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Sample records for cross-linked hyaluronan gel

  1. Self-reinforcement and protein sustained delivery of hyaluronan hydrogel by tailoring a dually cross-linked network.

    PubMed

    Luo, Chunhong; Xu, Guoguang; Wang, Xinghui; Tu, Mei; Zeng, Rong; Rong, Jianhua; Zhao, Jianhao

    2015-01-01

    A series of self-reinforcing hyaluronan hydrogels were developed to improve mechanical properties and protein sustained delivery thanks to a dually cross-linked network. Hyaluronan gel particles (HGPs, 1-5 μm in diameter) with different cross-linking densities, i.e. HGPs-1.5, HGPs-3 and HGPs-15, were prepared in an inverse emulsion system and used as the reinforcing phase after glycidyl methacrylation, while glycidyl methacrylated hyaluronan with a substitution degree of 45.2% was synthesized as the matrix phase. These two phases were cross-linked under ultraviolet irradiation to form self-reinforcing hyaluronan hydrogels (srHAs) that showed typical cross-linked structure of HGPs connecting the matrix phase by cross-section observation. In comparison to hyaluronan bulk gels and their blends with HGPs, srHAs distinctly enhanced the mechanical properties and BSA long-term sustained delivery, especially srHA-1.5 showed the highest compressive modulus of 220±15 kPa and the slowest BSA delivery (67% release at 14 d). The 3T3 fibroblast cell culture showed that all the srHAs had no cytotoxicity. PMID:25491993

  2. 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 35C. 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.

  3. Enzymatically cross-linked gelatin/chitosan hydrogels: tuning gel properties and cellular response.

    PubMed

    da Silva, Marcelo A; Bode, Franziska; Drake, Alex F; Goldoni, Silvia; Stevens, Molly M; Dreiss, Ccile A

    2014-06-01

    This work investigates the effect of combining physical and chemical gelation processes in a biopolymer blend: chitosan and tilapia fish gelatin. Chemical (C) gels are obtained by cross-linking with the microbial enzyme transglutaminase at 37?C. Hybrid physical-co-chemical (PC) gels are cross-linked at 21?C, below gelatin gelation temperature. These protocols provide two microenvironments for the gelation process: in C gels, both gelatin and chitosan are present as single strands; in PC gels, cross-linking proceeds within a transient physical gel of gelatin, filled by chitosan strands. The chitosan/gelatin chemical networks generated in PC gels show a consistently higher shear modulus than pure C gels; they are also less turbid than their C gels counterparts, suggesting a more homogeneous network. Finally, chitosan enhances the gels' shear modulus in all gels. Proliferation assays show that MC3T3 cells proliferate in these mixed, hybrid gels and better so on PC gels than in C mixed gels. PMID:24550134

  4. Ionically cross-linked hyaluronic acid: wetting, lubrication, and viscoelasticity of a modified adhesion barrier gel

    PubMed Central

    Vorvolakos, Katherine; Isayeva, Irada S; Luu, Hoan-My Do; Patwardhan, Dinesh V; Pollack, Steven K

    2011-01-01

    Hyaluronic acid (HA), in linear or cross-linked form, is a common component of cosmetics, personal care products, combination medical products, and medical devices. In all cases, the ability of the HA solution or gel to wet surfaces and/or disrupt and lubricate interfaces is a limiting feature of its mechanism of action. We synthesized ferric ion–cross-linked networks of HA based on an adhesion barrier, varied the degree of cross-linking, and performed wetting goniometry, viscometry, and dynamic mechanical analysis. As cross-linking increases, so do contact angle, viscosity, storage modulus, and loss modulus; thus, wetting and lubrication are compromised. These findings have implications in medical device materials, such as adhesion barriers and mucosal drug delivery vehicles. PMID:22915924

  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. 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 provide evidence that collagen telopeptide sites play a role in collagen gel lattice contraction.

  7. Preparation of single or double-network chitosan/poly(vinyl alcohol) gel films through selective cross-linking method

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A selective cross-linking method was developed to create single or double network chitosan/poly(vinyl alcohol) gel films. The cross-linking is based on the hydrogen bonding between PVA and borate and the strong electrostatic interaction between chitosan and tripolyphosphate. The resultant gel films ...

  8. 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. PMID:25266064

  9. A gel network constituted by rigid schizophyllan chains and nonpermanent cross-links.

    PubMed

    Fang, Yapeng; Takahashi, Rheo; Nishinari, Katsuyoshi

    2004-01-01

    This work reports a gel network formed by rigid schizophyllan (SPG) chains with Borax as a cross-linking agent. The formed cross-links are non-permanent and somewhat dynamic in nature because the cross-linking reaction is governed by a complexation equilibrium. Gelation processes are traced by dynamic viscoelastic measurements to examine the effects of Borax content, SPG concentration, temperature, salt concentration, salt type, and strain. The first-order kinetic model containing three parameters, t(0) (induction time), 1/tau(c) (gelation rate), and (saturated storage modulus), is successfully applied to describe the gelation of the SPG-Borax system. Gelation occurs faster at higher Borax content, higher SPG concentration, higher salt concentration, or lower temperature. Moreover the gelation is cation-type-specific. Storage modulus is a linear function of both Borax content and SPG concentration. The linear relationship between storage modulus and Borax content can be explained by a modified ideal rubber elasticity theory with a front factor alpha to take into account the presence of ineffective cross-links and the effect of SPG chain rigidity. On the other hand, the linear dependence of storage modulus on SPG concentration could be explained on the basis of chain-chain contacting behavior of extended SPG chains. Apparent activation energy and cross-linking enthalpy are calculated to be -74.5 and -32.4 kJ/mol for the present system. Strain sweep measurements manifest that the elasticity behavior of this gel starts to deviate from Gaussian-chain network at a small strain of 10%. PMID:14715018

  10. Phase diagram of selectively cross-linked block copolymers shows chemically microstructured gel.

    PubMed

    von der Heydt, Alice; Zippelius, Annette

    2015-02-01

    We study analytically the intricate phase behavior of cross-linked AB diblock copolymer melts, which can undergo two main phase transitions due to quenched random constraints. Gelation, i.e., spatially random localisation of polymers forming a system-spanning cluster, is driven by increasing the number parameter ? of irreversible, type-selective cross-links between random pairs of A blocks. Self-assembly into a periodic pattern of A/B-rich microdomains (microphase separation) is controlled by the AB incompatibility ? inversely proportional to temperature. Our model aims to capture the system's essential microscopic features, including an ensemble of random networks that reflects spatial correlations at the instant of cross-linking. We identify suitable order parameters and derive a free-energy functional in the spirit of Landau theory that allows us to trace a phase diagram in the plane of ? and ?. Selective cross-links promote microphase separation at higher critical temperatures than in uncross-linked diblock copolymer melts. Microphase separation in the liquid state facilitates gelation, giving rise to a novel gel state whose chemical composition density mirrors the periodic AB pattern. PMID:25662662

  11. Oxidation increases mucin polymer cross-links to stiffen airway mucus gels

    PubMed Central

    Yuan, Shaopeng; Hollinger, Martin; Lachowicz-Scroggins, Marrah E.; Kerr, Sheena C.; Dunican, Eleanor M.; Daniel, Brian M.; Ghosh, Sudakshina; Erzurum, Serpel C.; Willard, Belinda; Hazen, Stanley L.; Huang, Xiaozhu; Carrington, Stephen D.; Oscarson, Stefan; Fahy, John V.

    2015-01-01

    Airway mucus in cystic fibrosis (CF) is highly elastic, but the mechanism behind this pathology is unclear. We hypothesized that the biophysical properties of CF mucus are altered because of neutrophilic oxidative stress. Using confocal imaging, rheology, and biochemical measures of inflammation and oxidation, we found that CF airway mucus gels have a molecular architecture characterized by a core of mucin covered by a web of DNA and a rheological profile characterized by high elasticity that can be normalized by chemical reduction. We also found that high levels of reactive oxygen species in CF mucus correlated positively and significantly with high concentrations of the oxidized products of cysteine (disulfide cross-links). To directly determine whether oxidation can cross-link mucins to increase mucus elasticity, we exposed induced sputum from healthy subjects to oxidizing stimuli and found a marked and thiol-dependent increase in sputum elasticity. Targeting mucin disulfide cross-links using current thiol-amino structures such as N-acetylcysteine (NAC) requires high drug concentrations to have mucolytic effects. We therefore synthesized a thiol-carbohydrate structure (methyl 6-thio-6-deoxy-α-D-galactopyranoside) and found that it had stronger reducing activity than NAC and more potent and fast-acting mucolytic activity in CF sputum. Thus, oxidation arising from airway inflammation or environmental exposure contributes to pathologic mucus gel formation in the lung, which suggests that it can be targeted by thiol-modified carbohydrates. PMID:25717100

  12. Oxidation increases mucin polymer cross-links to stiffen airway mucus gels.

    PubMed

    Yuan, Shaopeng; Hollinger, Martin; Lachowicz-Scroggins, Marrah E; Kerr, Sheena C; Dunican, Eleanor M; Daniel, Brian M; Ghosh, Sudakshina; Erzurum, Serpel C; Willard, Belinda; Hazen, Stanley L; Huang, Xiaozhu; Carrington, Stephen D; Oscarson, Stefan; Fahy, John V

    2015-02-25

    Airway mucus in cystic fibrosis (CF) is highly elastic, but the mechanism behind this pathology is unclear. We hypothesized that the biophysical properties of CF mucus are altered because of neutrophilic oxidative stress. Using confocal imaging, rheology, and biochemical measures of inflammation and oxidation, we found that CF airway mucus gels have a molecular architecture characterized by a core of mucin covered by a web of DNA and a rheological profile characterized by high elasticity that can be normalized by chemical reduction. We also found that high levels of reactive oxygen species in CF mucus correlated positively and significantly with high concentrations of the oxidized products of cysteine (disulfide cross-links). To directly determine whether oxidation can cross-link mucins to increase mucus elasticity, we exposed induced sputum from healthy subjects to oxidizing stimuli and found a marked and thiol-dependent increase in sputum elasticity. Targeting mucin disulfide cross-links using current thiol-amino structures such as N-acetylcysteine (NAC) requires high drug concentrations to have mucolytic effects. We therefore synthesized a thiol-carbohydrate structure (methyl 6-thio-6-deoxy-α-D-galactopyranoside) and found that it had stronger reducing activity than NAC and more potent and fast-acting mucolytic activity in CF sputum. Thus, oxidation arising from airway inflammation or environmental exposure contributes to pathologic mucus gel formation in the lung, which suggests that it can be targeted by thiol-modified carbohydrates. PMID:25717100

  13. Comparison of agarose and cross-linked protein gels as magnetic resonance imaging phantoms.

    PubMed

    Mendelson, D A; Heinsbergen, J F; Kennedy, S D; Szczepaniak, L S; Lester, C C; Bryant, R G

    1991-01-01

    Measurements of the magnetic field dependence of spin-lattice relaxation rates and the response of the water-proton signal intensity to off-resonance radio frequency fields show that the commonly used agarose phantom provides a less faithful representation for the magnetic response of tissue than does a cross-linked protein system. The origin of these differences lies in the structure and intramolecular dynamics of the macromolecular system used to make the gel. These distinctions will also cause differences in the magnetic response of the water spin system when paramagnetic relaxation agents or contrast agents are incorporated. Use of a thermally cross-linked bovine serum albumin phantom is suggested. PMID:1766326

  14. The enhanced stability of the cross-linked hylan structure to hydroxyl (OH) radicals compared with the uncross-linked hyaluronan

    NASA Astrophysics Data System (ADS)

    Al-Assaf, Saphwan; Phillips, Glyn O.; Deeble, D. J.; Parsons, Barry; Starnes, Hazel; Von Sonntag, C.

    1995-08-01

    A comparison has been made of the relative stabilities of hyaluronan and hylan to degradation by OH radicals produced by ?-irradiation of aqueous solutions in N 2O, when G (yield per 100 eV) for OH radicals is 5.6 and H atoms 0.6. Using low angle light scattering and viscometric methods, the change in molecular weight of the polysaccharides was measured with increasing dose. From the yield/dose curves (expressed as breaks per molecule), the initial G value for hyaluronan degradation is 4. A further slow post-irradiation decrease in molecular weight is observed, which can be brought to completion by incubating the solutions for 1 h at 60C. Thereafter, the G value for degradation is 6. A similar post-irradiation degradation was found for hylan. A technique using tetranitromethane (TNM) has been used to distinguish between two types of radicals formed on the hyaluronan backbone. Radicals of the 1-hydroxy-2-alkoxy type (C-2, C-4, C-2 and C3 of the glucuronic acid) would induce strand breakage by alkoxy elimination. For the equivalent alkoxy radical at C6 of the acetamido monosaccharide, ring opening would occur with formation of a hemi-acetal, leading also to strand breakage. The C-2 and C-3 radicals would eliminate water rather than produce breaks by ?-alkoxy elimination. Thus three out of the initially formed radicals would produce breaks by ?-alkoxy formation. These can be stabilised with TNM and distinguished. It is concluded that these are the radicals involved in the post-irradiation thermal degradation process. Comparison of hylan and hyaluronan is, therefore, most valid when this post-irradiation process has been completed. Therefore, all G values for degradation were measured after incubation for 1 h at 60. This investigation establishes the greater stability of hylan ( Gdegradation = 2) compared to hylan ( Gdegradation = 6). Therefore, in an environment such as supplementation of an inflammed joint where OH radicals are released, hylan is able to retain its integrity as a viscoelastic macromolecule three times better than hyaluronan. Its potential as a viscosupplementation material, or as an inflammatory drug release matrix inserted within the joint is, therefore, greater than non-cross-linked hyaluronan.

  15. 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. PMID:12454437

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

    PubMed Central

    Hall, Kristina K.; Gatts-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

  17. 1,4-Butanediol diglycidyl ether-cross-linked hyaluronan inhibits fibrosis in rat primary tenocytes by down-regulating autophagy modulation.

    PubMed

    Hsu, Dur-Zong; Jou, I-Ming

    2016-05-01

    Epidural fibrosis, an inevitable part of the postoperative healing process, is one of the important causes of failed back surgery syndrome after spinal surgery. The aim of this study was to examine the inhibitory effect of a novel material 1,4-butanediol diglycidyl ether-cross-linked hyaluronan (cHA) on fibrosis in primary tenocytes. cHA inhibited migration, cell proliferation, and suppressed the expression of fibronectin, but not transforming growth factor-β, in primary tenocytes. cHA significantly increased matrix metalloproteinase-3 but decreased collagen-1 and microtubule-associated protein light chain 3-II expression in a dose-dependent manner compared with control groups. We therefore concluded that suppressing autophagy activity may be involved in the anti-fibrotic effect of cHA in primary tenocytes. Further, cHA may have the potential for preventing epidural fibrosis and subsequent failed back syndrome in patients with laminectomy in the future. PMID:26968759

  18. Citrate cross-linked gels with strain reversibility and viscoelastic behavior accelerate healing of osteochondral defects in a rabbit model.

    PubMed

    Ghosh, Paulomi; Rameshbabu, Arun Prabhu; Dhara, Santanu

    2014-07-22

    Most living tissues are viscoelastic in nature. Self-repair due to the dissipation of energy by reversible bonds prevents the rupture of the molecular backbone in these tissues. Recent studies, therefore, have aimed to synthesize biomaterials that approximate the mechanical performance of biological materials with self-recovery properties. We report an environmentally friendly method for the development of ionotropically cross-linked viscoelastic chitosan gels with a modulus comparable to that of living tissues. The strain recovery property was found to be highest for the gels with the lowest cross-linking density. The force-displacement curve showed significant hysteresis due to the presence of reversible bonds in the cross-linked gels. Nanoindentation studies demonstrated the creep phenomenon for the cross-linked chitosan gels. Creep, hysteresis, and plasticity index confirmed the viscoelastic behavior of the cross-linked gels. The viscoelastic gels were implanted at osteochondral defect sites to assess the tissue regeneration ability. In vivo results demonstrated early cartilage formation and woven bone deposition for defects filled with the gels compared to nontreated defects. PMID:24971647

  19. Generalized structural description of calcium-sodium aluminosilicate hydrate gels: the cross-linked substituted tobermorite model.

    PubMed

    Myers, Rupert J; Bernal, Susan A; San Nicolas, Rackel; Provis, John L

    2013-04-30

    Structural models for the primary strength and durability-giving reaction product in modern cements, a calcium (alumino)silicate hydrate gel, have previously been based solely on non-cross-linked tobermorite structures. However, recent experimental studies of laboratory-synthesized and alkali-activated slag (AAS) binders have indicated that the calcium-sodium aluminosilicate hydrate [C-(N)-A-S-H] gel formed in these systems can be significantly cross-linked. Here, we propose a model that describes the C-(N)-A-S-H gel as a mixture of cross-linked and non-cross-linked tobermorite-based structures (the cross-linked substituted tobermorite model, CSTM), which can more appropriately describe the spectroscopic and density information available for this material. Analysis of the phase assemblage and Al coordination environments of AAS binders shows that it is not possible to fully account for the chemistry of AAS by use of the assumption that all of the tetrahedral Al is present in a tobermorite-type C-(N)-A-S-H gel, due to the structural constraints of the gel. Application of the CSTM can for the first time reconcile this information, indicating the presence of an additional activation product that contains highly connected four-coordinated silicate and aluminate species. The CSTM therefore provides a more advanced description of the chemistry and structure of calcium-sodium aluminosilicate gel structures than that previously established in the literature. PMID:23534827

  20. Enzymatic Cross-Linking of Alkali Extracted Arabinoxylans: Gel Rheological and Structural Characteristics

    PubMed Central

    Berlanga-Reyes, Claudia M.; Carvajal-Millan, Elizabeth; Lizardi-Mendoza, Jaime; Islas-Rubio, Alma R.; Rascón-Chu, Agustin

    2011-01-01

    Ferulated arabinoxylans were alkali-extracted from wheat bran at different incubation times (0.0, 0.5, 1.0, 1.5 and 2.0 h). Wheat bran ferulated arabinoxylans (WBAX) arabinose-to-xylose ratio, ferulic acid content, intrinsic viscosity and viscosimetric molecular weight values decreased as the incubation time of extraction increased. WBAX enzymatic cross-linking capability was affected by incubation time while an increase in WBAX concentration from 5 to 6% (w/v) favored gelation. The WBAX gels formed presented a macroporous structure with mesh size ranging from 40 to 119 nm and hardness values varying from 1.7 to 5 N. PMID:22016631

  1. Austin Chalk: cross-link gel water, slick water fracs common in Giddings field

    SciTech Connect

    Mickey, V.

    1981-05-01

    Of the approx. 600 wells drilled in the Giddings field, Austin Chalk trend in the last year, most were probably completed by cross-link gel fracs or slick water fracs. Although open hole completions are not uncommon in the chalk, they are not really practical in most situations, given the 600 to 800 ft of gross pay usually encountered in the Austin Chalk trend. Most of the open hole completions occur when an operator takes a kick while drilling and decides to drop a mud pill over the zone, run casing above it, cement it in and produce the interval open hole. Chalk wells are subject to kick at any time during drilling because of natural fracturing systems throughout the trend. Mud weights are generally fairly light for faster drilling, approx. 9.4 to 9.8 lb, depending on the particular area.

  2. Cross-linked gel polymer electrolyte containing multi-wall carbon nanotubes for application in dye-sensitized solar cells

    NASA Astrophysics Data System (ADS)

    Benedetti, Joo E.; Corra, Aline A.; Carmello, Mayara; Almeida, Luiz C. P.; Gonalves, Agnaldo S.; Nogueira, Ana F.

    2012-06-01

    A cross-linked gel polymer electrolyte composed of poly (ethylene oxide-co-2-(2-methoxyethoxy) ethyl glycidyl ether-co-allyl glycidyl ether, ?-butyrolactone, LiI and I2, mixed with functionalized multi-wall carbon nanotubes (MWCNT), was applied in dye-sensitized solar cells (DSSC). The electrolyte was characterized by differential scanning calorimetry, conductivity measurements and Raman spectroscopy. Due to its nature, the terpolymer was chemically cross-linked during DSSC assembly to give rise to quasi-solid state solar cells. The gel polymer electrolyte containing 1 wt% of MWCNT exhibited conductivity values higher than 10-3 S cm-1. An increase in MWCNT content provided a smaller concentration of polyiodide species, as confirmed by Raman spectroscopy. The highest photocurrent (8.74 mA cm-2) was achieved by DSSC based on the cross-linked gel polymer electrolyte containing 1 wt% of MWCNT compared to DSSC based on non-cross-linked gel polymer electrolytes (7.93 mA cm-2). The incorporation of 3 wt% of MWCNT into the gel polymer electrolyte promoted a decrease in both Jsc and Voc, probably due to poorer light transmittance of the electrolyte in the visible range and higher charge recombination losses, respectively. Thus, the highest efficiency in this work was achieved by using electrolyte containing 1 wt% of MWCNT. After the cross-linking process, the dimensional stability of the gel composite electrolyte was improved and the conversion efficiency of DSSC was only slightly affected, changing from 3.37% (before cross-linking) to 3.35%.

  3. 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 165103, 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 2035% decrease in the amount of soluble protein with a Mr of 40165 (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 15103 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

  4. Cooperative effect of stress and ion displacement on the dynamics of cross-link unzipping and rupture of alginate gels.

    PubMed

    Baumberger, T; Ronsin, O

    2010-06-14

    We study the effect of nonbinding Na(+) ions on the kinetics of rupture of alginate gels cross-linked by Ca(2+). Wetting a crack tip with a saline solution at physiological concentrations is found to be able to induce a quasi-instantaneous, 10-fold velocity jump. This effect is analyzed with a phenomenological model for the rate-dependent fracture energy in physical gels, extended here to account for the role of ions on the rate of cross-link "unzipping". Ionic interaction is found to act cooperatively with mechanical tension, leading to an enhanced rate of rupture. The kinetics turns out to be second order in counterion concentration. The definition of the reference state requires to take into account counterion condensation due to long-range interactions in the polyelectrolyte gel. Surprisingly, the contribution of the Na(+) ions to the free energy of the activated state is essentially entropic, suggesting that the displacement of Ca(2+) is primarily a steric process, electrostatic interactions being reduced to the constraint of charge conservation. This phenomenon may have important consequences on the rate of degradation of alginate based scaffolds for in vivo tissue regeneration. PMID:20499914

  5. Preparation of porous, chemically cross-linked, PVdF-based gel polymer electrolytes for rechargeable lithium batteries

    NASA Astrophysics Data System (ADS)

    Cheng, C. L.; Wan, C. C.; Wang, Y. Y.

    This study reports the development of a new system of porous, chemically cross-linked, gel polymer electrolytes based on poly(vinylidene fluoride-co-hexafluoropropylene) (PVdF-HFP) copolymer as a polymer matrix, polyethylene glycol (PEG) as a plasticizer, and polyethylene glycol dimethacrylate (PEGDMA) as a chemical cross-linking oligomer. The electrolytes are prepared by a combination of controlled evaporation and thermal polymerization of PEGDMA. PVdF-HFP/PEG/PEGDMA gel polymer electrolytes with a composition of 5/3/2 exhibit both high ambient ionic conductivity, viz., >1 mS cm -1, and a high tensile modulus of 52 MPa, because of their porous and network structures. All the blends of electrolytes are electrochemically stable up to 5 V versus Li/Li + in the presence of 1 M LiPF 6/ethylene carbonate-diethyl carbonate (EC-DEC). With these polymer electrolytes, rechargeable lithium batteries composed of carbon anode and LiCoO 2 cathode have acceptable cycleability and a good rate capability.

  6. Cross-linked redox gels containing glucose oxidase for amperometric biosensor applications

    SciTech Connect

    Gregg, B.A.; Heller, A. )

    1990-02-01

    Oxidoreductases, such as glucose oxidase, can be electrically wired to electrodes by electrostatic complexing or by covalent binding of redox polymers so that the electrons flow from the enzyme, through the polymer, to the electrode. We describe two materials for amperometric biosensors based on a cross-linkable poly(vinylpyridine) complex of (Os-(bpy){sub 2}Cl){sup +/2+} that communicates electrically with flavin adenine dinucleiotide redox centers of enzymes such as glucose oxidase. The uncomplexed pyridines of the poly(vinylpyridine) are quaternized with two types of groups, one promoting hydrophilicity (2-bromoethanol or 3-bromopropionic acid), the other containing an active ester (N-hydroxysuccinimide) that forms amide bonds with both lysines on the enzyme surface and with an added polyamine cross-linking agent (tri-ethylenetetraamine, trien). In the presence of glucose oxidase and trien this polymer forms rugged, cross-linked, electroactive films on the surface of electrodes, thereby eliminating the requirement for a membrane for containing the enzyme and redox couple. The glucose response time of the resulting electrodes is less than 10 s. The glucose response under N{sub 2} shows an apparent Michaelis constant, K{sub m}{prime} = 7.3 mM, and limiting current densities, j{sub max}, between 100 and 800 {mu}A/cm{sup 2}. Currents are decreased by 30-50% in air-saturated solutions because of competition between O{sub 2} and the Os(III) complex for electrons from the reduced enzyme. Rotating ring disk experiments in air-saturated solutions containing 10 mM glucose show that about 20% of the active enzyme is electrooxidized via the Os(III) complex, while the rest is oxidized by O{sub 2}. These results suggest that only part of the active enzyme is in electrical contact with the electrode.

  7. Design principles for cytokine-neutralizing gels: Cross-linking effects

    PubMed Central

    Sun, Liang Tso; Bencherif, Sidi A.; Gilbert, Thomas W.; Lotze, Michael T.; Washburn, Newell R.

    2012-01-01

    Constructs composed of cytokine-neutralizing antibodies conjugated to high-molecular-weight hyaluronic acid have been shown to be effective at controlling inflammatory responses in vivo. A critical question in the development of this new class of biomaterial is whether crosslinked conjugates have similar anti-inflammatory effects, which would open up a broad range of tissue engineering applications in which the material would have intrinsic inflammation-controlling function. To test this, high-molecular-weight hyaluronic acid was conjugated with monoclonal antibodies to the pro-inflammatory cytokines interleukin-1? and tumor necrosis factor-? in two forms of the material: viscous, non-crosslinked polymerantibody conjugates and crosslinked, elastomeric polymerantibody conjugates. The cytokine affinities of both constructs were validated using molecular characterization methods, and the biological activities were tested through subcutaneous implantation in SpragueDawley rats. In vitro, both forms of these constructs are capable of binding cytokines, but in vivo only the non-crosslinked polymer significantly reduces markers of acute inflammation compared to controls that lack the antibodies. We propose that these materials function by retarding cytokine diffusion, with the non-crosslinked polymers being capable of retarding the diffusion of cytokines in the extracellular matrix and preventing engagement with receptors. In contrast, crosslinked materials have long diffusion lengths into the gel compared with those between cells on the surface of the material, which may make them ineffective at sequestering pro-inflammatory cytokines on biologically relevant timescales. These results suggest an important design principle for preparing cytokine-regulating materials based on consideration of transport phenomena. PMID:20601239

  8. Topical diclofenac in hyaluronan gel for the treatment of solar keratoses.

    PubMed

    Gebauer, Kurt; Brown, Pam; Varigos, George

    2003-02-01

    This randomized, double-blind, placebo-controlled study assessed the efficacy and safety of a topical gel containing 3% diclofenac in 2.5% hyaluronan in 150 patients with solar keratoses (SK). The active treatment was compared with the vehicle only, hyaluronan gel, as placebo over a 12-week period. Patients in both groups applied the active treatment or placebo to a targeted area of skin (0.25 g b.d.). At 12 weeks the mean lesion-count reduction in the targeted area was not significantly different between treatments. However, at post-termination follow up (16 weeks), there was a highly significant decrease in the number of lesions, 6.2 +/- 7.5 standard deviations (SD) (56.1% reduction) in the active treatment group compared with 2.4 +/- 4.3 SD (23.6% reduction) in the placebo group (P < 0.001). Other efficacy measures (complete lesion resolution, >50% lesion reduction) were also significantly different (P < 0.01) between treatments at 16 weeks. In conclusion, topical 3% diclofenac in 2.5% hyaluronan gel was effective and well tolerated in this study, suggesting a role for this therapy in the treatment of SK. PMID:12581080

  9. Applicability of a particularly simple model to nonlinear elasticity of slide-ring gels with movable cross-links as revealed by unequal biaxial deformation

    NASA Astrophysics Data System (ADS)

    Kondo, Yuuki; Urayama, Kenji; Kidowaki, Masatoshi; Mayumi, Koichi; Takigawa, Toshikazu; Ito, Kohzo

    2014-10-01

    The strain energy density function (F) of the polyrotaxane-based slide-ring (SR) gels with movable cross-links along the network strands is characterized by unequal biaxial stretching which can achieve various types of deformation. The SR gels as prepared without any post-preparation complication exhibit considerably smaller values of the ratio of the stresses (?y/?x) in the stretched (x) and constrained (y) directions in planar extension than classical chemical gels with heterogeneous and nearly homogeneous network structures do. This feature of the SR gels leads to the peculiar characteristic that the strain energy density function (F) has no explicit cross term of strains in different directions, which is in contrast to F with explicit strain cross terms for most chemical gels and elastomers. The biaxial stress-strain data of the SR gels are successfully described by F of the Gent model with only two parameters (small-strain shear modulus and a parameter representing ultimate elongation), which introduces the finite extensibility effect into the neo-Hookean model with no explicit cross term of strain. The biaxial data of the deswollen SR gels examined in previous study, which underwent a considerable reduction in volume from the preparation state, are also well described by the Gent model, which is in contrast to the case of the classical chemical gels that the stress-strain relations before and after large deswelling are not described by a common type of F due to a significant degree of collapse of the network strands in the deswollen state. These intriguing features of nonlinear elasticity of the SR gels originate from a novel function of the slidable cross-links that can maximize the arrangement entropy of cross-linked and non-cross-linked cyclic molecules in the deformed networks.

  10. Cross-Linking the Fibers of Supramolecular Gels Formed from a Tripodal Terpyridine Derived Ligand with d-Block Metal Ions.

    PubMed

    Kotova, Oxana; Daly, Ronan; dos Santos, Cidália M G; Kruger, Paul E; Boland, John J; Gunnlaugsson, Thorfinnur

    2015-08-17

    The tripodal terpyridine ligand, L, forms 1D helical supramolecular polymers/gels in H2O-CH3OH solution mediated through hydrogen bonding and π-π interactions. These gels further cross-link into 3D supramolecular metallogels with a range of metal ions (M) such as Fe(II), Ni(II), Cu(II), Zn(II), and Ru(III); the cross-linking resulting in the formation of colored or colorless gels. The fibrous morphology of these gels was confirmed using scanning electron microscopy (SEM); while the self-assembly processes between L and M were investigated by absorbance and emission spectroscopy from which their binding constants were determined by using a nonlinear regression analysis. PMID:26222397

  11. Case study: Hydraulic fracturing with cross-linked gels in the Oriskany formation to improve gas storage deliverability

    SciTech Connect

    Stiles, E.K.; Reese, R.R.

    1995-12-31

    During the summer and fall of 1994, CNG Transmission Corporation fracture-stimulated thirty (30) gas storage wells with a cross-linked gel fluid system. The work was done to increase late season field deliverability. Individual well tests taken to date have shown that, on average, per-well deliverability has increased by a factor of five. The text that follows will detail the engineering and geologic aspects, procedures, results, and analysis of the project. A discussion of the strategic placement of the storage fields involved will be presented, along with a review of FERC Order 636 and it`s impact on the current gas storage business mechanics. Overviews are presented on the geology, reservoir properties, and characteristics of the storage fields involved in the project. General well design and well completion configuration are discussed. Fracturing/refracturing candidate selection criteria are presented. Basic fracturing design criteria are discussed including: fluid parameters, geometry, proppant selection, and concentration. Discussions of observations and minor changes in treatment designs which occurred between the different fields are discussed. The treatments are summarized and compared on a pre and post fracturing deliverability test analysis. Explanations are offered for wells with exceptional results, as well as marginally successful treatments.

  12. 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 -1 to 0.36 -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 as evidenced by a reduction in intensity in the lower Q region.

  13. Cross-linking properties of alginate gels determined by using advanced NMR imaging and Cu(2+) as contrast agent.

    PubMed

    Manz, B; Hillgrtner, M; Zimmermann, H; Zimmermann, D; Volke, F; Zimmermann, U

    2004-02-01

    The entrapment of enzymes, drugs, cells or tissue fragments in alginates cross-linked with Ca(2+) or Ba(2+) has great potential in basic research, biotechnology and medicine. The swelling properties and, in turn, the mechanical stability are key factors in designing an optimally cross-linked hydrogel matrix. These parameters depend critically on the cross-linking process and seemingly minor modifications in manufacture have a large impact. Thus, sensitive and non-invasive tools are required to determine the spatial homogeneity and efficacy of the cross-linking process. Here, we show for alginate microcapsules (between 400 microm and 600 microm in diameter) that advanced (1)H NMR imaging, along with paramagnetic Cu(2+) as contrast agent, can be used to validate the cross-linking process. Two- and three-dimensional images and maps of the spin-lattice relaxation time T(1) of Ba(2+) cross-linked microcapsules exposed to external Cu(2+) yielded qualitative as well as quantitative information about the accumulation of Cu(2+) within and removal from microcapsules upon washing with Cu(2+) free saline solution. The use of Cu(2+) (having a slightly higher affinity constant to alginate than Ba(2+)) for gelling gave a complementary insight into the spatial homogeneity of the cross-linking process together with information about the mechanical stability of the microcapsules. The potential of this technique was demonstrated for alginates extracted from two different algal sources and cross-linked either externally by the conventional air-jet dropping method or internally by the "crystal gun" method. PMID:13680210

  14. CO2/light gas separation performance of cross-linked poly(vinylimidazolium) gel membranes as a function of ionic liquid loading and cross-linker content

    SciTech Connect

    Carlisle, TK; Nicodemus, GD; Gin, DL; Noble, RD

    2012-04-15

    A series of cross-linked poly(vinylimidazolium)-RTIL gel membranes was synthesized and evaluated for room-temperature, ideal CO2/N-2, CO2/CH4, and CO2/H-2 separation performance. The membranes were formed by photo-polymerization of oligo(ethylene glycol)-functionalized cross-linking (i.e., di-functional) and non-cross-linking (i.e., mono-functional) vinylimidazolium RTIL monomers with nonpolymerizable, "free RTIL." The effect of free RTIL ([emim][Tf2N]) loading on CO2 separation performance was evaluated by varying RTIL loading at three levels (45, 65, and 75 wt.%). The effect of cross-linker content on CO2 separation performance was also evaluated by varying the copolymer composition of cross-linked membranes from 5 to 100 mol% di-functional monomer. The substituent on the monofunctional RTIL monomer was also varied to investigate the effect of substituent structure and chemistry on CO2 separation performance. CO2 permeability was dramatically increased with higher loading of free RTIL. Increased RTIL loading had no effect on CO2/N-2 or CO2/CH4 permeability selectivity, but significantly improved CO2/H-2 permeability selectivity. Reducing the cross-linking monomer concentration generally improved CO2 permeability. However, anomalous permeability and selectivity behavior was observed below critical concentrations of cross-linker. The effect of the substituent on the monofunctional monomer on CO2 separation performance was minimal compared to the effects of RTIL loading and copolymer composition. (C) 2012 Elsevier B.V. All rights reserved.

  15. Gel electrophoretic studies of photochemical cross-linking of type I collagen with brominated 1,8-naphthalimide dyes and visible light

    NASA Astrophysics Data System (ADS)

    Judy, Millard M.; Fuh, L.; Matthews, James Lester; Lewis, David E.; Utecht, Ronald E.

    1994-09-01

    Insoluble Type I collagen from bovine Achilles tendon (Sigma C9879) was suspended in a 3 mM solution of the dye diEd66Br dissolved in Cremophor ELR (BASF) to give a molecular concentration ratio. Fifty-microliter aliquots in 5-mm-diameter wells were exposed to 458 J/cm2 (225 mW/cm2, 1800 sec) of 457.9-nm light from an argon ion laser; similar aliquots with and without dye were kept in the dark to serve as controls. Following pelleting of the collagen by centrifugation and 3x washing in phosphate-buffered saline, aliquots of light-treated and control sample pellets were (1) digested in collagenase (Sigma C9891) or (2) extracted in 0.5 M acetic acid, followed by centrifugative ultrafiltration (10-kd cutoff) in 0.01 M acetic acid. Aliquots of the supernatant of the acid-extracted collagen also were digested in pepsin. Electrophoretic protein migration in 8% to 25% gradient polyacrylamide gels following SDS solubilization disclosed numerous, densely packed, essentially contiguous protein bands. These studies indicate that the dye and light treatment of insoluble Type I collagen (1) results in cross-linking of collagen molecules and (2) does not denature the trimer conformation sufficiently to enable significant digestion by pepsin.

  16. Chemical cross-linking of Chlamydia trachomatis.

    PubMed Central

    Birkelund, S; Lundemose, A G; Christiansen, G

    1988-01-01

    Purified elementary bodies (EBs) of Chlamydia trachomatis serovar L2 were analyzed by chemical cross-linking with disuccinimidyl selenodipropionate. The effect of the cross-linking was analyzed by immunoblotting sodium dodecyl sulfate-polyacrylamide gel electrophoresis-separated components which were reacted with monoclonal antibodies against major outer membrane protein (MOMP) and lipopolysaccharide (LPS). It was shown that in EBs, MOMP was cross-linked to the LPS component of the outer membrane. Migration analysis of the cross-linked components showed that with extensive cross-linking, most of the MOMP became cross-linked to LPS, changing the migration rate from 40 to 42.5 kilodaltons. A small fraction of MOMP associated with LPS was shown to be present in bands with migration rates of 100 and 110 kilodaltons. No association of MOMP or LPS to other proteins, or to dimer or multimer forms of MOMP without LPS, was observed. A totally different membrane structure must be present in reticulate bodies, since there, MOMP was so heavily cross-linked that it did not enter the polyacrylamide gel and thus became impossible to analyze. Furthermore, the monoclonal antibody, which reacted with LPS associated with MOMP in the cross-linked EBs, did not react with reticulate bodies. Images PMID:2449399

  17. Corneal cross-linking.

    PubMed

    Randleman, J Bradley; Khandelwal, Sumitra S; Hafezi, Farhad

    2015-01-01

    Since its inception in the late 1990s, corneal cross-linking has grown from an interesting concept to a primary treatment for corneal ectatic disease worldwide. Using a combination of ultraviolet-A light and a chromophore (vitamin B2, riboflavin), the cornea can be stiffened, usually with a single application, and progressive thinning diseases such as keratoconus arrested. Despite being in clinical use for many years, some of the underlying processes, such as the role of oxygen and the optimal treatment times, are still being worked out. More than a treatment technique, corneal cross-links represent a physiological principle of connective tissue, which may explain the enormous versatility of the method. We highlight the history of corneal cross-linking, the scientific underpinnings of current techniques, evolving clinical treatment parameters, and the use of cross-linking in combination with refractive surgery and for the treatment of infectious keratitis. PMID:25980780

  18. Application of NMR spectroscopy and multidimensional imaging to the gelcasting process and in-situ real-time monitoring of cross-linking polyacrylamide gels

    SciTech Connect

    Ahuja, S.; Dieckman, S.L.; Gopalsami, N.

    1995-04-01

    In the gelcasting process, a slurry of ceramic powder in a solution of organic monomers is cast in a mold. The process is different from injection molding in that it separates mold-filling from setting during conversion of the ceramic slurry to a formed green part. In this work, NMR spectroscopy and imaging have been conducted for in-situ monitoring of the gelation process and for mapping the polymerization. {sup 1}H nuclear magnetic resonance spectra have been obtained during polymerization of a premix of soluble reactive methacrylamide (monomer) and N, N`-methylene bisacrylamide (cross-linking molecules). The premix was polymerized by adding ammonium persulfate (initiator) and tetramethyl-ethylene-diamine (accelerator) to form long-chain, cross-linked polymers. The time-varying spin-lattice relaxation times T{sub 1} during polymerization have been studied at 25 and 35{degrees}C, and the variation of spectra and T{sub 1} with respect to extent of polymerization has been determined. To verify homogeneous polymerization, multidimensional NMR imaging was utilized for in-situ monitoring of the process. The intensities from the images are modeled and the correspondence shows a direct extraction of T{sub 1} data from the images.

  19. Topologically controlled hyaluronan-based gel coatings of hydrophobic grid-like scaffolds to modulate drug delivery.

    PubMed

    Arnal-Pastor, M; Pérez-Garnes, M; Monleón Pradas, M; Vallés Lluch, A

    2016-04-01

    Scaffolds based on poly(ethyl acrylate) having interwoven channels were coated with a hyaluronan (HA) hydrogel to be used in tissue engineering applications. Controlled typologies of coatings evolving from isolated aggregates to continuous layers, which eventually clog the channels, were obtained by using hyaluronan solutions of different concentrations. The efficiency of the HA loading was determined using gravimetric and thermogravimetric methods, and the hydrogel loss during the subsequent crosslinking process was quantified, seeming to depend on the mass fraction of hyaluronan initially incorporated to the pores. The effect of the topologically different coatings on the scaffolds, in terms of mechanical properties and swelling at equilibrium under different conditions was evaluated and correlated with the hyaluronan mass fraction. The potential of these hydrogel coatings as vehicle for controlled drug release from the scaffolds was validated using a protein model. PMID:26780254

  20. 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. PMID:26426303

  1. Kinetics of the swelling of a gel of cross-linked polyvinyl alcohol during the synthesis of copper-containing composite based on it

    NASA Astrophysics Data System (ADS)

    Khudyakova, S. N.; Tokmachev, M. G.; Ferapontov, N. B.

    2013-07-01

    The kinetics of processes that occur during the chemical reduction of copper in a gel of polyvinyl alcohol (PVA) are described. It is shown that we can assess the processes occurring in the polymer from changes in the degree of swelling. The effects the nature of the precursor (precursor compound) has on the conditions of reduction are analyzed. It is found that crystals of reduced copper do not change the volume of PVA gel, which has a positive effect on the mechanical stability of the resulting composite.

  2. Radiation cross-linked carboxymethyl sago pulp hydrogels loaded with ciprofloxacin: Influence of irradiation on gel fraction, entrapped drug and in vitro release

    NASA Astrophysics Data System (ADS)

    Lam, Yi Lyn; Muniyandy, Saravanan; Kamaruddin, Hashim; Mansor, Ahmad; Janarthanan, Pushpamalar

    2015-01-01

    Carboxymethyl sago pulp (CMSP) with 0.4 DS, viscosity 184 dl/g and molecular weight 76,000 g/mol was synthesized from sago waste. 10 and 20% w/v solutions of CMSP were irradiated at 10-30 kGy to form hydrogels and were characterized by % gel fraction (GF). Irradiation of 20% CMSP using 25 kGy has produced stable hydrogels with the highest % GF and hence loaded with ciprofloxacin HCl. Drug-loaded hydrogels were produced by irradiating the mixture of drug and 20% CMSP solution at 25 kGy. After irradiation, the hydrogels were cut into circular discs with a diameter of 6±1 mm and evaluated for physicochemical properties as well as drug release kinetics. The ciprofloxacin loading in the disc was 14.7%±1 w/w with an entrapment efficiency of 73.5% w/w. The low standard deviation of drug-loaded discs indicated uniform thickness (1.5±0.3 mm). The unloaded discs were thinner (1±0.4 mm) and more brittle than the drug-loaded discs. FESEM, FT-IR, XRD, DSC and TGA analysis revealed the absence of polymer-drug interaction and transformation of crystalline to amorphous form of ciprofloxacin in the discs. The disc sustained the drug release in phosphate buffer pH 7.4 over 36 h in a first-order manner. The mechanism of the drug release was found to be swelling controlled diffusion and matrix erosion. The anti-bacterial effect of ciprofloxacin was retained after irradiation and CMSP disc could be a promising device for ocular drug delivery.

  3. Effects of shear and proppant on the viscosity of cross-linked fracturing fluids

    SciTech Connect

    Gardner, D.C.; Eikerts, J.V.

    1982-01-01

    Several frequently used cross-linked fracturing fluids (gels) have been evaluated as to the effects that shear and 20/40 mesh sand proppant have on their rheological properties. A closed-loop pipe viscometer was used to evaluate the shear resistance and post-shear recovery of various gels. The aqueous-base gels tested included those made with guar, hydroxypropyl guar (HPG), and carboxymethyl hydroxyethyl cellulose (CMHEC) polymers. Test results indicate that when cross-linked with borate ions, guar and HPG gels have the greatest resistance to shear. Yet when cross-linked with various titanates, HPG gels showed significantly lower resistance to induced shear. Tests on CMHEC gels which were cross-linked with trivalent aluminum (Al/sup +3/) indicate a shear resistance between that of the HPG-borate gels and the CMHEC-Al/sup +3/ gels demonstrated considerably higher post-shear recoveries than those attained with HPG-titanate gels.

  4. Irradiated cross-linked wire

    NASA Astrophysics Data System (ADS)

    Uda, I.; Ueno, K.; Tada, S.; Suzuki, S.

    Generally speaking, polyurethane has excellent mechanical properties but poor water resistance. It is rather difficult to obtain flame retardant polyurethane while maintaining water resistance. We ascertained that improvement in water resistance could be achieved on polyurethane by high energy electron beam irradiation. The irradiated cross-linked polyurethane is very useful as jacketing material for automobile cables.

  5. Cross-linking chitosan nanofibers.

    PubMed

    Schiffman, Jessica D; Schauer, Caroline L

    2007-02-01

    In the present study, we have electrospun various grades of chitosan and cross-linked them using a novel method involving glutaraldehyde (GA) vapor, utilizing a Schiff base imine functionality. Chemical, structural, and mechanical analyses have been conducted by Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), and Kawabata microtensile testing, respectively. Additionally, the solubilities of the as-spun and cross-linked chitosan mats have been evaluated;solubility was greatly improved after cross-linking. SEM images displayed evidence that unfiltered low, medium, and high molecular weight chitosans, as well as practical-grade chitosan, can be electrospun into nanofibrous mats. The as-spun medium molecular weight chitosan nanofibers have a Young's modulus of 154.9 +/- 40.0 MPa and display a pseudo-yield point that arose due to the transition from the pulling of a fibrous mat with high cohesive strength to the sliding and elongation of fibers. As-spun mats were highly soluble in acidic and aqueous solutions. After cross-linking, the medium molecular weight fibers increased in diameter by an average of 161 nm, have a decreased Young's modulus of 150.8 +/- 43.6 MPa, and were insoluble in basic, acidic, and aqueous solutions. Though the extent to which GA penetrates into the chitosan fibers is currently unknown, it is evident that the cross-linking resulted in increased brittleness, a color change, and the restriction of fiber sliding that resulted in the loss of a pseudo-yield point. PMID:17291083

  6. Collagen cross linking: current perspectives.

    PubMed

    Rao, Srinivas K

    2013-08-01

    Keratoconus is a common ectatic disorder occurring in more than 1 in 1,000 individuals. The condition typically starts in adolescence and early adulthood. It is a disease with an uncertain cause and its progression is unpredictable, but in extreme cases, vision deteriorates and can require corneal transplant surgery. Corneal collagen cross-linking (CCL) with riboflavin (C3R) is a recent treatment option that can enhance the rigidity of the cornea and prevent disease progression. Since its inception, the procedure has evolved with newer instrumentation, surgical techniques, and is also now performed for expanded indications other than keratoconus. With increasing experience, newer guidelines regarding optimization of patient selection, the spectrum of complications and their management, and combination procedures are being described. This article in conjunction with the others in this issue, will try and explore the uses of collagen cross-linking (CXL) in its current form. PMID:23925327

  7. Hyaluronan in human malignancies

    SciTech Connect

    Sironen, R.K.; Department of Pathology, Kuopio University Hospital, P.O. Box 1777, FI-70211 Kuopio ; Tammi, M.; Tammi, R.; Auvinen, P.K.; Anttila, M.; Department of Gynecology and Obstetrics, Kuopio University Hospital, P.O. Box 1777, FI-70211 Kuopio ; Kosma, V-M.

    2011-02-15

    Hyaluronan, a major macropolysaccharide in the extracellular matrix of connective tissues, is intimately involved in the biology of cancer. Hyaluronan accumulates into the stroma of various human tumors and modulates intracellular signaling pathways, cell proliferation, motility and invasive properties of malignant cells. Experimental and clinicopathological evidence highlights the importance of hyaluronan in tumor growth and metastasis. A high stromal hyaluronan content is associated with poorly differentiated tumors and aggressive clinical behavior in human adenocarcinomas. Instead, the squamous cell carcinomas and malignant melanomas tend to have a reduced hyaluronan content. In addition to the stroma-cancer cell interaction, hyaluronan can influence stromal cell recruitment, tumor angiogenesis and epithelial-mesenchymal transition. Hyaluronan receptors, hyaluronan synthases and hyaluronan degrading enzymes, hyaluronidases, are involved in the modulation of cancer progression, depending on the tumor type. Furthermore, intracellular signaling and angiogenesis are affected by the degradation products of hyaluronan. Hyaluronan has also therapeutic implications since it is involved in multidrug resistance.

  8. Microrheology of cross-linked polyacrylamide networks.

    PubMed

    Dasgupta, Bivash R; Weitz, D A

    2005-02-01

    Experiments investigating the local viscoelastic properties of a chemically cross-linked polymer are performed on polyacrylamide solutions in the sol and the gel regimes using polystyrene beads of varying sizes and surface chemistry as probes. The thermal motions of the probes are measured to obtain the elastic and viscous moduli of the sample. Probe dynamics are measured using two different dynamic light scattering techniques, diffusing wave spectroscopy (DWS) and quasielastic light scattering (QELS) as well as video-based particle tracking. Diffusing wave spectroscopy probes the short-time dynamics of the scatterers while QELS measures the dynamics at larger times. Video-based particle tracking provides a way to investigate the local environment of the individual probe particles. A combination of all the techniques results in a larger range of frequencies that can be probed compared to conventional bulk measurements while providing local information at the level of individual probes. A modified algebraic form of the generalized Stokes-Einstein equation is used to calculate the frequency-dependent moduli. A comparison of microrheological measurements with bulk rheology exhibits striking similarity, confirming the applicability of microrheology for chemically cross-linked polymeric systems. PMID:15783330

  9. Microrheology of cross-linked polyacrylamide networks

    NASA Astrophysics Data System (ADS)

    Dasgupta, Bivash R.; Weitz, D. A.

    2005-02-01

    Experiments investigating the local viscoelastic properties of a chemically cross-linked polymer are performed on polyacrylamide solutions in the sol and the gel regimes using polystyrene beads of varying sizes and surface chemistry as probes. The thermal motions of the probes are measured to obtain the elastic and viscous moduli of the sample. Probe dynamics are measured using two different dynamic light scattering techniques, diffusing wave spectroscopy (DWS) and quasielastic light scattering (QELS) as well as video-based particle tracking. Diffusing wave spectroscopy probes the short-time dynamics of the scatterers while QELS measures the dynamics at larger times. Video-based particle tracking provides a way to investigate the local environment of the individual probe particles. A combination of all the techniques results in a larger range of frequencies that can be probed compared to conventional bulk measurements while providing local information at the level of individual probes. A modified algebraic form of the generalized Stokes-Einstein equation is used to calculate the frequency-dependent moduli. A comparison of microrheological measurements with bulk rheology exhibits striking similarity, confirming the applicability of microrheology for chemically cross-linked polymeric systems.

  10. A method for the isolation of cross-linked nucleosides from DNA: application to cross-links induced by nitrous acid.

    PubMed

    Dubelman, S; Shapiro, R

    1977-06-01

    A procedure is reported for the isolation of cross-linked nucleosides from nitrous acid-treated calf thymus DNA. Cross-linked DNA was hydrolyzed enzymatically with deoxyribonuclease I and snake venom phosphodiesterase and fractionated on a DEAE-Sephadex column. After desalting, the fractions were characterized by ultraviolet spectroscopy, anion exchange high pressure liquid chromatography, gel filtration, and two dimensional thin layer chromatography. A cross-linked dinucleotide, and a series of oligonucleotides were isolated. The oligomers, which had resisted digestion by the above enzyme system, were digested to the nucleoside level by a spleen phosphodiesterase-alkaline phosphatase combination. A second cross-linked product was isolated from this mixture. The cross-linked nucleosides were less than 0.17% of the total nucleotides of the DNA. The methods developed here are recommended for the isolation of products from DNA treated with other cross-linking agents. PMID:197494

  11. Electrospinning formaldehyde cross-linked zein solutions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In order to develop zein fibers with improved physical properties and solvent resistance, formaldehyde was used as the cross-linking reagent before spinning. The cross-linking reaction was carried out in either acetic acid or ethanolic-HCl where the amount of cross-linking reagent was between 1 and...

  12. Cross-linking of dark-adapted frog photoreceptor disk membranes. Evidence for monomeric rhodopsin.

    PubMed Central

    Downer, N W

    1985-01-01

    A model for random cross-linking of identical monomers diffusing in a membrane was formulated to test whether rhodopsin's cross-linking behavior was quantitatively consistent with a monomeric structure. Cross-linking was performed on rhodopsin both in intact retinas and in isolated rod outer segment (ROS) membranes using the reagent glutaraldehyde. The distribution of covalent oligomers formed was analyzed by SDS-polyacrylamide gel electrophoresis and compared to predictions for the random model. A similar analysis was made for ROS membranes cross-linked by diisocyanatohexane and retinas cross-linked by cupric ion complexed with o-phenanthroline. Patterns of cross-linking produced by these three reagents are reasonably consistent with the monomer model. Glutaraldehyde was also used to cross-link the tetrameric protein aldolase in order to verify that cross-linking of a stable oligomer, under conditions comparable to those used for ROS, yielded the pattern predicted for a tetrameric protein having D2 symmetry. This pattern is markedly different from the one for a random-collision model. Moreover, a comparison of rates showed that aldolase cross-linking with glutaraldehyde is significantly faster than cross-linking of membrane-bound rhodopsin. It is concluded that rhodopsin is monomeric in dark-adapted photoreceptor membranes and that the observed cross-linking results from collisions between diffusing rhodopsin molecules. PMID:3919779

  13. Electrospun cross linked rosin fibers

    NASA Astrophysics Data System (ADS)

    Baek, Woo-il; Nirmala, R.; Barakat, Nasser A. M.; El-Newehy, Mohamed H.; Al-Deyab, Salem S.; Kim, Hak Yong

    2011-12-01

    In this study, we describe the first reported preparation of rosin in fiber form through use of an electrospinning technique utilizing various solvent systems. The polymer concentration of the formed fiber was studied by using various solvents such as chloroform, ethanol, N-N dimethylformamide (DMF), tetrahydrofuran (THF), acetone, and methylene chloride (MC). An electrospray of the solution resulted in the beaded form of the rosin. By varying the polymer concentration with MC, we were then able to obtain uniform fibers. However, the fibers exhibited large diameter. We believe that it is possible to reduce the diameter of the rosin fibers through appropriate selection of electrospinning parameters. In addition, the morphological transitions from beads, to beaded fiber, to fiber were studied at different polymer concentrations. We propose a possible physical cross linking mechanism for the formation of rosin fibers during the electrospinning process. Our results demonstrate the feasibility of producing fiber nanostructures of rosin by using an electrospinning technique.

  14. Molecular structure of citric acid cross-linked starch films.

    PubMed

    Menzel, Carolin; Olsson, Erik; Plivelic, Tomás S; Andersson, Roger; Johansson, Caisa; Kuktaite, Ramune; Järnström, Lars; Koch, Kristine

    2013-07-01

    The effect of citric acid (CA) on starch films has been examined. A new method to detect cross-linking of starch by CA in solution-cast films by molecular weight measurements is described. Furthermore, we managed to distinguished between free, mono- and di-esterified CA and quantify di-ester content within starch films by using a modification in the method of complexometric titration with copper(II)-sulfate. Cross-linking of starch by CA occurred at low temperature, 70°C, which we assumed is so far the lowest temperature reported where cross-linking reaction occurred. This is essential for starch coating applications within paper industry since no high temperatures for curing will be required. However, curing at 150°C and high CA concentrations, 30 pph, increased cross-linking reaction. Furthermore, the physical properties like water solubility, gel content and glass transition temperature, were highly reflected by changes in the molecular structure i.e. cross-linking and hydrolysis, as well as CA content and curing temperature. PMID:23688480

  15. Cross-Linked Antioxidant Nanozymes for Improved Delivery to CNS

    PubMed Central

    Klyachko, Natalia L.; Manickam, Devika S.; Brynskikh, Anna M.; Uglanova, Svetlana V.; Li, Shu; Higginbotham, Sheila M.; Bronich, Tatiana K.; Batrakova, Elena V.; Kabanov, Alexander V.

    2011-01-01

    Formulations of antioxidant enzymes, superoxide dismutase 1 (SOD1, also known as Cu/Zn SOD) and catalase were prepared by electrostatic coupling of enzymes with cationic block copolymers, polyethyleneimine-poly(ethylene glycol) or poly(L-lysine)-poly(ethylene glycol), followed by covalent cross-linking to stabilize nanoparticles. Different cross-linking strategies (using glutaraldehyde, bis-(sulfosuccinimidyl)suberate sodium salt or 1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride with N-hydroxysulfosuccinimide) and reaction conditions (pH and polycation/protein charge ratio) were investigated that allowed immobilizing active enzymes in cross-linked nanoparticles, termed nanozymes. Bi-enzyme nanoparticles, containing both SOD1 and catalase were also formulated. Formation of complexes was confirmed using denaturing gel electrophoresis and western blotting and physicochemical characterization was conducted using dynamic light scattering and atomic force microscopy. In vivo studies of 125I-labeled SOD1-containing nanozymes in mice demonstrated its increased stability in both blood and brain and increased accumulation in brain tissues, compared to non-cross-linked complexes and native SOD1. Future studies will evaluate potential of these formulations for delivery of antioxidant enzymes to central nervous system to attenuate oxidative stress associated with neurological diseases. PMID:21703990

  16. 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.

  17. 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.

  18. Highly cross-linked nanoporous polymers

    DOEpatents

    Steckle, Jr., Warren P. (Los Alamos, NM); Apen, Paul G. (Los Alamos, NM); Mitchell, Michael A. (Los Alamos, NM)

    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.

  19. Highly cross-linked nanoporous polymers

    DOEpatents

    Steckle, Jr., Warren P. (Los Alamos, NM); Apen, Paul G. (Los Alamos, NM); Mitchell, Michael A. (Los Alamos, NM)

    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.

  20. 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. PMID:24564765

  1. Characterization of Hyaluronan-Protein Microstructures and Polymer Solutions

    NASA Astrophysics Data System (ADS)

    Curtis, J. E.; McLane, L.; Bedoya, M.; Beatty, R.; Kramer, A.; Boehm, H.; Scrimgeour, J.

    2010-03-01

    Evidence is mounting that mechanical and topographical features of biomaterials can be as critical for cellular behavior as chemical properties. A case in point is hyaluronan (HA), a large polysaccharide with unique mechanical and hydrodynamic properties, found in many tissues and bodily fluids. Thanks to a large variety of accessible conformations and aggregation states, this remarkable polymer can impart on its biological environment a diverse range of structural and viscoelastic properties with far-reaching consequences for cell physiology (migration, inflammation, cancer). Supramolecular assembly of HA is typically mediated by HA-binding proteins. These specialized molecules are known to assist the formation of organized structures, such as cross-linked bundles, gels, or the all-important pericellular coat, a polymer network anchored to many cell surfaces. Precisely how the material properties of HA-rich matrices and aggregates are modified by the associated proteins, however, is largely a matter of speculation. We will present new insights concerning the cell coat and HA-protein solutions characterized using passive microrheology, fluorescence recovery after photobleaching (FRAP), and optical force probe microscopy.

  2. Rheology Modification of Hydrogen Peroxide-based Applications Using Cross-linked Polyacrylic Acid Polymers.

    PubMed

    Schmucker-Castner, J; Desai, D

    1999-10-01

    A fundamental study of the stability of hydrogen peroxide-based gels, thickened using cross-linked polyacrylic acid polymers, will be presented. Recent systematic experiments will demonstrate that excellent long term stability of such gels can be achieved. The study evaluates different types of cross-linked polyacrylic acid polymers (as well as an associative acrylic polymer), several commercial sources of hydrogen peroxide, hydrogen peroxide concentrations and the effect of pH. Resulting viscosities from combinations of these variables will be presented, as well as the effects of accelerated aging on percent active peroxide and viscosity of the thickened peroxide gels. The results of this study will enable the formulator to determine the appropriate polymer and conditions necessary for this optimal stability and rheological properties for hydrogen peroxide applications. This study will demonstrate that some cross-linked polyacrylic acid polymers are very compatible and stable with H _ 2O _ 2 and that these polymers do not affect the H _ 2O _ 2 stability under certain conditions. In addition, this study will show that certain cross-linked polyacrylic acid polymers can produce very viscous gels and yield higher viscosities in typical permanent hair color formulas compared to many commercially available products. Because of the excellent compatibility and stability demonstrated, cross-linked acrylic acid polymers are ideal rheological modifiers for hydrogen peroxide-based applications. The performance benefits of using a cross-linked acrylic acid polymer in two-part hair color formulations will be described in detail. PMID:18503448

  3. Chemistry and physical properties of melt-processed and solution-cross-linked corn zein.

    PubMed

    Sessa, David J; Mohamed, Abdellatif; Byars, Jeffrey A

    2008-08-27

    Corn zein was cross-linked with glutaraldehyde (GDA) using glacial acetic acid (HAc) as catalyst. The objectives are to evaluate the swelling characteristics of GDA cross-linked zein gels in water, ethanol, and their combinations. Similar formulations, upon solvent evaporation, form films. The mechanical properties of the films are compared to compression molded tensile bars from GDA melt-processed zein as a second objective. Chemistry of the cross-linking reaction was based on the aldehyde binding characteristics defined by use of fluorescence spectroscopy; sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) to demonstrate the cross-linking reaction; FTIR to observe absorption differences of the cross-linked product; differential scanning calorimetry, dynamic mechanical analysis and thermogravimetric analysis to assess thermal properties; and the use of Instron Universal Testing Machine to evaluate mechanical properties. A reaction mechanism for acid catalyzed GDA cross-linking of zein is proposed. Thermal and mechanical properties of tensile bars cut from either film or formed by compression molding were similar, where both showed increased tensile strengths, ductility and stiffness when compared with unmodified controls. Samples that were reacted with 8% GDA by weight based on weight of zein from either process retained their integrity when tensile bars from each were subjected to boiling water for 10 min or soaking in either water or HAc for 24 h. The melt-processed, cross-linked zein is a more environmentally friendly method that would eliminate the need for HAc recovery. PMID:18636736

  4. Composition of cross-linked 125I-follitropin-receptor complexes

    SciTech Connect

    Shin, J.; Ji, T.H.

    1985-10-15

    Both of the alpha and beta subunits of intact human follitropin (FSH) were radioiodinated with SVI-sodium iodide and chloramine-T and could be resolved on sodium dodecyl sulfate-polyacrylamide gels. Radioiodinated FSH was affinity-cross-linked with a cleavable (nondisulfide) homobifunctional reagent to its membrane receptor on the porcine granulosa cell surface as well as to a Triton X-100-solubilized form of the receptor. Cross-linked samples revealed three additional bands of slower electrophoretic mobility, corresponding to 65, 83, and 117 kDa, in addition to the hormone bands. The hormone alpha beta dimer band corresponded to 43 kDa. Formation of the three bands requires the SVI-hormone to bind specifically to the receptor with subsequent cross-linking. Binding was prevented by an excess of the native hormone but not by other hormones. A monofunctional analog of the cross-linking reagent failed to produce the three bands. Reagent concentration-dependent cross-linking revealed that their formation was sequential; smaller complexes formed first and then larger ones. When gels of cross-linked complexes were treated to cleave covalent cross-links and then electrophoresed in a second dimension, 18-, 22-, and 34-kDa components were released, in addition to the alpha and beta subunits of the hormone.

