link formation by a bifunctional aryl derivative is through a bisquinone methide. DNA cross-linking via a bisarylcarbocation remains a less explored area. We designed and synthesized a series of naphthalene boronates that produce DNA interstrand cross-links via a carbocation upon UV irradiation. A free radical was generated from the naphthalene boronates with 350 nm irradiation and further converted to a carbocation by electron transfer. The activation mechanism was determined using the orthogonal traps, 2,2,6,6-tetramethylpiperidin-1-oxyl (TEMPO) and methoxyamine that react with either the free radical or the carbocation but not both. This represents a novel example of photoinduced DNA cross-link formation via carbocations generated from a bisaryl derivative. This work provides information useful for the design of novel photoactivated DNA cross-linking agents. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Study of the effect of mixing approach on cross-linking efficiency of hyaluronic acid-based hydrogel cross-linked with 1,4-butanediol diglycidyl ether.

    PubMed

    Al-Sibani, Mohammed; Al-Harrasi, Ahmed; Neubert, Reinhard H H

    2016-08-25

    Regardless of various strategies reported for cross-linking hyaluronic acid (HA) with 1,4-butanediol diglycidyl ether (BDDE), seeking new strategies that enhance cross-linking efficiency with a low level of cross-linker is essential. In this work, we studied the influence of mixing approach on two cross-linked BDDE-HA hydrogels prepared by two different mixing approaches; the large-batch mixing approach in which the hydrogel quantities were all mixed as a single lump in one container (hydrogel 1), and the small-batches mixing approach in which the hydrogel quantities were divided into smaller batches, mixed separately at various HA/BDDE ratios then combined in one reaction mixture (hydrogel 2). The result showed that the cross-linking reaction was mixing process-dependent. Degradation tests proved that, in relation to hydrogel 1, hydrogel 2 was more stable, and exhibited a higher resistance towards hyaluronidase activity. The swelling ratio of hydrogel 1 was significantly higher than that of hydrogel 2 in distilled water; however, in phosphate buffer saline, both hydrogels showed no significant difference. SEM images demonstrated that hydrogel 2 composite showed a denser network structure and smaller pore-size than hydrogel 1. In comparison to native HA, the occurrence of chemical modification in the cross-linked hydrogels was confirmed by FTIR and NMR distinctive peaks. These peaks also provided evidence that hydrogel 2 exhibited a higher degree of modification than hydrogel 1. In conclusion, the small-batches mixing approach proved to be more effective than large-batch mixing in promoting HA-HA entanglement and increasing the probability of BDDE molecules for binding with HA chains. Copyright © 2016 Elsevier B.V. All rights reserved.

  2. Induction of DNA-protein cross-links by platinum compounds.

    PubMed

    Woźniak, K; Walter, Z

    2000-01-01

    The differences between cis- and trans-diamminedichloroplatinum II (DDP) in forming DNA-protein cross-links in isolated human lymphocytes were investigated. Both cis- and trans-DDP can induce DNA-protein cross-links. We show that cis-DDP forms complexes between DNA and proteins faster than trans-DDP. This results from an increase in the quantity of DNA and platinum together with an increase in drug concentration. Under the same conditions trans-DDP causes a decrease in DNA-forming complexes with proteins. After a 12 h incubation of lymphocytes we observe a similar level of DNA in DNA-protein cross-links induced by DDP isomers, but more platinum appears in complexes induced by trans-DDP. The results obtained demonstrate that the antitumor drug - cis-DDP and the clinically ineffective trans-DDP induce links between DNA and proteins in a different manner. We suggest that the therapeutic activity of cis-DDP can in part arise from rapidly forming DNA-protein complexes which can destroy the most important cellular processes, such as replication and transcription.

  3. [Complications and postoperative therapeutic strategies in cross-linking].

    PubMed

    Kohlhaas, M

    2017-08-01

    The reduced corneal mechanical stability in keratoconus and similar collagen diseases can lead to a progressive and irregular corneal shape and decrease of visual acuity. A progression of keratectatic diseases can be shown with corneal topography. Keratoconus can be treated by photo-oxidative cross-linking of the corneal collagen. In order to achieve a high absorption of irradiation energy in the cornea, riboflavin at a concentration of 0.1% and UVA light at a wavelength of 370 nm corresponding to the relative maximum absorption of riboflavin (vitamin B2) are used. Evidence for corneal cross-linking are the increase of biomechanical stiffness, the increased resistance against enzymatic degradation, a higher shrinkage temperature, a lower swelling rate and an increased diameter of collagen fibers. The currently available data demonstrate that the therapeutic cross-linking procedure is safe when respecting the important theoretical and clinical parameters and that a progression of the keratoconus can be avoided. In 80% of cases an average levelling of the curvature of approximately 2 dpt can be achieved, which leads not only to stabilization but also to an increase in visual acuity of approximately 1.2 lines. In a Cochrane review from 2015 publications about complications and results were reviewed. Complication rates ranged from 1-10% depending on the initial situation, comorbidities and stage of the keratoconus. The most important complications are early epithelial wound healing problems as well as extremely rare perforations. Corneal cross-linking is a well-established and safe procedure but is not free of complications.

  4. Tissue-specific effects of aldose reductase inhibition on fluorescence and cross-linking of extracellular matrix in chronic galactosemia. Relationship to pentosidine cross-links.

    PubMed

    Richard, S; Tamas, C; Sell, D R; Monnier, V M

    1991-08-01

    Chronic experimental hyperglycemia mediated by galactose has been shown to induce browning and cross-linking of rat tail tendon collagen that could be duplicated in vitro by nonenzymatic galactosylation. To investigate the nature of these changes, Sprague-Dawley rats were placed on a 33% galactose diet without and with sorbinil for 6 and 12 mo. Collagen-linked fluorescence and pentosidine cross-links increased with age and galactosemia in tail tendons (P less than 0.001) and skin but were essentially unresponsive to aldose reductase inhibition (ARI). In contrast, tendon breaking time in urea, a likely parameter of cross-linking, was markedly improved (P less than 0.001) by ARI. Fluorescence that was inhibited by sorbinil treatment was increased in pepsin and proteinase K digest of aortic tissue from galactosemic rats (P less than 0.001), but impaired enzymatic digestibility was not observed. Systolic blood pressure as potential consequence of aortic stiffening was not increased in galactosemia. These data suggest that fluorescence in skin and tendon might be in part due to advanced glycosylation and pentosidine formation because these were not decreased by ARI. However, they also suggest that nonfluorescent cross-links may also be forming because, in contrast to fluorescence, tail tendon breaking time was partly corrected by ARI. Thus, it appears that extracellular matrix changes in chronic galactosemia are complex, being partly attributable to advanced glycosylation and partly to polyol-pathway activation.

  5. Application of a fast sorting algorithm to the assignment of mass spectrometric cross-linking data.

    PubMed

    Petrotchenko, Evgeniy V; Borchers, Christoph H

    2014-09-01

    Cross-linking combined with MS involves enzymatic digestion of cross-linked proteins and identifying cross-linked peptides. Assignment of cross-linked peptide masses requires a search of all possible binary combinations of peptides from the cross-linked proteins' sequences, which becomes impractical with increasing complexity of the protein system and/or if digestion enzyme specificity is relaxed. Here, we describe the application of a fast sorting algorithm to search large sequence databases for cross-linked peptide assignments based on mass. This same algorithm has been used previously for assigning disulfide-bridged peptides (Choi et al., ), but has not previously been applied to cross-linking studies. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. CD44S-hyaluronan interactions protect cells resulting from EMT against anoikis

    PubMed Central

    Cieply, Benjamin; Koontz, Colton; Frisch, Steven M.

    2016-01-01

    The detachment of normal epithelial cells from matrix triggers an apoptotic response known as anoikis, during homeostatic turnover. Metastatic tumor cells evade anoikis, by mechanisms that are only partly characterized. In particular, the epithelial–mesenchymal transition (EMT) in a subset of invasive tumor cells confers anoikis-resistance. In some cases, EMT up-regulates the cancer stem cell marker CD44S and the enzyme hyaluronic acid synthase-2 (HAS2). CD44S is the major receptor for hyaluronan in the extracellular matrix. Herein, we demonstrate that CD44S, unlike the CD44E isoform expressed in normal epithelial cells, contributes to the protection against anoikis. This protection requires the interaction of CD44S with hyaluronan (HA). CD44S–HA interaction is proposed to play an important role in tumor metastasis through enhanced cell survival under detached conditions. PMID:25937513

  7. Computational investigation of kinetics of cross-linking reactions in proteins: importance in structure prediction.

    PubMed

    Bandyopadhyay, Pradipta; Kuntz, Irwin D

    2009-01-01

    The determination of protein structure using distance constraints is a new and promising field of study. One implementation involves attaching residues of a protein using a cross-linking agent, followed by protease digestion, analysis of the resulting peptides by mass spectroscopy, and finally sequence threading to detect the protein folds. In the present work, we carry out computational modeling of the kinetics of cross-linking reactions in proteins using the master equation approach. The rate constants of the cross-linking reactions are estimated using the pKas and the solvent-accessible surface areas of the residues involved. This model is tested with fibroblast growth factor (FGF) and cytochrome C. It is consistent with the initial experimental rate data for individual lysine residues for cytochrome C. Our model captures all observed cross-links for FGF and almost 90% of the observed cross-links for cytochrome C, although it also predicts cross-links that were not observed experimentally (false positives). However, the analysis of the false positive results is complicated by the fact that experimental detection of cross-links can be difficult and may depend on specific experimental conditions such as pH, ionic strength. Receiver operator characteristic plots showed that our model does a good job in predicting the observed cross-links. Molecular dynamics simulations showed that for cytochrome C, in general, the two lysines come closer for the observed cross-links as compared to the false positive ones. For FGF, no such clear pattern exists. The kinetic model and MD simulation can be used to study proposed cross-linking protocols.

  8. Gamma-irradiated cross-linked LDPE foams: Characteristics and properties

    NASA Astrophysics Data System (ADS)

    Cardoso, E. C. L.; Scagliusi, S. R.; Parra, D. F.; Lugão, A. B.

    2013-03-01

    Foamed polymers are future materials, as they are increasingly considered "green materials" due to their interesting properties at very low consumption of raw materials. They can be used to improve appearance of insulation structures, thermal and acoustic insulation, core materials for sandwich panels, fabrication of furniture and flotation materials or to reduce costs involving materials. Low-density polyethylene is widely used because of its excellent properties, such as softness, elasticity, processibility and insulation. In general, cross-linking is often applied to improve the thermal and mechanical properties of polyethylene products, due to the formation of a three-dimensional network. In particular for the production of PE foams, cross-linking is applied prior the expansion to control bubble formation, cell characteristics and final properties of the foam. However, the usual production process of PE foams is a process in which a gaseous blowing agent is injected into a melted thermoplastic polymer, under pressure, to form a solution between blowing agent and melted polymer. An extrusion system is provided for foaming the polymer, supplied to an extruder and moving through a rotating screw. The pressure must be high enough to keep the gas blowing agent (or foaming agent) in the solution with the melt. The foaming agent is then diffused and dissolved in the molten material to form a single-phase solution. In the present work carbon dioxide was used as the bowing agent, a chemically stable and non-toxic gas, with good diffusion coefficient; gas pressure used varied within a 20-40 bar range. Some requirements for physical foaming are required, as low friction heat generation, homogeneous melt temperature distribution, melt temperature at die exit just above crystallization temperature (die) and high melt strength during expansion. This work studied foams properties gamma-irradiated within 0, 10, 15, 20, 25, and 30 kGy, from a LDPE exhibiting 2.6 g/10 min Melt

  9. Investigating the importance of flow when utilizing hyaluronan scaffolds for tissue engineering.

    PubMed

    Donegan, Gail C; Hunt, John A; Rhodes, Nicholas

    2010-02-01

    Esterified hyaluronan scaffolds offer significant advantages for tissue engineering. They are recognized by cellular receptors, interact with many other extracellular matrix proteins and their metabolism is mediated by intrinsic cellular pathways. In this study differences in the viability and structural integrity of vascular tissue models cultured on hyaluronan scaffolds under laminar flow conditions highlighted potential differences in the biodegradation kinetics, processes and end-products, depending on the culture environment. Critical factors are likely to include seeding densities and the duration and magnitude of applied biomechanical stress. Proteomic evaluation of the timing and amount of remodelling protein expression, the resulting biomechanical changes arising from this response and metabolic cell viability assay, together with examination of tissue morphology, were conducted in vascular tissue models cultured on esterified hyaluronan felt and PTFE mesh scaffolds. The vascular tissue models were derived using complete cell sheets derived from harvested and expanded umbilical cord vein cells. This seeding method utilizes high-density cell populations from the outset, while the cells are already supported by their own abundant extracellular matrix. Type I and type IV collagen expression in parallel with MMP-1 and MMP-2 expression were monitored in the tissue models over a 10 day culture period under laminar flow regimes using protein immobilization technologies. Uniaxial tensile testing and scanning electron microscopy were used to compare the resulting effects of hydrodynamic stimulation upon structural integrity, while viability assays were conducted to evaluate the effects of shear on metabolic function. The proteomic results showed that the hyaluronan felt-supported tissues expressed higher levels of all remodelling proteins than those cultured on PTFE mesh. Overall, a 21% greater expression of type I collagen, 24% higher levels of type IV collagen

  10. Cross-Linked Amylose Bio-Plastic: A Transgenic-Based Compostable Plastic Alternative

    PubMed Central

    Sagnelli, Domenico; Kemmer, Gerdi Christine; Holse, Mette; Hebelstrup, Kim H.; Bao, Jinsong; Stelte, Wolfgang; Bjerre, Anne-Belinda; Blennow, Andreas

    2017-01-01

    Bio-plastics and bio-materials are composed of natural or biomass derived polymers, offering solutions to solve immediate environmental issues. Polysaccharide-based bio-plastics represent important alternatives to conventional plastic because of their intrinsic biodegradable nature. Amylose-only (AO), an engineered barley starch with 99% amylose, was tested to produce cross-linked all-natural bioplastic using normal barley starch as a control. Glycerol was used as plasticizer and citrate cross-linking was used to improve the mechanical properties of cross-linked AO starch extrudates. Extrusion converted the control starch from A-type to Vh- and B-type crystals, showing a complete melting of the starch crystals in the raw starch granules. The cross-linked AO and control starch specimens displayed an additional wide-angle diffraction reflection. Phospholipids complexed with Vh-type single helices constituted an integrated part of the AO starch specimens. Gas permeability tests of selected starch-based prototypes demonstrated properties comparable to that of commercial Mater-Bi© plastic. The cross-linked AO prototypes had composting characteristics not different from the control, indicating that the modified starch behaves the same as normal starch. The data shows the feasibility of producing all-natural bioplastic using designer starch as raw material. PMID:28973963

  11. Cross-Linked Amylose Bio-Plastic: A Transgenic-Based Compostable Plastic Alternative.

    PubMed

    Sagnelli, Domenico; Hooshmand, Kourosh; Kemmer, Gerdi Christine; Kirkensgaard, Jacob J K; Mortensen, Kell; Giosafatto, Concetta Valeria L; Holse, Mette; Hebelstrup, Kim H; Bao, Jinsong; Stelte, Wolfgang; Bjerre, Anne-Belinda; Blennow, Andreas

    2017-09-30

    Bio-plastics and bio-materials are composed of natural or biomass derived polymers, offering solutions to solve immediate environmental issues. Polysaccharide-based bio-plastics represent important alternatives to conventional plastic because of their intrinsic biodegradable nature. Amylose-only (AO), an engineered barley starch with 99% amylose, was tested to produce cross-linked all-natural bioplastic using normal barley starch as a control. Glycerol was used as plasticizer and citrate cross-linking was used to improve the mechanical properties of cross-linked AO starch extrudates. Extrusion converted the control starch from A-type to Vh- and B-type crystals, showing a complete melting of the starch crystals in the raw starch granules. The cross-linked AO and control starch specimens displayed an additional wide-angle diffraction reflection. Phospholipids complexed with Vh-type single helices constituted an integrated part of the AO starch specimens. Gas permeability tests of selected starch-based prototypes demonstrated properties comparable to that of commercial Mater-Bi © plastic. The cross-linked AO prototypes had composting characteristics not different from the control, indicating that the modified starch behaves the same as normal starch. The data shows the feasibility of producing all-natural bioplastic using designer starch as raw material.

  12. Effect of Rubber Polarity on Cluster Formation in Rubbers Cross-Linked with Diels–Alder Chemistry

    PubMed Central

    2017-01-01

    Diels–Alder chemistry has been used for the thermoreversible cross-linking of furan-functionalized ethylene/propylene (EPM) and ethylene/vinyl acetate (EVM) rubbers. Both furan-functionalized elastomers were successfully cross-linked with bismaleimide to yield products with a similar cross-link density. NMR relaxometry and SAXS measurements both show that the apolar EPM-g-furan precursor contains phase-separated polar clusters and that cross-linking with polar bismaleimide occurs in these clusters. The heterogeneously cross-linked network of EPM-g-furan contrasts with the homogeneous network in the polar EVM-g-furan. The heterogeneous character of the cross-links in EPM-g-furan results in a relatively high Young’s modulus, whereas the more uniform cross-linking in EVM-g-furan results in a higher tensile strength and elongation at break. PMID:29213149

  13. Hyaluronan mediates airway hyperresponsiveness in oxidative lung injury

    PubMed Central

    Lazrak, Ahmed; Creighton, Judy; Yu, Zhihong; Komarova, Svetlana; Doran, Stephen F.; Aggarwal, Saurabh; Emala, Charles W.; Stober, Vandy P.; Trempus, Carol S.; Garantziotis, Stavros

    2015-01-01

    Chlorine (Cl2) inhalation induces severe oxidative lung injury and airway hyperresponsiveness (AHR) that lead to asthmalike symptoms. When inhaled, Cl2 reacts with epithelial lining fluid, forming by-products that damage hyaluronan, a constituent of the extracellular matrix, causing the release of low-molecular-weight fragments (L-HA, <300 kDa), which initiate a series of proinflammatory events. Cl2 (400 ppm, 30 min) exposure to mice caused an increase of L-HA and its binding partner, inter-α-trypsin-inhibitor (IαI), in the bronchoalveolar lavage fluid. Airway resistance following methacholine challenge was increased 24 h post-Cl2 exposure. Intratracheal administration of high-molecular-weight hyaluronan (H-HA) or an antibody against IαI post-Cl2 exposure decreased AHR. Exposure of human airway smooth muscle (HASM) cells to Cl2 (100 ppm, 10 min) or incubation with Cl2-exposed H-HA (which fragments it to L-HA) increased membrane potential depolarization, intracellular Ca2+, and RhoA activation. Inhibition of RhoA, chelation of intracellular Ca2+, blockade of cation channels, as well as postexposure addition of H-HA, reversed membrane depolarization in HASM cells. We propose a paradigm in which oxidative lung injury generates reactive species and L-HA that activates RhoA and Ca2+ channels of airway smooth muscle cells, increasing their contractility and thus causing AHR. PMID:25747964

  14. Identification of Pyridinoline Trivalent Collagen Cross-Links by Raman Microspectroscopy.

    PubMed

    Gamsjaeger, Sonja; Robins, Simon P; Tatakis, Dimitris N; Klaushofer, Klaus; Paschalis, Eleftherios P

    2017-06-01

    Intermolecular cross-linking of bone collagen is intimately related to the way collagen molecules are arranged in a fibril, imparts certain mechanical properties to the fibril, and may be involved in the initiation of mineralization. Raman microspectroscopy allows the analysis of minimally processed bone blocks and provides simultaneous information on both the mineral and organic matrix (mainly type I collagen) components, with a spatial resolution of ~1 μm. The aim of the present study was to validate Raman spectroscopic parameters describing one of the major mineralizing type I trivalent cross-links, namely pyridinoline (PYD). To achieve this, a series of collagen cross-linked peptides with known PYD content (as determined by HPLC analysis), human bone, porcine skin, predentin and dentin animal model tissues were analyzed by Raman microspectroscopy. The results of the present study confirm that it is feasible to monitor PYD trivalent collagen cross-links by Raman spectroscopic analysis in mineralized tissues, exclusively through a Raman band ~1660 wavenumbers. This allows determination of the relative PYD content in undecalcified bone tissues with a spatial resolution of ~1 μm, thus enabling correlations with histologic and histomorphometric parameters.

  15. Functional role of R462 in the degradation of hyaluronan catalyzed by hyaluronate lyase from Streptococcus pneumoniae.

    PubMed

    Li, Fengxue; Xu, Dingguo

    2015-08-01

    Hyaluronan lyase from Streptococcus pneumoniae can degrade hyaluronic acid, which is one of the major components in the extracellular matrix. Hyaluronan can regulate water balance, osmotic pressure, and act as an ion exchange resin. Followed by our recent work on the catalytic reaction mechanism and substrate binding mode, we in this work further investigate the functional role of active site arginine residue, R462, in the degradation of hyaluronan. The site directed mutagenesis simulation of R462A and R462Q were modeled using a combined quantum mechanical and molecular mechanical method. The overall substrate binding features upon mutations do not have significant changes. The energetic profiles for the reaction processes are essentially the same as that in wild type enzyme, but significant activation barrier height changes can be observed. Both mutants were shown to accelerate the overall enzymatic activity, e.g., R462A can reduce the barrier height by about 2.8 kcal mol(-1), while R462Q reduces the activation energy by about 2.9 kcal mol(-1). Consistent with the active site model calculated using density functional theory, our results can support that the positive charge on R462 guanidino side chain group plays a negative role in the catalysis. Finally, the functional role of R462 was proposed to facilitate the formation of initial enzyme-substrate complex, but not in the subsequent catalytic degradation reaction. Graphical Abstract Degradation of hyaluronan catalyzed by hyaluronate lyase from Streptococcus pneumoniae.

  16. No influence of exogenous hyaluronan on the behavior of human cancer cells or endothelial cell capillary formation.

    PubMed

    Seino, Satoshi; Takeshita, Fumitaka; Asari, Akira; Masuda, Yasunobu; Kunou, Masaaki; Ochiya, Takahiro

    2014-07-01

    Hyaluronan (HA), a type of glycosaminoglycan used to construct the extracellular matrix, is involved in the proliferation and motility of cells, including cancer cells. The aim of this study was to determine whether exogenous HA has an influence on cancer in vitro and in vivo. High-molecular-weight HA (900 kDa) and low-molecular-weight HA (10 kDa) were added to several types of cancer cell lines in vitro, and proliferation and invasion were assessed. The effect of HA on capillary formation by human umbilical vein endothelial cells was also analyzed. The results showed that both types of HA had no apparent effect on cellular proliferation, invasion, or capillary formation. In an animal study, the 2 types of HA were orally administered to tumor-bearing mice at a dosage of 200 mg/kg/d for 4 wk. Analysis using an in vivo imaging system revealed that tumor proliferation and metastasis were not greatly altered by HA administration. Furthermore, CD31 immunohistochemical staining revealed no obvious change in tumor microvessels. Taken together, these results demonstrate that exogenously administered HA has little effect on cancer. This study may support the safety of various forms of HA administration, including oral intake. Orally administered hyaluronan was recently found to have beneficial effects. However, the effect of exogenous hyaluronan on cancer remains unclear. Our findings support the safety of orally administered hyaluronan and its use as a functional food ingredient. © 2014 Institute of Food Technologists®

  17. Radiation cross-linking in ultra-high molecular weight polyethylene for orthopaedic applications

    NASA Astrophysics Data System (ADS)

    Oral, Ebru; Muratoglu, Orhun K.

    2007-12-01

    The motivation for radiation cross-linking of ultra-high molecular weight polyethylene (UHMWPE) is to increase its wear resistance to be used as bearing surfaces for total joint arthroplasty. However, radiation also leaves behind long-lived residual free radicals in this polymer, the reactions of which can detrimentally affect mechanical properties. In this review, we focus on the radiation cross-linking and oxidative stability of first and second generation highly cross-linked UHMWPEs developed in our laboratory.

  18. Motion in partially and fully cross-linked F-actin networks

    NASA Astrophysics Data System (ADS)

    Morris, Eliza; Ehrlicher, Allen; Weitz, David

    2012-02-01

    Single molecule experiments have measured stall forces and procession rates of molecular motors on isolated cytoskeletal fibers in Newtonian fluids. But in the cell, these motors are transporting cargo through a highly complex cytoskeletal network. To compare these single molecule results to the forces exerted by motors within the cell, an evaluation of the response of the cytoskeletal network is needed. Using magnetic tweezers and fluorescence confocal microscopy we observe and quantify the relationship between bead motion and filament response in F-actin networks both partially and fully cross-linked with filamin We find that when the transition from full to partial cross-linking is brought about by a decrease in cross-linker concentration there is a simultaneous decline in the elasticity of the network, but the response of the bead remains qualitatively similar. However, when the cross-linking is reduced through a shortening of the F-actin filaments the bead response is completely altered. The characteristics of the altered bead response will be discussed here.

  19. Interrelation of electret properties of polyethylene foam from the method of cross-linking

    NASA Astrophysics Data System (ADS)

    Gilmanov, I. R.; Galikhanov, M. F.; Gilmanova, A. R.

    2017-09-01

    The electret properties of chemically cross-linked polyethylene foam and physically cross-linked polyethylene foam have been studied. It has been shown that chemically cross-linked polyethylene foam has higher surface potential, effective surface charge density and electric field strength compared to physically bonded polyethylene foam. This is due to the presence of molecules and fragments of dicumyl peroxide, which can play the role of traps for injection charge carriers, a greater degree of cross-linking and with the oxidation of polyethylene, which occurs during irradiation during physical cross-linking. When the foam is deformed, its electret properties are reduced, and when the volume is relaxed, they are restored. This is due to the partial mutual compensation of homo- and heterocharge during compression and the return of the structure of the gas-filled polymer to its former position when the load is removed.

  20. Gel-forming reagents and uses thereof for preparing microarrays

    DOEpatents

    Golova, Julia; Chernov, Boris; Perov, Alexander

    2010-11-09

    New gel-forming reagents including monomers and cross-linkers, which can be applied to gel-drop microarray manufacturing by using co-polymerization approaches are disclosed. Compositions for the preparation of co-polymerization mixtures with new gel-forming monomers and cross-linker reagents are described herein. New co-polymerization compositions and cross-linkers with variable length linker groups between unsaturated C.dbd.C bonds that participate in the formation of gel networks are disclosed.