  5. Enzymatically cross-linked hyperbranched polyglycerol hydrogels as scaffolds for living cells.

    PubMed

    Wu, Changzhu; Strehmel, Christine; Achazi, Katharina; Chiappisi, Leonardo; Dernedde, Jens; Lensen, Marga C; Gradzielski, Michael; Ansorge-Schumacher, Marion B; Haag, Rainer

    2014-11-10

    Although several strategies are now available to enzymatically cross-link linear polymers to hydrogels for biomedical use, little progress has been reported on the use of dendritic polymers for the same purpose. Herein, we demonstrate that horseradish peroxidase (HRP) successfully catalyzes the oxidative cross-linking of a hyperbranched polyglycerol (hPG) functionalized with phenol groups to hydrogels. The tunable cross-linking results in adjustable hydrogel properties. Because the obtained materials are cytocompatible, they have great potential for encapsulating living cells for regenerative therapy. The gel formation can be triggered by glucose and controlled well under various environmental conditions. PMID:25144348

  6. The effect of substrate stiffness, thickness, and cross-linking density on osteogenic cell behavior.

    PubMed

    Mullen, Conleth A; Vaughan, Ted J; Billiar, Kristen L; McNamara, Laoise M

    2015-04-01

    Osteogenic cells respond to mechanical changes in their environment by altering their spread area, morphology, and gene expression profile. In particular, the bulk modulus of the substrate, as well as its microstructure and thickness, can substantially alter the local stiffness experienced by the cell. Although bone tissue regeneration strategies involve culture of bone cells on various biomaterial scaffolds, which are often cross-linked to enhance their physical integrity, it is difficult to ascertain and compare the local stiffness experienced by cells cultured on different biomaterials. In this study, we seek to characterize the local stiffness at the cellular level for MC3T3-E1 cells plated on biomaterial substrates of varying modulus, thickness, and cross-linking concentration. Cells were cultured on flat and wedge-shaped gels made from polyacrylamide or cross-linked collagen. The cross-linking density of the collagen gels was varied to investigate the effect of fiber cross-linking in conjunction with substrate thickness. Cell spread area was used as a measure of osteogenic differentiation. Finite element simulations were used to examine the effects of fiber cross-linking and substrate thickness on the resistance of the gel to cellular forces, corresponding to the equivalent shear stiffness for the gel structure in the region directly surrounding the cell. The results of this study show that MC3T3 cells cultured on a soft fibrous substrate attain the same spread cell area as those cultured on a much higher modulus, but nonfibrous substrate. Finite element simulations predict that a dramatic increase in the equivalent shear stiffness of fibrous collagen gels occurs as cross-linking density is increased, with equivalent stiffness also increasing as gel thickness is decreased. These results provide an insight into the response of osteogenic cells to individual substrate parameters and have the potential to inform future bone tissue regeneration strategies that can optimize the equivalent stiffness experienced by a cell. PMID:25863052

  7. Cross-linking reconsidered: binding and cross-linking fields and the cellular response.

    PubMed Central

    Sulzer, B; De Boer, R J; Perelson, A S

    1996-01-01

    We analyze a model for the reversible cross-linking of cell surface receptors by a collection of bivalent ligands with different affinities for the receptor as would be found in a polyclonal anti-receptor serum. We assume that the amount of cross-linking determines, via a monotonic function, the rate at which cells become activated and divide. In addition to the density of receptors on the cell surface, two quantities, the binding field and the cross-linking field, are needed to characterize the cross-linking curve, i.e., the equilibrium concentration of cross-linked receptors plotted as a function of the total ligand site concentration. The binding field is the sum of all ligand site concentrations weighted by their respective binding affinities, and the cross-linking field is the sum of all ligand site concentrations weighted by the product of their respective binding and cross-linking affinity and the total receptor density. Assuming that the cross-linking affinity decreases if the binding affinity decreases, we find that the height of the cross-linking curve decreases, its width narrows, and its center shifts to higher ligand site concentrations as the affinities decrease. Moreover, when we consider cross-linking-induced proliferation, we find that there is a minimum cross-linking affinity that must be surpassed before a clone can expand. We also show that under many circumstances a polyclonal antiserum would be more likely than a monoclonal antibody to lead to cross-linking-induced proliferation. Images FIGURE 1 FIGURE 2 FIGURE 5 PMID:8785275

  8. 2-(2-methoxyethoxy)ethyl methacrylate hydrogels with gradient of cross-link density

    NASA Astrophysics Data System (ADS)

    Kadlubowski, Slawomir; Matusiak, Malgorzata; Adamus, Agnieszka; Olejniczak, Magdalena N.; Kozanecki, Marcin

    2016-01-01

    Electron beam irradiation of 2-(2-methoxyethoxy)ethyl methacrylate and ethylene glycol dimethacrylate mixtures leads to the formation of cross-linked structures that exhibit a gradient of cross-link density, as demonstrated by gel fraction, swelling and Differential Scanning Calorimetry analysis. The reason for observed phase separation is formation of the high molecular weight clusters and its precipitation before gelation dose. This effect can be controlled/influenced by absorbed dose and cross-linker concentration.

  9. 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.

  10. Cross-linking chemistry of squid beak.

    PubMed

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

    2010-12-01

    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 ((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. PMID:20870720

  11. 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

  12. Tracer diffusion through F-actin: effect of filament length and cross-linking.

    PubMed Central

    Jones, J D; Luby-Phelps, K

    1996-01-01

    We have determined diffusion coefficients for small (50- to 70-nm diameter) fluorescein-thiocarbamoyl-labeled Ficoll tracers through F-actin as a function of filament length and cross-linking. fx45 was used to regulate filament length and avidin/biotinylated actin or ABP-280 was used to prepare cross-linked actin gels. We found that tracer diffusion was generally independent of filament length in agreement with theoretical predictions for diffusion through solutions of rods. However, in some experiments diffusion was slower through short (< or = 1.0 micron) filaments, although this result was not consistently reproducible. Measured diffusion coefficients through unregulated F-actin and filaments of lengths > 1.0 micron were more rapid than predicted by theory for tracer diffusion through rigid, random networks, which was consistent with some degree of actin bundling. Avidin-induced cross-linking of biotinylated F-actin did not affect diffusion through unregulated F-actin, but in cases where diffusion was slower through short filaments this cross-linking method resulted in enhanced tracer diffusion rates indistinguishable from unregulated F-actin. This finding, in conjunction with increased turbidity of 1.0-micron filaments upon avidin cross-linking, indicated that this cross-linking method induces F-actin bundling. By contrast, ABP-280 cross-linking retarded diffusion through unregulated F-actin and decreased turbidity. Tracer diffusion under these conditions was well approximated by the diffusion theory. Both cross-linking procedures resulted in gel formation as determined by falling ball viscometry. These results demonstrate that network microscopic geometry is dependent on the cross-linking method, although both methods markedly increase F-actin macroscopic viscosity. PMID:8913611

  13. The effects of shear and proppant on the viscosity of cross-linked fracturing fluids

    SciTech Connect

    Gardner, D.

    1982-09-01

    Several frequently used cross-linked fracturing fluids, hereafter referred to as gels, have been evaluated as to the effects that shear and that 20/40 mesh sand proppant have on their rheological properties. A closed-loop pipe viscometer was used to evaluate the shear resistance and post-shear recovery of various gels as a function of pH, cross-linker type, polymer concentration, shear intensity (shear stress and shear rate), and shear duration. The aqueous-base gels that were tested included those made with guar, hydroxypropyl guar (HPG), and carboxymethyl hydroxyethyl cellulose (CMHEC) polymers. Selected aqueous-base gels were also evaluated as to the effect on slurry viscosity of varying concentrations of 20/40 mesh sand proppant. Test results indicate that when cross-linked with borate ions, guar and HPG gels have the greatest resistance to shear. Yet when cross-linked with various titanates, HPG gels showed significantly lower resistance to induced shear. Tests on CMHEC gels which were cross-linked with trivalent aluminum (A1+++) indicate a shear resistance between that of the HPG-borate gels and that of the HPG-titanate gels. Also tested was post-shear recovery or the percentage of viscosity increase following high shear exposure as correlated to field tubular pumping conditions. The HPG-borate gels and the CMHEC-A1+++ gels demonstrated considerably higher post-shear recoveries than those attained with HPG-titanate gels. In addition, the effect of 20/40 mesh sand on the viscosity of gels is a larger increase in slurry viscosity in the fracture than that predicted from Newtonian slurry mathematical models.

  14. Cross-linked structure of network evolution

    SciTech Connect

    Bassett, Danielle S.; Wymbs, Nicholas F.; Grafton, Scott T.; Porter, Mason A.; CABDyN Complexity Centre, University of Oxford, Oxford, OX1 1HP ; Mucha, Peter J.; Department of Applied Physical Sciences, University of North Carolina, Chapel Hill, North Carolina 27599

    2014-03-15

    We study the temporal co-variation of network co-evolution via the cross-link structure of networks, for which we take advantage of the formalism of hypergraphs to map cross-link structures back to network nodes. We investigate two sets of temporal network data in detail. In a network of coupled nonlinear oscillators, hyperedges that consist of network edges with temporally co-varying weights uncover the driving co-evolution patterns of edge weight dynamics both within and between oscillator communities. In the human brain, networks that represent temporal changes in brain activity during learning exhibit early co-evolution that then settles down with practice. Subsequent decreases in hyperedge size are consistent with emergence of an autonomous subgraph whose dynamics no longer depends on other parts of the network. Our results on real and synthetic networks give a poignant demonstration of the ability of cross-link structure to uncover unexpected co-evolution attributes in both real and synthetic dynamical systems. This, in turn, illustrates the utility of analyzing cross-links for investigating the structure of temporal networks.

  15. Electrospinning covalently cross-linking biocompatible hydrogelators

    PubMed Central

    Schultz, Kelly M.; Campo-Deao, Laura; Baldwin, Aaron D.; Kiick, Kristi L.; Clasen, Christian; Furst, Eric M.

    2012-01-01

    Many hydrogel materials of interest are homogeneous on the micrometer scale. Electrospinning, the formation of sub-micrometer to micrometer diameter fibers by a jet of fluid formed under an electric field, is one process being explored to create rich microstructures. However, electrospinning a hydrogel system as it reacts requires an understanding of the gelation kinetics and corresponding rheology near the liquid-solid transition. In this study, we correlate the structure of electrospun fibers of a covalently cross-linked hydrogelator with the corresponding gelation transition and kinetics. Polyethylene oxide (PEO) is used as a carrier polymer in a chemically cross-linking poly(ethylene glycol)-high molecular weight heparin (PEG-HMWH) hydrogel. Using measurements of gelation kinetics from multiple particle tracking microrheology (MPT), we correlate the material rheology with the the formation of stable fibers. An equilibrated, cross-linked hydrogel is then spun and the PEO is dissolved. In both cases, microstructural features of the electrospun fibers are retained, confirming the covalent nature of the network. The ability to spin fibers of a cross-linking hydrogel system ultimately enables the engineering of materials and microstructural length scales suitable for biological applications. PMID:23459473

  16. Electrospinning covalently cross-linking biocompatible hydrogelators.

    PubMed

    Schultz, Kelly M; Campo-Deao, Laura; Baldwin, Aaron D; Kiick, Kristi L; Clasen, Christian; Furst, Eric M

    2013-01-01

    Many hydrogel materials of interest are homogeneous on the micrometer scale. Electrospinning, the formation of sub-micrometer to micrometer diameter fibers by a jet of fluid formed under an electric field, is one process being explored to create rich microstructures. However, electrospinning a hydrogel system as it reacts requires an understanding of the gelation kinetics and corresponding rheology near the liquid-solid transition. In this study, we correlate the structure of electrospun fibers of a covalently cross-linked hydrogelator with the corresponding gelation transition and kinetics. Polyethylene oxide (PEO) is used as a carrier polymer in a chemically cross-linking poly(ethylene glycol)-high molecular weight heparin (PEG-HMWH) hydrogel. Using measurements of gelation kinetics from multiple particle tracking microrheology (MPT), we correlate the material rheology with the the formation of stable fibers. An equilibrated, cross-linked hydrogel is then spun and the PEO is dissolved. In both cases, microstructural features of the electrospun fibers are retained, confirming the covalent nature of the network. The ability to spin fibers of a cross-linking hydrogel system ultimately enables the engineering of materials and microstructural length scales suitable for biological applications. PMID:23459473

  17. Cross-linked structure of network evolution

    NASA Astrophysics Data System (ADS)

    Bassett, Danielle S.; Wymbs, Nicholas F.; Porter, Mason A.; Mucha, Peter J.; Grafton, Scott T.

    2014-03-01

    We study the temporal co-variation of network co-evolution via the cross-link structure of networks, for which we take advantage of the formalism of hypergraphs to map cross-link structures back to network nodes. We investigate two sets of temporal network data in detail. In a network of coupled nonlinear oscillators, hyperedges that consist of network edges with temporally co-varying weights uncover the driving co-evolution patterns of edge weight dynamics both within and between oscillator communities. In the human brain, networks that represent temporal changes in brain activity during learning exhibit early co-evolution that then settles down with practice. Subsequent decreases in hyperedge size are consistent with emergence of an autonomous subgraph whose dynamics no longer depends on other parts of the network. Our results on real and synthetic networks give a poignant demonstration of the ability of cross-link structure to uncover unexpected co-evolution attributes in both real and synthetic dynamical systems. This, in turn, illustrates the utility of analyzing cross-links for investigating the structure of temporal networks.

  18. 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.

  19. Arabinosylation Plays a Crucial Role in Extensin Cross-linking In Vitro

    PubMed Central

    Chen, Yuning; Dong, Wen; Tan, Li; Held, Michael A; Kieliszewski, Marcia J

    2015-01-01

    Extensins (EXTs) are hydroxyproline-rich glycoproteins (HRGPs) that are structural components of the plant primary cell wall. They are basic proteins and are highly glycosylated with carbohydrate accounting for >50% of their dry weight. Carbohydrate occurs as monogalactosyl serine and arabinosyl hydroxyproline, with arabinosides ranging in size from ~1 to 4 or 5 residues. Proposed functions of EXT arabinosylation include stabilizing the polyproline II helix structure and facilitating EXT cross-linking. Here, the involvement of arabinosylation in EXT cross-linking was investigated by assaying the initial cross-linking rate and degree of cross-linking of partially or fully de-arabinosylated EXTs using an in vitro cross-linking assay followed by gel permeation chromatography. Our results indicate that EXT arabinosylation is required for EXT cross-linking in vitro and the fourth arabinosyl residue in the tetraarabinoside chain, which is uniquely ?-linked, may determine the initial cross-linking rate. Our results also confirm the conserved structure of the oligoarabinosides across species, indicating an evolutionary significance for EXT arabinosylation. PMID:26568683

  20. Encapsulation of volatiles by homogenized partially-cross linked alginates.

    PubMed

    Inguva, Pavan K; Ooi, Shing Ming; Desai, Parind M; Heng, Paul W S

    2015-12-30

    Cross-linked calcium alginate gels are too viscous to be efficaciously incorporated into spray dried formulations. Thus, viscosity reduction is essential to ensure the processability of calcium alginate gels to be sprayed. Viscosity reduction by high pressure homogenization can open new formulation possibilities. Presently, testing of microcapsule integrity is also limited because either single particle tests neglect collective particle behaviours in bulk or bulk testing methods are often associated with single compressions which may not fully characterize individual particle strengths. The aim of this study was sub-divided into three objectives. First objective was to evaluate the impact of high pressure homogenization on gel viscosity. Second objective was to explore the use of the homogenized gels with modified starch for microencapsulation by spray drying. The final objective was to develop a stamping system as microcapsule strength tester that can assess microcapsules in bulk and evaluate the impact of multiple compressions. Collectively, this study would lead towards developing a pressure-activated patch of microcapsules with encapsulated volatiles and the method to assess the patch efficacy. The alginate gels largely experienced an exponential decay in viscosity when homogenized. Furthermore, the homogenized gels were successfully incorporated in spray drying formulations for microencapsulation. The custom-designed microcapsule strength tester was successfully used and shown to possess the required sensitivity to discern batches of microcapsules containing volatiles to have different release profiles. Addition of homogenized gels strengthened the microcapsules only at high wall to core ratios with low mass-load alginate gels. High mass-load gels weaken the microcapsules, exhibiting a higher release at low stamping pressures and wrinkling on the microcapsules surface. PMID:26581772

  1. 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.

  2. 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 reactive group that can be photoactivated at a specific point in the nucleation or crystal growth process to "capture" protein molecules bound within reach of the crosslinking agent. If those bound protein molecules have a defined geometric relationship with the capturing molecule, such as would be found in a crystal, then the photoreacted cross-linking site should be consistent. Random protein interactions, typical of an amorphous precipitate or interaction, would show a random cross-linking reaction. The results of these and other experiments will be presented.

  3. Ultrathin cross-linked nanoparticle membranes.

    PubMed

    Lin, Yao; Skaff, Habib; Bker, Alexander; Dinsmore, A D; Emrick, Todd; Russell, Thomas P

    2003-10-22

    The fabrication of functional nanostructured materials for sensing, encapsulation and delivery requires practical approaches to self-assembly on multiple length scales and the synthesis of tough yet permeable structures. Here, the self-assembly of functionalized, photoluminescent nanoparticles at liquid interfaces, followed by cross-linking of the associated ligands, affords robust membranes that maintain their integrity even when they are removed from the interface. These composite membranes, nanometers in thickness, are elastic yet permeable and have potential applications involving controlled permeability and diffusion. Cadmium selenide (CdSe) nanoparticles are used, since their inherent photoluminescence offers a direct way to probe the spatial organization of the particles. Functionalized ligands attached to the nanoparticles provide an effective means to stabilize the interfacial assembly by cross-linking. The concepts shown are adaptable to other type of nanoparticles, ligands, and solvent combinations. PMID:14558800

  4. Positive tone cross-linked resists based on photoacid inhibition of cross linking

    NASA Astrophysics Data System (ADS)

    Lawson, Richard A.; Chun, Jun Sung; Neisser, Mark; Tolbert, Laren M.; Henderson, Clifford L.

    2014-03-01

    A resist imaging design that utilizes photoacid inhibition of cationic polymerization and cross-linking during a postexposure bake step has been studied. The key to the design approach is the use of two different polymerization catalysts/initiators: (1) a photoacid produced from a photoacid generator (PAG) upon exposure of the resist that can result in polymerization and cross-linking of the resist matrix and (2) a thermal cross-linking catalyst (TCC) designed to thermally catalyze epoxide-phenol cross-linking. The TCC can be chosen from a variety of compounds such as triphenylphosphine (TPP) or imidazole. When only one of these catalysts (e.g TPP or photoacid) is present in an epoxide and phenol containing resist matrix, it will individually catalyze cross-linking. When they are present together, they effectively quench one another and little to no cross-linking occurs. This approach can be used to switch the tone of a resist from negative (photoacid catalyzed) to positive (TCC catalyzed and photoacid inhibited). The effect of the ratio of TCC:PAG was examined and the optimal ratio for positive tone behavior was determined. Resist contrast can be modified by optimization of epoxide:phenol ratio in the formulation. Dual tone behavior with positive tone at low dose and negative tone at higher doses can be observed in certain formulation conditions. Initial EUV patterning shows poor results, but the source of the poor imaging is not yet understood.

  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. Enzymatically cross-linked alginic-hyaluronic acid composite hydrogels as cell delivery vehicles.

    PubMed

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

    2013-04-01

    An injectable composite gel was developed from alginic and hyaluronic acid. The enzymatically 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 tyraminated polymer solutions in 10U HRP and treating with 1.0% H2O2. The properties of the alginate gels were significantly 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

  7. Reaction of nucleic acids with cis-diamminedichloroplatinum(II): interstrand cross-links

    SciTech Connect

    Rahmouni, A.; Leng, M.

    1987-11-17

    In the reaction of cis-diamminedichloroplatinum(II) (cis-DDP) with double-helical (dC-dG)/sub 4/-(dC-dG)/sub 4/ or (dC-dG)/sub 5/-(dC-dG)/sub 5/, intrastrand and interstrand cross-links between two guanine residues are formed. This is shown by gel electrophoresis in denaturing conditions of the reaction products and by high-performance liquid chromatography (HPLC) analysis of the products digested with nuclease P1. In the reaction of cis-DDP and poly(dG-dC)-poly(dG-dC), at relatively low levels of platination, it is mainly interstrand cross-links between two guanine residues that are formed. This is shown by HPLC analysis of the nuclease P1 digest and by gel electrophoresis in denaturing and nondenaturing conditions of the platinated polymer after cleavage with the restriction enzyme HhaI. Moreover, the antibodies to platinated poly(dG-dC)-poly(dG-dC) cross-react with the interstrand cross-linked (dC-dG)/sub 4/ or (dC-dG)/sub 5/ but not with the intrastrand cross-linked (dG-dC)/sub 4/ or (dC-dG)/sub 5/. These antibodies cross-react with platinated natural DNA. The amount of interstrand cross-links deduced from radioimmunoassays (0.5% of the total bound platinum) is lower than that (2%) deduced by gel electrophoresis in denaturing conditions of a platinated DNA restriction fragment. By gel electrophoresis, it is also shown that in vitro the isomer trans-DDP is more efficient in forming interstrand cross-links than cis-DDP.

  8. 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

  9. Covalently Cross-Linked Arabinoxylans Films for Debaryomyces hansenii Entrapment.

    PubMed

    Gonzlez-Estrada, Ramss; Caldern-Santoyo, Montserrat; Carvajal-Millan, Elizabeth; Ascencio Valle, Felipe de Jess; Ragazzo-Snchez, Juan Arturo; Brown-Bojorquez, Francisco; Rascn-Chu, Agustn

    2015-01-01

    In the present study, wheat water extractable arabinoxylans (WEAX) were isolated and characterized, and their capability to form covalently cross-linked films in presence of Debaryomyces hansenii was evaluated. WEAX presented an arabinose to xylose ratio of 0.60, a ferulic acid and diferulic acid content of 2.1 and 0.04 g?mg(-1) WEAX, respectively and a Fourier Transform Infra-Red (FT-IR) spectrum typical of WEAX. The intrinsic viscosity and viscosimetric molecular weight values for WEAX were 3.6 dL?g(-1) and 440 kDa, respectively. The gelation of WEAX (1% w/v) with and without D. hansenii (1 10(7) CFU?cm(-2)) was rheologically investigated by small amplitude oscillatory shear. The entrapment of D. hansenii decreased gel elasticity from 1.4 to 0.3 Pa, probably by affecting the physical interactions between WEAX chains. Covalently cross-linked WEAX films containing D. hansenii were prepared by casting. Scanning electron microscopy images show that WEAX films containing D. hansenii were porous and consisted of granular-like and fibre microstructures. Average tensile strength, elongation at break and Young's modulus values dropped when D. hansenii was present in the film. Covalently cross-lined WEAX containing D. hansenii could be a suitable as a functional entrapping film. PMID:26102070

  10. 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. PMID:21294517

  11. Ferredoxin Cross-Links to a 22 kD Subunit of Photosystem I 1

    PubMed Central

    Zilber, April L.; Malkin, Richard

    1988-01-01

    We have used a cross-linking approach to study the interaction of ferredoxin (Fd) with photosystem I (PSI). The cross-linking reagent N-ethyl-3-(3-dimethylaminopropyl) carbodiimide was found to cross-link spinach Fd to a 22 kilodalton subunit of PSI in both isolated spinach (Spinacia oleracea) PSI complexes and spinach thylakoid membranes. The product had an apparent molecular weight of 38 kilodaltons on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and was identified as a cross-linked product using specific antibodies to Fd and the 22 kilodalton subunit. In both a native PSI complex (200 Chl/P700) and a PSI core complex (100 Chl/P700), a second cross-linked product at 36 kilodaltons was seen. The latter cross-reacted with an antibody to Fd but did not cross-react with antibodies directed against the 24.3, 22, 19, 17.3 or 8.5 kilodalton, or psaC subunits of PSI. Its composition remains to be determined. In thylakoids only the 38 kilodalton product was observed along with a cross-linked complex of Fd and Fd:NADP+ reductase. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:16666389

  12. 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 (?Hm). 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. PMID:23713446

  13. 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 ?-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 ?-sheet formation. Genipin cross-linking before ?-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. PMID:25978549

  14. Chicken Corneocyte Cross-Linked Proteome

    PubMed Central

    Rice, Robert H.; Winters, Brett R.; Durbin-Johnson, Blythe P.; Rocke, David M.

    2013-01-01

    Shotgun proteomic analysis was performed of epidermal scale, feather, beak and claw from the domestic chicken. To this end, the samples were separated first into solubilized and particulate fractions, the latter enriched in isopeptide cross-linking, by exhaustive extraction in sodium dodecyl sulfate under reducing conditions. Among the 205 proteins identified were 17 keratins (types α and β), 51 involved in protein synthesis, 8 junctional, 8 histone, 5 heat shock and 5 14-3-3 proteins. Considerable overlap among the beak, claw, feather and scale samples was observed in protein profiles, but those from beak and claw were the most similar. Scale and feather profiles were the most distinctive, each exhibiting specific proteins. Less than 20% of the proteins were found only in the detergent solubilized fraction, while 34-57% were found only in the particulate fraction, depending on the source, and the rest in both fractions. The results provide the first comprehensive analysis of the content of these cornified structures, reveal the efficient use of available proteins in conferring mechanical and chemical stability to them and emphasize the importance of isopeptide cross-linking in avian epithelial cornification. PMID:23256538

  15. Corneal collagen cross-linking: A review

    PubMed Central

    O’Brart, David P.S.

    2014-01-01

    The aim was to review the published literature on corneal collagen cross-linking. The emphasis was on the seminal publications, systemic reviews, meta-analyses and randomized controlled trials. Where such an evidence did not exist, selective large series cohort studies, case controlled studies and case series with follow-up preferably greater than 12 months were included. Riboflavin/Ultraviolet A (UVA) corneal collagen cross-linking appears to be the first treatment modality to halt the progression of keratoconus and other corneal ectatic disorders with improvement in visual, keratometric and topographic parameters documented by most investigators. Its precise mechanism of action at a molecular level is as yet not fully determined. Follow-up is limited to 4–6 years at present but suggests continued stability and improvement in corneal shape with time. Most published data are with epithelium-off techniques. Epithelium-on studies suggest some efficacy but less than with the epithelium-off procedures and long-term data are not currently available. The use of Riboflavin/UVA CXL for the management of infectious and non-infectious keratitis appears very promising. Its use in the management of bullous keratopathy is equivocal. Investigation of other methodologies for CXL are under investigation. PMID:25000866

  16. Glycosylation and cross-linking in bone type I collagen.

    PubMed

    Terajima, Masahiko; Perdivara, Irina; Sricholpech, Marnisa; Deguchi, Yoshizumi; Pleshko, Nancy; Tomer, Kenneth B; Yamauchi, Mitsuo

    2014-08-15

    Fibrillar type I collagen is the major organic component in bone, providing a stable template for mineralization. During collagen biosynthesis, specific hydroxylysine residues become glycosylated in the form of galactosyl- and glucosylgalactosyl-hydroxylysine. Furthermore, key glycosylated hydroxylysine residues, ?1/2-87, are involved in covalent intermolecular cross-linking. Although cross-linking is crucial for the stability and mineralization of collagen, the biological function of glycosylation in cross-linking is not well understood. In this study, we quantitatively characterized glycosylation of non-cross-linked and cross-linked peptides by biochemical and nanoscale liquid chromatography-high resolution tandem mass spectrometric analyses. The results showed that glycosylation of non-cross-linked hydroxylysine is different from that involved in cross-linking. Among the cross-linked species involving ?1/2-87, divalent cross-links were glycosylated with both mono- and disaccharides, whereas the mature, trivalent cross-links were primarily monoglycosylated. Markedly diminished diglycosylation in trivalent cross-links at this locus was also confirmed in type II collagen. The data, together with our recent report (Sricholpech, M., Perdivara, I., Yokoyama, M., Nagaoka, H., Terajima, M., Tomer, K. B., and Yamauchi, M. (2012) Lysyl hydroxylase 3-mediated glucosylation in type I collagen: molecular loci and biological significance. J. Biol. Chem. 287, 22998-23009), indicate that the extent and pattern of glycosylation may regulate cross-link maturation in fibrillar collagen. PMID:24958722

  17. Glycosylation and Cross-linking in Bone Type I Collagen*

    PubMed Central

    Terajima, Masahiko; Perdivara, Irina; Sricholpech, Marnisa; Deguchi, Yoshizumi; Pleshko, Nancy; Tomer, Kenneth B.; Yamauchi, Mitsuo

    2014-01-01

    Fibrillar type I collagen is the major organic component in bone, providing a stable template for mineralization. During collagen biosynthesis, specific hydroxylysine residues become glycosylated in the form of galactosyl- and glucosylgalactosyl-hydroxylysine. Furthermore, key glycosylated hydroxylysine residues, ?1/2-87, are involved in covalent intermolecular cross-linking. Although cross-linking is crucial for the stability and mineralization of collagen, the biological function of glycosylation in cross-linking is not well understood. In this study, we quantitatively characterized glycosylation of non-cross-linked and cross-linked peptides by biochemical and nanoscale liquid chromatography-high resolution tandem mass spectrometric analyses. The results showed that glycosylation of non-cross-linked hydroxylysine is different from that involved in cross-linking. Among the cross-linked species involving ?1/2-87, divalent cross-links were glycosylated with both mono- and disaccharides, whereas the mature, trivalent cross-links were primarily monoglycosylated. Markedly diminished diglycosylation in trivalent cross-links at this locus was also confirmed in type II collagen. The data, together with our recent report (Sricholpech, M., Perdivara, I., Yokoyama, M., Nagaoka, H., Terajima, M., Tomer, K. B., and Yamauchi, M. (2012) Lysyl hydroxylase 3-mediated glucosylation in type I collagen: molecular loci and biological significance. J. Biol. Chem. 287, 2299823009), indicate that the extent and pattern of glycosylation may regulate cross-link maturation in fibrillar collagen. PMID:24958722

  18. Gauging and Tuning Cross-Linking Kinetics of Catechol-PEG Adhesives via Catecholamine Functionalization.

    PubMed

    Paez, Julieta I; Ustahseyin, Oya; Serrano, Cristina; Ton, Xuan-Anh; Shafiq, Zahid; Auernhammer, Gnter K; d'Ischia, Marco; Del Campo, Arnzazu

    2015-12-14

    The curing time of an adhesive material is determined by the polymerization and cross-linking kinetics of the adhesive formulation and needs to be optimized for the particular application. Here, we explore the possibility of tuning the polymerization kinetics and final mechanical properties of tissue-adhesive PEG gels formed by polymerization of end-functionalized star-PEGs with catecholamines with varying substituents. We show strong differences in cross-linking time and cohesiveness of the final gels among the catecholamine-PEG variants. Installation of an electron-withdrawing but ?-electron donating chloro substituent on the catechol ring resulted in faster and more efficient cross-linking, while opposite effects were observed with the strongly electron-withdrawing nitro group. Chain substitution slowed down the kinetics and hindered cross-linking due either to chain breakdown (?-OH group, in norepinephrine) or intramolecular cyclization (?-carboxyl group, in DOPA). Interesting perspectives derive from use of mixtures of catecholamine-PEG precursors offering further opportunities for fine-tuning of the curing parameters. These are interesting properties for the application of catecholamine-PEG gels as tissue glues or biomaterials for cell encapsulation. PMID:26583428

  19. 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

    2016-04-01

    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. PMID:26733331

  20. Genipin cross-linked alginate-chitosan microcapsules: membrane characterization and optimization of cross-linking reaction.

    PubMed

    Chen, Hongmei; Ouyang, Wei; Lawuyi, Bisi; Prakash, Satya

    2006-07-01

    The genipin cross-linked alginate-chitosan (GCAC) microcapsule, composed of an alginate core and a genipin cross-linked chitosan membrane, was recently proposed for live cell encapsulation and other delivery applications. This article for the first time describes the details of the microcapsule membrane characterization using a noninvasive and in situ method without any physical or chemical modifications on the samples. Results showed that the cross-linking reaction generated the fluorescent chitosan-genipin conjugates. The cross-linked chitosan membrane was clearly visualized by confocal laser scanning microscopy (CLSM). A straightforward assessment on the membrane thickness and relative intensity was successfully achieved. CLSM studies showed that the shell-like cross-linked chitosan membranes of approximately 37 microm in thickness were formed surrounding the microcapsule. The reaction variables, including cross-linking temperature and time significantly affected the fluorescence intensity of the membranes. Elevating the cross-linking temperature from 4 to 37 degrees C drastically intensified the membrane fluorescence, suggesting the attainment of a high degree of cross-linking on the chitosan membrane. Extended cross-linking time altered the cross-linked membranes in modulation. Although genipin concentration and cross-linking time had little effects on the membrane thickness, cross-linking at higher temperatures tended to form relatively thinner membranes. PMID:16827575

  1. Synthesis of acrylic and allylic bifunctional cross-linking monomers derived from PET waste

    NASA Astrophysics Data System (ADS)

    Cruz-Aguilar, A.; Herrera-Gonzlez, A. M.; Vzquez-Garca, R. A.; Navarro-Rodrguez, D.; Coreo, J.

    2013-06-01

    An acrylic and two novel allylic monomers synthesized from bis (hydroxyethyl) terephthalate, BHET, are reported. This was obtained by glycolysis of post-consumer PET with boiling ethylene glycol. The bifunctional monomer bis(2-(acryloyloxy)ethyl) terephthalate was obtained from acryloyl chloride, while the allylic monomers 2-(((allyloxi)carbonyl)oxy) ethyl (2-hydroxyethyl) terephthalate and bis(2-(((allyloxi)carbonyl)oxy)ethyl) terephthalate, from allyl chloroformate. Cross-linking was studied in bulk polymerization using two different thermal initiators. Monomers were analyzed by means of 1H NMR and the cross-linked polymers by infrared spectroscopy. Gel content higher than 90% was obtained for the acrylic monomer. In the case of the mixture of the allylic monomers, the cross-linked polymer was 80 % using BPO initiator, being this mixture 24 times less reactive than the acrylic monomer.

  2. Kojak: efficient analysis of chemically cross-linked protein complexes.

    PubMed

    Hoopmann, Michael R; Zelter, Alex; Johnson, Richard S; Riffle, Michael; MacCoss, Michael J; Davis, Trisha N; Moritz, Robert L

    2015-05-01

    Protein chemical cross-linking and mass spectrometry enable the analysis of protein-protein interactions and protein topologies; however, complicated cross-linked peptide spectra require specialized algorithms to identify interacting sites. The Kojak cross-linking software application is a new, efficient approach to identify cross-linked peptides, enabling large-scale analysis of protein-protein interactions by chemical cross-linking techniques. The algorithm integrates spectral processing and scoring schemes adopted from traditional database search algorithms and can identify cross-linked peptides using many different chemical cross-linkers with or without heavy isotope labels. Kojak was used to analyze both novel and existing data sets and was compared to existing cross-linking algorithms. The algorithm provided increased cross-link identifications over existing algorithms and, equally importantly, the results in a fraction of computational time. The Kojak algorithm is open-source, cross-platform, and freely available. This software provides both existing and new cross-linking researchers alike an effective way to derive additional cross-link identifications from new or existing data sets. For new users, it provides a simple analytical resource resulting in more cross-link identifications than other methods. PMID:25812159

  3. 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.

  4. 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.

  5. Ehrlich chromogens, probable cross-links in elastin and collagen.

    PubMed Central

    Kemp, P D; Scott, J E

    1988-01-01

    (1) Proteolytic digests of tissue elastin contain material which reacts with dimethylaminobenzaldehyde in acid solution (Ehrlich's reagent) to give a cherry-pink colour. This Ehrlich chromogen(s) [EC(s)] is similar to but not identical with EC(s) previously demonstrated in tissue collagens [Scott, Hughes & Shuttleworth (1979) Biosci. Rep. 1, 611-618]. Both ECs react with diazonium salts in acid to give coloured products. (2) Diazobenzene linked via a phenolic ester to polyacrylamide beads (Biogel P10) has been used to absorb ECs specifically and almost quantitatively from proteolytic digests. The coupled deeply coloured azo-EC-peptides were then recovered after mild alkaline cleavage from the support and purified by gel chromatography. (3) Using 15N-labelled NaNO2, the collagen azo-EC-peptides were prepared, and 15N abundance measured therein. The molar absorption coefficient of the azo-EC group was calculated (18,700) based on the assumption that each azo-EC group contained one 15N atom. (4) Collagen azo-EC-peptides contained glucose and galactose, whereas elastin azo-EC peptides did not. The amino acid patterns of the two peptides were quite different, the former being rich in polar amino acids, the latter containing much alanine. The patterns were compatible with an origin from the cross-linking regions of collagen and elastin respectively. (5) Quantitative (molar) comparisons of the azo-EC group content with amino acid, amino end-group and sugar contents, and azo-EC peptide molecular mass, suggest that a structure is present in the collagen azo-EC-peptides containing two EC groups shared between four peptide chains. Three peptide chains probably meet at each (cross-linking) EC group. (6) Based on this structure, about 15% of adult bovine skin collagen contains EC groups. PMID:3415661

  6. 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.

  7. Vitamin C hinders radiation cross-linking in aqueous poly(vinyl alcohol) solutions

    NASA Astrophysics Data System (ADS)

    Oral, Ebru; Bodugoz-Senturk, Hatice; Macias, Celia; Muratoglu, Orhun K.

    2007-12-01

    Poly(vinyl alcohol) (PVA) is a promising semi-crystalline material for biomedical applications. It is soluble in water and can be formed into hydrogels by freezing and thawing or crystallizing from an aqueous theta solution such as that of polyethylene glycol (PEG). Radiation cross-linking caused by sterilization or high dose irradiation of concentrated PVA solutions could compromise some properties of these hydrogels. Therefore, we hypothesized that radiation cross-linking of PVA solutions and PVA-PEG theta gels could be prevented by using the antioxidant vitamin C as an anticross-linking agent. Our hypothesis tested positive. Vitamin C concentrations of 0.75 and 4.5 mol/mol of PVA repeating unit could prevent cross-linking in 17.5 wt/v% PVA solutions made with PVA molecular weight of 115,000 g/mol irradiated to 25 and 100 kGy, respectively. Vitamin C also prevented cross-linking in 25 kGy irradiated PVA-PEG theta gels containing up to 5 wt% PEG and decreased the viscosity of those up to 39 wt%.

  8. Nonlinear elasticity of cross-linked networks

    NASA Astrophysics Data System (ADS)

    John, Karin; Caillerie, Denis; Peyla, Philippe; Raoult, Annie; Misbah, Chaouqi

    2013-04-01

    Cross-linked semiflexible polymer networks are omnipresent in living cells. Typical examples are actin networks in the cytoplasm of eukaryotic cells, which play an essential role in cell motility, and the spectrin network, a key element in maintaining the integrity of erythrocytes in the blood circulatory system. We introduce a simple mechanical network model at the length scale of the typical mesh size and derive a continuous constitutive law relating the stress to deformation. The continuous constitutive law is found to be generically nonlinear even if the microscopic law at the scale of the mesh size is linear. The nonlinear bulk mechanical properties are in good agreement with the experimental data for semiflexible polymer networks, i.e., the network stiffens and exhibits a negative normal stress in response to a volume-conserving shear deformation, whereby the normal stress is of the same order as the shear stress. Furthermore, it shows a strain localization behavior in response to an uniaxial compression. Within the same model we find a hierarchy of constitutive laws depending on the degree of nonlinearities retained in the final equation. The presented theory provides a basis for the continuum description of polymer networks such as actin or spectrin in complex geometries and it can be easily coupled to growth problems, as they occur, for example, in modeling actin-driven motility.

  9. Cross-Linking Aromatic Polymers With Ionizing Radiation

    NASA Technical Reports Server (NTRS)

    Bell, Vernon L.; Havens, Stephen J.

    1987-01-01

    Resistance to heat and solvents increased. Certain aromatic polymers containing radiation-sensitive methylene groups cross-linked through methylene groups upon exposure to ionizing radiation. Cross-linked polymers resistant to most organic solvents and generally more resistant to high temperatures, with less tendency to creep under load. No significant embrittlement of parts fabricated from these polymers when degree of cross-linking, as controlled by irradiation dose, kept at moderate level.

  10. The Role of Nonenzymatic Glycation and Carbonyls in Collagen Cross-Linking for the Treatment of Keratoconus

    PubMed Central

    Littlechild, Stacy; McCall, Scott; Zhang, Yuntao; Conrad, Gary W.

    2011-01-01

    Purpose. Corneal cross-linking (CXL) is a treatment for keratoconus that eliminates the need for keratoplasty in most patients. However, its molecular mechanisms remain under study. Advanced glycation end products (AGEs) have been suggested by many studies as the causative strengthening agent during CXL, though no studies to date have directly tested this hypothesis. Methods. Corneas of young rabbits and sharks were pretreated with pyridoxal hydrochloride and copper ions before CXL. Two known inhibitors of AGE formation, aminoguanidine and rifampicin, were applied during CXL in the treatment solution. Tensile strength tests were conducted after these experiments to detect diminished or accentuated corneal stiffening after CXL. SDS-PAGE was performed on type I collagen cross-linked in the absence and presence of AGE inhibitors. Results. Pretreatment with pyridoxal hydrochloride resulted in significantly higher corneal stiffening after CXL. AGE inhibitors significantly diminished cross-linking as detected by both tensile strength measurements using whole corneas and gel electrophoresis of in vitro cross-linking of type I collagen in solution, in the presence and absence of the inhibitors. Rifampicin inhibited CXL more significantly than aminoguanidine in gel electrophoresis and tensile strength tests, confirming recent findings on its efficacy as an AGE inhibitor. Conclusions. Data presented here suggest that CXL is carbonyl dependent and involves the formation of AGE cross-links. Six possible cross-linking mechanisms are discussed. PMID:21724915

  11. Enzyme stabilization via cross-linked enzyme aggregates.

    PubMed

    Gupta, Munishwar N; Raghava, Smita

    2011-01-01

    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 both conventional aqueous media as well as nonaqueous media. These are also stable at fairly high temperatures. CLEAs having 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. PMID:20865393

  12. Intermolecular Peptide Cross-Linking by Using Diaminodicarboxylic Acids.

    PubMed

    Kamalov, Meder; Kaur, Harveen; Brimble, Margaret A

    2016-03-01

    Synthetic methods aimed at preparing peptides cross-linked by diaminodiacids remain an important chemical challenge. These cross-links are known to play a crucial role on the activity, structural stability, and folding of the host peptides and proteins. Recent developments in the syntheses of such systems have led to intriguing advances in the understanding of intermolecular side-chain cross-linking and the role that these structural motifs play in the biochemistry of proteins. Herein we provide an overview of the existing synthetic methodology that has been developed to effect protein cross-linking using diaminodiacids. PMID:26749083

  13. 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).

  14. Cross-linking of smooth muscle caldesmon to the NH2-terminal region of skeletal F-actin.

    PubMed

    Bartegi, A; Fattoum, A; Kassab, R

    1990-02-01

    The cross-linking of the F-actin-caldesmon complex with 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide in the presence of N-hydroxysuccinimide generated four major adducts which were identified on polyacrylamide gels. By cross-linking 3H-actin to 14C-caldesmon, these were found to represent 1:1 cross-linked complexes of actin and caldesmon displaying different electrophoretic mobilities. Tropomyosin did not noticeably affect the cross-linking process. The same four fluorescent species resulting from the cross-linking of caldesmon to F-actin labeled with N-[7-(dimethylamino)-4-methyl-3-coumarinyl]maleimide were subjected separately to partial cleavages with hydroxylamine or cyanogen bromide. These treatments yielded fluorescent 41- and 37-kDa fragments, respectively, from each cross-linked entity indicating unambiguously that caldesmon was cross-linked only to the NH2-terminal actin stretch of residues 1-12. This region is also known to serve for the carbodiimide-mediated cross-linking of the myosin subfragment-1 heavy chain (Sutoh, K. (1982) Biochemistry 21, 3654-3661). A covalent caldesmon-F-actin conjugate containing a protein molar ratio close to 1:19 was isolated following dissociation of uncross-linked caldesmon. It showed a low level of activation of the ATPase activity of skeletal myosin subfragment-1, and the binding of Ca2(+)-calmodulin to the derivative did not cause the reversal of the ATPase inhibition. In contrast, the reversible binding of caldesmon to F-actin cross-linked to myosin subfragment-1 did not inhibit the accelerated ATPase of the complex. The overall data point to the dual involvement of the actin's NH2 terminus in the inhibitory binding of caldesmon and in actomyosin interactions in the presence of ATP. PMID:2298747

  15. Hyaluronan-Dependent Pericellular Matrix

    PubMed Central

    Evanko, Stephen P.; Tammi, Markku I.; Tammi, Raija H.; Wight, Thomas N.

    2007-01-01

    Hyaluronan is a multifunctional glycosaminoglycan that forms the structural basis of the pericellular matrix. Hyaluronan is extruded directly through the plasma membrane by one of three hyaluronan synthases and anchored to the cell surface by the synthase or cell surface receptors such as CD44 or RHAMM. Aggregating proteoglycans and other hyaluronan-binding proteins, contribute to the material and biological properties of the matrix and regulate cell and tissue function. The pericellular matrix plays multiple complex roles in cell adhesion/de-adhesion, and cell shape changes associated with proliferation and locomotion. Time-lapse studies show that pericellular matrix formation facilitates cell detachment and mitotic cell rounding. Hyaluronan crosslinking occurs through various proteins, such as tenascin, TSG-6, inter-alpha-trypsin inhibitor, pentraxin and TSP-1. This creates higher order levels of structured hyaluronan that may regulate inflammation and other biological processes. Microvillous or filopodial membrane protrusions are created by active hyaluronan synthesis, and form the scaffold of hyaluronan coats in certain cells. The importance of the pericellular matrix in cellular mechanotransduction and the response to mechanical strain are also discussed. PMID:17804111

  16. A photolithographic approach to spatially resolved cross-linked nanolayers.

    PubMed

    Fuchise, Keita; Lindemann, Peter; Heiler, Stefan; Gliemann, Hartmut; Trouillet, Vanessa; Welle, Alexander; Berson, Jonathan; Walheim, Stefan; Schimmel, Thomas; Meier, Michael A R; Barner-Kowollik, Christopher

    2015-03-17

    The preparation of cross-linked nanosheets with 1-2 nm thickness and predefined shape was achieved by lithographic immobilization of trimethacryloyl thioalkanoates onto the surface of Si wafers, which were functionalized with 2-(phenacylthio)acetamido groups via a photoinduced reaction. Subsequent cross-linking via free radical polymerization as well as a phototriggered Diels-Alder reaction under mild conditions on the surface led to the desired nanosheets. Electrospray ionization mass spectrometry (ESI-MS), X-ray photoelectron spectroscopy (XPS), time-of-flight secondary ion mass spectrometry (ToF-SIMS), as well as infrared reflection-absorption spectroscopy (IRRAS) confirmed the success of individual surface-modification and cross-linking reactions. The thickness and lateral size of the cross-linked structures were determined by atomic force microscopy (AFM) for samples prepared on Si wafers functionalized with a self-assembled monolayer of 1H,1H,2H,2H-perfluorodecyl groups bearing circular pores obtained via a polymer blend lithographic approach, which led to the cross-linking reactions occurring in circular nanoareas (diameter of 50-640 nm) yielding an average thickness of 1.2 nm (radical cross-linking), 1.8 nm (radical cross-linking in the presence of 2,2,2-trifluoroethyl methacrylate as a comonomer), and 1.1 nm (photochemical cross-linking) of the nanosheets. PMID:25705846

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

    SciTech Connect

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

    1983-02-01

    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: PVA-dialdehyde blends post-treated with an acid or acid periodate solution and PVA-dialdehyde blends cross-linked during film formation (drying) by using a reagent with both aldehyde and acid functionality. Laboratory samples of each cross-linked type of film were prepared and evaluated in standard separator screening tests. Then pilotplant 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.

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

    SciTech Connect

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

    1982-03-01

    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.

  19. 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.

  20. 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.

  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. Controlled uptake and release of lysozyme from glycerol diglycidyl ether cross-linked oxidized starch microgel.

    PubMed

    Zhao, Luhai; Chen, Yuying; Li, Wei; Lu, Meiling; Wang, Shanshan; Chen, Xiaodong; Shi, Mengxuan; Wu, Jiande; Yuan, Qipeng; Li, Yuan

    2015-05-01

    A biodegradable microgel system based on glycerol-1,3-diglycidyl ether (GDGE) cross-linked TEMPO-oxidized potato starch polymers was developed for controlled uptake and release of proteins. A series of microgels were prepared with a wide range of charge density and cross-link density. We found both swelling capacity (SWw) and lysozyme uptake at saturation (?sat) increased with increasing degree of oxidation (DO) and decreasing cross-link density. Microgel of DO100% with a low cross-link density (RGDGE/polymer (w/w) of 0.025) was selected to be the optimum gel type for lysozyme absorption; ?sat increased with increasing pH and decreasing ionic strength. It suggests that the binding strength was the strongest at high pH and low ionic strength, which was recognized as the optimum absorption conditions. The lysozyme release was promoted at low pH and high ionic strength, which were considered to be the most suitable conditions for triggering protein release. These results may provide useful information for the controlled uptake and release of proteins by oxidized starch microgels. PMID:25659699

  3. Biology of hyaluronan: Insights from genetic disorders of hyaluronan metabolism.

    PubMed

    Triggs-Raine, Barbara; Natowicz, Marvin R

    2015-08-26

    Hyaluronan is a rapidly turned over component of the vertebrate extracellular matrix. Its levels are determined, in part, by the hyaluronan synthases, HAS1, HAS2, and HAS3, and three hyaluronidases, HYAL1, HYAL2 and HYAL3. Hyaluronan binding proteins also regulate hyaluronan levels although their involvement is less well understood. To date, two genetic disorders of hyaluronan metabolism have been reported in humans: HYAL1 deficiency (Mucopolysaccharidosis IX) in four individuals with joint pathology as the predominant phenotypic finding and HAS2 deficiency in a single person having cardiac pathology. However, inherited disorders and induced mutations affecting hyaluronan metabolism have been characterized in other species. Overproduction of hyaluronan by HAS2 results in skin folding and thickening in shar-pei dogs and the naked mole rat, whereas a complete deficiency of HAS2 causes embryonic lethality in mice due to cardiac defects. Deficiencies of murine HAS1 and HAS3 result in a predisposition to seizures. Like humans, mice with HYAL1 deficiency exhibit joint pathology. Mice lacking HYAL2 have variably penetrant developmental defects, including skeletal and cardiac anomalies. Thus, based on mutant animal models, a partial deficiency of HAS2 or HYAL2 might be compatible with survival in humans, while complete deficiencies of HAS1, HAS3, and HYAL3 may yet be recognized. PMID:26322170

  4. Biology of hyaluronan: Insights from genetic disorders of hyaluronan metabolism

    PubMed Central

    Triggs-Raine, Barbara; Natowicz, Marvin R

    2015-01-01

    Hyaluronan is a rapidly turned over component of the vertebrate extracellular matrix. Its levels are determined, in part, by the hyaluronan synthases, HAS1, HAS2, and HAS3, and three hyaluronidases, HYAL1, HYAL2 and HYAL3. Hyaluronan binding proteins also regulate hyaluronan levels although their involvement is less well understood. To date, two genetic disorders of hyaluronan metabolism have been reported in humans: HYAL1 deficiency (Mucopolysaccharidosis IX) in four individuals with joint pathology as the predominant phenotypic finding and HAS2 deficiency in a single person having cardiac pathology. However, inherited disorders and induced mutations affecting hyaluronan metabolism have been characterized in other species. Overproduction of hyaluronan by HAS2 results in skin folding and thickening in shar-pei dogs and the naked mole rat, whereas a complete deficiency of HAS2 causes embryonic lethality in mice due to cardiac defects. Deficiencies of murine HAS1 and HAS3 result in a predisposition to seizures. Like humans, mice with HYAL1 deficiency exhibit joint pathology. Mice lacking HYAL2 have variably penetrant developmental defects, including skeletal and cardiac anomalies. Thus, based on mutant animal models, a partial deficiency of HAS2 or HYAL2 might be compatible with survival in humans, while complete deficiencies of HAS1, HAS3, and HYAL3 may yet be recognized. PMID:26322170

  5. 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 ?-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. PMID:25431991

  6. Fabrication of cross-linked polyethyleneimine microfibers by reactive electrospinning with in situ photo-cross-linking by UV radiation.

    PubMed

    Xu, Xiaoming; Zhang, Jian-Feng; Fan, Yuwei

    2010-09-13

    The objective of this work is to demonstrate the feasibility of fabrication of cross-linked polyethyleneimine microfibers by a reactive photo-electrospinning technology. Linear polyethyleneimine (L-PEI) has been grafted with cross-linkable methacrylate moiety by reaction with glycidyl methacrylate (GMA), enabling the polymer to cross-link upon UV exposure. The photo-cross-linking reaction was characterized by a photo-rheometer. Neat L-PEI or methacrylated L-PEI tends to aggregate rendering it very difficult to electrospin into microfibers. A high molecular weight polyvinylpyrrolidone (PVP) is an efficient chain entanglement enhancer for both L-PEI and methacrylated L-PEI and helpful to maintain fibrous structure. An optimized composition consisted of 10% methacrylated L-PEI (less than 14.8% methacrylation of total L-PEI) combined with 2% PVP and 1% photoinitiator in ethanol was successfully electrospun into smooth cross-linked microfibers using the reactive electrospinning device. Diameters of cross-linked fibers can be controlled from 419 nm to 2 ?m depending on methacrylation degree and UV irradiation intensity. The resultant cross-linked L-PEI microfibers have demonstrated significantly improved solvent resistance, thermal stability, and mechanical properties. The distinguished characteristics of this novel reactive electrospinning technology are the high cross-linking efficiency and minimal toxic chemical residues in the products. The stability of the fibers can be readily modified and controlled by the cross-linking degree, which is of great importance for biomedical applications. PMID:20690594

  7. Biology and biotechnology of hyaluronan.

    PubMed

    Viola, Manuela; Vigetti, Davide; Karousou, Evgenia; D'Angelo, Maria Luisa; Caon, Ilaria; Moretto, Paola; De Luca, Giancarlo; Passi, Alberto

    2015-05-01

    The hyaluronan (HA) polymer is a critical component of extracellular matrix with a remarkable structure: is a linear and unbranched polymer without sulphate or phosphate groups. It is ubiquitous in mammals showing several biological functions, ranging from cell proliferation and migration to angiogenesis and inflammation. For its critical biological functions the amount of HA in tissues is carefully controlled by different mechanisms including covalent modification of the synthetic enzymes and epigenetic control of their gene expression. The concentration of HA is also critical in several pathologies including cancer, diabetes and inflammation. Beside these biological roles, the structural properties of HA allow it to take advantage of its capacity to form gels even at concentration of 1% producing scaffolds with very promising applications in regenerative medicine as biocompatible material for advanced therapeutic uses. In this review we highlight the biological aspects of HA addressing the mechanisms controlling the HA content in tissues as well as its role in important human pathologies. In the second part of the review we highlight the different use of HA polymers in the modern biotechnology. PMID:25971701

  8. Transglutaminase 2-Catalyzed Intramolecular Cross-Linking of Osteopontin.

    PubMed

    Christensen, Brian; Zachariae, Elias D; Scavenius, Carsten; Klverpris, Sren; Oxvig, Claus; Petersen, Steen V; Enghild, Jan J; Srensen, Esben S

    2016-01-19

    Osteopontin (OPN) is a multifunctional integrin-binding protein present in several tissues and body fluids. OPN is a substrate for the enzyme transglutaminase 2 (TG2), which catalyzes inter- and intramolecular cross-linking affecting the biological activity of the protein. Polymerization of OPN by intermolecular cross-linking has mostly been studied using relatively high TG2 concentrations, whereas the effect of lower concentrations of TG2 has remained unexplored. Here we show that TG2 at physiologically relevant concentrations predominantly catalyzes the formation of intramolecular cross-links in OPN. By site-directed mutagenesis and mass spectrometry, we demonstrate that Gln(42) and Gln(193) serve as the primary amine acceptor sites for isopeptide bond formation. We find that Gln(42) predominantly is linked to Lys(4) and that Gln(193) participates in a cross-link with Lys(154), Lys(157), or Lys(231). The formation of specific isopeptide bonds was not dependent on OPN phosphorylation, and similar patterns of cross-linking were observed in human and mouse OPN. Furthermore, we find that OPN purified from human urine contains the Lys(154)-Gln(193) isopeptide bond, indicating that intramolecular cross-linking of OPN occurs in vivo. Collectively, these data suggest that specific intramolecular cross-linking in the N- and C-terminal parts of OPN is most likely the dominant step in TG2-catalyzed modification of OPN. PMID:26678563

  9. Cross-linked carbon nanotubes buckygel actuators: an in-depth study

    NASA Astrophysics Data System (ADS)

    Gendron, David; Bubak, Grzegorz; Ceseracciu, Luca; Ansaldo, Alberto; Ricci, Davide

    2015-04-01

    Recently, materials that can convert electrical energy into mechanical work have drawn great attention. Applications in robotics, tactile or optical displays and microelectrochemical systems are currently investigated. Likewise, interest in actuators devices is increasing toward applications where low voltage and low weight properties are required. One way to achieve such prerequisites is to combine the mechanical and electronic properties of carbon nanotubes (CNTs) with the stability and conductivity of ionic liquids. Indeed, the CNTs can be dispersed in ionic liquids to form hybrid composites also named bucky gels, thanks to the non-covalent (?-? stacking and cation-?) interactions. In our previous studies, we demonstrated an improvement in actuator performance whilst using cross-linked CNTs. Indeed, our preliminary results showed an increase in the capacitance together with a faster response of the actuator. At the time, these results were explained by an actuation mechanism model. Herein, we designed new experiments in order to allow us to get a deeper insight in the effect the crosslinking process on the carbon nanotubes properties. Thus, we present a set of electromechanical and electrochemical data that shed light on the chemical modification of the CNTs, the different cross-linking strategies and also on the uses of cross-linked CNTS polymer blends. Finally, corresponding bucky gels actuators performances will also be discussed.

  10. Expression and purification of functionally active hyaluronan-binding domains from human cartilage link protein, aggrecan and versican: formation of ternary complexes with defined hyaluronan oligosaccharides.

    PubMed

    Seyfried, Nicholas T; McVey, Gillian F; Almond, Andrew; Mahoney, David J; Dudhia, Jayesh; Day, Anthony J

    2005-02-18

    The chondroitin sulfate proteoglycan aggrecan forms link protein-stabilized complexes with hyaluronan (HA), via its N-terminal G1-domain, that provide cartilage with its load bearing properties. Similar aggregates (potentially containing new members of the link protein family), in which other chondroitin sulfate proteoglycans (i.e. versican, brevican, and neurocan) substitute for aggrecan, may contribute to the structural integrity of many other tissues including skin and brain. In this study, cartilage link protein (cLP) and the G1-domains of aggrecan (AG1) and versican (VG1) were expressed in Drosophila S2 cells. The recombinant human proteins were found to have properties similar to those described for the native molecules (e.g. cLP was able to form oligomers, and HA decasaccharides were the minimum size that could compete effectively for their binding to polymeric HA). Gel filtration and protein cross-linking/matrix-assisted laser desorption ionization time-of-flight peptide fingerprinting showed that cLP and AG1 interact in the absence or presence of HA. Conversely, cLP and VG1 did not bind directly to each other in solution yet formed ternary complexes with HA24. N-linked glycosylation of AG1 and VG1 was demonstrated to be unnecessary for either HA binding or the formation of ternary complexes. Surprisingly, the length of HA required to accommodate two G1-domains was found to be significantly larger for aggrecan than versican, which may reflect differences in the conformation of HA stabilized on binding these proteins. PMID:15590670

  11. Mapping protein structural changes by quantitative cross-linking.

    PubMed

    Kukacka, Zdenek; Rosulek, Michal; Strohalm, Martin; Kavan, Daniel; Novak, Petr

    2015-11-01

    Chemical cross-linking is a promising technology for protein tertiary structure determination. Though the data has low spatial resolution, it is possible to obtain it at physiological conditions on proteins that are not amenable to standard high resolution techniques such as X-ray, NMR analysis and cryo-EM. Here we demonstrate the utilization of isotopically labeled chemical cross-linking to visualize protein conformation rearrangements. Since calmodulin exists in two distinct conformations (calcium-free and calcium-containing forms), we selected this protein for testing the potential and the limits of a new technique. After cross-linking of both calmodulin forms, the calcium-free and calcium-containing forms were mixed together and digested under different conditions and the products of proteolysis were monitored using high resolution mass spectrometry. Finally, the ratios of heavy/light cross-links were calculated by mMass open source platform. PMID:26048481

  12. Actin cross-linking and inhibition of the actomyosin motor.

    PubMed

    Kim, Eldar; Bobkova, Elena; Hegyi, Gyrgy; Muhlrad, Andras; Reisler, Emil

    2002-01-01

    Intrastrand cross-linking of actin filaments by ANP, N-(4-azido-2-nitrophenyl) putrescine, between Gln-41 in subdomain 2 and Cys-374 at the C-terminus, was shown to inhibit force generation with myosin in the in vitro motility assays [Kim et al. (1998) Biochemistry 37, 17801-17809]. To clarify the immobilization of which of these two sites inhibits the actomyosin motor, the properties of actins with partially overlapping cross-linked sites were examined. pPDM (N,N'-p-phenylenedimaleimide) and ABP [N-(4-azidobenzoyl) putrescine] were used to obtain actin filaments cross-linked ( approximately 50%) between Cys-374 and Lys-191 (interstrand) and Gln-41 and Lys-113 (intrastrand), respectively. ANP, ABP, and pPDM cross-linked filaments showed similar inhibition of their sliding speeds and force generation with myosin ( approximately 25%) in the in vitro motility assays. In analogy to ANP cross-linking of actin, pPDM and ABP cross-linkings did not change the strong S1 binding to actin and the V(max) and K(m) parameters of actomyosin ATPase. The similar effects of these three cross-linkings reveal the tight coupling between structural elements of the subdomain 2/subdomain 1 interface and show the importance of its dynamic flexibility to force generation with myosin. The possibility that actin cross-linkings inhibit rate-limiting steps in motion and force generation during myosin cross-bridge cycle was tested in stopped-flow experiments. Measurements of the rates of mantADP release from actoS1 and ATP-induced dissociation of actoS1 did not reveal any differences between un-cross-linked and ANP cross-linked actin in these complexes. These findings are discussed in terms of the uncoupling between force generation and other aspects of actomyosin interactions due to a constrained dynamic flexibility of the subdomain 2/subdomain 1 interface in cross-linked actin filaments. PMID:11772006

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

    SciTech Connect

    Hsu, L.; Philipp, W.H.; Sheibley, D.W.

    1981-06-09

    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, preferably 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. In that event, the mixture of polymer and cross-linking agent is formed into a sheet or film or the like and the film is cut to size and otherwise fabricated into a configuration suitable for a particular end use. The crosslinking reaction is then carried out to produce the final product.

  14. 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

    2016-03-01

    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. PMID:26862769

  15. 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.

  16. Enzymatically Cross-Linked Bovine Lactoferrin as Injectable Hydrogel for Cell Delivery

    PubMed Central

    Amini, Ashley A.; Kan, Ho-Man; Cui, Zhanwu; Maye, Peter

    2014-01-01

    Lactoferrin (LF), a 78 kDa glycoprotein, has recently been recognized as an effector molecule in the skeleton due to its ability to decrease osteoclastogenesis and increase osteoblast proliferation, survival, and differentiation. The objective of the study is to investigate the feasibility of developing an injectable hydrogel from bovine lactoferrin (bLF) as a cell delivery vehicle. The study demonstrated the feasibility of cross-linking tyramine substituted bLF in the presence of horse radish peroxidase and hydrogen peroxide (H2O2). The gel presented a mild environment to maintain mouse bone marrow-derived stromal cell (mBMSC) viability and proliferation. Stromal cells derived from multiple gene reporter transgenic mouse (Ibsp-Topaz/Dmp1-mCherry) line showed the ability of the cells to undergo osteogenic differentiation in the hydrogel when cultured in mineralization media. The cross-linked gel supported protein phosphorylation/de-phosphorylation in the encapsulated MC3T3-E1 cells. bLF and bLF gel also showed the ability to modulate growth factor production in mBMSCs. PMID:24802947

  17. Enzymatically cross-linked bovine lactoferrin as injectable hydrogel for cell delivery.

    PubMed

    Amini, Ashley A; Kan, Ho-Man; Cui, Zhanwu; Maye, Peter; Nair, Lakshmi S

    2014-11-01

    Lactoferrin (LF), a 78 kDa glycoprotein, has recently been recognized as an effector molecule in the skeleton due to its ability to decrease osteoclastogenesis and increase osteoblast proliferation, survival, and differentiation. The objective of the study is to investigate the feasibility of developing an injectable hydrogel from bovine lactoferrin (bLF) as a cell delivery vehicle. The study demonstrated the feasibility of cross-linking tyramine substituted bLF in the presence of horse radish peroxidase and hydrogen peroxide (H2O2). The gel presented a mild environment to maintain mouse bone marrow-derived stromal cell (mBMSC) viability and proliferation. Stromal cells derived from multiple gene reporter transgenic mouse (Ibsp-Topaz/Dmp1-mCherry) line showed the ability of the cells to undergo osteogenic differentiation in the hydrogel when cultured in mineralization media. The cross-linked gel supported protein phosphorylation/de-phosphorylation in the encapsulated MC3T3-E1 cells. bLF and bLF gel also showed the ability to modulate growth factor production in mBMSCs. PMID:24802947

  18. Moulded cross-linked chitosan matrix systems for controlled drug release.

    PubMed

    Tarirai, Clemence; Enslin, Gill M; Steenekamp, Jan H; Hamman, Josias H

    2011-08-01

    Matrix-type drug delivery systems were prepared by moulding and drying cross-linked chitosan gels in 24-well plates and they were evaluated in terms of their physical properties, drug content, surface morphology and swelling. Furthermore, the in vitro drug release profiles were subjected to kinetic modelling at two different pH values. In general, the moulded matrix systems showed statistically significantly slower drug release compared to immediate release tablets as measured by the mean dissolution time. Drug release from the moulded matrix systems prepared from chitosan cross-linked with tripolyphosphate was pH-dependent as can be seen from the release exponent value (n) of 0.75 at pH 5.8 (anomalous transport, erosion), while the n value was only 0.40 at pH 7.4 (Fickian diffusion). The matrix systems obtained from chitosan cross-linked with sodium lauryl sulphate showed higher swelling but mostly Fickian diffusional release (n?=?0.25 at pH 7.4, n?=?0.41 at pH 5.8). PMID:20214415

  19. The filamentous actin cross-linking/bundling activity of mammalian formins.

    PubMed

    Esue, Osigwe; Harris, Elizabeth S; Higgs, Henry N; Wirtz, Denis

    2008-12-12

    Formins are multidomain proteins that regulate actin filament dynamics and are defined by the formin homology 2 domain. Biochemical assays suggest that mammalian formins display actin-filament nucleation, severing, and bundling activities. Whether formins can cross-link actin filaments into viscoelastic arrays and the effectiveness of formins' bundling activity compared with that of important filamentous actin (F-actin) cross-linking/bundling proteins are unknown. Here, we used rigorous in vitro rheologic assays to deconvolve the dynamic cross-linking activity from the bundling activity of formin FRL1 and the closely related mDia1 and mDia2. In addition, we compared these formins with the canonical F-actin bundling protein fascin and cross-linking/bundling proteins alpha-actinin and filamin. We found that FRL1 and mDia2, but not mDia1, can help F-actin form highly elastic networks. FRL1 and mDia2 mediate the formation of highly elastic F-actin networks as effectively and rapidly as alpha-actinin and filamin but only past a relatively high actin-to-formin molar ratio of 50:1. Past that threshold molar ratio, the mechanical properties of F-actin/formin networks are independent of formin concentration, similar to fascin. Moreover, unlike those for alpha-actinin and filamin but similar to those for fascin, F-actin/formin networks show no strain-induced hardening. mDia1 cannot bundle F-actin but can weakly cross-link filaments at high concentrations. Point mutagenesis reveals that reducing the barbed-end binding activity of FRL1 and mDia2 greatly enhances the rate of formation of F-actin gels but does not significantly affect the mechanical properties of the resulting networks at steady state. Together, these results suggest that the mechanical behaviors of FRL1 and mDia2 are fundamentally different from those of cross-linking/bundling proteins alpha-actinin and filamin but qualitatively similar to the mechanical behavior of the bundling protein fascin, albeit with a dramatically increased (>10-fold) threshold concentration for transition to bundling, which nevertheless leads to much stiffer F-actin networks than fascin. PMID:18835565

  20. Collagen/elastin hydrogels cross-linked by squaric acid.

    PubMed

    Skopinska-Wisniewska, J; Kuderko, J; Bajek, A; Maj, M; Sionkowska, A; Ziegler-Borowska, M

    2016-03-01

    Hydrogels based on collagen and elastin are very valuable materials for medicine and tissue engineering. They are biocompatible; however their mechanical properties and resistance for enzymatic degradation need to be improved by cross-linking. Up to this point many reagents have been tested but more secure reactants are still sought. Squaric acid (SqAc), 3,4-dihydroxy 3-cyclobutene 1,2-dione, is a strong, cyclic acid, which reacts easily with amine groups. The properties of hydrogels based on collagen/elastin mixtures (95/5, 90/10) containing 5%, 10% and 20% of SqAc and neutralized via dialysis against deionized water were tested. Cross-linked, 3-D, transparent hydrogels were created. The cross-linked materials are stiffer and more resistant to enzymatic degradation than those that are unmodified. The pore size, swelling ability and surface polarity are reduced due to 5% and 10% of SqAc addition. At the same time, the cellular response is not significantly affected by the cross-linking. Therefore, squaric acid would be regarded as a safe, effective cross-linking agent. PMID:26706512

  1. 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.

  2. Boundary lubricant polymer films: effect of cross-linking.

    PubMed

    Giasson, Suzanne; Lagleize, Jeanne-Marie; Rodríguez-Hernández, Juan; Drummond, Carlos

    2013-10-22

    We have studied the adsorption and lubricant properties of a multifunctional triblock copolymer poly(L-lysine)-b-poly(acrylic acid)-b-poly(L-lysine). In particular, we investigated the nature of the layer adsorbed under different conditions of polymer and salt concentration and the lubricant properties of the polymer layer before and after its chemical cross-linking by bridging the poly(acrylic acid) blocks. We found that the amount of polymer adsorbed is controlled by the ionic strength and the polymer concentration in the solution. In all cases, the self-assembled polymer layer is a poor lubricant before cross-linking, but the cohesion and load-carrying ability of the layer are substantially improved by this reaction. However, the chemically cross-linked coating has a limited deformation capacity as a consequence of its permanent network nature, and irreversible damage is observed after excessive strain of the film. PMID:24053315

  3. Functional synergy of actin filament cross-linking proteins.

    PubMed

    Tseng, Yiider; Schafer, Benjamin W; Almo, Steven C; Wirtz, Denis

    2002-07-12

    The organization of filamentous actin (F-actin) in resilient networks is coordinated by various F-actin cross-linking proteins. The relative tolerance of cells to null mutations of genes that code for a single actin cross-linking protein suggests that the functions of those proteins are highly redundant. This apparent functional redundancy may, however, reflect the limited resolution of available assays in assessing the mechanical role of F-actin cross-linking/bundling proteins. Using reconstituted F-actin networks and rheological methods, we demonstrate how alpha-actinin and fascin, two F-actin cross-linking/bundling proteins that co-localize along stress fibers and in lamellipodia, could synergistically enhance the resilience of F-actin networks in vitro. These two proteins can generate microfilament arrays that "yield" at a strain amplitude that is much larger than each one of the proteins separately. F-actin/alpha-actinin/fascin networks display strain-induced hardening, whereby the network "stiffens" under shear deformations, a phenomenon that is non-existent in F-actin/fascin networks and much weaker in F-actin/alpha-actinin networks. Strain-hardening is further enhanced at high rates of deformation and high concentrations of actin cross-linking proteins. A simplified model suggests that the optimum results of the competition between the increased stiffness of bundles and their decreased density of cross-links. Our studies support a re-evaluation of the notion of functional redundancy among cytoskeletal regulatory proteins. PMID:12006593

  4. FTIR Spectroscopic Studies on Cross Linking of SU-8 Photoresist

    NASA Astrophysics Data System (ADS)

    Kalaiselvi, S. M. P.; Tan, T. L.; Rawat, R. S.; Lee, P.; Heussler, S. P.; Breese, M. B. H.

    2013-11-01

    The usage of chemically-amplified, negative tone SU-8 photoresist is numerous, spanning industrial, scientific and medical fields. Hence, in this study, some preliminary studies were conducted to understand the dosage and heat treatment requirements of the SU-8 photoresist essential for pattern generation using X-ray lithography. In this work, using Synchrotron as the X-ray source, SU-8 photoresist was characterized for X-ray lithography in terms of its process parameters such as X-ray exposure dose, post exposure bake (PEB) time and temperature for various photoresist thicknesses which is considered worthwhile in view of applications of SU-8 for the fabrication of very high aspect ratio micro structures. The process parameters were varied and the resultant cross linking of the molecular chains of the photoresist was accurately monitored using a Fourier Transform Infra-Red (FTIR) spectrometer and the results are discussed. The infrared absorption peak at 914 cm-1 in the spectrum of the SU-8 photoresist was found to be a useful indicator for the completion of cross linking in the SU-8 photoresist. Results show that the cross linking of the SU-8 photoresist is at a higher rate from 0 J/cm3 to 30 J/cm3 after which the peak almost saturates regardless of the PEB time. It is a good evidence for the validation of dosage requirement of SU-8 photoresist for effective completion of cross linking, which in turn is a requirement for efficient fabrication of micro and nano structures. An analogous behavior was also observed between the extent of cross linking and the PEB time and temperature. The rate of cross linking declines after a certain period of PEB time regardless of PEB temperature. The obtained results also show a definite relation between variation of the absorbance area of the peak at 914 cm-1 and the X-ray exposure dose.

  5. Enzymatic production of specifically distributed hyaluronan oligosaccharides.

    PubMed

    Yuan, Panhong; Lv, Mengxian; Jin, Peng; Wang, Miao; Du, Guocheng; Chen, Jian; Kang, Zhen

    2015-09-20

    High-molecular-mass hyaluronan (HA) was controllably depolymerized in pure aqueous solution with recombinant leech hyaluronidase (HAase). The HAase concentration per unit HA and hydrolysis time played important roles in molecular mass distribution. By modulating the concentrations of HAase and controlling the hydrolysis time, any molar-mass-defined HA oligomers could be efficiently and specifically produced on a large scale (40 g/L), such as HA oligosaccharides with weight-average molar mass of 4000, 10,000, and 30,000Da and end hydrolysates containing only HA6 and HA4. High performance liquid chromatography-size exclusion chromatography, polyacrylamide gel electrophoresis, capillary zone electrophoresis, and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry confirmed low polydispersity of the produced molar-mass-defined HA oligosaccharides. Therefore, large-scale production of defined HA oligosaccharides with narrow molecular mass distribution will significantly promote progress in related research and its potential applications. PMID:26050905

  6. 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.

  7. Advanced Corneal Cross-Linking System with Fluorescence Dosimetry

    PubMed Central

    Friedman, Marc D.; Pertaub, Radha; Usher, David; Sherr, Evan; Kamaev, Pavel; Muller, David

    2012-01-01

    Purpose. This paper describes an advanced system that combines corneal cross-linking with riboflavin with fluorescence dosimetry, the ability to measure riboflavin diffusion within the cornea both before and during UVA treatment. Methods and Results. A corneal cross-linking system utilizing a digital micromirror device (DMD) was assembled and used to measure diffusion coefficients of 0.1% riboflavin in 20% dextran in porcine eyes. A value of (3.3 ± 0.2) × 10−7 cm2/s was obtained for the stroma. Diffusion coefficients for the transepithelial formulation of 0.1% riboflavin in 0.44% saline and 0.02% BAK were also measured to be 4.7 ± 0.3 × 10−8 cm2/s for epithelium only and (4.6 ± 0.4) × 10−7 cm2/s for stroma only. Riboflavin consumption during a UVA treatment was also demonstrated. Conclusion. A new advanced corneal cross-linking system with fluorescence dosimetry of riboflavin has been demonstrated. It is hoped that this method may play a significant role in determining the underlying mechanisms of corneal cross-linking and assist with the development of additional riboflavin formulations. Moreover, dosimetry may prove valuable in providing a method to account for the biological differences between individuals, potentially informing cornea-specific UVA treatment doses in real time. PMID:22792444

  8. Molecular mechanisms in deformation of cross-linked hydrogel nanocomposite.

    PubMed

    Mathesan, Santhosh; Rath, Amrita; Ghosh, Pijush

    2016-02-01

    The self-folding behavior in response to external stimuli observed in hydrogels is potentially used in biomedical applications. However, the use of hydrogels is limited because of its reduced mechanical properties. These properties are enhanced when the hydrogels are cross-linked and reinforced with nanoparticles. In this work, molecular dynamics (MD) simulation is applied to perform uniaxial tension and pull out tests to understand the mechanism contributing towards the enhanced mechanical properties. Also, nanomechanical characterization is performed using quasi static nanoindentation experiments to determine the Young's modulus of hydrogels in the presence of nanoparticles. The stress-strain responses for chitosan (CS), chitosan reinforced with hydroxyapatite (HAP) and cross-linked chitosan are obtained from uniaxial tension test. It is observed that the Young's modulus and maximum stress increase as the HAP content increases and also with cross-linking process. Load displacement plot from pullout test is compared for uncross-linked and cross-linked chitosan chains on hydroxyapatite surface. MD simulation reveals that the variation in the dihedral conformation of chitosan chains and the evolution of internal structural variables are associated with mechanical properties. Additional results reveal that the formation of hydrogen bonds and electrostatic interactions is responsible for the above variations in different systems. PMID:26652360

  9. 21 CFR 177.1211 - Cross-linked polyacrylate copolymers.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... weight of aqueous sodium chloride solution at 20 °C for 24 hours. The low molecular weight extractives... applied mass). The solvent used shall be at least 60 milliliters aqueous sodium chloride solution per gram... polyacrylate copolymers consist of: (1) The grafted copolymer of cross-linked sodium polyacrylate identified...

  10. 21 CFR 177.1211 - Cross-linked polyacrylate copolymers.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ...). The solvent used shall be at least 60 milliliters aqueous sodium chloride solution per gram of... grafted copolymer of cross-linked sodium polyacrylate identified as 2-propenoic acid, polymers with N,N-di-2-propenyl-2-propen-1-amine and hydrolyzed polyvinyl acetate, sodium salts, graft (CAS Reg....

  11. 21 CFR 177.1211 - Cross-linked polyacrylate copolymers.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... weight of aqueous sodium chloride solution at 20 °C for 24 hours. The low molecular weight extractives... applied mass). The solvent used shall be at least 60 milliliters aqueous sodium chloride solution per gram... polyacrylate copolymers consist of: (1) The grafted copolymer of cross-linked sodium polyacrylate identified...

  12. 21 CFR 177.1211 - Cross-linked polyacrylate copolymers.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... weight of aqueous sodium chloride solution at 20 °C for 24 hours. The low molecular weight extractives... applied mass). The solvent used shall be at least 60 milliliters aqueous sodium chloride solution per gram... polyacrylate copolymers consist of: (1) The grafted copolymer of cross-linked sodium polyacrylate identified...

  13. 21 CFR 177.1211 - Cross-linked polyacrylate copolymers.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... weight of aqueous sodium chloride solution at 20 °C for 24 hours. The low molecular weight extractives... applied mass). The solvent used shall be at least 60 milliliters aqueous sodium chloride solution per gram... polyacrylate copolymers consist of: (1) The grafted copolymer of cross-linked sodium polyacrylate identified...

  14. Clinical fracture of cross-linked UHMWPE acetabular liners.

    PubMed

    Furmanski, Jevan; Anderson, Martin; Bal, Sonny; Greenwald, A Seth; Halley, David; Penenberg, Brad; Ries, Michael; Pruitt, Lisa

    2009-10-01

    Highly cross-linked ultrahigh molecular weight polyethylene (UHMWPE) is increasingly used as a bearing material in total hip replacements. Cross-linking of UHMWPE has been shown to increase wear resistance but decrease its fracture resistance. We analyzed the clinical fracture failure of four cross-linked UHMWPE total hip replacement components of four different designs via microscopic observation of the fracture surfaces, and found that in all cases fractures initiated at stress concentrations in an unsupported region of the component (termed the elevated rim). Finite element analyses (FEA) of each individual implant design were then conducted. Results from this analysis demonstrated that the predicted magnitude and orientation of maximum principal stress due to mechanical loading of the elevated rim was sufficient to propagate initiated fatigue cracks in each case. FEA also predicted that cracks may arrest after some amount of growth due to a steep stress gradient near the initiation site. Further, while anatomical positioning of the implant and material properties affect the risk of fracture, we examined whether these failures are strongly related to the notched elevated rim design feature that is common to the four failed cases presented here. We believe that cross-linked UHMWPE remains an excellent bearing material for total hip replacements but that designs employing this material should mitigate stress concentrations or other design features that increase the risk of fracture. PMID:19643471

  15. 21 CFR 177.2420 - Polyester resins, cross-linked.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Polyester resins, cross-linked. 177.2420 Section 177.2420 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN CONSUMPTION (CONTINUED) INDIRECT FOOD ADDITIVES: POLYMERS Substances for Use Only as Components of Articles Intended for...

  16. 21 CFR 177.2420 - Polyester resins, cross-linked.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Polyester resins, cross-linked. 177.2420 Section 177.2420 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN CONSUMPTION (CONTINUED) INDIRECT FOOD ADDITIVES: POLYMERS Substances for Use Only as Components of Articles Intended for...

  17. 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

  18. Viscoelastic Nanomechanics of Ionically Cross-linked Polyelectrolyte Networks

    NASA Astrophysics Data System (ADS)

    Han, Biao; Lee, Daeyeon; Han, Lin

    2015-03-01

    Understanding the mechanics of ionic polyelectrolyte networks is critical for applications where nm-to-um mechanics is the key to success. This study aims to reveal the roles of ionic cross-links and fixed charges in the viscoelasticity of layer-by-layer poly(allylamine hydrochloride)/poly(acrylic acid) microfilms, PAH/PAA, a complex held by pH-sensitive amine-carboxyl links. AFM-nanoindentation and force relaxation (tip R =12.5um) was performed at ionic strength(IS) =0.01-1.0M, pH =5.5-2.0 (pKa of PAA =2.3). When pH changes from 5.5 to 2.0, the films swell for 4x from densely linked, net neutral state to loosely linked, positively charged one. A >100x reduction in indentation modulus was observed at all IS, suggesting the dominance of decrease in cross-link density. In most states, more than 90% force relaxation was observed, where cross-link breaking/reformation likely dominates viscoelasticity. However, at pH =2.5 and IS =0.01M, when electrical double layer repulsion is important (Debye length =3nm), relaxation was about 60%, highlighting the contribution of fixed charges. In summary, this study revealed unique viscoelastic behaviors of PAH/PAA due to the pH- and IS-dependent cross-link and charge densities.

  19. Femtosecond laser collagen cross-linking without traditional photosensitizers

    NASA Astrophysics Data System (ADS)

    Guo, Yizang; Wang, Chao; Celi, Nicola; Vukelic, Sinisa

    2015-03-01

    Collagen cross-linking in cornea has the capability of enhancing its mechanical properties and thereby providing an alternative treatment for eye diseases such as keratoconus. Currently, riboflavin assisted UVA light irradiation is a method of choice for cross-link induction in eyes. However, ultrafast pulsed laser interactions may be a powerful alternative enabling in-depth treatment while simultaneously diminishing harmful side effects such as, keratocyte apoptosis. In this study, femtosecond laser is utilized for treatment of bovine cornea slices. It is hypothesized that nonlinear absorption of femtosecond laser pulses plays a major role in the maturation of immature cross-links and the promotion of their growth. Targeted irradiation with tightly focused laser pulses allows for the absence of a photosensitizing agent. Inflation test was conducted on half treated porcine cornea to identify the changes of mechanical properties due to laser treatment. Raman spectroscopy was utilized to study subtle changes in the chemical composition of treated cornea. The effects of treatment are analyzed by observing shifts in Amide I and Amide III bands, which suggest deformation of the collagen structure in cornea due to presence of newly formed cross-links.

  20. 21 CFR 177.2420 - Polyester resins, cross-linked.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Polyester resins, cross-linked. 177.2420 Section 177.2420 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN CONSUMPTION (CONTINUED) INDIRECT FOOD ADDITIVES: POLYMERS Substances for Use Only as Components of Articles Intended for...

  1. 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

  2. Structural characterization of Y1 and Y2 receptors for neuropeptide Y and peptide YY by affinity cross-linking

    SciTech Connect

    Sheikh, S.P.; Williams, J.A. )

    1990-05-15

    Pharmacological studies indicate that peptide YY (PYY) and neuropeptide Y interact with multiple binding sites, categorized as Y1 and Y2 subtypes. In order to identify and structurally characterize the Y1 and Y2 receptors we covalently cross-linked (125I-Tyr36)PYY to its receptors. The Y2 receptor in rat hippocampus and rabbit kidney membranes was affinity labeled using different homo- and heterobifunctional cross-linking reagents. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography resulted in a major labeled protein band of Mr = 50,000 in both hippocampal and kidney membranes, which was unaffected by reducing agents. The Y1 receptor was analyzed in membranes from the MC-IXC human neuroblastoma cell line. Autoradiography revealed two labeled bands at Mr = 70,000 and 45,000. As the intensity of the Mr = 45,000 band was reduced by protease inhibitors, it is likely that this band is a degradation product of the larger band. Labeling of these proteins was obtained only when N-5-azido-2-nitrobenzoyloxysuccinimide was employed for cross-linking followed by exposure to UV light. Labeling of the two cross-linked bands was unaffected by reducing agents. The binding of radiolabeled PYY and the intensity of the cross-linked bands, for both the Y1 and Y2 receptors, were inhibited similarly in a dose-dependent manner by increasing concentrations of unlabeled PYY. When exposed to agarose-coupled lectins, the detergent-solubilized Y1 receptor-hormone complex was completely adsorbed by wheat germ agglutinin and partially by ricin communis II. The cross-linked Y2 receptor was almost totally adsorbed by wheat germ agglutinin-agarose and partially adsorbed by concanavalin A. The adsorptions were in all cases blocked by the appropriate hapten sugar.

  3. Graft copolymerization onto polybutadiene: Cross-linking and thermal degradation of vinyl polymers and copolymers

    NASA Astrophysics Data System (ADS)

    Jiang, Dayue (David)

    This work consists of three parts. In Part I, the graft copolymerization of methyl methacrylate, methyl acrylate, methacylic acid and acrylic acid onto polybutadiene and its copolymers by benzoyl peroxide, BPO, or 2, 2'azobis(2-methylpropionitrile), AIBN, initiation were explored. The results show that these monomers can be grafted onto butadiene region of butadiene-containing polymers. The extent of both graft copolymerization and homopolymerization are dependent on the time and temperature of the reaction and the concentration of all of the reactants. One must specify the monomer, initiator and solvent for the efficient graft copolymerization. The methyl methacrylate adds directly to the radical sites which are formed on the backbone by the interaction of the polymer and the primary radical form the initiator, while for the other three monomers, the graft copolymerization occurs by addition of macro-radical to the double bonds. In Part II, the cross-linking of polybutadiene, butadiene-styrene copolymers, and polystyrene by irradiation, thermal and chemical processes, and Friedel-Crafts chemistry and the effect of cross-linking on the thermal stability were investigated. The proof of cross-linking of the polymer comes from the insolubility of the product after the cross-linking reaction and is characterized by gel content and swelling ratio. The results show that the thermal stability of the polymer can be improved by cross-linking. In Part III, the thermal degradation of three vinyl polymers, poly(vinylsulfonic acid) and its sodium salt and poly(vinylphosphonic acid) were studied by combination technique: TGA/FTIR. The results show that TGA/FTIR combined with analysis of residues provides an excellent opportunity to understand the degradation pathway of the compounds. The observation of foaming indicates that the char which is formed contains carbon as well as the inorganic salts which have been observed. The carbon is in a partially graphitized form. The salts produce a very large amount of char and may prove useful to impart increased flame retardance to other polymers if they can be incorporated in a convenient manner into these polymers.

  4. Effect of chemical cross-linking on gelatin membrane solubility with a non-toxic and non-volatile agent: terephthalaldehyde.

    PubMed

    Biscarat, Jennifer; Galea, Benjamin; Sanchez, Jos; Pochat-Bohatier, Celine

    2015-03-01

    In this paper, terephthalaldehyde (TPA) is proposed as non-toxic and non-volatile gelatin cross-linker. Optimal cross-linking parameters (TPA/gelatin ratio, temperature) were first determined from in situ rheological measurements on gelatin solutions and from chemical tests with 2,4,6-trinitrobenzenesulfonic acid (TNBS assays) on gelatin gel. The highest cross-linking ratio was achieved for a concentration of 0.005 g TPA/g gelatin at 60C. The impact of TPA cross-linking on gelatin membrane functional properties (water swelling ratio, water vapor sorption and mechanical properties) was measured. TPA cross-linking increased 17 times the liquid water resistance duration of gelatin films, and delayed the entry of vapor water in the polymer matrix for 7 days, indicating that TPA increased the hydrophobic character of the gelatin matrix. PMID:25478961

  5. Bacteriophage HK97 structure: wholesale covalent cross-linking between the major head shell subunits.

    PubMed Central

    Popa, M P; McKelvey, T A; Hempel, J; Hendrix, R W

    1991-01-01

    We describe initial genetic and structural characterizations of HK97, a temperate bacteriophage of Escherichia coli. We isolated 28 amber mutants, characterized them with respect to what phage-related structures they make, and mapped many of them to restriction fragments of genomic DNA. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of HK97 virions revealed nine different protein species plus a substantial amount of material that failed to enter the gel, apparently because it is too large. Five proteins are tail components and are assigned functions as tail fiber subunit, tail length template, and major shaft subunit (two and possibly three species). The four remaining proteins and the material that did not enter the gel are head components. One of these proteins is assigned as the portal subunit, and the remaining three head proteins in the gel and the material that did not enter the gel are components of the head shell. All of the head shell protein species have apparent molecular masses well in excess of 100 kDa; they share amino acid sequence with each other and also with a 42-kDa protein that is found in infected lysates and as the major component of prohead structures that accumulate in infections by one of the amber mutants. We propose that all of the head shell species found in mature heads are covalently cross-linked oligomers derived from the 42-kDa precursor during head shell maturation. Images PMID:1709700

  6. INJECTABLE IN SITU CROSS-LINKING HYDROGELS FOR LOCAL ANTIFUNGAL THERAPY

    PubMed Central

    Hudson, Sarah; Langer, Robert; Fink, Gerald R.; Kohane, Daniel S.

    2009-01-01

    Invasive fungal infections can be devastating, particularly in immunocompromised patients, and difficult to treat with systemic drugs. Furthermore, systemic administration of those medications can have severe side effects. We have developed an injectable local antifungal treatment for direct administration into existing or potential sites of fungal infection. Amphotericin B (AmB), a hydrophobic, potent, and broad-spectrum antifungal agent, was rendered water-soluble by conjugation to a dextran-aldehyde polymer. The dextran-aldehyde-AmB conjugate retained antifungal efficacy against C. albicans. Mixing carboxymethylcellulose-hydrazide with dextran-aldehyde formed a gel that cross-linked in situ by formation of hydrazone bonds. The gel provided in vitro release of antifungal activity for 11 days, and contact with the gel killed Candida for three weeks. There was no apparent tissue toxicity in the murine peritoneum and the gel caused no adhesions. Gels produced by entrapment of a suspension of AmB in CMC-dextran without conjugation of drug to polymers did not release fungicidal activity, but did kill on contact. Injectable systems of these types, containing soluble or insoluble drug formulations, could be useful for treatment of local antifungal infections, with or without concurrent systemic therapy. PMID:19942285

  7. Characterization of the somatogenic receptor in rat liver. Hydrodynamic properties and affinity cross-linking

    SciTech Connect

    Husman, B.; Haldosen, L.A.; Andersson, G.; Gustafsson, J.A.

    1988-03-15

    Rat liver somatogenic receptors have been characterized by gel permeation chromatography, sucrose density gradients in H/sub 2/O and D/sub 2/O, and affinity cross-linking using /sup 125/I-bovine growth hormone (bGH) as a specific somatogenic receptor ligand. Cross-linking of /sup 125/I-bovine growth hormone to a Triton X-100-treated low density fraction isolated from livers of late pregnant rats followed by sodium dodecylsulfate-polyacrylamide gel electrophoresis under reducing conditions showed three major binders with Mr 95,000, 86,000, and 43,000 and a minor binder of Mr 55,000, after correction for bound ligand assuming a 1:1 binding ratio of ligand-receptor. The Mr 86,000, 55,000, and 43,000 species were recovered in the detergent-soluble supernatant after high-speed centrifugation, whereas the Mr 95,000 species remained Triton X-100 insoluble. Detergent-soluble /sup 125/I-bGH-receptor complexes were further analyzed by sedimentation into sucrose density gradients. The sedimentation coefficient was S20,w = 5.2 S and the partial specific volume v = 0.72 ml/g. Gel permeation chromatography on a Sepharose S-400 column indicated a Stokes radius of 61 A for the /sup 125/I-bGH-receptor-Triton X-100 complex. Based on these figures, the molecular weight of the complex was calculated as 131,100. The molecular weight of the ligand-free receptor-Triton X-100 complex was calculated as Mr 109,100. Affinity cross-linking and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the 61 A peak from Sephacryl S-400 chromatography (cf. above) showed two binding entities, one major and one minor with Mr values 86,000 and 43,000, respectively, in the absence of reductant. When electrophoresis was run in the presence of reductant the Mr 43,000 species was the major binding entity.

  8. Hyaluronan in Tubular and Interstitial Nephrocalcinosis

    NASA Astrophysics Data System (ADS)

    Verkoelen, Carl F.

    2007-04-01

    Hyaluronan (HA) is the major glycosaminoglycan (GAG) component of the renal medullary interstitium. HA is extremely large (up to 104 kDa) and composed of thousands repeating disaccharides of glucuronic acid (GlcUA) and N-acetylglucosamine (GlcNAc). HA is synthesized by hyaluronan synthases (HASs) and degraded by hyaluronidases (Hyals). The production of HA by renomedullary interstitial cells is mediated by local osmolality. When excess water needs to be excreted, increased interstitial HA seems to antagonize water reabsorption, while the opposite occurs during water conservation. Hence, papillary interstitial HA is low and Hyal high during anti-diuresis, whereas during diuresis HA is high and Hyal low. The polyanion HA plays a role in the reabsorption of hypotonic fluid by immobilizing cations (Na+) via the carboxylate (COO-) groups of GlcUA. The binding of Ca2+ to anionic HA is probably also responsible for the fact that the papilla does not become a stone despite the extremely high interstitial phosphate and oxalate. HA is also an excellent crystal binding molecule. The expression of HA at the luminal surface of renal tubular cells leads to tubular nephrocalcinosis (tubular NC). Calcium staining methods (Von Kossa, Yasue) demonstrated that crystallization inhibitors cannot avoid the occasional precipitation of calcium phosphate in the papillary interstitium (interstitial NC). These crystals are probably immediately immobilized by the gel-like HA matrix. After ulcerating through the pelvic wall the calcified matrix becomes a Randall's plaque. The attachment of calcium oxalate crystals from the primary urine to plaque may ultimately lead to the development of clinical stones in the renal calyces (nephrolithiasis).

  9. Transparent Humidity Sensor Using Cross-Linked Polyelectrolyte Membrane

    SciTech Connect

    Zhang, Q.; Smith, James R.; Saraf, Laxmikant V.; Hua, Feng

    2009-07-02

    This paper describes the fabrication of a porous cross-linked polyelectrolyte membrane and the characterization of its humidity sensitivity performance. Electrostatic self-assembly, combined with acid treatment, and post-deposition annealing produced the membrane. The fabrication process offers the ability to control the thickness of the membrane, as well as enabling the engineering of the humidity sensitivity properties. A transparent humidity sensor was fabricated by integrating the membrane between two parallel electrodes. In order to improve the moisture absorption and diffusion, both the polyelectrolyte layer and the electrode were made porous. The membrane was cross-linked to enhance the durability in high humid environments. Such a polyelectrolyte membrane showed high sensitivity to relative humidity variation over a range of 25%99%. The see-through property of the structure adds extra features and benefits to the sensor.

  10. Hydration and swelling of amorphous cross-linked starch microspheres.

    PubMed

    Wojtasz, Joanna; Carlstedt, Jonas; Fyhr, Peter; Kocherbitov, Vitaly

    2016-01-01

    Hydration of cross-linked starch microspheres, commercially available as a medical device, was investigated using a multi-method approach. We found that the uptake of water is accompanied by substantial swelling and changes of the polymer structure. Sorption calorimetry provided information about thermodynamics of water sorption, revealed presence of isothermal glass transition and absence of hydration-induced crystallization, observed in non-cross linked starch material. The changes in the surface and bulk properties of microspheres at different water-starch concentrations were investigated using synchrotron radiation X-ray scattering and analyzed using concept of fractals. The obtained information, combined with the results of differential scanning calorimetry, was used to construct a phase diagram of the studied material. Finally, hydration induced evolution of polymer structure revealed by the X-ray scattering was linked to the changes observed during swelling with optical microscopy. PMID:26453872

  11. Diepoxybutane interstrand cross-links induce DNA bending.

    PubMed

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

    2012-02-01

    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

  12. Estimating the Degree of Cross-Linking in Rubber

    NASA Technical Reports Server (NTRS)

    Fedors, R. F.

    1983-01-01

    Degree of cross-linking or network chain concentration of rubber estimated with aid of new method. Quantity is needed in studies of mechanical behavior of rubber. New method is based on finding rubber follows different stress/ strain relationships in extension and retraction. When rubber specimen is stretched to given extension ration and released. Stress-vs-strain curve follows two paths: one for extension and other for retraction.

  13. Cytokines and growth factors cross-link heparan sulfate

    PubMed Central

    Migliorini, Elisa; Thakar, Dhruv; Khnle, Jens; Sadir, Rabia; Dyer, Douglas P.; Li, Yong; Sun, Changye; Volkman, Brian F.; Handel, Tracy M.; Coche-Guerente, Liliane; Fernig, David G.; Lortat-Jacob, Hugues; Richter, Ralf P.

    2015-01-01

    The glycosaminoglycan heparan sulfate (HS), present at the surface of most cells and ubiquitous in extracellular matrix, binds many soluble extracellular signalling molecules such as chemokines and growth factors, and regulates their transport and effector functions. It is, however, unknown whether upon binding HS these proteins can affect the long-range structure of HS. To test this idea, we interrogated a supramolecular model system, in which HS chains grafted to streptavidin-functionalized oligoethylene glycol monolayers or supported lipid bilayers mimic the HS-rich pericellular or extracellular matrix, with the biophysical techniques quartz crystal microbalance (QCM-D) and fluorescence recovery after photobleaching (FRAP). We were able to control and characterize the supramolecular presentation of HS chainstheir local density, orientation, conformation and lateral mobilityand their interaction with proteins. The chemokine CXCL12? (or SDF-1?) rigidified the HS film, and this effect was due to protein-mediated cross-linking of HS chains. Complementary measurements with CXCL12? mutants and the CXCL12? isoform provided insight into the molecular mechanism underlying cross-linking. Fibroblast growth factor 2 (FGF-2), which has three HS binding sites, was also found to cross-link HS, but FGF-9, which has just one binding site, did not. Based on these data, we propose that the ability to cross-link HS is a generic feature of many cytokines and growth factors, which depends on the architecture of their HS binding sites. The ability to change matrix organization and physico-chemical properties (e.g. permeability and rigidification) implies that the functions of cytokines and growth factors may not simply be confined to the activation of cognate cellular receptors. PMID:26269427

  14. Reversible PH Lability of Cross-Linked Vault Nanocapsules

    SciTech Connect

    Yu, M.; Ng, B.C.; Rome, L.H.; Tolbert, S.H.; Monbouquette, H.G.

    2009-05-28

    Vaults are ubiquitous, self-assembled protein nanocapsules with dimension in the sub-100 nm range that are conserved across diverse phyla from worms to humans. Their normal presence in humans at a copy number of over 10 000/cell makes them attractive as potential drug delivery vehicles. Toward this goal, bifunctional amine-reactive reagents are shown to be useful for the reversible cross-linking of recombinant vaults such that they may be closed and opened in a controllable manner.

  15. Utility of formaldehyde cross-linking and mass spectrometry in the study of protein-protein interactions.

    PubMed

    Sutherland, Brent W; Toews, Judy; Kast, Juergen

    2008-06-01

    For decades, formaldehyde has been routinely used to cross-link proteins in cells, tissue, and in some instances, even entire organisms. Due to its small size, formaldehyde can readily permeate cell walls and membranes, resulting in efficient cross-linking, i.e. the formation of covalent bonds between proteins, DNA, and other reactive molecules. Indeed, formaldehyde cross-linking is an instrumental component of many mainstream analytical/cell biology techniques including chromatin immunoprecipitation (ChIP) of protein-DNA complexes found in nuclei; immunohistological analysis of protein expression and localization within cells, tissues, and organs; and mass spectrometry (MS)-compatible silver-staining methodologies used to visualize low abundance proteins in polyacrylamide gels. However, despite its exquisite suitability for use in the analysis of protein environments within cells, formaldehyde has yet to be commonly employed in the directed analysis of protein-protein interactions and cellular networks. The general purpose of this article is to discuss recent advancements in the use of formaldehyde cross-linking in combination with MS-based methodologies. Key advantages and limitations to the use of formaldehyde over other cross-linkers and technologies currently used to study protein-protein interactions are highlighted, and formaldehyde-based experimental approaches that are proving very promising in their ability to accurately and efficiently identify novel protein-protein and multiprotein interaction complexes are presented. PMID:18438963

  16. Microscopic examination in vivo and in vitro of natural and cross-linked polyunsaturated mclPHA.

    PubMed

    Bassas, M; Diaz, J; Rodriguez, E; Espuny, M J; Prieto, M J; Manresa, A

    2008-03-01

    Pseudomonas aeruginosa 42A2 produces a polyunsaturated polyhydroxyalkanoates (PHA-L) when grown on linseed oil as a substrate. Its high unsaturation content (36.5%) provides highly reactive PHA-L, generating a cross-linked biopolymer after ultraviolet (UV) irradiation. Both PHAs (PHA-L and uvPHA-L) were characterized by nuclear magnetic resonance, Fourier transform infrared spectroscopy, gel permeation chromatography, gas chromatography-mass spectrometry and differential scanning calorimetry-thermogravimetric analysis. The structural analysis of the new polymer revealed a dramatic decrease in unsaturated monomer content (8.5%), due to the complete disappearance of the polyunsaturated monomers (C(12:2), C(14:2), and C(14:3)). The cross-linking reaction was also confirmed by atomic force microscopy (AFM) and transmission electron microscopy. AFM showed morphological changes in bacteria cells with and without PHA granules. The microscope techniques provided us with micrographs of the native and cross-linked polymers, showing the formation of a reticular structure as the consequence of the cross-linking reaction. PMID:18224316

  17. 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.

  18. 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.

  19. Magnetic macromolecular cross linked enzyme aggregates (CLEAs) of glucoamylase.

    PubMed

    Nadar, Shamraja S; Rathod, Virendra K

    2016-02-01

    This work illustrates the preparation of magnetic macromolecular glucoamylase CLEAs using dialdehydic pectin, as a cross linker instead of traditional glutaraldehyde. The effect of precipitators type and amount, cross linker concentration, cross linking time and amount of amino functionalized magnetic nanoparticles (AFMNs) on glucoamylase activity was studied. Glucoamylase magnetic macromolecular CLEAs prepared by precipitation in presence of AFMNs by ammonium sulfate were subsequently cross linked by dialdehydic pectin. After cross-linked by pectin, 95.4% activity recovery was achieved in magnetic macromolecular CLEAs, whereas in case of glutaraldehyde cross linker, 85.3% activity recovery was achieved. Magnetic macromolecular CLEAs showed 2.91 and 1.27 folds higher thermal stability as compared to free and magnetic glutaraldehyde CLEAs. In kinetics study, magnetic macromolecular CLEAs retained same Km values, whereas magnetic glutaraldehyde CLEAs showed higher Km value than free enzyme. The porous structure of magnetic macromolecular CLEAs was not only enhanced mass transfer toward macromolecular substrates, but also showed compression resistance for 5 consecutive cycles which was checked in terms of effectiveness factor. At the end, in reusability study; magnetic macromolecular CLEAs were retained 84% activity after 10(th) cycle without leaching of enzyme which is 22% higher than traditional magnetic CLEAs. PMID:26777253

  20. Homogeneous UVA system for corneal cross-linking treatment

    NASA Astrophysics Data System (ADS)

    Ayres Pereira, Fernando R.; Stefani, Mario A.; Otoboni, Jos A.; Richter, Eduardo H.; Ventura, Liliane

    2010-02-01

    The treatment of keratoconus and corneal ulcers by collagen cross-linking using ultraviolet type A irradiation, combined with photo-sensitizer Riboflavin (vitamin B2), is a promising technique. The standard protocol suggests instilling Riboflavin in the pre-scratched cornea every 5min for 30min, during the UVA irradiation of the cornea at 3mW/cm2 for 30 min. This process leads to an increase of the biomechanical strength of the cornea, stopping the progression, or sometimes, even reversing Keratoconus. The collagen cross-linking can be achieved by many methods, but the utilization of UVA light, for this purpose, is ideal because of its possibility of a homogeneous treatment leading to an equal result along the treated area. We have developed a system, to be clinically used for treatment of unhealthy corneas using the cross-linking technique, which consists of an UVA emitting delivery device controlled by a closed loop system with high homogeneity. The system is tunable and delivers 3-5 mW/cm2, at 365nm, for three spots (6mm, 8mm and 10mm in diameter). The electronics close loop presents 1% of precision, leading to an overall error, after the calibration, of less than 10% and approximately 96% of homogeneity.

  1. Analysis of growth hormone and lactogenic binding sites cross-linked to iodinated human growth hormone

    SciTech Connect

    Hughes, J.P.; Simpson, J.S.; Friesen, H.G.

    1983-06-01

    GH (GHR) and lactogenic receptors were analyzed after use of the cross-linking reagent ethylene glycol bis-(succinimidyl succinate) to attach covalently iodinated human GH (hGH) to binding proteins 1) on intact IM-9 lymphocytes, 2) in a partially purified GHR preparation from rabbit liver, and 3) in crude microsomal fractions from rabbit liver, rabbit mammary gland, and rat liver. The latter two microsomal preparations contain primarily lactogenic receptors, whereas in IM-9 lymphocytes and the rabbit liver preparations, GHR predominate. Cross-linked (125I)hGH-receptor complexes were solubilized, reduced, and separated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Analysis of proteins cross-linked to (125I)hGH in the microsomal fraction from rabbit liver showed a specifically labeled complex with an estimated molecular weight (mol wt) of 75K. A slightly lower mol wt (71K) was determined for the complex labeled in the purified GHR preparation. In contrast to the relatively low mol wt complexes in rabbit liver, a complex that migrated with an apparent mol wt of 130K was identified in IM-9 lymphocytes. Labeled complexes were identified at 66K from rat liver and 61K from rabbit mammary gland. If it is assumed that hGH contributes 21K to the mol wt of the radiolabeled complexes, then the approximate mol wts of hGH-binding sites are 50-54K from rabbit liver, 109K from IM-9 lymphocytes, 45K from rat liver, and 40K from rabbit mammary gland.

  2. Cross-linked dimers with nucleating activity in actin prepared from muscle acetone powder.

    PubMed

    Selden, L A; Kinosian, H J; Estes, J E; Gershman, L C

    2000-01-11

    A covalently linked actin dimer is identified in solutions of actin prepared from an acetone powder from skeletal muscle. This actin dimer acts as an actin nucleating factor (ANF), decreasing the half-time for spontaneous actin polymerization. ANF reacts with antibodies to both the N- and C-terminal portions of actin on Western blots and migrates during reduced polyacrylamide gel electrophoresis like actin cross-linked with N, N'-p-phenylenebismaleimide. The origin of the cross-linked dimer appears to be related to the presence of carbonyl groups in purified actin. A large number of carbonyls (approximately 0.3/actin) are introduced into actin during the prolonged treatment with acetone in the preparation of the muscle acetone powder from which actin is extracted. Actin extracted from acetone powder prepared by a single acetone wash and actin prepared from bovine spleen, which is not washed with acetone, both contain fewer carbonyl groups (approximately 0.05 carbonyl/actin). ANF forms spontaneously in solutions of polymer actin containing 0.3 carbonyl/actin. We speculate that a reaction between a carbonyl on one actin polymer subunit and a lysine on a neighboring subunit is responsible for ANF formation. The presence of cross-linked actin dimers in commonly used skeletal muscle actin preparations could certainly affect studies of actin polymerization and, particularly, studies of the nucleation reaction. The physiological relevance of ANF is not clear, but given the large cellular concentration of actin, similar reactions yielding ANF could occur in vivo when increased levels of reactive oxygen species are present. PMID:10625480

  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. PMID:23886705

  4. Cross linking molecular systems to form ultrathin dielectric layers

    NASA Astrophysics Data System (ADS)

    Feng, Danqin

    Dehydrogenation leads to cross linking of polymer or polymer like formation in very different systems: self-assembled monolayers and in closo -carboranes leading to the formation of semiconducting and dielectric boron carbide. We find evidence of intermolecular interactions for a self-assembled monolayer (SAM) formed from a large molecular adsorbate, [1,1';4',1"-terphenyl]-4,4"-dimethanethiol, from the dispersion of the molecular orbitals with changing the wave vector k and from the changes with temperature. With the formation self assembled molecular (SAM) layer, the molecular orbitals hybridize to electronic bands, with indications of significant band dispersion of the unoccupied molecular orbitals. Although organic adsorbates and thin films are generally regarded as "soft" materials, the effective Debye temperature, indicative of the dynamic motion of the lattice normal to the surface, can be very high, e.g. in the multilayer film formed from [1,1'-biphenyl]-4,4'-dimethanethiol (BPDMT). Depending on molecular orientation, the effective Debye temperature can be comparable to that of graphite due to the 'stiffness' of the benzene rings, but follows the expected Debye-Waller behavior for the core level photoemission intensities with temperature. This is not always the case. We find that a monomolecular film formed from [1,1';4',1"-terphenyl]-4,4"-dimethanethiol deviates from Debye-Waller temperature behavior and is likely caused by temperature dependent changes in molecular orientation. We also find evidence for the increase in dielectric character with polymerization (cross-linking) in spite of the decrease in the HOMO-LUMO gap upon irradiation of TPDMT. The changes in the HOMO-LUMO gap, with cross-linking, are roughly consistent with the band dispersion. The decomposition and cross-linking processes are also accompanied by changes in molecular orientation. The energetics of the three isomeric carborane cage compounds [ closo-1,2-orthocarborane, closo-1,7-metacarborane, closo-1,12-paracarborane (C2B10H12)] decomposition are investigated. Thermodynamic Born-Haber cycles are constructed for neutral and ionic species in an attempt to systemically characterize closo-carborane decomposition process. The decomposition processes are in favor of lower energy decomposition processes. Among the ionic species the photon induced decomposition is dominated by BH+ and BH 2+ fragment loss, and associated with core to bound excitations. It has been observed that dehydrogenation of the closo-carboranes leads to possible cross-linking and formation of a boron rich semiconductor with good dielectric properties.

  5. 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

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

    2013-01-23

    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 (NaCNBH(3), 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 the cross-link, once formed, was stable to a variety of workup 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 LC-MS/MS were consistent with a cross-linking mechanism involving reversible reaction of the Ap aldehyde residue with the N(2)-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 (NaCNBH(3)/pH 5.2) to a stable amine linkage. Further support for the importance of the exocyclic N(2)-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 NaCNBH(3). PMID:23215239

  6. Spontaneous Cross-linking for Fabrication of Nanohybrids Embedded with Size-Controllable Particles.

    PubMed

    Kang, Danmiao; Liu, Qinglei; Chen, Min; Gu, Jiajun; Zhang, Di

    2016-01-26

    This paper reports a versatile method to fabricate robust carbon/metal hybrids with ultrasmall particle and highly developed porous structure through a scalable and facile way. Alginate is used as the precursor for it could perform cross-linking reaction with different polyvalent metal ions to form gels. After simple freeze-drying and carbonization of the alginate-derived gels, we obtained the carbon/metal hybrids with fine nanostructure. Eleven kinds of metal ions were introduced to form gels and five kinds of the gels were carbonized to produce the carbon/metal hybrids. By adjusting the reaction condition, we could tune the size of the nanoparticles in the obtained hybrids. The obtained SnO2/C hybrid shows outstanding specific capacity, rate performance, and long cycle life when it is used as the anode materials of lithium ion batteries. The ultrasmall active nanoparticles were uniformly dispersed within an interconnected pore framework. It ensured a short diffusion and transportation distance of electrolyte ions to the surfaces of active nanoparticles. In addition, the robust carbon framework comprises of quasigraphitic carbon layers. It contributed to the high rate performance by providing excellent conductive pathways for electrons within the electrodes. This work provides a general method for fabrication of carbon/metal (oxide) hybrids with fine nanostructure for application in energy storage. PMID:26650902

  7. Genipin-cross-linked poly(L-lysine)-based hydrogels: synthesis, characterization, and drug encapsulation.

    PubMed

    Wang, Steven S S; Hsieh, Ping-Lun; Chen, Pei-Shan; Chen, Yu-Tien; Jan, Jeng-Shiung

    2013-11-01

    Genipin-cross-linked hydrogels composed of biodegradable and pH-sensitive cationic poly(L-lysine) (PLL), poly(L-lysine)-block-poly(L-alanine) (PLL-b-PLAla), and poly(L-lysine)-block-polyglycine (PLL-b-PGly) polypeptides were synthesized, characterized, and used as carriers for drug delivery. These polypeptide hydrogels can respond to pH-stimulus and their gelling and mechanical properties, degradation rate, and drug release behavior can be tuned by varying polypeptide composition and cross-linking degree. Comparing with natural polymers, the synthetic polypeptides with well-defined chain length and composition can warrant the preparation of the hydrogels with tunable properties to meet the criteria for specific biomedical applications. These hydrogels composed of natural building blocks exhibited good cell compatibility and enzyme degradability and can support cell attachment/proliferation. The evaluation of these hydrogels for in vitro drug release revealed that the controlled release profile was a biphasic pattern with a mild burst release and a moderate release rate thereafter, suggesting the drug molecules were encapsulated inside the gel matrix. With the versatility of polymer chemistry and conjugation of functional moieties, it is expected these hydrogels can be useful for biomedical applications such as polymer therapeutics and tissue engineering. PMID:23872465

  8. Stable biocompatible cross-linked fluorescent polymeric nanoparticles based on AIE dye and itaconic anhydride.

    PubMed

    Li, Haiyin; Zhang, Xiqi; Zhang, Xiaoyong; Yang, Bin; Wei, Yen

    2014-09-01

    Self-assembly of polymeric materials to form nanoparticles is a particularly promising strategy for various biomedical applications, however, these self-assembling systems often encounter the critical micelle concentration (CMC) issue, as the nanoparticles is usually unstable at low concentration. Therefore, stable cross-linked fluorescent polymeric nanoparticles (FPNs) were covalently constructed from an aggregation induced emission (AIE) dye, itaconic anhydride, poly(ethylene glycol) monomethyl ether methacylate and polyethylenimine. These obtained PhE-ITA-20%(80%) FPNs were fully characterized by a series of techniques including (1)H NMR spectra, UV-vis absorption spectra, fluorescence spectra, FT-IR spectra, transmission electron microscopy, gel permeation chromatography, and dynamic light scattering. Such FPNs emitted intense fluorescence due to the introduction of aggregation induced emission dye. More importantly, the FPNs were found extremely stable in physiological solution even below the CMC owing to their cross-linked architectures. Biocompatibility evaluation and cell uptake behavior of the FPNs were further investigated to explore their potential biomedical applications, the demonstrated excellent biocompatibility made them promising for cell imaging. PMID:24973146

  9. Atomic force spectroscopy of thermoresponsive photo-cross-linked hydrogel films.

    PubMed

    Junk, Matthias J N; Berger, Rdiger; Jonas, Ulrich

    2010-05-18

    Responsive hydrogel thin films are interesting materials as responsive adhesives or as an active matrix in actuators and sensing applications, and thus, knowledge about their structural and micromechanical properties is of high relevance. Using atomic force spectroscopy, temperature-induced structural and adhesive changes of thermoresponsive hydrogel layers with micrometer thickness based on photo-cross-linked N-isopropylacrylamide (NiPAAm) were investigated in the temperature range of 18-50 degrees C. Grafted onto flat surfaces, these hydrogel layers are restricted to a highly anisotropic swelling and deswelling predominantly perpendicular to the substrate surface, which was monitored and evaluated by force spectroscopy during vertical tip approach and retraction. Analyses of the tip penetration depth yielded quantitative information about the degree of swelling. As a second feature, the critical temperature was found to decrease with increasing cross-linking density. Temperature-dependent measurements with hydrophobic and hydrophilic atomic force microscopy (AFM) tips revealed a strong adhesion to the hydrogel layer in the swollen state, which was reduced upon the layer volume collapse. These observations on the micrometer-thick gel network layers are in contrast to previous reports on ultrathin pNiPAAm brushes and monolayers, which show no adhesion in the swollen state but only in the collapsed state. Furthermore, it was found that the hydrophobicity of the hydrogel probed with a hydrophobic tip continuously increases with temperature over a broad range of at least 30 K. PMID:20163151

  10. Hybrid hydrogels cross-linked by genetically engineered coiled-coil block proteins.

    PubMed

    Wang, C; Kopecek, J; Stewart, R J

    2001-01-01

    Hybrid hydrogels of hydrophilic synthetic polymers cross-linked by protein modules undergo externally triggered volume transitions as a result of protein conformational changes. To investigate the influence of coiled-coil protein structure and stability on hydrogel volume transition, a series of block proteins containing interspersed naturally derived recombinant coiled-coils was synthesized. Proteins were characterized using circular dichroism, size exclusion chromatography, gel electrophoresis, and analytical ultracentrifugation. The block proteins formed self-associating oligomers and displayed thermal unfolding profiles indicative of a hierarchic higher-order structure. Hybrid hydrogels were assembled from an N-(2-hydroxypropyl)-methacrylamide (HPMA) copolymer and His-tagged block proteins through metal complexation. A temperature-induced decrease in hydrogel swelling was observed, and the onset temperature of the volume transition corresponded to the onset temperature of protein unfolding. We conclude that stimuli-responsive properties of hybrid hydrogels can be tailored by engineering the structure and properties of protein cross-links. PMID:11710049

  11. Subunit interactions of ribulose bisphosphate carboxylase/oxygenase as determined by chemical cross-linking and site-directed mutagenesis

    SciTech Connect

    Lee, E.H.; Soper, T.S.; Hartman, F.C.

    1987-05-01

    Ribulose bisphosphate carboxylase/oxygenase from Rhodospirillum rubrum is a homodimer of 50-kDa subunits. Although a high-resolution 3D structure of the enzyme has not yet been reported, recent work from their laboratory with 4,4'-diisothiocyano-2,2'-disulfonate stilbene demonstrated an intrasubunit distance of 12 A between the active-site lysines at positions 166 and 329. To continue mapping distances between residues in the active-site region, they have modified the enzyme with 4,4'-difluoro-3,3'-dinitrophenyl sulfone, which spans 9 A. The inactivated carboxylase exhibits an unaltered molecular weight as judged by gel filtration, thereby excluding intermolecular cross-linking. However, in the presence of urea, gel filtration reveals a prominent species of 100-kDa, attributed to intersubunit cross-linking. The major chromophoric peptide has been isolated from a tryptic digest of the 100-kDa material. Sequence analyses reveal that the intersubunit cross-link occurs between Cys-58 and active-site Lys-166. In contrast to other substitutions by site-directed mutagenesis, replacement of Lys-166 with Asp prevents association of the two subunits, presumably as a consequence of repulsive forces between the thiolate and carboxylate anions. These observations suggest that each catalytic site may consist of residues from both subunits.

  12. Autoclavable highly cross-linked polyurethane networks in ophthalmology.

    PubMed

    Bruin, P; Meeuwsen, E A; van Andel, M V; Worst, J G; Pennings, A J

    1993-11-01

    Highly cross-linked aliphatic polyurethane networks have been prepared by the bulk step reaction of low molecular weight polyols and hexamethylenediisocyanate (HDI). These polyurethane networks are optically transparent, colourless and autoclavable amorphous glassy thermosets, which are suited for use in ophthalmic applications such as intraocular lenses and keratoprostheses. The properties of these glassy polyurethanes, obtained from the reaction of the low molecular weight polyols triisopropanolamine (TIPA) or tetrakis (2-hydroxypropyl)ethylenediamine (Quadrol) and HDI in stoichiometric proportions, have been investigated in more detail. The glassy Quadrol/HDI-based polyurethane exhibits a reduction in ultimate glass transition temperature from 85 to 48 degrees C by uptake of 1% of water, and good ultimate mechanical properties (tensile strength 80-85 MPa, elongation at break ca 15%, modulus ca 1.5 GPa). IR spectra of these hydrophobic polyurethane networks revealed the absence of an isocyanate absorption, indicating that all isocyanates, apparently, had reacted during the cross-linking reaction. The biocompatibility could be increased by grafting tethered polyacrylamide chains onto the surface during network formation. These transparent cross-linked polyurethanes did not transmit UV light up to 400 nm, by incorporation of a small amount of the UV absorbing chromophore Coumarin 102, and could be sterilized simply by autoclaving. They were implanted in rabbit eyes, either in the form of small circular disks or in the form of a keratoprosthesis (artificial cornea). It was shown that the material was well tolerated by the rabbit eyes. Serious opacification of the cornea, a direct result of an adverse reaction to the implant, was never seen. Even 1 yr after implantation of a polyurethane keratoprosthesis the eye was still 'quiet'. PMID:7508760

  13. Stability of casein micelles cross-linked by transglutaminase.

    PubMed

    Smiddy, M A; Martin, J-E G H; Kelly, A L; de Kruif, C G; Huppertz, T

    2006-06-01

    In this study, caseins micelles were internally cross-linked using the enzyme transglutaminase (TGase). The integrity of the micelles was examined on solubilization of micellar calcium phosphate (MCP) or on disruption of hydrophobic interactions and breakage of hydrogen bonds. The level of monomeric caseins, determined electrophoretically, decreased with increasing time of incubation with TGase at 30 degrees C; after incubation for 24 h, no monomeric beta- or kappa-caseins were detected, whereas only a small level of monomeric alphaS1-casein remained, suggesting near complete intramicellar cross-linking. The ability of casein micelles to maintain structural integrity on disruption of hydrophobic interactions (using urea, sodium dodecyl sulfate, or heating in the presence of ethanol), solubilization of MCP (using the calcium-chelating agent trisodium citrate) or high-pressure treatment was estimated by measurement of the L*-value of milk; i.e., the amount of back-scattered light. The amount of light scattered by casein micelles in noncross-linked milk was reduced by >95% on complete disruption of hydrophobic interactions or complete solubilization of MCP; treatment of milk with TGase increased the stability of casein micelles against disruption by all methods studied and stability increased progressively with incubation time. After 24 h of cross-linking, reductions in the extent of light scattering were still apparent in the presence of high levels of dissociating agents, possibly through citrate-induced removal of MCP nanoclusters from the micelles, or urea- or sodium dodecyl sulfate-induced increases in solvent refractive index, which reduce the extent of light-scattering. PMID:16702254

  14. LET dependence of DNA-protein cross-links

    SciTech Connect

    Blakely, E.A.; Chang, P.Y.; Bjornstad, K.A.

    1995-08-01

    We have preliminary data indicating a fluence-dependent yield of particle-induced protein cross-links (DPC`s) with a dependency on LET and particle residual energy. Our data indicate that the DPC yield for hamster fibroblasts in vitro irradiated at 32 keV/{mu}m is similar to that reported for hamster cells irradiated with cobalt-60 gamma rays. At 100-120 keV/{mu}m there is some evidence for an enhanced DPC yield with increasing particle fluence, but there are differences in the yields that are dependent on particle track structure.

  15. 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.

  16. Doubling the cross-linking interface of a rationally designed beta roll peptide for calcium-dependent proteinaceous hydrogel formation.

    PubMed

    Dooley, Kevin; Bulutoglu, Beyza; Banta, Scott

    2014-10-13

    We have rationally engineered a stimulus-responsive cross-linking domain based on a repeats-in-toxin (RTX) peptide to enable calcium-dependent formation of supramolecular hydrogel networks. The peptide isolated from the RTX domain is intrinsically disordered in the absence of calcium. In calcium rich environments, the peptide binds Ca(2+) ions and folds into a beta roll (?-roll) secondary structure composed to two parallel ?-sheet faces. Previously, we mutated one of the faces to contain solvent exposed leucine side chains which are localized only in the calcium-bound ?-roll conformation. We demonstrated the ability of this mutant peptide to self-assemble into hydrogels in the presence of calcium with the aid of additional peptide-based cross-linking moieties. Here, we have expanded this approach by engineering both ?-roll faces to contain leucine residues, thereby doubling the cross-linking interface for each monomeric building block. These leucine rich surfaces impart a hydrophobic driving force for self-assembly. Extensive characterization was performed on this double-faced mutant to ensure the retention of calcium affinity and subsequent structural rearrangement similar to that of the wild type domain. We genetically fused an ?-helical leucine zipper capable of forming tetrameric coiled-coil bundles to the peptide and the resulting chimeric protein self-assembles into a hydrogel only in calcium rich environments. Since this new mutant peptide enables cross-linking on both surfaces simultaneously, a higher oligomerization state was achieved. To further investigate the cross-linking capability, we constructed concatemers of the ?-roll with maltose binding protein (MBP), a monomeric globular protein, without the leucine zipper domains. These concatemers show a similar sol-gel transition in response to calcium. Several biophysical techniques were used to probe the structural and mechanical properties of the mutant ?-roll domain and the resulting supramolecular networks including circular dichroism, fluorescence resonance energy transfer, bis-ANS binding, and microrheology. These results demonstrate that the engineered ?-roll peptides can mediate calcium-dependent cross-linking for protein hydrogel formation without the need for any other cross-linking moieties. PMID:25226243

  17. 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. PMID:25482205

  18. Gelatin hydrogels cross-linked with bis(vinylsulfonyl)methane (BVSM): 1. The chemical networks.

    PubMed

    Hellio-Serughetti, Dominique; Djabourov, Madeleine

    2006-09-26

    This paper deals with chemical gelation of gelatin in the presence of a cross-linker, bis(vinylsulfonyl)methane (BVSM), which is able to create covalent C-N bonds with amine groups. The investigation is performed at 40 degrees C, where no triple helices are present. Gelatin is in random coil conformation. The influence of various parameters (gelatin concentration, cross-linker concentration, and pH (number of reacting sites along the gelatin chain)) was examined. Gel formation was followed by rheological and thermodynamic measurements (microcalorimetry) versus time (kinetic measurements). Furthermore, the storage moduli were compared to the number of links formed in the course of gelation. The experiments show that, within the experimental range investigated, a fully homogeneous network is not reached; the chemical gels, even upon completion of the reactions, are still in the critical domain, near the threshold. A power law behavior was put in evidence for the shear modulus versus the distance to the gel point, expressed as the concentration of links per gelatin chain. The exponent (f = 3.4 +/- 0.3) is close to that expected for the vulcanization of long chains. The storage moduli can be superposed on a single curve where the abscissa is the product of the number of C-N links per unit volume and the gelatin concentration at an exponent equal to -0.76 +/- 0.03. This exponent suggests the role of entanglements for interchain cross-linking. PMID:16981770

  19. Cross-linked comb-shaped anion exchange membranes with high base stability

    SciTech Connect

    Li, NW; Wang, LZ; Hickner, M

    2014-01-01

    A unique one-step cross-linking strategy that connects quaternary ammonium centers using Grubbs II-catalyzed olefin metathesis was developed. The cross-linked anion exchange membranes showed swelling ratios of less than 10% and hydroxide conductivities of 18 to 40 mS cm(- 1). Cross-linking improved the membranes' stability to hydroxide degradation compared to their non-cross-linked analogues.