  1. Microchannel gel electrophoretic separation systems and methods for preparing and using

    DOEpatents

    Herr, Amy E; Singh, Anup K; Throckmorton, Daniel J

    2015-02-24

    A micro-analytical platform for performing electrophoresis-based immunoassays was developed by integrating photopolymerized cross-linked polyacrylamide gels within a microfluidic device. The microfluidic immunoassays are performed by gel electrophoretic separation and quantifying analyte concentration based upon conventional polyacrylamide gel electrophoresis (PAGE). To retain biological activity of proteins and maintain intact immune complexes, native PAGE conditions were employed. Both direct (non-competitive) and competitive immunoassay formats are demonstrated in microchips for detecting toxins and biomarkers (cytokines, c-reactive protein) in bodily fluids (serum, saliva, oral fluids). Further, a description of gradient gels fabrication is included, in an effort to describe methods we have developed for further optimization of on-chip PAGE immunoassays. The described chip-based PAGE immunoassay method enables immunoassays that are fast (minutes) and require very small amounts of sample (less than a few microliters). Use of microfabricated chips as a platform enables integration, parallel assays, automation and development of portable devices.

  2. Microchannel gel electrophoretic separation systems and methods for preparing and using

    DOEpatents

    Herr, Amy; Singh, Anup K; Throckmorton, Daniel J

    2013-09-03

    A micro-analytical platform for performing electrophoresis-based immunoassays was developed by integrating photopolymerized cross-linked polyacrylamide gels within a microfluidic device. The microfluidic immunoassays are performed by gel electrophoretic separation and quantifying analyte concentration based upon conventional polyacrylamide gel electrophoresis (PAGE). To retain biological activity of proteins and maintain intact immune complexes, native PAGE conditions were employed. Both direct (non-competitive) and competitive immunoassay formats are demonstrated in microchips for detecting toxins and biomarkers (cytokines, c-reactive protein) in bodily fluids (serum, saliva, oral fluids). Further, a description of gradient gels fabrication is included, in an effort to describe methods we have developed for further optimization of on-chip PAGE immunoassays. The described chip-based PAGE immunoassay method enables immunoassays that are fast (minutes) and require very small amounts of sample (less than a few microliters). Use of microfabricated chips as a platform enables integration, parallel assays, automation and development of portable devices.

  3. RNA oxidation catalyzed by cytochrome c leads to its depurination and cross-linking, which may facilitate cytochrome c release from mitochondria

    PubMed Central

    Tanaka, Mikiei; Jaruga, Pawel; Küpfer, Pascal A.; Leumann, Christian J.; Dizdaroglu, Miral; Sonntag, William E.; Chock, P. Boon

    2015-01-01

    Growing evidence indicates that RNA oxidation is correlated with a number of age-related neurodegen-erative diseases, and RNA oxidation has also been shown to induce dysfunction in protein synthesis. Here we study in vitro RNA oxidation catalyzed by cytochrome c (cyt c)/H2O2 or by the Fe(II)/ascorbate/H2O2 system. Our results reveal that the products of RNA oxidation vary with the oxidant used. Guanosine residues are preferentially oxidized by cyt c/H2O2 relative to the Fe(II)/ascorbate/H2O2 system. GC/MS and LC/MS analyses demonstrated that the guanine base was not only oxidized but also depurinated to form an abasic sugar moiety. Results from gel electrophoresis and HPLC analyses show that RNA formed a cross-linked complex with cyt c in an H2O2 concentration-dependent manner. Furthermore, when cyt c was associated with liposomes composed of cardiolipin/phosphatidylcholine, and incubated with RNA and H2O2, it was found cross-linked with the oxidized RNA and dissociated from the liposome. Results of the quantitative analysis indicate that the release of the cyt c from the liposome is facilitated by the formation of an RNA–cyt c cross-linked complex. Thus, RNA oxidation may facilitate the release of cyt c from the mitochondrial membrane to induce apoptosis in response to oxidative stress. PMID:22683603

  4. Cross-linking reveals laminin coiled-coil architecture

    PubMed Central

    Armony, Gad; Jacob, Etai; Moran, Toot; Levin, Yishai; Mehlman, Tevie; Levy, Yaakov; Fass, Deborah

    2016-01-01

    Laminin, an ∼800-kDa heterotrimeric protein, is a major functional component of the extracellular matrix, contributing to tissue development and maintenance. The unique architecture of laminin is not currently amenable to determination at high resolution, as its flexible and narrow segments complicate both crystallization and single-particle reconstruction by electron microscopy. Therefore, we used cross-linking and MS, evaluated using computational methods, to address key questions regarding laminin quaternary structure. This approach was particularly well suited to the ∼750-Å coiled coil that mediates trimer assembly, and our results support revision of the subunit order typically presented in laminin schematics. Furthermore, information on the subunit register in the coiled coil and cross-links to downstream domains provide insights into the self-assembly required for interaction with other extracellular matrix and cell surface proteins. PMID:27815530

  5. Modeling formalin fixation and histological processing with ribonuclease A: effects of ethanol dehydration on reversal of formaldehyde cross-links.

    PubMed

    Fowler, Carol B; O'Leary, Timothy J; Mason, Jeffrey T

    2008-07-01

    Understanding the chemistry of protein modification by formaldehyde fixation and subsequent tissue processing is central to developing improved methods for antigen retrieval in immunohistochemistry and for recovering proteins from formalin-fixed, paraffin-embedded (FFPE) tissues for proteomic analysis. Our initial studies of single proteins, such as bovine pancreatic ribonuclease A (RNase A), in 10% buffered formalin solution revealed that upon removal of excess formaldehyde, monomeric RNase A exhibiting normal immunoreactivity could be recovered by heating at 60 degrees C for 30 min at pH 4. We next studied tissue surrogates, which are gelatin-like plugs of fixed proteins that have sufficient physical integrity to be processed using normal tissue histology. Following histological processing, proteins could be extracted from the tissue surrogates by combining heat, detergent, and a protein denaturant. However, gel electrophoresis revealed that the surrogate extracts contained a mixture of monomeric and multimeric proteins. This suggested that during the subsequent steps of tissue processing protein-formaldehyde adducts undergo further modifications that are not observed in aqueous proteins. As a first step toward understanding these additional modifications we have performed a comparative evaluation of RNase A following fixation in buffered formaldehyde alone and after subsequent dehydration in 100% ethanol by combining gel electrophoresis, chemical modification, and circular dichroism spectroscopic studies. Our results reveal that ethanol-induced rearrangement of the conformation of fixed RNase A leads to protein aggregation through the formation of large geometrically compatible hydrophobic beta-sheets that are likely stabilized by formaldehyde cross-links, hydrogen bonds, and van der Waals interactions. It requires substantial energy to reverse the formaldehyde cross-links within these sheets and regenerate protein monomers free of formaldehyde modifications

  6. Assessment of UVA-Riboflavin Corneal Cross-Linking Using Small Amplitude Oscillatory Shear Measurements.

    PubMed

    Aslanides, Ioannis M; Dessi, Claudia; Georgoudis, Panagiotis; Charalambidis, Georgios; Vlassopoulos, Dimitris; Coutsolelos, Athanassios G; Kymionis, George; Mukherjee, Achyut; Kitsopoulos, Theofanis N

    2016-04-01

    The effect of ultraviolet (UV)-riboflavin cross-linking (CXL) has been measured primarily using the strip extensometry technique. We propose a simple and reliable methodology for the assessment of CXL treatment by using an established rheologic protocol based on small amplitude oscillatory shear (SAOS) measurements. It provides information on the average cross-link density and the elastic modulus of treated cornea samples. Three fresh postmortem porcine corneas were used to study the feasibility of the technique, one serving as control and two receiving corneal collagen cross-linking treatment. Subsequently, five pairs of fresh postmortem porcine corneas received corneal collagen cross-linking treatment with riboflavin and UVA-irradiation (370 nm; irradiance of 3 mW/cm2) for 30 minutes (Dresden protocol); the contralateral porcine corneas were used as control samples. After the treatment, the linear viscoelastic moduli of the corneal samples were measured using SAOS measurements and the average cross-linking densities extracted. For all cases investigated, the dynamic moduli of the cross-linked corneas were higher compared to those of the corresponding control samples. The increase of the elastic modulus of the treated samples was between 122% and 1750%. The difference was statistically significant for all tested samples (P = 0.018, 2-tailed t-test). We report a simple and accurate methodology for quantifying the effects of cross-linking on porcine corneas treated with the Dresden protocol by means of SAOS measurements in the linear regime. The measured dynamic moduli, elastic and viscous modulus, represent the energy storage and energy dissipation, respectively. Hence, they provide a means to assess the changing physical properties of the cross-linked collagen networks after CXL treatment.

  7. On-Demand Dissolution of Chemically Cross-Linked Hydrogels.

    PubMed

    Konieczynska, Marlena D; Grinstaff, Mark W

    2017-02-21

    The formation and subsequent on-demand dissolution of chemically cross-linked hydrogels is of keen interest to chemists, engineers, and clinicians. In this Account, we summarize our recent advances in the area of dissolvable chemically cross-linked hydrogels and provide a comparative discussion of other recent hydrogel systems. Using biocompatible macromonomers, we developed a library of cross-linked dendritic hydrogels that possess favorable properties, including biocompatibility, tissue adhesion, and swelling. Additionally, these hydrogels possess the unique ability to dissolve on-demand via application of a biocompatible aqueous solution. Each of the three hydrogel systems described employs a thiol-thioester exchange reaction as the mechanism of dissolution. These new materials successfully decrease bleeding in in vivo models of hepatic and aortic hemorrhage and dissolve on-demand, providing easy removal. In addition, we evaluated these hydrogels as dressings for second-degree burn wounds and performed proof-of-concept in vivo studies. These hydrogel wound dressings provide a means of repeatedly changing the dressing in a minimally invasive and atraumatic manner while also serving as a protective barrier against bacterial infection. Finally, we highlight the seminal work of other researchers in the field of dissolvable chemically cross-linked hydrogels using thiol-disulfide exchange, retro-Michael-type, and retro-Diels-Alder reactions. These chemistries provide a versatile synthetic toolbox to dissolve hydrogels in a controlled manner on time scales from minutes to weeks. Continued investigation of these dissolution approaches as well as the development of new chemical reactions will open doors to other avenues of on-demand dissolution and expand the application space for these materials. In summary, the management and closure of wounds after traumatic injury or surgical intervention are of significant clinical importance. Stimuli-responsive hydrogels that

  8. Dependence of nanomechanical modification of polymers on plasma-induced cross-linking

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Tajima, S.; Komvopoulos, K.

    2007-01-01

    The nanomechanical properties of low-density polyethylene (LDPE) modified by inductively coupled, radio-frequency Ar plasma were investigated by surface force microscopy. The polymer surface was modified under plasma conditions of different ion energy fluences and radiation intensities obtained by varying the sample distance from the plasma power source. Nanoindentation results of the surface stiffness versus maximum penetration depth did not reveal discernible differences between untreated and plasma-treated LDPE, presumably due to the small thickness of the modified surface layer that resulted in a substrate effect. On the contrary, nanoscratching experiments demonstrated a significant increase in the surface shear resistance of plasma-modifiedmore » LDPE due to chain cross-linking. These experiments revealed an enhancement of cross-linking with increasing ion energy fluence and radiation intensity, and a tip size effect on the friction force and dominant friction mechanisms (adhesion, plowing, and microcutting). In addition, LDPE samples with a LiF crystal shield were exposed to identical plasma conditions to determine the role of vacuum ultraviolet (VUV) and ultraviolet (UV) radiation in the cross-linking process. The cross-linked layer of plasma-treated LDPE exhibited much higher shear strength than that of VUV/UV-treated LDPE. Plasma-induced surface modification of the nanomechanical properties of LDPE is interpreted in the context of molecular models of the untreated and cross-linked polymer surfaces derived from experimental findings.« less

  9. Poly(ethylene imine)-based granular sorbents by a new process of templated gel-filling. High capacity and selectivity of copper sorption in acidic and alkaline media

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Chanda, M.; Rempel, G.L.

    A new process has been developed for making granular gel-type sorbents from chelating resins using metal ion as template. Named as templated gel-filling, the process uses the chosen metal as templating host ion on high-surface-area silica to build a templated gel layer from a solution of the chelating resin in a suitable solvent in which the resin is soluble but its metal complex is insoluble. After cross-linking the templated gel layer, the silica support is removed by alkali to produce a hollow shell of the templated gel. The shells are then soaked in a concentrated aqueous solution of the samemore » metal ion and suspended in the same resin solution to afford gel-filling. The shells thus filled with metal-templated gel are treated with cross-linking agent, followed by acid to remove the template ion and activate the resin for metal sorption. Poly(ethyleneimine) and its partially ethylated derivative have been used to produce granular gel-type sorbents by this process, with Cu(II) as the template ion. These sorbents are found to offer high capacity and selectivity for copper over nickel, cobalt, and zinc in both acidic and alkaline media. Containing a relatively high fraction of imbibed water, the sorbents exhibit markedly enhanced rate behavior, in both sorption and stripping.« less

  10. Fiber optic immunosensor for cross-linked fibrin concentration

    NASA Astrophysics Data System (ADS)

    Moskowitz, Samuel E.

    2000-08-01

    Working with calcium ions in the blood, platelets produce thromboplastin which transforms prothrombin into thrombin. Removing peptides, thrombin changes fibrinogen into fibrin. Cross-linked insoluble fibrin polymers are solubilized by enzyme plasmin found in blood plasma. Resulting D-dimers are elevated in patients with intravascular coagulation, deep venous thrombosis, pulmonary embolism, myocardial infarction, multiple trauma, cancer, impaired renal and liver functions, and sepsis. Consisting principally of a NIR 780 nm GaAlAs laser diode and a 800 nm avalanche photodiode (APD), the fiber-optic immunosensor can determined D-dimer concentration to levels <0.1 ng/ml. A capture monoclonal antibody to the antigen soluble cross-linked fibrin is employed. Immobilized at the tip of an optical fiber by avidin-biotin, the captured antigen is detected by a second antibody which is labeled with NN 382 fluorescent dye. An evanescent wave traveling on an excitation optical fiber excites the antibody-antigen fluorophore complex. Concentration of cross-linked fibrin is directly proportional to the APD measured intensity of fluorescence. NIR fluorescence has advantages of low background interference, short fluorescence lifetime, and large difference between excitation and emission peaks. Competitive ELISA test for D-dimer concentration requires trained personnel performing a time consuming operation.

  11. Role of special cross-links in structure formation of bacterial DNA polymer

    NASA Astrophysics Data System (ADS)

    Agarwal, Tejal; Manjunath, G. P.; Habib, Farhat; Lakshmi Vaddavalli, Pavana; Chatterji, Apratim

    2018-01-01

    Using data from contact maps of the DNA-polymer of Escherichia coli (E. Coli) (at kilobase pair resolution) as an input to our model, we introduce cross-links between monomers in a bead-spring model of a ring polymer at very specific points along the chain. Via suitable Monte Carlo simulations, we show that the presence of these cross-links leads to a particular organization of the chain at large (micron) length scales of the DNA. We also investigate the structure of a ring polymer with an equal number of cross-links at random positions along the chain. We find that though the polymer does get organized at the large length scales, the nature of the organization is quite different from the organization observed with cross-links at specific biologically determined positions. We used the contact map of E. Coli bacteria which has around 4.6 million base pairs in a single circular chromosome. In our coarse-grained flexible ring polymer model, we used 4642 monomer beads and observed that around 80 cross-links are enough to induce the large-scale organization of the molecule accounting for statistical fluctuations caused by thermal energy. The length of a DNA chain even of a simple bacterial cell such as E. Coli is much longer than typical proteins, hence we avoided methods used to tackle protein folding problems. We define new suitable quantities to identify the large scale structure of a polymer chain with a few cross-links.

  12. Hyaluronan and hyaluronectin in the extracellular matrix of human brain tumour stroma.

    PubMed

    Delpech, B; Maingonnat, C; Girard, N; Chauzy, C; Maunoury, R; Olivier, A; Tayot, J; Creissard, P

    1993-01-01

    Hyaluronan (HA) and the hyaluronan-binding glycoprotein hyaluronectin (HN) were measured in 23 gliomas and 8 meningiomas and their location was revisited in 35 tumours. A clear-cut difference was found in the HN/HA ratio values of glioblastomas (below 0.5) and that of astrocytomas (above 0.5 P < 0.001). Besides their location in the intercellular part of gliomas, HA and HN displayed a perivascular location in 1/3 astrocytomas, 17/24 glioblastomas, and 3/7 meningiomas, suggesting they could be produced also by the vascular stroma of tumours and that they would characterise the neoangiogenesis. All cultivated glioma cells tested produced HA in vitro, whereas only 1/11 cell lines produced HN, at a low level. The results obtained suggest that glioma HA and HN are produced by both cancer cells and vascular stroma cells, which contribute to the edification of the extracellular matrix. In meningiomas only the stroma would be responsible for HA and HN production.

  13. Competition between dewetting and cross-linking in poly(N-vinylpyrrolidone)/polystyrene bilayer films.

    PubMed

    Telford, Andrew M; Thickett, Stuart C; James, Michael; Neto, Chiara

    2011-12-06

    We investigated the dewetting of metastable poly(N-vinylpyrrolidone) (PNVP) thin films (45 nm) on top of polystyrene (PS) thin films (58 nm) as a function of annealing temperature and molecular weight of PS (96 and 6850 kg/mol). We focused on the competition between dewetting, occurring as a result of unfavorable intermolecular interactions at the PNVP/PS interface, and spontaneous cross-linking of PNVP, occurring during thermal annealing, as we recently reported (Telford, A. M.; James, M.; Meagher, L.; Neto, C. ACS Appl. Mater. Interfaces 2010, 2, 2399-2408). Using optical microscopy, we studied how the dewetting morphology and dynamics at different temperatures depended on the relative viscosity of the top PNVP film, which increased with cross-linking time, and of the bottom PS film. In the PNVP/PS96K system, cross-linking dominated over dewetting at temperatures below 180 °C, reducing drastically nucleated hole density and their maximum size, while above 180 °C the two processes reversed, with complete dewetting occurring at 200 °C. On the other hand, the PNVP/PS6850K system never achieved advanced dewetting stages as the dewetting was slower than cross-linking in the investigated temperature range. In both systems, dewetting of the PNVP films could be avoided altogether by thermally annealing the bilayers at temperatures where cross-linking dominated. The cross-linking was characterized quantitatively using neutron reflectometry, which indicated shrinkage and densification of the PNVP film, and qualitatively through selective removal of the bottom PS film. A simple model accounting for progressive cross-linking during the dewetting process predicted well the observed hole growth profiles and produced estimates of the PNVP cross-linking rate coefficients and of the activation energy of the process, in good agreement with literature values for similar systems. © 2011 American Chemical Society

  14. Microstructure and rheology of thermoreversible nanoparticle gels.

    PubMed

    Ramakrishnan, S; Zukoski, C F

    2006-08-29

    Naïve mode coupling theory is applied to particles interacting with short-range Yukawa attractions. Model results for the location of the gel line and the modulus of the resulting gels are reduced to algebraic equations capturing the effects of the range and strength of attraction. This model is then applied to thermo reversible gels composed of octadecyl silica particles suspended in decalin. The application of the model to the experimental system requires linking the experimental variable controlling strength of attraction, temperature, to the model strength of attraction. With this link, the model predicts temperature and volume fraction dependencies of gelation and modulus with five parameters: particle size, particle volume fraction, overlap volume of surface hairs, and theta temperature. In comparing model predictions with experimental results, we first observe that in these thermal gels there is no evidence of clustering as has been reported in depletion gels. One consequence of this observation is that there are no additional adjustable parameters required to make quantitative comparisons between experimental results and model predictions. Our results indicate that the naïve mode coupling approach taken here in conjunction with a model linking temperature to strength of attraction provides a robust approach for making quantitative predictions of gel mechanical properties. Extension of model predictions to additional experimental systems requires linking experimental variables to the Yukawa strength and range of attraction.

  15. Hyaluronan as a propellant for epithelial movement: the development of semicircular canals in the inner ear of Xenopus.

    PubMed

    Haddon, C M; Lewis, J H

    1991-06-01

    The membranous labyrinth of the inner ear, with its three semicircular canals, originates from a simple spheroidal otic vesicle. The process is easily observed in Xenopus. The vesicle develops three dorsal outpocketings; from the two opposite faces of each outpocketing pillars of tissue are protruded into the lumen; and these paired 'axial protrusions' eventually meet and fuse, to form a column of tissue spanning the lumen of the outpocketing like the hub of a wheel, with a tube of epithelium forming the semicircular canal around the periphery. Each axial protrusion consists of epithelium encasing a core of largely cell-free extracellular matrix that stains strongly with alcian blue. In sections, at least 60% of the stainable material is removed by treatment with Streptomyces hyaluronidase. When Streptomyces hyaluronidase is microinjected into the core of a protrusion in vivo, the protrusion collapses and the corresponding semicircular canal fails to form. Hyaluronan (hyaluronic acid) in the core of the protrusion therefore seems to be essential in driving the extension of the protrusion. Autoradiography with tritiated glucosamine indicates that the hyaluronan-rich matrix is synthesised by the epithelium covering the tip of the protrusion; the basal lamina here appears to be discontinuous. These findings indicate that the epithelium of the axial protrusion propels itself into the lumen of the otocyst by localised synthesis of hyaluronan. Hyaluronan may be used in a similar way in the development of other organs, such as the heart and the secondary palate.

  16. Durability and mechanical properties of silane cross-linked wood thermoplastic composites

    Treesearch

    Magnus Bengtsson; Nicole M. Stark; Kristiina Oksman

    2007-01-01

    In this study, silane cross-linked wood–polyethylene composite profiles were manufactured by reactive extrusion. These composites were evaluated regarding their durability and mechanical properties in comparison with two non-cross-linked wood– polyethylene composites. An addition of only 2% w/w of silane solution during manufacturing was enough to achieve almost 60%...

  17. Cross-Linked Nanohybrid Polymer Electrolytes With POSS Cross-Linker for Solid-State Lithium Ion Batteries.

    PubMed

    Zhang, Jinfang; Li, Xiaofeng; Li, Ying; Wang, Huiqi; Ma, Cheng; Wang, Yanzhong; Hu, Shengliang; Wei, Weifeng

    2018-01-01

    A new class of freestanding cross-linked hybrid polymer electrolytes (HPEs) with POSS as the cross-linker was prepared by a one-step free radical polymerization reaction. Octavinyl octasilsesquioxane (OV-POSS) with eight functional corner groups was used to provide cross-linking sites for the connection of polymer segments and the required mechanical strength to separate the cathode and anode. The unique cross-linked structure offers additional free volume for the motion of EO chains and provides fast and continuously interconnected ion-conducting channels along the nanoparticles/polymer matrix interface. The HPE exhibits the highest ionic conductivity of 1.39 × 10 -3 S cm -1 , as well as excellent interfacial compatibility with the Li electrode at 80°C. In particular, LiFePO 4 /Li cells based on the HPE deliver good rate capability and long-term cycling performance with an initial discharge capacity of 152.1 mAh g -1 and a capacity retention ratio of 88% after 150 cycles with a current density of 0.5 C at 80°C, demonstrating great potential application in high-performance LIBs at elevated temperatures.

  18. Cross-linked Nanohybrid Polymer Electrolytes with POSS Cross-linker for Solid-state Lithium Ion Batteries

    NASA Astrophysics Data System (ADS)

    Zhang, Jinfang; Li, Xiaofeng; Li, Ying; Wang, Huiqi; Ma, Cheng; Wang, Yanzhong; Hu, Shengliang; Wei, Weifeng

    2018-05-01

    A new class of freestanding cross-linked hybrid polymer electrolytes (HPEs) with POSS as the cross-linker was prepared by a one-step free radical polymerization reaction. Octavinyl octasilsesquioxane (OV-POSS) with eight functional corner groups was used to provide cross-linking sites for the connection of polymer segments and the required mechanical strength to separate the cathode and anode. The unique cross-linked structure offers additional free volume for the motion of EO chains and provides fast and continuously interconnected ion-conducting channels along the nanoparticles/polymer matrix interface. The HPE exhibits the highest ionic conductivity of 1.39×10-3 S cm-1, as well as excellent interfacial compatibility with the Li electrode at 80 oC. In particular, LiFePO4/Li cells based on the HPE deliver good rate capability and long-term cycling performance with an initial discharge capacity of 152.1 mAh g-1 and a capacity retention ratio of 88% after 150 cycles with a current density of 0.5 C at 80 oC, demonstrating great potential application in high-performance LIBs at elevated temperatures.

  19. The Preparation and Properties of Thermo-reversibly Cross-linked Rubber Via Diels-Alder Chemistry.

    PubMed

    Polgar, Lorenzo Massimo; van Duin, Martin; Picchioni, Francesco

    2016-08-25

    A method for using Diels Alder thermo-reversible chemistry as cross-linking tool for rubber products is demonstrated. In this work, a commercial ethylene-propylene rubber, grafted with maleic anhydride, is thermo-reversibly cross-linked in two steps. The pending anhydride moieties are first modified with furfurylamine to graft furan groups to the rubber backbone. These pendant furan groups are then cross-linked with a bis-maleimide via a Diels-Alder coupling reaction. Both reactions can be performed under a broad range of experimental conditions and can easily be applied on a large scale. The material properties of the resulting Diels-Alder cross-linked rubbers are similar to a peroxide-cured ethylene/propylene/diene rubber (EPDM) reference. The cross-links break at elevated temperatures (> 150 °C) via the retro-Diels-Alder reaction and can be reformed by thermal annealing at lower temperatures (50-70 °C). Reversibility of the system was proven with infrared spectroscopy, solubility tests and mechanical properties. Recyclability of the material was also shown in a practical way, i.e., by cutting a cross-linked sample into small parts and compression molding them into new samples displaying comparable mechanical properties, which is not possible for conventionally cross-linked rubbers.