  20. Status of hyaluronan supplementation therapy in osteoarthritis.

    PubMed

    Altman, Roy D

    2003-02-01

    Hyaluronans are polysaccharide molecules that occur naturally in synovial fluid; they help to create a viscous environment, cushion joints, and maintain normal function. The American College of Rheumatology recommends intra-articular injection of hyaluronans, which are available as several distinct therapeutic products, as an alternative to oral analgesics or nonsteroidal anti-inflammatory drugs for the symptomatic treatment of pain associated with osteoarthritis of the knee. A large body of literature supports the clinical efficacy and safety of this therapeutic class for this indication, although there are differences between the marketed products and they should be evaluated independently. Preliminary work investigating the use of hyaluronans for osteoarthritis in joints other than the knee has also produced promising results. There is growing evidence that hyaluronans, the biology of which is complex, may also have structure-modifying activity. Thus, compared with currently approved nonoperative therapies for osteoarthritis, hyaluronans may also have beneficial effects on the disease process in osteoarthritis. PMID:12590879

  1. Matrix cross-linkingmediated mechanotransduction promotes posttraumatic osteoarthritis

    PubMed Central

    Kim, Jin-Hong; Lee, Gyuseok; Won, Yoonkyung; Lee, Minju; Kwak, Ji-Sun; Chun, Churl-Hong; Chun, Jang-Soo

    2015-01-01

    Osteoarthritis (OA) is characterized by impairment of the load-bearing function of articular cartilage. OA cartilage matrix undergoes extensive biophysical remodeling characterized by decreased compliance. In this study, we elucidate the mechanistic origin of matrix remodeling and the downstream mechanotransduction pathway and further demonstrate an active role of this mechanism in OA pathogenesis. Aging and mechanical stress, the two major risk factors of OA, promote cartilage matrix stiffening through the accumulation of advanced glycation end-products and up-regulation of the collagen cross-linking enzyme lysyl oxidase, respectively. Increasing matrix stiffness substantially disrupts the homeostatic balance between chondrocyte catabolism and anabolism via the RhoRho kinasemyosin light chain axis, consequently eliciting OA pathogenesis in mice. Experimental enhancement of nonenzymatic or enzymatic matrix cross-linking augments surgically induced OA pathogenesis in mice, and suppressing these events effectively inhibits OA with concomitant modulation of matrix degrading enzymes. Based on these findings, we propose a central role of matrix-mediated mechanotransduction in OA pathogenesis. PMID:26170306

  2. Optimization model for UV-Riboflavin corneal cross-linking

    NASA Astrophysics Data System (ADS)

    Schumacher, S.; Wernli, J.; Scherrer, S.; Bueehler, M.; Seiler, T.; Mrochen, M.

    2011-03-01

    Nowadays UV-cross-linking is an established method for the treatment of keraectasia. Currently a standardized protocol is used for the cross-linking treatment. We will now present a theoretical model which predicts the number of induced crosslinks in the corneal tissue, in dependence of the Riboflavin concentration, the radiation intensity, the pre-treatment time and the treatment time. The model is developed by merging the difussion equation, the equation for the light distribution in dependence on the absorbers in the tissue and a rate equation for the polymerization process. A higher concentration of Riboflavin solution as well as a higher irradiation intensity will increase the number of induced crosslinks. However, performed stress-strain experiments which support the model showed that higher Riboflavin concentrations (> 0.125%) do not result in a further increase in stability of the corneal tissue. This is caused by the inhomogeneous distribution of induced crosslinks throughout the cornea due to the uneven absorption of the UV-light. The new model offers the possibility to optimize the treatment individually for every patient depending on their corneal thickness in terms of efficiency, saftey and treatment time.

  3. Damage and fatigue in cross-linked rubbers

    NASA Astrophysics Data System (ADS)

    Melnikov, Alexei

    Damage and fatigue of elastomers have not been fundamentally understood because of the complex nature of these materials. All currently existing models are completely phenomenological. Therefore two problems have been investigated in this research to address those fundamental issues. The first problem was creating an innovative concept with a mathematical modeling, which would be able to describe the damage using molecular characteristics of elastomers. The second problem is developing new approaches to study fatigue, and especially impact fatigue of elastomers. The following results have been obtained in this research. A theoretical model of damage has been developed which involves the basic molecular characteristics of cross-linked elastomers and takes into account the effects of viscoelasticity and stress-induced crystallization. This model was found very reliable and successful in description of numerous quasi-static simple extension experiments for monotonous and repeating loadings. It also roughly predicts in molecular terms the failure of elastomers with various degrees of cross-linking. Quasi-impact fatigue tests with different geometry of an indenter have also been performed. Some microscopic features of rubber damage have been investigated using optical microscopy and SEM. In particular, the accumulation of a completely de-vulcanized, liquid-like substance was observed under intense, multi-cycle impacts. All the findings discovered in quasi-impact experiments are consistent with the damage model predictions.

  4. Matrix cross-linking-mediated mechanotransduction promotes posttraumatic osteoarthritis.

    PubMed

    Kim, Jin-Hong; Lee, Gyuseok; Won, Yoonkyung; Lee, Minju; Kwak, Ji-Sun; Chun, Churl-Hong; Chun, Jang-Soo

    2015-07-28

    Osteoarthritis (OA) is characterized by impairment of the load-bearing function of articular cartilage. OA cartilage matrix undergoes extensive biophysical remodeling characterized by decreased compliance. In this study, we elucidate the mechanistic origin of matrix remodeling and the downstream mechanotransduction pathway and further demonstrate an active role of this mechanism in OA pathogenesis. Aging and mechanical stress, the two major risk factors of OA, promote cartilage matrix stiffening through the accumulation of advanced glycation end-products and up-regulation of the collagen cross-linking enzyme lysyl oxidase, respectively. Increasing matrix stiffness substantially disrupts the homeostatic balance between chondrocyte catabolism and anabolism via the Rho-Rho kinase-myosin light chain axis, consequently eliciting OA pathogenesis in mice. Experimental enhancement of nonenzymatic or enzymatic matrix cross-linking augments surgically induced OA pathogenesis in mice, and suppressing these events effectively inhibits OA with concomitant modulation of matrix degrading enzymes. Based on these findings, we propose a central role of matrix-mediated mechanotransduction in OA pathogenesis. PMID:26170306

  5. Collagen Cross-Linking: Current Status and Future Directions

    PubMed Central

    Hovakimyan, Marine; Guthoff, Rudolf F.; Stachs, Oliver

    2012-01-01

    Collagen cross-linking (CXL) using UVA light and riboflavin (vitamin B2) was introduced as a clinical application to stabilize the cornea by inducing cross-links within and between collagen fibers. CXL has been investigated extensively and has been shown clinically to arrest the progression of keratoconic or post-LASIK ectasia. With its minimal cost, simplicity, and proven positive clinical outcome, CXL can be regarded as a useful approach to reduce the number of penetrating keratoplasties performed. Small case series have also indicated that CXL is beneficial in corneal edema by reducing stromal swelling behavior and in keratitis by inhibiting pathogen growth. Despite these encouraging results, CXL remains a relatively new method that is potentially associated with complications. Aspects such as side effects and recurrence rates have still to be elucidated. In light of the growing interest in CXL, our paper summarizes present knowledge about this promising approach. We have intentionally endeavored to include the more relevant studies from the recent literature to provide an overview of the current status of CXL. PMID:22288005

  6. Chemical cross-linking/mass spectrometry targeting acidic residues in proteins and protein complexes

    PubMed Central

    Leitner, Alexander; Joachimiak, Lukasz A.; Unverdorben, Pia; Walzthoeni, Thomas; Frydman, Judith; Frster, Friedrich; Aebersold, Ruedi

    2014-01-01

    The study of proteins and protein complexes using chemical cross-linking followed by the MS identification of the cross-linked peptides has found increasingly widespread use in recent years. Thus far, such analyses have used almost exclusively homobifunctional, amine-reactive cross-linking reagents. Here we report the development and application of an orthogonal cross-linking chemistry specific for carboxyl groups. Chemical cross-linking of acidic residues is achieved using homobifunctional dihydrazides as cross-linking reagents and a coupling chemistry at neutral pH that is compatible with the structural integrity of most protein complexes. In addition to cross-links formed through insertion of the dihydrazides with different spacer lengths, zero-length cross-link products are also obtained, thereby providing additional structural information. We demonstrate the application of the reaction and the MS identification of the resulting cross-linked peptides for the chaperonin TRiC/CCT and the 26S proteasome. The results indicate that the targeting of acidic residues for cross-linking provides distance restraints that are complementary and orthogonal to those obtained from lysine cross-linking, thereby expanding the yield of structural information that can be obtained from cross-linking studies and used in hybrid modeling approaches. PMID:24938783

  7. Effects of processing conditions on the reliability of cross-linked polyethylene cable insulation. Progress report

    SciTech Connect

    Phillips, P.J.

    1981-03-01

    Crystallization and morphology were investigated in cross-linked PE. /sup 13/C NMR was used to quantify the cross-links. Production of cable is being studied. Dielectric constant and loss of cross-linked PE are being measured. (DLC)

  8. Exercise increases pyridinoline cross-linking and counters the mechanical effects of concurrent lathyrogenic treatment.

    PubMed

    McNerny, Erin M B; Gardinier, Joseph D; Kohn, David H

    2015-12-01

    The collagen cross-link profile of bone, associated with bone strength and fracture toughness, is tightly regulated (affecting cross-link quantity, type, lysine hydroxylation and maturity) and may contribute to the improvements in bone quality during exercise. We hypothesized that 1) exercise promotes mature cross-link formation, 2) increased mature cross-linking is accompanied by shifts in lysine hydroxylation, and 3) these changes in collagen cross-link profile have positive effects on mechanical properties. Growing male C57Bl6 mice were treated with 30 min/day of running exercise, 350 mg/kg/day ?-aminopropionitrile (BAPN) injected subcutaneously to inhibit enzymatic collagen cross-linking, or both exercise and BAPN, from 5 to 8 weeks of age. Bone collagen cross-linking profile, mechanical properties, morphology, and mineralization were measured from the tibiae. Cross-link measures, including immature, pyridinoline, pyrrole and pentosidine cross-links, ratios reflecting cross-link maturity and hydroxylation, and mineralization were tested for their importance to mechanical properties across 8 week groups through correlation analyses and step-wise linear regressions. BAPN treatment significantly reduced lysylpyridinoline, pyrrole, hydroxylysinorleucine, and total mature collagen cross-linking, resulting in decreased bone elastic modulus and increased yield strain despite a marginal increase in TMD. Exercise caused a shift toward pyridinoline cross-linking, with increased hydroxylysylpyridinoline and decreased pyrrole cross-linking resulting in total mature cross-linking and estimated tissue level mechanical properties matching sedentary control levels. Exercise superimposed on BAPN treatment increased total mature cross-linking from BAPN to control levels, but did so by increasing pyridinoline, not pyrrole, cross-links. Exercise also counteracted the BAPN effects on modulus and strain, without a change in TMD. Pyrrole cross-linking was the strongest correlate of modulus (r=0.470, p<0.01) and yield strain (r=-0.467, p<0.01). Cross-links with similar levels of telopeptide lysine hydroxylation to pyrrole (lysylpyridinoline and hydroxylysinorleucine) also correlated with modulus and strain to a lesser extent. In conclusion, exercise in growing mice promotes pyridinoline collagen cross-linking in bone, the resulting increase in total mature cross-linking is sufficient to counteract the mechanical effects of concurrent cross-link inhibition, and this responsiveness to loading is a potential means by which exercise might improve bone quality in diseased or otherwise compromised bone. PMID:26211995

  9. Comparison of Cross-linked and NonCross-linked Acellular Porcine Dermal Scaffolds for Long-term Full-Thickness Hernia Repair in a Small Animal Model

    PubMed Central

    Mestak, Ondrej; Spurkova, Zuzana; Benkova, Kamila; Vesely, Pavel; Hromadkova, Veronika; Miletin, Jakub; Juzek, Robert; Molitor, Martin; Sukop, Andrej

    2014-01-01

    Background: This study compared the strength of incorporation and biocompatibility of 2 porcine-derived grafts (cross-linked and noncross-linked) in a rat hernia model. Methods: A standardized 2 4 cm2 fascial defect was created in 30 Wistar rats and repaired with either a cross-linked or a noncross-linked graft. The rats were killed 3, 6, and 12 months later. The strength of incorporation, vascularization, cellular invasion, foreign body reaction, and capsule formation were evaluated. Results: Both graft materials showed cellular ingrowth and neovascularization by 3 months postimplantation. The average level of cellularization was significantly higher in the noncross-linked grafts than in the cross-linked grafts at 6 months (2 vs 1; P = .029). Vascularization was significantly higher in the noncross-linked grafts than in the cross-linked grafts at 6 months postimplantation (2 vs 1; P = .029) and insignificant at 3 months (2 vs 1.75; P = .311) and 12 months (1 vs 0.67; P = 1). The maximum load and breaking strength of both biomaterials increased during the study period. Overall, the strength of incorporation of the noncross-linked grafts increased from 3 months (0.75 MPa) to 12 months (3.06 MPa) postimplantation. The strength of incorporation of the cross-linked grafts also increased from 3 months (0.59 MPa) to 12 months (1.58 MPa) postimplantation. Conclusions: The results of our study suggest that noncross-linked grafts may be slightly more biocompatible and allow a more rapid and higher degree of cellular penetration and vascularization, resulting in stronger attachment to the tissues. PMID:24966996

  10. The Leaving Group Strongly Affects H2O2-Induced DNA Cross-Linking by Arylboronates

    PubMed Central

    Cao, Sheng; Wang, Yibin; Peng, Xiaohua

    2014-01-01

    We evaluated the effects of the benzylic leaving group and core structure of arylboronates on H2O2-induced formation of bisquinone methides for DNA interstrand cross-linking. The mechanism of DNA cross-linking induced by these arylboronates involves generation of phenol intermediates followed by departure of benzylic leaving group leading to QMs which directly cross-link DNA via alkylation. The QM formation is the rate-determining step for DNA cross-linking. A better leaving group (Br) and stepwise bisquinone methide formation increased interstrand cross-linking efficiency. These findings provide essential guidelines for designing novel anticancer prodrugs. PMID:24378073

  11. Effect of modified starch and nanoclay particles on biodegradability and mechanical properties of cross-linked poly lactic acid.

    PubMed

    Shayan, M; Azizi, H; Ghasemi, I; Karrabi, M

    2015-06-25

    Mechanical properties and biodegradation of cross-linked poly(lactic acid) (PLA)/maleated thermoplastic starch (MTPS)/montmorillonite (MMT) nanocomposite were studied. Crosslinking was carried out by adding di-cumyl peroxide (DCP) in the presence of triallyl isocyanurate (TAIC) as coagent. At first, MTPS was prepared by grafting maleic anhydride (MA) to thermoplastic starch in internal mixer. Experimental design was performed by using Box-Behnken method at three variables: MTPS, nanoclay and TAIC at three levels. Results showed that increasing TAIC amount substantially increased the gel fraction, enhanced tensile strength, and caused a decrease in elongation at break. Biodegradation was prevented by increasing TAIC amount in nanocomposite. Increasing MTPS amount caused a slight increase in gel fraction and decreased the tensile strength of nanocomposite. Also, MTPS could increase the elongation at break of nanocomposite and improve the biodegradation. Nanoclay had no effect on the gel fraction, but it improved tensile strength. PMID:25839817

  12. Influence of Chemical Cross-Linking on the Over Voltage Positive Temperature Coefficient of Linear Low Density Polyethylene/carbon Black/aluminum Hydroxide Nano Composites

    NASA Astrophysics Data System (ADS)

    Huang, C. Y.; Tsai, C. S.; Tsao, K. Y.

    2008-08-01

    The effect of cross-linking of blends on the stability of positive temperature coefficient (PTC) and the elimination of negative temperature coefficient (NTC) were investigated. Linear low density polyethylene (LLDPE) was chemically cross-linking with various amounts of dicumyl peroxide (DCP). The resulting of cross-linking structure of composites was analyzed by gel content, differential scanning calorimetry (DSC), and dynamic mechanical analysis (DMA). By the effect of DCP, not only a high PTC intensity appeared, but also was the NTC effect of the composite eliminated. The PTC intensity reached six orders of magnitude for the LLDPE/DCP(0.10 wt%)/CB/Al(OH)3 composite, and successfully passed the test of over-voltage resistance.

  13. Studies on N-vinylformamide cross-linked copolymers

    NASA Astrophysics Data System (ADS)

    Świder, Joanna; Tąta, Agnieszka; Sokołowska, Katarzyna; Witek, Ewa; Proniewicz, Edyta

    2015-12-01

    Copolymers of N-vinylformamide (NVF) cross-linked with three multifunctional monomers, including divinylbenzene (DVB), ethylene glycol dimethacrylate (EGDMA), and N,N‧-methylenebisacrylamide (MBA) were synthetized by a three-dimensional free radical polymerization in inverse suspension using 2,2‧-azobis(2-methylpropionamide) dihydrochloride (AIBA) as an initiator. Methyl silicon oil was used as the continuous phase during the polymerization processes. Fourier-transform adsorption infrared (FT-IR) spectra revealed the presence of silicone oil traces and suggested that silicone oil strongly interacted with the copolymers surface. Purification procedure allowed to completely remove the silicon oil traces from P(NVF-co-DVB) only. The morphology and the structure of the investigated copolymers were examined by optical microscopy, FT-IR, and FT-Raman (Fourier-transform Raman spectroscopy) methods.

  14. Encapsulation of cobalt nanoparticles in cross-linked-polymer cages

    NASA Astrophysics Data System (ADS)

    Hatamie, Shadie; Dhole, S. D.; Ding, J.; Kale, S. N.

    2009-07-01

    Nanoparticles embedded in polymeric cages give rise to interesting applications ranging from nanocatalysis to drug-delivery systems. In this context, we report on synthesis of cobalt (Co) nanoparticles trapped in polyvinyl alcohol (PVA) matrix to yield self-supporting magnetic films in PVA slime. A 20 nm, Co formed in FCC geometry encapsulated with a weak citrate coat when caged in PVA matrix exhibited persistence of magnetism and good radio-frequency response. Cross-linking of PVA chains to form cage-like structures to arrest Co nanoparticles therein, is believed to be the reason for oxide-free nature of Co, promising applications in biomedicine as well as in radio-frequency shielding.

  15. DNA interstrand cross-link repair requires replication fork convergence

    PubMed Central

    Zhang, Jieqiong; Dewar, James M.; Budzowska, Magda; Motnenko, Anna; Cohn, Martin A.; Walter, Johannes C.

    2014-01-01

    DNA interstrand cross-links (ICLs) prevent strand separation during DNA replication and transcription and are therefore extremely cytotoxic. In metazoans, a major pathway of ICL repair is coupled to DNA replication and requires the Fanconi anemia pathway. In most current models, collision of a single DNA replication fork with an ICL is sufficient to initiate repair. In contrast, we show here that in Xenopus egg extracts, two DNA replication forks must converge on an ICL to trigger repair. When only one fork reaches the ICL, the replicative CMG helicase fails to unload from the stalled fork, and repair is blocked. Arrival of a second fork, even when substantially delayed, rescues repair. We conclude that ICL repair requires a replication-induced X-shaped DNA structure surrounding the lesion, and we speculate how this requirement helps maintain genomic stability in S phase. PMID:25643322

  16. Dynamic Role of Cross-Linking Proteins in Actin Rheology

    PubMed Central

    Kim, Taeyoon; Hwang, Wonmuk; Kamm, RogerD.

    2011-01-01

    We develop a computational model to compare the relative importance of unbinding and unfolding of actin cross-linking proteins (ACPs) in the dynamic properties of the actin cytoskeleton. We show that in the strain-stiffening regime with typical physiological and experimental strain rates, unbinding events are predominant with negligible unfolding. ACPs unbound by greater forces experience larger displacements, with a tendency to rebind to different filaments. At constant strain, stress relaxes to physiological levels by unbinding onlynot unfoldingof ACPs, which is consistent with experiments. Also, rebinding of ACPs dampens full relaxation of stress. When the network is allowed to return to a stress-free state after shear deformation, plastic deformation is observed only with unbinding. These results suggest that despite the possibility of unfolding, unbinding of ACPs is the major determinant for the rheology of the actin network. PMID:21961585

  17. Thermal fluctuations in chemically cross-linked polymers of cyclodextrins.

    PubMed

    Rossi, Barbara; Venuti, Valentina; Paciaroni, Alessandro; Mele, Andrea; Longeville, Stéphane; Natali, Francesca; Crupi, Vincenza; Majolino, Domenico; Trotta, Francesco

    2015-03-21

    The extent and nature of thermal fluctuations in the innovative class of cross-linked polymers called cyclodextrin nanosponges (CDNS) are investigated, on the picosecond time scale, through elastic and quasielastic neutron scattering experiments. Nanosponges are complex 3D polymer networks where covalent bonds connecting different cyclodextrin (CD) units and intra- and inter-molecular hydrogen-bond interactions cooperate to define the molecular architecture and fast dynamics of the polymer. The study presented here aims to clarify the nature of the conformational rearrangements activated by increasing temperature in the nanosponge polymer, and the constraints imposed by intra- and inter-molecular hydrogen-bond patterns on the internal dynamics of the macromolecule. The results suggest a picture, in which conformational rearrangements involving the torsion of the OH groups around the C-O bonds dominate the internal dynamics of the polymer over the picosecond time scale. Moreover, the estimated values of mean square displacements reveal that the motions of the hydrogen atoms in the nanosponges are progressively hampered as the cross-linking degree of the polymer is increased. Finally, the study of the molecular relaxations suggests a dynamical rearrangement of the hydrogen-bond networks, which is characterized by a jump diffusion motion of the more mobile hydrogen atoms belonging to the OH groups of the CD units. All these findings add further contribution to the rational comprehensive view of the dynamics of these macromolecules, which may be particularly beneficial in designing new drug-delivery systems with tuneable inclusion/release properties. PMID:25639345

  18. Cross-linked polyethylenimine-tripolyphosphate nanoparticles for gene delivery.

    PubMed

    Huang, Xianzhang; Shen, Sujing; Zhang, Zhanfeng; Zhuang, Junhua

    2014-01-01

    The high transfection efficiency of polyethylenimine (PEI) makes it an attractive potential nonviral genetic vector for gene delivery and therapy. However, the highly positive charge of PEI leads to cytotoxicity and limits its application. To reduce the cytotoxicity of PEI, we prepared anion-enriched nanoparticles that combined PEI with tripolyphosphate (TPP). We then characterized the PEI-TPP nanoparticles in terms of size, zeta potential, and Fourier-transform infrared (FTIR) spectra, and assessed their transfection efficiency, cytotoxicity, and ability to resist deoxyribonuclease (DNase) I digestion. The cellular uptake of PEI-TPP with phosphorylated internal ribosome entry site-enhanced green fluorescent protein C1 or FAM (fluorouracil, Adriamycin [doxorubicin] and mitomycin)-labeled small interfering ribonucleic acids (siRNAs) was monitored by fluorescence microscopy and confocal laser microscopy. The efficiency of transfected delivery of plasmid deoxyribonucleic acid (DNA) and siRNA in vitro was 1.11- to 4.20-fold higher with the PEI-TPP particles (7.6% cross-linked) than with the PEI, at all N:P ratios (nitrogen in PEI to phosphorus in DNA) tested. The cell viability of different cell lines was more than 90% at the chosen N:P ratios of PEI-TPP/DNA complexes. Moreover, PEI-TPP nanoparticles resisted digestion by DNase I for more than 2 hours. The time-dependent absorption experiment showed that 7.6% of cross-linked PEI-TPP particles were internalized by 293T cells within 1 hour. In summary, PEI-TPP nanoparticles effectively transfected cells while conferring little or no toxicity, and thus have potential application in gene delivery. PMID:25342902

  19. The wear of cross-linked polyethylene against itself.

    PubMed

    Joyce, T J; Ash, H E; Unsworth, A

    1996-01-01

    Cross-linked polyethylene (XLPE) may have an application as a material for an all-plastic surface replacement finger joint. It is inexpensive, biocompatible and can be injection-moulded into the complex shapes that are found on the ends of the finger bones. Further, the cross-linking of polyethylene has significantly improved its mechanical properties. Therefore, the opportunity exists for an all-XLPE joint, and so the wear characteristics of XLPE sliding against itself have been investigated. Wear tests were carried out on both reciprocating pin-on-plate machines and a finger function simulator. The reciprocating pin-on-plate machines had pins loaded at 10 N and 40 N. All pin-on-plate tests show wear factors from the plates very much greater than those of the pins. After 349 km of sliding, a mean wear factor of 0.46 x 10(-6) mm3/N m was found for the plates compared with 0.021 x 10(-6) mm3/N m for the pins. A fatigue mechanism may be causing this phenomenon of greater plate wear. Tests using the finger function simulator give an average wear rate of 0.22 x 10(-6) mm3/N m after 368 km. This sliding distance is equivalent to 12.5 years of use in vivo. The wear factors found were comparable with those of ultra-high molecular weight polyethylene (UHMWPE) against a metallic counterface and, therefore, as the loads across the finger joint are much less than those across the knee or the hip, it is probable that an all-XLPE finger joint will be viable from a wear point of view. PMID:8663888

  20. Measuring the Formaldehyde ProteinDNA Cross-Link Reversal Rate

    PubMed Central

    2015-01-01

    ProteinDNA binding interactions play critical roles in important cellular processes such as gene expression, cell division, and chromosomal organization. Techniques to identify and characterize these interactions often utilize formaldehyde cross-linking for stabilization of the complexes. Advantages of formaldehyde as a cross-linking reagent include cell permeability, relatively fast cross-linking kinetics, and short cross-linker length. In addition, formaldehyde cross-links are reversible, which has the advantage of allowing complexes to be dissociated if desired but may also present a problem if undesired dissociation occurs in the course of an experiment. While the kinetics of formaldehyde cross-link formation have been well-established in numerous studies, there have been no reports of the rate of cross-link dissociation, even though it is clearly a critical variable when developing a biochemical protocol involving formaldehyde cross-linking. We present here a method for measurement of the rate of formaldehyde cross-link reversal based upon the Formaldehyde-Assisted Isolation of Regulatory Elements (FAIRE) procedure and use it to determine the rate of cross-link reversal for cross-linked proteinDNA complexes from yeast cell lysate. The half-life of the proteinDNA cross-links varies from 179 h at 4 C to 11.3 h at 47 C, with a rate that increases exponentially with temperature and is independent of salt concentration. PMID:24848408

  1. 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. PMID:21560767

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

    PubMed

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

    2013-11-26

    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 (M w 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:25411511

  3. 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:25411511

  4. Anisotropic Thermal Conductivity Measurements on Cross-Linked Polybutadienes in Uniaxial Elongation

    NASA Astrophysics Data System (ADS)

    Venerus, David C.; Kolev, Dimitre

    2008-07-01

    Cross-linked elastomers have numerous applications including automobiles, sporting goods, and biomedical devices. During both their processing and application, these materials experience large mechanical stresses and thermal gradients. In this study, we investigate the mechanical and thermal transport behavior of cross-linked polybutadienes. These materials have been prepared by cross linking well-entangled polybutadienes using an organic peroxide cross-linking agent at low concentration. Samples obtained after nearly complete conversion of the cross-linking agent, which can be characterized as lightly cross-linked (i.e., more than 10 entanglements per cross-link), were subjected to a series of large strain, uniaxial deformations. Measurements of the tensile stress and two components of the thermal conductivity tensor will be reported as a function of elongation. These data are also used to examine the stress-thermal rule in which the stress and thermal conductivity tensors are linearly related.

  5. Effect of cross-linking on physicochemical properties of tapioca starch and its application in soup product.

    PubMed

    Wongsagonsup, Rungtiwa; Pujchakarn, Thamonwan; Jitrakbumrung, Suparat; Chaiwat, Weerawut; Fuongfuchat, Asira; Varavinit, Saiyavit; Dangtip, Somsak; Suphantharika, Manop

    2014-01-30

    Physicochemical properties of cross-linked tapioca starch (CLTS) with different cross-linking levels and their application as a thickening agent in soups were studied. The CLTS was prepared by cross-linking native tapioca starch suspended in alkaline solution (41.67% (w/w), pH 11) using a mixture (99:1 (w/w) ratio) of sodium trimetaphosphate (STMP) and sodium tripolyphosphate (STPP) at different concentrations ranged from 0.25% to 6.0% (w/w of starch) at 45C for 3h. Starch paste clarity decreased with increasing concentration of STMP/STPP mixture. Variations of swelling power, solubility, pasting, gelatinization, and rheological properties of the CLTS were found. Thermogravimetric analysis exhibited higher thermal stability for the CLTS granules compared to the native one. Among the samples, the CLTS prepared using 1.0% STMP/STPP (1.0%-CLTS) and soup containing the 1.0%-CLTS exhibited the strongest gel characteristic and the greatest shear resistant properties. The 1.0%-CLTS improved the textural properties and sensory quality of soups. PMID:24299823

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

    SciTech Connect

    Nishida, Yoshihiro . E-mail: ynishida@med.nagoya-u.ac.jp; Knudson, Warren; Knudson, Cheryl B.; Ishiguro, Naoki

    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 in 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.

  7. 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.

  8. 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.

  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 HAp-Gemosil ceramics. PMID:23139457

  10. Synthesis of poly(N-isopropylacrylamide) hydrogels by radiation polymerization and cross-linking

    SciTech Connect

    Nagaoka, Noriyasu; Kubota, Hitoshi; Katakai, Ryoichi; Safranj, Agneza; Yoshida, Masaru; Omichi, Hideki

    1993-12-20

    Poly(N-isopropylacrylamide) [poly(NIPAAm)] shows a typical thermal reversibility of phase transition in aqueous solutions. That is, it precipitates from solution above a critical temperature called the lower critical solution temperature (LCST) and dissolves below this temperature. When it is cross-linked, the obtained hydrogel collapses above LCST, while it swells and expands below LCST. This hydrogel has received much attention recently and has been used as a model system to demonstrate the validity of theories describing the coil-globule transition, swelling of networks, and folding and unfolding of biopolymers. It has also been proposed for various applications ranging from controlled drug delivery to solute separation. Poly(NIPAAm) hydrogel is usually synthesized at room temperature from an aqueous solution of the monomer by using a redox initiator composed of ammonium persulfate and N,N,N{prime},N{prime}-tetramethylethylenediamine in the presence of N,N{prime}-methylenebisacrylamide as a cross-linker. Since the LCST of poly(NIPAAm) is around 32 C, the polymerization at room temperature proceeds in a homogeneous solution. Recently, poly(NIPAAm) hydrogels were synthesized by starting the polymerization below the LCST and then elevating the temperature above it, by which method macroporous gels with fast temperature response were obtained. The idea is to apply a radiation--induced polymerization method for the synthesis of poly(NIPAAm) hydrogels. This method offers unique advantages for synthesis: it is a simple and additive-free process at all temperatures, and the degree of cross-linking can be easily controlled by irradiation conditions. Therefore, radiation methods are especially attractive for the synthesis of hydrogels with potential biomedical application where the residual chemical initiators may contaminate the product. It is possible to combine into one step the synthesis and sterilization of the product, and it is economically competitive.

  11. 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.

  12. Physicochemical properties and bioactivity of nisin-containing cross-linked hydroxypropylmethylcellulose films.

    PubMed

    Sebti, Issam; Delves-Broughton, John; Coma, Vronique

    2003-10-22

    Cross-linked hydroxypropylmethylcellulose (HPMC) cast films with citric acid as polycarboxylic cross-linker were elaborated to study the effect of cross-linking level on various properties. Increased amounts of cross-linking agent were not connected to statistically different tensile strength and Young's modulus. Whatever the cross-linking level of the film was, the ultimate elongation parameter decreased by approximately 60% compared to the HMPC control film. Moisture sorption isotherms and water contact angle meter showed that the effect of cross-linking degree tends to reduce the hygroscopic and hydrophilic characteristics of films. In addition, to control bacteria growth on food surfaces, the antimicrobial activity of both 98% cross-linked HPMC-nisin and control HPMC-nisin films was tested on Micrococcus luteus. Despite the incorporation of a significant content of nisin, cross-linked HPMC-nisin films were completely inactive on the microbial strain compared to the HPMC-nisin control films. Cross-linking conditions likely either denatured the nisin or irreversibly bound nisin to the cross-linked HPMC. However, nisin adsorbed into films made from previously cross-linked HPMC maintained its activity. PMID:14558764

  13. Secondary cytotoxicity of cross-linked dermal sheep collagens during repeated exposure to human fibroblasts.

    PubMed

    van Luyn, M J; van Wachem, P B; Olde Damink, L H; Dijkstra, P J; Feijen, J; Nieuwenhuis, P

    1992-01-01

    We investigated commercially available dermal sheep collagen either cross-linked with hexamethylenediisocyanate, or cross-linked with glutaraldehyde. In previous in vitro studies we could discriminate primary, i.e. extractable, and secondary cytotoxicity, due to cell-biomaterial interactions, i.e. enzymatic actions. To develop dermal sheep collagen for clinical applications, we focused in this study on the release, e.g. elimination, of secondary cytotoxicity over time. We used the universal 7 d methylcellulose cell culture with human skin fibroblasts as a test system. Hexamethylenediisocyanate-cross-linked dermal sheep collagen and glutaraldehyde-cross-linked dermal sheep collagen were tested, with intervals of 6 d, over a culture period of 42 d. With hexamethylenediisocyanate-cross-linked dermal sheep collagen, cytotoxicity, i.e. cell growth inhibition and deviant cell morphology, was eliminated after 18 d of exposure. When testing glutaraldehyde-cross-linked dermal sheep collagen, the bulk of cytotoxic products was released after 6 d, but a continuous low secondary cytotoxicity was measured up to 42 d. As a control, non-cross-linked dermal-sheep collagen was tested over a period of 36 d, but no secondary cytotoxic effects were observed. The differences in release of secondary cytotoxicity between hexamethylenediisocyanate-cross-linked dermal sheep collagen, glutaraldehyde-cross-linked dermal sheep collagen and non-cross-linked dermal sheep collagen are explained from differences in cross-linking agents and cross-links obtained. We hypothesize that secondary cytotoxicity results from enzymatic release of pendant molecules from hexamethylene-diisocyanate-cross-linked dermal sheep collagen, e.g. formed after reaction of hydrolysis products of hexamethylenediisocyanate with dermal sheep collagen.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1472588

  14. Characterization of Interstrand DNA-DNA Cross-Links Using the ?-Hemolysin Protein Nanopore.

    PubMed

    Zhang, Xinyue; Price, Nathan E; Fang, Xi; Yang, Zhiyu; Gu, Li-Qun; Gates, Kent S

    2015-12-22

    Nanopore-based sensors have been studied extensively as potential tools for DNA sequencing, characterization of epigenetic modifications such as 5-methylcytosine, and detection of microRNA biomarkers. In the studies described here, the ?-hemolysin protein nanopore embedded in a lipid bilayer was used for the detection and characterization of interstrand cross-links in duplex DNA. Interstrand cross-links are important lesions in medicinal chemistry and toxicology because they prevent the strand separation that is required for read-out of genetic information from DNA in cells. In addition, interstrand cross-links are used for the stabilization of duplex DNA in structural biology and materials science. Cross-linked DNA fragments produced unmistakable current signatures in the nanopore experiment. Some cross-linked substrates gave irreversible current blocks of >10 min, while others produced long current blocks (10-100 s) before the double-stranded DNA cross-link translocated through the ?-hemolysin channel in a voltage-driven manner. The duration of the current block for the different cross-linked substrates examined here may be dictated by the stability of the duplex region left in the vestibule of the nanopore following partial unzipping of the cross-linked DNA. Construction of calibration curves measuring the frequency of cross-link blocking events (1/?on) as a function of cross-link concentration enabled quantitative determination of the amounts of cross-linked DNA present in samples. The unique current signatures generated by cross-linked DNA in the ?-HL nanopore may enable the detection and characterization of DNA cross-links that are important in toxicology, medicine, and materials science. PMID:26563913

  15. A role for hyaluronan in joint development.

    PubMed Central

    Craig, F M; Bayliss, M T; Bentley, G; Archer, C W

    1990-01-01

    Hyaluronan, a common connective tissue component, influences cell adhesion, migration and cytodifferentiation in vitro, and because of these properties has been postulated to have a role in morphogenesis. Its role in the development of cartilage-associated structures, such as joints, has yet to be defined. Using a biotinylated hyaluronan-binding region-link protein complex, free hyaluronan binding sites have been localised in the joint region concomitant with the first signs of cavitation (Stage 37), whereafter it is localised in the joint space and is maintained here as this enlarges. The application of our results is discussed in the context of a primary role for hyaluronan in joint cavity formation. Images Fig. 1 Fig. 2 PMID:1707045

  16. Energy harvesting from vibration with cross-linked polypropylene piezoelectrets

    NASA Astrophysics Data System (ADS)

    Zhang, Xiaoqing; Wu, Liming; Sessler, Gerhard M.

    2015-07-01

    Piezoelectret films are prepared by modification of the microstructure of polypropylene foam sheets cross-linked by electronic irradiation (IXPP), followed by proper corona charging. Young's modulus, relative permittivity, and electromechanical coupling coefficient of the fabricated films, determined by dielectric resonance spectra, are about 0.7 MPa, 1.6, and 0.08, respectively. Dynamic piezoelectric d33 coefficients up to 650 pC/N at 200 Hz are achieved. The figure of merit (FOM, d33 ? g33) for a more typical d33 value of 400 pC/N is about 11.2 GPa-1. Vibration-based energy harvesting with one-layer and two-layer stacks of these films is investigated at various frequencies and load resistances. At an optimum load resistance of 9 M? and a resonance frequency of 800 Hz, a maximum output power of 120 ?W, referred to the acceleration g due to gravity, is obtained for an energy harvester consisting of a one-layer IXPP film with an area of 3.14 cm2 and a seismic mass of 33.7 g. The output power can be further improved by using two-layer stacks of IXPP films in electric series. IXPP energy harvesters could be used to energize low-power electronic devices, such as wireless sensors and LED lights.

  17. Profile of Microbial Keratitis after Corneal Collagen Cross-Linking

    PubMed Central

    Shetty, Rohit; Kaweri, Luci; Nuijts, Rudy M. M. A.; Nagaraja, Harsha; Arora, Vishal; Kumar, Rajesh S.

    2014-01-01

    Purpose. To report the profile of microbial keratitis occurring after corneal collagen cross-linking (CXL) in keratoconus patients. Methods. A retrospective analysis of 2350 patients (1715 conventional CXL, 310 transepithelial CXL, and 325 accelerated CXL) over 7 years (from January 2007 to January 2014) of progressive keratoconus, who underwent CXL at a tertiary eye care centre, was performed. Clinical findings, treatment, and course of disease of four eyes that developed postprocedural moxifloxacin resistant Staphylococcus aureus (MXRSA) infectious keratitis are highlighted. Results. Four eyes that underwent CXL (0.0017%) had corneal infiltrates. All eyes that developed keratitis had conventional CXL. Corneal infiltrates were noted on the third postoperative day. Gram's stain as well as culture reported MXRSA as the causative agent in all cases. Polymerase chain reaction (PCR) in each case was positive for eubacterial genome. All patients were treated with fortified antibiotic eye drops, following which keratitis resolved over a 6-week period with scarring. All these patients were on long-term preoperative oral/topical steroids for chronic disorders (chronic vernal keratoconjunctivitis, bronchial asthma, and chronic eczema). Conclusion. The incidence of infectious keratitis after CXL is a rare complication (0.0017%). MXRSA is a potential organism for causing post-CXL keratitis and should be identified early and treated aggressively with fortified antibiotics. PMID:25302296

  18. Cross-linking of polytetrafluoroethylene during room-temperature irradiation

    SciTech Connect

    Pugmire, David L; Wetteland, Chris J; Duncan, Wanda S; Lakis, Rollin E; Schwartz, Daniel S

    2008-01-01

    Exposure of polytetrafluoroethylene (PTFE) to {alpha}-radiation was investigated to detennine the physical and chemical effects, as well as to compare and contrast the damage mechanisms with other radiation types ({beta}, {gamma}, or thermal neutron). A number of techniques were used to investigate the chemical and physical changes in PTFE after exposure to {alpha}-radiation. These techniques include: Fourier transform infrared spectroscopy (FTIR), differential scanning calorimetry (DSC), and fluorescence spectroscopy. Similar to other radiation types at low doses, the primary damage mechanism for the exposure of PTFE to {alpha}-radiation appears to be chain scission. Increased doses result in a change-over of the damage mechanism to cross-linking. This result is not observed for any radiation type other than {alpha} when irradiation is performed at room temperature. Finally, at high doses, PTFE undergoes mass-loss (via smallfluorocarbon species evolution) and defluorination. The amount and type of damage versus sample depth was also investigated. Other types of radiation yield damage at depths on the order of mm to cm into PTFE due to low linear energy transfer (LET) and the correspondingly large penetration depths. By contrast, the {alpha}-radiation employed in this study was shown to only induce damage to a depth of approximately 26 {mu}m, except at very high doses.

  19. Zinc cross-linked hydroxamated alginates for pulsed drug release

    PubMed Central

    Raut, Neha S; Deshmukh, Prasad R; Umekar, Milind J; Kotagale, Nandkishor R

    2013-01-01

    Introduction: Alginates can be tailored chemically to improve solubility, physicochemical, and biological properties and its complexation with metal ion is useful for controlling the drug release. Materials And Methods: Synthesized N,O-dimethyl, N-methyl, or N-Benzyl hydroxylamine derivatives of sodium alginate were subsequently complexed with zinc to form beads. Hydroxamation of sodium alginate was confirmed by Fourier transform infra-red spectroscopy (FTIR) and differential scanning calorimetry (DSC). Results: The synthesized polymeric material exhibited reduced aqueous, HCl and NaOH solubility. The hydroxamated derivatives demonstrated pulsed release where change in pH of the dissolution medium stimulated the atenolol release. Conclusion: Atenolol loaded Zn cross-linked polymeric beads demonstrated the sustained the plasma drug levels with increased half-life. Although the synthesized derivatives greatly altered the aqueous solubility of sodium alginate, no significant differences in in vitro and in vivo atenolol release behavior amongst the N,O-dimethyl, N-methyl, or N-Benzyl hydroxylamine derivatives of sodium alginate were observed. PMID:24350039

  20. Hyaluronan-oligosaccharide-induced transcription of metalloproteases.

    PubMed

    Fieber, Christina; Baumann, Petra; Vallon, Rüdiger; Termeer, Christian; Simon, Jan C; Hofmann, Martin; Angel, Peter; Herrlich, Peter; Sleeman, Jonathan P

    2004-01-15

    Activated dendritic epidermal Langerhans cells and metastatic tumour cells share many properties. Both cell types can invade the surrounding tissue, enter the lymphatic system and travel to regional lymph nodes. We have recently shown that fragments of the extracellular matrix component hyaluronan, which are typically produced at sites of inflammation, can activate dendritic cells. Upon activation, dendritic cells upregulate expression of matrix metalloproteases (MMPs). These observations prompted us to investigate whether exposure to hyaluronan fragments also induces MMP expression in tumour cells. Here, we report that MMP-9, MMP-13 and urokinase plasminogen activator are upregulated in murine 3LL tumour cells after exposure to mixed-size hyaluronan. Similarly upregulated MMP-9 and MMP-13 expression was observed in primary fibroblasts. By using size-fractionated hyaluronan preparations, we show that the enhanced expression of MMP-9 and MMP-13 is only induced by small hyaluronan (HA) fragments. Although our data suggest that HA-fragment-induced MMP-9 and MMP-13 expression is receptor mediated, they rule out an involvement of the hyaluronan receptors CD44, RHAMM/IHAP and TLR-4. Finally, we show that HA fragment-induced MMP-9 transcription is mediated via NF-kappa B. Our results suggest that the metastasis-associated HA degradation in tumours might promote invasion by inducing MMP expression. PMID:14657275

  1. Characterization of the DeoxyguanosineLysine 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 DNAprotein 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 N?-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

  2. Effect of glucose content on thermally cross-linked fibrous gelatin scaffolds for tissue engineering.

    PubMed

    Siimon, Kaido; Reemann, Paula; Pder, Annika; Pook, Martin; Kangur, Triin; Kingo, Klli; Jaks, Viljar; Meorg, Uno; Jrveklg, Martin

    2014-09-01

    Thermally cross-linked glucose-containing electrospun gelatin meshes were studied as possible cell substrate materials. FTIR analysis was used to study the effect of glucose on cross-linking reactions. It was found that the presence of glucose increases the extent of cross-linking of fibrous gelatin scaffolds, which in return determines scaffold properties and their usability in tissue engineering applications. Easy to handle fabric-like scaffolds were obtained from blends containing up to 15% glucose. Maximum extent of cross-linking was reached at nearly 20% glucose content. Cross-linking effectively resulted in decreased solubility and increased resistance to enzymatic degradation. Preliminary short-term cell culture experiments indicate that such thermally cross-linked gelatin-glucose scaffolds are suitable for tissue engineering applications. PMID:25063151

  3. Cross-linked Polymer-Blend Gate Dielectrics through Thermal Click Chemistry.

    PubMed

    Li, Shengxia; Tang, Wei; Zhang, Weimin; Guo, Xiaojun; Zhang, Qing

    2015-12-01

    New cross-linking reagents were synthesized and mixed with polystyrene (PS) in solution to form a blend. Thin-films were spin-coated from the blend and then cross-linked by thermal activation at relatively low temperature (100?C) to form cross-linked gate dielectrics. This new method is compatible with plastic substrates in flexible electronics. The azide and alkyne cross-linking reagents are kinetically stable at room temperature, so any premature cross-linking is avoided during processing. This method also significantly improved the dielectric performances of PS thin films. Solution-processed top-gate organic field-effect transistor devices with indacenodithiophene-benzothiadiazole copolymer as semiconductor layer and the cross-linked PS blend as dielectric layer showed improved performances with lower gate leakages and higher operation stabilities than devices with neat PS film as dielectric layer. PMID:26477514

  4. 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.

  5. Peroxidase induced oligo-tyrosine cross-links during polymerization of ?-lactalbumin.

    PubMed

    Dhayal, Surender Kumar; Sforza, Stefano; Wierenga, Peter A; Gruppen, Harry

    2015-12-01

    Horseradish peroxidase (HRP) induced cross-linking of proteins has been reported to proceed through formation of di-tyrosine cross-links. In the case of low molar mass phenolic substrates, the enzymatic oxidation is reported to lead to polymerization of the phenols. The aim of this work was to investigate if during oxidative cross-linking of proteins oligo-tyrosine cross-links are formed in addition to dityrosine. To this end, ?-lactalbumin (?-LA) was cross-linked using horseradish peroxidase (HRP) and hydrogen peroxide (H?O?). The reaction products were acid hydrolysed, after which the cross-linked amino acids were investigated by LC-MS and MALDI-MS. To test the effect of the size of the substrate, the cross-linking reaction was also performed with L-tyrosine, N-acetyl L-tyrosinamide and angiotensin. These products were analyzed by LC-MS directly, as well as after acid hydrolysis. In the acid hydrolysates of all samples oligo-tyrosine (Yn, n=3-8) was found in addition to di-tyrosine (Y2). Two stages of cross-linking of ?-LA were identified: a) 1-2 cross-links were formed per monomer until the monomers were converted into oligomers, and b) subsequent cross-linking of oligomers formed in the first stage to form nanoparticles containing 3-4 cross-links per monomer. The transition from first stage to the second stage coincided with the point where di-tyrosine started to decrease and more oligo-tyrosines were formed. In conclusion, extensive polymerization of ?-LA using HRP via oligo-tyrosine cross-links is possible, as is the case for low molar mass tyrosine containing substrates. PMID:26282909

  6. Effect of radiation cross-linking on the abrasive wear behaviour of polyethylenes

    NASA Astrophysics Data System (ADS)

    Gul, Rizwan M.; Khan, Tahir I.

    2014-06-01

    This study explores the differences in the dry abrasive wear behavior of different polyethylenes, and compares the effect of radiation cross-linking on the wear behavior. Four different types of polyethylenes: LDPE, LLDPE, HDPE and UHMWPE were studied. Cross-linking was carried out by high energy electron beam with radiation dose of 200 kGy. The results show that in unirradiated state UHMWPE has excellent wear resistance, with HDPE showing comparable wear properties; both LDPE and LLDPE exhibit high wear rate. Cross-linking improves wear rate of LDPE and UHMWPE, however, the wear rate of HDPE and LLDPE increases with cross-linking.

  7. Effect of cross-linking and enzymatic hydrolysis composite modification on the properties of rice starches.

    PubMed

    Xiao, Huaxi; Lin, Qinlu; Liu, Gao-Qiang

    2012-01-01

    Native rice starch lacks the versatility necessary to function adequately under rigorous industrial processing, so modified starches are needed to meet the functional properties required in food products. This work investigated the impact of enzymatic hydrolysis and cross-linking composite modification on the properties of rice starches. Rice starch was cross-linked with epichlorohydrin (EPI) with different concentrations (0.5%, 0.7%, 0.9% w/w, on a dry starch basis), affording cross-linked rice starches with the three different levels of cross-linking that were named R?, R?, and R?, respectively. The cross-linked rice starches were hydrolyzed by ?-amylase and native, hydrolyzed, and hydrolyzed cross-linked rice starches were comparatively studied. It was found that hydrolyzed cross-linked rice starches showed a lower the degree of amylase hydrolysis compared with hydrolyzed rice starch. The higher the degree of cross-linking, the higher the capacity to resist enzyme hydrolysis. Hydrolyzed cross-linked rice starches further increased the adsorptive capacities of starches for liquids and decreased the trend of retrogradation, and it also strengthened the capacity to resist shear compared to native and hydrolyzed rice starches. PMID:22772809

  8. Viscoelastic Properties of Hyaluronan in Physiological Conditions

    PubMed Central

    Cowman, Mary K.; Schmidt, Tannin A.; Raghavan, Preeti; Stecco, Antonio

    2015-01-01

    Hyaluronan (HA) is a high molecular weight glycosaminoglycan of the extracellular matrix (ECM), which is particularly abundant in soft connective tissues. Solutions of HA can be highly viscous with non-Newtonian flow properties. These properties affect the movement of HA-containing fluid layers within and underlying the deep fascia. Changes in the concentration, molecular weight, or even covalent modification of HA in inflammatory conditions, as well as changes in binding interactions with other macromolecules, can have dramatic effects on the sliding movement of fascia. The high molecular weight and the semi-flexible chain of HA are key factors leading to the high viscosity of dilute solutions, and real HA solutions show additional nonideality and greatly increased viscosity due to mutual macromolecular crowding. The shear rate dependence of the viscosity, and the viscoelasticity of HA solutions, depend on the relaxation time of the molecule, which in turn depends on the HA concentration and molecular weight. Temperature can also have an effect on these properties. High viscosity can additionally affect the lubricating function of HA solutions. Immobility can increase the concentration of HA, increase the viscosity, and reduce lubrication and gliding of the layers of connective tissue and muscle. Over time, these changes can alter both muscle structure and function. Inflammation can further increase the viscosity of HA-containing fluids if the HA is modified via covalent attachment of heavy chains derived from Inter-α-Inhibitor. Hyaluronidase hydrolyzes HA, thus reducing its molecular weight, lowering the viscosity of the extracellular matrix fluid and making outflow easier. It can also disrupt any aggregates or gel-like structures that result from HA being modified. Hyaluronidase is used medically primarily as a dispersion agent, but may also be useful in conditions where altered viscosity of the fascia is desired, such as in the treatment of muscle stiffness. PMID:26594344

  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. Integrated Cryogenic Satellite Communications Cross-Link Receiver Experiment

    NASA Technical Reports Server (NTRS)

    Romanofsky, R. R.; Bhasin, K. B.; Downey, A. N.; Jackson, C. J.; Silver, A. H.; Javadi, H. H. S.

    1995-01-01

    An experiment has been devised which will validate, in space, a miniature, high-performance receiver. The receiver blends three complementary technologies; high temperature superconductivity (HTS), pseudomorphic high electron mobility transistor (PHEMT) monolithic microwave integrated circuits (MMIC), and a miniature pulse tube cryogenic cooler. Specifically, an HTS band pass filter, InP MMIC low noise amplifier, HTS-sapphire resonator stabilized local oscillator (LO), and a miniature pulse tube cooler will be integrated into a complete 20 GHz receiver downconverter. This cooled downconverter will be interfaced with customized signal processing electronics and integrated onto the space shuttle's 'HitchHiker' carrier. A pseudorandom data sequence will be transmitted to the receiver, which is in low Earth orbit (LEO), via the Advanced Communication Technology Satellite (ACTS) on a 20 GHz carrier. The modulation format is QPSK and the data rate is 2.048 Mbps. The bit error rate (BER) will be measured in situ. The receiver is also equipped with a radiometer mode so that experiment success is not totally contingent upon the BER measurement. In this mode, the receiver uses the Earth and deep space as a hot and cold calibration source, respectively. The experiment closely simulates an actual cross-link scenario. Since the receiver performance depends on channel conditions, its true characteristics would be masked in a terrestrial measurement by atmospheric absorption and background radiation. Furthermore, the receiver's performance depends on its physical temperature, which is a sensitive function of platform environment, thermal design, and cryocooler performance. This empirical data is important for building confidence in the technology.

  11. Corneal cross-linking in 9 horses with ulcerative keratitis

    PubMed Central

    2013-01-01

    Background Corneal ulcers are one of the most common eye problems in the horse and can cause varying degrees of visual impairment. Secondary infection and protease activity causing melting of the corneal stroma are always concerns in patients with corneal ulcers. Corneal collagen cross-linking (CXL), induced by illumination of the corneal stroma with ultraviolet light (UVA) after instillation of riboflavin (vitamin B2) eye drops, introduces crosslinks which stabilize melting corneas, and has been used to successfully treat infectious ulcerative keratitis in human patients. Therefore we decided to study if CXL can be performed in sedated, standing horses with ulcerative keratitis with or without stromal melting. Results Nine horses, aged 1 month to 16 years (median 5 years) were treated with a combination of CXL and medical therapy. Two horses were diagnosed with mycotic, 5 with bacterial and 2 with aseptic ulcerative keratitis. A modified Dresden-protocol for CXL could readily be performed in all 9 horses after sedation. Stromal melting, diagnosed in 4 horses, stopped within 24 h. Eight of nine eyes became fluorescein negative in 13.5 days (median time; range 426 days) days after CXL. One horse developed a bacterial conjunctivitis the day after CXL, which was successfully treated with topical antibiotics. One horse with fungal ulcerative keratitis and severe uveitis was enucleated 4 days after treatment due to panophthalmitis. Conclusions CXL can be performed in standing, sedated horses. We did not observe any deleterious effects attributed to riboflavin or UVA irradiation per se during the follow-up, neither in horses with infectious nor aseptic ulcerative keratitis. These data support that CXL can be performed in the standing horse, but further studies are required to compare CXL to conventional medical treatment in equine keratitis and to optimize the CXL protocol in this species. PMID:23803176

  12. Encoding Hydrogel Mechanics via Network Cross-Linking Structure

    PubMed Central

    2015-01-01

    The effects of mechanical cues on cell behaviors in 3D remain difficult to characterize as the ability to tune hydrogel mechanics often requires changes in the polymer density, potentially altering the material’s biochemical and physical characteristics. Additionally, with most PEG diacrylate (PEGDA) hydrogels, forming materials with compressive moduli less than ∼10 kPa has been virtually impossible. Here, we present a new method of controlling the mechanical properties of PEGDA hydrogels independent of polymer chain density through the incorporation of additional vinyl group moieties that interfere with the cross-linking of the network. This modification can tune hydrogel mechanics in a concentration dependent manner from <1 to 17 kPa, a more physiologically relevant range than previously possible with PEG-based hydrogels, without altering the hydrogel’s degradation and permeability. Across this range of mechanical properties, endothelial cells (ECs) encapsulated within MMP-2/MMP-9 degradable hydrogels with RGDS adhesive peptides revealed increased cell spreading as hydrogel stiffness decreased in contrast to behavior typically observed for cells on 2D surfaces. EC-pericyte cocultures exhibited vessel-like networks within 3 days in highly compliant hydrogels as compared to a week in stiffer hydrogels. These vessel networks persisted for at least 4 weeks and deposited laminin and collagen IV perivascularly. These results indicate that EC morphogenesis can be regulated using mechanical cues in 3D. Furthermore, controlling hydrogel compliance independent of density allows for the attainment of highly compliant mechanical regimes in materials that can act as customizable cell microenvironments. PMID:26082943

  13. Transepithelial corneal collagen cross-linking in ultrathin keratoconic corneas

    PubMed Central

    Spadea, Leopoldo; Mencucci, Rita

    2012-01-01

    Background The purpose of this paper was to report the results of transepithelial corneal collagen cross-linking (CXL) with modified riboflavin and ultraviolet A irradiation in patients affected by keratoconus, each with thinnest pachymetry values of less than 400 μm (with epithelium) and not treatable using standard de-epithelialization techniques. Methods Sixteen patients affected by progressive keratoconus with thinnest pachymetry values ranging from 331 μm to 389 μm underwent transepithelial CXL in one eye using a riboflavin 0.1% solution in 15% Dextran T500 containing ethylenediamine tetra-acetic acid 0.01% and trometamol to enhance epithelial penetration. The patients underwent complete ophthalmological examination, including endothelial cell density measurements and computerized videokeratography, before CXL and at one day, one week, and one, 6, and 12 months thereafter. Results Epithelial healing was complete in all patients after one day of use of a soft bandage contact lens. No side effects or damage to the limbal region was observed during the follow-up period. All patients showed slightly improved uncorrected and spectacle-corrected visual acuity; keratometric astigmatism showed reductions (up to 5.3 D) and apical ectasia power decreased (Kmax values reduced up to 4.3 D). Endothelial cell density was unchanged. Conclusion Application of transepithelial CXL using riboflavin with substances added to enhance epithelial permeability was safe, seemed to be moderately effective in keratoconic eyes with ultrathin corneas, and applications of the procedure could be extended to patients with advanced keratoconus. PMID:23152657

  14. Influence of vitamin D status on hyaluronan localization in bone.

    PubMed

    Genever, P G; Dickson, I R

    1996-05-01

    The distribution of hyaluronan was investigated in the proximal tibiotarsal bones of normal (vitamin D-treated) chicks, and chicks with rachitic lesions induced by vitamin D deficiency. Localization studies using a biotinylated hyaluronan-binding probe revealed that in vitamin D-treated chicks, a high level of hyaluronan staining was present in upper proliferative zone cartilage and upper hypertrophic zone cartilage. Hyaluronan staining was greatly reduced in the zone of provisional calcification. In the metaphyses and diaphyses of normal chicks hyaluronan was predominantly localized to the non-bone-forming surfaces of osteoblasts but was also present on the basolateral surfaces of lining cells and osteoclasts. Marked changes in hyaluronan distribution were observed in vitamin D-deficient chicks. The amount of hyaluronan present in proliferative zone growth cartilage was similar to control chicks, although with a more widespread distribution, extending into lower proliferative zone cartilage. In the zone of hypertrophy/calcification, biochemical analyses revealed that hyaluronan levels in rachitic chicks were about 3.6 times greater than in vitamin D-treated chicks; localization studies demonstrated that this increase was associated with the presence of hyaluronan-positive spindle-shaped cells in the metaphyseal vascular spaces. Intense hyaluronan staining was also associated with abundant spindle-shaped cells occupying the marrow spaces of rachitic diaphyseal bone. The distribution of hyaluronan in vitamin D-treated chick bone, and the alterations observed in rachitic tissue suggests a role for hyaluronan in endochondral bone formation. PMID:8739900

  15. Response of biopolymer networks governed by the physical properties of cross-linking molecules.

    PubMed

    Wei, Xi; Zhu, Qian; Qian, Jin; Lin, Yuan; Shenoy, V B

    2016-02-23

    In this study, we examine how the physical properties of cross-linking molecules affect the bulk response of bio-filament networks, an outstanding question in the study of biological gels and the cytoskeleton. We show that the stress-strain relationship of such networks typically undergoes linear increase - strain hardening - stress serration - total fracture transitions due to the interplay between the bending and stretching of individual filaments and the deformation and breakage of cross-linkers. Interestingly, the apparent network modulus is found to scale with the linear and rotational stiffness of the crosslinks to a power exponent of 0.78 and 0.13, respectively. In addition, the network fracture energy will reach its minimum at intermediate rotational compliance values, reflecting the fact that most of the strain energy will be stored in the distorted filaments with rigid cross-linkers while the imposed deformation will be "evenly" distributed among significantly more crosslinking molecules with high rotational compliance. PMID:26760315

  16. 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. PMID:23399178

  17. Use of Hyaluronan-Derived Hydrogels for Three-Dimensional Cell Culture and Tumor Xenografts

    PubMed Central

    Serban, Monica A.; Scott, Anna; Prestwich, Glenn D.

    2012-01-01

    The practice of in vitro three-dimensional (3-D) cell culture has lagged behind the realization that classical two-dimensional (2-D) culture on plastic surfaces fails to mirror normal cell biology. Biologically, a complex network of proteins and proteoglycans that constitute the extracellular matrix (ECM) surrounds every cell. To recapitulate the normal cellular behavior, scaffolds (ECM analogs) that reconstitute the essential biological cues are required. This unit describes the 3-D cell culture and tumor engineering applications of Extracel, a novel semisynthetic ECM (sECM), based on cross-linked derivatives of hyaluronan and gelatin. A simplified cell encapsulation and pseudo-3-D culturing (on top of hydrogels) protocol is provided. In addition, the use of this sECM as a vehicle to obtain tumor xenografts with improved take rates and tumor growth is presented. These engineered tumors can be used to evaluate anticancer therapies under physiologically relevant conditions. PMID:18819087

  18. Investigation of anisotropic thermal transport in cross-linked polymers

    NASA Astrophysics Data System (ADS)

    Simavilla, David Nieto

    Thermal transport in lightly cross-linked polyisoprene and polybutadine subjected to uniaxial elongation is investigated experimentally. We employ two experimental techniques to assess the effect that deformation has on this class of materials. The first technique, which is based on Forced Rayleigh Scattering (FRS), allows us to measure the two independent components of the thermal diffusivity tensor as a function of deformation. These measurements along with independent measurements of the tensile stress and birefringence are used to evaluate the stress-thermal and stress-optic rules. The stress-thermal rule is found to be valid for the entire range of elongations applied. In contrast, the stress-optic rule fails for moderate to large stretch ratios. This suggests that the degree of anisotropy in thermal conductivity depends on both orientation and tension in polymer chain segments. The second technique, which is based on infrared thermography (IRT), allows us to measure anisotropy in thermal conductivity and strain induced changes in heat capacity. We validate this method measurements of anisotropic thermal conductivity by comparing them with those obtained using FRS. We find excellent agreement between the two techniques. Uncertainty in the infrared thermography method measurements is estimated to be about 2-5 %. The accuracy of the method and its potential application to non-transparent materials makes it a good alternative to extend current research on anisotropic thermal transport in polymeric materials. A second IRT application allows us to investigate the dependence of heat capacity on deformation. We find that heat capacity increases with stretch ratio in polyisoprene specimens under uniaxial extension. The deviation from the equilibrium value of heat capacity is consistent with an independent set of experiments comparing anisotropy in thermal diffusivity and conductivity employing FRS and IRT techniques. We identify finite extensibility and strain-induced crystallization as the possible causes explaining our observations and evaluate their contribution making use of classical rubber elasticity results. Finally, we study of the role of evaporation-induced thermal effects in the well-know phenomena of the tears of wine. We develop a transport model and support its predictions by experimentally measuring the temperature gradient present in wine and cognac films using IRT. Our results demonstrate that the Marangoni flow responsible for wine tears results from both composition and temperature gradients, whose relative contribution strongly depends on the thermodynamic properties of ethanol-water mixtures. The methods developed here can be used to obtain a deeper understanding of Marangoni flows, which are ubiquitous in nature and modern technology.

  19. Chemistry and Physical Properties of Melt Processed- and Solution- Cross Linked Corn Zein

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Corn zein was cross linked with the glutaraldehyde (GDA) using glacial acetic acid (HAc) as catalyst. The objectives are to enhance the mechanical properties of poured films and to compare them with compression molded tensile bars from melt processed zein. Chemistry of the cross linking reaction w...

  20. Electrospun zein fibers using glutaraldehyde as the cross-linking reagent

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Glutaraldehyde was used as a cross-linking reagent for zein (corn protein) to provide fibers with improved physical properties and solvent resistance. Glutaraldehyde was used at levels between 2 - 8%. The cross-linking reaction was carried out in acetic acid for twenty hours at room temperature. ...

  1. 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...

  2. Lamb and Cow Performance when Fed Corn Silage that has Reduced Ferulate Cross Linking

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ferulate-mediated lignin/hemicellulose cross linking in grasses reduces in vitro NDF digestibility (IVNDFD). Impact of ferulate cross linking on animal performance was examined in lamb digestibility and dairy cow performance trials using the seedling ferulate ester (sfe) corn mutant that reduces cro...

  3. PREPARATION OF NOVEL METALLIC AND BIMETALLIC CROSS-LINKED POLY (VINYL ALCOHOL) NANOCOMPOSITES UNDER MICROWAVE IRRADIATION

    EPA Science Inventory

    A facile method utilizing microwave irradiation is described that accomplishes the cross-linking reaction of PVA with metallic and bimetallic systems. Nanocomposites of PVA-cross-linked metallic systems such as Pt, Cu, and In and bimetallic systems such as Pt-In, Ag-Pt, Pt-Fe, Cu...

  4. Electrospun zein fibers using glyoxal or formaldehyde as the cross-linking reagent

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Glyoxal or formaldehyde was used as a cross-linking reagent for zein (corn protein) to provide electrospun fibers with improved physical properties and solvent resistance. These reagents were used between 2 and 6%. The cross-linking reaction was carried out in acetic acid for various lengths of ti...

  5. Cross-Linking Electrochemical Mass Spectrometry for Probing Protein Three-Dimensional Structures

    PubMed Central

    2015-01-01

    Chemical cross-linking combined with mass spectrometry (MS) is powerful to provide protein three-dimensional structure information but difficulties in identifying cross-linked peptides and elucidating their structures limit its usefulness. To tackle these challenges, this study presents a novel cross-linking MS in conjunction with electrochemistry using disulfide-bond-containing dithiobis[succinimidyl propionate] (DSP) as the cross-linker. In our approach, electrolysis of DSP-bridged protein/peptide products, as online monitored by desorption electrospray ionization mass spectrometry is highly informative. First, as disulfide bonds are electrochemically reducible, the cross-links are subject to pronounced intensity decrease upon electrolytic reduction, suggesting a new way to identify cross-links. Also, mass shift before and after electrolysis suggests the linkage pattern of cross-links. Electrochemical reduction removes disulfide bond constraints, possibly increasing sequence coverage for tandem MS analysis and yielding linear peptides whose structures are more easily determined than their cross-linked precursor peptides. Furthermore, this cross-linking electrochemical MS method is rapid, due to the fast nature of electrochemical conversion (much faster than traditional chemical reduction) and no need for chromatographic separation, which would be of high value for structural proteomics research. PMID:25141260

  6. A novel cross-linked human amniotic membrane for corneal implantations.

    PubMed

    Sekar, S; Sasirekha, K; Krishnakumar, S; Sastry, T P

    2013-03-01

    The aim of this study was to evaluate the efficacy of Al2(SO4)3 cross-linked human amniotic membrane for ocular surface reconstruction using tissue culture techniques. The human amniotic membrane was cross-linked with Al2(SO4)3, and the cross-linked human amniotic membrane was characterized for its mechanical properties, percentage of swelling in water, sterility, infrared spectroscopy and scanning electron microscopy. The potential of cross-linked human amniotic membrane to support the attachment and proliferation of corneal limbal epithelial cells was assessed in vitro, using static culture system. About 125% increase in the tensile strength was observed in the cross-linked human amniotic membrane compared to human amniotic membrane. Infrared spectroscopy studies have confirmed the cross-linking of human amniotic membrane with Al2(SO4)3. The cross-linked human amniotic membrane was found to be sterile up to 1 year. In culture studies, confluent sheets of epithelial cells were seen at the end of 14th day resembling the morphological features of limbal epithelia. The cross-linked human amniotic membrane has exhibited improved mechanical properties, and the tissue culture studies have shown its feasibility to be used as a limbal transplant. It was concluded that the crosslinked human amniotic membrane with its improved mechanical properties could be used on par with human amniotic membrane. PMID:23662337

  7. Ionically Cross-Linked Polymer Networks for the Multiple-Month Release of Small Molecules.

    PubMed

    Lawrence, Patrick G; Patil, Pritam S; Leipzig, Nic D; Lapitsky, Yakov

    2016-02-24

    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

  8. Ionically cross-linked alginate hydrogels as tissue engineering scaffolds

    NASA Astrophysics Data System (ADS)

    Kuo, Catherine Kyleen

    Generation of living tissues through tissue engineering can be achieved via incorporation of cells into synthetic scaffolds designed to facilitate new tissue formation. Necessary characteristics of a scaffold include biocompatibility, high porosity with controllable pore size and interconnectivity, moldability, chemical and mechanical stability, and structural homogeneity. Hydrogels often possess many of the necessary characteristics and thus are favorable candidates for scaffolding. Alginate hydrogels are commonly made by ionically crosslinking with calcium ions from CaCl2 or CaSO4. These hydrogels are favored for their mild gel formation, however the gelation rate is rapid and uncontrollable (fast-gelation), resulting in varying crosslinking density throughout the gel. In this work, structurally homogeneous calcium alginate hydrogels were formed via a slow-gelation system that utilizes uniform mixing of CaCO3 with sodium alginate solution, and the addition of slowly hydrolyzing D-gluconic acid lactone to slowly release calcium ions for crosslinking. Homogeneity and mechanical properties of these hydrogels were shown to be superior to those of fast-gelled hydrogels. Gelation rate was controlled through the incorporation of CaSO4, and by varying total calcium content, polymer concentration and gelation temperature. Control over mechanical properties and diffusivity was demonstrated in the homogeneous hydrogels by adjusting compositional variables. Consistent control over solute diffusivity through gel discs reflected the structural homogeneity of the gels. To overcome the instability of ionically crosslinked gels in tissue culture medium, a method was developed to control the hydrogel dimensions by adjusting the ionic concentration of the medium. Stability of the hydrogels in this controlled environment was characterized through swelling experiments and mechanical testing. To provide for scaffold degradation and thereby promote tissue growth, alginate lyase was incorporated into the system induce enzymatically controlled alginate degradation. This alginate hydrogel system is novel in that cells are directly and uniformly incorporated into a hydrogel with controlled gelation rate and material properties. The injectable alginate hydrogels can be molded into complex shapes or injected into the body to avoid invasive surgery. Further, the ability to tailor their gelation rate, material properties and degradation enable these alginate hydrogels to be used for a variety of biomedical applications including drug or biofactor delivery.

  9. 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.

  10. 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. PMID:26750612

  11. Cross-Linking the Surface of Cured Polydimethylsiloxane via Hyperthemal Hydrogen Projectile Bombardment.

    PubMed

    Bao, Chao; Xu, Ke-Qin; Tang, Chang-Yu; Lau, Woon-ming; Yin, Cong-Bin; Zhu, Yan; Mei, Jun; Lee, Jonathan; Hui, David; Nie, Heng-Yong; Liu, Yu

    2015-04-29

    Cross-linking of polydimethylsiloxane (PDMS) is increasingly important with recent focus on its top surface stiffness. In this paper, we demonstrate that hyperthermal hydrogen projectile bombardment, a surface sensitive cross-linking technology, is superior in enhancing the mechanical properties of a cured PDMS surface without significantly degrading its hydrophobicity. Both water contact angle measurements and time-of-flight secondary ion mass spectrometry are used to investigate the variations in surface chemistry and structure upon cross-linking. Using nanoindentation and atomic force microscopy, we confirm that the thickness of the cross-linked PDMS is controllable by the bombardment time, which opens opportunities for tuning cross-linking degree in compliance with arising requirements from the practice. PMID:25849306

  12. Modified dextran cross-linked electrospun gelatin nanofibres for biomedical applications.

    PubMed

    Jalaja, K; Kumar, P R Anil; Dey, Tuli; Kundu, Subhas C; James, Nirmala R

    2014-12-19

    Electrospun gelatin nanofibres attract attention of bioengineering arena because of its excellent biocompatibility and structural resemblance with native extracellular matrix. In this study, we have developed gelatin nanofibres using an innovative cross-linking approach to minimize cytotoxic effects. Gelatin was dissolved in water:acetic acid (8:2, v/v) solution and electrospun to form nanofibres with diameter in the range of 156 30 nm. The nanofibres were cross-linked with a modified polysaccharide, namely, dextran aldehyde (DA). Cross-linking with DA could be achieved without compromising the fibrous architecture. DA cross-linked gelatin nanofibres maintained the fibrous morphology in aqueous medium. These mats exhibit improved mechanical properties and gradual degradation behaviour. The nanofibres were evaluated for cytotoxicity, cell adhesion, viability, morphology and proliferation using L-929 fibroblast cells. The results confirmed that DA cross-linked mats were non cytotoxic towards L-929 cells with good cell adhesion, spreading and proliferation. PMID:25263915

  13. Comparison of chitosan/starch composite film properties before and after cross-linking.

    PubMed

    Li, Haihong; Gao, Xiaochen; Wang, Yan; Zhang, Xiaobo; Tong, Zhiwei

    2013-01-01

    Unmodified and cross-linked chitosan/starch composite films were prepared using the solvent evaporation method. The properties of the films were studied to obtain useful information about the possible applications of composite films. FT-IR, SEM, and swelling property investigations show that the cross-linking agent glutaraldehyde reacts in the chitosan and starch blend. The compatibility of chitosan and starch blends before and after cross-linking was studied by UV-vis spectroscopy. The compatibility of the blends deteriorated after cross-linking. This finding was confirmed by the results of mechanical properties. The films show improved water barrier performance after cross-linking. The use of trace concentrations of glutaraldehyde in chitosan/starch films allows for possible application in the biomedical field. PMID:23107802

  14. Hydroxyl radical induced cross-linking between phenylalanine and 2-deoxyribose

    SciTech Connect

    Farahani, M.; Simic, M.G.

    1988-06-28

    Hydroxy radicals induce cross-linking between phenylalanine (Phe) and 2-deoxyribose (dR) via formation of corresponding free radical intermediates. The cross-linked products were separated and identified by capillary gas chromatography-mass spectrometry. When phenylalanine and 2-deoxyribose radicals were generated in a 1:1 ratio, the predominant interaction was between Phe and dR radicals while the Phe-Phe and dR-dR cross-links were less abundant. The newly discovered cross-linked between 2-deoxyribose and phenylalanine may serve as a model for radiation or free radical induced cross-linking between DNA and proteins and in general between sugar moieties and amino acids.

  15. Mechanical characterization of electrospun gelatin scaffolds cross-linked by glucose.

    PubMed

    Siimon, Kaido; Siimon, Hele; Jrveklg, Martin

    2015-01-01

    Nanofibrous gelatin scaffolds were prepared by electrospinning from aqueous acetic acid and cross-linked thermally by glucose. The effect of the amount of glucose used as cross-linking agent on the mechanical properties of gelatin fibres was studied in this paper. The elastic modulus of gelatin fibres cross-linked by glucose was determined by modelling the behaviour of the meshes during tensile test. The model draws connections between the elastic moduli of a fibrous mesh and the fibre material and allows evaluation of elastic modulus of the fibre material. It was found that cross-linking by glucose increases the elastic modulus of gelatin fibres from 0.3 GPa at 0 % glucose content to 1.1 GPa at 15 % glucose content. This makes fibrous gelatin scaffolds cross-linked by glucose a promising material for biomedical applications. PMID:25578715

  16. Analysis of protein-RNA interactions in CRISPR proteins and effector complexes by UV-induced cross-linking and mass spectrometry.

    PubMed

    Sharma, Kundan; Hrle, Ajla; Kramer, Katharina; Sachsenberg, Timo; Staals, Raymond H J; Randau, Lennart; Marchfelder, Anita; van der Oost, John; Kohlbacher, Oliver; Conti, Elena; Urlaub, Henning

    2015-11-01

    Ribonucleoprotein (RNP) complexes play important roles in the cell by mediating basic cellular processes, including gene expression and its regulation. Understanding the molecular details of these processes requires the identification and characterization of protein-RNA interactions. Over the years various approaches have been used to investigate these interactions, including computational analyses to look for RNA binding domains, gel-shift mobility assays on recombinant and mutant proteins as well as co-crystallization and NMR studies for structure elucidation. Here we report a more specialized and direct approach using UV-induced cross-linking coupled with mass spectrometry. This approach permits the identification of cross-linked peptides and RNA moieties and can also pin-point exact RNA contact sites within the protein. The power of this method is illustrated by the application to different single- and multi-subunit RNP complexes belonging to the prokaryotic adaptive immune system, CRISPR-Cas (CRISPR: clustered regularly interspaced short palindromic repeats; Cas: CRISPR associated). In particular, we identified the RNA-binding sites within three Cas7 protein homologs and mapped the cross-linking results to reveal structurally conserved Cas7 - RNA binding interfaces. These results demonstrate the strong potential of UV-induced cross-linking coupled with mass spectrometry analysis to identify RNA interaction sites on the RNA binding proteins. PMID:26071038

  17. Carbodiimide cross-linking of amniotic membranes in the presence of amino acid bridges.

    PubMed

    Lai, Jui-Yang

    2015-06-01

    The purpose of this study was to investigate the carbodiimide cross-linking of amniotic membrane (AM) in the presence of amino acid bridges. The biological tissues were treated with glycine, lysine, or glutamic acid and chemically cross-linked to examine the role of amino acid types in collagenous biomaterial processing. Results of zeta potential measurements showed that the use of uncharged, positively and negatively charged amino acids dictates the charge state of membrane surface. Tensile strength and water content measurements demonstrated that the addition of lysine molecules to the cross-linking system can increase the cross-linking efficiency and dehydration degree while the introduction of glutamic acid in the AM samples decreases the number of cross-links per unit mass of chemically modified tissue collagen. The differences in the cross-linking density further determined the thermal and biological stability by differential scanning calorimetry and in vitro degradation tests. As demonstrated in matrix permeability studies, the improved formation of covalent cross-linkages imposed by lysine facilitated construction of stronger cross-linking structures. In contrast, the added glycine molecules were insufficient to enhance the resistances of the proteinaceous matrices to thermal denaturation and enzymatic degradation. The cytocompatibility of these biological tissue membranes was evaluated by using human corneal epithelial cell cultures. Results of cell viability, metabolic activity, and pro-inflammatory gene expression level showed that the AM materials cross-linked with carbodiimide in the presence of different types of amino acids are well tolerated without evidence of detrimental effect on cell growth. In addition, the amino acid treated and carbodiimide cross-linked AM implants had good biocompatibility in the anterior chamber of the rabbit eye model. Our findings suggest that amino acid type is a very important engineering parameter to mediate carbodiimide cross-linking of AM collagen. PMID:25842104

  18. Characterization of the bombesin receptor on mouse pancreatic acini by chemical cross-linking

    SciTech Connect

    Huang, S.C.; Yu, D.H.; Wank, S.A.; Gardner, J.D.; Jensen, R.T. )

    1990-11-01

    Bombesin (BN), gastrin-releasing peptide (GRP) and GRP(18-27) (neuromedin C) were equipotent and 30-fold more potent than neuromedin B (NMB) in inhibiting binding of {sup 125}I-GRP to and in stimulating amylase release from mouse pancreatic acini. In the present study we used {sup 125}I-GRP and chemical cross-linking techniques to characterize the mouse pancreatic BN receptor. After binding of {sup 125}I-GRP to membranes, and incubation with various chemical cross-linking agents, cross-linked radioactivity was analyzed by SDS-PAG electrophoresis and autoradiography. With each of 4 different chemical cross-linking agents, there was a single broad polypeptide band of Mr 80,000. Cross-linking did not occur in the absence of the cross-linking agent. Cross-linking was inhibited only by peptides that interact with the BN receptor such as GRP, NMB, GRP(18-27) or BN. Dose-inhibition curves for the ability of BN or NMB to inhibit binding of {sup 125}I-GRP to membranes or cross-linking to the 80,000 polypeptide demonstrated for both that BN was 15-fold more potent than NMB. The apparent molecular weight of the cross-linked polypeptide was unchanged by adding dithiothreitol. N-Glycanase treatment reduced the molecular weight of the cross-linked peptide to 40,000. The present results indicate that the BN receptor on mouse pancreatic acinar cell membranes resembles that recently described on various tumor cells in being a single glycoprotein with a molecular weight of 76,000. Because dithiothreitol had no effect, this glycoprotein is not a subunit of a larger disulfide-linked structure.

  19. Stabilization of native amyloid ?-protein oligomers by Copper and Hydrogen peroxide Induced Cross-linking of Unmodified Proteins (CHICUP).

    PubMed

    Williams, Thomas L; Serpell, Louise C; Urbanc, Brigita

    2016-03-01

    Oligomeric assemblies are postulated to be proximate neurotoxic species in human diseases associated with aberrant protein aggregation. Their heterogeneous and transient nature makes their structural characterization difficult. Size distributions of oligomers of several amyloidogenic proteins, including amyloid ?-protein (A?) relevant to Alzheimer's disease (AD), have been previously characterized in vitro by photo-induced cross-linking of unmodified proteins (PICUP) followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Due to non-physiological conditions associated with the PICUP chemistry, A? oligomers cross-linked by PICUP may not be representative of in vivo conditions. Here, we examine an alternative Copper and Hydrogen peroxide Induced Cross-linking of Unmodified Proteins (CHICUP), which utilizes naturally occurring divalent copper ions and hydrogen peroxide and does not require photo activation. Our results demonstrate that CHICUP and PICUP applied to the two predominant A? alloforms, A?40 and A?42, result in similar oligomer size distributions. Thioflavin T fluorescence data and atomic force microscopy images demonstrate that both CHICUP and PICUP stabilize A? oligomers and attenuate fibril formation. Relative to noncross-linked peptides, CHICUP-treated A?40 and A?42 cause prolonged disruption to biomimetic lipid vesicles. CHICUP-stabilized A? oligomers link the amyloid cascade, metal, and oxidative stress hypotheses of AD into a more comprehensive understanding of the molecular basis of AD pathology. Because copper and hydrogen peroxide are elevated in the AD brain, CHICUP-stabilized A? oligomers are biologically relevant and should be further explored as a new therapeutic target. PMID:26699836

  20. Corneal changes following collagen cross linking and simultaneous topography guided photoablation with collagen cross linking for keratoconus

    PubMed Central

    Padmanabhan, Prema; Radhakrishnan, Aishwaryah; Venkataraman, Abinaya Priya; Gupta, Nidhi; Srinivasan, Bhaskar

    2014-01-01

    Purpose: To compare the outcome of Collagen cross-linking (CXL) with that following topography-guided customized ablation treatment (T-CAT) with simultaneous CXL in eyes with progressive keratoconus. Materials and Methods: This was a prospective, non-randomized single centre study of 66 eyes with progressive keratoconus. Of these, 40 eyes underwent CXL and 26 eyes underwent T-CAT + CXL. The refractive, topographic, tomographic and aberrometric changes measured at baseline, 1, 3 and 6 months post-operatively were compared between both groups. Results: After a mean follow-up of 7.7 1.3 months, the mean retinoscopic cylinder decreased by 1.02 3.16 D in the CXL group (P = 0.1) and 2.87 3.22 D in the T-CAT + CXL group (P = 0.04). The Best corrected visual acuity increased by 2 lines or more in 10% of eyes in the CXL group and in 23.3% of eyes in the T-CAT + CXL group. The mean steepest-K reduced by 0.40 3.71 D (P = 0.77) in the CXL group and by 2.91 2.01D (P = 0.03) in the T-CAT + CXL group. The sag factor and surface asymmetry index showed no significant change in the CXL group but reduced by 3.59 5.94 D (P = 0.01) and 0.72 1.18 (P = 0.02) respectively in the T-CAT + CXL group. There was a significant increase in the highest posterior corneal elevation in both groups (9.57 14.93 ? in the CXL group and 7.85 9.25 ? in the T-CAT + CXL group, P ? 0.001 for both). There was significantly greater reduction of mean coma (P < 0.001) and mean higher-order aberrations (P = 0.01) following T-CAT + CXL compared to CXL. Conclusions: CAT + CXL is an effective approach to confer biomechanical stability and to improve the corneal contour in eyes with keratoconus and results in better refractive, topographic and aberrometric outcomes than CXL alone. PMID:23619500

  1. 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

  2. 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

  3. Determination of protein conformation by isotopically labelled cross-linking and dedicated software

    NASA Astrophysics Data System (ADS)

    Nielsen, Tina; Thaysen-Andersen, Morten; Larsen, Nanna; Jørgensen, Flemming S.; Houen, Gunnar; Højrup, Peter

    2007-12-01

    Chemical cross-linking in conjunction with mass spectrometry (MS) can be used for sensitive and rapid investigation of the three-dimensional structure of proteins at low resolution. However, the resulting data are very complex, and on the bioinformatic side, there still exists an urgent need for improving computer software for (semi-) automated cross-linking data analysis. In this study, we have developed dedicated software for rapid and confident identification and validation of cross-linked species using an isotopic labelled cross-linker approach in combination with MS. Deuterated (+4 Da) and non-deuterated (+0 Da) bis(sulfosuccinimidyl)suberate, BS3, was used as homobifunctional cross-linker to tag the cross-linked regions. Peptides generated from proteolysis were separated using high performance liquid chromatography, and peptide mass fingerprinting was obtained for the individual fractions using matrix-assisted laser-desorption ionisation time-of-flight (MALDI TOF) MS. The resulting peptide mass lists were combined and transferred to the program, ProteinXXX, which generated the theoretical mass values of all combinations of cross-linked peptides and dead-end cross-links and compared this to the obtained mass lists. In addition, screening for 4 Da-separated signals aided the identification of potential cross-linked species. Sequence information of these candidates was then obtained using MALDI TOF TOF. The cross-linked peptides could then be validated based on the match of the fragmentation pattern and the theoretical values produced by ProteinXXX. This semi-automated interpretation provided a high analysis speed of cross-linking data, with efficient and confident identification of cross-linked species. Four experiments using different conditions showed a high degree of reproducibility as only 1 and 2 cross-links out of 36 identified was not observed in two experiments. The method was tested using human placenta calreticulin (CRT). Based on the identified cross-links, a few corrections to a model of calreticulin obtained by homology modelling using calnexin as template can be suggested. Furthermore, the cross-links show that the C-terminal of the protein continues along the core region opposite the P-domain for at least 11 residues beyond the known structure. In addition, it was observed that the conformation of CRT does not change significantly in the presence or absence of the divalent ions, Ca2+ and Zn2+.

  4. Characterization of polyelectrolyte behavior of the polysaccharides chitosan, heparin, and hyaluronan, by light scattering and viscometry.

    NASA Astrophysics Data System (ADS)

    Boddohi, Soheil; Yonemura, Susan; Kipper, Matt

    2008-03-01

    This study on the polyelectrolyte behavior of polysaccharides in solution is motivated by our recent work in development of nanostructured polysaccharide-based surface coatings. Chitosan behaves as a weak polycation, and hyaluronan behaves as a weak polyanion, while heparin behaves as a strong polyanion. The ability to control the conformation of these polysaccharides in solution, by changing the solution ionic strength and pH may offer the opportunity to further tune the nanoscale features of polysaccharide-based surface coatings assembled from solution. In the work reported here, the solution conformation of these polymers is determined from gel permeation chromatography coupled to differential refractive index, light scattering, and viscometry detection. These results are related to the nanostructure of chitosan-heparin and chitosan-hyaluronan surface coatings based on polyelectrolyte multilayers.

  5. Highly efficient copper(II) ion sorbents obtained by calcium carbonate mineralization on functionalized cross-linked copolymers.

    PubMed

    Mihai, Marcela; Bunia, Ion; Doroftei, Florica; Varganici, Cristian-Dragos; Simionescu, Bogdan C

    2015-03-23

    A new type of Cu(II) ion sorbents is presented. These are obtained by CaCO3 mineralization from supersaturated solutions on gel-like cross-linked polymeric beads as insoluble templates. A divinylbenzene-ethylacrylate-acrylonitrile cross-linked copolymer functionalized with weakly acidic, basic, or amphoteric functional groups has been used, as well as different initial inorganic concentrations and addition procedures for CaCO3 crystal growth. The morphology of the new composites was investigated by SEM and compared to that of the unmodified beads, and the polymorph content was established by X-ray diffraction. The beads, before and after CaCO3 mineralization, were tested as sorbents for Cu(II) ions. The newly formed patterns on the bead surface after Cu(II) sorption were observed by SEM, and the elemental distribution on the composites and the chemical structure of crystals after interaction with Cu(II) were investigated by EDAX elemental mapping and by FTIR-ATR spectroscopy, respectively. The sorption capacity increased significantly after CaCO3 crystals growth on the weak anionic bead surface (up to 1041.5 mg Cu(II) /g sample) compared to that of unmodified beads (491.5 mg Cu(II) /g sample). PMID:25675892

  6. The RecQ helicase RECQL5 participates in psoralen-induced interstrand cross-link repair

    PubMed Central

    Bohr, Vilhelm A.

    2013-01-01

    Interstrand cross-links (ICLs) are very severe lesions as they are absolute blocks of replication and transcription. This property of interstrand cross-linking agents has been exploited clinically for the treatment of cancers and other diseases. ICLs are repaired in human cells by specialized DNA repair pathways including components of the nucleotide excision repair pathway, double-strand break repair pathway and the Fanconi anemia pathway. In this report, we identify the role of RECQL5, a member of the RecQ family of helicases, in the repair of ICLs. Using laser-directed confocal microscopy, we demonstrate that RECQL5 is recruited to ICLs formed by trioxalen (a psoralen-derived compound) and ultraviolet irradiation A. Using single-cell gel electrophoresis and proliferation assays, we identify the role of RECQL5 in the repair of ICL lesions. The domain of RECQL5 that recruits to the site of ICL was mapped to the KIX region between amino acids 500 and 650. Inhibition of transcription and of topoisomerases did not affect recruitment, which was inhibited by DNA-intercalating agents, suggesting that the DNA structure itself may be responsible for the recruitment of RECQL5 to the sites of ICLs. PMID:23715498

  7. The effect of cross-linking on latex film formation

    NASA Astrophysics Data System (ADS)

    Mohd. Ghazaly, Hafsah

    The effect of the crosslinker chain lengths in the copolymerization reaction with n-BMA and the formation of the resulting latex films was investigated. Three crosslinkers; ethylene glycol dimethacrylate (EGDMA-198 g/mol), Aliphatic Urethane Acrylate (AUA-3728 g/mol) and a novel Macromonomer Crosslinker (Mac-3480 g/mol) were compared. Copolymerization was achieved via a miniemulsion process. The amount of the crosslinkers employed in the copolymers was 0.2 mol%. When Mac was used as the crosslinker in the latex, high toughness was retained which is comparable to the toughness of the PBMA homopolymer latex films. The P(BMA-co-Mac) films contained 60% gel (crosslinked network) prepared using persulfate (KPS, water-soluble) to >95% gel prepared using azo (AMBN, oil-soluble) initiators. For the P(BMA-co-AUA) copolymer latex films prepared using either of the initiators, no appreciable toughness was recorded at all. For the P(BMA-co-EGDMA) copolymer latex films, a dependence on the type of initiator was obtained. The persulfate initiator gave P(BMA-co-EGDMA) copolymer latex films appreciable toughness while the AMBN initiator gave brittle films. These copolymer latexes have gel contents >95%. In films derived from crosslinked latex particles, the interface or the continuous matrix (continuum) of the film is crucial for the film to be useful mechanically. The toughness in the PBMA homopolymer and P(BMA-co-Mac) copolymer latex films, was due to the long polymer linear chains that can interdiffused and entangled with chains in the adjacent neighboring particles. The network in P(BMA-co-Mac) copolymer latex films, are flexible and are able to deform upon load application such that these films have high extensibility. Lightly crosslinked PBMA polymer chains which are attached to the network chains may exist at the interface of the particles rendering the P(BMA-co-EGDMA) copolymer films the toughness as observed for the persulfate, and not in the AMBN-initiated system. Phase separations were postulated to occur during the synthesis of the copolymers. The phase separation is hypothesized to determine the particle morphology and hence the resulting film morphology and therefore the film properties.

  8. Shell and core cross-linked poly(L-lysine)/poly(acrylic acid) complex micelles.

    PubMed

    Hsieh, Yi-Hsuan; Hsiao, Yung-Tse; Jan, Jeng-Shiung

    2014-12-21

    We report the versatility of polyion complex (PIC) micelles for the preparation of shell and core cross-linked (SCL and CCL) micelles with their surface properties determined by the constituent polymer composition and cross-linking agent. The negatively and positively charged PIC micelles with their molecular structure and properties depending on the mixing weight percentage and polymer molecular weight were first prepared by mixing the negatively and positively charged polyions, poly(acrylic acid) (PAA) and poly(L-lysine) (PLL). The feasibility of preparing SCL micelles was demonstrated by cross-linking the shell of the negatively and positively charged micelles using cystamine and genipin, respectively. The core of the micelles can be cross-linked by silica deposition to stabilize the assemblies. The shell and/or core cross-linked micelles exhibited excellent colloid stability upon changing solution pH. The drug release from the drug-loaded SCL micelles revealed that the controllable permeability of the SCL micelles can be achieved by tuning the cross-linking degree and the SCL micelles exhibited noticeable pH-responsive behavior with accelerated release under acidic conditions. With the versatility of cross-linking strategies, it is possible to prepare a variety of SCL and CCL micelles from PIC micelles. PMID:25357089

  9. Anomalous normal stresses in biopolymer networks with compliant cross-links

    NASA Astrophysics Data System (ADS)

    Chen, P.

    2014-02-01

    Cross-linked biopolymer filament networks that constitute the cytoskeleton in living cells show rich mechanical response including large strain stiffening and negative normal stresses. Here, we have studied these phenomena in the case where the cross-links act as flexible worm-like chain springs. We find that in contrast to the well-studied case of sparse networks with rigid cross-links that display negative normal stresses when subject to shear loading, flexible cross-links give rise to positive normal stresses during early stages of deformation. With increased loading, we observe a transition where the normal stresses become negative. In this regime, the ratio of the normal stresses to the shear stress is found to increase with increase in the density of cross-links, in distinct contrast to the behavior observed for rigid cross-links. We show that these anomalous properties can be understood by considering the way in which the applied load is shared by the extension of the cross-links and the bending and stretching of the filaments.

  10. Characterization of protein cross-links via mass spectrometry and an open-modification search strategy.

    PubMed

    Singh, Pragya; Shaffer, Scott A; Scherl, Alexander; Holman, Carol; Pfuetzner, Richard A; Larson Freeman, Theodore J; Miller, Samuel I; Hernandez, Patricia; Appel, Ron D; Goodlett, David R

    2008-11-15

    Protein-protein interactions are key to function and regulation of many biological pathways. To facilitate characterization of protein-protein interactions using mass spectrometry, a new data acquisition/analysis pipeline was designed. The goal for this pipeline was to provide a generic strategy for identifying cross-linked peptides from single LC/MS/MS data sets, without using specialized cross-linkers or custom-written software. To achieve this, each peptide in the pair of cross-linked peptides was considered to be "post-translationally" modified with an unknown mass at an unknown amino acid. This allowed use of an open-modification search engine, Popitam, to interpret the tandem mass spectra of cross-linked peptides. False positives were reduced and database selectivity increased by acquiring precursors and fragments at high mass accuracy. Additionally, a high-charge-state-driven data acquisition scheme was utilized to enrich data sets for cross-linked peptides. This open-modification search based pipeline was shown to be useful for characterizing both chemical as well as native cross-links in proteins. The pipeline was validated by characterizing the known interactions in the chemically cross-linked CYP2E1-b5 complex. Utility of this method in identifying native cross-links was demonstrated by mapping disulfide bridges in RcsF, an outer membrane lipoprotein involved in Rcs phosphorelay. PMID:18947195

  11. 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 aldehydederived collagen cross-links (HLCCs) and lower levels of lysine aldehydederived 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

  12. DNA cross-linking by intermediates in the mitomycin activation cascade

    SciTech Connect

    Cera, C.; Egbertson, M.; Teng, S.P.; Crothers, D.M.; Danishefsky, S.J. )

    1989-06-27

    The authors have assayed the cross-linking of oligonucleotides containing repeated mitomycin-reactive CpG sites in order to assess the factors that enhance activation of the carbamoyl function at C{sub 10}, yielding efficient mitomycin cross-linking. Drugs studied include mitomycin C (MC), N-methylmitomycin A (NMA), and the aziridinomitosene of NMA (MS). Drugs were reduced both by catalytic hydrogenation and by dithionite. They find that cross-linking by fully reduced NMA can be increased severalfold by addition of either excess dithionite reductant or the oxidant FeCl{sub 3}. Enhancement by FeCl{sub 3} is not seen with MC or MS, but excess dithionite increases cross-linking by all three compounds. They explain the action of Fe{sup 3+} by postulating production of the semiquinone of the monoadduct of mitomycin reacted at the C{sub 1}-position; according to this mechanism, departure of the carbamate from C{sub 10} is more efficient for the semiquinone than for the hydroquinone. However, the results imply that the hydroquinone can also function as a cross-linking agent. Excess dithionite beyond that required for stoichiometric reduction, activates the carbamate 2-3-fold for cross-linking. They find that the fully reduced leucoaziridinomitosene is highly unstable in solution, yet it produces efficient cross-linking. Hence, this compound is highly reactive in DNA alkylation and a good candidate for the role of primary alkylating agent.

  13. 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. PMID:25937394

  14. Proteome-wide profiling of protein assemblies by cross-linking mass spectrometry.

    PubMed

    Liu, Fan; Rijkers, Dirk T S; Post, Harm; Heck, Albert J R

    2015-12-01

    We describe an integrated workflow that robustly identifies cross-links from endogenous protein complexes in human cellular lysates. Our approach is based on the application of mass spectrometry (MS)-cleavable cross-linkers, sequential collision-induced dissociation (CID)-tandem MS (MS/MS) and electron-transfer dissociation (ETD)-MS/MS acquisitions, and a dedicated search engine, XlinkX, which allows rapid cross-link identification against a complete human proteome database. This approach allowed us to detect 2,179 unique cross-links (1,665 intraprotein cross-links at a 5% false discovery rate (FDR) and 514 interprotein cross-links at 1% FDR) in HeLa cell lysates. We validated the confidence of our cross-linking results by using a target-decoy strategy and mapping the observed cross-link distances onto existing high-resolution structures. Our data provided new structural information about many protein assemblies and captured dynamic interactions of the ribosome in contact with different elongation factors. PMID:26414014

  15. Electrospun gelatin nanofibers: a facile cross-linking approach using oxidized sucrose.

    PubMed

    Jalaja, K; James, Nirmala R

    2015-02-01

    Gelatin nanofibers were fabricated via electrospinning with minimal toxicity from solvents and cross-linking agents. Electrospinning was carried out using a solvent system based on water and acetic acid (8:2, v/v). Acetic acid concentration was kept as minimum as possible to reduce the toxic effects. Electrospun gelatin nanofibers were cross-linked with oxidized sucrose. Sucrose was oxidized by periodate oxidation to introduce aldehyde functionality. Cross-linking with oxidized sucrose could be achieved without compromising the nanofibrous architecture. Cross-linked gelatin nanofibers maintained the fibrous morphology even after keeping in contact with aqueous medium. The morphology of the cross-linked nanofibrous mats was examined by scanning electron microscopy (SEM). Oxidized sucrose cross-linked gelatin nanofibers exhibited improved thermal and mechanical properties. The nanofibrous mats were evaluated for cytotoxicity and cell viability using L-929 fibroblast cells. The results confirmed that oxidized sucrose cross-linked gelatin nanofibers were non-cytotoxic towards L-929 cells with good cell viability. PMID:25478965

  16. Genipin cross-linked nanocomposite films for the immobilization of antimicrobial agent.

    PubMed

    Khan, Avik; Salmieri, Stphane; Fraschini, Carole; Bouchard, Jean; Riedl, Bernard; Lacroix, Monique

    2014-09-10

    Cellulose nanocrystal (CNC) reinforced chitosan based antimicrobial films were prepared by immobilizing nisin on the surface of the films. Nanocomposite films containing 18.65 ?g/cm(2) of nisin reduced the count of L. monocytogenes by 6.73 log CFU/g, compared to the control meat samples (8.54 log CFU/g) during storage at 4 C in a Ready-To-Eat (RTE) meat system. Film formulations containing 9.33 ?g/cm(2) of nisin increased the lag phase of L. monocytogenes on meat by more than 21 days, whereas formulations with 18.65 ?g/cm(2) completely inhibited the growth of L. monocytogenes during storage. Genipin was used to cross-link and protect the activity of nisin during storage. Nanocomposite films cross-linked with 0.05% w/v genipin exhibited the highest bioactivity (10.89 ?g/cm(2)) during the storage experiment, as compared to that of the un-cross-linked films (7.23 ?g/cm(2)). Genipin cross-linked films were able to reduce the growth rate of L. monocytogenes on ham samples by 21% as compared to the un-cross-linked films. Spectroscopic analysis confirmed the formation of genipin-nisin-chitosan heterocyclic cross-linked network. Genipin cross-linked films also improved the swelling, water solubility, and mechanical properties of the nanocomposite films. PMID:25140839

  17. Novel antimicrobial superporous cross-linked chitosan/pyromellitimide benzoyl thiourea hydrogels.

    PubMed

    Mohamed, Nadia A; El-Ghany, Nahed A Abd; Fahmy, Mona M

    2016-01-01

    In this work, chitosan (CS) was cross-linked with different amounts of pyromellitimide benzoyl thiourea moieties. The structure of the cross-linked CS was confirmed by elemental analyses, FTIR and (1)H- NMR spectroscopy. The cross-linking process proceeds via reacting of the amino groups of CS with the isothiocyanate groups of the N,N'-bis [4-(isothiocyanate carbonyl)phenyl] pyromellitimide cross-linker. The amount of the cross-linker was varied with respect to CS to produce four new pyromellitimide benzoyl thiourea cross-linked CS (PIBTU-CS) hydrogels designated as PIBTU-CS-1, PIBTU-CS-2, PIBTU-CS-3, and PIBTU-CS-4 of increasing cross-linking degree percent of 11, 22, 44 and 88%, respectively. The scanning electron microscopy observation indicates the extremely porous structure of the hydrogels. XRD results showed that the crystallinity of CS was decreased upon cross-linking. The four hydrogels exhibit a higher antibacterial activity on Bacillus subtilis and Streptococcus pneumoniae as Gram positive bacteria and against Escherichia coli as Gram negative bacteria and higher antifungal activity on Aspergillus fumigatus, Syncephalastrum racemosum and Geotricum candidum than that of the parent CS as shown from their higher inhibition zone diameters and their lower MIC values. The swell ability of the hydrogel as well as their antimicrobial activity increased with increasing cross-linking density. PMID:26388182

  18. Microencapsulation of protein into biodegradable matrix: a smart solution cross-linking technique.

    PubMed

    Bejugam, Naveen K; Gayakwad, Sanjay G; Uddin, Akm N; D'Souza, Martin J

    2013-01-01

    Sustained-release albumin microspheres (MSs) can be obtained by chemically cross-linking albumin. However, a significant challenge is preventing the cross-linking of the active pharmaceutical (protein or small molecule) ingredient (API) with the MS matrix. To prevent cross-linking of the API with the albumin matrix, a smart "solution cross-linking-microencapsulation" method was developed which involves cross-linking albumin solution with glutaraldehyde first, neutralizing any excess glutaraldehyde with sodium bisulphite, followed by the addition of API and finally spray drying. Using lysozyme as model API, MS formulations FL1 and FL2 were prepared and characterized. Physicochemical characterization using FT-IR and bioactivity evaluation indicate that microencapsulated API did not undergo any significant change in its native structure and the bioactivity was preserved during the formulation processing. Preliminary immunogenicity potential of the cross-linked albumin matrix determined by invivo studies did not show any significant increase in antigen-specific serum-IgG levels, implying safety and biocompatibility of the cross-linked albumin matrix. PMID:23098577

  19. Lysyl hydroxylase 2 induces a collagen cross-link switch in tumor stroma.

    PubMed

    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

    2015-03-01

    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

  20. Aliphatic beta-nitro alcohols for non-enzymatic collagen cross-linking of scleral tissue.

    PubMed

    Paik, David C; Wen, Quan; Airiani, Suzanna; Braunstein, Richard E; Trokel, Stephen L

    2008-09-01

    The success of riboflavin photochemical cross-linking of the cornea in treating keratoconus and post-surgical keratectasia has prompted interest in cross-linking scleral tissue with a potential application to stabilize myopic progression. Applying an UVA light source to the sclera is difficult, particularly in the posterior region. An alternate pharmacologic approach to scleral cross-linking may be possible. The present study was undertaken in order to identify nitrite related compounds capable of inducing scleral tissue cross-linking and to gain information regarding the possible chemical mechanisms involved. 8x4 mm strips of porcine and human sclera were incubated in various concentrations of nitrite related agents (1-100mM) at 37 degrees C. pH 7.4 was used for all experiments except those involving NaNO(2). Following a 24-96 h incubation period, the samples were tested for cross-linking effects using thermal shrinkage temperature (T(s)) analysis. Several compounds were studied including NaNO(2), 2-nitroethanol, 2-nitro-1-propanol, 3-nitro-2-pentanol, 2-nitrophenol, 2-nitroethane, 2-aminoethanol, isopentyl nitrite, DPTA/NO, DETA/NO, and urea, a nitrous acid trap. The results indicate that short chain aliphatic beta-nitro alcohols (2-nitroethanol, 2-nitro-1-propanol, and 3-nitro-2-pentanol) are particularly effective cross-linking agents at pH 7.4, showing both time and concentration dependent effects. Furthermore, nitrosation does not appear to induce tissue cross-linking. In conclusion, aliphatic beta-nitro alcohols can cross-link scleral tissue at physiologic pH and temperature. Since beta-nitro alcohols are known to have reasonable toxicity profiles, these agents could find utility as pharmacologic cross-linking agents for scleral thinning disease. PMID:18616942

  1. Riboflavin/UVA Collagen Cross-Linking-Induced Changes in Normal and Keratoconus Corneal Stroma

    PubMed Central

    Hayes, Sally; Boote, Craig; Kamma-Lorger, Christina S.; Rajan, Madhavan S.; Harris, Jonathan; Dooley, Erin; Hawksworth, Nicholas; Hiller, Jennifer; Terill, Nick J.; Hafezi, Farhad; Brahma, Arun K.; Quantock, Andrew J.; Meek, Keith M.

    2011-01-01

    Purpose To determine the effect of Ultraviolet-A collagen cross-linking with hypo-osmolar and iso-osmolar riboflavin solutions on stromal collagen ultrastructure in normal and keratoconus ex vivo human corneas. Methods Using small-angle X-ray scattering, measurements of collagen D-periodicity, fibril diameter and interfibrillar spacing were made at 1 mm intervals across six normal post-mortem corneas (two above physiological hydration (swollen) and four below (unswollen)) and two post-transplant keratoconus corneal buttons (one swollen; one unswollen), before and after hypo-osmolar cross-linking. The same parameters were measured in three other unswollen normal corneas before and after iso-osmolar cross-linking and in three pairs of swollen normal corneas, in which only the left was cross-linked (with iso-osmolar riboflavin). Results Hypo-osmolar cross-linking resulted in an increase in corneal hydration in all corneas. In the keratoconus corneas and unswollen normal corneas, this was accompanied by an increase in collagen interfibrillar spacing (p<0.001); an increase in fibril diameter was also seen in two out of four unswollen normal corneas and one unswollen keratoconus cornea (p<0.001). Iso-osmolar cross-linking resulted in a decrease in tissue hydration in the swollen normal corneas only. Although there was no consistent treatment-induced change in hydration in the unswollen normal samples, iso-osmolar cross-linking of these corneas did result in a compaction of collagen fibrils and a reduced fibril diameter (p<0.001); these changes were not seen in the swollen normal corneas. Collagen D-periodicity was not affected by either treatment. Conclusion The observed structural changes following Ultraviolet-A cross-linking with hypo-osmolar or iso-osmolar riboflavin solutions are more likely a consequence of treatment-induced changes in tissue hydration rather than cross-linking. PMID:21850225

  2. 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.

  3. 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.

  4. Kinetically controlled patterning of highly cross-linked phosphonium photopolymers using simple anion exchange.

    PubMed

    Guterman, Ryan; Gillies, Elizabeth R; Ragogna, Paul J

    2015-05-12

    A phosphonium salt possessing three methacrylate groups has been incorporated into a photopolymeric system to generate highly cross-linked polyelectrolyte networks. Emergent chemical and physical properties in the polymers were observed and attributed to the tandem increase in cross-link density and ion-content upon incorporation of the self-cross-linking cation. Anion-exchange with bis(trifluoromethylsulfonyl)imide or dodecylbenzenesulfonate resulted in significant differences in wettability and ion-exchange behavior. The passivating effects of dodecylbenzenesulfonate were utilized to selectively pattern fluorescein dye into the polymer network, highlighting a new patterning procedure using ionic-bond forming reactions. PMID:25896478

  5. Cross-linked polyvinyl alcohols and oil reservoir permeability control therewith

    SciTech Connect

    Hoskin, D.H.; Shu, P.

    1990-01-30

    This patent describes an improved method of recovering oil under steam flooding conditions from an oil-bearing subterranean formation penetrated by an injection well and a production well, in which an aqueous fluid is injected into the formation through the injection well to displace oil to the production well. The improvement comprising employing, in at least a portion of the aqueous fluid injected into the formation, a cross-linked polymer obtained by cross-linking polyvinyl alcohol with a cross-linking agent which is a mixture of a phenolic component and an aldehyde or a mixture of a naphtholic component and an aldehyde.

  6. Radiation cross-linking in ultra-high molecular weight polyethylene for orthopaedic applications

    PubMed Central

    Oral, Ebru; Muratoglu, Orhun K.

    2007-01-01

    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. PMID:19050735

  7. Cross-linkage and cross-linking of peptidoglycan in Escherichia coli: definition, determination, and implications.

    PubMed

    Driehuis, F; de Jonge, B; Nanninga, N

    1992-03-01

    The glycan chains in peptidoglycan or murein are cross-linked by transpeptidation of the peptide side chains. To assess the fraction of side chains involved in cross-bridges, distinction has been made between cross-linkage and cross-linking. The first expression refers to the situation in unlabeled (or fully labeled) peptidoglycan, and the second refers to pulse-labeled peptidoglycan. It is argued that for the determination of the cross-linking value, the mode of insertion as denoted by the so-called acceptor/donor radioactivity ratio should be taken into account. PMID:1548240

  8. Effector-sensitive cross-linking of phosphorylase b kinase by the novel cross-linker 4-phenyl-1,2,4-triazoline-3,5-dione.

    PubMed Central

    Ayers, N A; Nadeau, O W; Read, M W; Ray, P; Carlson, G M

    1998-01-01

    The dienophile 4-phenyl-1,2,4-triazoline-3,5-dione (PTD) was identified as a novel protein cross-linker, and utilized as a conformational probe of phosphorylase b kinase (PhK), a hexadecameric enzyme with the subunit composition (alphabetagammadelta)4. In its reaction with this enzyme, PTD produced five major cross-linked conjugates as resolved by denaturing gel electrophoresis: alphabeta, betagammagamma, alphagamma and a doublet of differently migrating homodimers, betabeta1 and betabeta2. Cross-linking in the presence of six different activators of the kinase targeted to its various subunits caused substantial changes in the amounts of three of the conjugates. The formation of alphagamma was increased by all of the activators but the largest enhancement was caused by exogenous Ca2+/calmodulin. All except one of the activators decreased the amount of betagammagamma formed, with Mg2+ having the greatest effect, and all except two increased the amount of betabeta1, with Mg2+ again having the largest influence. From the overall similarity of the changes in cross-linking by PTD induced by the various activators, we conclude that, even though they are targeted to different sites and subunits, they induce activated conformations of PhK that have certain structural features in common. Regarding the mechanism of cross-linking by PTD, its reaction with a model nucleophile suggests that its initial reaction with a side chain nucleophile of PhK involves a 1,4-conjugate addition to form a urazole adduct, with the secondary cross-linking reaction occurring through an as yet unknown pathway. PMID:9512471

  9. Production of Cisplatin-Incorporating Hyaluronan Nanogels via Chelating Ligand-Metal Coordination.

    PubMed

    Ohta, Seiichi; Hiramoto, Syota; Amano, Yuki; Sato, Mayu; Suzuki, Yukimitsu; Shinohara, Marie; Emoto, Shigenobu; Yamaguchi, Hironori; Ishigami, Hironori; Sakai, Yasuyuki; Kitayama, Joji; Ito, Taichi

    2016-03-16

    Hyaluronan (HA) is a promising drug carrier for cancer therapy because of its CD44 targeting ability, good biocompatibility, and biodegradability. In this study, cisplatin (CDDP)-incorporating HA nanogels were fabricated through a chelating ligand-metal coordination cross-linking reaction. We conjugated chelating ligands, iminodiacetic acid or malonic acid, to HA and used them as a precursor polymer. By mixing the ligand-conjugated HA with CDDP, cross-linking occurred via coordination of the ligands with the platinum in CDDP, resulting in the spontaneous formation of CDDP-loaded HA nanogels. The nanogels showed pH-responsive release of CDDP, because the stability of the ligand-platinum complex decreases in an acidic environment. Cell viability assays for MKN45P human gastric cancer cells and Met-5A human mesothelial cells revealed that the HA nanogels selectively inhibited the growth of gastric cancer cells. In vivo experiments using a mouse model of peritoneal dissemination of gastric cancer demonstrated that HA nanogels specifically localized in peritoneal nodules after the intraperitoneal administration. Moreover, penetration assays using multicellular tumor spheroids indicated that HA nanogels had a significantly higher ability to penetrate tumors than conventional, linear HA. These results suggest that chelating-ligand conjugated HA nanogels will be useful for targeted cancer therapy. PMID:26781684

  10. Hydrophilic films based on poly(acrylic acid)-poly(vinyl methyl ether) blends cross-linked by gamma-radiation

    NASA Astrophysics Data System (ADS)

    Nurkeeva, Zauresh S.; Mun, Grigoriy A.; Khutoryanskiy, Vitaliy V.; Dzhusupbekova, Aizhana B.

    2004-02-01

    Hydrophilic polymeric films based on blends of poly(acrylic acid) and poly(vinyl methyl ether) (PVME) were prepared by casting technique and were cross-linked by gamma-radiation. The films are soft and elastic in a dry state and form hydrogels upon immersion in water. Effect of absorbed dose on the gel fraction as well as on the swelling of the films in aqueous solutions of different pH is studied. It was found that addition of lower molecular weight PVME decreases the gelation dose, which is likely related to a decrease in glass transition temperature of the blends. In acidic media the films have low swelling degree because of suppression of carboxylic groups ionisation and formation of additional physical cross-links via interpolymer hydrogen bonding.

  11. Psoralen covalently linked to oligodeoxyribonucleotides: synthesis, sequence specific recognition of DNA and photo-cross-linking to pyrimidine residues of DNA.

    PubMed

    Pieles, U; Englisch, U

    1989-01-11

    The psoralen derivative 4,5',8-trimethylpsoralen was covalently linked to the 5'-terminus of an 18mer oligodeoxyribonucleotide in the course of solid phase synthesis using phosphoroamidite chemistry. The derivative was introduced as a phosphitylation compound in the last cycle of the oligomer synthesis. The reagent was prepared by 4'-chloromethylation of 4,5',8-trimethylpsoralen, introduction of a linker by ethanediol and phosphitylation with chloro-[(beta-cyanoethoxy)-N,N-diisopropylamino]-phosphine. After oxydation and deprotection the 5'-psoralen modified oligodeoxyribonucleotide was characterised by HPLC. Hybridisation of the psoralen-modified oligomer to a complementary single stranded 21mer followed by irradiation at 350 nm revealed a photo-cross-linked double-stranded DNA fragment analysed on denaturing polyacrylamide gels. The cross-link could be reversed upon irradiation at 254nm. PMID:2911468

  12. Psoralen covalently linked to oligodeoxyribonucleotides: synthesis, sequence specific recognition of DNA and photo-cross-linking to pyrimidine residues of DNA.

    PubMed Central

    Pieles, U; Englisch, U

    1989-01-01

    The psoralen derivative 4,5',8-trimethylpsoralen was covalently linked to the 5'-terminus of an 18mer oligodeoxyribonucleotide in the course of solid phase synthesis using phosphoroamidite chemistry. The derivative was introduced as a phosphitylation compound in the last cycle of the oligomer synthesis. The reagent was prepared by 4'-chloromethylation of 4,5',8-trimethylpsoralen, introduction of a linker by ethanediol and phosphitylation with chloro-[(beta-cyanoethoxy)-N,N-diisopropylamino]-phosphine. After oxydation and deprotection the 5'-psoralen modified oligodeoxyribonucleotide was characterised by HPLC. Hybridisation of the psoralen-modified oligomer to a complementary single stranded 21mer followed by irradiation at 350 nm revealed a photo-cross-linked double-stranded DNA fragment analysed on denaturing polyacrylamide gels. The cross-link could be reversed upon irradiation at 254nm. Images PMID:2911468

  13. Molybdate sorption by cross-linked chitosan beads: Dynamic studies

    SciTech Connect

    Guibal, E.; Milot, C.; Roussy, J.

    1999-01-01

    Recent trends in environmental monitoring have induced increasing development of new wastewater treatment techniques. Membrane processes, electrochemical techniques, or ion-exchange systems are widely used, but biosorption has been recognized in the last 30 years as a promising way to reduce the contamination of surface water issued from industrial effluent. Chitosan, a biopolymer extracted from crustacean shells, exhibits high sorption capacities for metal ion recovery. Sorption efficiency and removal rates are controlled by several diffusion mechanisms. Chitosan gel beads have been prepared and have shown enhanced sorption performance in batch systems. This study shows that, in continuous systems, sorption capacities can reach 700 mg/g, a level close to that obtained in batch studies. The effects of metal concentration, flow velocity, and column size are investigated and demonstrate that, because of diffusion mechanisms, the optimum concentration range is approximately 50 to 100 mg/L. In column systems, the Biot number, though greater than 1, is lower than the Biot number obtained in batch systems, indicating that external mass transfer influences mass transfer at the low superficial velocity investigated in this work.

  14. Protein cross-linking and oligomerization through dityrosine formation upon exposure to ozone

    NASA Astrophysics Data System (ADS)

    Liu, Fobang; Kampf, Christopher; Reinmuth-Selzle, Kathrin; Berkemeier, Thomas; Shiraiwa, Manabu; Pöschl, Ulrich

    2015-04-01

    Air pollution is a potential factor for the increasing prevalence of allergic diseases. Airborne allergenic proteins can be directly exposed to air pollution promoting post-translational modifications, which can enhance the allergenic potential of proteins. The formation of dimers or oligomers of allergenic proteins has been reported to result in an enhanced allergenicity. However, the oligomerization process for proteins at atmospherically relevant concentration of O3 is still largely unknown. In this study, the kinetics and reaction mechanism of protein oligomerization upon ozone exposure were studied at atmospherically relevant ozone concentrations and relative humidity (RH) in coated-wall flow tube experiments. Bovine Serum Albumin (BSA) was used as a model protein. Protein ozone exposure was studied for different protein phase-states, i.e. amorphous solid (45% RH experiments), semi-solid (96% RH experiments) and liquid (bulk solution experiments) to account for the differences of phase in atmospheric particulates, e.g., aerosol particles and cloud droplets. Product analysis was performed using a size exclusion chromatography-high performance liquid chromatography-diode array detector (SEC-HPLC-DAD). We demonstrate that protein cross-linking upon ozone exposure can be attributed to the formation of covalent intermolecular dityrosine species by gel electrophoretic and fluorescence spectroscopic methods. The exposure experiments indicate that in addition to ozone concentration, the oligomerization process was depending on the phase-state of protein. In liquid-phase experiments, dimer formation was significantly enhanced, thus indicating a potential relevance of in-cloud processes for protein oligomerization. The reactive turnover is higher at 96% RH compared to 45% RH, indicating a higher bulk diffusion coefficient at high RH, which is explicitly resolved by kinetic modeling. Further, the reactive turnover showed a strong correlation to particle surface-to-volume ratio, confirming the bulk diffusion limitation.

  15. Chemosensitivity of primary human fibroblasts with defective unhooking of DNA interstrand cross-links

    SciTech Connect

    Clingen, Peter H. . E-mail: p.clingen@ucl.ac.uk; Arlett, Colin F.; Hartley, John A.; Parris, Christopher N.

    2007-02-15

    Xeroderma pigmentosum (XP) is characterised by defects in nucleotide excision repair, ultraviolet (UV) radiation sensitivity and increased skin carcinoma. Compared to other complementation groups, XP-F patients show relatively mild cutaneous symptoms. DNA interstrand cross-linking agents are a highly cytotoxic class of DNA damage induced by common cancer chemotherapeutics such as cisplatin and nitrogen mustards. Although the XPF-ERCC1 structure-specific endonuclease is required for the repair of ICLs cellular sensitivity of primary human XP-F cells has not been established. In clonogenic survival assays, primary fibroblasts from XP-F patients were moderately sensitive to both UVC and HN2 compared to normal cells (2- to 3-fold and 3- to 5-fold, respectively). XP-A fibroblasts were considerably more sensitive to UVC (10- to 12-fold) but not sensitive to HN2. The sensitivity of XP-F fibroblasts to HN2 correlated with the defective incision or 'unhooking' step of ICL repair. Using the comet assay, XP-F cells exhibited only 20% residual unhooking activity over 24 h. Over the same time, normal and XP-A cells unhooked greater than 95% and 62% of ICLs, respectively. After HN2 treatment, ICL-associated DNA double-strand breaks (DSBs) are detected by pulse field gel electrophoresis in dividing cells. Induction and repair of DNA DSBs was normal in XP-F fibroblasts. These findings demonstrate that in primary human fibroblasts, XPF is required for the unhooking of ICLs and not for the induction or repair of ICL-associated DNA DSBs induced by HN2. In terms of cancer chemotherapy, people with mild DNA repair defects affecting ICL repair may be more prevalent in the general population than expected. Since cellular sensitivity of primary human fibroblasts usually reflects clinical sensitivity such patients with cancer would be at risk of increased toxicity.

  16. 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

  17. Cross-linking proteins with bimetallic tetracarboxylate compounds of transition metals

    DOEpatents

    Kostic, Nenad M.; Chen, Jian

    1991-03-05

    Stable cross-linked complexes of transition-metal tetracarboxylates and proteins are formed. The preferred transition-metal is rhodium. The protein may be collagen or an enzyme such as a proteolytic enzyme.

  18. Cross-linking proteins with bimetallic tetracarboxylate compounds of transition metals

    DOEpatents

    Kostic, N.M.; Chen, J.

    1991-03-05

    Stable cross-linked complexes of transition-metal tetracarboxylates and proteins are formed. The preferred transition-metal is rhodium. The protein may be collagen or an enzyme such as a proteolytic enzyme. No Drawings

  19. Effect of cross-link density on carbon dioxide separation in polydimethylsiloxane-norbornene membranes

    DOE PAGESBeta

    Hong, Tao; Niu, Zhenbin; Hu, Xunxiang; Gmernicki, Kevin; Cheng, Shiwang; Fan, Fei; Johnson, J. Casey; Hong, Eunice; Mahurin, Shannon; Jiang, De -en; et al

    2015-11-02

    The development of high performance materials for CO2 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 (CO2 permeability ~ 6800 Barrer and CO2/N2 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, which provides lightly cross-linked membranes.more » 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

  20. Effect of Cross-Link Density on Carbon Dioxide Separation in Polydimethylsiloxane-Norbornene Membranes.

    PubMed

    Hong, Tao; Niu, Zhenbin; Hu, Xunxiang; Gmernicki, Kevin; Cheng, Shiwang; Fan, Fei; Johnson, J Casey; Hong, Eunice; Mahurin, Shannon; Jiang, De-en; Long, Brian; Mays, Jimmy; Sokolov, Alexei; Saito, Tomonori

    2015-11-01

    The development of high-performance materials for carbon dioxide separation and capture will significantly contribute to a solution for climate change. Herein, (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 (CO2 permeability?6800?Barrer; CO2 /N2 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 for difunctional PDMS macromonomers, which provides lightly cross-linked membranes. By combining positron annihilation lifetime spectroscopy, broadband dielectric spectroscopy, and gas solubility measurements, key parameters necessary for achieving excellent performance have been elucidated. PMID:26482115

  1. Biological relevance and consequences of chemical- or metal-induced DNA cross-linking

    SciTech Connect

    Paustenbach, D.J.; Finley, B.L.

    1996-03-01

    A vast number of chemicals are known to induce mutagenesis and/or carcinogenesis in mammals. Although disruption of cellular nuclear material resulting ultimately in mutagenesis/carcinogenesis can be accomplished by various mechanisms, the search for biomarkers of chemical-induced toxicity continues. This review focuses on the ability of certain metals or chemicals to bind to DNA in a cross-link fashion in whole animal as well as under in vitro conditions. The methodologies currently used to determine DNA cross-linking are described. The biological relevance of the presence of chemical- or metal-induced DNA cross-linking as a measure of carcinogenesis in humans is still under debate, as there is no clear correlation between the disease and the DNA cross-link reaction. 62 refs., 3 tabs.

  2. 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

  3. Inverted bulk-heterojunction solar cell with cross-linked hole-blocking layer.

    PubMed

    Udum, Yasemin; Denk, Patrick; Adam, Getachew; Apaydin, Dogukan H; Nevosad, Andreas; Teichert, Christian; S White, Matthew; S Sariciftci, Niyazi; Scharber, Markus C

    2014-05-01

    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

  4. Effect of cross-link density on carbon dioxide separation in polydimethylsiloxane-norbornene membranes

    SciTech Connect

    Hong, Tao; Niu, Zhenbin; Hu, Xunxiang; Gmernicki, Kevin; Cheng, Shiwang; Fan, Fei; Johnson, J. Casey; Hong, Eunice; Mahurin, Shannon; Jiang, De -en; Long, Brian; Mays, Jimmy; Sokolov, Alexei; Saito, Tomonori

    2015-11-02

    The development of high performance materials for CO2 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 (CO2 permeability ~ 6800 Barrer and CO2/N2 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, which 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.

  5. Cross-Linked Conjugated Polymer Fibrils: Robust Nanowires from Functional Polythiophene Diblock Copolymers

    SciTech Connect

    Hammer, Brenton A. G.; Bokel, Felicia A.; Hayward, Ryan C.; Emrick, Todd

    2011-09-27

    A series of poly(3-hexyl thiophene) (P3HT)-based diblock copolymers were prepared and examined in solution for their assembly into fibrils, and post-assembly cross-linking into robust nanowire structures. P3HT-b-poly(3-methanol thiophene) (P3MT), and P3HT-b-poly(3-aminopropyloxymethyl thiophene) (P3AmT) diblock copolymers were synthesized using Grignard metathesis (GRIM) polymerization. Fibrils formed from solution assembly of these copolymers are thus decorated with hydroxyl and amine functionality, and cross-linking is achieved by reaction of diisocyanates with the hydroxyl and amine groups. A variety of cross-linked structures, characterized by transmission electron microscopy (TEM), were produced by this method, including dense fibrillar sheets, fibril bundles, or predominately individual fibrils, depending on the chosen reaction conditions. In solution, the cross-linked fibrils maintained their characteristic vibronic structure in solvents that would normally disrupt (dissolve) the structures.

  6. 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

  7. A thermodynamic model of physical gels

    NASA Astrophysics Data System (ADS)

    An, Yonghao; Solis, Francisco J.; Jiang, Hanqing

    2010-12-01

    Physical gels are characterized by dynamic cross-links that are constantly created and broken, changing its state between solid and liquid under influence of environmental factors. This restructuring ability of physical gels makes them an important class of materials with many applications, such as in drug delivery. In this article, we present a thermodynamic model for physical gels that considers both the elastic properties of the network and the transient nature of the cross-links. The cross-links' reformation is captured through a connectivity tensor M at the microscopic level. The macroscopic quantities, such as the volume fraction of the monomer ?, number of monomers per cross-link s, and the number of cross-links per volume q, are defined by statistic averaging. A mean-field energy functional for the gel is constructed based on these variables. The equilibrium equations and the stress are obtained at the current state. We study the static thermodynamic properties of physical gels predicted by the model. We discuss the problems of un-constrained swelling and stress driven phase transitions of physical gels and describe the conditions under which these phenomena arise as functions of the bond activation energy Ea, polymer/solvent interaction parameter ?, and external stress p.

  8. Visible-Light Photocatalyzed Cross-Linking of Diacetylene Ligands by Quantum Dots to Improve Their Aqueous Colloidal Stability

    PubMed Central

    2015-01-01

    Ligand cross-linking is known to improve the colloidal stability of nanoparticles, particularly in aqueous solutions. However, most cross-linking is performed chemically, in which it is difficult to limit interparticle cross-linking, unless performed at low concentrations. Photochemical cross-linking is a promising approach but usually requires ultraviolet (UV) light to initiate. Using such high-energy photons can be harmful to systems in which the ligand–nanoparticle bond is fairly weak, as is the case for the commonly used semiconductor quantum dots (QDs). Here, we introduce a novel approach to cross-link thiolated ligands on QDs by utilizing the photocatalytic activity of QDs upon absorbing visible light. We show that using visible light leads to better ligand cross-linking by avoiding the problem of ligand dissociation that occurs upon UV light exposure. Once cross-linked, the ligands significantly enhance the colloidal stability of those same QDs that facilitated cross-linking. PMID:25036275

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

    SciTech Connect

    Kruppa, Gary Hermann; Young, Malin M.; Novak, Petr; Schoeniger, Joseph S.