  20. Preparation of Nanocellulose Reinforced Chitosan Films, Cross-Linked by Adipic Acid

    PubMed Central

    Falamarzpour, Pouria; Behzad, Tayebeh; Zamani, Akram

    2017-01-01

    Adipic acid, an abundant and nontoxic compound, was used to dissolve and cross-link chitosan. After the preparation of chitosan films through casting technique, the in situ amidation reaction was performed at 80–100 °C as verified by Fourier transform infrared (FT-IR). The reaction was accompanied by the release of water which was employed to investigate the reaction kinetics. Accordingly, the reaction rate followed the first-order model and Arrhenius equation, and the activation energy was calculated to be 18 kJ/mol. Furthermore, the mechanical properties of the chitosan films were comprehensively studied. First, optimal curing conditions (84 °C, 93 min) were introduced through a central composite design. In order to evaluate the effects of adipic acid, the mechanical properties of physically cross-linked (uncured), chemically cross-linked (cured), and uncross-linked (prepared by acetic acid) films were compared. The use of adipic acid improved the tensile strength of uncured and chemically cross-linked films more than 60% and 113%, respectively. Finally, the effect of cellulose nanofibrils (CNFs) on the mechanical performance of cured films, in the presence of glycerol as a plasticizer, was investigated. The plasticized chitosan films reinforced by 5 wt % CNFs showed superior properties as a promising material for the development of chitosan-based biomaterials. PMID:28208822

  1. Preparation of Nanocellulose Reinforced Chitosan Films, Cross-Linked by Adipic Acid.

    PubMed

    Falamarzpour, Pouria; Behzad, Tayebeh; Zamani, Akram

    2017-02-13

    Adipic acid, an abundant and nontoxic compound, was used to dissolve and cross-link chitosan. After the preparation of chitosan films through casting technique, the in situ amidation reaction was performed at 80-100 °C as verified by Fourier transform infrared (FT-IR). The reaction was accompanied by the release of water which was employed to investigate the reaction kinetics. Accordingly, the reaction rate followed the first-order model and Arrhenius equation, and the activation energy was calculated to be 18 kJ/mol. Furthermore, the mechanical properties of the chitosan films were comprehensively studied. First, optimal curing conditions (84 °C, 93 min) were introduced through a central composite design. In order to evaluate the effects of adipic acid, the mechanical properties of physically cross-linked (uncured), chemically cross-linked (cured), and uncross-linked (prepared by acetic acid) films were compared. The use of adipic acid improved the tensile strength of uncured and chemically cross-linked films more than 60% and 113%, respectively. Finally, the effect of cellulose nanofibrils (CNFs) on the mechanical performance of cured films, in the presence of glycerol as a plasticizer, was investigated. The plasticized chitosan films reinforced by 5 wt % CNFs showed superior properties as a promising material for the development of chitosan-based biomaterials.

  2. Fluorescent molecularly imprinted polymers as plastic antibodies for selective labeling and imaging of hyaluronan and sialic acid on fixed and living cells.

    PubMed

    Panagiotopoulou, Maria; Kunath, Stephanie; Medina-Rangel, Paulina Ximena; Haupt, Karsten; Tse Sum Bui, Bernadette

    2017-02-15

    Altered glycosylation levels or distribution of sialic acids (SA) or hyaluronan in animal cells are indicators of pathological conditions like infection or malignancy. We applied fluorescently-labeled molecularly imprinted polymer (MIP) particles for bioimaging of fixed and living human keratinocytes, to localize hyaluronan and sialylation sites. MIPs were prepared with the templates D-glucuronic acid (GlcA), a substructure of hyaluronan, and N-acetylneuraminic acid (NANA), the most common member of SA. Both MIPs were found to be highly selective towards their target monosaccharides, as no cross-reactivity was observed with other sugars like N-acetyl-D-glucosamine, N-acetyl-D-galactosamine, D-glucose and D-galactose, present on the cell surface. The dye rhodamine and two InP/ZnS quantum dots (QDs) emitting in the green and in the red regions were used as fluorescent probes. Rhodamine-MIPGlcA and rhodamine-MIPNANA were synthesized as monodispersed 400nm sized particles and were found to bind selectively their targets located in the extracellular region, as imaged by epifluorescence and confocal microscopy. In contrast, when MIP-GlcA and MIP-NANA particles with a smaller size (125nm) were used, the MIPs being synthesized as thin shells around green and red emitting QDs respectively, it was possible to stain the intracellular and pericellular regions as well. In addition, simultaneous dual-color imaging with the two different colored QDs-MIPs was demonstrated. Importantly, the MIPs were not cytotoxic and did not affect cell viability; neither was the cells morphology affected as demonstrated by live cell imaging. These synthetic receptors could offer a new and promising imaging tool to monitor disease progression. Copyright © 2016 Elsevier B.V. All rights reserved.

  3. Photochromic cross-link polymer for color changing and sensing surface

    NASA Astrophysics Data System (ADS)

    Fu, Richard; Shi, Jianmin; Forsythe, Eric; Srour, Merric

    2016-12-01

    Photochromic cross-link polymers were developed using patented ultraviolet (UV) photoinitiator and commercial photochromic dyes. The photochromic dyes have been characterized by measuring absorbance before and after UV activation using UV-visible (Vis) spectrometry with varying activation intensities and wavelengths. Photochromic cross-link polymers were characterized by a dynamic xenon and UV light activation and fading system. The curing processes on cloth were established and tested to obtain effective photochromic responses. Both PulseForge photonic curing and PulseForge plus heat surface curing processes had much better photochromic responses (18% to 19%, 16% to 25%, respectively) than the xenon lamp treatment (8%). The newly developed photochromic cross-link polymer showed remarkable coloration contrasts and fast and comparable coloration and fading rates. Those intelligent, controlled color changing and sensing capabilities will be used on flexible and "drapeable" surfaces, which will incorporate ultra-low power sensors, sensor indicators, and identifiers.

  4. In vitro progesterone release from γ-irradiated cross-linked polydimethylsiloxane

    NASA Astrophysics Data System (ADS)

    Mashak, Arezou; Taghizadeh, S. Mojtaba

    2006-02-01

    Instead of conventional method such as thermal cross-linking method, γ-irradiation is used to improve the properties of polydimethylsiloxane (PDMS) as a matrix containing progesterone. The thermal cross-linking of PDMS monolithic systems containing drug is deleterious to the drug. Usually, all drugs are unstable both at high vulcanizing temperature and in the presence of peroxide catalysts. This novel method is found to be effective for the stability of the controlled drug delivery systems. The PDMS (three medical grades) matrices were exposed to γ-irradiation in ambient conditions with total doses of 50, 75 and 100 kGy. The mechanical properties confirmed that the samples are cross-linked. It is found that the progesterone release rate is affected by irradiation treatment. It is deduced, however that there is no significant difference in the release profile of progesterone by increasing the irradiation dose from 50 to 100 kGy.

  5. A theory for fracture of polymeric gels

    NASA Astrophysics Data System (ADS)

    Mao, Yunwei; Anand, Lallit

    2018-06-01

    A polymeric gel is a cross-linked polymer network swollen with a solvent. If the concentration of the solvent or the deformation is increased to substantial levels, especially in the presence of flaws, then the gel may rupture. Although various theoretical aspects of coupling of fluid permeation with large deformation of polymeric gels are reasonably well-understood and modeled in the literature, the understanding and modeling of the effects of fluid diffusion on the damage and fracture of polymeric gels is still in its infancy. In this paper we formulate a thermodynamically-consistent theory for fracture of polymeric gels - a theory which accounts for the coupled effects of fluid diffusion, large deformations, damage, and also the gradient effects of damage. The particular constitutive equations for fracture of a gel proposed in our paper, contain two essential new ingredients: (i) Our constitutive equation for the change in free energy of a polymer network accounts for not only changes in the entropy, but also changes in the internal energy due the stretching of the Kuhn segments of the polymer chains in the network. (ii) The damage and failure of the polymer network is taken to occur by chain-scission, a process which is driven by the changes in the internal energy of the stretched polymer chains in the network, and not directly by changes in the configurational entropy of the polymer chains. The theory developed in this paper is numerically implemented in an open-source finite element code MOOSE, by writing our own application. Using this simulation capability we report on our study of the fracture of a polymeric gel, and some interesting phenomena which show the importance of the diffusion of the fluid on fracture response of the gel are highlighted.

  6. Optimization of Formaldehyde Cross-Linking for Protein Interaction Analysis of Non-Tagged Integrin β1

    PubMed Central

    Klockenbusch, Cordula; Kast, Juergen

    2010-01-01

    Formaldehyde cross-linking of protein complexes combined with immunoprecipitation and mass spectrometry analysis is a promising technique for analysing protein-protein interactions, including those of transient nature. Here we used integrin β1 as a model to describe the application of formaldehyde cross-linking in detail, particularly focusing on the optimal parameters for cross-linking, the detection of formaldehyde cross-linked complexes, the utility of antibodies, and the identification of binding partners. Integrin β1 was found in a high molecular weight complex after formaldehyde cross-linking. Eight different anti-integrin β1 antibodies were used for pull-down experiments and no loss in precipitation efficiency after cross-linking was observed. However, two of the antibodies could not precipitate the complex, probably due to hidden epitopes. Formaldehyde cross-linked complexes, precipitated from Jurkat cells or human platelets and analyzed by mass spectrometry, were found to be composed of integrin β1, α4 and α6 or β1, α6, α2, and α5, respectively. PMID:20634879

  7. Fabrication of patterned calcium cross-linked alginate hydrogel films and coatings through reductive cation exchange.

    PubMed

    Bruchet, Marion; Melman, Artem

    2015-10-20

    Calcium cross-linked alginate hydrogels are widely used in targeted drug delivery, tissue engineering, wound treatment, and other biomedical applications. We developed a method for preparing homogeneous alginate hydrogels cross-linked with Ca(2+) cations using reductive cation exchange in homogeneous iron(III) cross-linked alginate hydrogels. Treatment of iron(III) cross-linked alginate hydrogels with calcium salts and sodium ascorbate results in reduction of iron(III) cations to iron(II) that are instantaneously replaced with Ca(2+) cations, producing homogeneous ionically cross-linking hydrogels. Alternatively, the cation exchange can be performed by photochemical reduction in the presence of calcium chloride using a sacrificial photoreductant. This approach allows fabrication of patterned calcium alginate hydrogels through photochemical patterning of iron(III) cross-linked alginate hydrogel followed by the photochemical reductive exchange of iron cations to calcium. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. Hydroxyl radical and ferryl-generating systems promote gel network formation of myofibrillar protein.

    PubMed

    Xiong, Youling L; Blanchard, Suzanne P; Ooizumi, Tooru; Ma, Yuanyuan

    2010-03-01

    The objective of the study was to examine how oxidatively induced protein cross-linking would influence the gelation properties of myofibrillar protein (MP) under meat processing conditions. MP suspensions in 0.6 M NaCl at pH 6 were treated with an iron-catalyzed oxidizing system (IOS: 10 microM FeCl(3), 0.1 mM ascorbic acid, 0.05 to 5 mM H(2)O(2)) or a H(2)O(2)-activated metmyoglobin oxidizing system (MOS: 0.01 to 0.1 mM metmyoglobin/H(2)O(2)) that produced hydroxyl radical and ferryl species, respectively. Both oxidizing systems promoted MP thermal gelation, which was evidenced by rapid protein-protein interaction and the enhancement in storage modulus (elasticity) of the gel network as revealed by dynamic rheological testing in the 20 to 74 degrees C temperature range. This gelation-enhancing effect was attributed to the shift of myosin aggregation in the early stage of heating from predominantly head-head association (nonoxidized control samples) to prevalently tail-tail cross-linking through disulfide bonds. However, both hardness and water-holding capacity of chilled gels tended to decline when MP was exposed to >or=1 mM H(2)O(2) in IOS and to all concentrations of metmyoglobin in MOS. Microscopic examination confirmed a more porous structure in oxidized gels when compared with nonoxidized protein gels. The results demonstrated that mild oxidation altered the mode of myosin aggregation in favor of an elastic gel network formation, but it did not improve or had a negative effect on water-binding properties of MP gels. Mild oxidation promotes protein network formation and enhances gelation of myofibrillar protein under normal salt and pH conditions used in meat processing. This oxidative effect, which involves disulfide linkages, is somewhat similar to that in bakery product processing where oxidants are used to improve dough performance through gluten protein interaction.

  9. Ionically Cross-Linked Polymer Networks for the Multiple-Month Release of Small Molecules

    PubMed Central

    2016-01-01

    Long-term (multiple-week or -month) release of small, water-soluble molecules from hydrogels remains a significant pharmaceutical challenge, which is typically overcome at the expense of more-complicated drug carrier designs. Such approaches are payload-specific and include covalent conjugation of drugs to base materials or incorporation of micro- and nanoparticles. As a simpler alternative, here we report a mild and simple method for achieving multiple-month release of small molecules from gel-like polymer networks. Densely cross-linked matrices were prepared through ionotropic gelation of poly(allylamine hydrochloride) (PAH) with either pyrophosphate (PPi) or tripolyphosphate (TPP), all of which are commonly available commercial molecules. The loading of model small molecules (Fast Green FCF and Rhodamine B dyes) within these polymer networks increases with the payload/network binding strength and with the PAH and payload concentrations used during encapsulation. Once loaded into the PAH/PPi and PAH/TPP ionic networks, only a few percent of the payload is released over multiple months. This extended release is achieved regardless of the payload/network binding strength and likely reflects the small hydrodynamic mesh size within the gel-like matrices. Furthermore, the PAH/TPP networks show promising in vitro cytocompatibility with model cells (human dermal fibroblasts), though slight cytotoxic effects were exhibited by the PAH/PPi networks. Taken together, the above findings suggest that PAH/PPi and (especially) PAH/TPP networks might be attractive materials for the multiple-month delivery of drugs and other active molecules (e.g., fragrances or disinfectants). PMID:26811936

  10. Single-Molecule Unbinding Forces between the Polysaccharide Hyaluronan and Its Binding Proteins.

    PubMed

    Bano, Fouzia; Tammi, Markku I; Kang, David W; Harris, Edward N; Richter, Ralf P

    2018-06-19

    The extracellular polysaccharide hyaluronan (HA) is ubiquitous in all vertebrate tissues, where its various functions are encoded in the supramolecular complexes and matrices that it forms with HA-binding proteins (hyaladherins). In tissues, these supramolecular architectures are frequently subjected to mechanical stress, yet how this affects the intermolecular bonding is largely unknown. Here, we used a recently developed single-molecule force spectroscopy platform to analyze and compare the mechanical strength of bonds between HA and a panel of hyaladherins from the Link module superfamily, namely the complex of the proteoglycan aggrecan and cartilage link protein, the proteoglycan versican, the inflammation-associated protein TSG-6, the HA receptor for endocytosis (stabilin-2/HARE), and the HA receptor CD44. We find that the resistance to tensile stress for these hyaladherins correlates with the size of the HA-binding domain. The lowest mean rupture forces are observed for members of the type A subgroup (i.e., with the shortest HA-binding domains; TSG-6 and HARE). In contrast, the mechanical stability of the bond formed by aggrecan in complex with cartilage link protein (two members of the type C subgroup, i.e., with the longest HA-binding domains) and HA is equal or even superior to the high affinity streptavidin⋅biotin bond. Implications for the molecular mechanism of unbinding of HA⋅hyaladherin bonds under force are discussed, which underpin the mechanical properties of HA⋅hyaladherin complexes and HA-rich extracellular matrices. Copyright © 2018 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  11. Let there be light: photo-cross-linked block copolymer nanoparticles.

    PubMed

    Roy, Debashish; Sumerlin, Brent S

    2014-01-01

    Polymeric nanoparticles are prepared by selectively cross-linking a photo-sensitive dimethylmaleimide-containing block of a diblock copolymer via UV irradiation. A well-defined photo-cross-linkable block copolymer is prepared via reversible addition-fragmentation chain transfer (RAFT) polymerization of a dimethylmaleimide-functional acrylamido monomer containing photoreactive pendant groups with a poly(N,N-dimethylacrylamide) (PDMA) macro-chain transfer agent. The resulting amphiphilic block copolymers form micelles in water with a hydrophilic PDMA shell and a hydrophobic photo-cross-linkable dimethylmaleimide-containing core. UV irradiation results in photodimerization of the dimethylmaleimide groups within the micelle cores to yield core-cross-linked aggregates. Alternatively, UV irradiation of homogeneous solutions of the block copolymer in a non-selective solvent leads to in situ nanoparticle formation. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Polymeric micelles with ionic cores containing biodegradable cross-links for delivery of chemotherapeutic agents.

    PubMed

    Kim, Jong Oh; Sahay, Gaurav; Kabanov, Alexander V; Bronich, Tatiana K

    2010-04-12

    Novel functional polymeric nanocarriers with ionic cores containing biodegradable cross-links were developed for delivery of chemotherapeutic agents. Block ionomer complexes (BIC) of poly(ethylene oxide)-b-poly(methacylic acid) (PEO-b-PMA) and divalent metal cations (Ca(2+)) were utilized as templates. Disulfide bonds were introduced into the ionic cores by using cystamine as a biodegradable cross-linker. The resulting cross-linked micelles with disulfide bonds represented soft, hydrogel-like nanospheres and demonstrated a time-dependent degradation in the conditions mimicking the intracellular reducing environment. The ionic character of the cores allowed to achieve a very high level of doxorubicin (DOX) loading (50% w/w) into the cross-linked micelles. DOX-loaded degradable cross-linked micelles exhibited more potent cytotoxicity against human A2780 ovarian carcinoma cells as compared to micellar formulations without disulfide linkages. These novel biodegradable cross-linked micelles are expected to be attractive candidates for delivery of anticancer drugs.

  13. The effect of ionotropic gelation residence time on alginate cross-linking and properties.

    PubMed

    Patel, Mitulkumar A; AbouGhaly, Mohamed H H; Schryer-Praga, Jacqueline V; Chadwick, Keith

    2017-01-02

    The ability to engineer biocompatible polymers with controllable properties is highly desirable. One such approach is to cross-link carbohydrate polymers using ionotropic gelation (IG). Previous studies have investigated the effect of curing time on alginate cross-linking. Herein, we discuss a novel study detailing the effect of IG residence time (IGRT) on the cross-linking of alginate with calcium ions (Ca 2+ ) along with water migration (syneresis) and their subsequent impact on the pharmaceutical properties of alginate particles. IGRT was shown to have a significant effect on particle size, porosity, density, mechanical strength and swelling of calcium alginate particles as well as drug release mechanism. Furthermore, we describe a novel application of electron dispersive spectroscopy (EDS), in conjunction with Fourier Transform- infra red (FT-IR) spectroscopy, to analyze and monitor the changes in Ca 2+ concentration during cross-linking. A simple procedure to determine the concentration and distribution of the surface and internal Ca 2+ involved in alginate cross-linking was successfully developed. Copyright © 2016 Elsevier Ltd. All rights reserved.

  14. Electrochemical and spectroscopic characterization of surface sol-gel processes.

    PubMed

    Chen, Xiaohong; Wilson, George S

    2004-09-28

    (3-Mercaptopropyl)trimethoxysilane (MTS) forms a unique film on a platinum substrate by self-assembly and sol-gel cross-linking. The gelating and drying states of the self-assembled MTS sol-gel films were probed by use of electrochemical and spectroscopic methods. The thiol moiety was the only active group within the sol-gel network. Gold nanoparticles were employed to detect the availability of the thiol group and their interaction further indicated the physicochemical states of the sol-gel inner structure. It was found that the thiol groups in the open porous MTS aerogel matrix were accessible to the gold nanoparticles while thiol groups in the compact MTS xerogel network were not accessible to the gold nanoparticles. The characteristics of the sol-gel matrix change with time because of its own irreversible gelating and drying process. The present work provides direct evidence of gold nanoparticle binding with thiol groups within the sol-gel structures and explains the different permeability of "aerogel" and "xerogel" films of MTS on the basis of electrochemical and spectroscopic results. Two endogenous species, hydrogen peroxide and ascorbic acid, were used to test the permeability of the self-assembled sol-gel film in different states. The MTS xerogel film on the platinum electrode was extremely selective against ascorbic acid while maintaining high sensitivity to hydrogen peroxide in contrast to the relatively high permeability of ascorbic acid in the MTS aerogel film. This study showed the potential of the MTS sol-gel film as a nanoporous material in biosensor development.

  15. An Evaluation of Lysyl Oxidase–Derived Cross-Linking in Keratoconus by Liquid Chromatography/Mass Spectrometry

    PubMed Central

    Takaoka, Anna; Babar, Natasha; Hogan, Julia; Kim, MiJung; Price, Marianne O.; Price, Francis W.; Trokel, Stephen L.; Paik, David C.

    2016-01-01

    Purpose Current literature contains scant information regarding the extent of enzymatic collagen cross-linking in the keratoconus (KC) cornea. The aim of the present study was to examine levels of enzymatic lysyl oxidase–derived cross-links in stromal collagen in KC tissue, and to correlate the cross-link levels with collagen fibril stability as determined by thermal denaturation temperature (Tm). Methods Surgical KC samples (n = 17) and Eye-Bank control (n = 11) corneas of age 18 to 68 years were analyzed. The samples were defatted, reduced (NaBH4), hydrolyzed (6N HCl at 110°C for 18 hours), and cellulose enriched before analysis by C8 high-performance liquid chromatography equipped with parallel fluorescent and mass detectors in selective ion monitoring mode (20 mM heptafluorobutyric acid/methanol 70:30 isocratic at 1 mL/min). Nine different cross-links were measured, and the cross-link density was determined relative to collagen content (determined colorimetrically). The Tm was determined by differential scanning calorimetry. Results Cross-links detected were dihydroxylysinonorleucine (DHLNL), hydroxylysinonorleucine, lysinonorleucine (LNL), and histidinohydroxylysinonorleucine in both control and KC samples. Higher DHLNL levels were detected in KC, whereas the dominant cross-link, LNL, was decreased in KC samples. Decreased LNL levels were observed among KC ≤ 40 corneas. There was no difference in total cross-link density between KC samples and the controls. Pyridinolines, desmosines, and pentosidine were not detected. There was no notable correlation between cross-link levels with fibril instability as determined by Tm. Conclusions Lower levels of LNL in the KC cornea suggest that there might be a cross-linking defect either in fibrillar collagen or the microfibrillar elastic network composed of fibrillin. PMID:26780316

  16. Oral hyaluronan relieves knee pain: a review.

    PubMed

    Oe, Mariko; Tashiro, Toshiyuki; Yoshida, Hideto; Nishiyama, Hiroshi; Masuda, Yasunobu; Maruyama, Koh; Koikeda, Takashi; Maruya, Reiko; Fukui, Naoshi

    2016-01-27

    Hyaluronan (HA) is a component that is particularly abundant in the synovial fluid. Randomized, double-blinded, placebo-controlled trials carried out between 2008 and 2015 have proven the effectiveness of HA for the treatment of symptoms associated with synovitis, and particularly, knee pain, relief of synovial effusion or inflammation, and improvement of muscular knee strength. The mechanism by which HA exerts its effects in the living body, specifically receptor binding in the intestinal epithelia, has gradually been clarified. This review examines the effects of HA upon knee pain as assessed in clinical trials, as well as the mechanism of these effects and the safety of HA.

  17. The counterbend dynamics of cross-linked filament bundles and flagella

    PubMed Central

    Coy, Rachel

    2017-01-01

    Cross-linked filament bundles, such as in cilia and flagella, are ubiquitous in biology. They are considered in textbooks as simple filaments with larger stiffness. Recent observations of flagellar counterbend, however, show that induction of curvature in one section of a passive flagellum instigates a compensatory counter-curvature elsewhere, exposing the intricate role of the diminutive cross-linking proteins at large scales. We show that this effect, a material property of the cross-linking mechanics, modifies the bundle dynamics and induces a bimodal L2 − L3 length-dependent material response that departs from the Euler–Bernoulli theory. Hence, the use of simpler theories to analyse experiments can result in paradoxical interpretations. Remarkably, the counterbend dynamics instigates counter-waves in opposition to driven oscillations in distant parts of the bundle, with potential impact on the regulation of flagellar bending waves. These results have a range of physical and biological applications, including the empirical disentanglement of material quantities via counterbend dynamics. PMID:28566516

  18. 21 CFR 177.2420 - Polyester resins, cross-linked.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... following prescribed conditions: (a) The cross-linked polyester resins are produced by the condensation of... fiber Polyester fiber produced by the condensation of one or more of the acids listed in paragraph (a)(1...

  19. Automated Assignment of MS/MS Cleavable Cross-Links in Protein 3D-Structure Analysis

    NASA Astrophysics Data System (ADS)

    Götze, Michael; Pettelkau, Jens; Fritzsche, Romy; Ihling, Christian H.; Schäfer, Mathias; Sinz, Andrea

    2015-01-01

    CID-MS/MS cleavable cross-linkers hold an enormous potential for an automated analysis of cross-linked products, which is essential for conducting structural proteomics studies. The created characteristic fragment ion patterns can easily be used for an automated assignment and discrimination of cross-linked products. To date, there are only a few software solutions available that make use of these properties, but none allows for an automated analysis of cleavable cross-linked products. The MeroX software fills this gap and presents a powerful tool for protein 3D-structure analysis in combination with MS/MS cleavable cross-linkers. We show that MeroX allows an automatic screening of characteristic fragment ions, considering static and variable peptide modifications, and effectively scores different types of cross-links. No manual input is required for a correct assignment of cross-links and false discovery rates are calculated. The self-explanatory graphical user interface of MeroX provides easy access for an automated cross-link search platform that is compatible with commonly used data file formats, enabling analysis of data originating from different instruments. The combination of an MS/MS cleavable cross-linker with a dedicated software tool for data analysis provides an automated workflow for 3D-structure analysis of proteins. MeroX is available at www.StavroX.com .

  20. CLMSVault: A Software Suite for Protein Cross-Linking Mass-Spectrometry Data Analysis and Visualization.

    PubMed

    Courcelles, Mathieu; Coulombe-Huntington, Jasmin; Cossette, Émilie; Gingras, Anne-Claude; Thibault, Pierre; Tyers, Mike

    2017-07-07

    Protein cross-linking mass spectrometry (CL-MS) enables the sensitive detection of protein interactions and the inference of protein complex topology. The detection of chemical cross-links between protein residues can identify intra- and interprotein contact sites or provide physical constraints for molecular modeling of protein structure. Recent innovations in cross-linker design, sample preparation, mass spectrometry, and software tools have significantly improved CL-MS approaches. Although a number of algorithms now exist for the identification of cross-linked peptides from mass spectral data, a dearth of user-friendly analysis tools represent a practical bottleneck to the broad adoption of the approach. To facilitate the analysis of CL-MS data, we developed CLMSVault, a software suite designed to leverage existing CL-MS algorithms and provide intuitive and flexible tools for cross-platform data interpretation. CLMSVault stores and combines complementary information obtained from different cross-linkers and search algorithms. CLMSVault provides filtering, comparison, and visualization tools to support CL-MS analyses and includes a workflow for label-free quantification of cross-linked peptides. An embedded 3D viewer enables the visualization of quantitative data and the mapping of cross-linked sites onto PDB structural models. We demonstrate the application of CLMSVault for the analysis of a noncovalent Cdc34-ubiquitin protein complex cross-linked under different conditions. CLMSVault is open-source software (available at https://gitlab.com/courcelm/clmsvault.git ), and a live demo is available at http://democlmsvault.tyerslab.com/ .