    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 the 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 information can be obtained by exploiting new cross-linker chemistry, increasing the probability that the protein target of choice will yield sufficient distance constraints to develop a structural model.

  10. A novel fibre-ensemble level constitutive model for exogenous cross-linked collagenous tissues.

    PubMed

    Sacks, Michael S; Zhang, Will; Wognum, Silvia

    2016-02-01

    Exogenous cross-linking of soft collagenous tissues is a common method for biomaterial development and medical therapies. To enable improved applications through computational methods, physically realistic constitutive models are required. Yet, despite decades of research, development and clinical use, no such model exists. In this study, we develop the first rigorous full structural model (i.e. explicitly incorporating various features of the collagen fibre architecture) for exogenously cross-linked soft tissues. This was made possible, in-part, with the use of native to cross-linked matched experimental datasets and an extension to the collagenous structural constitutive model so that the uncross-linked collagen fibre responses could be mapped to the cross-linked configuration. This allowed us to separate the effects of cross-linking from kinematic changes induced in the cross-linking process, which in turn allowed the non-fibrous tissue matrix component and the interaction effects to be identified. It was determined that the matrix could be modelled as an isotropic material using a modified Yeoh model. The most novel findings of this study were that: (i) the effective collagen fibre modulus was unaffected by cross-linking and (ii) fibre-ensemble interactions played a large role in stress development, often dominating the total tissue response (depending on the stress component and loading path considered). An important utility of the present model is its ability to separate the effects of exogenous cross-linking on the fibres from changes due to the matrix. Applications of this approach include the utilization in the design of novel chemical treatments to produce specific mechanical responses and the study of fatigue damage in bioprosthetic heart valve biomaterials. PMID:26855761

  11. Computational exploration of polymer nanocomposite mechanical property modification via cross-linking topology

    SciTech Connect

    Lacevic, N; Gee, R; Saab, A; Maxwell, R

    2008-04-24

    Molecular dynamics simulations have been performed in order to study the effects of nanoscale filler cross-linking topologies and loading levels on the mechanical properties of a model elastomeric nanocomposite. The model system considered here is constructed from octa-functional polyhedral oligomeric silsesquioxane (POSS) dispersed in a poly(dimethylsiloxane) (PDMS) matrix. Shear moduli, G, have been computed for pure and for filled and unfilled PDMS as a function of cross-linking density, POSS fill loading level, and polymer network topology. The results reported here show that G increases as the cross-linking (covalent bonds formed between the POSS and the PDMS network) density increases. Further, G is found to have a strong dependence on cross-linking topology. The increase in shear modulus, G, for POSS filled PDMS is significantly higher than that for unfilled PDMS cross-linked with standard molecular species, suggesting an enhanced reinforcement mechanism for POSS. In contrast, in blended systems (POSS/PDMS mixture with no cross-linking) G was not observed to significantly increase with POSS loading. Finally, we find intriguing differences in the structural arrangement of bond strains between the cross-linked and the blended systems. In the unfilled PDMS the distribution of highly strained bonds appears to be random, while in the POSS filled system, the strained bonds form a net-like distribution that spans the network. Such a distribution may form a structural network 'holding' the composite together and resulting in increases in G compared to an unfilled, cross-linked system. These results are of importance for engineering of new POSS-based multifunctional materials with tailor-made mechanical properties.

  12. Ruthenium-catalyzed photo cross-linking of fibrin-based engineered tissue.

    PubMed

    Bjork, Jason W; Johnson, Sandra L; Tranquillo, Robert T

    2011-04-01

    Most cross-linking methods utilize chemistry or physical processes that are detrimental to cells and tissue development. Those that are not as harmful often do not provide a level of strength that ultimately meets the required application. The purpose of this work was to investigate the use of a ruthenium-sodium persulfate cross-linking system to form dityrosine in fibrin-based engineered tissue. By utilizing the tyrosine residues inherent to fibrin and cell-deposited proteins, at least 3-fold mechanical strength increases and 10-fold stiffness increases were achieved after cross-linking. This strengthening and stiffening effect was found to increase with culture duration prior to cross-linking such that physiologically relevant properties were obtained. Fibrin was not required for this effect as demonstrated by testing with collagen-based engineered tissue. Cross-linked tissues were implanted subcutaneously and shown to have minimal inflammation after 30 days, similar to non-cross-linked controls. Overall, the method employed is rapid, non-toxic, minimally inflammatory, and is capable of increasing strength and stiffness of engineered tissues to physiological levels. PMID:21196047

  13. Enabling Thermoreversible Physically Cross-Linked Polymerized Colloidal Array Photonic Crystals

    PubMed Central

    Asher, Sanford A.; Kimble, Kyle W.; Walker, Jeremy P.

    2009-01-01

    We physically cross-linked a thermoreversible poly(vinyl alcohol) (PVA) hydrogel (TG) within a crystalline colloidal array (CCA) to form an enabling photonic crystal material. The TG consists of a physically cross-linked network formed in a process reminiscent of the well-known freeze-thaw physically cross-linking process, but which avoids solvent freezing which invariably disorders the CCA. These TGCCA can be inexpensively fabricated in any large volume and shape by avoiding the previous covalently polymerized CCA constraints that required thin sheet geometries to enable penetration of the UV light used to photopolymerize the system. This TG hydrogel enables rigidificaton of CCA crystals and subsequent chemical functionalization. In addition, an additional interpenetrating hydrogel can be polymerized within the TGPCCA. The TG can then be dissolved away by simply increasing the temperature. The TGCCA photonic crystal diffraction is highly efficient and similar to previously demonstrated PCCA with covalent cross-links. These TGCCA are stable for weeks or longer at room temperature and can be utilized as photonic crystal materials. They also can be irreversibly covalently cross-linked by using gluteraldehyde. These gluteraldehyde cross-linked TGCCA can be made into chemically responsive sensor photonic crystals by functionalizing the PVA hydroxyl groups with chemical recognition agents. We demonstrate low and high pH sensing by functionalizing with carboxylates and phenol derivatives, respectively. PMID:19966904

  14. Corneal melting after collagen cross-linking for keratoconus: a case report

    PubMed Central

    2011-01-01

    Introduction Corneal collagen cross-linking is a rather new technique that uses riboflavin and ultraviolet A light for collagen fiber stabilization in keratoconus corneas. Other than reversible side effects, the preliminary results of corneal collagen cross-linking studies suggest that it is a rather safe technique. In this report, we demonstrate a case of corneal melting after corneal collagen cross-linking for keratoconus corneas associated with an acute inflammatory response. Case presentation A 23-year-old Caucasian man with keratoconus cornea stage 1 to 2 underwent uneventful corneal collagen cross-linking treatment according to the Dresden protocol. The next day the patient had intense photophobia, watering and redness of the eye, and his visual acuity was limited to counting fingers. Slit lamp biomicroscopy revealed severe corneal haze accompanied by non-specific endothelial precipitates following an acute inflammatory response. Mild inflammation could be detected in the anterior chamber. Moreover, the re-epithelialization process could barely be detected. His corneal state gradually deteriorated, resulting in descemetocele and finally perforation. Conclusion In this report, we present a case of a patient with corneal melting after standard corneal collagen cross-linking treatment for keratoconus corneas following an acute inflammatory response. Despite modifying postoperative treatment, elaboration of all apparent associated causes by the treating physicians and undergoing extensive laboratory testing, the patient developed descemetocele, which led to perforation. Our report suggests that further research is necessary regarding the safety of corneal collagen cross-linking in keratoconus corneas. PMID:21496288

  15. Bifunctional electrophiles cross-link thioredoxins with redox relay partners in cells.

    PubMed

    Naticchia, Matthew R; Brown, Haley A; Garcia, Francisco J; Lamade, Andrew M; Justice, Samantha L; Herrin, Rachelle P; Morano, Kevin A; West, James D

    2013-03-18

    Thioredoxin protects cells against oxidative damage by reducing disulfide bonds in improperly oxidized proteins. Previously, we found that the baker's yeast cytosolic thioredoxin Trx2 undergoes cross-linking to form several protein-protein complexes in cells treated with the bifunctional electrophile divinyl sulfone (DVSF). Here, we report that the peroxiredoxin Tsa1 and the thioredoxin reductase Trr1, both of which function in a redox relay network with thioredoxin, become cross-linked in complexes with Trx2 upon DVSF treatment. Treatment of yeast with other bifunctional electrophiles, including diethyl acetylenedicarboxylate (DAD), mechlorethamine (HN2), and 1,2,3,4-diepoxybutane (DEB), resulted in the formation of similar cross-linked complexes. Cross-linking of Trx2 and Tsa1 to other proteins by DVSF and DAD is dependent on modification of the active site Cys residues within these proteins. In addition, the human cytosolic thioredoxin, cytosolic thioredoxin reductase, and peroxiredoxin 2 form cross-linked complexes to other proteins in the presence of DVSF, although each protein shows different susceptibilities to modification by DAD, HN2, and DEB. Taken together, our results indicate that bifunctional electrophiles potentially disrupt redox homeostasis in yeast and human cells by forming cross-linked complexes between thioredoxins and their redox partners. PMID:23414292

  16. Bifunctional Electrophiles Cross-Link Thioredoxins with Redox Relay Partners in Cells

    PubMed Central

    Naticchia, Matthew R.; Brown, Haley A.; Garcia, Francisco J.; Lamade, Andrew M.; Justice, Samantha L.; Herrin, Rachelle P.; Morano, Kevin A.; West, James D.

    2013-01-01

    Thioredoxin protects cells against oxidative damage by reducing disulfide bonds in improperly oxidized proteins. Previously, we found that the baker's yeast cytosolic thioredoxin Trx2 undergoes cross-linking to form several protein-protein complexes in cells treated with the bifunctional electrophile divinyl sulfone (DVSF). Here, we report that the peroxiredoxin Tsa1 and the thioredoxin reductase Trr1, both of which function in a redox relay network with thioredoxin, become cross-linked in complexes with Trx2 upon DVSF treatment. Treatment of yeast with other bifunctional electrophiles, including diethyl acetylenedicarboxylate (DAD), mechlorethamine (HN2), and 1,2,3,4-diepoxybutane (DEB), resulted in the formation of similar cross-linked complexes. Cross-linking of Trx2 and Tsa1 to other proteins by DVSF and DAD is dependent on modification of the active site Cys residues within these proteins. In addition, the human cytosolic thioredoxin, cytosolic thioredoxin reductase, and peroxiredoxin 2 form cross-linked complexes to other proteins in the presence of DVSF, although each protein shows different susceptibilities to modification by DAD, HN2, and DEB. Taken together, our results indicate that bifunctional electrophiles potentially disrupt redox homeostasis in yeast and human cells by forming cross-linked complexes between thioredoxins and their redox partners. PMID:23414292

  17. SiRNA delivery systems based on neutral cross-linked dendrimers.

    PubMed

    Liu, Jie; Zhou, Jihan; Luo, Ying

    2012-02-15

    A neutral cross-linked dendritic system is described for use of delivering small interfering RNA (siRNA) for targeted gene silencing. By replacing the terminal amines with hydrazide groups and N-acetylgalactosamine (GalNAc) ligands, cationic polyamidoamine (PAMAM) dendrimers were transformed into neutral glycosylated carriers for siRNA delivery. Mainly owing to the pH sensitivity and the proton-absorption capability of the tertiary amines within the interior branches, these PAMAM derivatives showing neutrality under physiological conditions (pH 7.4) formed complexes with siRNA through electrostatic interactions at pH 5. Cross-linking procedures via reactions with glutaraldehyde were established, and cytocompatible cross-linked systems loaded with siRNA obtained, with the particulate properties variable with the cross-linking condition and the GalNAc level in the dendritic carrier. In vitro cellular uptake and RNAi experiments showed that the cross-linked dendritic systems with an appropriate level of GalNAc composition effectively interacted with HepG2 cells and inhibited the expression of a luciferase reporter gene. Neutral cross-linked dendritic systems provide a new paradigm for designing siRNA delivery systems with biocompatibility and targeting capability. PMID:22292572

  18. Enhanced Cross-Linking of Diazirine-Modified Sialylated Glycoproteins Enabled through Profiling of Sialidase Specificities.

    PubMed

    McCombs, Janet E; Zou, Chunxia; Parker, Randy B; Cairo, Christopher W; Kohler, Jennifer J

    2016-01-15

    Sialic-acid-mediated interactions play critical roles on the cell surface, providing an impetus for the development of methods to study this important monosaccharide. In particular, photo-cross-linking sialic acids incorporated onto cell surfaces have allowed covalent capture of transient interactions between sialic acids and sialic-acid-recognizing proteins via cross-linking. However, natural sialic acids also present on the cell surface compete with photo-cross-linking sialic acids in binding events, limiting cross-linking yields. In order to improve the utility of one such photo-cross-linking sialic acid, SiaDAz, we examined a number of sialidases, enzymes that remove sialic acids from glycoconjugates, to find one that would cleave natural sialic acids but remain inactive toward SiaDAz. Using this sialidase, we improved SiaDAz-mediated cross-linking of an antisialyl Lewis X antibody and of endoglin. This protocol can be applied generally to sialic-acid-mediated interactions and will facilitate identification of sialic acid binding partners. PMID:26541974

  19. Cross-linked chitosan improves the mechanical properties of calcium phosphate-chitosan cement.

    PubMed

    Aryaei, Ashkan; Liu, Jason; Jayatissa, Ahalapitiya H; Champa Jayasuriya, A

    2015-09-01

    Calcium phosphate (CaP) cements are highly applicable and valuable materials for filling bone defects by minimally invasive procedures. The chitosan (CS) biopolymer is also considered as one of the promising biomaterial candidates in bone tissue engineering. In the present study, some key features of CaP-CS were significantly improved by developing a novel CaP-CS composite. For this purpose, CS was the first cross-linked with tripolyphosphate (TPP) and then mixed with CaP matrix. A group of CaP-CS samples without cross-linking was also prepared. Samples were fabricated and tested based on the known standards. Additionally, the effect of different powder (P) to liquid (L) ratios was also investigated. Both cross-linked and uncross-linked CaP-CS samples showed excellent washout resistance. The most significant effects were observed on Young's modulus and compressive strength in wet condition as well as surface hardness. In dry conditions, the Young's modulus of cross-linked samples was slightly improved. Based on the presented results, cross-linking does not have a significant effect on porosity. As expected, by increasing the P/L ratio of a sample, ductility and injectability were decreased. However, in the most cases, mechanical properties were enhanced. The results have shown that cross-linking can improve the mechanical properties of CaP-CS and hence it can be used for bone tissue engineering applications. PMID:26046262

  20. Dimethyl suberimidate cross-linked pericardium tissue: Raman spectroscopic and atomic force microscopy investigations

    NASA Astrophysics Data System (ADS)

    Jastrzebska, Maria; Zalewska-Rejdak, Justyna; Wrzalik, Roman; Kocot, Antoni; Barwi?ski, Bogdan; Mrz, Iwona; Cwalina, Beata

    2005-06-01

    Chemically stabilized pericardium tissue is widely used as a tissue-derived biomaterial for the preparation of bioprostheses such as heart valves or vascular grafts. The bifunctional imidoester dimethyl suberimidate (DMS) belongs to the wide class of the cross-linking reagents and is often used to cross-link a variety of proteins, including collagen matrices and collagen-based tissues. Raman spectroscopy in the wide frequency range 200-4000 cm -1 and contact mode atomic force microscopy (AFM) have been employed to investigate the structural changes and chemical bonds in DMS cross-linked porcine pericardium tissue. It has been found, that in addition to the commonly accepted reaction with the ?-amine groups of lysine or hydroxylysine residues, DMS may interact also with the carbonyl CO and amide NH groups of the peptide bond in collagen. Our paper presents for the first time spectral evidence for the peptide contribution to the formation of DMS-collagen cross-links. The results confirm also possible competition between the hydrolysis of the free imidoester group and cross-linking reaction. Products of the partial alkaline hydrolysis of DMS have been found in the spectra. The observed changes in the surface topography of the fibrils as well as in their spatial organization in the tissue support the formation of both intra- and interfibrillar cross-links in DMS-stabilized tissue.

  1. Antifouling coatings based on covalently cross-linked agarose film via thermal azide-alkyne cycloaddition.

    PubMed

    Xu, Li Qun; Pranantyo, Dicky; Neoh, Koon-Gee; Kang, En-Tang; Teo, Serena Lay-Ming; Fu, Guo Dong

    2016-05-01

    Coatings based on thin films of agarose-poly(ethylene glycol) (Agr-PEG) cross-linked systems are developed as environmentally-friendly and fouling-resistant marine coatings. The Agr-PEG cross-linked systems were prepared via thermal azide-alkyne cycloaddition (AAC) using azido-functionalized Agr (AgrAz) and activated alkynyl-containing poly(2-propiolamidoethyl methacrylate-co-poly(ethylene glycol)methyl ether methacrylate) P(PEMA-co-PEGMEMA) random copolymers as the precursors. The Agr-PEG cross-linked systems were further deposited onto a SS surface, pre-functionalized with an alkynyl-containing biomimetic anchor, dopamine propiolamide, to form a thin film after thermal treatment. The thin film-coated SS surfaces can effectively reduce the adhesion of marine algae and the settlement of barnacle cyprids. Upon covalent cross-linking, the covalently cross-linked Agr-PEG films coated SS surfaces exhibit good stability in flowing artificial seawater, and enhanced resistance to the settlement of barnacle cyprids, in comparison to that of the surfaces coated with physically cross-linked AgrAz films. PMID:26836479

  2. Nanopod Formation through Gold Nanoparticle Templated and Catalyzed Cross-linking of Polymers Bearing Pendant Propargyl Ethers

    SciTech Connect

    Zhang, Ke; Cutler, Joshua I.; Zhang, Jian A.; Zheng, Dan; Auyeung, Evelyn; Mirkin, Chad A.

    2010-11-03

    A novel method for synthesizing polymer nanopods from a linear polymer bearing pendant propargyl ether groups, using gold nanoparticles as both the template and the catalyst for the cross-linking reaction, is reported. The transformations involved in the cross-linking process are unprecedented on the surface of a gold particle. A tentative cross-linking mechanism is proposed.

  3. Structural Investigation of Supertough Polymer Gels by Small-Angle Neutron Scattering Measurement

    NASA Astrophysics Data System (ADS)

    Shibayama, Mitsuhiro

    2009-04-01

    Various types of supertough polymer gels capable of high recoverable deformability and/or with high shear/compressive moduli are investigated by small-angle neutron scattering (SANS) and mechanical measurements. These gels have unique structures, such as sliding cross-links (slide-ring gels), plane cross-links by clay platelets (nanocomposite gels; NC gels), and tetrahedral networks (tetra-PEG gels). One of the common features of these gels is that frozen inhomogeneities inherent to polymer gels are negligible. This fact, observed by SANS measurements, indicates that cross-links are introduced very effectively. Hence, the mechanical properties of these gels are expected to be well predicted by the theories of rubber elasticity. In this review, we discuss the relationship between the structure and mechanical properties of supertough polymer gels. Furthermore, we address the necessary conditions for high-performance polymer gels in the analogy of rubber.

  4. Mass spectrometry evidence for cisplatin as a protein cross-linking reagent

    PubMed Central

    Li, Huilin; Zhao, Yao; Phillips, Hazel I. A.; Qi, Yulin; Lin, Tzu-Yung; Sadler, Peter J.; OConnor, Peter B.

    2011-01-01

    Cisplatin is a potent anti-cancer drug, which functions by cross-linking adjacent DNA guanine residues. However within one day of injection, 65~98% of the platinum in the blood plasma is protein-bound. It is generally accepted that cisplatin binds to methionine and histidine residues, but what is often underappreciated is that platinum from cisplatin has a 2+ charge and can form up to four bonds. Thus, it has the potential to function as a cross-linker. In this report, the cross-linking ability of cisplatin is demonstrated by Fourier transform ion cyclotron resonance (FTICR) mass spectrometry (MS) with the use of standard peptides, the 16.8 kDa protein calmodulin (CaM), but was unsuccessful for the 64 kDa protein hemoglobin. The high resolution and mass accuracy of FTICR MS along with the high degree of fragmentation of large peptides afforded by collisionally activated dissociation (CAD) and electron capture dissociation (ECD) are shown to be a valuable means of characterizing cross-linking sites. Cisplatin is different from current cross-linking reagents by targeting new functional groups, thioethers, and imidazoles groups, which provides complementarity with existing cross-linkers. In addition, platinum(II) inherently has two positive charges which enhance the detection of cross-linked products. Higher charge states not only promote the detection of cross-linking products with less purification, but result in more comprehensive MS/MS fragmentation and can assist the assignment of modification sites. Moreover, the unique isotopic pattern of platinum flags cross-linking products and modification sites by mass spectrometry. PMID:21591778

  5. Pharmacologic Alternatives to Riboflavin Photochemical Corneal Cross-Linking: A Comparison Study of Cell Toxicity Thresholds

    PubMed Central

    Kim, MiJung; Takaoka, Anna; Hoang, Quan V.; Trokel, Stephen L.; Paik, David C.

    2014-01-01

    Purpose. The efficacy of therapeutic cross-linking of the cornea using riboflavin photochemistry (commonly abbreviated as CXL) has caused its use to become widespread. Because there are known chemical agents that cross-link collagenous tissues, it may be possible to cross-link tissue pharmacologically. The present study was undertaken to compare the cell toxicity of such agents. Methods. Nine topical cross-linking agents (five nitroalcohols, glyceraldehyde [GLYC], genipin [GP], paraformaldehyde [FA], and glutaraldehyde [GLUT]) were tested with four different cell lines (immortalized human corneal epithelial cells, human skin fibroblasts, primary bovine corneal endothelial cells, and immortalized human retinal pigment epithelial cells [ARPE-19]). The cells were grown in planar culture and exposed to each agent in a range of concentrations (0.001 mM to 10 mM) for 24 hours followed by a 48-hour recovery phase. Toxicity thresholds were determined by using the trypan blue exclusion method. Results. A semiquantitative analysis using five categories of toxicity/fixation was carried out, based on plate attachment, uptake of trypan blue stain, and cellular fixation. The toxicity levels varied by a factor of 103 with the least toxic being mononitroalcohols and GLYC, intermediate toxicity for a nitrodiol and nitrotriol, and the most toxic being GLUT, FA, GP, and bronopol, a brominated nitrodiol. When comparing toxicity between different cell lines, the levels were generally in agreement. Conclusions. There are significant differences in cell toxicity among potential topical cross-linking compounds. The balance between cross-linking of tissue and cell toxicity should be borne in mind as compounds and strategies to improve mechanical tissue properties through therapeutic tissue cross-linking continue to develop. PMID:24722697

  6. A comparative cross-linking strategy to probe conformational changes in protein complexes

    PubMed Central

    Schmidt, Carla; Robinson, Carol V.

    2014-01-01

    Chemical cross-linking, together with mass spectrometry, is established as a powerful combination for probing subunit interactions within static protein assemblies. To probe conformational changes in response to stimuli we have developed a comparative cross-linking strategy, employing lysine-specific deuterated and non-deuterated BS3 cross-linking reagents. Here we describe the experimental procedures as well as the data analysis, validation and interpretation. Briefly the protocol involves first assigning cross-linked peptides in the complex without ligand binding, or with post-translational modifications (PTMs) at natural abundance, using a standard procedure employing labeled cross-linkers, proteolysis and subsequent assignment of cross-linked peptides following LC-MS/MS and database searching. An aliquot of the protein complex is then exposed to the stimulus, either ligand binding, or incubation with a phosphatase or kinase to bring about changes in PTMs. Two solutions, one containing the apo/untreated complex the other the enzymatically modified/ligand-bound complex, are then cross-linked independently. Typically non-deuterated BS3-d0 is used or the untreated complex and deuterated BS3-d4 is used for the experiment. The two aliquots are then incubated at equal concentration, digested and processed as before. The ratios of labeled and unlabeled cross-linked peptides then provide a direct readout of the effect of the stimulus. We exemplify our method by quantifying changes in subunit interactions induced by dephosphorylation of an ATP synthase. The protocol is however widely applicable for determining conformational changes in protein complexes induced by various stimuli including ligand/drug binding, oligomerization and other PTMs. Application of the established protocol takes ~9 days, including protein complex purification. PMID:25144272

  7. Conformational Transitions of the Cross-linking Domains of Elastin during Self-assembly*

    PubMed Central

    Reichheld, Sean E.; Muiznieks, Lisa D.; Stahl, Richard; Simonetti, Karen; Sharpe, Simon; Keeley, Fred W.

    2014-01-01

    Elastin is the intrinsically disordered polymeric protein imparting the exceptional properties of extension and elastic recoil to the extracellular matrix of most vertebrates. The monomeric precursor of elastin, tropoelastin, as well as polypeptides containing smaller subsets of the tropoelastin sequence, can self-assemble through a colloidal phase separation process called coacervation. Present understanding suggests that self-assembly is promoted by association of hydrophobic domains contained within the tropoelastin sequence, whereas polymerization is achieved by covalent joining of lysine side chains within distinct alanine-rich, ?-helical cross-linking domains. In this study, model elastin polypeptides were used to determine the structure of cross-linking domains during the assembly process and the effect of sequence alterations in these domains on assembly and structure. CD temperature melts indicated that partial ?-helical structure in cross-linking domains at lower temperatures was absent at physiological temperature. Solid-state NMR demonstrated that ?-strand structure of the cross-linking domains dominated in the coacervate state, although ?-helix was predominant after subsequent cross-linking of lysine side chains with genipin. Mutation of lysine residues to hydrophobic amino acids, tyrosine or alanine, leads to increased propensity for ?-structure and the formation of amyloid-like fibrils, characterized by thioflavin-T binding and transmission electron microscopy. These findings indicate that cross-linking domains are structurally labile during assembly, adapting to changes in their environment and aggregated state. Furthermore, the sequence of cross-linking domains has a dramatic effect on self-assembly properties of elastin-like polypeptides, and the presence of lysine residues in these domains may serve to prevent inappropriate ordered aggregation. PMID:24550393

  8. Cross-linking of wheat gluten proteins during production of hard pretzels.

    PubMed

    Rombouts, Ine; Lagrain, Bert; Brijs, Kristof; Delcour, Jan A

    2012-06-01

    The impact of the hot alkaline dip, prior to pretzel-baking, on the types and levels of cross-links between wheat proteins was studied. Protein extractability of pretzel dough in sodium dodecyl sulfate containing buffer decreased during alkaline dipping [45s, 1.0% (w/v) NaOH, 90C], and even more during baking (3min at 250C) and drying (10min at 135C). Reducing agent increased the extractability partly, indicating that both reducible (disulfide, SS) and non-reducible (non-SS) protein cross-links had been formed. The decrease in cystine levels suggested ?-elimination of cystine releasing Cys and dehydroalanine (DHA). Subsequent reaction of DHA with Lys and Cys, induced the unusual and potentially cross-linking amino acids lysinoalanine (LAL) and lanthionine (LAN), respectively, in alkaline dipped dough (7?mol LAN/g protein) and in the end product (9?mol LAL and 50?mol LAN/g protein). The baking/drying step increased sample redness, decreased Lys levels more than expected based on LAL formation (57?mol/g protein), and induced a loss of reducing sugars (99?mol/g protein), which suggested the potential contribution of Maillard-derived cross-links to the observed extractability loss. However, levels of Maillard products which possibly cross-link proteins, are small compared to DHA-derived cross-links. Higher dipping temperatures, longer dipping times, and higher NaOH concentrations increased protein extractability losses and redness, as well as LAL and LAN levels in the end product. No indications for Maillard-derived cross-links or LAL in pretzel dough immediately after dipping were found, even when severe dipping conditions were used. PMID:21822731

  9. Cross-linking methionine and amine residues with reactive halogen species.

    PubMed

    Ronsein, Graziella E; Winterbourn, Christine C; Di Mascio, Paolo; Kettle, Anthony J

    2014-05-01

    Irreversible cross-links are increasingly being recognized as important posttranslational oxidative protein modifications that contribute to tissue injury during oxidative stress and inflammation. They also have a structural function in extracellular matrix proteins such as collagen IV. Likely contenders for forming such cross-links are the reactive halogen species that are generated by neutrophils and eosinophils, including hypochlorous acid, hypobromous acid, and their related haloamines. Methionine residues are kinetically preferred targets for these oxidants and oxidation can potentially result in sulfilimine (>S=N-) bonds with amines. Therefore, we investigated whether oxidation of methionine in the model peptide formyl-Met-Leu-Phe-Lys (fMLFK) produces cross-links with lysine residues, using mass spectrometry to characterize the products. As expected, the sulfoxide was the major product with each reactive halogen species. However, intra- and intermolecular cross-linked products were also formed. Isomers of an intramolecular sulfilimine were readily produced by hypobromous acid and bromamines, with hypochlorous acid forming lesser amounts. The predominant cross-link with chloramines was an intermolecular bond between the sulfur of fMLFK and the amine derived from the chloramine. Reactive halogen species also formed these sulfilimine cross-links in other peptides that contain methionine. We propose that protein cross-links involving methionine and amine residues will form via this mechanism when granulocytes are activated at sites of inflammation. Our results also support the proposal that reactive halogen species generated by the peroxidase peroxidasin could be responsible for the sulfilimine bonds that are integral to the structure of collagen IV. PMID:24486343

  10. Characterization of the Enzymatic Activity of the Actin Cross-Linking Domain from the Vibrio cholerae MARTXVc Toxin

    PubMed Central

    Kudryashov, Dmitri S.; Cordero, Christina L.; Reisler, Emil; Fullner Satchell, Karla J.

    2008-01-01

    Vibrio cholerae is a Gram-negative bacterial pathogen that exports enterotoxins which alter host cells through a number of mechanisms resulting in diarrheal disease. Among the secreted toxins is the multifunctional, autoprocessing RTX toxin (MARTXVc), which disrupts actin cytoskeleton by covalently cross-linking actin monomers into oligomers. The region of the toxin responsible for cross-linking activity is the actin cross-linking domain (ACD). In this study, we demonstrate unambiguously that ACD utilizes G- and not F-actin as a substrate for the cross-linking reaction and hydrolyzes one molecule of ATP per cross-linking event. Furthermore, major actin binding proteins that regulate actin cytoskeleton in vivo do not block the cross-linking reaction in vitro. Cofilin inhibits the cross-linking of G- and F-actin at high mole ratio to actin, but accelerates F-actin cross-linking at low mole ratios. DNase I blocks completely the cross-linking of actin, likely due to steric hindrance with one of the cross-linking sites on actin. In the context of the holotoxin, the inhibition of Rho by the Rho-inactivating domain of MARTXVc (Sheahan, K.L., Satchell, K.J.F. 2007 Cellular Microbiology 9:1324-1335) would accelerate F-actin depolymerization and provide G-actin, alone or in complex with actin binding proteins, for cross-linking by ACD, ultimately leading to the observed rapid cell rounding. PMID:17951576

  11. Vitamin E-diffused highly cross-linked UHMWPE particles induce less osteolysis compared to highly cross-linked virgin UHMWPE particles in vivo.

    PubMed

    Bichara, David A; Malchau, Erik; Sillesen, Nanna H; Cakmak, Selami; Nielsen, G Petur; Muratoglu, Orhun K

    2014-09-01

    Recent in vitro findings suggest that UHMWPE wear particles containing vitamin E (VE) may have reduced biologic activity and decreased osteolytic potential. We hypothesized that particles from VE-stabilized, radiation cross-linked UHMWPE would cause less osteolysis in a murine calvarial bone model when compared to virgin gamma irradiated cross-linked UHMWPE. Groups received equal amount of particulate debris overlaying the calvarium for 10 days. Calvarial bone was examined using high resolution micro-CT and histomorphometric analyses. There was a statistically significant difference between virgin (12.2%8%) and VE-UHMWPE (3%1.4%) groups in regards to bone resorption (P=0.005) and inflammatory fibrous tissue overlaying the calvaria (0.48 vs. 0.20, P<0.0001). These results suggest that VE-UHMWPE particles have reduced osteolytic potential in vivo when compared to virgin UHMWPE. PMID:24998319

  12. Protein oxidation at different salt concentrations affects the cross-linking and gelation of pork myofibrillar protein catalyzed by microbial transglutaminase.

    PubMed

    Li, Chunqiang; Xiong, Youling L; Chen, Jie

    2013-06-01

    In a fabricated then restructured meat product, protein gelation plays an essential role in producing desirable binding and fat-immobilization properties. In the present study, myofibrillar protein (MFP) suspended in 0.15, 0.45, and 0.6 M NaCl was subjected to hydroxyl radical stress for 2 or 24 h and then treated with microbial transglutaminase (MTGase) in 0.6 M NaCl (E?:?S = 1?:?20) at 4 and 15 C for 2 h. Protein cross-linking and dynamic rheological tests were performed to assess the efficacy of MTGase for mediating the gelation of oxidized MFP. MTGase treatments affected more remarkable polymerization of myosin in oxidized MFP than in nonoxidized, especially for samples oxidized at 0.6 M NaCl. Notably, the extent of MTGase-induced myosin cross-linking at 15 C in oxidized MFP improved up to 46.8%, compared to 31.6% in nonoxidized MFP. MTGase treatment at 4 C for MFP oxidized in 0.6 M NaCl, but not MFP oxidized in 0.15 M NaCl, produced stronger gels than nonoxidized MFP (P < 0.05). The final (75 C) storage modulus (G') of oxidized MFP gels was significantly greater than that of nonoxidized, although the G' of the transient peak (?44.5 C) showed the opposite trend. Overall, oxidation at high salt concentrations significantly improved MTGase-mediated myosin cross-linking and MFP gelation. This might be because under this condition, MTGase had an increased accessibility to glutamine and lysine residues to effectively initiate protein-protein interactions and gel network formation. PMID:23627930

  13. Intersubunit interaction at the active site of D-ribulose-1,5-bisphosphate carboxylase/oxygenase as revealed by cross-linking and site-directed mutagenesis

    SciTech Connect

    Lee, E.H.; Soper, T.S.; Mural, R.J.; Stringer, C.D.; Hartman, F.C.

    1987-07-28

    For measurement of distances between active-site residues of ribulose-1,5-bisphosphate carboxylase/oxygenase from Rhodospirillum rubrum (a homodimer of 50.5-kDa subunits), the reaction of the enzyme with 4,4'-difluoro-3,3'-dinitrodiphenyl sulfone (which spans 9 A) has been explored. Inactivation of the enzyme by the bifunctional reagent is not associated with an increase in apparent molecular weight, thereby excluding intermolecular cross-linking. However, in the presence of urea, gel filtration of the inactivated enzyme reveals a prominent dimeric species attributed to intersubunit cross-linking. The major chromophoric peptide has been isolated from a tryptic digest of the dimer; sequence analysis of this peptide reveals that the intersubunit cross-link occurs between Cys-58 and active site Lys-166. In contrast to previous substitutions for Lys-166 introduced by site-directed mutagenesis, replacement by aspartic acid prevents association of the two subunits. In addition to identifying an intersubunit contact, these observations suggest that the catalytic site of the carboxylase is positioned at an interface between subunits and that segments of both subunits may be required for catalytic competence.

  14. Synthesis of a new potential biodegradable disulfide containing poly(ethylene imine)-poly(ethylene glycol) copolymer cross-linked with click cluster for gene delivery.

    PubMed

    Zhao, Nan; Roesler, Susanne; Kissel, Thomas

    2011-06-15

    Poly(ethylene glycol)-grafted-polyethylenimine (PEG-PEI) are promising non-viral gene delivery systems. Herein, we aimed to synthesize a biodegradable disulfide containing PEGylated PEI to attempt to reduce its cytotoxicity and enhance the gene transfer activity. Using click chemistry, low Mw PEI (br. 2 kDa) and short chain length PEG (tetraethylene glycol, TEG) were cross-linked to a high Mw PEG-PEI copolymer (∼ 22 kDa). The chemical structure of the copolymer was characterized using (1)H NMR and GPC. The degradation behavior was investigated under in vitro conditions in the presence of 1,4-dithiothreitol (DTT). The gel retardation assay, dynamic light scattering and atomic force microscopy showed good DNA condensation ability by forming polyplexes with small particle size and positive zeta potential. In particular, MTT assay indicated that this PEG-PEI polymer is about 22-fold less toxic than PEI 25k and only 2-fold more toxic than PEI 2k in L929 cell line. After coupling of small PEG chains and cross-linking by disulfide bridges, the transfection efficiency is increased approximately 6-fold in comparison to PEI 2k and still reaches approximately 17% of PEI 25k. Hence, this click cluster cross-linked disulfide containing PEG-PEI copolymer could be an attractive cationic polymer for non-viral gene delivery. PMID:21439364

  15. Bcr is a substrate for Transglutaminase 2 cross-linking activity

    PubMed Central

    2011-01-01

    Background Breakpoint cluster region (Bcr) is a multi-domain protein that contains a C-terminal GTPase activating protein (GAP) domain for Rac. Transglutaminase 2 (TG2) regulates Bcr by direct binding to its GAP domain. Since TG2 has transglutaminase activity that has been implicated in the response to extreme stress, we investigated if Bcr can also act as a substrate for TG2. Results We here report that activation of TG2 by calcium caused the formation of covalently cross-linked Bcr. Abr, a protein related to Bcr but lacking its N-terminal oligomerization domain, was not cross-linked by TG2 even though it forms a complex with it. A Bcr mutant missing the first 62 amino acid residues remained monomeric in the presence of activated TG2, showing that this specific domain is necessary for the cross-linking reaction. Calcium influx induced by a calcium ionophore in primary human endothelial cells caused cross-linking of endogenous Bcr, which was inhibited by the TG2 inhibitor cystamine. Treatment of cells with cobalt chloride, a hypoxia-mimetic that causes cellular stress, also generated high molecular weight Bcr complexes. Cross-linked Bcr protein appeared in the TritonX-100-insoluble cell fraction and further accumulated in cells treated with a proteasome inhibitor. Conclusions Bcr thus represents both an interacting partner under non-stressed conditions and a target of transglutaminase activity for TG2 during extreme stress. PMID:21310073

  16. Detection of the cross-linking amino acid, histidinoalanine, in human brown cataractous lens protein.

    PubMed

    Kanayama, T; Miyanaga, Y; Horiuchi, K; Fujimoto, D

    1987-02-01

    In human nuclear cataract, with its progression, high polymers protein mass, which is aggregated and strengthened by disulfide cross-links and other covalent cross-links, accumulates in the nuclear region to cause hardening of the lens nucleus. However, the structure of non-disulfide covalent cross-links has not been clearly confirmed yet. Recently a cross-linking amino acid, histidinoalanine, was discovered in connective tissue and is thought to cause hardening of the tissue with aging. The content of this amino acid in the lens nucleus was studied. Nuclear cataract was classified on the basis of nuclear color as described by Pirie (1986). The content of histidinoalanine was found to be 0.05 +/- 0.03 nmol mg-1 in normal nucleus; 0.06 +/- 0.03 nmol mg-1 in grade I (uniform pale yellow); 0.05 +/- 0.03 nmol mg-1 in grade II (deep yellow nucleus); 0.38 +/- 0.07 nmol mg-1 (P less than 0.01) in grade III (hazel-brown nucleus) and 3.35 +/- 2.76 nmol mg-1 (P less than 0.01) in grade IV (deep brown nucleus). The result suggests that histidinoalanine is one of the non-disulfide cross-links of lens proteins in nuclear cataract and is closely related to the grade of cataract. PMID:3582505

  17. Biodegradable Chitosan-Based Ambroxol Hydrochloride Microspheres: Effect of Cross-Linking Agents

    PubMed Central

    Gangurde, HH; Chavan, NV; Mundada, AS; Derle, DV; Tamizharasi, S

    2011-01-01

    The objective of this study was to investigate the influence of type of cross-linking method used on the properties of ambroxol hydrochloride microspheres such as encapsulation efficiency, particle size, and drug release. Microspheres were prepared by solvent evaporation technique using chitosan as a matrix-forming agent and cross-linked using formaldehyde and heat treatment. Morphological and physicochemical properties of microspheres were then investigated by scanning electron microscopy (SEM), X-ray diffractometry (XRD), differential scanning calorimetry (DSC), and Fourier-transform infrared spectroscopy (FTIR) spectroscopy. The cross-linking of chitosan takes place at the free amino group because of formation of imine bond as evidenced by FTIR. The DSC, XRD, and FTIR analysis showed that chitosan microspheres cross linked by heating were superior in properties and performance as compared to the microspheres cross-linked using formaldehyde. SEM results revealed that heat-treated microspheres were spherical, discrete having smooth, and porous structure. The particle size and encapsulation efficiencies of the prepared chitosan microspheres ranged between 10.83–24.11 μm and 39.73μ80.56%, respectively. The drug release was extended up to 12 h, and the kinetics of the drug release was obeying Higuchi kinetic proving diffusion-controlled drug release. PMID:21607049

  18. Synthesis of surface protein-imprinted nanoparticles endowed with reversible physical cross-links.

    PubMed

    Yang, Chongchong; Yan, Xianming; Guo, Hao; Fu, Guoqi

    2016-01-15

    Researches on protein molecularly imprinted polymers have been challenged by the difficulties in facilitating biomacromolecular transfer, in particular upon the template removal step, and enhancing their recognition performance. Addressing these issues, herein we report synthesis of coreshell structured surface protein-imprinted nanoparticles with reversible physical cross-links formed in the imprinted nanoshells. The imprinted layers over nanoparticle supports are fabricated via aqueous precipitation polymerization (PP) of di(ethylene glycol) methyl ether methacrylate (MEO2MA), a thermo-responsive monomer bearing no strong H-bond donor, and other functional and cross-linking monomers. During polymerization, physical cross-links together with chemical cross-links are in site produced within the imprinted shells based on hydrophobic association among the PMEO2MA, favoring formation of high-quality imprints. While cooled appropriately below the polymerization temperature, these physical cross-links can be dissociated rapidly, thus facilitating removal of the embedded template. For proof of this concept, lysozyme-imprinted nanoparticles were synthesized at 37 C over the nanoparticles functionalized with carboxylic and vinyl groups. The template removal from the imprinted nanoparticles was readily achieved by washing with a dilute acidic detergent solution at 4 C. As-prepared imprinted nanoparticles showed greatly higher imprinting factor and specific rebinding than obtained with the same recipe but by solution polymerization (SP). Moreover, such imprinted nanomaterials exhibited satisfactory rebinding selectivity, kinetics and reusability. PMID:26313422

  19. Cornifin, a cross-linked envelope precursor in keratinocytes that is down-regulated by retinoids.

    PubMed Central

    Marvin, K W; George, M D; Fujimoto, W; Saunders, N A; Bernacki, S H; Jetten, A M

    1992-01-01

    In this study, we have characterized the cDNA clone SQ37 that was isolated previously from a rabbit squamous cell library. The gene encodes a 14-kDa protein that appears to function as a component of the cross-linked envelope in squamous differentiating cells. The protein, which has been named cornifin, has a high content of proline (31%), glutamine (20%), and cysteine (11%) and contains 13 repeats of an octapeptide (consensus sequence, EPCQPKVP) at its C terminus. SQ37 mRNA and protein are induced during squamous differentiation of rabbit tracheal (RbTE) cells and human epidermal keratinocytes. This induction is repressed by retinoids. Immunohistochemical studies reveal SQ37 immunoreactivity in fragmented cross-linked envelopes from squamous-differentiated RbTE cells and in the suprabasal layers of the epidermis. In situ hybridization analysis showed that the presence of SQ37 mRNA is restricted to the suprabasal layers. Treatment of RbTE cells with a Ca2+ ionophore induces cross-linking of the SQ37 protein into higher molecular weight complexes. This cross-linking reaction appears to be mediated by transglutaminase type I. Our observations suggest that the protein encoded by SQ37 participates in the assembly of the cross-linked envelope. Images PMID:1438308

  20. Nanoparticle cross-linked collagen shields for sustained delivery of pilocarpine hydrochloride.

    PubMed

    Agban, Yosra; Lian, Jiaxin; Prabakar, Sujay; Seyfoddin, Ali; Rupenthal, Ilva D

    2016-03-30

    Glaucoma is a common progressive eye disorder which remains the second leading cause of blindness worldwide. Current therapy involves frequent administration of eye drops which often results in poor patient adherence and therapeutic outcomes. The aim of this study was to overcome these limitations by developing a novel nanoparticle cross-linked collagen shield for sustained delivery of pilocarpine hydrochloride (PHCl). Three metal oxide nanoparticles (NPs); titanium dioxide (TiO2), zinc oxide (ZnO) and polyvinylpyrrolidone (PVP) capped zinc oxide (ZnO/PVP), were evaluated for their cytotoxicity as well as shield transparency before selecting ZnO/PVP NPs as the ideal candidate. Cross-linked collagen shields were then characterized for their mechanical strength, swelling capacity and bioadhesive properties, with ZnO/PVP NP cross-linked shields showing the most favorable characteristics compared to plain films. The shield with the best properties was then loaded with PHCl and in vitro release of zinc ions as well as PHCl was measured without and with further cross-linking by ultraviolet irradiation. The concentration of zinc ions released was well below the IC50 rendering them safe for ocular use. Moreover, collagen shields cross-linked with ZnO/PVP NPs released PHCl over a period of 14 days offering a promising sustained release treatment option for glaucoma. PMID:26828672

  1. Cross-Linking Poly(lactic acid) Film Surface by Neutral Hyperthermal Hydrogen Molecule Bombardment.

    PubMed

    Du, Wangli; Shao, Hong; He, Zhoukun; Tang, Changyu; Liu, Yu; Shen, Tao; Zhu, Yan; Lau, Woon-Ming; Hui, David

    2015-12-16

    Constructing a dense cross-linking layer on a polymer film surface is a good way to improve the water resistance of poly(lactic acid) (PLA). However, conventional plasma treatments have failed to achieve the aim as a result of the unavoidable surface damage arising from the charged species caused by the uncontrolled high energy coming from colliding ions and electrons. In this work, we report a modified plasma method called hyperthermal hydrogen-induced cross-linking (HHIC) technology to construct a dense cross-linking layer on PLA film surfaces. This method produces energy-controlled neutral hyperthermal hydrogen, which selectively cleaves C-H bonds by molecule collision from the PLA film without breaking other bonds (e.g., C-C bonds in the polymer backbone), and results in subsequent cross-linking of the carbon radicals generated from the organic molecules. The formation of a dense cross-linking layer can serve as a barrier layer to significantly improve both the hydrophobicity and water vapor barrier property of the PLA film. Because of the advantage of selective cleavage of C-H bonds by HHIC treatment, the original physical properties (e.g., mechanical strength and light transmittance) of the PLA films are well-preserved. PMID:26594874

  2. Polyimide aerogels with amide cross-links: a low cost alternative for mechanically strong polymer aerogels.

    PubMed

    Meador, Mary Ann B; Alemn, Christian R; Hanson, Katrina; Ramirez, Nakaira; Vivod, Stephanie L; Wilmoth, Nathan; McCorkle, Linda

    2015-01-21

    Polyimide aerogels combine high porosity, low thermal conductivity, flexibility, and low density with excellent mechanical properties. However, previously used cross-linkers, such as 1,3,5-triaminophenoxybenzene (TAB), 2,4,6-tris(4-aminophenyl)pyridine (TAPP), or octa(aminophenoxy)silsesquioxane (OAPS), either are not commercially available or are prohibitively expensive. Finding more cost efficient cross-linkers that are commercially available to synthesize these aerogels is crucial for making large scale manufacturing attractive. Herein, we describe an approach to making polyimide aerogels starting with amine capped oligomers that are cross-linked with 1,3,5-benzenetricarbonyl trichloride (BTC). BTC is a lower cost, commercially available alternative to TAB, TAPP, or OAPS. Aerogels made in this way have the same or higher modulus and higher surface area compared to those previously reported with either TAB or OAPS cross-links at the same density. While the cross-link structure is an amide, the thermal stability is not compromised most likely because the cross-link is only a small part of the composition of the aerogel. Onset of decomposition depends primarily on the backbone chemistry with 4,4'-oxidianiline (ODA) being more thermally stable than 2,2'-dimethylbenzidine (DMBZ), similar to those previously reported with other cross-links. PMID:25564878

  3. Dependence of nanomechanical modification of polymers on plasma-induced cross-linking

    NASA Astrophysics Data System (ADS)

    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-modified 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.

  4. Availability of fluorescence spectroscopic in the accompaniment of formation of corneal cross-linking

    NASA Astrophysics Data System (ADS)

    Costa, M. M.; Kurachi, C.; Bagnato, V. S.; Faria e Sousa, S. J.; Ventura, L.

    2010-02-01

    The corneal cross-linking is a method that associates riboflavin and ultraviolet light to induce a larger mechanical resistance at cornea. This method has been used for the treatment of Keratoconus. Since cross-linking is recent as treatment, there is a need to verify the effectiveness of the method. Therefore, the viability of the fluorescence spectroscopy technique to follow the cross-linking formation at cornea was studied. Corneas were divided in two measuring procedures: M1 (cornea + riboflavin), and M2 (cornea + riboflavina + light irradiation, 365nm). For fluorescence measurements, a spectrofluorimeter was used, where several wavelengths were selected (between 320nm and 370nm) for cornea excitation. Several fluorescence spectra were collected, at 10 min-interval, during 60 min. Spectra allowed one to observe two very well defined bands of fluorescence: the first one at 400nm (collagen), and the second one at 520nm (riboflavin). After spectra analyses, a decrease of collagen fluorescence was observed for both groups. For riboflavin, on the other hand, there was a fluorescence increase for M1, and a decrease for M2. Thus, it is possible to conclude that it this technique is sensitive for the detection of tissue structural changes during cross-linking treatment, encouraging subsequent studies on quantification of cross-linking promotion in tissue.

  5. Distortion-dependent unhooking of interstrand cross-links in mammalian cell extracts

    PubMed Central

    Smeaton, Michael B.; Hlavin, Erica M.; Mason, Tracey McGregor; Noronha, Anne M.; Wilds, Christopher J.; Miller, Paul S.

    2009-01-01

    Interstrand cross-links (ICLs) are formed by many chemotherapeutic agents and may also arise endogenously. The mechanisms used to repair these lesions remain unclear in mammalian cells. Repair in Escherichia coli and Saccharomyces cerevisiae requires an initial unhooking step to release the tethered DNA strands. We used a panel of linear substrates containing different site-specific ICLs in order to characterize how structure affects ICL processing in mammalian cell extracts. We demonstrate that ICL-induced distortions affect NER-dependent and independent processing events. The NER-dependent pathway produces dual incisions 5 to the site of the ICL as described previously [Bessho et. al. (1997) Mol Cell Bio 17(12): 6822-30] but does not release the cross-link. Surprisingly, we also found that the interstrand cross-linked duplexes were unhooked in mammalian cell extracts independent of the NER pathway. Unhooking occurred identically in extracts prepared from human and rodent cells and is dependent on ATP hydrolysis and metal ions. The structure of the unhooked product was characterized and was found to contain the remnant of the cross-link. Both the NER mediated dual 5 incisions and unhooking reactions were greatly stimulated by ICL-induced distortions including increased local flexibility and disruption of base-pairs surrounding the site of the ICL. These results suggest that in DNA not undergoing transcription or replication, distortions induced by the presence of an ICL could contribute significantly to initial cross-link recognition and processing. PMID:18702509

  6. Cross-linking connectivity in bone collagen fibrils: the COOH-terminal locus of free aldehyde

    NASA Technical Reports Server (NTRS)

    Otsubo, K.; Katz, E. P.; Mechanic, G. L.; Yamauchi, M.

    1992-01-01

    Quantitative analyses of the chemical state of the 16c residue of the alpha 1 chain of bone collagen were performed on samples from fetal (4-6-month embryo) and mature (2-3 year old) bovine animals. All of this residue could be accounted for in terms of three chemical states, in relative amounts which depended upon the age of the animal. Most of the residue was incorporated into either bifunctional or trifunctional cross-links. Some of it, however, was present as free aldehyde, and the content increased with maturation. This was established by isolating and characterizing the aldehyde-containing peptides generated by tryptic digestion of NaB3H4-reduced mature bone collagen. We have concluded that the connectivity of COOH-terminal cross-linking in bone collagen fibrils changes with maturation in the following way: at first, each 16c residue in each of the two alpha 1 chains of the collagen molecule is incorporated into a sheet-like pattern of intermolecular iminium cross-links, which stabilizes the young, nonmineralized fibril as a whole. In time, some of these labile cross-links maturate into pyridinoline while others dissociate back to their precursor form. The latter is likely due to changes in the molecular packing brought about by the mineralization of the collagen fibrils. The resultant reduction in cross-linking connectivity may provide a mechanism for enhancing certain mechanical characteristics of the skeleton of a mature animal.

  7. 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

  8. Usage of polarization-sensitive optical coherence tomography for investigation of collagen cross-linking

    NASA Astrophysics Data System (ADS)

    Ju, Myeong Jin; Tang, Shuo

    2015-04-01

    To investigate morphological alternation in corneal stroma induced by collagen cross-linking (CXL) treatment, polarization-sensitive optical coherence tomography (PS-OCT) capable of providing scattering, phase retardation, and degree of polarization uniformity (DOPU) images were employed on fresh bovine cornea. Significant corneal thickness reduction was observed after the CXL procedure, and its variation was quantitatively analyzed. From the scattering contrast, a hyperscattering region was observed in the anterior of the cornea immediately after the CXL procedure and its range increased with time. Within the scattering region, a slow increase was observed in the phase retardation image, and a discriminable characteristic was found in the DOPU image. A global threshold value was empirically determined from the averaged DOPU depth profile in order to locate the effective cross-linking depth. In addition to the standard protocol, an accelerated CXL procedure shortening the treatment time with higher intensity of ultraviolet-A (UV-)A power was also performed. From the measurement results after the two different CXL protocols, different cross-linking aspects were found and their difference was discussed in terms of the effectiveness of cross-linking. Based on this study, we believe that PS-OCT could be a promising optical imaging modality to evaluate the progression and effectiveness of the riboflavin/UV-A induced corneal collagen cross-linking.

  9. Orientation birefringence of cross-linked rubber containing low-mass compound

    NASA Astrophysics Data System (ADS)

    Kiyama, Ayumi; Nobukawa, Shogo; Yamauchi, Masayuki

    2015-05-01

    Molecular orientation of low-mass compounds (LMCs) in a cross-linked rubber is studied in order to obtain the basic information on the dynamics of LMC molecules in a polymer beyond the glass transition temperature. A small amount of LMCs such as 4-cyano-4'-pentylbiphenyl (5CB), tricresylphosphate (TCP), and styrene-based tackifier (TF) is added into polybutadiene rubber (BR). After cross-linking reaction, the sheet samples are used to evaluate the orientation birefringence during stretching and stress relaxation. The rectangular films, cut out from the cross-linked sheets, are set in a uniaxial stretching machine equipped with an optical system to measure both birefringence and tensile stress simultaneously. It is confirmed that orientation birefringence is proportional to the stress for not only pure cross-linked BR, but also cross-linked BR containing an LMC in a wide range of strain. Even after stretching, the birefringence does not change as far as the sample is kept at a constant strain. The results suggest that the LMC molecules are forced to orient with polymer chains by the strong intermolecular orientation correlation. Because of the LMC orientation, the stress-optical coefficient CR is enhanced by the addition of 5CB and TCP, but depressed by TF. Therefore, the LMC doping can be used to control the birefringence of a retardation film.

  10. Mechanical and biocompatible characterization of a cross-linked collagen-hyaluronic acid wound dressing.

    PubMed

    Kirk, James F; Ritter, Gregg; Finger, Isaac; Sankar, Dhyana; Reddy, Joseph D; Talton, James D; Nataraj, Chandra; Narisawa, Sonoko; Millán, José Luis; Cobb, Ronald R

    2013-01-01

    Collagen scaffolds have been widely employed as a dermal equivalent to induce fibroblast infiltrations and dermal regeneration in the treatment of chronic wounds and diabetic foot ulcers. Cross-linking methods have been developed to address the disadvantages of the rapid degradation associated with collagen-based scaffolds. To eliminate the potential drawbacks associated with glutaraldehyde cross-linking, methods using a water soluble carbodiimide have been developed. In the present study, the glycosaminoglycan (GAG) hyaluronic acid (HA), was covalently attached to an equine tendon derived collagen scaffold using 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC) to create ntSPONGE The HA was shown to be homogeneously distributed throughout the collagen matrix. In vitro analyses of the scaffold indicated that the cross-linking enhanced the biological stability by decreasing the enzymatic degradation and increasing the thermal denaturation temperature. The material was shown to support the attachment and proliferation of mouse L929 fibroblast cells. In addition, the cross-linking decreased the resorption rate of the collagen as measured in an intramuscular implant model in rabbits. The material was also shown to be biocompatible in a variety of in vitro and in vivo assays. These results indicate that this cross-linked collagen-HA scaffold, ntSPONGE has the potential for use in chronic wound healing. PMID:23896569

  11. Nanomechanics of collagen fibrils under varying cross-link densities: atomistic and continuum studies.

    PubMed

    Buehler, Markus J

    2008-01-01

    Collagen is a protein material with intriguing mechanical properties - it is highly elastic, shows large fracture strength and plays a crucial role in making Nature's structural materials tough. Collagen based tissues consist of collagen fibrils, each of which is composed out of a staggered array of ultra-long tropocollagen molecules extending to several hundred nanometers. Albeit the macroscopic properties of collagen based tissues have been studied extensively, less is known about the nanomechanical properties of tropocollagen molecules and collagen fibrils, their elementary building blocks. In particular, the relationship between molecular properties and tissue properties remains a scarcely explored aspect of the science of collagen materials. Results of molecular multi-scale modeling of the nanomechanical properties of the large-strain deformation regime of collagen fibrils under varying cross-link densities are reported in this paper. The results confirm the significance of cross-links in collagen fibrils in improving its mechanical strength. Further, it is found that cross-links influence the nature of its large-deformation and fracture behavior. Cross-link deficient collagen fibrils show a highly dissipative deformation behavior with large yield regimes. Increasing cross-link densities lead to stronger fibrils that display an increasingly brittle deformation character. The simulation results are compared with recent nanomechanical experiments at the scale of tropocollagen molecules and collagen fibrils. PMID:19627772

  12. Knee-simulator testing of conventional and cross-linked polyethylene tibial inserts.

    PubMed

    Muratoglu, Orhun K; Bragdon, Charles R; Jasty, Murali; O'Connor, Daniel O; Von Knoch, Rebecca S; Harris, William H

    2004-10-01

    We compared the resistance to delamination and to adhesive/abrasive wear of conventional and highly cross-linked polyethylene tibial inserts of a cruciate-retaining total knee design using a knee simulator. Both groups were tested after aggressive, accelerated aging, and 1 set of conventional inserts was studied without aging. Aging oxidized the conventional, but not the highly cross-linked, inserts. The simulated normal gait testing lasted for 5 and 10 million cycles for the conventional and highly cross-linked inserts, respectively. Aged conventional inserts showed delaminations, whereas none were observed in the unaged conventional and aged cross-linked inserts. Wear rates measured by the gravimetric method were 9 +/- 2 mm3, 10 +/- 4 mm3, and 1 +/- 0 mm3 per million cycles; by the metrologic method, they were 8 +/- 1 mm3, 9 +/- 2 mm3, and 3 +/- 0 mm3 for the unaged conventional, aged conventional, and aged highly crosslinked inserts, respectively. In the test model used, oxidation led to delamination, whereas increased cross-link density resulted in reduced adhesive/abrasive wear of tibial inserts. PMID:15483806

  13. Cross-linked collagen sponges loaded with plant polyphenols with inhibitory activity towards chronic wound enzymes.

    PubMed

    Antonio, Francesko; Guillem, Rocasalbas; Sonia, Tourio; Clara, Mattu; Piergiorgio, Gentile; Valeria, Chiono; Gianluca, Ciardelli; Tzanov, Tzanko

    2011-10-01

    Collagen sponges loaded with polyphenols from Hamamelis virginiana were investigated as active materials for chronic wound dressings, evaluating in vitro the inhibition of two major enzymes that impair the wound healing process - myeloperoxidase (MPO) and collagenase. Prior to polyphenols loading, collagen was cross-linked with genipin to improve its biostability. The effect of genipin cross-linking and polyphenol concentration in the development of mechanically and enzymatically stable sponges was studied. The tensile strength of the cross-linked collagen increased with the increase of the cross-linking degree, coupled to decrease in the elongation and the swelling capacity of the sponges. The stability of the sponges to collagenase digestion reached maximum when 1 mM genipin was used. However, the biostability decreased more than 10-fold after loading the sponges with polyphenols (0.5 mg/mL), nevertheless, this effect was partially overcome using higher concentration of polyphenols (1 and 2 mg/mL) to inhibit collagenase. Moreover, the polyphenols released from the sponges were sufficient for complete inhibition of MPO activity. No considerable cytotoxicity of the genipin cross-linked collagen loaded with polyphenols was observed evaluating the NIH 3T3 fibroblasts viability. PMID:21805643

  14. Cross-linked acrylic hydrogel for the controlled delivery of hydrophobic drugs in cancer therapy

    PubMed Central

    Deepa, G; Thulasidasan, Arun Kumar T; Anto, Ruby John; Pillai, J Jisha; Kumar, GS Vinod

    2012-01-01

    Objective: To investigate cross-linked hydrogels prepared via inverse emulsion polymerization to entrap poorly aqueous soluble drugs. Polyethylene glycol cross-linked acrylic polymers were synthesized and the loading and release of curcumin, a model hydrophobic drug, was investigated. Methods: Physicochemical characteristics of hydrogels were studied with 13C nuclear magnetic resonance, Fourier transform infrared spectroscopy, transmission electron microscopy, scanning electron microscopy, differential scanning calorimetry, and swelling. Polymerization of the acrylic acid with cross-linked polyethylene glycol diacrylate was characterized with 13C nuclear magnetic resonance imaging and Fourier transform infrared spectroscopy. Results: The in vitro release rate of curcumin showed that there was a sustained release from the hydrogel with increased cross-linking; the release rate depended on the pH of the releasing medium. Intracellular and cytotoxicity studies were carried out in human cervical cancer cell lines. Conclusion: The results suggest cross-linked acrylic polymers can be used as efficient vectors for pH-sensitive, controlled delivery of hydrophobic drugs. PMID:22888244

  15. Effects of partial hydrolysis and subsequent cross-linking on wheat gluten physicochemical properties and structure.

    PubMed

    Wang, Kaiqiang; Luo, Shuizhong; Cai, Jing; Sun, Qiaoqiao; Zhao, Yanyan; Zhong, Xiyang; Jiang, Shaotong; Zheng, Zhi

    2016-04-15

    The rheological behavior and thermal properties of wheat gluten following partial hydrolysis using Alcalase and subsequent microbial transglutaminase (MTGase) cross-linking were investigated. The wheat gluten storage modulus (G') and thermal denaturation temperature (Tg) were significantly increased from 2.26 kPa and 54.43°C to 7.76 kPa and 57.69°C, respectively, by the combined action of partial hydrolysis (DH 0.187%) and cross-linking. The free SH content, surface hydrophobicity, and secondary structure analysis suggested that an appropriate degree of Alcalase-based hydrolysis allowed the compact wheat gluten structure to unfold, increasing the β-sheet content and surface hydrophobicity. This improved its molecular flexibility and exposed additional glutamine sites for MTGase cross-linking. SEM images showed that a compact 3D network formed, while SDS-PAGE profiles revealed that excessive hydrolysis resulted in high-molecular-weight subunits degrading to smaller peptides, unsuitable for cross-linking. It was also demonstrated that the combination of Alcalase-based partial hydrolysis with MTGase cross-linking might be an effective method for modifying wheat gluten rheological behavior and thermal properties. PMID:26616937

  16. Characterization of Interstrand DNA-DNA Cross-Links Derived from Abasic Sites Using Bacteriophage ?29 DNA Polymerase.