  1. Conformational analysis of the Streptococcus pneumoniae hyaluronate lyase and characterization of its hyaluronan-specific carbohydrate-binding module.

    PubMed

    Suits, Michael D L; Pluvinage, Benjamin; Law, Adrienne; Liu, Yan; Palma, Angelina S; Chai, Wengang; Feizi, Ten; Boraston, Alisdair B

    2014-09-26

    For a subset of pathogenic microorganisms, including Streptococcus pneumoniae, the recognition and degradation of host hyaluronan contributes to bacterial spreading through the extracellular matrix and enhancing access to host cell surfaces. The hyaluronate lyase (Hyl) presented on the surface of S. pneumoniae performs this role. Using glycan microarray screening, affinity electrophoresis, and isothermal titration calorimetry we show that the N-terminal module of Hyl is a hyaluronan-specific carbohydrate-binding module (CBM) and the founding member of CBM family 70. The 1.2 Å resolution x-ray crystal structure of CBM70 revealed it to have a β-sandwich fold, similar to other CBMs. The electrostatic properties of the binding site, which was identified by site-directed mutagenesis, are distinct from other CBMs and complementary to its acidic ligand, hyaluronan. Dynamic light scattering and solution small angle x-ray scattering revealed the full-length Hyl protein to exist as a monomer/dimer mixture in solution. Through a detailed analysis of the small angle x-ray scattering data, we report the pseudoatomic solution structures of the monomer and dimer forms of the full-length multimodular Hyl. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  2. Kinetics of swelling of polyelectrolyte gels: Fixed degree of ionization

    NASA Astrophysics Data System (ADS)

    Sen, Swati; Kundagrami, Arindam

    2015-12-01

    The swelling kinetics of uncharged and charged polymer (polyelectrolyte) gels in salt-free conditions is studied in one dimension by solving the constitutive equation of motion (Newton's law for the elementary gel volume) of the displacement variable by two theoretical methods: one in which the classical definition of stress is used with the bulk modulus taken as a parameter, and the other in which a phenomenological expression of the osmotic stress as a function of polymer density and degree of ionization is taken as an input to the dynamics. The time-evolution profiles for spatially varying polymer density and stress, along with the location of the gel-solvent interface, are obtained from the two methods. We show that both the polymer density (volume fraction) and stress inside the gel follow expected behaviours of being maximum for the uniformly shrunken gel, and relaxing slowly to the lowest values as the gel approaches equilibrium. We further show that, by comparing the temporal profiles of the gel-solvent interface and other variables between the two methods, one may attempt to assign an effective bulk modulus to the polyelectrolyte gel as a function of the degree of ionization and other parameters of the gel such as hydrophobicity, cross-link density, and the temperature. The major result we get is that the effective bulk modulus of a polyelectrolyte gel increases monotonically with its degree of ionization. In the process of identifying the parameters for a monotonic swelling, we calculated using a well-known expression of the free energy the equilibrium results of two-phase co-existence and the critical point of a polyelectrolyte gel with a fixed degree of ionization.

  3. Kinetics of swelling of polyelectrolyte gels: Fixed degree of ionization.

    PubMed

    Sen, Swati; Kundagrami, Arindam

    2015-12-14

    The swelling kinetics of uncharged and charged polymer (polyelectrolyte) gels in salt-free conditions is studied in one dimension by solving the constitutive equation of motion (Newton's law for the elementary gel volume) of the displacement variable by two theoretical methods: one in which the classical definition of stress is used with the bulk modulus taken as a parameter, and the other in which a phenomenological expression of the osmotic stress as a function of polymer density and degree of ionization is taken as an input to the dynamics. The time-evolution profiles for spatially varying polymer density and stress, along with the location of the gel-solvent interface, are obtained from the two methods. We show that both the polymer density (volume fraction) and stress inside the gel follow expected behaviours of being maximum for the uniformly shrunken gel, and relaxing slowly to the lowest values as the gel approaches equilibrium. We further show that, by comparing the temporal profiles of the gel-solvent interface and other variables between the two methods, one may attempt to assign an effective bulk modulus to the polyelectrolyte gel as a function of the degree of ionization and other parameters of the gel such as hydrophobicity, cross-link density, and the temperature. The major result we get is that the effective bulk modulus of a polyelectrolyte gel increases monotonically with its degree of ionization. In the process of identifying the parameters for a monotonic swelling, we calculated using a well-known expression of the free energy the equilibrium results of two-phase co-existence and the critical point of a polyelectrolyte gel with a fixed degree of ionization.

  4. A general method for targeted quantitative cross-linking mass spectrometry

    USDA-ARS?s Scientific Manuscript database

    Chemical cross-linking mass spectrometry (XL-MS) provides protein structural information by identifying covalently linked proximal amino acid residues on protein surfaces. The information gained by this technique is complementary to other structural biology methods such as x-ray crystallography, NM...

  5. Highly Conductive Ionic-Liquid Gels Prepared with Orthogonal Double Networks of a Low-Molecular-Weight Gelator and Cross-Linked Polymer.

    PubMed

    Kataoka, Toshikazu; Ishioka, Yumi; Mizuhata, Minoru; Minami, Hideto; Maruyama, Tatsuo

    2015-10-21

    We prepared a heterogeneous double-network (DN) ionogel containing a low-molecular-weight gelator network and a polymer network that can exhibit high ionic conductivity and high mechanical strength. An imidazolium-based ionic liquid was first gelated by the molecular self-assembly of a low-molecular-weight gelator (benzenetricarboxamide derivative), and methyl methacrylate was polymerized with a cross-linker to form a cross-linked poly(methyl methacrylate) (PMMA) network within the ionogel. Microscopic observation and calorimetric measurement revealed that the fibrous network of the low-molecular-weight gelator was maintained in the DN ionogel. The PMMA network strengthened the ionogel of the low-molecular-weight gelator and allowed us to handle the ionogel using tweezers. The orthogonal DNs produced ionogels with a broad range of storage elastic moduli. DN ionogels with low PMMA concentrations exhibited high ionic conductivity that was comparable to that of a neat ionic liquid. The present study demonstrates that the ionic conductivities of the DN and single-network, low-molecular-weight gelator or polymer ionogels strongly depended on their storage elastic moduli.

  6. CrossLink: a novel method for cross-condition classification of cancer subtypes.

    PubMed

    Ma, Chifeng; Sastry, Konduru S; Flore, Mario; Gehani, Salah; Al-Bozom, Issam; Feng, Yusheng; Serpedin, Erchin; Chouchane, Lotfi; Chen, Yidong; Huang, Yufei

    2016-08-22

    We considered the prediction of cancer classes (e.g. subtypes) using patient gene expression profiles that contain both systematic and condition-specific biases when compared with the training reference dataset. The conventional normalization-based approaches cannot guarantee that the gene signatures in the reference and prediction datasets always have the same distribution for all different conditions as the class-specific gene signatures change with the condition. Therefore, the trained classifier would work well under one condition but not under another. To address the problem of current normalization approaches, we propose a novel algorithm called CrossLink (CL). CL recognizes that there is no universal, condition-independent normalization mapping of signatures. In contrast, it exploits the fact that the signature is unique to its associated class under any condition and thus employs an unsupervised clustering algorithm to discover this unique signature. We assessed the performance of CL for cross-condition predictions of PAM50 subtypes of breast cancer by using a simulated dataset modeled after TCGA BRCA tumor samples with a cross-validation scheme, and datasets with known and unknown PAM50 classification. CL achieved prediction accuracy >73 %, highest among other methods we evaluated. We also applied the algorithm to a set of breast cancer tumors derived from Arabic population to assign a PAM50 classification to each tumor based on their gene expression profiles. A novel algorithm CrossLink for cross-condition prediction of cancer classes was proposed. In all test datasets, CL showed robust and consistent improvement in prediction performance over other state-of-the-art normalization and classification algorithms.

  7. Hyaluronan and N-ERC/mesothelin as key biomarkers in a specific two-step model to predict pleural malignant mesothelioma.

    PubMed

    Mundt, Filip; Nilsonne, Gustav; Arslan, Sertaç; Csürös, Karola; Hillerdal, Gunnar; Yildirim, Huseyin; Metintas, Muzaffer; Dobra, Katalin; Hjerpe, Anders

    2013-01-01

    Diagnosis of malignant mesothelioma is challenging. The first available diagnostic material is often an effusion and biochemical analysis of soluble markers may provide additional diagnostic information. This study aimed to establish a predictive model using biomarkers from pleural effusions, to allow early and accurate diagnosis. Effusions were collected prospectively from 190 consecutive patients at a regional referral centre. Hyaluronan, N-ERC/mesothelin, C-ERC/mesothelin, osteopontin, syndecan-1, syndecan-2, and thioredoxin were measured using ELISA and HPLC. A predictive model was generated and validated using a second prospective set of 375 effusions collected consecutively at a different referral centre. Biochemical markers significantly associated with mesothelioma were hyaluronan (odds ratio, 95% CI: 8.82, 4.82-20.39), N-ERC/mesothelin (4.81, 3.19-7.93), CERC/mesothelin (3.58, 2.43-5.59) and syndecan-1 (1.34, 1.03-1.77). A two-step model using hyaluronan and N-ERC/mesothelin, and combining a threshold decision rule with logistic regression, yielded good discrimination with an area under the ROC curve of 0.99 (95% CI: 0.97-1.00) in the model generation dataset and 0.83 (0.74-0.91) in the validation dataset, respectively. A two-step model using hyaluronan and N-ERC/mesothelin predicts mesothelioma with high specificity. This method can be performed on the first available effusion and could be a useful adjunct to the morphological diagnosis of mesothelioma.

  8. Vision Restoration with a Collagen Cross-linked Boston Keratoprosthesis Unit

    DTIC Science & Technology

    2017-09-01

    Tissue Bank International, the tissue bank used for this study changed their company name to KeraLink International. The name was changed to better...solution to KeraLink International, and Dr. Joseph Ciolino traveled to the tissue bank to train staff on the Cross-Linking procedure. KeraLink is now... tissue by collagen-crosslinking the cornea graft ex vivo using vitamin B2 (riboflavin) and ultraviolet light. The overall objective of this study is

  9. Viscosupplementation for treatment of osteoarthritis: from initial discovery to current status and results.

    PubMed

    Balazs, Endre A

    2004-01-01

    was introduced, the same highly elastoviscous hyaluronan solutions also were applied in ophthalmic surgery as viscosurgical tools to protect sensitive tissues in the eye during surgery and to be used as soft instruments for tissue manipulation. Modified hyaluronan products (gels) also were introduced for augmentation of the intercellular matrix in tissues (viscoaugmentation) and for separating tissues to prevent adhesions and excessive scar formation (viscoseparation). Hyaluronan and its derivatives (gels) also have been used for drug delivery. The therapeutic use of highly elastoviscous solutions and gels of hyaluronan and its derivatives to build intercellular matrices for supplementation, regeneration, and developing new tissues introduced the concept of matrix engineering into medical practice.

  10. Absorption of Orally Administered Hyaluronan.

    PubMed

    Kimura, Mamoru; Maeshima, Takuya; Kubota, Takumi; Kurihara, Hitoshi; Masuda, Yasunobu; Nomura, Yoshihiro

    2016-12-01

    Hyaluronan (HA) has been utilized as a supplement. However, the absorption of orally administrated HA remains controversial. The degradation and absorption of HA in the intestine were investigated in this study. HA excretion into the feces, degradation in the intestinal tract, absorption through the large intestine, and translocation to the blood and skin were examined. HA administered orally was not detected in rat feces. HA was degraded by cecal content, but not by artificial gastric juice and intestinal juice. Oligosaccharide HA passed through excised large intestine sacs. Furthermore, disaccharides, tetrasaccharides, and polysaccharides HA were distributed to the skin of rats following oral administration of high molecular weight HA (300 kDa). The results of the study suggest that orally administered HA is degraded to oligosaccharides by intestinal bacteria, and oligosaccharide HA is absorbed in the large intestine and is subsequently distributed throughout the tissues, including the skin.

  11. Analysis of splicing complexes on native gels.

    PubMed

    Ares, Manuel

    2013-10-01

    Splicing requires a complex set of ATP-dependent macromolecular associations that lead to the rearrangement of just a few covalent bonds in the pre-mRNA substrate. Seeing only the covalent bonds breaking and forming is seeing only a very small part of the process. Analysis of native splicing complexes into which the radiolabeled substrate has been assembled, but not necessarily completely reacted, has provided a good understanding of the process. This protocol describes a gel method for detecting and analyzing yeast splicing complexes formed in vitro on a radiolabeled pre-mRNA substrate. Complexes formed during the splicing reaction are quenched by dilution and addition of an excess of RNA, which is thought to strip nonspecifically bound proteins from the labeled substrate RNA. After loading on a low-percentage, low-cross-linking ratio composite agarose-acrylamide gel (in 10% glycerol), labeled bands are detected. These can be extracted and shown to contain small nuclear RNAs (snRNAs) and partly reacted pre-mRNA.

  12. Cross-linked compared with historical polyethylene in THA: an 8-year clinical study.

    PubMed

    Geerdink, Carel H; Grimm, Bernd; Vencken, Wendy; Heyligers, Ide C; Tonino, Alphons J

    2009-04-01

    Wear particle-induced osteolysis is a major cause of aseptic loosening in THA. Increasing wear resistance of polyethylene (PE) occurs by increasing the cross-link density and early reports document low wear rates with such implants. To confirm longer-term reductions in wear we compared cross-linked polyethylene (irradiation in nitrogen, annealing) with historical polyethylene (irradiation in air) in a prospective, randomized clinical study involving 48 patients who underwent THAs with a minimum followup of 7 years (mean, 8 years; range, 7-9 years). The insert material was the only variable. The Harris hip score, radiographic signs of osteolysis, and polyethylene wear were recorded annually. Twenty-three historical and 17 moderately cross-linked polyethylene inserts were analyzed (five patients died, three were lost to followup). At 8 years, the wear rate was lower for cross-linked polyethylene (0.088 +/- 0.03 mm/year) than for the historical polyethylene (0.142 +/- 0.07 mm/year). This reduction (38%) did not diminish with time (33% at 5 years). Acetabular cyst formation was less frequent (39% versus 12%), affected fewer DeLee and Charnley zones (17% versus 4%), and was less severe for the cross-linked polyethylene. The only revision was for an aseptically loose cup in the historical polyethylene group. Moderately cross-linked polyethylene maintained its wear advantage with time and produced less osteolysis, showing no signs of aging at mid-term followup. Level I, therapeutic study. See Guidelines for Authors for a complete description of levels of evidence.

  13. Multiple repair pathways mediate tolerance to chemotherapeutic cross-linking agents in vertebrate cells.

    PubMed

    Nojima, Kuniharu; Hochegger, Helfrid; Saberi, Alihossein; Fukushima, Toru; Kikuchi, Koji; Yoshimura, Michio; Orelli, Brian J; Bishop, Douglas K; Hirano, Seiki; Ohzeki, Mioko; Ishiai, Masamichi; Yamamoto, Kazuhiko; Takata, Minoru; Arakawa, Hiroshi; Buerstedde, Jean-Marie; Yamazoe, Mitsuyoshi; Kawamoto, Takuo; Araki, Kasumi; Takahashi, Jun A; Hashimoto, Nobuo; Takeda, Shunichi; Sonoda, Eiichiro

    2005-12-15

    Cross-linking agents that induce DNA interstrand cross-links (ICL) are widely used in anticancer chemotherapy. Yeast genetic studies show that nucleotide excision repair (NER), Rad6/Rad18-dependent postreplication repair, homologous recombination, and cell cycle checkpoint pathway are involved in ICL repair. To study the contribution of DNA damage response pathways in tolerance to cross-linking agents in vertebrates, we made a panel of gene-disrupted clones from chicken DT40 cells, each defective in a particular DNA repair or checkpoint pathway, and measured the sensitivities to cross-linking agents, including cis-diamminedichloroplatinum (II) (cisplatin), mitomycin C, and melphalan. We found that cells harboring defects in translesion DNA synthesis (TLS), Fanconi anemia complementation groups (FANC), or homologous recombination displayed marked hypersensitivity to all the cross-linking agents, whereas NER seemed to play only a minor role. This effect of replication-dependent repair pathways is distinctively different from the situation in yeast, where NER seems to play a major role in dealing with ICL. Cells deficient in Rev3, the catalytic subunit of TLS polymerase Polzeta, showed the highest sensitivity to cisplatin followed by fanc-c. Furthermore, epistasis analysis revealed that these two mutants work in the same pathway. Our genetic comprehensive study reveals a critical role for DNA repair pathways that release DNA replication block at ICLs in cellular tolerance to cross-linking agents and could be directly exploited in designing an effective chemotherapy.

  14. Effect of cross-link density on carbon dioxide separation in polydimethylsiloxane-norbornene membranes

    DOE PAGES

    Hong, Tao; Niu, Zhenbin; Hu, Xunxiang; ...

    2015-10-20

    The development of high performance materials for CO 2 separation and capture will significantly contribute to a solution for climate change. In this work, (bicycloheptenyl) ethyl terminated polydimethylsiloxane (PDMSPNB) membranes with varied cross-link densities were synthesized via ring-opening metathesis polymerization. The developed polymer membranes show higher permeability and better selectivity than those of conventional cross-linked PDMS membrane. The achieved performance (CO 2 permeability ~ 6800 Barrer and CO 2/N 2 selectivity ~ 14) is very promising for practical applications. The key to achieving this high performance is the use of an in-situ cross-linking method of the difunctional PDMS macromonomers, whichmore » provides lightly cross-linked membranes. By combining positron annihilation lifetime spectroscopy, broadband dielectric spectroscopy and gas solubility measurements, we have elucidated the key parameters necessary for achieving their excellent performance.« less

  15. Impact of styrenic polymer one-step hyper-cross-linking on volatile organic compound adsorption and desorption performance.

    PubMed

    Ghafari, Mohsen; Atkinson, John D

    2018-06-05

    A novel one-step hyper-cross-linking method, using 1,2-dichloroethane (DCE) and 1,6-dichlorohexane (DCH) cross-linkers, expands the micropore volume of commercial styrenic polymers. Performance of virgin and modified polymers was evaluated by measuring hexane, toluene, and methyl-ethyl-ketone (MEK) adsorption capacity, adsorption/desorption kinetics, and desorption efficiency. Hyper-cross-linked polymers have up to 128% higher adsorption capacity than virgin polymers at P/P 0  = 0.05 due to micropore volume increases up to 330%. Improvements are most pronounced with the DCE cross-linker. Hyper-cross-linking has minimal impact on hexane adsorption kinetics, but adsorption rates for toluene and MEK decrease by 6-41%. Desorption rates decreased (3-36%) for all materials after hyper-cross-linking, with larger decreases for DCE hyper-cross-linked polymers due to smaller average pore widths. For room temperature desorption, 20-220% more adsorbate remains in hyper-cross-linked polymers after regeneration compared to virgin materials. DCE hyper-cross-linked polymers have 13-92% more residual adsorbate than DCH counterparts. Higher temperatures were required for DCE hyper-cross-linked polymers to completely desorb VOCs compared to the DCH hyper-cross-linked and virgin counterparts. Results show that the one-step hyper-cross-linking method for modifying styrenic polymers improves adsorption capacity because of added micropores, but decreases adsorption/desorption kinetics and desorption efficiency for large VOCs due to a decrease in average pore width. Copyright © 2018 Elsevier B.V. All rights reserved.

  16. Radiological properties of normoxic polymer gel dosimeters

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Venning, A.J.; Nitschke, K.N.; Keall, P.J.

    2005-04-01

    The radiological properties of the normoxic polymer gel dosimeters MAGIC, MAGAS, and MAGAT [methacrylic and ascorbic acid in gelatin initiated by copper; methacrylic acid gelatine gel with ascorbic acid; and methacrylic acid gelatine and tetrakis (hydroxymethyl) phosphonium chloride, respectively] have been investigated. The radiological water equivalence was determined by comparing the polymer gel macroscopic photon and electron interaction cross sections over the energy range from 10 keV to 20 MeV and by Monte Carlo modeling of depth doses. Normoxic polymer gel dosimeters have a high gelatine and monomer concentration and therefore mass density (kg m{sup -3}) up to 3.8% highermore » than water. This results in differences between the cross-section ratios of the normoxic polymer gels and water of up to 3% for the attenuation, energy absorption, and collision stopping power coefficient ratios through the Compton dominant energy range. The mass cross-section ratios were within 2% of water except for the mass attenuation and energy absorption coefficients ratios, which showed differences with water of up to 6% for energies less than 100 keV. Monte Carlo modeling was undertaken for the polymer gel dosimeters to model the electron and photon transport resulting from a 6 MV photon beam. The absolute percentage differences between gel and water were within 1% and the relative percentage differences were within 3.5%. The results show that the MAGAT gel formulation is the most radiological water equivalent of the normoxic polymer gel dosimeters investigated due to its lower mass density measurement compared with MAGAS and MAGIC gels.« less

  17. Preparation and characterization of cross-linked excipient of coprocessed xanthan gum-acacia gum as matrix for sustained release tablets

    NASA Astrophysics Data System (ADS)

    Surini, Silvia; Wati, Dina Risma; Syahdi, Rezi Riadhi

    2018-02-01

    Sustained release tablet is solid dosage form which is designed to release drugs slowly in the body. This research was intended to prepare and characterize the cross-linked excipients of co-processed xanthan gum-acacia gum (CL-Co-XGGA) as matrices for sustained release tablets with gliclazide as a model drug. CL-Co-XGGA excipients were cross-linked materials of co-processed excipients of xanthan gum-acacia gum (Co-XGGA) using sodium trimetaphosphate. Co-processed excipients of xanthan gum-acacia gum were prepared in the ratio of each excipient 1:2, 1:1 and 2:1. Co-XGGA and CL-Co-XGGA excipients were characterized physically, chemically and functionally. Then, the sustained release (SR) tablets were formulated by wet granulation method using CL-Co-XGGA excipients as matrices. Also, the dissolution study of the gliclazide SR tablets was carried out in phosphate buffer medium pH 7,4 containing sodium lauryl sulphate 0.2% for 12 hours. The results showed that the degree of substitution (DS) of CL-Co-XGGA 1:2, 1:1, 2:1 excipients were respectively 0.067, 0.082 and 0.08. Besides that, the excipients gel strengths were 14.03, 17.27 and 20,70 gF, respectively. The cross-linked excipients had improved flow properties and swelling capability compared to the Co-XGGA excipients. The results of the gliclazide SR tablets evaluations showed that all tablets were passed all tablet requirements. Moreover, the gliclazide release from SR tablets F1 - F6 revealed the sustained release profile, which was following zero order kinetics (F1, F2, F3, F6) and Higuchi kinetics (F4 and F5). It could be concluded that the obtained CL-Co-XGGA excipients might be used as matrices for sustained release tablets and could retard drug release up to 8 until 32 hours.

  18. The mechanism of collagen cross-linking in diabetes: a puzzle nearing resolution.

    PubMed

    Monnier, V M; Glomb, M; Elgawish, A; Sell, D R

    1996-07-01

    Considerable interest has been focused in recent years on the mechanism of collagen cross-linking by high glucose in vitro and in vivo. Experiments in both diabetic humans and in animals have shown that over time collagen becomes less soluble, less digestible by collagenase, more stable to heat-induced denaturation, and more glycated. In addition, collagen becomes more modified by advanced products of the Maillard reaction, i.e., immunoreactive advanced glycation end products and the glycoxidation markers carboxymethyllysine and pentosidine. Mechanistic studies have shown that collagen cross-linking in vitro can be uncoupled from glycation by the use of antioxidants and chelating agents. Experiments in the authors' laboratory revealed that approximately 50% of carboxymethyllysine formed in vitro originates from pathways other than oxidation of Amadori products, i.e., most likely the oxidation of Schiff base-linked glucose. In addition, the increase in thermal stability of rat tail tendons exposed to high glucose in vitro or in vivo was found to strongly depend on H2O2 formation. The final missing piece of the puzzle is that of the structure of the major cross-link. We speculate that it is a nonfluorescent nonultraviolet active cross-link between two lysine residues, which includes a fragmentation product of glucose linked in a nonreducible bond labile to both strong acids and bases.

  19. Case series: evaluation of a liquid silicone gel on scar appearance following excisional surgery--a pilot study.

    PubMed

    Spencer, James M

    2010-07-01

    Efforts to improve the size and appearance of scars have included therapies as varied as laser treatments and onion extract gels. Silicone gel sheeting is well know to improve the appearance of hypertrophic scars, and may have a role in the management of routine surgical and traumatic scars. By varying the degree of cross linking, silicone elastomer can be a solid sheet or a liquid gel. In this pilot series, seven patients applied a liquid silicone gel twice a day to one half of a new surgical scar for three months. At the end of this time, the treated side was noticeably better in appearance in five of seven patients while two of seven had no difference. In no patient was the silicone treated side worse in appearance.

  20. Carbon Nanofiber Incorporated Silica Based Aerogels with Di-Isocyanate Cross-Linking

    NASA Technical Reports Server (NTRS)

    Vivod, Stephanie L.; Meador, Mary Ann B.; Capadona, Lynn A.; Sullivan, Roy M.; Ghosn, Louis J.; Clark, Nicholas; McCorkle, Linda

    2008-01-01

    Lightweight materials with excellent thermal insulating properties are highly sought after for a variety of aerospace and aeronautic applications. (1) Silica based aerogels with their high surface area and low relative densities are ideal for applications in extreme environments such as insulators for the Mars Rover battery. (2) However, the fragile nature of aerogel monoliths prevents their widespread use in more down to earth applications. We have shown that the fragile aerogel network can be cross-linked with a di-isocyanate via amine decorated surfaces to form a conformal coating. (3) This coating reinforces the neck regions between secondary silica particles and significantly strengthens the aerogels with only a small effect on density or porosity. Scheme 1 depicts the cross-linking reaction with the di-isocyanate and exhibits the stages that result in polymer cross-linked aerogel monoliths.

  1. Experimental scleral cross-linking increases glaucoma damage in a mouse model

    PubMed Central

    Kimball, Elizabeth C.; Nguyen, Cathy; Steinhart, Matthew R.; Nguyen, Thao D.; Pease, Mary E.; Oglesby, Ericka N.; Oveson, Brian C.; Quigley, Harry A.

    2014-01-01

    The purpose of this study was to assess the effect of a scleral cross-linking agent on susceptibility to glaucoma damage in a mouse model. CD1 mice underwent 3 subconjunctival injections of 0.5 M glyceraldehyde (GA) in 1 week, then had elevated intraocular pressure (IOP) induced by bead injection. Degree of cross-linking was measured by enzyme-linked immunosorbent assay (ELISA), scleral permeability was measured by fluorescence recovery after photobleaching (FRAP), and the mechanical effects of GA exposure were measured by inflation testing. Control mice had buffer injection or no injection in 2 separate glaucoma experiments. IOP was monitored by Tonolab and retinal ganglion cell (RGC) loss was measured by histological axon counting. To rule out undesirable effects of GA, we performed electroretinography and detailed histology of the retina. GA exposure had no detectable effects on RGC number, retinal structure or function either histologically or electrophysiologically. GA increased cross-linking of sclera by 37% in an ELISA assay, decreased scleral permeability (FRAP, p = 0.001), and produced a steeper pressure—strain behavior by in vitro inflation testing. In two experimental glaucoma experiments, GA-treated eyes had greater RGC axon loss from elevated IOP than either buffer-injected or control eyes, controlling for level of IOP exposure over time (p = 0.01, and 0.049, multivariable regression analyses). This is the first report that experimental alteration of the sclera, by cross-linking, increases susceptibility to RGC damage in mice. PMID:25285424

  2. Interstrand cross-links arising from strand breaks at true abasic sites in duplex DNA

    PubMed Central

    Yang, Zhiyu; Price, Nathan E.; Johnson, Kevin M.

    2017-01-01

    Abstract Interstrand cross-links are exceptionally bioactive DNA lesions. Endogenous generation of interstrand cross-links in genomic DNA may contribute to aging, neurodegeneration, and cancer. Abasic (Ap) sites are common lesions in genomic DNA that readily undergo spontaneous and amine-catalyzed strand cleavage reactions that generate a 2,3-didehydro-2,3-dideoxyribose sugar remnant (3’ddR5p) at the 3’-terminus of the strand break. Interestingly, this strand scission process leaves an electrophilic α,β-unsaturated aldehyde residue embedded within the resulting nicked duplex. Here we present evidence that 3’ddR5p derivatives generated by spermine-catalyzed strand cleavage at Ap sites in duplex DNA can react with adenine residues on the opposing strand to generate a complex lesion consisting of an interstrand cross-link adjacent to a strand break. The cross-link blocks DNA replication by ϕ29 DNA polymerase, a highly processive polymerase enzyme that couples synthesis with strand displacement. This suggests that 3’ddR5p-derived cross-links have the potential to block critical cellular DNA transactions that require strand separation. LC-MS/MS methods developed herein provide powerful tools for studying the occurrence and properties of these cross-links in biochemical and biological systems. PMID:28531327

  3. Target-responsive DNAzyme cross-linked hydrogel for visual quantitative detection of lead.

    PubMed

    Huang, Yishun; Ma, Yanli; Chen, Yahong; Wu, Xuemeng; Fang, Luting; Zhu, Zhi; Yang, Chaoyong James

    2014-11-18

    Because of the severe health risks associated with lead pollution, rapid, sensitive, and portable detection of low levels of Pb(2+) in biological and environmental samples is of great importance. In this work, a Pb(2+)-responsive hydrogel was prepared using a DNAzyme and its substrate as cross-linker for rapid, sensitive, portable, and quantitative detection of Pb(2+). Gold nanoparticles (AuNPs) were first encapsulated in the hydrogel as an indicator for colorimetric analysis. In the absence of lead, the DNAzyme is inactive, and the substrate cross-linker maintains the hydrogel in the gel form. In contrast, the presence of lead activates the DNAzyme to cleave the substrate, decreasing the cross-linking density of the hydrogel and resulting in dissolution of the hydrogel and release of AuNPs for visual detection. As low as 10 nM Pb(2+) can be detected by the naked eye. Furthermore, to realize quantitative visual detection, a volumetric bar-chart chip (V-chip) was used for quantitative readout of the hydrogel system by replacing AuNPs with gold-platinum core-shell nanoparticles (Au@PtNPs). The Au@PtNPs released from the hydrogel upon target activation can efficiently catalyze the decomposition of H2O2 to generate a large volume of O2. The gas pressure moves an ink bar in the V-chip for portable visual quantitative detection of lead with a detection limit less than 5 nM. The device was able to detect lead in digested blood with excellent accuracy. The method developed can be used for portable lead quantitation in many applications. Furthermore, the method can be further extended to portable visual quantitative detection of a variety of targets by replacing the lead-responsive DNAzyme with other DNAzymes.

  4. A structural and kinetic study on myofibrils prevented from shortening by chemical cross-linking.

    PubMed

    Herrmann, C; Sleep, J; Chaussepied, P; Travers, F; Barman, T

    1993-07-20

    In previous work, we studied the early steps of the Mg(2+)-ATPase activity of Ca(2+)-activated myofibrils [Houadjeto, M., Travers, F., & Barman, T. (1992) Biochemistry 31, 1564-1569]. The myofibrils were free to contract, and the results obtained refer to the ATPase cycle of myofibrils contracting with no external load. Here we studied the ATPase of myofibrils contracting isometrically. To prevent shortening, we cross-linked them with 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (EDC). SDS-PAGE and Western blot analyses showed that the myosin rods were extensively cross-linked and that 8% of the myosin heads were cross-linked to the thin filament. The transient kinetics of the cross-linked myofibrils were studied in 0.1 M potassium acetate, pH 7.4 and 4 degrees C, by the rapid-flow quench method. The ATP binding steps were studied by the cold ATP chase and the cleavage and release of products steps by the Pi burst method. In Pi burst experiments, the sizes of the bursts were equal within experimental error to the ATPase site concentrations (as determined by the cold ATP chase methods) for both cross-linked (isometric) and un-cross-linked (isotonic) myofibrils. This shows that in both cases the rate-limiting step is after the cleavage of ATP. When cross-linked, the kcat of Ca(2+)-activated myofibrils was reduced from 1.7 to 0.8 s-1. This is consistent with the observation that fibers shortening at moderate velocity have a higher ATPase activity than isometric fibers.(ABSTRACT TRUNCATED AT 250 WORDS)

  5. Cross Country MetroLink Segment I Business Plan

    DOT National Transportation Integrated Search

    1997-12-02

    In the St. Louis, Missouri metropolitan area, the East-West Gateway Coordinating Council decided the route for the first MetroLink extension in the Cross-County Corridor in September 1997. The next phase, reflected in this paper is develop, during th...

  6. Bypass of a psoralen DNA interstrand cross-link by DNA polymerases beta, iota, and kappa in vitro

    PubMed Central

    Smith, Leigh A.; Makarova, Alena V.; Samson, Laura; Thiesen, Katherine E.; Dhar, Alok; Bessho, Tadayoshi

    2012-01-01

    Repair of DNA inter-strand cross-links in mammalian cells involves several biochemically distinctive processes, including the release of one of the cross-linked strands and translesion DNA synthesis (TLS). In this report, we investigated in vitro TLS activity of psoralen DNA inter-strand cross-link by three DNA repair polymerases, DNA polymerase beta, kappa and iota. DNA polymerase beta is capable of bypassing a psoralen cross-link with a low efficiency. Cell extracts prepared from DNA polymerase beta knockout mouse embryonic fibroblast showed a reduced bypass activity of the psoralen cross-link and purified DNA polymerase beta restored the bypass activity. In addition, DNA polymerase iota mis-incorporated thymine across the psoralen cross-link and DNA polymerase kappa extended these mis-paired primer ends, suggesting that DNA polymerase iota may serve as an inserter and DNA polymerase kappa may play a role as an extender in the repair of psoralen DNA inter-strand cross-links. The results demonstrated here indicate that multiple DNA polymerases could participate in TLS steps in mammalian DNA inter-strand cross-link repair. PMID:23106263

  7. Strain-enhanced stress relaxation impacts nonlinear elasticity in collagen gels

    PubMed Central

    Nam, Sungmin; Hu, Kenneth H.; Chaudhuri, Ovijit

    2016-01-01

    The extracellular matrix (ECM) is a complex assembly of structural proteins that provides physical support and biochemical signaling to cells in tissues. The mechanical properties of the ECM have been found to play a key role in regulating cell behaviors such as differentiation and malignancy. Gels formed from ECM protein biopolymers such as collagen or fibrin are commonly used for 3D cell culture models of tissue. One of the most striking features of these gels is that they exhibit nonlinear elasticity, undergoing strain stiffening. However, these gels are also viscoelastic and exhibit stress relaxation, with the resistance of the gel to a deformation relaxing over time. Recent studies have suggested that cells sense and respond to both nonlinear elasticity and viscoelasticity of ECM, yet little is known about the connection between nonlinear elasticity and viscoelasticity. Here, we report that, as strain is increased, not only do biopolymer gels stiffen but they also exhibit faster stress relaxation, reducing the timescale over which elastic energy is dissipated. This effect is not universal to all biological gels and is mediated through weak cross-links. Mechanistically, computational modeling and atomic force microscopy (AFM) indicate that strain-enhanced stress relaxation of collagen gels arises from force-dependent unbinding of weak bonds between collagen fibers. The broader effect of strain-enhanced stress relaxation is to rapidly diminish strain stiffening over time. These results reveal the interplay between nonlinear elasticity and viscoelasticity in collagen gels, and highlight the complexity of the ECM mechanics that are likely sensed through cellular mechanotransduction. PMID:27140623

  8. Liquid behavior of cross-linked actin bundles.

    PubMed

    Weirich, Kimberly L; Banerjee, Shiladitya; Dasbiswas, Kinjal; Witten, Thomas A; Vaikuntanathan, Suriyanarayanan; Gardel, Margaret L

    2017-02-28

    The actin cytoskeleton is a critical regulator of cytoplasmic architecture and mechanics, essential in a myriad of physiological processes. Here we demonstrate a liquid phase of actin filaments in the presence of the physiological cross-linker, filamin. Filamin condenses short actin filaments into spindle-shaped droplets, or tactoids, with shape dynamics consistent with a continuum model of anisotropic liquids. We find that cross-linker density controls the droplet shape and deformation timescales, consistent with a variable interfacial tension and viscosity. Near the liquid-solid transition, cross-linked actin bundles show behaviors reminiscent of fluid threads, including capillary instabilities and contraction. These data reveal a liquid droplet phase of actin, demixed from the surrounding solution and dominated by interfacial tension. These results suggest a mechanism to control organization, morphology, and dynamics of the actin cytoskeleton.

  9. Alkaline battery containing a separator of a cross-linked copolymer of vinyl alcohol and unsaturated carboxylic acid

    NASA Technical Reports Server (NTRS)

    Hsu, L. C.; Philipp, W. H.; Sheibley, D. W.; Gonzalez-Sanabria, O. D. (Inventor)

    1985-01-01

    A battery separator for an alkaline battery is described. The separator comprises a cross linked copolymer of vinyl alcohol units and unsaturated carboxylic acid units. The cross linked copolymer is insoluble in water, has excellent zincate diffusion and oxygen gas barrier properties and a low electrical resistivity. Cross linking with a polyaldehyde cross linking agent is preferred.

  10. Mixed-Isotope Labeling with LC-IMS-MS for Characterization of Protein–Protein Interactions by Chemical Cross-Linking

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Merkley, Eric D.; Baker, Erin S.; Crowell, Kevin L.

    2013-02-20

    Chemical cross-linking of proteins followed by proteolysis and mass spectrometric analysis of the resulting cross-linked peptides can provide insights into protein structure and protein-protein interactions. However, cross-linked peptides are by necessity of low stoichometry and have different physicochemical properties than linear peptides, routine unambiguous identification of the cross-linked peptides has remained difficult. To address this challenge, we demonstrated the use of liquid chromatography and ion mobility separations coupled with mass spectrometry in combination with a heavy-isotope labeling method. The combination of mixed-isotope cross-linking and ion mobility provided unique and easily interpretable spectral multiplet features for the intermolecular cross-linked peptides. Applicationmore » of the method to two different homodimeric proteins - SrfN, a virulence factor from Salmonella Typhimurium and SO_2176, a protein of unknown function from Shewanella oneidensis- revealed several cross-linked peptides from both proteins that were identified with a low false discovery rate (estimated using a decoy approach). A greater number of cross-linked peptides were identified using ion mobility drift time information in the analysis than when the data were summed across the drift time dimension before analysis. The identified cross-linked peptides migrated more quickly in the ion mobility drift tube than the unmodified peptides.« less

  11. Collagen type IX from human cartilage: a structural profile of intermolecular cross-linking sites.

    PubMed Central

    Diab, M; Wu, J J; Eyre, D R

    1996-01-01

    Type IX collagen, a quantitatively minor collagenous component of cartilage, is known to be associated with and covalently cross-linked to type II collagen fibrils in chick and bovine cartilage. Type IX collagen molecules have also been shown to form covalent cross-links with each other in bovine cartilage. In the present study we demonstrate by structural analysis and location of cross-linking sites that, in human cartilage, type IX collagen is covalently cross-linked to type II collagen and to other molecules of type IX collagen. We also present evidence that, if the proteoglycan form of type IX collagen is present in human cartilage, it can only be a minor component of the matrix, similar to findings with bovine cartilage. PMID:8660302

  12. ProXL (Protein Cross-Linking Database): A Platform for Analysis, Visualization, and Sharing of Protein Cross-Linking Mass Spectrometry Data

    PubMed Central

    2016-01-01

    ProXL is a Web application and accompanying database designed for sharing, visualizing, and analyzing bottom-up protein cross-linking mass spectrometry data with an emphasis on structural analysis and quality control. ProXL is designed to be independent of any particular software pipeline. The import process is simplified by the use of the ProXL XML data format, which shields developers of data importers from the relative complexity of the relational database schema. The database and Web interfaces function equally well for any software pipeline and allow data from disparate pipelines to be merged and contrasted. ProXL includes robust public and private data sharing capabilities, including a project-based interface designed to ensure security and facilitate collaboration among multiple researchers. ProXL provides multiple interactive and highly dynamic data visualizations that facilitate structural-based analysis of the observed cross-links as well as quality control. ProXL is open-source, well-documented, and freely available at https://github.com/yeastrc/proxl-web-app. PMID:27302480

  13. ProXL (Protein Cross-Linking Database): A Platform for Analysis, Visualization, and Sharing of Protein Cross-Linking Mass Spectrometry Data.

    PubMed

    Riffle, Michael; Jaschob, Daniel; Zelter, Alex; Davis, Trisha N

    2016-08-05

    ProXL is a Web application and accompanying database designed for sharing, visualizing, and analyzing bottom-up protein cross-linking mass spectrometry data with an emphasis on structural analysis and quality control. ProXL is designed to be independent of any particular software pipeline. The import process is simplified by the use of the ProXL XML data format, which shields developers of data importers from the relative complexity of the relational database schema. The database and Web interfaces function equally well for any software pipeline and allow data from disparate pipelines to be merged and contrasted. ProXL includes robust public and private data sharing capabilities, including a project-based interface designed to ensure security and facilitate collaboration among multiple researchers. ProXL provides multiple interactive and highly dynamic data visualizations that facilitate structural-based analysis of the observed cross-links as well as quality control. ProXL is open-source, well-documented, and freely available at https://github.com/yeastrc/proxl-web-app .

  14. Nanomechanics of layer-by-layer polyelectrolyte complexes: a manifestation of ionic cross-links and fixed charges.

    PubMed

    Han, Biao; Chery, Daphney R; Yin, Jie; Lu, X Lucas; Lee, Daeyeon; Han, Lin

    2016-01-28

    This study investigates the roles of two distinct features of ionically cross-linked polyelectrolyte networks - ionic cross-links and fixed charges - in determining their nanomechanical properties. The layer-by-layer assembled poly(allylamine hydrochloride)/poly(acrylic acid) (PAH/PAA) network is used as the model material. The densities of ionic cross-links and fixed charges are modulated through solution pH and ionic strength (IS), and the swelling ratio, elastic and viscoelastic properties are quantified via an array of atomic force microscopy (AFM)-based nanomechanical tools. The roles of ionic cross-links are underscored by the distinctive elastic and viscoelastic nanomechanical characters observed here. First, as ionic cross-links are highly sensitive to solution conditions, the instantaneous modulus, E0, exhibits orders-of-magnitude changes upon pH- and IS-governed swelling, distinctive from the rubber elasticity prediction based on permanent covalent cross-links. Second, ionic cross-links can break and self-re-form, and this mechanism dominates force relaxation of PAH/PAA under a constant indentation depth. In most states, the degree of relaxation is >90%, independent of ionic cross-link density. The importance of fixed charges is highlighted by the unexpectedly more elastic nature of the network despite low ionic cross-link density at pH 2.0, IS 0.01 M. Here, the complex is a net charged, loosely cross-linked, where the degree of relaxation is attenuated to ≈50% due to increased elastic contribution arising from fixed charge-induced Donnan osmotic pressure. In addition, this study develops a new method for quantifying the thickness of highly swollen polymer hydrogel films. It also underscores important technical considerations when performing nanomechanical tests on highly rate-dependent polymer hydrogel networks. These results provide new insights into the nanomechanical characters of ionic polyelectrolyte complexes, and lay the ground for further

  15. The Synergy of Double Cross-linking Agents on the Properties of Styrene Butadiene Rubber Foams

    PubMed Central

    Shao, Liang; Ji, Zhan-You; Ma, Jian-Zhong; Xue, Chao-Hua; Ma, Zhong-Lei; Zhang, Jing

    2016-01-01

    Sulfur (S) cross-linking styrene butadiene rubber (SBR) foams show high shrinkage due to the cure reversion, leading to reduced yield and increased processing cost. In this paper, double cross-linking system by S and dicumyl peroxide (DCP) was used to decrease the shrinkage of SBR foams. Most importantly, the synergy of double cross-linking agents was reported for the first time to our knowledge. The cell size and its distribution of SBR foams were investigated by FESEM images, which show the effect of DCP content on the cell structure of the SBR foams. The relationships between shrinkage and crystalline of SBR foams were analyzed by the synergy of double cross-linking agents, which were demonstrated by FTIR, Raman spectra, XRD, DSC and TGA. When the DCP content was 0.6 phr, the SBR foams exhibit excellent physical and mechanical properties such as low density (0.223 g/cm3), reduced shrinkage (2.25%) and compression set (10.96%), as well as elevated elongation at break (1.78 × 103%) and tear strength (54.63 N/mm). The results show that these properties are related to the double cross-linking system of SBR foams. Moreover, the double cross-linking SBR foams present high electromagnetic interference (EMI) shielding properties compared with the S cross-linking SBR foams. PMID:27841307

  16. The Synergy of Double Cross-linking Agents on the Properties of Styrene Butadiene Rubber Foams

    NASA Astrophysics Data System (ADS)

    Shao, Liang; Ji, Zhan-You; Ma, Jian-Zhong; Xue, Chao-Hua; Ma, Zhong-Lei; Zhang, Jing

    2016-11-01

    Sulfur (S) cross-linking styrene butadiene rubber (SBR) foams show high shrinkage due to the cure reversion, leading to reduced yield and increased processing cost. In this paper, double cross-linking system by S and dicumyl peroxide (DCP) was used to decrease the shrinkage of SBR foams. Most importantly, the synergy of double cross-linking agents was reported for the first time to our knowledge. The cell size and its distribution of SBR foams were investigated by FESEM images, which show the effect of DCP content on the cell structure of the SBR foams. The relationships between shrinkage and crystalline of SBR foams were analyzed by the synergy of double cross-linking agents, which were demonstrated by FTIR, Raman spectra, XRD, DSC and TGA. When the DCP content was 0.6 phr, the SBR foams exhibit excellent physical and mechanical properties such as low density (0.223 g/cm3), reduced shrinkage (2.25%) and compression set (10.96%), as well as elevated elongation at break (1.78 × 103%) and tear strength (54.63 N/mm). The results show that these properties are related to the double cross-linking system of SBR foams. Moreover, the double cross-linking SBR foams present high electromagnetic interference (EMI) shielding properties compared with the S cross-linking SBR foams.

  17. Isocyanate Cross-Linked Silica: Structurally Strong Aerogels

    NASA Technical Reports Server (NTRS)

    Leventis, Nicholas; Sotiriou-Leventis, Chariklia; Zhang, Guo-Hui; Rawashdeh, Abdel-Monem M.

    2002-01-01

    Molecular-level synergism between the silica nanoparticles of pre-formed monoliths and molecular cross-linkers inverts the relative host-guest roles in glass-polymer composites, leading to new strong low-density materials. Attempts to load gels with variable amounts of polyurethane precursors such as di-ISO and diol end-capped polybutylene adipate followed by heat treatment, washing, and supercritical drying led to opaque materials, somewhat stronger than silica but still quite brittle and much inferior to the materials described above. Direct mixing of a diisocyanate and an alcohol-free sol has been attempted recently by Yim et al. Reportedly, that procedure leads to week-long gelation times and requires an at least equally long aging period. In our attempt to add various amounts of di-ISO in a base-catalyzed sol in PC, we also noticed a week-long gelation time. The resulting aerogels were translucent but no less brittle than native silica. According to more recent studies, if propylene carbonate is replaced with acetone, it leads not only to shorter processing times, but also to much stronger gels that can tolerate loads in excess of 40 kg in the arrangement presented. We attribute that behavior to the lower viscosity of acetone, that allows faster diffusion of the di-ISO solution within the pores before di-ISO has time to react with the surface of silica. Further studies are underway to vary the chemical identity of the diisocyanate, as well as the composition and density of silica.

  18. Ingested hyaluronan moisturizes dry skin.

    PubMed

    Kawada, Chinatsu; Yoshida, Takushi; Yoshida, Hideto; Matsuoka, Ryosuke; Sakamoto, Wakako; Odanaka, Wataru; Sato, Toshihide; Yamasaki, Takeshi; Kanemitsu, Tomoyuki; Masuda, Yasunobu; Urushibata, Osamu

    2014-07-11

    Hyaluronan (HA) is present in many tissues of the body and is essential to maintain moistness in the skin tissues, which contain approximately half the body's HA mass. Due to its viscosity and moisturizing effect, HA is widely distributed as a medicine, cosmetic, food, and, recently marketed in Japan as a popular dietary supplement to promote skin moisture. In a randomized, double-blind, placebo-controlled clinical study it was found that ingested HA increased skin moisture and improved treatment outcomes for patients with dry skin. HA is also reported to be absorbed by the body distributed, in part, to the skin. Ingested HA contributes to the increased synthesis of HA and promotes cell proliferation in fibroblasts. These effects show that ingestion of HA moisturizes the skin and is expected to improve the quality of life for people who suffer from dry skin. This review examines the moisturizing effects of dry skin by ingested HA and summarizes the series of mechanisms from absorption to pharmacological action.

  19. Interstrand cross-links arising from strand breaks at true abasic sites in duplex DNA.

    PubMed

    Yang, Zhiyu; Price, Nathan E; Johnson, Kevin M; Wang, Yinsheng; Gates, Kent S

    2017-06-20

    Interstrand cross-links are exceptionally bioactive DNA lesions. Endogenous generation of interstrand cross-links in genomic DNA may contribute to aging, neurodegeneration, and cancer. Abasic (Ap) sites are common lesions in genomic DNA that readily undergo spontaneous and amine-catalyzed strand cleavage reactions that generate a 2,3-didehydro-2,3-dideoxyribose sugar remnant (3'ddR5p) at the 3'-terminus of the strand break. Interestingly, this strand scission process leaves an electrophilic α,β-unsaturated aldehyde residue embedded within the resulting nicked duplex. Here we present evidence that 3'ddR5p derivatives generated by spermine-catalyzed strand cleavage at Ap sites in duplex DNA can react with adenine residues on the opposing strand to generate a complex lesion consisting of an interstrand cross-link adjacent to a strand break. The cross-link blocks DNA replication by ϕ29 DNA polymerase, a highly processive polymerase enzyme that couples synthesis with strand displacement. This suggests that 3'ddR5p-derived cross-links have the potential to block critical cellular DNA transactions that require strand separation. LC-MS/MS methods developed herein provide powerful tools for studying the occurrence and properties of these cross-links in biochemical and biological systems. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  20. Facile fabrication of core cross-linked micelles by RAFT polymerization and enzyme-mediated reaction.

    PubMed

    Wu, Yukun; Lai, Quanyong; Lai, Shuqi; Wu, Jing; Wang, Wei; Yuan, Zhi

    2014-06-01

    Polymeric micelles formed in aqueous solution by assembly of amphiphilic block copolymers have been extensively investigated due to their great potential as drug carriers. However, the stability of polymeric assembly is still one of the major challenges in delivering drugs to tissues and cells. Here, we report a facile route to fabricate core cross-linked (CCL) micelles using an enzymatic polymerization as the cross-linking method. We present synthesis of poly(ethylene glycol)-block-poly(N-isopropyl acrylamide-co-N-(4-hydroxyphenethyl) acrylamide) diblock copolymer PEG-b-P(NIPAAm-co-NHPAAm) via reversible addition-fragmentation chain transfer (RAFT) polymerization. The diblock copolymer was then self-assembled into non-cross-linked (NCL) micelles upon heating above the lower critical solution temperature (LCST), and subsequently cross-linked using horseradish peroxidase (HRP) and hydrogen peroxide (H2O2) as enzyme and oxidant. The characterization of the diblock copolymer and micelles were studied by NMR, DLS, UV-vis, and fluorescence spectroscopy. The fluorescence study reveals that the cross-linking process endows the micelles with much lower critical micelle concentration (CMC). In addition, the drug release study shows that the CCL micelles have lower release amount of doxorubicin (DOX) than the NCL micelles due to the enhanced stability of the CCL micelles by core cross-linking process. Copyright © 2014 Elsevier B.V. All rights reserved.

  1. Noninvasive Assessment of Collagen Gel Microstructure and Mechanics Using Multiphoton Microscopy

    PubMed Central

    Raub, Christopher B.; Suresh, Vinod; Krasieva, Tatiana; Lyubovitsky, Julia; Mih, Justin D.; Putnam, Andrew J.; Tromberg, Bruce J.; George, Steven C.