    PubMed

    Yang, Zhiyu; Price, Nathan E; Johnson, Kevin M; Gates, Kent S

    2015-07-14

    Interstrand cross-links in cellular DNA are highly deleterious lesions that block transcription and replication. We recently characterized two new structural types of interstrand cross-links derived from the reaction of abasic (Ap) sites with either guanine or adenine residues in duplex DNA. Interestingly, these Ap-derived cross-links are forged by chemically reversible processes, in which the two strands of the duplex are joined by hemiaminal, imine, or aminoglycoside linkages. Therefore, understanding the stability of Ap-derived cross-links may be critical in defining the potential biological consequences of these lesions. Here we employed bacteriophage ?29 DNA polymerase, which can couple DNA synthesis and strand displacement, as a model system to examine whether dA-Ap cross-links can withstand DNA-processing enzymes. We first demonstrated that a chemically stable interstrand cross-link generated by hydride reduction of the dG-Ap cross-link completely blocked primer extension by ?29 DNA polymerase at the last unmodified nucleobase preceding cross-link. We then showed that the nominally reversible dA-Ap cross-link behaved, for all practical purposes, like an irreversible, covalent DNA-DNA cross-link. The dA-Ap cross-link completely blocked progress of the ?29 DNA polymerase at the last unmodified base before the cross-link. This suggests that Ap-derived cross-links have the power to block various DNA-processing enzymes in the cell. In addition, our results reveal ?29 DNA polymerase as a tool for detecting the presence and mapping the location of interstrand cross-links (and possibly other lesions) embedded within regions of duplex DNA. PMID:26103998

  17. CD44 and hyaluronan expression in human cutaneous scar fibroblasts.

    PubMed Central

    Messadi, D. V.; Bertolami, C. N.

    1993-01-01

    Fibrotic disorders of skin and other organs are typically associated with an abnormal accumulation of extracellular matrix. This study focuses on a matrix constituent, hyaluronan-which is known to be altered in fibrotic disorders of skin- and on CD44, a cell adhesion molecule and putative receptor for hyaluronan. Tissue samples were obtained from biopsies of human normal skin, normal cutaneous scar; and hypertrophic cutaneous scar. After culturing, cells were studied by single- and double-labeling immunohistochemistry using the two anti-CD44 monoclonal antibodies, BU-52 and J173, and a biotinylated hyaluronan binding complex probe, b-HABR. Certain cultures were pretreated with Streptomyces hyaluronidase to assess the dependency of CD44 expression on the presence of endogenous hyaluronan. CD44 expression, both in the presence and the absence of exogenous hyaluronan, was quantitated by radioimmunobinding assay. Overall glycosaminoglycan synthesis and identification of hyaluronan were accomplished by precursor incorporation assays and by quantitative cellulose acetate electrophoresis. CD44 was found to be a normal human adult fibroblastic antigen whose expression is markedly increased for hypertrophic scar fibroblasts compared with normal skin fibroblasts. Although hyaluronan was found to be the predominant glycosaminoglycan constituent of the pericellular matrix for these fibroblasts, CD44 attachment to the cell surface is neither mediated by hyaluronan nor is the presence of hyaluronan a prerequisite for CD44 expression. Exogenous hyaluronan induced a decline in measurable CD44 expression for normal skin fibroblasts but not for hypertrophic scar fibroblasts. These observations are compatible with current understanding of the way cells manage the hyaluronan economy of the extracellular matrix and emphasize phenotypic heterogeneities between fibroblasts derived from normal versus scar tissues. Images Figure 1 Figure 4 PMID:8475990

  18. Radiation cross-linking of ethylene vinyl alcohol copolymer functionalized with m-isopropenyl-[alpha],[alpha]-dimethyl benzyl isocyanate

    SciTech Connect

    Ekman, K.B.; Naesman, J.H. . Lab. of Polymer Technology)

    1993-10-10

    An ethylene vinyl alcohol copolymer was functionalized with m-isopropenyl-[alpha],[alpha]-dimethyl benzyl isocyanate using reactive processing in a mixer. The functionalization introduces pendant unsaturation to the polymer, which allows radiation cross-linked to gel contents >70% at radiation doses below 100 kGy. Unfunctionalized ethylene vinyl alcohol copolymer, on the other hand, forms no gel upon irradiation. The functionalization was completed within a few minutes of reactive mixing, which was confirmed with both FTIR and [sup 13]C-NMR measurements. The oxygen permeability of ethylene vinyl alcohol copolymer increased with increasing degree of functionalization, and irradiation of the samples formed trapped radicals, which act as oxygen scavengers. Consequently no oxygen permeability was detected. However, radical activity was inhibited by annealing the samples at 110 C resulting in a 24% higher oxygen permeability value for the irradiated unfunctionalized copolymer. The oxygen permeability values of the irradiated functionalized samples were approximately 13% lower. Laminates of m-isopropenyl-[alpha],[alpha]-dimethyl benzyl isocyanate functionalized ethylene vinyl alcohol copolymer and m-isopropenyl-[alpha],[alpha]-dimethyl benzyl isocyanate functionalized ethylene hydroxyethyl methacrylate copolymer acquired improved adhesive strength both at dry and wet conditions as well as at elevated temperature upon exposure to radiation.

  19. Enhanced apoptotic effects of dihydroartemisinin-aggregated gelatin and hyaluronan nanoparticles on human lung cancer cells.

    PubMed

    Sun, Qian; Teong, Benjamin; Chen, I-Fen; Chang, Shwu Jen; Gao, Jimin; Kuo, Shyh-Ming

    2014-04-01

    Recent studies suggest that dihydroartemisinin (DHA), a derivative of artemisinin isolated from the traditional Chinese herb Artemisia annua L., has anticancer properties. Due to poor water solubility, poor oral activity, and a short plasma half-life, large doses of DHA have to be injected to achieve the necessary bioavailability. This study examined increasing DHA bioavailability by encapsulating DHA within gelatin (GEL) or hyaluronan (HA) nanoparticles via an electrostatic field system. Observations from transmission electron microscopy show that DHA in GEL and HA nanoparticles formed GEL/DHA and HA/DHA aggregates that were approximately 30-40 nm in diameter. The entrapment efficiencies for DHA were approximately 13 and 35% for the GEL/DHA and HA/DHA aggregates, respectively. The proliferation of A549 cells was inhibited by the GEL/DHA and HA/DHA aggregates. Fluorescent annexin V-fluorescein isothiocyanate (FITC) and propidium iodide (PI) staining displayed low background staining with annexin V-FITC or PI on DHA-untreated cells. In contrast, annexin V-FITC and PI stains dramatically increased when the cells were incubated with GEL/DHA and HA/DHA aggregates. These results suggest that DHA-aggregated GEL and HA nanoparticles exhibit higher anticancer proliferation activities than DHA alone in A549 cells most likely due to the greater aqueous dispersion after hydrophilic GEL or HA nanoparticles aggregation. These results demonstrate that DHA can aggregate with nanoparticles in an electrostatic field environment to form DHA nanosized aggregates. PMID:24039154

  20. DNA Cross-linking, Double-Strand Breaks, and Apoptosis in Corneal Endothelial Cells after a Single Exposure to Mitomycin C

    PubMed Central

    Roh, Danny S.; Cook, Amanda L.; Rhee, Steven S.; Joshi, Amar; Kowalski, Regis; Dhaliwal, Deepinder K.; Funderburgh, James L.

    2010-01-01

    PURPOSE To investigate the cellular effects of mitomycin C (MMC) treatment on corneal endothelial (CE) cells at clinically relevant applications and dosages. METHODS Radial and posterior diffusion of MMC was determined by an Escherichia coli growth inhibition bioassay. A modified version of the comet assay (single cell gel electrophoresis) was used to detect DNA cross-linking. Immunostaining detected the nuclear phosphorylated histone variant H2AX (?-H2AX) indicating DNA double-strand breaks. Apoptosis in MMC-treated cells was detected with annexin V staining. RESULTS Topical application of 0.02% MMC to intact goat globes resulted in MMC in the CE at 0.37 g/mL and produced a significant increase in CE DNA cross-linking with as little as 6 seconds of topical MMC treatment. DNA cross-linking was also demonstrated in cultured CE cells by using MMC exposures similar to those detected in CE of intact eyes. Such MMC treatment of CE produced elevated and persistent ?-H2AX-positive cells indicative of DNA double-strand breaks. Similarly, there was an increase in the proportion of apoptotic CE cells, evidenced by positive annexin V staining. CONCLUSIONS The results demonstrate that exposure to MMC at times and concentrations commonly used in refractive surgery produces cross-linking of corneal endothelial DNA, persistent DNA damage, and endothelial death via apoptosis. Current practices of MMC application during refractive surgeries may increase the potential for long-term and permanent deleterious effects on the health of the corneal endothelium. PMID:18658091

  1. Design and synthesis of curcumin analogues for in vivo fluorescence imaging and inhibiting copper-induced cross-linking of amyloid beta species in Alzheimer's disease.

    PubMed

    Zhang, Xueli; Tian, Yanli; Li, Zeng; Tian, Xiaoyu; Sun, Hongbin; Liu, Hong; Moore, Anna; Ran, Chongzhao

    2013-11-01

    In this article, we first designed and synthesized curcumin-based near-infrared (NIR) fluorescence imaging probes for detecting both soluble and insoluble amyloid beta (Aβ) species and then an inhibitor that could attenuate cross-linking of Aβ induced by copper. According to our previous results and the possible structural stereohindrance compatibility of the Aβ peptide and the hydrophobic/hydrophilic property of the Aβ13-20 (HHQKLVFF) fragment, NIR imaging probe CRANAD-58 was designed and synthesized. As expected CRANAD-58 showed significant fluorescence property changes upon mixing with both soluble and insoluble Aβ species in vitro. In vivo NIR imaging revealed that CRANAD-58 was capable of differentiating transgenic and wild-type mice as young as 4 months old, the age that lacks apparently visible Aβ plaques and Aβ is likely in its soluble forms. According to our limited studies on the interaction mechanism between CRANAD-58 and Aβ, we also designed CRANAD-17 to attenuate the cross-linking of Aβ42 induced by copper. It is well-known that the coordination of copper with imidazoles on Histidine-13 and 14 (H13, H14) of Aβ peptides could initialize covalent cross-linking of Aβ. In CRANAD-17, a curcumin scaffold was used as an anchoring moiety to usher the designed compound to the vicinity of H13 and H14 of Aβ, and imidazole rings were incorporated to compete with H13/H14 for copper binding. The results of SDS-PAGE gel and Western blot indicated that CRANAD-17 was capable of inhibiting Aβ42 cross-linking induced by copper. This raises a potential for CRANAD-17 to be considered for AD therapy. PMID:24116384

  2. Direct cross-linking of snRNP proteins F and 70K to snRNAs by ultra-violet radiation in situ.

    PubMed Central

    Woppmann, A; Rinke, J; Lhrmann, R

    1988-01-01

    Protein-RNA interactions in small nuclear ribonucleoproteins (UsnRNPs) from HeLa cells were investigated by irradiation of purified nucleoplasmic snRNPs U1 to U6 with UV light at 254 nm. The cross-linked proteins were analyzed on one- and two-dimensional gel electrophoresis systems, and the existence of a stable cross-linkage was demonstrated by isolating protein-oligonucleotide complexes from snRNPs containing 32P-labelled snRNAs after exhaustive digestion with a mixture of RNases of different specificities. The primary target of the UV-light induced cross-linking reaction between protein and RNA was protein F. It was also found to be cross-linked to U1 snRNA in purified U1 snRNPs. Protein F is known to be one of the common snRNP proteins, which together with D, E and G protect a 15-25 nucleotide long stretch of snRNAs U1, U2, U4 and U5, the so-called domain A or Sm binding site against nuclease digestion (Liautard et al., 1982). It is therefore likely that the core-protein may bind directly and specifically to the common snRNA domain A, or else to a sub-region of this. The second protein which was demonstrated to be cross-linked to snRNA was the U1 specific protein 70K. Since it has been shown that binding of protein 70K to U1 RNP requires the presence of the 5' stem and loop of U1 RNA (Hamm et al., 1987) it is likely that the 70K protein directly interacts with a sub-region of the first stem loop structure. Images PMID:2974540

  3. xiNET: Cross-link Network Maps With Residue Resolution*

    PubMed Central

    Combe, Colin W.; Fischer, Lutz; Rappsilber, Juri

    2015-01-01

    xiNET is a visualization tool for exploring cross-linking/mass spectrometry results. The interactive maps of the cross-link network that it generates are a type of node-link diagram. In these maps xiNET displays: (1) residue resolution positional information including linkage sites and linked peptides; (2) all types of cross-linking reaction product; (3) ambiguous results; and, (4) additional sequence information such as domains. xiNET runs in a browser and exports vector graphics which can be edited in common drawing packages to create publication quality figures. Availability: xiNET is open source, released under the Apache version 2 license. Results can be viewed by uploading data to http://crosslinkviewer.org/ or by downloading the software from http://github.com/colin-combe/crosslink-viewer and running it locally. PMID:25648531

  4. Fracture Behavior of High-Toughness, Ionically Cross-linked Triblock Copolymer Hydrogels

    NASA Astrophysics Data System (ADS)

    Henderson, Kevin; Otim, Kathryn; Shull, Kenneth

    2011-03-01

    Mechanisms for enhancing energy dissipation and hence toughness are important for the generation of robust synthetic soft materials for biomedical applications. Ionic cross-linking in particular has been explored in triblock copolymer hydrogels and affords a remarkable change in mechanical performance comparable to non-cross-linked analogs. Here we employ a physically associated base triblock copolymer network composed of hydrophobic poly(methyl methacrylate) endblocks and a hydrophilic poly(methacrylic acid) midblock capable of complexing with divalent cations. Increases in stiffness and strength have previously been reported, with the extent dependent upon the identity, concentration, and pH of a cross-linking cation solution. We delineate the measured toughness in such systems using tensile tear tests and relate the mechanical performance to a damage zone model reminiscent of loading behavior observed in double network hydrogels.

  5. Comparison of Wear and Oxidation in Retrieved Conventional and Highly Cross-Linked UHMWPE Tibial Inserts.

    PubMed

    Currier, Barbara H; Currier, John H; Franklin, Katherine J; Mayor, Michael B; Reinitz, Steven D; Van Citters, Douglas W

    2015-12-01

    Two groups of retrieved tibial inserts from one manufacturer's knee system were analyzed to evaluate the effect of a highly cross-linked bearing surface on wear and in vivo oxidation. The two groups ((1) conventional gamma-inert sterilized and (2) highly cross-linked, coupled with the same rough (Ra=0.25) Ti-6Al-4V tray) were matched with statistically similar in vivo duration and patient variables. The retrieved inserts were analyzed for ketone oxidation and wear in the form of dimensional change. The difference in oxidation rate between highly cross-linked and conventional gamma-inert sterilized inserts did not reach statistical significance. Observations suggest that the majority of wear can be accounted for by the backside interface with the rough Ti-6Al-4V tray; however, wear measured by thickness-change rate was statistically indistinguishable between the two bearing materials. PMID:26143237

  6. Bioinert solution-cross-linked hydrogen-bonded multilayers on colloidal particles.

    PubMed

    Yang, Sung Yun; Lee, Daeyeon; Cohen, Robert E; Rubner, Michael F

    2004-07-01

    Bioinert polyelectrolyte multilayers comprised of poly(acrylic acid) and polyacrylamide were deposited on colloidal particles (1.7 microm in diameter) at low pH conditions by layer-by-layer assembly using hydrogen-bonding interactions. The multilayer films were coated uniformly on the colloidal particles without causing any flocculation of the colloids, and the deposited films were subsequently cross-linked by a single treatment of a carbodiimide aqueous solution. The lightly cross-linked multilayer films show excellent stability at physiological conditions (pH 7.4, phosphate-buffered saline), whereas untreated multilayer films dissolved. The multilayer-coated surfaces, both on flat substrates and on colloidal particles, exhibit excellent resistance toward mammalian cell adhesion. With this new solution-based cross-linking method, bioinert H-bonded multilayer coatings offer potential for biomedical applications. PMID:16459619

  7. In vitro calcification and in vivo biocompatibility of the cross-linked polypentapeptide of elastin

    SciTech Connect

    Wood, S.A.; Lemons, J.E.; Prasad, K.U.; Urry, D.W.

    1986-03-01

    The in vitro calcifiability and molecular weight dependence of calcification of the polypentapeptide, (L X Val1-L X Pro2-Gly3-L X Val4-Gly5)n, which had been gamma-irradiation cross-linked have been determined when exposed to dialyzates of normal, nonaugmented fetal bovine serum. The material was found to calcify: calcifiability was found to be highly molecular weight dependent and to be most favored when the highest molecular weight polymers (n approximately equal to 240) had been used for cross-linking. The in vivo biocompatibility, biodegradability, and calcifiability of the gamma-irradiation cross-linked polypentapeptide were examined in rabbits in both soft and hard tissue sites. The material was found to be biocompatible irrespective of its physical form and to be biodegradable but with n of 200 or less it was not shown to calcify or ossify in the rabbit tibial nonunion model.

  8. [Cross-Linking in Keratoconus: "Epi-off" or "Epi-on"?].

    PubMed

    Raiskup, F; Velik, V; Vesel, M; Sprl, E

    2015-12-01

    Corneal cross-linking can halt the progression of keratoconus, but what is the best approach for treatment? There are a number of treatment options for keratoconus, but only corneal cross-linking (CXL) appears to halt the progression of the disease. To guarantee effective cross-linking, CXL treatment involves removal of the corneal epithelium prior to riboflavin application and ultraviolet light illumination - "epi-off" CXL. Several methods of "epi-on" (transepithelial) CXL have been proposed, such as keeping the corneal epithelium intact which should be less painful and help avoid other CXL-associated adverse events. The evidence so far is that epi-off CXL remains the most effective method of strengthening the cornea and slowing keratoconus progression - but transepithelial methods are gaining ground. PMID:26678902

  9. Three-dimensional multimodal microscopy of rabbit cornea after cross-linking treatment

    NASA Astrophysics Data System (ADS)

    Krger, A.; Hovakimyan, M.; Ramrez, D. F.; Lorbeer, R.-A.; Krger, M.; Stachs, O.; Wree, A.; Guthoff, R. F.; Lubatschowski, H.; Heisterkamp, A.

    2010-02-01

    Cross-linking of stromal collagen with Riboflavin and UVA radiation is an alternative treatment of keratoconus. After the cross-linking a wound healing process starts with the regeneration of the abraded epithelial layer and the stromal keratocyte-network. To clarify possible side effects by visualization we established an imaging platform for the multimodal three-dimensional imaging of the cornea and looked for differences between normal and cross-linked rabbit corneae. The microscopy system utilizes femtosecond laser light for two photon excitation of autofluorescent metabolic compounds, second harmonic imaging in forward and backward direction for the study of stromal collagen-I structure and confocal detection of the backscattered femtosecond laser light for cell detection. Preliminary results show signatures of treatment 5 weeks after the intervention in all imaging modalities.

  10. Preparation of cross-linked maize (Zea mays L.) starch in different reaction media.

    PubMed

    Hong, Jung Sun; Gomand, Sara V; Delcour, Jan A

    2015-06-25

    Granular normal maize starch was reacted with sodium trimetaphosphate in deionized water ( [Formula: see text] ), aqueous sodium sulfate solution ( [Formula: see text] ), aqueous ethanol (MSethanol) or aqueous acetone (MSacetone) under otherwise identical reaction conditions. Analysis of the resultant starches by Rapid Visco Analysis (RVA) showed that the starch was cross-linked to a higher degree in aqueous ethanol or aqueous acetone than in water or sodium sulfate solution, and with minimal starch leaching. While MSacetone and MSethanol had incorporated similar levels of phosphorous, RVA analysis and microscopic analysis showed that MSacetone granules were more effectively stabilized by cross-linking than MSethanol granules. Cross-linking in aqueous acetone is believed to either contain the greater numbers of distarch monophosphate (versus monostarch monophosphate), or occur more intensively at the granule outer layers than that in aqueous ethanol and, at the same time, to account for the greater granular strength of MSethanol than that of MSacetone. PMID:25839824

  11. Pinpointing RNA-Protein Cross-Links with Site-Specific Stable Isotope-Labeled Oligonucleotides.

    PubMed

    Lelyveld, Victor S; Bjrkbom, Anders; Ransey, Elizabeth M; Sliz, Piotr; Szostak, Jack W

    2015-12-16

    High affinity RNA-protein interactions are critical to cellular function, but directly identifying the determinants of binding within these complexes is often difficult. Here, we introduce a stable isotope mass labeling technique to assign specific interacting nucleotides in an oligonucleotide-protein complex by photo-cross-linking. The method relies on generating site-specific oxygen-18-labeled phosphodiester linkages in oligonucleotides, such that covalent peptide-oligonucleotide cross-link sites arising from ultraviolet irradiation can be assigned to specific sequence positions in both RNA and protein simultaneously by mass spectrometry. Using Lin28A and a let-7 pre-element RNA, we demonstrate that mass labeling permits unambiguous identification of the cross-linked sequence positions in the RNA-protein complex. PMID:26583201

  12. Synthesis and evaluation of physicochemical properties of cross-linked sago starch.

    PubMed

    Singh, Akhilesh V; Nath, Lila K

    2012-01-01

    Highly substituted sago starch phosphate was synthesized using POCl(3) as cross-linking reagent. Titrimetric and Fourier transform infra red (FT-IR) spectral analysis were used to characterize the substitution. Studying the different factors affecting the reaction parameters showed that the optimal conditions for starch phosphorylation were: 4h reaction time and reagent concentration 1.5% (w/w). The physicochemical properties of cross-linked sago starch (CLSS) were done using Scanning electron micrograph (SEM), X-ray powder diffractometer (XRD and Thermogravimetric analysis (TGA). The results revealed that crystalline nature of native sago starch was transformed after cross-linking. TGA report exhibited higher thermal stability, which makes it suitable for various industrial applications. Swelling behavior showed high swelling at low temperature (30 and 60C) as compared to high temperature (90C). PMID:21971581

  13. Third-body wear of highly cross-linked polyethylene in a hip simulator.

    PubMed

    Bragdon, Charles R; Jasty, Murali; Muratoglu, Orhun K; O'Connor, Daniel O; Harris, William H

    2003-08-01

    The wear performance of a radiation cross-linked melted ultrahigh-molecular-weight polyethylene (UHMWPE) articulating against 28-mm cobalt chrome femoral heads in the presence of third-body particulate debris was investigated in a hip simulator and compared with the wear of conventional UHMWPE. Particles of aluminum oxide or bone cement containing barium sulfate were added to the serum. In the presence of aluminum oxide particles, the incremental wear rates of conventional UHMWPE averaged as high as 149 +/- 116 mg/million cycles compared with 37 +/- 38 mg/million cycles for the highly cross-linked components. The difference in the average weight loss was statistically significant at P <.01. With bone cement particles, the conventional UHMWPE components had an average incremental wear rate of 19 +/- 5mg/million cycles, and the wear rate of the highly cross-linked UHMWPE components was 0.5 +/- 0.7 mg/million cycles. PMID:12934205

  14. Optimizing end-group cross-linking polymer electrolytes for fuel cell applications

    SciTech Connect

    Kim, Yu Seung; Lee, Kwan Soo; Jeong, Myung - Hwan; Lee, Jae - Suk

    2009-01-01

    This paper demonstrates the optimization of proton conductivity and water uptake for cross-linkable polymer electrolytes through synthesis and characterization of end-group cross-linkable sulfonated poly(arylene ether) copolymers (ESF-BPs). The extent of reaction of cross-linking was controlled by reaction time resulting in a series of polymers with two, independent tunable parameters, degree of sulfonation (DS) and degree of cross-linking (DC). For the polymers presented, cross-linking improved proton conductivity while reducing water uptake, an uncommon trend in polymer electrolytes where water is critical for proton conduction. Other trends relating to changes are reported and the results yield insight into the role of DS and DC and how to optimize electrochemical properties and performance of polymer electrolytes through these tunable parameters. Select polymer electrolytes were tested in fuel cells where performance and durability with accelerated relative humidity cycling were compared with Nafion{reg_sign}.

  15. Pinpointing RNA-Protein Cross-Links with Site-Specific Stable Isotope-Labeled Oligonucleotides

    PubMed Central

    2015-01-01

    High affinity RNA-protein interactions are critical to cellular function, but directly identifying the determinants of binding within these complexes is often difficult. Here, we introduce a stable isotope mass labeling technique to assign specific interacting nucleotides in an oligonucleotide-protein complex by photo-cross-linking. The method relies on generating site-specific oxygen-18-labeled phosphodiester linkages in oligonucleotides, such that covalent peptide-oligonucleotide cross-link sites arising from ultraviolet irradiation can be assigned to specific sequence positions in both RNA and protein simultaneously by mass spectrometry. Using Lin28A and a let-7 pre-element RNA, we demonstrate that mass labeling permits unambiguous identification of the cross-linked sequence positions in the RNA-protein complex. PMID:26583201

  16. 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.

  17. Size Matters: Molecular Weight Specificity of Hyaluronan Effects in Cell Biology

    PubMed Central

    Cyphert, Jaime M.; Trempus, Carol S.; Garantziotis, Stavros

    2015-01-01

    Hyaluronan signaling properties are unique among other biologically active molecules, that they are apparently not influenced by postsynthetic molecular modification, but by hyaluronan fragment size. This review summarizes the current knowledge about the generation of hyaluronan fragments of different size and size-dependent differences in hyaluronan signaling as well as their downstream biological effects. PMID:26448754

  18. Carbodiimide cross-linked amniotic membranes for cultivation of limbal epithelial cells.

    PubMed

    Ma, David Hui-Kang; Lai, Jui-Yang; Cheng, Hsiao-Yun; Tsai, Chen-Chi; Yeh, Lung-Kun

    2010-09-01

    In ophthalmic tissue engineering, amniotic membrane (AM) is one of the most prevalent natural matrices used for limbal epithelial cell (LEC) cultivation and transplantation. However, the application of AM as a scaffold is limited by its low biomechanical strength and rapid biodegradation. The present study reports the development of 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide hydrochloride (EDC)/N-hydroxysuccinimide (NHS) cross-linked AM as an LEC carrier. The collagenous tissue materials were modified with varying cross-linker concentrations (0-0.25 mmol EDC/mg AM) and were characterized by attenuated total reflection-Fourier transform infrared spectroscopy (ATR-FTIR), differential scanning calorimetry (DSC), ninhydrin assays, electron microscopy, light transmission measurements, mechanical and in vitro degradation tests, as well as diffusion permeability and cell culture studies. Our results showed that chemical cross-linking approaches saturation at concentrations of 0.05 mmol EDC/mg AM. The formation of cross-links (i.e., amide bonds) in the samples treated with 0.05 mmol EDC/mg AM may cause significant aggregation of tropocollagen molecules and collagen microfibrils without affecting cell morphology of biological tissues. With the optimum concentration of 0.05 mmol EDC/mg AM, chemical cross-linker could significantly enhance the mechanical and thermal stability, optical transparency, and resistance to collagenase digestion. Continuous permeation of albumin through the cross-linked AM would be helpful to cell growth over the matrix surface. In addition, the EDC cross-linked samples were able to support LEC proliferation and preserve epithelial progenitor cells in vitro and in vivo. It is concluded that the AM cross-linked with 0.05 mmol EDC/mg AM may be a potential biomaterial for regenerative medicine. PMID:20541801

  19. Use of vitamin E to protect cross-linked UHMWPE from oxidation.

    PubMed

    Lerf, Reto; Zurbrgg, Daniel; Delfosse, Daniel

    2010-05-01

    Wear and oxidative degradation may limit the life span of UHMWPE implants. Cross-linking and stabilisation by vitamin E are proposed to overcome wear and degradation. The present investigation takes a close look to the oxidative behaviour of cross-linked and stabilised UHMWPE. First, the consolidated vitamin E stabilised UHMWPE was qualified in terms of microstructure and homogeneity of the distribution of the additive to be suitable for oxidation profiles over the entire section. Then cross-linked samples with five different concentrations of vitamin E (nil to 1.0%) underwent two different ageing protocols. The first was under pressurized oxygen at 70 degrees C, as defined in the ASTM F 2003 standard with a prolonged period of 60 days, the second was in 5% aqueous hydrogen peroxide solution with iron (III) chloride as catalyst at 50 degrees C. The first accelerated ageing protocol showed that a vitamin E concentration as low as 0.05% is effective to protect irradiated highly cross-linked UHMWPE against oxidation when exposed direct to oxygen. Vitamin E stabilised, highly cross-linked UHMWPE exhibits therefore no oxidation potential origination from the irradiation treatment. Analysis of samples treated by the second chemical ageing yielded, that vitamin E is effective to prolong initial stability against a supplementary attack of hydrogen peroxide and reactive radicals. The time period of stability against the aggressive hydrogen peroxide solution increases with increasing vitamin E content. However, even 0.05% have a marked stabilisation effect. Therefore, such small additions of vitamin E are effective to protect the UHMWPE material against a supplementary exposure to in vivo oxidation after the irradiation treatment. In conclusion, vitamin E shields cross-linked UHMWPE for orthopaedic application against oxidation in the heat of consolidation, during irradiation treatment and finally while implanted in the human body. PMID:20144479

  20. Proteomic Analysis of DNA-Protein Cross-Linking by Antitumor Nitrogen Mustards

    PubMed Central

    Loeber, Rachel L.; Michaelson-Richie, Erin D.; Codreanu, Simona G.; Liebler, Daniel C.; Campbell, Colin R.; Tretyakova, Natalia Y.

    2009-01-01

    Nitrogen mustards are antitumor agents used clinically for the treatment of a variety of neoplastic conditions. The biological activity of these compounds is typically attributed to their ability to induce DNA-DNA cross-links. However, nitrogen mustards are able to produce a variety of other lesions, including DNA-protein cross-links (DPCs). DPCs induced by nitrogen mustards are not well characterized because of their structural complexity and the insufficient specificity and sensitivity of previously available experimental methodologies. In the present work, affinity capture methodology in combination with mass spectrometry-based proteomics was employed to identify mammalian proteins that form covalent cross-links to DNA in the presence of a simple nitrogen mustard, mechlorethamine. Following incubation of 5′-biotinylated DNA duplexes with nuclear protein extracts, DPCs were isolated by affinity capture on streptavidin beads, and the cross-linked proteins were identified by HPLC-ESI+-MS/MS of tryptic peptides. Mechlorethamine treatment resulted in the formation of DPCs with nuclear proteins involved in chromatin regulation, DNA replication and repair, cell cycle control, transcriptional regulation, and cell architecture. Western blot analysis was employed to confirm protein identification and to quantify the extent of drug-mediated cross-linking. Mass spectrometry of amino acid-nucleobase conjugates found in total proteolytic digests revealed that mechlorethamine-induced DPCs are formed via alkylation of the N7 position of guanine in duplex DNA and cysteine thiols within the proteins to give N-[2-[S-cysteinyl]ethyl]-N-[2-(guan-7-yl)ethyl]methylamine lesions. The results described herein suggest that cellular exposure to nitrogen mustards leads to cross-linking of a large spectrum of nuclear proteins to chromosomal DNA, potentially contributing to the cytotoxic and mutagenic effects of these drugs. PMID:19480393

  1. Hyperglycemia-Induced Changes in Hyaluronan Contribute to Impaired Skin Wound Healing in Diabetes: Review and Perspective

    PubMed Central

    Shakya, Sajina; Wang, Yan; Mack, Judith A.; Maytin, Edward V.

    2015-01-01

    Ulcers and chronic wounds are a particularly common problem in diabetics and are associated with hyperglycemia. In this targeted review, we summarize evidence suggesting that defective wound healing in diabetics is causally linked, at least in part, to hyperglycemia-induced changes in the status of hyaluronan (HA) that resides in the pericellular coat (glycocalyx) of endothelial cells of small cutaneous blood vessels. Potential mechanisms through which exposure to high glucose levels causes a loss of the glycocalyx on the endothelium and accelerates the recruitment of leukocytes, creating a proinflammatory environment, are discussed in detail. Hyperglycemia also affects other cells in the immediate perivascular area, including pericytes and smooth muscle cells, through exposure to increased cytokine levels and through glucose elevations in the interstitial fluid. Possible roles of newly recognized, cross-linked forms of HA, and interactions of a major HA receptor (CD44) with cytokine/growth factor receptors during hyperglycemia, are also discussed. PMID:26448756

  2. Changes in protein conformation and surface hydrophobicity upon peroxidase-catalyzed cross-linking of apo-?-lactalbumin.

    PubMed

    Saricay, Yunus; Wierenga, Peter A; de Vries, Renko

    2014-09-24

    In this study, we explore the effect of peroxidase-catalyzed cross-linking on the molecular conformation of apo-?-lactalbumin (apo-?-LA) and the resulting changes in protein surface hydrophobicity. In studying conformational changes, we distinguish between early stages of the reaction ("partial cross-linking"), in which only protein oligomers (10(6) Da > Mw ? 10(4) Da) are formed, and a later stage ("full cross-linking"), in which larger protein particles (Mw ? 10(6) Da) are formed. Partial cross-linking induces a moderate loss of ?-helical content. Surprisingly, further cross-linking leads to a partial return of ?-helices that are lost upon early cross-linking. At the same time, for partially and fully cross-linked apo-?-LA, almost all tertiary structure is lost. The protein surface hydrophobicity first increases for partial cross-linking, but then decreases again at full cross-linking. Our results highlight the subtle changes in protein conformation and surface hydrophobicity of apo-?-LA upon peroxidase-catalyzed cross-linking. PMID:25207800

  3. Bioregeneration of hyper-cross-linked polymeric resin preloaded with phenol.

    PubMed

    Ren, Jie; Yang, Wenlan; Hua, Ming; Pan, Bingcai; Zhang, Weiming

    2013-08-01

    In this study, the preliminary feasibility of bio-regeneration of a hyper-cross-linked polymeric resin NDA-802 preloaded with phenol was investigated. As compared to the abiotic experiments, phenol preloaded with NDA-802 could be effectively desorbed and biodegraded, and the bioregenerated NDA-802 could be employed for multiple use. The concentration gradient hypothesis could interpret such bioregeneration process reasonably. A slight drop in adsorption capacity of NDA-802 after bioregeneration possibly resulted from the mesopore blockage by microbial metabolic by-products. In general, bioregeneration could serve as a potential choice for the exhausted hyper-cross-linked polymeric resin in wastewater treatment. PMID:23727014

  4. Increasing Thermal Stability of Gelatin by UV-Induced Cross-Linking with Glucose.

    PubMed

    Masutani, Evan M; Kinoshita, Christopher K; Tanaka, Travis T; Ellison, Andrew K D; Yoza, Brandon A

    2014-01-01

    The effects of ultraviolet (254?nm) radiation on a hydrated gelatin-glucose matrix were investigated for the development of a physiologically thermostable substrate for potential use in cell scaffold production. Experiments conducted with a differential scanning calorimeter indicate that ultraviolet irradiation of gelatin-glucose hydrogels dramatically increases thermal stability such that no melting is observed at temperatures of at least 90C. The addition of glucose significantly increases the yield of cross-linked product, suggesting that glucose has a role in cross-link formation. Comparisons of lyophilized samples using scanning electron microscopy show that irradiated materials have visibly different densities. PMID:24963297

  5. Increasing Thermal Stability of Gelatin by UV-Induced Cross-Linking with Glucose

    PubMed Central

    Masutani, Evan M.; Kinoshita, Christopher K.; Tanaka, Travis T.; Ellison, Andrew K. D.; Yoza, Brandon A.

    2014-01-01

    The effects of ultraviolet (254?nm) radiation on a hydrated gelatin-glucose matrix were investigated for the development of a physiologically thermostable substrate for potential use in cell scaffold production. Experiments conducted with a differential scanning calorimeter indicate that ultraviolet irradiation of gelatin-glucose hydrogels dramatically increases thermal stability such that no melting is observed at temperatures of at least 90C. The addition of glucose significantly increases the yield of cross-linked product, suggesting that glucose has a role in cross-link formation. Comparisons of lyophilized samples using scanning electron microscopy show that irradiated materials have visibly different densities. PMID:24963297

  6. 1,3-diphenylethenylcarbazolyl-based monomer for cross-linked hole transporting layers.

    PubMed

    Daskeviciene, Maryte; Bubniene, Giedre; Malinauskas, Tadas; Jankauskas, Vygintas; Gaidelis, Valentas; Paulauskas, Valdas; Getautis, Vytautas

    2015-01-01

    A new cross-linkable monomer containing 1,3-diphenylethenylcarbazolyl-based hole-transporting moieties and four reactive epoxy groups, was prepared by a multistep synthesis route from 1,3-bis(2,2-diphenylethenyl)-9H-carbazol-2-ol and its application for the in situ formation of cross-linked hole transporting layers was investigated. A high concentration of flexible aliphatic epoxy chains ensures good solubility and makes this compound an attractive cross-linking agent. The synthesized compounds were characterized by various techniques, including differential scanning calorimetry, xerographic time of flight, and electron photoemission in air methods. PMID:25996215

  7. Dynamically vulcanized biobased polylactide/natural rubber blend material with continuous cross-linked rubber phase.

    PubMed

    Chen, Yukun; Yuan, Daosheng; Xu, Chuanhui

    2014-03-26

    We prepared a biobased material, dynamically vulcanized polylactide (PLA)/natural rubber (NR) blend in which the cross-linked NR phase owned a continuous network-like dispersion. This finding breaks the traditional concept of a sea-island morphology formed after dynamic vulcanization of the blends. The scan electron microscopy and dissolution/swell experiments provided the direct proof of the continuous cross-linked NR phase. This new biobased PLA/NR blend material with the novel structure is reported for the first time in the field of dynamic vulcanization and shows promise for development for various functional applications. PMID:24621374

  8. Immunogenic Display of Purified Chemically Cross-Linked HIV-1 Spikes

    PubMed Central

    Leaman, Daniel P.; Lee, Jeong Hyun; Ward, Andrew B.

    2015-01-01

    ABSTRACT HIV-1 envelope glycoprotein (Env) spikes are prime vaccine candidates, at least in principle, but suffer from instability, molecular heterogeneity and a low copy number on virions. We anticipated that chemical cross-linking of HIV-1 would allow purification and molecular characterization of trimeric Env spikes, as well as high copy number immunization. Broadly neutralizing antibodies bound tightly to all major quaternary epitopes on cross-linked spikes. Covalent cross-linking of the trimer also stabilized broadly neutralizing epitopes, although surprisingly some individual epitopes were still somewhat sensitive to heat or reducing agent. Immunodepletion using non-neutralizing antibodies to gp120 and gp41 was an effective method for removing non-native-like Env. Cross-linked spikes, purified via an engineered C-terminal tag, were shown by negative stain EM to have well-ordered, trilobed structure. An immunization was performed comparing a boost with Env spikes on virions to spikes cross-linked and captured onto nanoparticles, each following a gp160 DNA prime. Although differences in neutralization did not reach statistical significance, cross-linked Env spikes elicited a more diverse and sporadically neutralizing antibody response against Tier 1b and 2 isolates when displayed on nanoparticles, despite attenuated binding titers to gp120 and V3 crown peptides. Our study demonstrates display of cross-linked trimeric Env spikes on nanoparticles, while showing a level of control over antigenicity, purity and density of virion-associated Env, which may have relevance for Env based vaccine strategies for HIV-1. IMPORTANCE The envelope spike (Env) is the target of HIV-1 neutralizing antibodies, which a successful vaccine will need to elicit. However, native Env on virions is innately labile, as well as heterogeneously and sparsely displayed. We therefore stabilized Env spikes using a chemical cross-linker and removed non-native Env by immunodepletion with non-neutralizing antibodies. Fixed native spikes were recognized by all classes of known broadly neutralizing antibodies but not by non-neutralizing antibodies and displayed on nanoparticles in high copy number. An immunization experiment in rabbits revealed that cross-linking Env reduced its overall immunogenicity; however, high-copy display on nanoparticles enabled boosting of antibodies that sporadically neutralized some relatively resistant HIV-1 isolates, albeit at a low titer. This study describes the purification of stable and antigenically correct Env spikes from virions that can be used as immunogens. PMID:25878116

  9. Accumulation of Extracellular Hyaluronan by Hyaluronan Synthase 3 Promotes Tumor Growth and Modulates the Pancreatic Cancer Microenvironment

    PubMed Central

    Zhao, Chunmei; Singha, Netai C.; Osgood, Ryan J.; Symons, Rebecca; Jiang, Ping; Li, Xiaoming; Thompson, Curtis B.; Infante, Jeffrey R.; Jacobetz, Michael A.; Tuveson, David A.; Frost, Gregory I.; Shepard, H. Michael; Huang, Zhongdong

    2014-01-01

    Extensive accumulation of the glycosaminoglycan hyaluronan is found in pancreatic cancer. The role of hyaluronan synthases 2 and 3 (HAS2, 3) was investigated in pancreatic cancer growth and the tumor microenvironment. Overexpression of HAS3 increased hyaluronan synthesis in BxPC-3 pancreatic cancer cells. In vivo, overexpression of HAS3 led to faster growing xenograft tumors with abundant extracellular hyaluronan accumulation. Treatment with pegylated human recombinant hyaluronidase (PEGPH20) removed extracellular hyaluronan and dramatically decreased the growth rate of BxPC-3 HAS3 tumors compared to parental tumors. PEGPH20 had a weaker effect on HAS2-overexpressing tumors which grew more slowly and contained both extracellular and intracellular hyaluronan. Accumulation of hyaluronan was associated with loss of plasma membrane E-cadherin and accumulation of cytoplasmic ?-catenin, suggesting disruption of adherens junctions. PEGPH20 decreased the amount of nuclear hypoxia-related proteins and induced translocation of E-cadherin and ?-catenin to the plasma membrane. Translocation of E-cadherin was also seen in tumors from a transgenic mouse model of pancreatic cancer and in a human non-small cell lung cancer sample from a patient treated with PEGPH20. In conclusion, hyaluronan accumulation by HAS3 favors pancreatic cancer growth, at least in part by decreasing epithelial cell adhesion, and PEGPH20 inhibits these changes and suppresses tumor growth. PMID:25147816

  10. Use of Proteinase K Nonspecific Digestion for Selective and Comprehensive Identification of Interpeptide Cross-links: Application to Prion Proteins*

    PubMed Central

    Petrotchenko, Evgeniy V.; Serpa, Jason J.; Hardie, Darryl B.; Berjanskii, Mark; Suriyamongkol, Bow P.; Wishart, David S.; Borchers, Christoph H.

    2012-01-01

    Chemical cross-linking combined with mass spectrometry is a rapidly developing technique for structural proteomics. Cross-linked proteins are usually digested with trypsin to generate cross-linked peptides, which are then analyzed by mass spectrometry. The most informative cross-links, the interpeptide cross-links, are often large in size, because they consist of two peptides that are connected by a cross-linker. In addition, trypsin targets the same residues as amino-reactive cross-linkers, and cleavage will not occur at these cross-linker-modified residues. This produces high molecular weight cross-linked peptides, which complicates their mass spectrometric analysis and identification. In this paper, we examine a nonspecific protease, proteinase K, as an alternative to trypsin for cross-linking studies. Initial tests on a model peptide that was digested by proteinase K resulted in a family of related cross-linked peptides, all of which contained the same cross-linking sites, thus providing additional verification of the cross-linking results, as was previously noted for other post-translational modification studies. The procedure was next applied to the native (PrPC) and oligomeric form of prion protein (PrP?). Using proteinase K, the affinity-purifiable CID-cleavable and isotopically coded cross-linker cyanurbiotindipropionylsuccinimide and MALDI-MS cross-links were found for all of the possible cross-linking sites. After digestion with proteinase K, we obtained a mass distribution of the cross-linked peptides that is very suitable for MALDI-MS analysis. Using this new method, we were able to detect over 60 interpeptide cross-links in the native PrPC and PrP? prion protein. The set of cross-links for the native form was used as distance constraints in developing a model of the native prion protein structure, which includes the 90124-amino acid N-terminal portion of the protein. Several cross-links were unique to each form of the prion protein, including a Lys185Lys220 cross-link, which is unique to the PrP? and thus may be indicative of the conformational change involved in the formation of prion protein oligomers. PMID:22438564

  11. Ultraviolet cross-linking of helical oligonucleotides to two monoclonal MRL-1pr/1pr anti-DNA autoantibodies. Variations in H and L chain binding to DNA

    SciTech Connect

    Jang, Y.J.; Stollar, B.D. )

    1990-11-15

    Experiments were performed to determine whether both H and L chains of different anti-native DNA autoantibodies are uniformly involved in binding to DNA. Two purified monoclonal mouse (MRL-1pr/1pr) IgG autoantibodies, H241 and 2C10, were tested. They both bound synthetic helical oligonucleotides of 10 to 20 base pairs in a gel electrophoresis retardation assay but differed in their preferences for given base sequences. Exposure of antibody-radiolabeled oligonucleotide mixtures to UV light (254 nm) for 10 min led to specific covalent cross-linking of oligonucleotide to both the H and the L chains of H241 but only to the H chain of 2C10. Single labeling events were detected without higher aggregation. The oligonucleotides were not cross-linked to unrelated IgG, even after 2 h of irradiation. Cross-linked (radioactively labeled) H and L chains of H241 and 2C10 were isolated from denaturing electrophoresis gels and digested with lysyl endopeptidase and/or staphylococcal V8 protease. H241 and 2C10 H chains each yielded a major labeled peptide fragment, but the peptides from the two antibodies were different. These experiments measured only some of the antibody-DNA interactions, probably with bases in the major groove of the DNA. They indicated that two MRL-1pr/1pr IgG anti-native DNA antibodies differ in their H and L chain contacts with DNA and provide an approach to identifying affinity-labeled binding sites in the antibodies.

  12. Photo-cross-linked PLA-PEO-PLA hydrogels from self-assembled physical networks: mechanical properties and influence of assumed constitutive relationships.

    PubMed

    Sanabria-DeLong, Naomi; Crosby, Alfred J; Tew, Gregory N

    2008-10-01

    Poly(lactide)-block-poly(ethylene oxide)-block-poly(lactide) (PLA-PEO-PLA) triblock copolymers are known to form physical hydrogels in water as a result of the polymer's amphiphilicity. Their mechanical properties, biocompatibility, and biodegradability have made them attractive for use as soft tissue scaffolds. However, the network junction points are not covalently cross-linked, and in a highly aqueous environment these hydrogels adsorb more water, transform from gel to sol, and lose the designed mechanical properties. In this article, a hydrogel was formed by the use of a novel two-step approach. In the first step, the end-functionalized PLA-PEO-PLA triblock was self-assembled into a physical hydrogel through hydrophobic micelle network junctions, and in the second step, this self-assembled physical network structure was locked into place by photo-cross-linking the terminal acrylate groups. In contrast with physical hydrogels, the photo-cross-linked gels remained intact in phosphate-buffered solution at body temperature. The swelling, degradation, and mechanical properties were characterized, and they demonstrated an extended degradation time (approximately 65 days), an exponential decrease in modulus with degradation time, and a tunable shear modulus (1.6-133 kPa). We also discuss the various constitutive relationships (Hookean, neo-Hookean, and Mooney-Rivlin) that can be used to describe the stress-strain behavior of these hydrogels. The chosen model and assumptions used for data fitting influenced the obtained modulus values by as much as a factor of 3.5, which demonstrates the importance of clearly stating one's data fitting parameters so that accurate comparisons can be made within the literature. PMID:18817440

  13. Baking Performance of Phosphorylated Cross-Linked Resistant Starch in Low-Moisture Bakery Goods

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Phosphorylated cross-linked resistant starch (RS) is a type 4 RS, which can be used for enhancing the benefits of dietary fiber. The baking performance of the RS was explored using wire-cut cookie baking and benchtop chemically-leavened cracker baking methods to produce low-moisture baked goods (coo...

  14. Transglutaminase C in cerebellar granule neurons: regulation and localization of substrate cross-linking.

    PubMed

    Perry, M J; Mahoney, S A; Haynes, L W

    1995-04-01

    Covalent cross-linking of cell surface proteins by the calcium-dependent enzyme transglutaminase C may be implicated in cell-cell interactions and growth regulation. We demonstrate the presence of the enzyme in rat cerebellar cortex during postnatal development. Transglutaminase C was induced in cerebellar granule neurons in culture by retinoic acid, dibutyryl- and 8-bromo-cyclic AMP analogues and by cultivation on a biomatrix substratum. Cyclic AMP analogues stimulated transglutaminase activity in protein synthesis-dependent and -independent phases. The enzyme was distributed at focal adhesion sites on the axon. By calcium-dependent covalent incorporation of the primary amine acceptor substrate, 5-(biotinamido)pentylamine, an increase in the Ca(2+)-dependent cross-linking of at least 11 substrate proteins in the presence of retinoic acid and dibutyryl-cyclic AMP was detected. Of these substrates, a subset was labelled on the surface of living granule neurons. A low-molecular-weight substrate, p18, was tentatively identified as the retinoic acid-inducible neurite-promoting factor, midkine. Transglutaminase-mediated amine incorporation, midkine and isopeptide cross-links were co-localized to axonal adhesion sites. The results provide evidence of transglutaminase C-catalysed protein cross-linking activity in cerebellar granule neurons and its possible implication in cell-substratum interactions. PMID:7617162

  15. CHEMISTRY AND PHYSICAL PROPERTIES OF MELT PROCESSED- AND SOLUTION-CROSS LINKED CORN ZEIN

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Corn zein was cross linked with glutaraldehyde (GDA) and with glacial acetic acid (HAc) as catalyst with the objective to enhance the mechanical properties of poured films which were compared with the physical properties of compression molded tensile bars from melt processed zein with GDA. A reacti...

  16. Is dialdehyde starch a valuable cross-linking agent for collagen/elastin based materials?

    PubMed

    Skopinska-Wisniewska, J; Wegrzynowska-Drzymalska, K; Bajek, A; Maj, M; Sionkowska, A

    2016-04-01

    Collagen and elastin are the main structural proteins in mammal bodies. They provide mechanical support, strength, and elasticity to various organs and tissues, e.g. skin, tendons, arteries, and bones. They are readily available, biodegradable, biocompatible and they stimulate cell growth. The physicochemical properties of collagen and elastin-based materials can be modified by cross-linking. Glutaraldehyde is one of the most efficient cross-linking agents. However, the unreacted molecules can be released from the material and cause cytotoxic reactions. Thus, the aim of our work was to investigate the influence of a safer, macromolecular cross-linking agent-dialdehyde starch (DAS). The properties of hydrogels based on collagen/elastin mixtures (95/5, 90/10) containing 5 and 10 % of DAS and neutralized via dialysis against deionized water were tested. The homogenous, transparent, stiff hydrogels were obtained. The DAS addition causes the formation of intermolecular cross-linking bonds but does not affect the secondary structure of the proteins. As a result, the thermal stability, mechanical strength, and, surprisingly, swelling ability increased. At the same time, the surface properties test and in vitro study show that the materials are attractive for 3T3 cells. Moreover, the materials containing 10 % of DAS are more resistant to enzymatic degradation. PMID:26886815

  17. Cross-Linked Nanoporous Materials from Reactive and Multifunctional Block Polymers

    SciTech Connect

    Seo, Myungeun; Amendt, Mark A.; Hillmyer, Marc A.

    2012-10-10

    Polylactide-b-poly(styrene-co-2-hydroxyethylmethacrylate) (PLA-b-P(S-co-HEMA)) and polylactide-b-poly(styrene-co-2-hydroxyethylacrylate) (PLA-b-P(S-co-HEA)) were synthesized by combination of ring-opening polymerization and reversible addition-fragmentation chain transfer polymerization. {sup 1}H nuclear magnetic resonance spectroscopy and size exclusion chromatography data indicated that the polymerizations were controlled and that hydroxyl groups were successfully incorporated into the block polymers. The polymers were reacted with 4,4{prime}-methylenebis(phenyl isocyanate) (MDI) to form the corresponding cross-linked materials. The materials were annealed at 150 C to complete the coupling reaction. Robust nanoporous materials were obtained from the cross-linked polymers by treatment with aqueous base to hydrolyze the PLA phase. Small-angle X-ray scattering study combined with scanning electron microscopy showed that MDI-cross-linked PLA-b-P(S-co-HEMA)/PLA-b-P(S-co-HEA) can adopt lamellar, hexagonally perforated lamellar, and hexagonally packed cylindrical morphologies after annealing. In particular, the HPL morphology was found to evolve from lamellae due to increase in volume fraction of PS phase as MDI reacted with hydroxyl groups. The reaction also kinetically trapped the morphology by cross-linking. Bicontinuous morphologies were also observed when dibutyltin dilaurate was added to accelerate reaction between the polymer and MDI.

  18. Water Processable Polythiophene Nanowires by Photo-Cross-Linking and Click-Functionalization.

    PubMed

    Kim, Hyeong Jun; Skinner, Matthew; Yu, Hojeong; Oh, Joon Hak; Briseno, Alejandro L; Emrick, Todd; Kim, Bumjoon J; Hayward, Ryan C

    2015-09-01

    Replacing or minimizing the use of halogenated organic solvents in the processing and manufacturing of conjugated polymer-based organic electronics has emerged as an important issue due to concerns regarding toxicity, environmental impact, and high cost. To date, however, the processing of well-ordered conjugated polymer nanostructures has been difficult to achieve using environmentally benign solvents. In this work, we report the development of water and alcohol processable nanowires (NWs) with well-defined crystalline nanostructure based on the solution assembly of azide functionalized poly(3-hexylthiophene) (P3HT-azide) and subsequent photo-cross-linking and functionalization of these NWs. The solution-assembled P3HT-azide NWs were successfully cross-linked by exposure to UV light, yielding good thermal and chemical stability. Residual azide units on the photo-cross-linked NWs were then functionalized with alkyne terminated polyethylene glycol (PEG-alkyne) using copper catalyzed azide-alkyne cycloaddition chemistry. PEG functionalization of the cross-linked P3HT-azide NWs allowed for stable dispersion in alcohols and water, while maintaining well-ordered NW structures with electronic properties suitable for the fabrication of organic field effect transistors (OFETs). PMID:26289785

  19. 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 pressurestrain 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

  20. Effect of cross-linking ultrahigh molecular weight polyethylene: Surface molecular orientation and wear characteristics

    SciTech Connect

    Sambasivan, Sharadha; Fischer, Daniel A.; Hsu, Stephen M.

    2007-07-15

    Molecular orientation at the surface layer of cross-linked ultrahigh molecular weight polyethylene (UHMWPE) has been examined. Molecular orientation has been shown to affect the wear resistance and surface mechanical properties of UHMWPE under biomechanical loading conditions. This study utilizes a nondestructive synchrotron based soft x-ray technique; near edge x-ray absorption fine structure at the carbon K-edge to examine the degree of surface molecular orientation of UHMWPE subjected to various cross-linking/sterilization techniques as a function of stress and wear. UHMWPE samples prepared under gamma irradiation, ethylene-oxide (EtO) treatment, and electron beam irradiation were worn in a wear tester systematically. Results suggest that the cross-linking resists surface orientation when the samples were under tensile and biomechanical stresses. The molecular orientation in the C-C chains in the polymer showed a monotonic decrease with an increase in gamma irradiation dosage levels. EtO sterilized samples showed more C-C chain orientation than the electron beam irradiated samples, but lower than the 30 kGy gamma irradiated samples. Ordered C-C chains in UHMWPE samples have been associated with more crystallinity or large strain plastic deformation of the polymer. Higher levels of gamma irradiation appear to induce cross-linking of C-C chains and render a polymer with more amorphous phase which resists orientation after wear and imparts wear resistance to the polymer.

  1. A robust and reproducible procedure for cross-linking thermoset polymers using molecular simulation.

    PubMed

    Demir, Baris; Walsh, Tiffany R

    2016-02-17

    Molecular simulation can provide valuable guidance in establishing clear links between structure and function to enable the design of new polymer-based materials. However, molecular simulation of thermoset polymers in particular, such as epoxies, present specific challenges, chiefly in the credible preparation of polymerised samples. Despite this need, a comprehensive, reproducible and robust process for accomplishing this using molecular simulation is still lacking. Here, we introduce a clear and reproducible cross-linking protocol to reliably generate three dimensional epoxy cross-linked polymer structures for use in molecular simulations. This protocol is sufficiently detailed to allow complete reproduction of our results, and is applicable to any general thermoset polymer. Amongst our developments, key features include a reproducible procedure for calculation of partial atomic charges, a reliable process for generating and validating an equilibrated liquid precursor mixture, and establishment of a novel, robust and reproducible protocol for generating the three-dimensional cross-linked solid polymer. We use these structures as input to subsequent molecular dynamics simulations to calculate a range thermo-mechanical properties, which compare favourably with experimental data. Our general protocol provides a benchmark for the process of simulating epoxy polymers, and can be readily translated to prepare and model epoxy samples that are dynamically cross-linked in the presence of surfaces and nanostructures. PMID:26822527

  2. Temperature dependence of creep compliance of highly cross-linked epoxy: A molecular simulation study

    SciTech Connect

    Khabaz, Fardin Khare, Ketan S. Khare, Rajesh

    2014-05-15

    We have used molecular dynamics (MD) simulations to study the effect of temperature on the creep compliance of neat cross-linked epoxy. Experimental studies of mechanical behavior of cross-linked epoxy in literature commonly report creep compliance values, whereas molecular simulations of these systems have primarily focused on the Young’s modulus. In this work, in order to obtain a more direct comparison between experiments and simulations, atomistically detailed models of the cross-linked epoxy are used to study their creep compliance as a function of temperature using MD simulations. The creep tests are performed by applying a constant tensile stress and monitoring the resulting strain in the system. Our results show that simulated values of creep compliance increase with an increase in both time and temperature. We believe that such calculations of the creep compliance, along with the use of time temperature superposition, hold great promise in connecting the molecular insight obtained from molecular simulation at small length- and time-scales with the experimental behavior of such materials. To the best of our knowledge, this work is the first reported effort that investigates the creep compliance behavior of cross-linked epoxy using MD simulations.

  3. DNA-Protein Cross-Linking by 1,2,3,4-Diepoxybutane

    PubMed Central

    Michaelson-Richie, Erin D.; Loeber, Rachel L.; Codreanu, Simona G.; Ming, Xun; Liebler, Daniel C.; Campbell, Colin; Tretyakova, Natalia Y.

    2010-01-01

    1,2,3,4-Diepoxybutane (DEB) is a strongly genotoxic diepoxide hypothesized to be the ultimate carcinogenic metabolite of the common industrial chemical and environmental carcinogen 1,3-butadiene. DEB is a bis-electrophile capable of cross-linking cellular biomolecules to form DNA-DNA and DNA-protein cross-links (DPCs), which are thought to play a central role in its biological activity. Previous studies with recombinant proteins have shown that the biological outcomes of DEB-induced DPCs are strongly influenced by protein identities. The present work combines affinity capture methodology with mass spectrometry-based proteomics and immunological detection to identify the proteins which form DPCs in nuclear extracts from human cervical carcinoma (HeLa) cells. We identified 39 human proteins that form covalent DPCs in the presence of DEB. DNA-protein cross-linking efficiency following treatment with 25 mM DEB was 212%, depending on protein identity. HPLC-ESI+-MS/MS analysis of the total proteolytic digests of cross-linked proteins revealed the presence of 1-(S-cysteinyl)-4-(guan-7-yl)-2,3-butanediol conjugates, suggesting that DEB forms DPCs between cysteine thiols within proteins and the N-7 guanine positions within DNA. PMID:20666492

  4. Identification of disulfide cross-linked tau dimer responsible for tau propagation

    PubMed Central

    Kim, Dohee; Lim, Sungsu; Haque, Md. Mamunul; Ryoo, Nayeon; Hong, Hyun Seok; Rhim, Hyewhon; Lee, Dong-Eun; Chang, Young-Tae; Lee, Jun-Seok; Cheong, Eunji; Kim, Dong Jin; Kim, Yun Kyung

    2015-01-01

    Recent evidence suggests that tau aggregates are not only neurotoxic, but also propagate in neurons acting as a seed for native tau aggregation. Prion-like tau transmission is now considered as an important pathogenic mechanism driving the progression of tau pathology in the brain. However, prion-like tau species have not been clearly characterized. To identify infectious tau conformers, here we prepared diverse tau aggregates and evaluated the effect on inducing intracellular tau-aggregation. Among tested, tau dimer containing P301L-mutation is identified as the most infectious form to induce tau pathology. Biochemical analysis reveals that P301L-tau dimer is covalently cross-linked with a disulfide bond. The relatively small and covalently cross-linked tau dimer induced tau pathology efficiently in primary neurons and also in tau-transgenic mice. So far, the importance of tau disulfide cross-linking has been overlooked in the study of tau pathology. Here our results suggested that tau disulfide cross-linking might play critical role in tau propagation by producing structurally stable and small tau conformers. PMID:26470054

  5. Energy dissipation and recovery in a simple model with reversible cross-links

    NASA Astrophysics Data System (ADS)

    Nabavi, S. Soran; Fratzl, Peter; Hartmann, Markus A.

    2015-03-01

    Reversible cross-linking is a method of enhancing the mechanical properties of polymeric materials. The inspiration for this kind of cross-linking comes from nature, which uses this strategy in a large variety of biological materials to dramatically increase their toughness. Recently, first attempts were made to transfer this principle to technological applications. In this study, Monte Carlo simulations are used to investigate the effect of the number and the topology of reversible cross-links on the mechanical performance of a simple model system. Computational cyclic loading tests are performed, and the work to fracture and the energy dissipation per cycle are determined, which both increase when the density of cross-links is increased. Furthermore, a different topology of the bonds may increase the work to fracture by a factor of more than 2 for the same density. This dependence of the mechanical properties on the topology of the bonds has important implications on the self-healing properties of such systems, because only a fast return of the system to its unloaded state after release of the load ensures that the optimal topology may form.

  6. Can para-aryl-dithiols cross-link two plasmonic noble nanoparticles as monolayer dithiolate spacers

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Para-aryl-dithiols (PADTs, HS-(C6H4)n-SH, n = 1, 2, and 3) have been used extensively in molecular electronics, surface-enhanced Raman spectroscopy (SERS), and quantum electron tunneling between two gold or silver nanoparticles (AuNPs and AgNPs). One popular belief is that these dithiols cross-link ...

  7. Self-assembly of biopolymeric structures below the threshold of random cross-link percolation.

    PubMed Central

    San Biagio, P L; Bulone, D; Emanuele, A; Palma, M U

    1996-01-01

    Self-assembly of extended structures via cross-linking of individual biomolecules often occurs in solutions at concentrations well below the estimated threshold for random cross-link percolation. This requires solute-solute correlations. Here we study bovine serum albumin. Its unfolding causes the appearance of an instability region of the sol, not observed for native bovine serum albumin. As a consequence, spinodal demixing of the sol is observed. The thermodynamic phase transition corresponding to this demixing is the determinative symmetry-breaking step allowing the subsequent occurrence of (correlated) cross-linking and its progress up to the topological phase transition of gelation. The occurrence of this sequence is of marked interest to theories of spontaneous symmetry-breaking leading to morphogenesis, as well as to percolation theories. The present results extend the validity of conclusions drawn from our previous studies of other systems, by showing in one single case, system features that we have hitherto observed separately in different systems. Time-resolved experimental observations of the present type also bring kinetic and diffusional processes and solute-solvent interactions into the picture of cross-link percolation. Images FIGURE 3 PMID:8770227

  8. Effect of dual modification with hydroxypropylation and cross-linking on physicochemical properties of taro starch.

    PubMed

    Hazarika, Bidyut Jyoti; Sit, Nandan

    2016-04-20

    Dual modification of taro starch by hydroxypropylation and cross-linking was carried out and the properties of the modified starches were investigated. Two different levels of hydroxypropylation (5 and 10%) and cross-linking (0.05 and 0.10%) were used in different sequences. The amylose contents of the starch decreased due to single and dual modification. For the dual-modified starches, the swelling, solubility and clarity was found to increase with level of hydroxypropylation and decrease with level of cross-linking. The freeze-thaw stability of the dual-modified starches was also affected by the sequence of modification. The viscosities of the cross-linked and dual-modified starches were more than native and hydroxypropylated starches. The firmness of the dual-modified starches was also higher than native and single modified starches. The dual-modified starches have benefits of both type of modifications and could be used for specific purposes e.g. food products requiring high viscosity as well as freeze-thaw stability. PMID:26876854

  9. Endogenous and enhanced oxidative cross-linking in wheat flour mill streams

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The oxidative cross-linking of arabinoxylan and protein polymers is partially responsible for variation in end-use quality of wheat flour; specifically, differences in batter viscosity as well as variation in bread and cookie quality. A better understanding of the variation in oxidative cross-linkin...

  10. Light-scattering thermal cross-linking material using morphology of nanoparticle free polymer blends

    NASA Astrophysics Data System (ADS)

    Takei, Satoshi

    2015-03-01

    A newly light-scattering thermal cross-linking material based on self-assembly for forming the morphology of nanoparticle free polymer blends was reported. The material design concept to use light-scattering thermal cross-linking material with high uniformity of light on display panel from LED for high quality such as brightness and evenness, mechanical properties, and gas and water barrier properties. The high light scattering rate of 8 % at 350-450 nm of wavelength, fast cure film at 140 ºC and 120 s, and thermal stability at 190 ºC in bake condition for high productivity were indicated in the light-scattering thermal cross-linking material using the nanoparticle free polymers with carboxylic acid functional groups. These novel system using morphology of nanoparticle free polymer blends in light-scattering package material for a LCD using LED was a valuable approach to the design of material formulations for newly light-scattering thermal cross-linking material.