    2007-01-01

    Multiphoton microscopy of collagen hydrogels produces second harmonic generation (SHG) and two-photon fluorescence (TPF) images, which can be used to noninvasively study gel microstructure at depth (∼1 mm). The microstructure is also a primary determinate of the mechanical properties of the gel; thus, we hypothesized that bulk optical properties (i.e., SHG and TPF) could be used to predict bulk mechanical properties of collagen hydrogels. We utilized polymerization temperature (4–37°C) and glutaraldehyde to manipulate collagen hydrogel fiber diameter, space-filling properties, and cross-link density. Multiphoton microscopy and scanning electron microscopy reveal that as polymerization temperature decreases (37–4°C) fiber diameter and pore size increase, whereas hydrogel storage modulus (G′, from 23 ± 3 Pa to 0.28 ± 0.16 Pa, respectively, mean ± SE) and mean SHG decrease (minimal change in TPF). In contrast, glutaraldehyde significantly increases the mean TPF signal (without impacting the SHG signal) and the storage modulus (16 ± 3.5 Pa before to 138 ± 40 Pa after cross-linking, mean ± SD). We conclude that SHG and TPF can characterize differential microscopic features of the collagen hydrogel that are strongly correlated with bulk mechanical properties. Thus, optical imaging may be a useful noninvasive tool to assess tissue mechanics. PMID:17172303

  2. Changes in the myosin secondary structure and shrimp surimi gel strength induced by dense phase carbon dioxide.

    PubMed

    Guo, Minghui; Liu, Shucheng; Ismail, Marliya; Farid, Mohammed M; Ji, Hongwu; Mao, Weijie; Gao, Jing; Li, Chengyong

    2017-07-15

    Dense phase carbon dioxide (DPCD) could induce protein conformation changes. Myosin and shrimp surimi from Litopenaeus vannamei were treated with DPCD at 5-25MPa and 40-60°C for 20min. Myosin secondary structure was investigated by circular dichroism and shrimp surimi gel strength was determined using textural analysis to develop correlations between them. DPCD had a greater effect on secondary structure and gel strength than heating. With increasing pressure and temperature, the α-helix content of DPCD-treated myosin decreased, while the β-sheet, β-turn and random coil contents increased, and the shrimp surimi gel strength increased. The α-helix content was negatively correlated with gel strength, while the β-sheet, β-turn and random coil contents were positively correlated with gel strength. Therefore, when DPCD induced myosin to form a gel, the α-helix of myosin was unfolded and gradually converted to a β-sheet. Such transformations led to protein-protein interactions and cross-linking, which formed a three-dimensional network to enhance the gel strength. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. Cross-linked polyethylene does not reduce wear in total knee arthroplasty.

    PubMed

    Lasurt-Bachs, S; Torner, P; Maculé, F; Prats, E; Menéndez-García, F; Ríos-Guillermo, J; Torrents, A

    To compare two different types of inserts: Ultra-high molecular weight polyethylene (UHMWPE) and cross-linked polyethylene with a quantitative and qualitative study of polyethylene wear particles in synovial fluid 3 years after total knee arthroplasty. A prospective, randomized, controlled cohort study with blinded evaluation was carried out on 25 patients undergoing staged bilateral total knee replacement, 6 months apart. Knee arthrocentesis was performed on 12 patients 3 years after surgery, and the polyethylene particles were analyzed. No significant differences were found in the number of particles generated by the two different types of inserts at 3 years from total knee arthroplasty (3,000×: x¯ cross-linked=849.7; x¯ UHMWPE=796.9; P=.63; 20,000×: x¯ cross-linked=66.3; x¯ UHMWPE=73.1; P=.76). Likewise, no differences in the probability of finding elongated (χ 2 =0.19; P=.66) or rounded (χ 2 =1.44; P=.23) particles in both types of inserts were observed. However, the probability of finding fibrillar particles is 3.08 times greater in UHMWPE. Cross-linked polyethylene does not significantly reduce the generation of polyethylene particles in patients with total knee arthroplasty, 3 years after the surgical procedure. Copyright © 2018 SECOT. Publicado por Elsevier España, S.L.U. All rights reserved.

  4. Collagen and elastin cross-linking is altered during aberrant late lung development associated with hyperoxia.

    PubMed

    Mižíková, Ivana; Ruiz-Camp, Jordi; Steenbock, Heiko; Madurga, Alicia; Vadász, István; Herold, Susanne; Mayer, Konstantin; Seeger, Werner; Brinckmann, Jürgen; Morty, Rory E

    2015-06-01

    Maturation of the lung extracellular matrix (ECM) plays an important role in the formation of alveolar gas exchange units. A key step in ECM maturation is cross-linking of collagen and elastin, which imparts stability and functionality to the ECM. During aberrant late lung development in bronchopulmonary dysplasia (BPD) patients and animal models of BPD, alveolarization is blocked, and the function of ECM cross-linking enzymes is deregulated, suggesting that perturbed ECM cross-linking may impact alveolarization. In a hyperoxia (85% O2)-based mouse model of BPD, blunted alveolarization was accompanied by alterations to lung collagen and elastin levels and cross-linking. Total collagen levels were increased (by 63%). The abundance of dihydroxylysinonorleucine collagen cross-links and the dihydroxylysinonorleucine-to-hydroxylysinonorleucine ratio were increased by 11 and 18%, respectively, suggestive of a profibrotic state. In contrast, insoluble elastin levels and the abundance of the elastin cross-links desmosine and isodesmosine in insoluble elastin were decreased by 35, 30, and 21%, respectively. The lung collagen-to-elastin ratio was threefold increased. Treatment of hyperoxia-exposed newborn mice with the lysyl oxidase inhibitor β-aminopropionitrile partially restored normal collagen levels, normalized the dihydroxylysinonorleucine-to-hydroxylysinonorleucine ratio, partially normalized desmosine and isodesmosine cross-links in insoluble elastin, and partially restored elastin foci structure in the developing septa. However, β-aminopropionitrile administration concomitant with hyperoxia exposure did not improve alveolarization, evident from unchanged alveolar surface area and alveoli number, and worsened septal thickening (increased by 12%). These data demonstrate that collagen and elastin cross-linking are perturbed during the arrested alveolarization of developing mouse lungs exposed to hyperoxia. Copyright © 2015 the American Physiological Society.

  5. Physicochemical properties and micro-structural characteristics in starch from kudzu root as affected by cross-linking.

    PubMed

    Chen, Boru; Dang, Leping; Zhang, Xiao; Fang, Wenzhi; Hou, Mengna; Liu, Tiankuo; Wang, Zhanzhong

    2017-03-15

    Kudzu starch was cross-linked with sodium trimetaphosphate (STMP) at different temperatures, time and of STMP concentrations in this work. The cross-linked starches (CLSs) were fractionated further into cross-linked amylose and amylopectin in order to compare the effect of cross-linking on the microstructure. According to scanning electron microscope (SEM), CLSs displayed the resemble appearance of spherical and polygonal shapes like NS. X-ray diffraction (XRD) revealed that amylose of native starch (A), NS and CLS displayed a combination of A-type and B-type structure, while that was not found in amylose of cross-linked starch (CLA). The deconvoluted fourier transform infrared (FT-IR) indicated that crystal structure of kudzu starch was losing with the proceeding of cross-linking reaction. The CLSs exhibited a higher retrogradation and freeze-thaw stability than NS. This was accompanied by a significant decrease in sedimentation, transparency, swelling power and solubility. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. CD44 mediated hyaluronan adhesion of Toxoplasma gondii-infected leukocytes.

    PubMed

    Hayashi, Takeshi; Unno, Akihiro; Baba, Minami; Ohno, Tamio; Kitoh, Katsuya; Takashima, Yasuhiro

    2014-04-01

    Toxoplasma gondii is an obligate intracellular apicomplexan parasite that infects humans and animals. Ingested parasites cross the intestinal epithelium, invade leukocytes and are then disseminated to peripheral organs. However, the mechanism of extravasation of the infected leukocytes remains poorly understood. In this study, we demonstrate that T. gondii-invaded human and mouse leukocytes express higher level of CD44, a ligand of hyaluronan (HA), and its expression on myeloid and non-myeloid leukocytes causes T. gondii-invaded human and mouse leukocyte to adhere to HA more effectively than non-invaded leukocytes. The specific adherence of parasite-invaded leukocytes was inhibited by anti CD44 antibody. Leukocytes of CD44 knockout mice did not show parasite-invaded leukocyte specific adhesion. Our results indicate that parasite-invaded leukocytes, regardless of whether myeloid or not, gain higher ability to adhere to HA than non-invaded leukocytes, via upregulation of CD44 expression and/or selective invasion to CD44 highly expressing cells. The difference in ability to adhere to HA between parasite-invaded cells and non-invaded neighboring cells might facilitate effective delivery of parasite-invaded leukocytes to the HA-producing endothelial cell surface and/or HA-rich extra cellular matrix. © 2013.

  7. Role of solvent-mediated carbodiimide cross-linking in fabrication of electrospun gelatin nanofibrous membranes as ophthalmic biomaterials.

    PubMed

    Chou, Shih-Feng; Luo, Li-Jyuan; Lai, Jui-Yang; Ma, David Hui-Kang

    2017-02-01

    Due to their ability to mimic the structure of extracellular matrix, electrospun gelatin nanofibers are promising cell scaffolding materials for tissue engineering applications. However, the hydrophilic gelatin molecules usually need stabilization before use in aqueous physiological environment. Considering that biomaterials cross-linked via film immersion technique may have a more homogeneous cross-linked structure than vapor phase cross-linking, this work aims to investigate the chemical modification of electrospun gelatin nanofibrous membranes by liquid phase carbodiimide in the presence of ethanol/water co-solvents with varying ethanol concentrations ranging from 80 to 99.5vol%. The results of characterization showed that increasing water content in the binary reaction solvent system increases the extent of cross-linking of gelatin nanofibers, but simultaneously promotes the effect of biopolymer swelling and distortion in fiber mat structure. As compared to non-cross-linked counterparts, carbodiimide treated gelatin nanofibrous mats exhibited better thermal and biological stability where the shrinkage temperature and resistance to enzymatic degradation varied in response to ethanol/water solvent composition-mediated generation of cross-links. Irrespective of their cross-linking density, all studied membrane samples did not induce any responses in ocular epithelial cell cultures derived from cornea, lens, and retina. Unlike many other cross-linking agents and/or methods (e.g., excessive vapor phase cross-linking) that may pose a risk of toxicity, our study demonstrated that these nanofibrous materials are well tolerated by anterior segment tissues. These findings also indicate the safety of using ethanol/water co-solvents for chemical cross-linking of gelatin to engineer nanofibrous materials with negligible biological effects. In summary, the present results suggest the importance of solvent-mediated carbodiimide cross-linking in modulating structure

  8. Collagen cross-linking: insights on the evolution of metazoan extracellular matrix.

    PubMed

    Rodriguez-Pascual, Fernando; Slatter, David Anthony

    2016-11-23

    Collagens constitute a large family of extracellular matrix (ECM) proteins that play a fundamental role in supporting the structure of various tissues in multicellular animals. The mechanical strength of fibrillar collagens is highly dependent on the formation of covalent cross-links between individual fibrils, a process initiated by the enzymatic action of members of the lysyl oxidase (LOX) family. Fibrillar collagens are present in a wide variety of animals, therefore often being associated with metazoan evolution, where the emergence of an ancestral collagen chain has been proposed to lead to the formation of different clades. While LOX-generated collagen cross-linking metabolites have been detected in different metazoan families, there is limited information about when and how collagen acquired this particular modification. By analyzing telopeptide and helical sequences, we identified highly conserved, potential cross-linking sites throughout the metazoan tree of life. Based on this analysis, we propose that they have importantly contributed to the formation and further expansion of fibrillar collagens.

  9. Tuning the density profile of surface-grafted hyaluronan and the effect of counter-ions.

    PubMed

    Berts, Ida; Fragneto, Giovanna; Hilborn, Jöns; Rennie, Adrian R

    2013-07-01

    The present paper investigates the structure and composition of grafted sodium hyaluronan at a solid-liquid interface using neutron reflection. The solvated polymer at the surface could be described with a density profile that decays exponentially towards the bulk solution. The density profile of the polymer varied depending on the deposition protocol. A single-stage deposition resulted in denser polymer layers, while layers created with a two-stage deposition process were more diffuse and had an overall lower density. Despite the diffuse density profile, two-stage deposition leads to a higher surface excess. Addition of calcium ions causes a strong collapse of the sodium hyaluronan chains, increasing the polymer density near the surface. This effect is more pronounced on the sample prepared by two-stage deposition due to the initial less dense profile. This study provides an understanding at a molecular level of how surface functionalization alters the structure and how surface layers respond to changes in calcium ions in the solvent.

  10. Drug-Triggered and Cross-Linked Self-Assembling Nanofibrous Hydrogels

    PubMed Central

    Kumar, Vivek A.; Shi, Siyu; Wang, Benjamin K.; Li, I-Che; Jalan, Abhishek A.; Sarkar, Biplab; Wickremasinghe, Navindee C.; Hartgerink, Jeffrey D.

    2015-01-01

    Self-assembly of multidomain peptides (MDP) can be tailored to carry payloads that modulate the extracellular environment. Controlled release of growth factors, cytokines, and small-molecule drugs allows for unique control of in vitro and in vivo responses. In this study, we demonstrate this process of ionic cross-linking of peptides using multivalent drugs to create hydrogels for sustained long-term delivery of drugs. Using phosphate, heparin, clodronate, trypan, and suramin, we demonstrate the utility of this strategy. Although all multivalent anions result in good hydrogel formation, demonstrating the generality of this approach, suramin led to the formation of the best hydrogels per unit concentration and was studied in greater detail. Suramin ionically cross-linked MDP into a fibrous meshwork as determined by scanning and transmission electron microscopy. We measured material storage and loss modulus using rheometry and showed a distinct increase in G′ and G″ as a function of suramin concentration. Release of suramin from scaffolds was determined using UV spectroscopy and showed prolonged release over a 30 day period. Suramin bioavailability and function were demonstrated by attenuated M1 polarization of THP-1 cells compared to positive control. Overall, this design strategy has allowed for the development of a novel class of polymeric delivery vehicles with generally long-term release and, in the case of suramin, cross-linked hydrogels that can modulate cellular phenotype. PMID:25831137

  11. Cross-modal links among vision, audition, and touch in complex environments.

    PubMed

    Ferris, Thomas K; Sarter, Nadine B

    2008-02-01

    This study sought to determine whether performance effects of cross-modal spatial links that were observed in earlier laboratory studies scale to more complex environments and need to be considered in multimodal interface design. It also revisits the unresolved issue of cross-modal cuing asymmetries. Previous laboratory studies employing simple cues, tasks, and/or targets have demonstrated that the efficiency of processing visual, auditory, and tactile stimuli is affected by the modality, lateralization, and timing of surrounding cues. Very few studies have investigated these cross-modal constraints in the context of more complex environments to determine whether they scale and how complexity affects the nature of cross-modal cuing asymmetries. Amicroworld simulation of battlefield operations with a complex task set and meaningful visual, auditory, and tactile stimuli was used to investigate cuing effects for all cross-modal pairings. Significant asymmetric performance effects of cross-modal spatial links were observed. Auditory cues shortened response latencies for collocated visual targets but visual cues did not do the same for collocated auditory targets. Responses to contralateral (rather than ipsilateral) targets were faster for tactually cued auditory targets and each visual-tactile cue-target combination, suggesting an inhibition-of-return effect. The spatial relationships between multimodal cues and targets significantly affect target response times in complex environments. The performance effects of cross-modal links and the observed cross-modal cuing asymmetries need to be examined in more detail and considered in future interface design. The findings from this study have implications for the design of multimodal and adaptive interfaces and for supporting attention management in complex, data-rich domains.

  12. MDA-MB-231 breast cancer cell viability, motility and matrix adhesion are regulated by a complex interplay of heparan sulfate, chondroitin-/dermatan sulfate and hyaluronan biosynthesis.

    PubMed

    Viola, Manuela; Brüggemann, Kathrin; Karousou, Evgenia; Caon, Ilaria; Caravà, Elena; Vigetti, Davide; Greve, Burkhard; Stock, Christian; De Luca, Giancarlo; Passi, Alberto; Götte, Martin

    2017-06-01

    Proteoglycans and glycosaminoglycans modulate numerous cellular processes relevant to tumour progression, including cell proliferation, cell-matrix interactions, cell motility and invasive growth. Among the glycosaminoglycans with a well-documented role in tumour progression are heparan sulphate, chondroitin/dermatan sulphate and hyaluronic acid/hyaluronan. While the mode of biosynthesis differs for sulphated glycosaminoglycans, which are synthesised in the ER and Golgi compartments, and hyaluronan, which is synthesized at the plasma membrane, these polysaccharides partially compete for common substrates. In this study, we employed a siRNA knockdown approach for heparan sulphate (EXT1) and heparan/chondroitin/dermatan sulphate-biosynthetic enzymes (β4GalT7) in the aggressive human breast cancer cell line MDA-MB-231 to study the impact on cell behaviour and hyaluronan biosynthesis. Knockdown of β4GalT7 expression resulted in a decrease in cell viability, motility and adhesion to fibronectin, while these parameters were unchanged in EXT1-silenced cells. Importantly, these changes were associated with a decreased expression of syndecan-1, decreased signalling response to HGF and an increase in the synthesis of hyaluronan, due to an upregulation of the hyaluronan synthases HAS2 and HAS3. Interestingly, EXT1-depleted cells showed a downregulation of the UDP-sugar transporter SLC35D1, whereas SLC35D2 was downregulated in β4GalT7-depleted cells, indicating an intricate regulatory network that connects all glycosaminoglycans synthesis. The results of our in vitro study suggest that a modulation of breast cancer cell behaviour via interference with heparan sulphate biosynthesis may result in a compensatory upregulation of hyaluronan biosynthesis. These findings have important implications for the development of glycosaminoglycan-targeted therapeutic approaches for malignant diseases.

  13. Effect of growth factors on hyaluronan production by canine vocal fold fibroblasts.

    PubMed

    Hirano, Shigeru; Bless, Diane M; Heisey, Dennis; Ford, Charles N

    2003-07-01

    Hyaluronan (HYA) is considered to be a crucial factor in scarless wound healing and in maintaining tissue viscosity of the vocal fold lamina propria. In this study focusing on the effects of growth factors, we examined how HYA is produced and controlled in canine cultured vocal fold fibroblasts. Fibroblasts were taken from the lamina propria of the vocal folds of 8 dogs and cultured with and without growth factors. The production of HYA in the supernatant culture was quantitatively examined by enzyme-linked immunosorbent assay. Hepatocyte growth factor, epidermal growth factor, basic fibroblast growth factor, and transforming growth factor beta1 all stimulated HYA synthesis from vocal fold fibroblasts. These effects differed with the concentration of growth factors and the incubation period. We also examined how frequently the growth factors had to be administered in order to maintain appropriate levels of HYA. A single administration was sufficient to maintain appropriate HYA levels for at least 7 days. The present studies have demonstrated positive effects of growth factors in stimulating HYA production. Further in vivo study is needed to clarify the usefulness of these growth factors in the management of vocal fold scarring.

  14. The Non-catalytic B Subunit of Coagulation Factor XIII Accelerates Fibrin Cross-linking*

    PubMed Central

    Souri, Masayoshi; Osaki, Tsukasa; Ichinose, Akitada

    2015-01-01

    Covalent cross-linking of fibrin chains is required for stable blood clot formation, which is catalyzed by coagulation factor XIII (FXIII), a proenzyme of plasma transglutaminase consisting of catalytic A (FXIII-A) and non-catalytic B subunits (FXIII-B). Herein, we demonstrate that FXIII-B accelerates fibrin cross-linking. Depletion of FXIII-B from normal plasma supplemented with a physiological level of recombinant FXIII-A resulted in delayed fibrin cross-linking, reduced incorporation of FXIII-A into fibrin clots, and impaired activation peptide cleavage by thrombin; the addition of recombinant FXIII-B restored normal fibrin cross-linking, FXIII-A incorporation into fibrin clots, and activation peptide cleavage by thrombin. Immunoprecipitation with an anti-fibrinogen antibody revealed an interaction between the FXIII heterotetramer and fibrinogen mediated by FXIII-B and not FXIII-A. FXIII-B probably binds the γ-chain of fibrinogen with its D-domain, which is near the fibrin polymerization pockets, and dissociates from fibrin during or after cross-linking between γ-chains. Thus, FXIII-B plays important roles in the formation of a ternary complex between proenzyme FXIII, prosubstrate fibrinogen, and activator thrombin. Accordingly, congenital or acquired FXIII-B deficiency may result in increased bleeding tendency through impaired fibrin stabilization due to decreased FXIII-A activation by thrombin and secondary FXIII-A deficiency arising from enhanced circulatory clearance. PMID:25809477

  15. Two-photon induced collagen cross-linking in bioartificial cardiac tissue

    NASA Astrophysics Data System (ADS)

    Kuetemeyer, Kai; Kensah, George; Heidrich, Marko; Meyer, Heiko; Martin, Ulrich; Gruh, Ina; Heisterkamp, Alexander

    2011-08-01

    Cardiac tissue engineering is a promising strategy for regenerative therapies to overcome the shortage of donor organs for transplantation. Besides contractile function, the stiffness of tissue engineered constructs is crucial to generate transplantable tissue surrogates with sufficient mechanical stability to withstand the high pressure present in the heart. Although several collagen cross-linking techniques have proven to be efficient in stabilizing biomaterials, they cannot be applied to cardiac tissue engineering, as cell death occurs in the treated area. Here, we present a novel method using femtosecond (fs) laser pulses to increase the stiffness of collagen-based tissue constructs without impairing cell viability. Raster scanning of the fs laser beam over riboflavin-treated tissue induced collagen cross-linking by two-photon photosensitized singlet oxygen production. One day post-irradiation, stress-strain measurements revealed increased tissue stiffness by around 40% being dependent on the fibroblast content in the tissue. At the same time, cells remained viable and fully functional as demonstrated by fluorescence imaging of cardiomyocyte mitochondrial activity and preservation of active contraction force. Our results indicate that two-photon induced collagen cross-linking has great potential for studying and improving artificially engineered tissue for regenerative therapies.

  16. Collagen cross-link excretion during space flight and bed rest

    NASA Technical Reports Server (NTRS)

    Smith, S. M.; Nillen, J. L.; Leblanc, A.; Lipton, A.; Demers, L. M.; Lane, H. W.; Leach, C. S.; LeBlanc, A. (Principal Investigator)

    1998-01-01

    Extended exposure to weightlessness results in bone loss. However, little information exists as to the precise nature or time course of this bone loss. Bone resorption results in the release of collagen breakdown products, including N-telopeptide and the pyridinium (PYD) cross-links, pyridinoline and deoxypyridinoline. Urinary pyridinoline and deoxypyridinoline are known to increase during bed rest. We assessed excretion of PYD cross-links and N-telopeptide before, during, and after long (28-day, 59-day, and 84-day) Skylab missions, as well as during short (14-day) and long (119-day) bed-rest studies. During space flight, the urinary cross-link excretion level was twice those observed before flight. Urinary excretion levels of the collagen breakdown products were also 40-50% higher, during short and long bed rest, than before. These results clearly show that the changes in bone metabolism associated with space flight involve increased resorption. The rate of response (i.e. within days to weeks) suggests that alterations in bone metabolism are an early effect of weightlessness. These studies are important for a better understanding of bone metabolism in space crews and in those who are bedridden.

  17. Gel performance in rheology and profile control under low-frequency vibration: coupling application of physical and chemical EOR techniques.

    PubMed

    Zheng, Li Ming; Pu, Chun Sheng; Liu, Jing; Ma, Bo; Khan, Nasir

    2017-01-01

    Flowing gel plugging and low-frequency vibration oil extraction technology have been widely applied in low-permeability formation. High probability of overlapping in action spheres of two technologies might lead to poor operating efficiency during gel injection. Study on flowing gel rheological properties under low-frequency vibration was essential, which was carried out indoor with viscosity measurement. Potential dynamic mechanisms were analyzed for the rheological variation. Under low-frequency vibration, gel rheological properties were found to be obviously influenced, with vibration delaying gel cross-linking in induction period, causing a two-stage gel viscosity change in acceleration period, and decreasing gel strength in stable period. Surface of gel system under vibration presented different fluctuating phenomenon from initial harmonic vibrating to heterogeneous fluctuating (droplet separation might appear) to final harmonic vibrating again. Dynamic displacement in unconsolidated sand pack revealed that low-frequency vibration during gel injection might be a measure to achieve deep profile control, with the gel injection depth increased by 65.8 % compared with the vibration-free sample. At last, suggestions for field test were given in the paper to achieve lower injection friction and better gel plugging efficiency.

  18. Occurrence and structural characterization of versican-like proteoglycan in human vitreous.

    PubMed

    Theocharis, Achilleas D; Papageorgakopoulou, Nickoletta; Feretis, Elias; Theocharis, Dimitrios A

    2002-12-01

    Human vitreous gel is a special type of extracellular matrix, in which interpenetrating networks of collagen fibrils and hyaluronan are found. In this study, we report that apart from significant amounts of collagen, hyaluronan and sialylated glycoproteins, it was found that the human vitreous gel also contained low amounts of versican-like proteoglycan. The concentration of versican-like proteoglycan in the whole vitreous is 0.06 mg protein/ml of vitreous gel and represents a small percentage (about 5%) of the total protein content. The versican-like proteoglycan has a molecular mass of 380 kDa, as estimated by gel chromatography. Its core protein is substituted by chondroitin sulphate side chains (average molecular weight 37 kDa), in which 6-sulphated disaccharides predominated. According to the physicochemical data, the number of chondroitin sulphate chains is likely to be 5-7 per molecule. These proteoglycan monomers form large aggregates with endogenous hyaluronan. Versican, which is able to bind lectins via its C-terminal region, may bridge or interconnect various constituents of the extracellular matrix via its terminal domains in order to stabilize large supramolecular complexes at the vitreous, contributing towards the integrity and specific properties of the tissue.

  19. Multi-stimuli-responsive organometallic gels based on ferrocene-linked poly(aryl ether) dendrons: reversible redox switching and Pb2+-ion sensing.

    PubMed

    Lakshmi, Neelakandan Vidhya; Mandal, Dipendu; Ghosh, Sundargopal; Prasad, Edamana

    2014-07-14

    We describe the design, synthesis, and "stimuli-responsive" study of ferrocene-linked Fréchet-type [poly(aryl ether)]-dendron-based organometallic gels, in which the ferrocene moiety is attached to the dendron framework through an acyl hydrazone linkage. The low-molecular-weight gelators (LMWGs) form robust gels in both polar and non-polar solvent/solvent mixtures. The organometallic gels undergo stimuli-responsive behavior through 1) thermal, 2) chemical, and 3) electrochemical methods. Among them, conditions 1 and 3 lead to seamlessly reversible with repeated cycles of identical efficiency. Results indicate that the flexible nature of the poly(aryl ether) dendron framework plays a key role in retaining the reversible electrochemical behavior of ferrocene moiety in the LMWGs. Further, the organometallic gelators have exhibited unique selectivity towards Pb(2+) ions (detection limit ≈10(-8)  M). The metal ion-sensing results in a gel-sol phase transition associated with a color change visible to the naked eye. Most importantly, decomplexing the metal ion from the system leads to the regeneration of the initial gel morphology, indicating the restoring ability of the organometallic gel. The metal-ligand binding nature has been analyzed by using (1)H NMR spectroscopy, mass spectrometry, and DFT calculations. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Thermal conductivity of cross-linked polyethylene from molecular dynamics simulation

    NASA Astrophysics Data System (ADS)

    Xiong, Xue; Yang, Ming; Liu, Changlin; Li, Xiaobo; Tang, Dawei

    2017-07-01

    The thermal conductivity of cross-linked bulk polyethylene is studied using molecular dynamics simulation. The atomic structure of the cross-linked polyethylene (PEX) is generated through simulated bond formation using LAMMPS. The thermal conductivity of PEX is studied with different degrees of crosslinking, chain length, and tensile strain. Generally, the thermal conductivity increases with the increasing degree of crosslinking. When the length of the primitive chain increases, the thermal conductivity increases linearly. When the polymer is stretched along one direction, the thermal conductivity increases in the stretched direction and decreases in the direction perpendicular to it. However, the thermal conductivity varies slightly when the polymer is stretched in three directions simultaneously.

  1. Large-Volume Crystalloid Fluid Is Associated with Increased Hyaluronan Shedding and Inflammation in a Canine Hemorrhagic Shock Model.

    PubMed

    Smart, Lisa; Boyd, C J; Claus, M A; Bosio, E; Hosgood, G; Raisis, A

    2018-05-04

    Shedding of the endothelial glycocalyx precedes leukocyte activation and adherence in acute inflammation. Rapid administration of crystalloid or colloid fluids for treating hemorrhagic shock may cause endothelial glycocalyx shedding, thereby increasing inflammation. This study aimed to compare the effect of different fluid treatments in a canine shock model on glycocalyx biomarker, hyaluronan, and inflammatory biomarkers. Greyhound dogs under general anesthesia subject to hemorrhage for 60 min were given 20 mL kg -1 of either fresh whole blood (FWB), hydroxyethyl starch (HES) 130/0.4, 4% succinylated gelatin (GELO), or 80 mL kg -1 of isotonic crystalloid (CRYST) over 20 min (n = 6 per group). Plasma biomarkers hyaluronan, interleukin (IL) 6, 8, 10, tumor necrosis factor-α, monocyte chemoattractant protein-1, keratinocyte chemokine-like, and atrial natriuretic peptide were measured at baseline, end of hemorrhage (Shock), end of fluid administration (T20), and then 40 (T60), 100 (T120), and 160 (T180) minutes later. Biomarker concentrations were compared between groups using the Kruskal-Wallis test or Fisher's exact test (measurable versus unmeasurable) (significance set at P < 0.05). Hyaluronan concentration peaked early in the CRYST group at T20, compared to HES (P = 0.005) and GELO (P = 0.018), and later in the GELO group at T60, compared to FWB (P < 0.001). The CRYST group had significantly more samples with measurable IL6 at T180 (P = 0.015), compared to GELO, and IL10 at T60, T120, and T180 (all P = 0.015), compared to FWB. There were no significant differences in other biomarker concentrations. In conclusion, rapid large-volume crystalloid administered for hemorrhagic shock was associated with increased hyaluronan and a greater inflammatory response.

  2. Dehydration of an ethanol/water azeotrope through alginate-DNA membranes cross-linked with metal ions by pervaporation.

    PubMed

    Uragami, Tadashi; Banno, Masashi; Miyata, Takashi

    2015-12-10

    To obtain high dehydration membranes for an ethanol/water azeotrope, dried blend membranes prepared from mixtures of sodium alginate (Alg-Na) and sodium deoxyribonucleate (DNA-Na) were cross-linked by immersing in a methanol solution of CaCl2 or MaCl2. In the dehydration of an ethanol/water azeotropic mixture by pervaporation, the effects of immersion time in methanol solution of CaCl2 or MaCl2 on the permeation rate and water/ethanol selectivity through Alg-DNA/Ca(2+) and Alg-DNA/Mg(2+) cross-linked membranes were investigated. Alg-DNA/Mg(2+) cross-linked membrane immersed for 12h in methanol solution of MaCl2 exhibited the highest water/ethanol selectivity. This results from depressed swelling of the membranes by formation of a cross-linked structure. However, excess immersion in solution containing cross-linker led to an increase in the hydrophobicity of cross-linked membrane. Therefore, the water/ethanol selectivity of Alg-DNA/Mg(2+) cross-linked membranes with an excess immersion in cross-linking solution was lowered. The relationship between the structure of Alg-DNA/Ca(2+) and Alg-DNA/Mg(2+) cross-linked membranes and their permeation and separation characteristics during pervaporation of an ethanol/water azeotropic mixture is discussed in detail. Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. Towards a fully synthetic substitute of alginate: optimization of a thermal gelation/chemical cross-linking scheme ("tandem" gelation) for the production of beads and liquid-core capsules.

    PubMed

    Cellesi, F; Weber, W; Fussenegger, M; Hubbell, J A; Tirelli, N

    2004-12-20

    Fully synthetic polymers were used for the preparation of hydrogel beads and capsules, in a processing scheme that, originally designed for calcium alginate, was adapted to a "tandem" process, that is the combination a physical gelation with a chemical cross-linking. The polymers feature a Tetronic backbone (tetra armed Pluronics), which exhibits a reverse thermal gelation in water solutions within a physiological range of temperatures and pHs. The polymers bear terminal reactive groups that allow for a mild, but effective chemical cross-linking. Given an appropriate temperature jump, the thermal gelation provides a hardening kinetics similar to that of alginate. With slower kinetics, the chemical cross-linking then develops an irreversible and elastic gel structure, and determines its transport properties. In the present article this process has been optimized for the production of monodisperse, high elastic, hydrogel microbeads, and liquid-core microcapsules. We also show the feasibility of the use of liquid-core microcapsules in cell encapsulation. In preliminary experiments, CHO cells have been successfully encapsulated preserving their viability during the process and after incubation. The advantages of this process are mainly in the use of synthetic polymers, which provide great flexibility in the molecular design. This, in principle, allows for a precise tailoring of mechanical and transport properties and of bioactivity of the hydrogels, and also for a precise control in material purification.

  4. A contact photo-cross-linking investigation of the active site of the 8-17 deoxyribozyme.

    PubMed

    Liu, Yong; Sen, Dipankar

    2008-09-12

    The small RNA-cleaving 8-17 deoxyribozyme (DNAzyme) has been the subject of extensive mechanistic and structural investigation, including a number of recent single-molecule studies of its global folding. Little detailed insight exists, however, into this DNAzyme's active site; for instance, the identity of specific nucleotides that are proximal to or in contact with the scissile site in the substrate. Here, we report a systematic replacement of a number of bases within the magnesium-folded DNAzyme-substrate complex with thio- and halogen-substituted base analogues, which were then photochemically activated to generate contact cross-links within the complex. Mapping of the cross-links revealed a striking pattern of DNAzyme-substrate cross-links but an absence of significant intra-DNAzyme cross-links. Notably, the two nucleotides directly flanking the scissile phosphodiester cross-linked strongly with functionally important elements within the DNAzyme, the thymine of a G.T wobble base pair, a WCGR bulge loop, and a terminal AGC loop. Mutation of the wobble base pair to a G-C pair led to a significant folding instability of the DNAzyme-substrate complex. The cross-linking patterns obtained were used to generate a model for the DNAzyme's active site that had the substrate's scissile phosphodiester sandwiched between the DNAzyme's wobble thymine and its AGC and WCGR loops.

  5. The counterbend phenomenon in flagellar axonemes and cross-linked filament bundles

    PubMed Central

    Gadêlha, Hermes; Gaffney, Eamonn A.; Goriely, Alain

    2013-01-01

    Recent observations of flagellar counterbend in sea urchin sperm show that the mechanical induction of curvature in one part of a passive flagellum induces a compensatory countercurvature elsewhere. This apparent paradoxical effect cannot be explained using the standard elastic rod theory of Euler and Bernoulli, or even the more general Cosserat theory of rods. Here, we develop a geometrically exact mechanical model to describe the statics of microtubule bundles that is capable of predicting the curvature reversal events observed in eukaryotic flagella. This is achieved by allowing the interaction of deformations in different material directions, by accounting not only for structural bending, but also for the elastic forces originating from the internal cross-linking mechanics. Large-amplitude static configurations can be described analytically, and an excellent match between the model and the observed counterbend deformation was found. This allowed a simultaneous estimation of multiple sperm flagellum material parameters, namely the cross-linking sliding resistance, the bending stiffness, and the sperm head junction compliance ratio. We further show that small variations on the empirical conditions may induce discrepancies for the evaluation of the flagellar material quantities, so that caution is required when interpreting experiments. Finally, our analysis demonstrates that the counterbend emerges as a fundamental property of sliding resistance in cross-linked filamentous polymer bundles, which also suggests that cross-linking proteins may contribute to the regulation of the flagellar waveform in swimming sperm via counterbend mechanics. PMID:23824293

  6. Enzymic cross-linkage of monomeric extensin precursors in vitro. [Lycopersicon esculentum

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Everdeen, D.S.; Kiefer, S.; Willard, J.J.

    Rapidly growing tomato (Lycopersicon esculentum) cell suspension cultures contain transiently high levels of cell surface, salt-elutable, monomeric precursors to the covalently cross-linked extensin network of the primary cell wall. Thus, the authors purified a highly soluble monomeric extensin substrate from rapidly growing cells, and devised a soluble in vitro cross-linking assay based on Superose-6 fast protein liquid chromatography separation, which resolved extensin monomers from the newly formed oligomers within 25 minutes. Salt elution of slowly growing (early stationary phase) cells yielded little or no extensin monomers but did give a highly active enzymic preparation that specifically cross-linked extensin monomers inmore » the presence of hydrogen peroxide, judging from: (a) a decrease in the extensin monomer peak on fast protein liquid chromatography gel filtration, (b) appearance of oligomeric peaks, and (c) direct electron microscopical observation of the cross-linked oligomers. The cross-linking reaction had a broad pH optimum between 5.5 and 6.5. An approach to substrate saturation of the enzyme required extensin monomer concentrations of 20 to 40 milligrams per milliliter. Preincubation with catalase completely inhibited the cross-linking reaction, which was highly dependent on hydrogen peroxide and optimal at 15 to 50 micromolar. They therefore identified the cross-linking activity as extensin peroxidase.« less

  7. Novel hyaluronic acid dermal filler: dermal gel extra physical properties and clinical outcomes.

    PubMed

    Monheit, Gary D; Baumann, Leslie S; Gold, Michael H; Goldberg, David J; Goldman, Mitchel P; Narins, Rhoda S; Bachtell, Nathan; Garcia, Emily; Kablik, Jeffrey; Gershkovich, Julia; Burkholder, David

    2010-11-01

    Dermal gel extra (DGE) is a new, tightly cross-linked hyaluronic acid (HA)-based dermal filler containing lidocaine engineered to resist gel deformation and degradation. To develop a firmer gel product (DGE) and compare the efficacy and safety of DGE with nonanimal stabilized HA (NASHA) for correction of nasolabial folds (NLFs). DGE physical properties were characterized, and 140 subjects with moderate to deep NLFs were treated with DGE and NASHA in a randomized, multicenter, split-face design study. Efficacy, pain, and satisfaction were measured using appropriate standard instruments. Adverse events were monitored throughout the study. DGE has a higher modulus and a higher gel:fluid ratio than other HA fillers. Similar optimal correction was observed with DGE and NASHA through 36 weeks (9 months). Study subjects required less volume (p<.001) and fewer touch-ups (p=.005) and reported less injection pain (p<.001) with DGE treatment. Most adverse events were mild to moderate skin reactions. DGE is a firm HA gel that required significantly less volume and fewer touch-ups to provide equivalent efficacy to NASHA for NLF correction; both dermal gels were well tolerated. DGE will provide a comfortable and cost-effective dermal filler option for clinicians and patients. © 2010 by the American Society for Dermatologic Surgery, Inc.

  8. Cell protein cross-linking by erbstatin and related compounds | Center for Cancer Research

    Cancer.gov

    The scheme depicts a possible mechanism of cross-linking by erbstatin and related analogues. A mechanism of action is proposed which involves initial oxidation to reactive quinone intermediates that subsequently cross-link protein nucleophiles via multiple 1,4-Michael-type additions. Similar alkylation of protein by protein-tyrosine kinase inhibitors, such as herbimycin A, has

  9. New Gel-Like Polymers as Selective Weak-Base Anion Exchangers

    PubMed Central

    Gierczyk, Błażej; Cegłowski, Michał; Zalas, Maciej

    2015-01-01

    A group of new anion exchangers, based on polyamine podands and of excellent ion-binding capacity, were synthesized. The materials were obtained in reactions between various poly(ethyleneamines) with glycidyl derivatives of cyclotetrasiloxane. The final polymeric, strongly cross-linked materials form gel-like solids. Their structures and interactions with anions adsorbed were studied by spectroscopic methods (CP-MAS NMR, FR-IR, UV-Vis). The sorption isotherms and kinetic parameters were determined for 29 anions. Materials studied show high ion capacity and selectivity towards some important anions, e.g., selenate(VI) or perrhenate. PMID:25946220

  10. N1N8-bis(gamma-glutamyl)spermidine cross-linking in epidermal-cell envelopes. Comparison of cross-link levels in normal and psoriatic cell envelopes.

    PubMed Central

    Martinet, N; Beninati, S; Nigra, T P; Folk, J E

    1990-01-01

    N1N8-Bis(gamma-glutamyl)spermidine was found in exhaustive proteolytic digests of isolated cell envelopes from human epidermis at levels comparable with those of epsilon-(gamma-glutamyl)lysine. Significantly higher than normal amounts of these compounds, particularly the bis(gamma-glutamyl)polyamine, were observed in envelopes from afflicted areas (scales) of psoriatic patients. These findings support the notions that N1N8-bis(gamma-glutamyl)spermidine, like epsilon-(gamma-glutamyl)lysine, functions in cell envelopes as an enzyme-generated protein cross-link and stabilizing force and that individuals with the chronic, recurrent skin disease, psoriasis, exhibit in involved epidermis abnormal cell-envelope-protein cross-linking. PMID:2241917

  11. Studies of the permeation properties of glomerular basement membrane: cross-linking renders glomerular basement membrane permeable to protein.

    PubMed

    Walton, H A; Byrne, J; Robinson, G B

    1992-03-20

    Cross-linking glomerular basement membrane (GBM) has been shown to render it more permeable to protein. Isolated pig GBM was cross-linked with dimethylmalonimidate which reacts selectively with lysine epsilon-NH2 groups or with glutaraldehyde, a less selective cross-linking agent. Studies of the ultrafiltration properties of these materials in vitro using cytochrome c, myoglobin, bovine serum albumin and immunoglobulin showed that cross-linking had markedly increased solvent and protein fluxes as compared with native membranes particularly at higher pressures. Filtration studies with serum demonstrated that the cross-linked membranes were more permeable to serum proteins. Thickness measurements under pressure indicated that cross-linked membrane was less compressed than native membrane as pressure was increased. Pore theory did not provide a suitable model for analysis of the results, but analysis of the results using the fibre-matrix hypothesis indicated that cross-linking had the effect of bundling together the fibres (type IV collagen) in the GBM matrix. The effect of cross-linking on filtration could be explained by a combination of contraction of the membrane, fibre bundling and increased rigidity compared with native membrane. Cross-linking of GBM might lead to long-term damage of the glomerular capillary wall in nephritis, so promoting proteinuria.

  12. Mocaf cross-linking with gluten to improve the quality of mocaf dough

    NASA Astrophysics Data System (ADS)

    Raharja, Sapta; Udin, Faqih; Suparno, Ono; Febrianti, Faricha Helfi; Nuraisyah, Ani

    2017-03-01

    Crosslink between mocaf and gluten is conducted to increase the using of mocaf which has very big potential in Indonesia. The effort of cross-linking between mocaf and gluten is to get mocaf flour with better dough quality. This study aims to produce a cross-linked mocaf-gluten flour and to evaluate the influence of heating temperature (X1) and the addition of gluten concentration (X2) using completely randomized design factorial (RAFL). The cross-linking is carried out in alkaline solution with 10%, 20%, and 30% gluten addition and heating temperature at 50, 55, and 60 °C. The result showed that mocaf - gluten flour with the treatment of 30% gluten addition at 55 °C had the largest amount of protein and baking expansion (i.e 19.77% and 2.78 mL/g). Swelling power of the flour was increasing along with the increasing of water absorbing capacity of the mocaf - gluten flour. Birefringence properties of mocaf - gluten flour tended to be reduced as the increasing heating temperature. FTIR analysis of mocaf - gluten flour showed that there was peak strengthening of the infrared spectrum of the C - N bond at 1167-1159 cm-1 which was presumably resulted from the gluten addition and the cross-linking properties.

  13. Cisplatin Cross-Linked Multifunctional Nanodrugplexes for Combination Therapy.

    PubMed

    Zhang, Weiqi; Tung, Ching-Hsuan

    2017-03-15

    Combination therapy efficiently tackles cancer by hitting multiple action mechanisms. However, drugs administered, simultaneously or sequentially, may not reach the targeted sites with the desired dose and ratio. The outcomes of combination therapy could be improved with a polymeric nanoparticle, which can simultaneously transport an optimal combination of drugs. We have demonstrated a simple one-pot strategy to formulate nanomedicines based on platinum coordination and the noncovalent interactions of the drugs. A naturally occurring polymer, hyaluronan (HA), was chosen as the building scaffold to form a nanodrugplex with cisplatin and aromatic-cationic drugs. The platinum coordination between cisplatin and HA induces the formation of a nanocomplex. The aromatic-cationic drugs are tightly packed by an electrostatic interaction and π-π stacking. The nanodrugplex bears excellent flexibility in drug combination and size control. It is stable in storage and has favorable release kinetics and targeting capabilities toward CD44, a receptor for HA that is highly expressed on many types of cancer cells.

  14. In Vivo Immune Responses of Cross-Linked Electrospun Tilapia Collagen Membrane.

    PubMed

    Hassanbhai, Ammar Mansoor; Lau, Chau Sang; Wen, Feng; Jayaraman, Praveena; Goh, Bee Tin; Yu, Na; Teoh, Swee-Hin

    2017-10-01

    Collagen has been used extensively in tissue engineering applications. However, the source of collagen has been primarily bovine and porcine. In view of the potential risk of zoonotic diseases and religious constraints associated with bovine and porcine collagen, fish collagen was examined as an alternative. The aim of this study is to use tilapia fish collagen to develop a cross-linked electrospun membrane to be used as a barrier membrane in guided bone regeneration. As there is limited data available on the cytotoxicity and immunogenicity of cross-linked tilapia collagen, in vitro and in vivo tests were performed to evaluate this in comparison to the commercially available Bio-Gide ® membrane. In this study, collagen was extracted and purified from tilapia skin and electrospun into a nanofibrous membrane. The resultant membrane was cross-linked to obtain a cross-linked electrospun tilapia collagen (CETC) membrane, which was characterized by scanning electron microscopy (SEM), differential scanning calorimetry (DSC), degradation studies, cytotoxicity studies, and cell proliferation studies. The membranes were also implanted subcutaneously in rats and the host immune responses were examined. The DSC data showed that cross-linking increased the denaturation temperature of tilapia collagen to 58.3°C ± 1.4°C. The in vitro tests showed that CETC exhibited no cytotoxicity toward murine fibroblast L929 cells, and culture of murine preosteoblast MC3T3-E1 cells demonstrated better proliferation on CETC as compared to Bio-Gide. When implanted in rats, CETC caused a higher production of interleukin IL-6 at early time points as compared to Bio-Gide, but there was no long-term inflammatory responses after the acute inflammation phase. This finding was supported with histology data, which clearly illustrated that CETC has a decreased inflammatory response comparable to the benchmark control group. In all, this study demonstrated the viability for the use of CETC as a

  15. Influence of cross-link structure, density and mechanical properties in the mesoscale deformation mechanisms of collagen fibrils

    PubMed Central

    Depalle, Baptiste; Qin, Zhao; Shefelbine, Sandra J.; Buehler, Markus J.

    2015-01-01

    Collagen is a ubiquitous protein with remarkable mechanical properties. It is highly elastic, shows large fracture strength and enables substantial energy dissipation during deformation. Most of the connective tissue in humans consists of collagen fibrils composed of a staggered array of tropocollagen molecules, which are connected by intermolecular cross-links. In this study, we report a three-dimensional coarse-grained model of collagen and analyze the influence of enzymatic cross-links on the mechanics of collagen fibrils. Two representatives immature and mature cross-links are implemented in the mesoscale model using a bottom-up approach. By varying the number, type and mechanical properties of cross-links in the fibrils and performing tensile test on the models, we systematically investigate the deformation mechanisms of cross-linked collagen fibrils. We find that cross-linked fibrils exhibit a three phase behavior, which agrees closer with experimental results than what was obtained using previous models. The fibril mechanical response is characterized by: (i) an initial elastic deformation corresponding to the collagen molecule uncoiling, (ii) a linear regime dominated by molecule sliding and (iii) the second stiffer elastic regime related to the stretching of the backbone of the tropocollagen molecules until the fibril ruptures. Our results suggest that both cross-link density and type dictate the stiffness of large deformation regime by increasing the number of interconnected molecules while cross-links mechanical properties determine the failure strain and strength of the fibril. These findings reveal that cross-links play an essential role in creating an interconnected fibrillar material of tunable toughness and strength. PMID:25153614

  16. The process of EDC-NHS cross-linking of reconstituted collagen fibres increases collagen fibrillar order and alignment

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Shepherd, D. V., E-mail: dvs23@cam.ac.uk; Shepherd, J. H.; Cameron, R. E.

    We describe the production of collagen fibre bundles through a multi-strand, semi-continuous extrusion process. Cross-linking using an EDC (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide), NHS (N-hydroxysuccinimide) combination was considered. Atomic Force Microscopy and Raman spectroscopy focused on how cross-linking affected the collagen fibrillar structure. In the cross-linked fibres, a clear fibrillar structure comparable to native collagen was observed which was not observed in the non-cross-linked fibre. The amide III doublet in the Raman spectra provided additional evidence of alignment in the cross-linked fibres. Raman spectroscopy also indicated no residual polyethylene glycol (from the fibre forming buffer) or water in any of the fibres.

  17. Amino Acid Proximities in Two Sup35 Prion Strains Revealed by Chemical Cross-linking*

    PubMed Central

    Wong, Shenq-Huey; King, Chih-Yen

    2015-01-01

    Strains of the yeast prion [PSI] are different folding patterns of the same Sup35 protein, which stacks up periodically to form a prion fiber. Chemical cross-linking is employed here to probe different fiber structures assembled with a mutant Sup35 fragment. The photo-reactive cross-linker, p-benzoyl-l-phenylalanine (pBpa), was biosynthetically incorporated into bacterially prepared recombinant Sup(1–61)-GFP, containing the first 61 residues of Sup35, followed by the green fluorescent protein. Four methionine substitutions and two alanine substitutions were introduced at fixed positions in Sup(1–61) to allow cyanogen bromide cleavage to facilitate subsequent mass spectrometry analysis. Amyloid fibers of pBpa and Met/Ala-substituted Sup(1–61)-GFP were nucleated from purified yeast prion particles of two different strains, namely VK and VL, and shown to faithfully transmit specific strain characteristics to yeast expressing the wild type Sup35 protein. Intra- and intermolecular cross-linking were distinguished by tandem mass spectrometry analysis on fibers seeded from solutions containing equal amounts of 14N- and 15N-labeled protein. Fibers propagating the VL strain type exhibited intra- and intermolecular cross-linking between amino acid residues 3 and 28, as well as intra- and intermolecular linking between 32 and 55. Inter- and intramolecular cross-linking between residues 32 and 55 were detected in fibers propagating the VK strain type. Adjacencies of amino acid residues in space revealed by cross-linking were used to constrain possible chain folds of different [PSI] strains. PMID:26265470

  18. Determination of Hyaluronan Molecular Mass Distribution in Human Breast Milk

    PubMed Central

    Yuan, Han; Amin, Ripal; Ye, Xin; De La Motte, Carol A.; Cowman, Mary K.

    2015-01-01

    Hyaluronan (HA) in human milk mediates host responses to microbial infection, via TLR4- and CD44-dependent signaling. Signaling by HA is generally size-specific. Because pure HA with average molecular mass (M) of 35 kDa can elicit a protective response in intestinal epithelial cells, it has been proposed that human milk HA may have a bioactive low M component. Here we report the size distribution of HA in human milk samples from twenty unique donors. A new method for HA analysis, employingion exchange (IEX) chromatography to fractionate HA by size, and specific quantification of each size fraction by competitive Enzyme Linked Sorbent Assay (ELSA), was developed. When separated into four fractions, milk HA with M ≤ 20 kDa, M ≈20-60 kDa, and M ≈ 60-110 kDa comprised an average of 1.5%, 1.4% and 2% of the total HA, respectively. The remaining 95% was HA with M≥110 kDa. Electrophoretic analysis of the higher M HA from thirteen samples showed nearly identical M distributions, with an average M of ∼440 kDa. This higher M HA component in human milk is proposed to bind to CD44 and to enhance human beta defensin 2 (HBD2) induction by the low M HA components. PMID:25579786

  19. EDC Cross-linking of Decellularized Tissue: A Promising Approach?

    PubMed

    Lehmann, Nadine; Christ, Torsten; Daugs, Aila; Bloch, Oliver; Holinski, Sebastian

    2017-07-01

    Decellularization of xenogenous cardiovascular structures is a promising approach to create scaffolds for tissue engineering. Unfortunately, handling and pliability of the unfixed tissue is challenging. N-(3-dimethylaminopropyl)-N9-ethylcarbodiimide (EDC) is an alternative cross-linking agent to glutaraldehyde (GA). Applied in native tissue, it provides biocompatibility and shows no potential for calcification. In addition, EDC can be used to link growth factors (GFs) to tissue scaffolds after decellularization. EDC cross-linking could thereby help to improve decellularized tissue without the toxicity of GA. Porcine aortic wall tissue specimens (TS) were decellularized, treated with EDC, and coated with fibroblast growth factor (FGF) or vascular endothelial growth factor (VEGF). Afterward, TS were subcutaneously implanted in 36 Lewis rats along with one decellularized TS without EDC treatment. After 2, 4, and 6 weeks TS were explanted from 12 rats, respectively. Vital cells were evaluated by RNA quantification, general cellular infiltration by hematoxylin and eosin staining (H&E), macrophage infiltration by CD68 staining, calcification by Von-Kossa staining, and tissue degradation by measurement of TS thickness. Quantification of vital cells showed reduced reseeding of EDC-treated TS compared to noncross-linked TS after 2 (p < 0.05) and 4 weeks (p < 0.05), while after 6 weeks only EDC+VEGF showed fewer viable cells (p < 0.01). Histological evaluation confirmed a reduced infiltration of EDC-treated TS. Macrophage infiltration decreased in all groups from 2 to 6 weeks, with the smallest population in EDC+VEGF-treated TS (p > 0.05). In EDC+FGF-treated TS, macrophages were reduced after 2 weeks compared to noncross-linked TS (p < 0.05), while after 4 and 6 weeks no significant difference was found (p > 0.05). Von-Kossa staining revealed no calcification in any of the specimens. Thickness of noncross-linked and EDC+FGF-treated TS was not

  20. Cross-linked polyimides for integrated optics

    NASA Astrophysics Data System (ADS)

    Singer, Kenneth D.; Kowalczyk, Tony C.; Nguyen, Hung D.; Beuhler, Allyson J.; Wargowski, David A.

    1997-01-01

    We have investigated a promising class of polyimide materials for both passive and active electro-optic devices, namely crosslinkable polyimides. These fluorinated polyimides are soluble in the imidized form and are both thermally and photo-crosslinkable leading to easy processability into waveguide structures and the possibility of stable electro-optic properties. We have fabricated channel and slab waveguides and investigated the mechanism of optical propagation loss using photothermal deflection spectroscopy and waveguide loss spectroscopy, and found the losses to arise from residual absorption due to the formation of charge transfer states. The absorption is inhibited by fluorination leading to propagation losses as low as 0.3 dB/cm in the near infrared. Because of the ability to photocrosslink, channel waveguides are fabricated using a simple wet-etch process. Channel waveguides so formed are observed to have no excess loss over slab structures. Solubility followed by thermal cross-linking allows the formation of multilayer structures. We have produced electro-optic polymers by doping with the nonlinear optical chromophores, DCM and DADC; and a process of concurrent poling and thermal crosslinking. Multilayer structures have been investigated and poling fields optimized in the active layer by doping the cladding with an anti-static agent. The high glass-transition temperature and cross-linking leads to very stable electro-optic properties. We are currently building electro-optic modulators based on these materials. Progress and results in this area also are reported.

  1. Durability improvements of two-dimensional metal nanoparticle sheets by molecular cross-linked structures between nanoparticles

    NASA Astrophysics Data System (ADS)

    Saito, Noboru; Ryuzaki, Sou; Wang, Pangpang; Park, Susie; Sakai, Nobuyuki; Tatsuma, Tetsu; Okamoto, Koichi; Tamada, Kaoru

    2018-03-01

    The durability of two-dimensional metal nanoparticle sheets is a crucial factor for realizing next-generation optoelectronic devices based on plasmonics such as organic light-emitting diodes. Here, we report improvements in the durability of Ag nanoparticle sheets by forming alkanedithiol (DT16) cross-linked structures between the nanoparticles. The cross-linked structures in a sheet were fabricated by the self-assembly of DT16-capped Ag nanoparticles with 10% coverage (AgDT16). The durabilities for thermal, organic solvent, and oxidation reactions of AgDT16 sheets were found to be improved owing to the cross-linked structures by comparing Ag nanoparticle sheets without the cross-linked structures. The absorbance spectra revealed that the Ag nanoparticle sheets without the structure are markedly damaged by each durability test, whereas the AgDT16 sheets remain. The molecular cross-linked structures between nanoparticles in two-dimansional metal nanoparticle sheets were found to have the potential to play a key role in the realization of plasmonic optoelectronic devices including metal nanoparticles.

  2. Controlled drug release properties of ionically cross-linked chitosan beads: the influence of anion structure.

    PubMed

    Shu, X Z; Zhu, K J

    2002-02-21

    By adopting a novel chitosan cross-linked method, i.e. chitosan/gelatin droplet coagulated at low temperature and then cross-linked by anions (sulfate, citrate and tripolyphosphate (TPP)), the chitosan beads were prepared. Scanning electron microscopy (SEM) observation showed that sulfate/chitosan and citrate/chitosan beads usually had a spherical shape, smooth surface morphology and integral inside structure. Cross-sectional analysis indicated that the cross-linking process of sulfate and citrate to chitosan was much faster than that of TPP due to their smaller molecular size. But, once completely cross-linked, TPP/chitosan beads possessed much better mechanical strength and the force to break the beads was approximately ten times higher than that of sulfate/chitosan or citrate/chitosan beads. Release media pH and ionic strength seriously influenced the controlled drug release properties of the beads, which related to the strength of electrostatic interaction between anions and chitosan. Sulfate and citrate cross-linked chitosan beads swelled and even dissociated in simulated gastric fluid (SGF) and hence, model drug (riboflavin) released completely in 5 h; while in simulated intestinal fluid (SIF), beads remained in a shrinkage state and drug released slowly (release % usually <70% in 24 h). However, swelling and drug release of TPP/chitosan bead was usually insensitive to media pH. Chitosan beads, cross-linked by a combination of TPP and citrate (or sulfate) together, not only had a good shape, but also improved pH-responsive drug release properties. Salt weakened the interaction of citrate, especially sulfate with chitosan and accelerated beads swelling and hence drug release rate, but it was insensitive to that of TPP/chitosan. These results indicate that ionically cross-linked chitosan beads may be useful in stomach specific drug delivery.

  3. Immobilization of cross-linked tannase enzyme on multiwalled carbon nanotubes and its catalytic behavior.

    PubMed

    Ong, Chong-Boon; Annuar, Mohamad S M

    2018-02-07

    Immobilization of cross-linked tannase on pristine multiwalled carbon nanotubes (MWCNT) was successfully performed. Cross-linking of tannase molecules was made through glutaraldehyde. The immobilized tannase exhibited significantly improved pH, thermal, and recycling stability. The optimal pH for both free and immobilized tannase was observed at pH 5.0 with optimal operating temperature at 30°C. Moreover, immobilized enzyme retained greater biocatalytic activities upon 10 repeated uses compared to free enzyme in solution. Immobilization of tannase was accomplished by strong hydrophobic interaction most likely between hydrophobic amino acid moieties of the glutaraldehyde-cross-linked tannase to the MWCNT.

  4. Chemical cross-linking of polypropylenes towards new shape memory polymers.

    PubMed

    Raidt, Thomas; Hoeher, Robin; Katzenberg, Frank; Tiller, Joerg C

    2015-04-01

    In this work, syndiotactic polypropylene (sPP) as well as isotactic polypropylene (iPP) are cross-linked to gain a shape memory effect. Both prepared PP networks exhibit maximum strains of 700%, stored strains of up to 680%, and recoveries of nearly 100%. While x-iPP is stable for many cycles, x-sPP ruptures after the first shape-memory cycle. It is shown by wide-angle X-ray scattering (WAXS) experiments that cross-linked iPP exhibits homoepitaxy in the temporary, stretched shape but in contrast to previous reports it contains a higher amount of daughter than mother crystals. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. An atomistic model for cross-linked HNBR elastomers used in seals

    NASA Astrophysics Data System (ADS)

    Molinari, Nicola; Sutton, Adrian; Stevens, John; Mostofi, Arash

    2015-03-01

    Hydrogenated nitrile butadiene rubber (HNBR) is one of the most common elastomeric materials used for seals in the oil and gas industry. These seals sometimes suffer ``explosive decompression,'' a costly problem in which gases permeate a seal at the elevated temperatures and pressures pertaining in oil and gas wells, leading to rupture when the seal is brought back to the surface. The experimental evidence that HNBR and its unsaturated parent NBR have markedly different swelling properties suggests that cross-linking may occur during hydrogenation of NBR to produce HNBR. We have developed a code compatible with the LAMMPS molecular dynamics package to generate fully atomistic HNBR configurations by hydrogenating initial NBR structures. This can be done with any desired degree of cross-linking. The code uses a model of atomic interactions based on the OPLS-AA force-field. We present calculations of the dependence of a number of bulk properties on the degree of cross-linking. Using our atomistic representations of HNBR and NBR, we hope to develop a better molecular understanding of the mechanisms that result in explosive decompression.

  6. Thermally Conductive-Silicone Composites with Thermally Reversible Cross-links.

    PubMed

    Wertz, J T; Kuczynski, J P; Boday, D J

    2016-06-08

    Thermally conductive-silicone composites that contain thermally reversible cross-links were prepared by blending diene- and dienophile-functionalized polydimethylsiloxane (PDMS) with an aluminum oxide conductive filler. This class of thermally conductive-silicones are useful as thermal interface materials (TIMs) within Information Technology (IT) hardware applications to allow rework of valuable components. The composites were rendered reworkable via retro Diels-Alder cross-links when temperatures were elevated above 130 °C and required little mechanical force to remove, making them advantageous over other TIM materials. Results show high thermal conductivity (0.4 W/m·K) at low filler loadings (45 wt %) compared to other TIM solutions (>45 wt %). Additionally, the adhesion of the material was found to be ∼7 times greater at lower temperatures (25 °C) and ∼2 times greater at higher temperatures (120 °C) than commercially available TIMs.

  7. Investigation of cross-linking characteristics of novel hole-transporting materials for solution-processed phosphorescent OLEDs

    NASA Astrophysics Data System (ADS)

    Lee, Jaemin; Ameen, Shahid; Lee, Changjin

    2016-04-01

    After the success of commercialization of the vacuum-evaporated organic light-emitting diodes (OLEDs), solutionprocessing or printing of OLEDs are currently attracting much research interests. However, contrary to various kinds of readily available vacuum-evaporable OLED materials, the solution-processable OLED materials are still relatively rare. Hole-transporting layer (HTL) materials for solution-processed OLEDs are especially limited, because they need additional characteristics such as cross-linking to realize multilayer structures in solution-processed OLEDs, as well as their own electrically hole-transporting characteristics. The presence of such cross-linking characteristics of solutionprocessable HTL materials therefore makes them more challenging in the development stage, and also makes them essence of solution-processable OLED materials. In this work, the structure-property relationships of thermally crosslinkable HTL materials were systematically investigated by changing styrene-based cross-linking functionalities and modifying the carbazole-based hole-transporting core structures. The temperature dependency of the cross-linking characteristics of the HTL materials was systematically investigated by the UV-vis. absorption spectroscopy. The new HTL materials were also applied to green phosphorescent OLEDs, and their device characteristics were also investigated based on the chemical structures of the HTL materials. The device configuration was [ITO / PEDOT:PSS / HTL / EML / ETL / CsF / Al]. We found out that the chemical structures of the cross-linking functionalities greatly affect not only the cross-linking characteristics of the resultant HTL materials, but also the resultant OLED device characteristics. The increase of the maximum luminance and efficiency of OLEDs was evident as the cross-linking temperature decreases from higher than 200°C to at around 150°C.

  8. NOVEL METALLIC AND BIMETALLIC CROSS-LINKED POLY (VINYL ALCOHOL) NANOCOMPOSITES PREPARED UNDER MICROWAVE IRRADIATION

    EPA Science Inventory

    A facile microwave irradiation approach that results in a cross-linking reaction of poly (vinyl alcohol) (PVA) with metallic and bimetallic systems is described. Nanocomposites of PVA cross-linked metallic systems such as Pt, Cu, and In and bimetallic systems such as Pt-In, Ag-P...

  9. Genipin Cross-Linked Glucose Oxidase and Catalase Multi-enzyme for Gluconic Acid Synthesis.

    PubMed

    Cui, Caixia; Chen, Haibin; Chen, Biqiang; Tan, Tianwei

    2017-02-01

    In this work, glucose oxidase (GOD) and catalase (CAT) were used simultaneously to produce gluconic acid from glucose. In order to reduce the distance between the two enzymes, and therefore improve efficiency, GOD and CAT were cross-linked together using genipin. Improvements in gluconic acid production were due to quick removal of harmful intermediate hydrogen peroxide by CAT. GOD activity was significantly affected by the proportion of CAT in the system, with GOD activity in the cross-linked multi-enzyme (CLME) being 10 times higher than that in an un-cross-linked GOD/CAT mixture. The glucose conversion rate after 15 h using 15 % glucose was also 10 % higher using the CLME than was measured using a GOD/CAT mixture.

  10. Effects of chebulic acid on advanced glycation endproducts-induced collagen cross-links.

    PubMed

    Lee, Ji-Young; Oh, Jun-Gu; Kim, Jin Sook; Lee, Kwang-Won

    2014-01-01

    Advanced glycation end-products (AGEs) have been implicated in the development of diabetic complications. We report the antiglycating activity of chebulic acid (CA), isolated from Terminalia chebula on breaking the cross-links of proteins induced by AGEs and inhibiting the formation of AGEs. Aminoguanidine (AG) reduced 50% of glycated bovine serum albumin (BSA) with glycolaldehyde (glycol-BSA)-induced cross-links of collagen at a concentration of 67.8 ± 2.5 mM, the level of CA required for exerting a similar antiglycating activity was 38.8 ± 0.5 µM. Also, the breaking activity on collagen cross-links induced by glycol-BSA was potent with CA (IC50=1.46 ± 0.05 mM), exhibiting 50-fold stronger breaking activity than with ALT-711, a well-known cross-link breaker (IC50=72.2 ± 2.4 mM). IC50 values of DPPH· scavenging activity for CA and ascorbic acid (AA) were 39.2 ± 4.9 and 19.0 ± 1.2 µg dry matter (DM) mL(-1), respectively, and ferric reducing and antioxidant power (FRAP) activities for CA and AA were 4.70 ± 0.06 and 11.4 ± 0.1 mmol/FeSO4·7H2O/g DM, respectively. The chelating activities of CA, AG and ALT711 on copper-catalyzed oxidation of AA were compared, and in increasing order, ALT-711 (IC50 of 1.92 ± 0.20 mM)cross-linking, the activity of which may be explained in large part by its chelating and antioxidant activities, suggesting that CA may constitute a promising antiglycating candidate in intervening AGE-mediated diabetic complications.

  11. Influence of cross-link structure, density and mechanical properties in the mesoscale deformation mechanisms of collagen fibrils.

    PubMed

    Depalle, Baptiste; Qin, Zhao; Shefelbine, Sandra J; Buehler, Markus J

    2015-12-01

    Collagen is a ubiquitous protein with remarkable mechanical properties. It is highly elastic, shows large fracture strength and enables substantial energy dissipation during deformation. Most of the connective tissue in humans consists of collagen fibrils composed of a staggered array of tropocollagen molecules, which are connected by intermolecular cross-links. In this study, we report a three-dimensional coarse-grained model of collagen and analyze the influence of enzymatic cross-links on the mechanics of collagen fibrils. Two representatives immature and mature cross-links are implemented in the mesoscale model using a bottom-up approach. By varying the number, type and mechanical properties of cross-links in the fibrils and performing tensile test on the models, we systematically investigate the deformation mechanisms of cross-linked collagen fibrils. We find that cross-linked fibrils exhibit a three phase behavior, which agrees closer with experimental results than what was obtained using previous models. The fibril mechanical response is characterized by: (i) an initial elastic deformation corresponding to the collagen molecule uncoiling, (ii) a linear regime dominated by molecule sliding and (iii) the second stiffer elastic regime related to the stretching of the backbone of the tropocollagen molecules until the fibril ruptures. Our results suggest that both cross-link density and type dictate the stiffness of large deformation regime by increasing the number of interconnected molecules while cross-links mechanical properties determine the failure strain and strength of the fibril. These findings reveal that cross-links play an essential role in creating an interconnected fibrillar material of tunable toughness and strength. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

  12. Hyaluronan--a review of the recent patent literature.

    PubMed

    Moore, A R

    2000-02-01

    A review of patent literature surrounding the use of hyaluronan (HA) and its derivatives, or agents capable of modulating endogenous levels, is presented. Indications are many and varied and include arthritis, abnormal wound healing, neoplasia, prevention of surgical adhesions and dermatological disorders. In many cases, it is the physicochemical and biological properties of HA itself that have been exploited. For example, HA and/or its derivatives have been suggested as a synovial fluid replacement, a vehicle for topical application of drugs, a matrix for sustained drug release and a putative drug delivery system (targeting HA receptors at sites of pathology). In each of these examples, the lack of toxicity of HA and its biocompatability are important considerations.

  13. Orally administered hyaluronan affects skin dryness and epidermal thickening in photoaged hairless mice.

    PubMed

    Kawada, Chinatsu; Kimura, Mamoru; Masuda, Yasunobu; Nomura, Yoshihiro

    2016-06-01

    The oral administration of hyaluronans (HAs) (molecular weight, 300k and less than 10k) to photoaged hairless mice increased the moisture content of the stratum corneum and decreased the epidermal thickness, respectively. Furthermore, orally administered HAs suppressed the low-molecular weight of HA content of the skin. This study indicates oral administered HAs may ameliorate the skin condition resulting from photoaging.

  14. Ingested hyaluronan moisturizes dry skin

    PubMed Central

    2014-01-01

    Hyaluronan (HA) is present in many tissues of the body and is essential to maintain moistness in the skin tissues, which contain approximately half the body’s HA mass. Due to its viscosity and moisturizing effect, HA is widely distributed as a medicine, cosmetic, food, and, recently marketed in Japan as a popular dietary supplement to promote skin moisture. In a randomized, double-blind, placebo-controlled clinical study it was found that ingested HA increased skin moisture and improved treatment outcomes for patients with dry skin. HA is also reported to be absorbed by the body distributed, in part, to the skin. Ingested HA contributes to the increased synthesis of HA and promotes cell proliferation in fibroblasts. These effects show that ingestion of HA moisturizes the skin and is expected to improve the quality of life for people who suffer from dry skin. This review examines the moisturizing effects of dry skin by ingested HA and summarizes the series of mechanisms from absorption to pharmacological action. PMID:25014997

  15. Probabilistic cross-link analysis and experiment planning for high-throughput elucidation of protein structure.

    PubMed

    Ye, Xiaoduan; O'Neil, Patrick K; Foster, Adrienne N; Gajda, Michal J; Kosinski, Jan; Kurowski, Michal A; Bujnicki, Janusz M; Friedman, Alan M; Bailey-Kellogg, Chris

    2004-12-01

    Emerging high-throughput techniques for the characterization of protein and protein-complex structures yield noisy data with sparse information content, placing a significant burden on computation to properly interpret the experimental data. One such technique uses cross-linking (chemical or by cysteine oxidation) to confirm or select among proposed structural models (e.g., from fold recognition, ab initio prediction, or docking) by testing the consistency between cross-linking data and model geometry. This paper develops a probabilistic framework for analyzing the information content in cross-linking experiments, accounting for anticipated experimental error. This framework supports a mechanism for planning experiments to optimize the information gained. We evaluate potential experiment plans using explicit trade-offs among key properties of practical importance: discriminability, coverage, balance, ambiguity, and cost. We devise a greedy algorithm that considers those properties and, from a large number of combinatorial possibilities, rapidly selects sets of experiments expected to discriminate pairs of models efficiently. In an application to residue-specific chemical cross-linking, we demonstrate the ability of our approach to plan experiments effectively involving combinations of cross-linkers and introduced mutations. We also describe an experiment plan for the bacteriophage lambda Tfa chaperone protein in which we plan dicysteine mutants for discriminating threading models by disulfide formation. Preliminary results from a subset of the planned experiments are consistent and demonstrate the practicality of planning. Our methods provide the experimenter with a valuable tool (available from the authors) for understanding and optimizing cross-linking experiments.

  16. A case of cervical cancer expressed three mRNA variant of Hyaluronan-mediated motility receptor

    PubMed Central

    Villegas-Ruíz, Vanessa; Salcedo, Mauricio; Zentella-Dehesa, Alejandro; de Oca, Edén V Montes; Román-Basaure, Edgar; Mantilla-Morales, Alejandra; Dávila-Borja, Víctor M; Juárez-Méndez, Sergio

    2014-01-01

    Cervical cancer is the second malignancy in Mexico, little is known about the prognostic factors associated with this disease. Several cellular components are important in their transformation and progression. Alternative mRNA splice is an important mechanism for generating protein diversity, nevertheless, in cancer unknown mRNA diversity is expressed. Hyaluronan-mediated motility receptor (HMMR, RHAMM, CD168) is a family member of proteins, hyaluronan acid dependent, and has been associated with different malignant processes such as: angiogenesis, cell invasiveness, proliferation, metastasis and poor outcome in some tumors. In the present study we identified expression of HMMR in cervical cancer by means of RT-PCR and sequencing. Our results indicate co-expression of two HMMR variants in all samples, and one case expressed three alternative HMMR splice transcripts. These results showed the heterogeneity of mRNA transcripts of HMMR that could express in cancer and the expression of HMMR could be marker of malignancy in CC. PMID:24966934

  17. Developing a Multiplexed Quantitative Cross-Linking Mass Spectrometry Platform for Comparative Structural Analysis of Protein Complexes.

    PubMed

    Yu, Clinton; Huszagh, Alexander; Viner, Rosa; Novitsky, Eric J; Rychnovsky, Scott D; Huang, Lan

    2016-10-18

    Cross-linking mass spectrometry (XL-MS) represents a recently popularized hybrid methodology for defining protein-protein interactions (PPIs) and analyzing structures of large protein assemblies. In particular, XL-MS strategies have been demonstrated to be effective in elucidating molecular details of PPIs at the peptide resolution, providing a complementary set of structural data that can be utilized to refine existing complex structures or direct de novo modeling of unknown protein structures. To study structural and interaction dynamics of protein complexes, quantitative cross-linking mass spectrometry (QXL-MS) strategies based on isotope-labeled cross-linkers have been developed. Although successful, these approaches are mostly limited to pairwise comparisons. In order to establish a robust workflow enabling comparative analysis of multiple cross-linked samples simultaneously, we have developed a multiplexed QXL-MS strategy, namely, QMIX (Quantitation of Multiplexed, Isobaric-labeled cross (X)-linked peptides) by integrating MS-cleavable cross-linkers with isobaric labeling reagents. This study has established a new analytical platform for quantitative analysis of cross-linked peptides, which can be directly applied for multiplexed comparisons of the conformational dynamics of protein complexes and PPIs at the proteome scale in future studies.

  18. A Modified ELISA Accurately Measures Secretion of High Molecular Weight Hyaluronan (HA) by Graves' Disease Orbital Cells

    PubMed Central

    Krieger, Christine C.

    2014-01-01

    Excess production of hyaluronan (hyaluronic acid [HA]) in the retro-orbital space is a major component of Graves' ophthalmopathy, and regulation of HA production by orbital cells is a major research area. In most previous studies, HA was measured by ELISAs that used HA-binding proteins for detection and rooster comb HA as standards. We show that the binding efficiency of HA-binding protein in the ELISA is a function of HA polymer size. Using gel electrophoresis, we show that HA secreted from orbital cells is primarily comprised of polymers more than 500 000. We modified a commercially available ELISA by using 1 million molecular weight HA as standard to accurately measure HA of this size. We demonstrated that IL-1β-stimulated HA secretion is at least 2-fold greater than previously reported, and activation of the TSH receptor by an activating antibody M22 from a patient with Graves' disease led to more than 3-fold increase in HA production in both fibroblasts/preadipocytes and adipocytes. These effects were not consistently detected with the commercial ELISA using rooster comb HA as standard and suggest that fibroblasts/preadipocytes may play a more prominent role in HA remodeling in Graves' ophthalmopathy than previously appreciated. PMID:24302624

  19. Molecular evolution of the hyaluronan synthase 2 gene in mammals: implications for adaptations to the subterranean niche and cancer resistance

    PubMed Central

    Faulkes, Christopher G.; Davies, Kalina T. J.; Rossiter, Stephen J.; Bennett, Nigel C.

    2015-01-01

    The naked mole-rat (NMR) Heterocephalus glaber is a unique and fascinating mammal exhibiting many unusual adaptations to a subterranean lifestyle. The recent discovery of their resistance to cancer and exceptional longevity has opened up new and important avenues of research. Part of this resistance to cancer has been attributed to the fact that NMRs produce a modified form of hyaluronan—a key constituent of the extracellular matrix—that is thought to confer increased elasticity of the skin as an adaptation for living in narrow tunnels. This so-called high molecular mass hyaluronan (HMM-HA) stems from two apparently unique substitutions in the hyaluronan synthase 2 enzyme (HAS2). To test whether other subterranean mammals with similar selection pressures also show molecular adaptation in their HAS2 gene, we sequenced the HAS2 gene for 11 subterranean mammals and closely related species, and combined these with data from 57 other mammals. Comparative screening revealed that one of the two putatively important HAS2 substitutions in the NMR predicted to have a significant effect on hyaluronan synthase function was uniquely shared by all African mole-rats. Interestingly, we also identified multiple other amino acid substitutions in key domains of the HAS2 molecule, although the biological consequences of these for hyaluronan synthesis remain to be determined. Despite these results, we found evidence of strong purifying selection acting on the HAS2 gene across all mammals, and the NMR remains unique in its particular HAS2 sequence. Our results indicate that more work is needed to determine whether the apparent cancer resistance seen in NMR is shared by other members of the African mole-rat clade. PMID:25948568

  20. Relationships between human cortical bone toughness and collagen cross-links on paired anatomical locations.

    PubMed

    Gauthier, Rémy; Follet, Hélène; Langer, Max; Gineyts, Evelyne; Rongiéras, Frédéric; Peyrin, Françoise; Mitton, David

    2018-07-01

    Human cortical bone fracture processes depend on the internal porosity network down to the lacunar length scale. Recent results show that at the collagen scale, the maturation of collagen cross-links may have a negative influence on bone mechanical behavior. While the effect of pentosidine on human cortical bone toughness has been studied, the influence of mature and immature enzymatic cross-links has only been studied in relation to strength and work of fracture. Moreover, these relationships have not been studied on different paired anatomical locations. Thus, the aim of the current study was to assess the relationships between both enzymatic and non-enzymatic collagen cross-links and human cortical bone toughness, on four human paired anatomical locations. Single Edge Notched Bending toughness tests were performed for two loading conditions: a quasi-static standard condition, and a condition representative of a fall. These tests were done with 32 paired femoral diaphyses, femoral necks and radial diaphyses (18 women, age 81 ± 12 y.o.; 14 men, age 79 ± 8 y.o.). Collagen enzymatic and non-enzymatic crosslinks were measured on the same bones. Maturation of collagen was defined as the ratio between immature and mature cross-links (CX). The results show that there was a significant correlation between collagen cross-link maturation and bone toughness when gathering femoral and radial diaphyses, but not when considering each anatomical location individually. These results show that the influence of collagen enzymatic and non-enzymatic cross-links is minor when considering human cortical bone crack propagation mechanisms. Copyright © 2018 Elsevier Inc. All rights reserved.