  11. Reversible Inter- and Intra-Microgel Cross-Linking using Disulfides

    PubMed Central

    Gaulding, Jeffrey C.; Smith, Michael H.; Hyatt, John S.; Fernandez-Nieves, Alberto; Lyon, L. Andrew

    2012-01-01

    Thermoresponsive hydrogel nanoparticles composed of poly(N-isopropylmethacrylamide) (pNIPMAm) and the disulfide-based cross-linker N,N’-bis(acryloyl)cystamine (BAC) have been prepared using a redox-initiated, aqueous precipitation polymerization approach, leading to improved stability of the disulfide bond compared to traditional thermally-initiated methods. The resultant particles demonstrate complete erosion in response to reducing conditions or thiol competition. This stands in contrast to the behavior of thermally-initiated particles, which retain a cross-linked network following disulfide cleavage due to uncontrolled chain-branching and self-cross-linking side reactions. The synthetic strategy has also been combined with the non-degradable cross-linker N,N-methylenebisacrylamide (BIS) to generate “co-cross-linked” pNIPMAm-BAC-BIS microgels. These particles are redox-responsive, swell upon BAC cross-link scission and present reactive thiols. This pendant thiol functionality was demonstrated to be useful for conjugation of thiol-reactive probes and in reversible network formation by assembling particles cross-linked by disulfide linkages. PMID:22287810

  12. In-vitro fermentability of cell walls as influenced by lignin composition and cross-linking.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We assessed how diverse modifications in lignin composition and reductions in ferulate-lignin cross-linking influence the degradability of cell walls. Cell walls from nonlignified maize cell suspensions were artificially lignified with varying ratios of normal monolignols (coniferyl and sinapyl alco...

  13. Cross-linking oppositely charged oil-in-water emulsions to enhance heteroaggregate stability.

    PubMed

    Maier, Christiane; Oechsle, Anja M; Weiss, Jochen

    2015-11-01

    The formation and subsequent enzymatic and chemical cross-linking of heteroaggregates from oppositely charged oil-in-water (O/W) emulsions was investigated. For this purpose, 10% (w/w) oil-in-water emulsions (d43<1?m) were prepared at pH 4 using a positively charged emulsifier (N?-lauroyl-L-arginine ethyl ester (LAE), cold water fish gelatin, or whey protein isolate) or a negatively charged one (sugar beet pectin or Quillaja saponins). The oppositely charged emulsions were then combined at a volume ratio of 1:1 and treated with laccase or glutaraldehyde in order to further stabilize the electrostatically attached aggregates by covalently cross-linking the oppositely charged membranes. Emulsions and heteroaggregates were characterized by their rheological properties, their surface charge, particle size distribution, and microstructure using dynamic and static light scattering as well as confocal laser scanning microscopy. Prior to cross-linking, the emulsifiers' stabilization mechanism were found to greatly influence the formation of heteroaggregates. Laccase treatment (1.34mU/mL) increased aggregate expansion by ca. 30% for the combined emulsions stabilized by Quillaja saponins/whey protein isolate, while combined Quillaja saponins/fish gelatin stabilized emulsions remained unaffected. When combined emulsions were treated with 50mM glutaraldehyde, aggregate size significantly increased 2- and 3-fold, respectively. Thus, our study provides novel insights into the enzymatic and chemical cross-linking of heteroaggregates composed of oppositely charged O/W emulsions. PMID:26298085

  14. Tip60 Is Required for DNA Interstrand Cross-link Repair in the Fanconi Anemia Pathway*

    PubMed Central

    Hejna, James; Holtorf, Megan; Hines, Jennie; Mathewson, Lauren; Hemphill, Aaron; Al-Dhalimy, Muhsen; Olson, Susan B.; Moses, Robb E.

    2008-01-01

    The disease Fanconi anemia is a genome instability syndrome characterized by cellular sensitivity to DNA interstrand cross-linking agents, manifest by decreased cellular survival and chromosomal aberrations after such treatment. There are at least 13 proteins acting in the pathway, with the FANCD2 protein apparently functioning as a late term effecter in the maintenance of genome stability. We find that the chromatin remodeling protein, Tip60, interacts directly with the FANCD2 protein in a yeast two-hybrid system. This interaction has been confirmed by co-immunoprecipitation and co-localization using both endogenous and epitope-tagged FANCD2 and Tip60 from human cells. The observation of decreased cellular survival after exposure to mitomycin C in normal fibroblasts depleted for Tip60 indicates a direct function in interstrand cross-link repair. The coincident function of Tip60 and FANCD2 in one pathway is supported by the finding that depletion of Tip60 in Fanconi anemia cells does not increase sensitivity to DNA cross-links. However, depletion of Tip60 did not reduce monoubiquitination of FANCD2 or its localization to nuclear foci following DNA damage. The observations indicate that Fanconi anemia proteins act in concert with chromatin remodeling functions to maintain genome stability after DNA cross-link damage. PMID:18263878

  15. ROMP-based thermosetting polymers from modified castor oil with various cross-linking agents

    NASA Astrophysics Data System (ADS)

    Ding, Rui

    Polymers derived from bio-renewable resources are finding an increase in global demand. In addition, polymers with distinctive functionalities are required in certain advanced fields, such as aerospace and civil engineering. In an attempt to meet both these needs, the goal of this work aims to develop a range of bio-based thermosetting matrix polymers for potential applications in multifunctional composites. Ring-opening metathesis polymerization (ROMP), which recently has been explored as a powerful method in polymer chemistry, was employed as a unique pathway to polymerize agricultural oil-based reactants. Specifically, a novel norbornyl-functionalized castor oil alcohol (NCA) was investigated to polymerize different cross-linking agents using ROMP. The effects of incorporating dicyclopentadiene (DCPD) and a norbornene-based crosslinker (CL) were systematically evaluated with respect to curing behavior and thermal mechanical properties of the polymers. Isothermal differential scanning calorimetry (DSC) was used to investigate the conversion during cure. Dynamic DSC scans at multiple heating rates revealed conversion-dependent activation energy by Ozawa-Flynn-Wall analysis. The glass transition temperature, storage modulus, and loss modulus for NCA/DCPD and NCA/CL copolymers with different cross-linking agent loading were compared using dynamic mechanical analysis. Cross-link density was examined to explain the very different dynamic mechanical behavior. Mechanical stress-strain curves were developed through tensile test, and thermal stability of the cross-linked polymers was evaluated by thermogravimetric analysis to further investigate the structure-property relationships in these systems.

  16. Spine fusion cross-link causing delayed dural erosion and CSF leak: case report.

    PubMed

    Rahmathulla, Gazanfar; Deen, H Gordon

    2015-04-01

    The past 2 decades have seen a considerable increase in the number of lumbar spinal fusion surgeries. To enhance spinal stabilization and fusion, make the construct resistant to or stiffer for axial stress loading, lateral bending, and torsional stresses, cross-links and connectors were designed and included in a rod-screw construct. The authors present the case of a 49-year-old woman who presented 11 years after undergoing an L4-5 decompression and fusion in which a pedicle screw-rod construct with an integrated cross-link was designed to attach onto the pedicle screws. The patient's response at the time to the initial surgery was excellent; however, at the time of presentation 11 years later, she had significant postural headaches, severe neurogenic claudication, and radiculopathy. Imaging revealed canal compression across the instrumented levels and a possible thickened adherent filum terminale. Reexploration of the level revealed a large erosive dural defect with a CSF leak, spinal canal compression, and a thickened filum at the level of the cross-link. To the author's knowledge, such complications have not been reported in literature. The authors discuss this rare complication of spinal fusion and the need to avoid dural compression when cross-links are used. PMID:25635637

  17. Preferential sites for intramolecular glucosepane cross-link formation in type I collagen: A thermodynamic study

    PubMed Central

    Collier, Thomas A.; Nash, Anthony; Birch, Helen L.; de Leeuw, Nora H.

    2015-01-01

    The extracellular matrix (ECM) undergoes progressive age-related stiffening and loss of proteolytic digestibility due to an increase in concentration of advanced glycation end products (AGEs). The most abundant AGE, glucosepane, accumulates in collagen with concentrations over 100 times greater than all other AGEs. Detrimental collagen stiffening properties are believed to play a significant role in several age-related diseases such as osteoporosis and cardiovascular disease. Currently little is known of the potential location of covalently cross-linked glucosepane formation within collagen molecules; neither are there reports on how the respective cross-link sites affect the physical and biochemical properties of collagen. Using fully atomistic molecular dynamics simulations (MD) we have identified six sites where the formation of a covalent intra-molecular glucosepane cross-link within a single collagen molecule in a fibrillar environment is energetically favourable. Identification of these favourable sites enables us to align collagen cross-linking with experimentally observed changes to the ECM. For example, formation of glucosepane was found to be energetically favourable within close proximity of the Matrix Metalloproteinase-1 (MMP1) binding site, which could potentially disrupt collagen degradation. PMID:26049074

  18. Effect of the cross-linking density on the thermoresponsive behavior of hollow PNIPAM microgels.

    PubMed

    Contreras-Cceres, Rafael; Schellkopf, Leonard; Fernndez-Lpez, Cristina; Pastoriza-Santos, Isabel; Prez-Juste, Jorge; Stamm, Manfred

    2015-01-27

    We report on the fabrication of thermally responsive hollow pNIPAM particles through the oxidation of the metal core in an Au@pNIPAM system. The selective oxidation of the Au core is achieved by addition of AuCl4(-) to an aqueous dispersion of Au@pNIPAM particles in the presence of cetyltrimethylammonium bromide (CTAB). We fabricate hollow pNIPAM particles with three cross-linking densities (N,N'-methylenebis(acrylamide), BA, at 5%, 10%, and 17.5%). The study of the effect of the amount of BA within the microgel network was performed by dynamic light scattering (DLS), transmission electron microscopy (TEM), and atomic force microscopy (AFM), showing its key role in determining the final hollow structure and thermal response. While the thermal responsiveness is largely achieved at low cross-linking densities, the hollow structure only remains at larger cross-linking densities. This was further confirmed by cryo-TEM analysis of hollow pNIPAM particles below and above the volume phase transition temperature (VPTT). Thus, it clearly shows (i) the shrinking of particle size with the temperature at low cross-linking density and (ii) the dependence of particle size on the amount of cross-linker for the final hollow pNIPAM structure. Observed differences in the hollow pNIPAM structure are attributed to different elastic contributions (?(elas)), showing higher elasticity for microgels synthesized at lower amount of BA. PMID:25526382

  19. Cross-linked smart poly(dimethylsiloxane) membranes for removal of volatile organic compounds in water

    NASA Astrophysics Data System (ADS)

    Ohshima, Tadahiro; Miyata, Takashi; Uragami, Tadashi; Berghmens, Hugo

    2005-04-01

    This paper focuses on the effects of fluorine cross-linker of the cross-linked poly(dimethylsiloxane) membranes from polydimethylsiloxane dimethylmethacrylate macromonomer (PDMSDMMA) and divinyl perfluoro- n-hexane (DVF) on the pervaporation characteristics of the removal of benzene from an aqueous solution of dilute benzene. When an aqueous solution of 0.05 wt% benzene was permeated through the cross-linked PDMSDMMA (PDMSDMMA-DVF) membranes, they showed a high benzene permselectivity and permeability of these membranes was enhanced with increasing DVF content significantly. The best normalized permeation rate, separation factor for benzene permselectivity, and pervaporation separation index (PSI) of a PDMSDMMA-DVF membrane were 1.7210 -5 kg m/m 2 h, 4316, and 7423, respectively. The best normalized permeation rate of a PDMSDMMA-DVF membrane was approximately same as the PDMSDMMA membranes cross-linked with other divinyl compounds, but the separation factor and PSI of the former membrane were greater than those of the latter ones. These pervaporation characteristics are discussed from the viewpoint of chemical and physical structure of the cross-linked PDMSDMMA-DVF membranes in detail.

  20. Nodular Epithelial Hyperplasia after Photorefractive Keratectomy Followed by Corneal Collagen Cross-Linking

    PubMed Central

    Salerno, Liberdade Cezaro; Ghanem, Vincius Coral; Ghanem, Ramon Coral

    2013-01-01

    This study describes a case of nodular epithelial hyperplasia and stromal alterations in a patient with keratoconus who was submitted to topography-guided photorefractive keratectomy (PRK) followed by corneal collagen cross-linking. Debridement of the epithelial nodule was performed. After a 2-year followup, a new topography-guided PRK was indicated. PMID:23691390

  1. Effect of different cross-linking methods and processing parameters on drug release from hydrogel beads.

    PubMed

    Mitra, Shataneek; Maity, Siddhartha; Sa, Biswanath

    2015-03-01

    The purpose of this work was to evaluate different methods of cross-linking for developing diltiazem-resin complex loaded carboxymethyl xanthan gum (CMXG) hydrogel beads to achieve highest possible drug entrapment and extended release for effective cardio-protection. The hydrogel beads were prepared by ionic cross-linking and dual cross-linking using simultaneous (SIM) and sequential (SEQ) methods. Among the three methods, SEQ method produced smaller sized beads having higher drug entrapment efficacy and prolonged release characteristics as evidenced from mean dissolution time and diffusion coefficient of drug. Keeping the concentration of ionic cross-linker constant, increase in the amount of covalent cross-linker and cross-linking time decreased the drug release. Higher release of the drug in acid solution was attributed to the higher solubility of the basic drug and higher swelling of the matrices in acid solution. Comparison of FTIR spectra, drug content and dissolution profiles indicated that the drug was stable in the beads when kept under stress condition up to 3 months. In conclusion, the sequential method was found superior for producing CMXG hydrogel beads as a prolonged release delivery device in cardiovascular diseases. PMID:25576745

  2. 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.

  3. Hybrid sol-gel optical materials

    DOEpatents

    Zeigler, John M. (Albuquerque, NM)

    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.

  4. Hybrid sol-gel optical materials

    DOEpatents

    Zeigler, John M. (Albuquerque, NM)

    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.

  5. Monitoring the internal structure of poly(N-vinylcaprolactam) microgels with variable cross-link concentration.

    PubMed

    Schneider, Florian; Balaceanu, Andreea; Feoktystov, Artem; Pipich, Vitaliy; Wu, Yaodong; Allgaier, Jrgen; Pyckhout-Hintzen, Wim; Pich, Andrij; Schneider, Gerald J

    2014-12-23

    The combination of a set of complementary techniques allows us to construct an unprecedented and comprehensive picture of the internal structure, temperature dependent swelling behavior, and the dependence of these properties on the cross-linker concentration of microgel particles based on N-vinylcaprolactam (VCL). The microgels were synthesized by precipitation polymerization using different amounts of cross-linking agent. Characterization was performed by small-angle neutron scattering (SANS) using two complementary neutron instruments to cover a uniquely broad Q-range with one probe. Additionally we used dynamic light scattering (DLS), atomic force microscopy (AFM), and differential scanning calorimetry (DSC). Previously obtained nuclear magnetic resonance spectroscopy (NMR) results on the same PVCL particles are utilized to round the picture off. Our study shows that both the particle radius and the cross-link density and therefore also the stiffness of the microgels rises with increasing cross-linker content. Hence, more cross-linker reduces the swelling capability distinctly. These findings are supported by SANS and AFM measurements. Independent DLS experiments also found the increase in particle size but suggest an unchanged cross-link density. The reason for the apparent contradiction is the indirect extraction of the parameters via a model in the evaluation of DLS measurements. The more direct approach in AFM by evaluating the cross section profiles of observed microgel particles gives evidence of significantly softer and more deformable particles at lower cross-linker concentrations and therefore verifies the change in cross-link density. DSC data indicate a minor but unexpected shift of the volume phase transition temperature (VPTT) to higher temperatures and exposes a more heterogeneous internal structure of the microgels with increasing cross-link density. Moreover, a change in the total energy transfer during the VPT gives evidence that the strength of hydrogen bonds is significantly affected by the cross-link density. A strong and reproducible deviation of the material density of the cross-linked microgel polymer chains toward a higher value compared to the respective linear chains has yet to be explained. PMID:25493607

  6. Tissue-specific distribution of cross-linked somatostatin receptor proteins in the rat.

    PubMed Central

    Srikant, C B; Murthy, K K; Patel, Y C

    1992-01-01

    Pharmacological studies have suggested that the somatostatin (SS) receptor is heterogeneous and exhibits SS-14-and SS-28-selective subtypes. Whether such subtypes arise from molecular heterogeneity of the receptor protein has not been definitively established. Previous reports characterizing the molecular properties of the SS receptor by the cross-linking approach have yielded divergent size estimates ranging from 27 kDa to 200 kDa. In order to resolve this discrepancy, as well as to determine whether SS-14 and SS-28 interact with specific receptor proteins, we have cross-linked radioiodinated derivatives of [125I-Tyr11]SS-14 (T*-SS-14) and [Leu8,D-Trp22,125I-Tyr25]SS-28 (LTT*-SS-28) to membrane SS receptors in rat brain, pituitary, exocrine pancreas and adrenal cortex using a number of chemical and photoaffinity cross-linking agents. The labelled cross-linked receptor proteins were analysed by SDS/PAGE under reducing conditions followed by autoradiography. Our findings indicate that the pattern of specifically labelled cross-linked SS receptor proteins is sensitive to the concentration of chemical cross-linking agents such as disuccinimidyl suberate and dithiobis-(succinimidyl propionate). Labelled high-molecular-mass complexes of cross-linked receptor-ligand proteins were observed only when high concentrations of these cross-linkers were employed. Using optimized low concentrations of cross-linkers, however, two major labelled bands of 58 +/- 3 kDa and 27 +/- 2 kDa were detected. These two bands were identified as specifically labelled SS receptor proteins subsequent to cross-linking with a number of photoaffinity cross-linking agents as well. We demonstrate here that the 58 kDa protein is the major SS receptor protein in the rat pituitary, adrenal and exocrine pancreas, whereas the 27 kDa moiety represents the principal form in the brain. Additionally, the presence of a minor specifically labelled band of 32 kDa was detected uniquely in the brain, and a minor labelled protein of 42 kDa was observed in the pancreas. The labelling pattern obtained with LTT*-SS-28 was identical to that observed with T*-SS-14. Labelling of the 27 kDa band by either ligand was inhibited by SS-14 and SS-28 in a dose-dependent manner. Densitometric quantification showed that SS-14 exhibited greater than 2-fold greater potency than SS-28 for inhibiting the labelling of the 27 kDa species. These findings emphasize the need for careful interpretation of cross-linking data obtained for SS receptors, and provide evidence for molecular heterogeneity and for a tissue-specific distribution of the two principal SS receptor proteins. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. Fig. 5. Fig. 6. PMID:1312325

  7. Enhancement of copolymerization of itaconic acid with N-vinyl 2-pyrrolidone by radiation in the presence of cross-linking agent

    NASA Astrophysics Data System (ADS)

    Şen, M.; Yakar, A.

    2005-06-01

    Enhancement of copolymerization of itaconic acid (IA) with N-vinyl 2-pyrrolidone (VP) by radiation in the presence of cross-linking agent was investigated. Hydrogels with varying IA content were prepared from the ternary systems N-vinyl 2-pyrrolidone/itaconic acid/water by irradiating with gamma-rays in the presence of a chemical cross-linker, ethylene glycol dimethacrylate (EGDMA) at ambient temperature. The incorporation of EGDMA into the ternary mixtures caused an increase in the amount of IA in the gel system up to a mole fraction of 13.7%. Hydrogels showed a typical pH response such as high pH swelling and low pH deswelling. Equilibrium volume swelling ratio at pH 7 was varied from 15-40 with changing the IA content in the gel system and irradiation dose. The equation recently modified by the authors for the determination of M is observed to describe the swelling behaviour of P(VP/IA/EGDMA) networks containing relatively higher amount of charged units very well. In addition to the evaluation of M from swelling data, the measurement of polymer solvent interaction parameter and the determination of the reaction yield of cross-links of the systems were examined.

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

    NASA Technical Reports Server (NTRS)

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

  9. Resistance of Corneal RFUVACross-Linked Collagens and Small Leucine-Rich Proteoglycans to Degradation by Matrix Metalloproteinases

    PubMed Central

    Zhang, Yuntao; Mao, Xiuli; Schwend, Tyler; Littlechild, Stacy; Conrad, Gary W.

    2013-01-01

    Purpose. Extracellular matrix metalloproteinases (MMPs) are thought to play a crucial role in corneal degradation associated with the pathological progression of keratoconus. Currently, corneal cross-linking by riboflavin and ultraviolet A (RFUVA) has received significant attention for treatment of keratoconus. However, the extent to which MMPs digest cross-linked collagen and small leucine-rich proteoglycans (SLRPs) remains unknown. In this study, the resistance of RFUVAcross-linked collagens and SLRPs to MMPs has been investigated. Methods. To investigate the ability of MMPs to digest cross-linked collagen and SLRPs, a model reaction system using purified collagen type I, type IV, and nonglycosylated, commercially available recombinant SLRPs, keratocan, lumican, mimecan, decorin, and biglycan in solution in vitro has been compared using reactions inside an intact bovine cornea, ex vivo. Results. Our data demonstrate that corneal cross-linked collagen type I and type IV are resistant to cleavage by MMP-1, MMP-2, MMP-9, and MMP-13, whereas noncrosslinked collagen I, IV, and natively glycosylated SLRPs are susceptible to degradation by MMPs. In addition, both cross-linked SLRPs themselves and cross-linked polymers of SLRPs and collagen appear able to resist degradation. These results suggest that the interactions between SLRPs and collagen caused by RFUVA protect both SLRPs and collagen fibrils from cleavage by MMPs. Conclusions. A novel approach for understanding the biochemical mechanism whereby RFUVA cross-linking stops keratoconus progression has been achieved. PMID:23322569

  10. Developing functional musculoskeletal tissues through hypoxia and lysyl oxidase-induced collagen cross-linking

    PubMed Central

    Makris, Eleftherios A.; Responte, Donald J.; Hu, Jerry C.; Athanasiou, Kyriacos A.

    2014-01-01

    The inability to recapitulate native tissue biomechanics, especially tensile properties, hinders progress in regenerative medicine. To address this problem, strategies have focused on enhancing collagen production. However, manipulating collagen cross-links, ubiquitous throughout all tissues and conferring mechanical integrity, has been underinvestigated. A series of studies examined the effects of lysyl oxidase (LOX), the enzyme responsible for the formation of collagen cross-links. Hypoxia-induced endogenous LOX was applied in multiple musculoskeletal tissues (i.e., cartilage, meniscus, tendons, ligaments). Results of these studies showed that both native and engineered tissues are enhanced by invoking a mechanism of hypoxia-induced pyridinoline (PYR) cross-links via intermediaries like LOX. Hypoxia was shown to enhance PYR cross-linking 1.4- to 6.4-fold and, concomitantly, to increase the tensile properties of collagen-rich tissues 1.3- to 2.2-fold. Direct administration of exogenous LOX was applied in native cartilage and neocartilage generated using a scaffold-free, self-assembling process of primary chondrocytes. Exogenous LOX was found to enhance native tissue tensile properties 1.9-fold. LOX concentration- and time-dependent increases in PYR content (?16-fold compared with controls) and tensile properties (approximately fivefold compared with controls) of neocartilage were also detected, resulting in properties on par with native tissue. Finally, in vivo subcutaneous implantation of LOX-treated neocartilage in nude mice promoted further maturation of the neotissue, enhancing tensile and PYR content approximately threefold and 14-fold, respectively, compared with in vitro controls. Collectively, these results provide the first report, to our knowledge, of endogenous (hypoxia-induced) and exogenous LOX applications for promoting collagen cross-linking and improving the tensile properties of a spectrum of native and engineered tissues both in vitro and in vivo. PMID:25349395

  11. Role of collagen content and cross-linking in large pulmonary arterial stiffening after chronic hypoxia

    PubMed Central

    Wang, Zhijie; Chesler, Naomi C.

    2011-01-01

    Chronic hypoxic pulmonary hypertension (HPH) is associated with large pulmonary artery (PA) stiffening, which is correlated with collagen accumulation. However, the mechanisms by which collagen contributes to PA stiffening remain largely unexplored. Moreover, HPH may alter mechanical properties other than stiffness, such as pulse damping capacity, which also affects ventricular workload but is rarely quantified. We hypothesized that collagen content and cross-linking differentially regulate the stiffness and damping capacity of large PAs during HPH progression. The hypothesis was tested with transgenic mice that synthesize collagen type I resistant to collagenase degradation (Col1a1R/R). These mice and littermate controls (Col1a1+/+) were exposed to hypoxia for 10 days; some were treated with ?-animopropionitrile (BAPN), which prevents new cross-link formation. Isolated PA dynamic mechanical tests were performed and collagen content and cross-linking were measured. In Col1a1+/+ mice, HPH increased both collagen content and cross-linking and BAPN treatment prevented these increases. Similar trends were observed in Col1a1R/R mice except that collagen content further increased with BAPN treatment. Mechanical tests showed that in Col1a1+/+ mice, HPH increased PA stiffness and damping capacity and these increases were impeded by BAPN treatment. In Col1a1R/R mice, HPH led to a smaller but significant increase in PA stiffness and a decrease in damping capacity. These mechanical changes were not affected by BAPN treatment. Vessel-specific correlations for each strain showed that the stiffness and damping capacity were correlated with the total content rather than cross-linking of collagen. Our results suggest that collagen total content is critical to extralobar PA stiffening during HPH. PMID:21538012

  12. Amino Acid Proximities in Two Sup35 Prion Strains Revealed by Chemical Cross-linking.

    PubMed

    Wong, Shenq-Huey; King, Chih-Yen

    2015-10-01

    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 (14)N- and (15)N-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

  13. Acoustic Radiation Force for Noninvasive Evaluation of Corneal Biomechanical Changes Induced by Cross-linking Therapy

    PubMed Central

    Urs, Raksha; Lloyd, Harriet O.; Silverman, Ronald H.

    2015-01-01

    Objectives To noninvasively measure changes in corneal biomechanical properties induced by ultraviolet-activated riboflavin cross-linking therapy using acoustic radiation force (ARF). Methods Cross-linking was performed on the right eyes of 6 rabbits, with the left eyes serving as controls. Acoustic radiation force was used to assess corneal stiffness before treatment and weekly for 4 weeks after treatment. Acoustic power levels were within US Food and Drug Administration guidelines for ophthalmic safety. Strain, determined from ARF-induced displacement of the front and back surfaces of the cornea, was fit to the Kelvin-Voigt model to determine the elastic modulus (E) and coefficient of viscosity (?). The stiffness factor, the ratio of E after treatment to E before treatment, was calculated for treated and control eyes. At the end of 4 weeks, ex vivo thermal shrinkage temperature analysis was performed for comparison with in vivo stiffness measurements. One-way analysis of variance and Student t tests were performed to test for differences in E, ?, the stiffness factor, and corneal thickness. Results Biomechanical stiffening was immediately evident in cross-linkingtreated corneas. At 4 weeks after treatment, treated corneas were 1.3 times stiffer and showed significant changes in E(P= .006) and ? (P= .007), with no significant effect in controls. Corneal thickness increased immediately after treatment but did not differ significantly from the pretreatment value at 4 weeks. Conclusions Our findings demonstrate a statistically significant increase in stiffness in cross-linkingtreated rabbit corneas based on in vivo axial stress/strain measurements obtained using ARF. The capacity to noninvasively monitor corneal stiffness offers the potential for clinical monitoring of cross-linking therapy. PMID:25063407

  14. Expression of Hyaluronan in human tumor progression

    PubMed Central

    Boregowda, Rajeev K; Appaiah, Hitesh N; Siddaiah, Manjunath; Kumarswamy, Sunil B; Sunila, Sunila; KN, Thimmaiah; Mortha, KarunaKumar; Toole, Bryan; Banerjee, Shib d

    2006-01-01

    Background The development and progression of human tumors is accompanied by various cellular, biochemical and genetic alterations. These events include tumor cells interaction with extracellular matrix molecules including hyaluronan (HA). Hyaluronan is a large polysaccharide associated with pericellular matrix of proliferating, migrating cells. Its implication in malignant transformation, tumor progression and with the degree of differentiation in various invasive tumors has well accepted. It has been well known the role HA receptors in tumor growth and metastasis in various cancer tissues. Previously we have observed the unified over expression of Hyaluronic Acid Binding Protein (HABP), H11B2C2 antigen by the tumor cells in various types progressing tumor tissues with different grades. However, the poor understanding of relation between HA and HA-binding protein expression on tumor cells during tumor progression as well as the asymmetric observations of the role of HA expression in tumor progression prompted us to examine the degree of HA expression on tumor cells vs. stroma in various types of human tumors with different grades. Methods In the present study clinically diagnosed tumor tissue samples of different grades were used to screen the histopathological expression of hyaluronan by using b-PG (biotinylated proteoglycan) as a probe and we compared the relative HA expression on tumor cells vs. stroma in well differentiated and poorly differentiated tumors. Specificity of the reaction was confirmed either by pre-digesting the tissue sections with hyaluronidase enzyme or by staining the sections with pre-absorbed complex of the probe and HA-oligomers. Results We show here the down regulation of HA expression in tumor cells is associated with progression of tumor from well differentiated through poorly differentiated stage, despite the constant HA expression in the tumor associated stroma. Conclusion The present finding enlighten the relative roles of HA expression on tumor vs. stroma during the progression of tumors. PMID:16401353

  15. Randomized, evaluator-blind, split-face comparison study of single cross-linked versus double cross-linked hyaluronic acid in the treatment of glabellar lines.

    PubMed

    Kono, Taro; Kinney, Brian M; Groff, William Frederick; Chan, Henry H; Ercocen, Ali Riza; Nozaki, Motohiro

    2008-06-01

    BACKGROUND At present, various hyaluronic acids are being used to rejuvenate facial skin. There is no comparative study of single cross-linked hyaluronic acid (SCHA) versus double cross-linked hyaluronic acid (DCHA). The objective of our study is to compare the effectiveness and complications of SCHA versus DCHA in the treatment of glabellar lines. METHODS Ten female patients were enrolled in this randomized, evaluator-blind study. One side (left vs. right) of each patient's glabellar lines was treated with SCHA and the other side was treated with DCHA. Two independent blinded observers reviewed the clinical photographs at 3, 6, 9, and 12 months after the treatment and assessed for degree of improvement as well as complications. RESULTS The two products were equally effective in producing an optimal cosmetic result, although at 6, 9, and 12 months posttreatment, a higher proportion of patients showed over 50% improvement with DCHA than with SCHA. At 12 months posttreatment, DCHA was considered superior in 70% of patients, whereas SCHA was superior in 10% of patients. CONCLUSIONS Both SCHA and DCHA are equally effective in producing an optimal cosmetic result. DCHA provides a more durable esthetic improvement when compared to SCHA in the treatment of glabellar lines. PMID:18547178

  16. Cancer Microenvironment and Inflammation: Role of Hyaluronan

    PubMed Central

    Nikitovic, Dragana; Tzardi, Maria; Berdiaki, Aikaterini; Tsatsakis, Aristidis; Tzanakakis, George N.

    2015-01-01

    The role of inflammation in the development of cancer was described as early as the nineteenth century. Abundant evidence supports the preposition that various cancers are triggered by infection and chronic inflammatory disease whereas, evading immune destruction has been proposed as one of the new “hallmarks of cancer.” Changes of the tumor microenvironment have been closely correlated to cancer-mediated inflammation. Hyaluronan (HA), an important extracellular matrices component, has become recognized as an active participant in inflammatory, angiogenic, fibrotic, and cancer promoting processes. This review discusses how HA and specific HA-binding proteins participate in and regulate cancer-related inflammatory processes. PMID:25926834

  17. Single-molecule imaging of hyaluronan in human synovial fluid

    NASA Astrophysics Data System (ADS)

    Kappler, Joachim; Kaminski, Tim P.; Gieselmann, Volkmar; Kubitscheck, Ulrich; Jerosch, Jrg

    2010-11-01

    Human synovial fluid contains a high concentration of hyaluronan, a high molecular weight glycosaminoglycan that provides viscoelasticity and contributes to joint lubrication. In osteoarthritis synovial fluid, the concentration and molecular weight of hyaluronan decrease, thus impairing shock absorption and lubrication. Consistently, substitution of hyaluronan (viscosupplementation) is a widely used treatment for osteoarthritis. So far, the organization and dynamics of hyaluronan in native human synovial fluid and its action mechanism in viscosupplementation are poorly characterized at the molecular level. Here, we introduce highly sensitive single molecule microscopy to analyze the conformation and interactions of fluorescently labeled hyaluronan molecules in native human synovial fluid. Our findings are consistent with a random coil conformation of hyaluronan in human synovial fluid, and point to specific interactions of hyaluronan molecules with the synovial fluid matrix. Furthermore, single molecule microscopy is capable of detecting the breakdown of the synovial fluid matrix in osteoarthritis. Thus, single molecule microscopy is a useful new method to probe the structure of human synovial fluid and its changes in disease states like osteoarthritis.

  18. Hyaluronan, CD44 and Emmprin: Partners in cancer cell chemoresistance

    PubMed Central

    Toole, Bryan P.; Slomiany, Mark G.

    2008-01-01

    Hyaluronan is not only an important structural component of extracellular matrices but also interacts with cells during dynamic cell processes such as occur in cancer. Consequently, interactions of hyaluronan with tumor cells play important cooperative roles in various aspects of malignancy. Hyaluronan binds to several cell surface receptors, including CD44, thus leading to co-regulation of signaling pathways that are important in regulation of multidrug resistance to anticancer drugs, in particular anti-apoptotic pathways induced by activation of receptor tyrosine kinases. Emmprin, a cell surface glycoprotein of the Ig superfamily, stimulates hyaluronan production and downstream signaling consequences. Emmprin and CD44 also interact with various multidrug transporters of the ABC family and monocarboxylate transporters associated with resistance to cancer therapies. Moreover, hyaluronan-CD44 interactions are critical to these properties in the highly malignant, chemotherapy-resistant cancer stem-like cells. Perturbations of the hyaluronan-CD44 interaction at the plasma membrane by various antagonists result in attenuation of receptor tyrosine kinase and transporter activities and inhibition of tumor progression in vivo. These antagonists, especially small hyaluronan oligomers, may be useful in therapeutic strategies aimed at preventing tumor refractoriness or recurrence due to drug-resistant sub-populations within malignant cancers. PMID:18490190

  19. A silver complex of hyaluronan-lipoate (SHLS12): Synthesis, characterization and biological properties.

    PubMed

    Sacco, Pasquale; Sechi, Alessandra; Trevisan, Antonia; Picotti, Fabrizio; Gianni, Rita; Stucchi, Luca; Fabbian, Matteo; Bosco, Marco; Paoletti, Sergio; Marsich, Eleonora

    2016-01-20

    In this study we present a novel silver complex of hyaluronan-lipoate (SHLS12) in a gel-state form. NMR analysis, conductometry and elemental analysis demonstrated stable non-covalent interactions between silver ions and the polysaccharide-lipoate backbone, whereas rheological investigations confirmed its gel-like physical-chemical behavior. Biological studies showed the ability of SHLS12 to exert a straightforward activity against different bacterial strains grown in sessile/planktonic state. The biocompatibility was also proved toward two eukaryotic cell lines. By considering both its ability to preserve antibacterial properties when exposed to the serum protein BSA and its low susceptibility to be degraded by hyaluronidase enzyme, this novel complex may be considered as a promising biomaterial for future in vivo applications. PMID:26572372

  20. Level and position of substituents in cross-linked and hydroxypropylated sweet potato starches using nuclear magnetic resonance spectroscopy.

    PubMed

    Zhao, Jianwei; Chen, Zhenghong; Jin, Zhengyu; de Waard, Pieter; Buwalda, Piet; Gruppen, Harry; Schols, Henk A

    2015-10-20

    Sweet potato starch was cross-linked using sodium trimetaphosphate and hydroxypropylated using propylene oxide. The level and position of phosphorus and hydroxypropyl groups within cross-linked and hydroxypropylated sweet potato starch was investigated by phosphorus and proton nuclear magnetic resonance spectroscopy ((31)P, (1)H NMR). The cross-linking reaction produced monostarch monophosphate and distarch monophosphate in a molar ratio of 1:1.03, indicating that only half of the introduced phosphorus resulted in a possible cross-link. One cross-link per approximately 2900 glucose residues was found. Phosphorylation leading to monostarch monophosphate mainly occurred at O-3 and O-6 (ratio 1:1). It was inferred that the majority of the cross-links formed in distarch monophosphate were between two glucose residues positioned in different starch chains, while a minor part of the cross-links may be formed between two glucose residues within the same starch chain. Hydroxypropylation under alkaline conditions resulted in the formation of intra-molecular phosphorus cross-links, subsequent hydroxypropylation following cross-linking lowered both the level of intra- and inter-molecular cross-linking. Using (1)H NMR the molar substitution of hydroxypropylation was determined to be 0.155-0.165. The hydroxypropylation predominantly occurred at O-2 (61%), and the level of substitution at O-6 (21%) was slightly higher than that at O-3 (17%). In dual modified starch, the preceding cross-linking procedure resulted in a slightly lower level of hydroxypropylation, where the substitution at O-6 decreased more compared to the substitution at O-2 and O-3. PMID:26256203

  1. Hyaluronan and synovial joint: function, distribution and healing

    PubMed Central

    2013-01-01

    Synovial fluid is a viscous solution found in the cavities of synovial joints. The principal role of synovial fluid is to reduce friction between the articular cartilages of synovial joints during movement. The presence of high molar mass hyaluronan (HA) in this fluid gives it the required viscosity for its function as lubricant solution. Inflammation oxidation stress enhances normal degradation of hyaluronan causing several diseases related to joints. This review describes hyaluronan properties and distribution, applications and its function in synovial joints, with short review for using thiol compounds as antioxidants preventing HA degradations under inflammation conditions. PMID:24678248

  2. Hyaluronan and synovial joint: function, distribution and healing.

    PubMed

    Tamer, Tamer Mahmoud

    2013-09-01

    Synovial fluid is a viscous solution found in the cavities of synovial joints. The principal role of synovial fluid is to reduce friction between the articular cartilages of synovial joints during movement. The presence of high molar mass hyaluronan (HA) in this fluid gives it the required viscosity for its function as lubricant solution. Inflammation oxidation stress enhances normal degradation of hyaluronan causing several diseases related to joints. This review describes hyaluronan properties and distribution, applications and its function in synovial joints, with short review for using thiol compounds as antioxidants preventing HA degradations under inflammation conditions. PMID:24678248

  3. Chemical Cross-Linking Stabilizes Native-Like HIV-1 Envelope Glycoprotein Trimer Antigens

    PubMed Central

    Schiffner, Torben; de Val, Natalia; Russell, Rebecca A.; de Taeye, Steven W.; de la Peña, Alba Torrents; Ozorowski, Gabriel; Kim, Helen J.; Nieusma, Travis; Brod, Florian; Cupo, Albert; Sanders, Rogier W.; Moore, John P.; Ward, Andrew B.

    2015-01-01

    ABSTRACT Major neutralizing antibody immune evasion strategies of the HIV-1 envelope glycoprotein (Env) trimer include conformational and structural instability. Stabilized soluble trimers such as BG505 SOSIP.664 mimic the structure of virion-associated Env but nevertheless sample different conformational states. Here we demonstrate that treating BG505 SOSIP.664 trimers with glutaraldehyde or a heterobifunctional cross-linker introduces additional stability with relatively modest effects on antigenicity. Thus, most broadly neutralizing antibody (bNAb) epitopes were preserved after cross-linking, whereas the binding of most weakly or nonneutralizing antibodies (non-NAb) was reduced. Cross-linking stabilized all Env conformers present within a mixed population, and individual conformers could be isolated by bNAb affinity chromatography. Both positive selection of cross-linked conformers using the quaternary epitope-specific bNAbs PGT145, PGT151, and 3BC315 and negative selection with non-NAbs against the V3 region enriched for trimer populations with improved antigenicity for bNAbs. Similar results were obtained using the clade B B41 SOSIP.664 trimer. The cross-linking method may, therefore, be useful for countering the natural conformational heterogeneity of some HIV-1 Env proteins and, by extrapolation, also vaccine immunogens from other pathogens. IMPORTANCE The development of a vaccine to induce protective antibodies against HIV-1 is of primary public health importance. Recent advances in immunogen design have provided soluble recombinant envelope glycoprotein trimers with near-native morphology and antigenicity. However, these trimers are conformationally flexible, potentially reducing B-cell recognition of neutralizing antibody epitopes. Here we show that chemical cross-linking increases trimer stability, reducing binding of nonneutralizing antibodies while largely maintaining neutralizing antibody binding. Cross-linking followed by positive or negative antibody affinity selection of individual stable conformational variants further improved the antigenic and morphological characteristics of the trimers. This approach may be generally applicable to HIV-1 Env and also to other conformationally flexible pathogen antigens. PMID:26512083

  4. Properties of Poly(ethylene glycol) Hydrogels Cross-Linked via Strain-Promoted Alkyne-Azide Cycloaddition (SPAAC).

    PubMed

    Hodgson, Sabrina M; Bakaic, Emilia; Stewart, S Alison; Hoare, Todd; Adronov, Alex

    2016-03-14

    A series of poly(ethylene glycol) (PEG) hydrogels was synthesized using strain-promoted alkyne-azide cycloaddition (SPAAC) between PEG chains terminated with either aza-dibenzocyclooctynes or azide functionalities. The gelation process was found to occur rapidly upon mixing the two components in aqueous solution without the need for external stimuli or catalysts, making the system a candidate for use as an injectable hydrogel. The mechanical and rheological properties of these hydrogels were found to be tunable by varying the polymer molecular weight and the number of cross-linking groups per chain. The gelation times of these hydrogels ranged from 10 to 60 s at room temperature. The mass-based swelling ratios varied from 45 to 76 at maximum swelling (relative to the dry state), while the weight percent of polymer in these hydrogels ranged from 1.31 to 2.05%, demonstrating the variations in amount of polymer required to maintain the structural integrity of the gel. Each hydrogel degraded at a different rate in PBS at pH = 7.4, with degradation times ranging from 1 to 35 days. By changing the composition of the two starting components, it was found that the Young's modulus of each hydrogel could be varied from 1 to 18 kPa. Hydrogel incubation with bovine serum albumin showed minimal protein adsorption. Finally, a cell cytotoxicity study of the precursor polymers with 3T3 fibroblasts demonstrated that the azide- and strained alkyne-functionalized PEGs are noncytotoxic. PMID:26842783

  5. Immobilization of Quantum Dots in the Photo-Cross-Linked Poly(ethylene glycol)-Based Hydrogel

    SciTech Connect

    Gattas-Asfura, Kerim M.; Zheng, Yujun; Micic, Miodrag ); Snedaker, Michael J.; Ji, Xiaojun; Sui, Guodong; Orbulescu, Jhony; Andreopoulos, Fotios M.; Pham, Si M.; Wang, Chong M.)

    2003-09-25

    An inorganic/organic composite hybrid nano-system has been successfully synthesized in which nanocrystalline quantum dots (QDs) were effectively immobilized within a photo-cross-linked poly(ethylene glycol) hydrogel. Organometallic synthesis of CdTe and CdSe QDs was accomplished with a trioctylphosphine oxide (TOPO) cap. Replacing the TOPO cap with mercaptoacetic acid groups further yielded modified water-soluble nanocrystals. The immobilization of these functionalized CdTe and CdSe QDs within PEG hydrogel network has been shown to be effective through utilization of physical trapping. The CdTe and CdSe QDs had a particle diameter of 4.5 and 2.5-6.0 nm, respectively. The most efficiently trapped QDs had a size of 4.5 nm or larger. Particle size determination was derived from spectroscopic (absorption and photoluminescence) and high-resolution transmission electron microscopic techniques. These QD-immobilized gel systems demonstrated photoluminescence characteristics unique to semiconductor QD nanocrystals. The authors have envisioned the utilization of the unique photophysical properties of this material as a convenient signal transducer for in vivo biosensing. The most promising application of the described QD/PEG-NC hybrid system is in the fields of in vivo fluorescence microscopy and as a monitoring system for drug delivery and wound healing.

  6. DHPLC and MS studies of a photoinduced intrastrand cross-link in DNA labeled with 5-bromo-2'-deoxyuridine.

    PubMed

    Wiczk, Justyna; Miloch, Justyna; Rak, Janusz

    2014-01-01

    It is well known that the replacement of thymidine with 5-bromo-2'-deoxyuridine (BrdU) in DNA sensitizes it to UVB light. Irradiation of a biopolymer substituted in such a way leads to manifold kinds of DNA damage, such as intrastrand cross-links, single- and double-strand breaks or alkali-labile sites that were studied in the past with a broad spectrum of analytical methods. Here, we demonstrate that completely denaturing high-performance liquid chromatography (DHPLC), underestimated so far in DNA damage studies, could act as an inexpensive, and high-resolution substitute for the commonly employed gel electrophoresis. We report on the DHPLC/mass spectrometry (MS) analyses of photolytes obtained with the UV irradiation of aqueous solutions containing 40 base pairs of a long, double-stranded oligonucleotide labeled with BrdU in one of its strands. The UV-product was detected by HPLC at a temperature of 70C. Subsequent MS analysis with electrospray ionization (ESI-MS) of the photolyte, enzymatic digestion of the irradiated material and HPLC and MS analysis (LC-MS) of the digest demonstrated unequivocally that an intrastrand covalent dimer, involving adenine and uracil, is formed in the irradiated system. PMID:24300995

  7. Hyaluronan stimulates pancreatic cancer cell motility.

    PubMed

    Cheng, Xiao-Bo; Kohi, Shiro; Koga, Atsuhiro; Hirata, Keiji; Sato, Norihiro

    2016-01-26

    Hyaluronan (HA) accumulates in pancreatic ductal adenocarcinoma (PDAC), but functional significance of HA in the aggressive phenotype remains unknown. We used different models to investigate the effect of HA on PDAC cell motility by wound healing and transwell migration assay. Changes in cell motility were examined in 8 PDAC cell lines in response to inhibition of HA production by treatment with 4-methylumbelliferone (4-MU) and to promotion by treatment with 12-O-tetradecanoyl-phorbol-13-acetate (TPA) or by co-culture with tumor-derived stromal fibroblasts. We also investigated changes in cell motility by adding exogenous HA. Additionally, mRNA expressions of hyaluronan synthases and hyaluronidases were examined using real time RT-PCR. Inhibition of HA by 4-MU significantly decreased the migration, whereas promotion of HA by TPA or co-culture with tumor-derived fibroblasts significantly increased the migration of PDAC cells. The changes in HA production by these treatments tended to be associated with changes in HAS3 mRNA expression. Furthermore, addition of exogenous HA, especially low-molecular-weight HA, significantly increased the migration of PDAC cells. These findings suggest that HA stimulates PDAC cell migration and thus represents an ideal therapeutic target to prevent invasion and metastasis. PMID:26684359

  8. 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 ?-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

  9. Kinetics of swelling of polyether-modified poly(acrylic acid) microgels with permanent and degradable cross-links.

    PubMed

    Bromberg, Lev; Temchenko, Marina; Alakhov, Valery; Hatton, T Alan

    2005-02-15

    Spherical particles of 50-100 mum size composed of poly(acrylic acid) networks covalently bonded to Pluronic polyether copolymers were tested for swelling in aqueous media. The microgels were cross-linked either by permanent ethylene glycol dimethacrylate (EGDMA) cross-links alone or by EDGMA together with reversible disulfide or biodegradable azoaromatic cross-links. Optimum conditions for a rapid, diffusion-limited swelling of the pH- and temperature-sensitive microgels with nondegradable cross-links were found. The microgels cross-linked by disulfide groups and equilibrium-swollen in the buffer solution exhibited degradation-limited kinetics of swelling under physiological conditions, with a first-order reaction constant, k(1), linearly proportional to the concentration of reducing agents such as dithiotreitol and tris(2-carboxyethyl)phosphine (TCEP). A severalfold faster swelling in the presence of more powerful reducing agent, TCEP, was observed, indicating the chemical specificity of the microgel swelling. The reoxidation of the thiol groups into disulfide cross-links by sodium hypochlorite led to the restoration of the microgels' diameter measured prior to the reduction-reoxidation cycle, which confirms the shape memory of the microgels. Enzymatically degradable azoaromatic cross-links enabled slow microgel swelling due to degradation of the cross-links by azoreductases from the rat intestinal cecum. The low rate of swelling of the Pluronic-containing microgels can enable sustained drug release in colon-specific drug delivery. PMID:15697312

  10. Expanding the Chemical Cross-Linking Toolbox by the Use of Multiple Proteases and Enrichment by Size Exclusion Chromatography*

    PubMed Central

    Leitner, Alexander; Reischl, Roland; Walzthoeni, Thomas; Herzog, Franz; Bohn, Stefan; Frster, Friedrich; Aebersold, Ruedi

    2012-01-01

    Chemical cross-linking in combination with mass spectrometric analysis offers the potential to obtain low-resolution structural information from proteins and protein complexes. Identification of peptides connected by a cross-link provides direct evidence for the physical interaction of amino acid side chains, information that can be used for computational modeling purposes. Despite impressive advances that were made in recent years, the number of experimentally observed cross-links still falls below the number of possible contacts of cross-linkable side chains within the span of the cross-linker. Here, we propose two complementary experimental strategies to expand cross-linking data sets. First, enrichment of cross-linked peptides by size exclusion chromatography selects cross-linked peptides based on their higher molecular mass, thereby depleting the majority of unmodified peptides present in proteolytic digests of cross-linked samples. Second, we demonstrate that the use of proteases in addition to trypsin, such as Asp-N, can additionally boost the number of observable cross-linking sites. The benefits of both SEC enrichment and multiprotease digests are demonstrated on a set of model proteins and the improved workflow is applied to the characterization of the 20S proteasome from rabbit and Schizosaccharomyces pombe. PMID:22286754

  11. Photouncaged Sequence-specific Interstrand DNA Cross-Linking with Photolabile 4-oxo-enal-modified Oligonucleotides

    NASA Astrophysics Data System (ADS)

    Sun, Jingjing; Tang, Xinjing

    2015-05-01

    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 ?-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. 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. PMID:26428097

  13. SJG-136 (NSC 694501), a novel rationally designed DNA minor groove interstrand cross-linking agent with potent and broad spectrum antitumor activity: part 1: cellular pharmacology, in vitro and initial in vivo antitumor activity.

    PubMed

    Hartley, John A; Spanswick, Victoria J; Brooks, Natalie; Clingen, Peter H; McHugh, Peter J; Hochhauser, Daniel; Pedley, R Barbara; Kelland, Lloyd R; Alley, Michael C; Schultz, Robert; Hollingshead, Melinda G; Schweikart, Karen M; Tomaszewski, Joseph E; Sausville, Edward A; Gregson, Stephen J; Howard, Philip W; Thurston, David E

    2004-09-15

    SJG-136 (NSC 694501) is a rationally designed pyrrolobenzodiazepine dimer that binds in the minor groove of DNA. It spans 6 bp with a preference for binding to purine-GATC-pyrimidine sequences. The agent has potent activity in the National Cancer Institute (NCI) anticancer drug screen with 50% net growth inhibition conferred by 0.14 to 320 nmol/L (7.4 nmol/L mean). Sensitive cell lines exhibit total growth inhibition and 50% lethality after treatment with as little as 0.83 and 7.1 nmol/L SJG-136, respectively. COMPARE and molecular target analysis of SJG-136 data versus that of >60,000 compounds tested in the NCI 60 cell line screen shows that, although the agent has similarity to other DNA binding agents, the pattern of activity for SJG-136 does not fit within the clusters of any known agents, suggesting that SJG-136 possesses a distinct mechanism of action. Testing in the NCI standard hollow fiber assay produced prominent growth inhibition in 20 of 24 i.p. and 7 of 24 s.c. test combinations with 5 of 12 cell lines exhibiting cell kill. In addition, SJG-136 produced antitumor activity in mice bearing CH1 and CH1cisR xenografts, a cisplatin-resistant human ovarian tumor model, and also in mice bearing LS174T xenografts, a human colon tumor model. SJG-136 produces DNA interstrand cross-links between two N-2 guanine positions on opposite strands and separated by 2 bp. In human tumor cell lines, the cross-links form rapidly and persist compared with those produced by conventional cross-linking agents such as nitrogen mustards. In mice bearing the LS174T human colon xenograft, DNA interstrand cross-links can be detected in tumor cells using a modification of the single cell gel electrophoresis (comet) assay after administration of a therapeutic dose. Cross-links in the tumor increase with dose and are clearly detectable at 1 hour after i.v. administration. The level of cross-linking persists over a 24-hour period in this tumor in contrast to cross-links produced by conventional cross-linking agents observed over the same time period. PMID:15374986

  14. Preparation of superhydrophobic cross-linked syndiotactic 1,2-polybutadiene membranes by electrospinning.

    PubMed

    Hao, Xiu-Feng; Du, Li-Ping; Cai, Hong-Guang; Zhang, Chun-Yu; Zhang, Xue-Quan; Zhang, He-Xin

    2012-10-01

    In this study, syndiotactic 1,2-polybutadiene (s-PB)/azobisisobutyronitrile (AIBN) membranes were prepared via electrospinning. The obtained membranes were intensively investigated by contact angle analyzer, differential scanning calorimetry (DSC) and environmental scanning electron microscope (ESEM). With increasing the concentration of spinning solution, the obtained membranes were changed from hydrophobic to surperhydrophobic and the contact angle (CA) as high as 157 degrees at higher s-PB concentration. In addition, the s-PB/AIBN membrane cross-linked completely through heat treatment and superhydrophobic property of s-PB/AIBN membrane has not changed. The diameter of cross-linked fiber-based membrane was much thinner than that of the uncross-linked fibers. PMID:23421181

  15. Elastin cross-linking in the skin from patients with amyotrophic lateral sclerosis

    NASA Technical Reports Server (NTRS)

    Ono, S.; Yamauchi, M.

    1994-01-01

    Two cross-links unique to elastin, desmosine and isodesmosine were measured and compared in skin tissue (left upper arm) from 10 patients with amyotrophic lateral sclerosis (ALS) and from seven age-matched controls. The contents of desmosine and isodesmosine were significantly decreased (p < 0.01 and p < 0.01, respectively) in patients with ALS compared with those of controls, and were negatively and significantly associated with duration of illness in ALS patients (r = -0.77, p < 0.01 and r = -0.65, p < 0.05, respectively). The decline in skin desmosine and isodesmosine is more rapid in ALS than in normal ageing. Thus cross-linking of skin elastin is affected in ALS.

  16. MeV ion beam interaction with polymer films containing cross-linking agents

    SciTech Connect

    Evelyn, A. L.

    1999-06-10

    Polymer films containing cross linking enhancers were irradiated with MeV alpha particles to determine the effects of MeV ion beam interaction on these materials. The contributed effects from the electronic and nuclear stopping powers were separated by irradiating stacked thin films of polyvinyl chloride (PVC), polystyrene (PS) and polyethersulfone (PES). This layered system allowed most of the effects of the electronic energy deposited to be experienced by the first layers and the last layers to receive most of the effects of the nuclear stopping power. RGA, Raman microprobe analysis, RBS and FTIR measured changes in the chemical structures of the irradiated films. The characterization resolved the effects of the stopping powers on the PVC, PS and PES and the results were compared with those from previously studied polymers that did not contain any cross linking agents.

  17. Novel terephthaloyl thiourea cross-linked chitosan hydrogels as antibacterial and antifungal agents.

    PubMed

    Mohamed, Nadia Ahmed; Al-mehbad, Noura Yahya

    2013-06-01

    Four novel terephthaloyl thiourea chitosan (TTU-chitosan) hydrogels were synthesized via a cross-linking reaction of chitosan with different concentrations of terephthaloyl diisothiocyanate. Their structures were investigated by elemental analyses, FTIR, SEM and X-ray diffraction. The antimicrobial activities of the hydrogels against three species of bacteria (Bacillis subtilis, Staphylococcus aureus and Escherichia coli) and three crop-threatening pathogenic fungi (Aspergillus fumigatus, Geotrichum candidum and Candida albicans) are much higher than that of the parent chitosan. The hydrogels were more potent in case of Gram-positive bacteria than Gram-negative bacteria. Increasing the degree of cross-linking in the hydrogels resulted in a stronger antimicrobial activity. PMID:23500435

  18. From epoxidized linseed oil to bioresin: an overall approach of epoxy/anhydride cross-linking.

    PubMed

    Pin, Jean-Mathieu; Sbirrazzuoli, Nicolas; Mija, Alice

    2015-04-13

    Biorenewable resources can be used as green monomers to design tailored structures for formulations that can play an important role as functional materials. The choice of optimal structures depends on the targeted properties and applications. This work focuses on the elaboration of biobased materials with toughened mechanical properties based on epoxidized linseed oil. This result was obtained by an overall approach of cross-linking process, that is, starting with the optimal choice of hardeners and finally favoring the side reactions of polymerization. Therefore, the anionic alternating copolymerization of epoxide with mono- and dianhydrides was investigated to tailor the parameters that led to maximal conversions and properties. The obtained highly cross-linked networks perform well, as demonstrated by good impact strengths, high glass transition temperatures, and excellent thermal stability, which opens up the possibility of using these emergent materials for industrial applications. PMID:25754910

  19. Stimuli-responsive cross-linked micelles for on-demand drug delivery against cancers.

    PubMed

    Li, Yuanpei; Xiao, Kai; Zhu, Wei; Deng, Wenbin; Lam, Kit S

    2014-02-01

    Stimuli-responsive cross-linked micelles (SCMs) represent an ideal nanocarrier system for drug delivery against cancers. SCMs exhibit superior structural stability compared to their non-cross-linked counterpart. Therefore, these nanocarriers are able to minimize the premature drug release during blood circulation. The introduction of environmentally sensitive cross-linkers or assembly units makes SCMs responsive to single or multiple stimuli present in tumor local microenvironment or exogenously applied stimuli. In these instances, the payload drug is released almost exclusively in cancerous tissue or cancer cells upon accumulation via enhanced permeability and retention effect or receptor mediated endocytosis. In this review, we highlight recent advances in the development of SCMs for cancer therapy. We also introduce the latest biophysical techniques, such as electron paramagnetic resonance (EPR) spectroscopy and fluorescence resonance energy transfer (FRET), for the characterization of the interactions between SCMs and blood proteins. PMID:24060922

  20. Biochemical properties of bioplastics made from wheat gliadins cross-linked with cinnamaldehyde.

    PubMed

    Balaguer, M Pau; Gómez-Estaca, Joaquín; Gavara, Rafael; Hernandez-Munoz, Pilar

    2011-12-28

    The aim of this work has been to study the modification of gliadin films with cinnamaldehyde as a potential cross-linker agent. The molecular weight profile and cross-linking density showed that cinnamaldehyde increased reticulation in the resulting films. The participation of free amino groups of the protein in the newly created entanglements could be a possible mechanism of connection between the polypeptidic chains. The combination of a Schiff base and a Michael addition is a feasible approach to understanding this mechanism. The protein solubility in different media pointed to lower participation by both noncovalent and disulfide bonds in stabilizing the structure of the cross-linked films. The new covalent bonds formed by the cinnamaldehyde treatment hampered water absorption and weight loss, leading to more water-resistant matrices which had not disintegrated after 5 months. The properties of this novel bioplastic could be modified to suit the intended application by using cinnamaldehyde, a naturally occurring compound. PMID:22047158

  1. 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. PMID:24127389

  2. Cross-linked sulfonated aromatic ionomers via SO2 bridges: Conductivity properties

    NASA Astrophysics Data System (ADS)

    Di Vona, M. L.; Pasquini, L.; Narducci, R.; Pelzer, K.; Donnadio, A.; Casciola, M.; Knauth, P.

    2013-12-01

    The proton conductivity of SPEEK membranes in situ cross-linked by thermal treatment at 180 C for various times was investigated by impedance spectroscopy. The conductivity measurements were made on fully humidified membranes between 25 and 65 C and on membranes exposed to different relative humidity between 80 and 140 C. The Ionic Exchange Capacity (IEC) was determined by acid-base titration and the water uptake by gravimetry. The proton conductivity was determined as function of temperature, IEC, degree of cross-linking and hydration number. A curve of proton conductivity vs. hydration number allows predicting that in order to reach a value of 0.1 S/cm at 100 C a hydration number above 20 is necessary. The measured conductivity at this temperature is 0.16 S/cm for a hydration number of 60.

  3. Multi-stimulus-responsive shape-memory polymer nanocomposite network cross-linked by cellulose nanocrystals.

    PubMed

    Liu, Ye; Li, Ying; Yang, Guang; Zheng, Xiaotong; Zhou, Shaobing

    2015-02-25

    In this study, we developed a thermoresponsive and water-responsive shape-memory polymer nanocomposite network by chemically cross-linking cellulose nanocrystals (CNCs) with polycaprolactone (PCL) and polyethylene glycol (PEG). The nanocomposite network was fully characterized, including the microstructure, cross-link density, water contact angle, water uptake, crystallinity, thermal properties, and static and dynamic mechanical properties. We found that the PEG[60]-PCL[40]-CNC[10] nanocomposite exhibited excellent thermo-induced and water-induced shape-memory effects in water at 37 C (close to body temperature), and the introduction of CNC clearly improved the mechanical properties of the mixture of both PEG and PCL polymers with low molecular weights. In addition, Alamar blue assays based on osteoblasts indicated that the nanocomposites possessed good cytocompatibility. Therefore, this thermoresponsive and water-responsive shape-memory nanocomposite could be potentially developed into a new smart biomaterial. PMID:25647407

  4. Cross-Linked Chitosan as an Efficient Binder for Si Anode of Li-ion Batteries.

    PubMed

    Chen, Chao; Lee, Sang Ha; Cho, Misuk; Kim, Jaehoon; Lee, Youngkwan

    2016-02-01

    We investigate the use of chitosan (CS) as a new cross-linkable and water-soluble binder for the Si anode of Li-ion batteries. In contrast to the traditional binder utilizing a hydrogen bond and/or van der Waals force-linked anode electrodes, CS can easily form a 3D network to limit the movement of Si particles through the cross-linking between the amino groups of CS and the dialdehyde of glutaraldehyde (GA). Chemical, mechanical, and morphological analyses are conducted by Fourier transform infrared spectroscopy, tensile testing, and scanning electron microscopy. The cross-linked Si/CS-GA anode exhibits an initial discharge capacity of 2782 mAh g(-1) with a high initial Coulombic efficiency of 89% and maintained a capacity of 1969 mAh g(-1) at the current density of 500 mA g(-1) over 100 cycles. PMID:26745390

  5. A supramolecular cross-linked conjugated polymer network for multiple fluorescent sensing.

    PubMed

    Ji, Xiaofan; Yao, Yong; Li, Jinying; Yan, Xuzhou; Huang, Feihe

    2013-01-01

    A supramolecular cross-linked network was fabricated and demonstrated to act as a multiple fluorescent sensor. It was constructed from a fluorescent conjugated polymer and a bisammonium salt cross-linker driven by dibenzo[24]crown-8/secondary ammonium salt host-guest interactions. Compared with the conjugated polymer, the network has weak fluorescence due to the aggregation of polymer chains. Thanks to the multiple stimuli-responsiveness of host-guest interactions, the fluorescence intensity of the system can be enhanced by four types of signals, including potassium cation, chloride anion, pH increase, and heating. Hence, the network can serve as a cation sensor, an anion sensor, a pH sensor, and a temperature sensor. It can be used in both solution and thin film. Interestingly, exposure of a film made from this supramolecular cross-linked network to ammonia leads to an increase of fluorescence, making it a good candidate for gas detection. PMID:23259828

  6. Dually cross-linked polymer electrolyte membranes for direct methanol fuel cells

    NASA Astrophysics Data System (ADS)

    Lee, Won Hyo; Lee, Kang Hyuck; Shin, Dong Won; Hwang, Doo Sung; Kang, Na Rae; Cho, Doo Hee; Kim, Ji Hoon; Lee, Young Moo

    2015-05-01

    End-group crosslinkable sulfonated poly(arylene ether sulfone) copolymer (ESPAES) and imidazolium poly(arylene ether sulfone) copolymer (IPAES) are synthesized as a proton exchange membrane and ionic crosslinker, respectively. A novel dually cross-linked membrane (DCM) based on ESPAES is similar to an inter-penetrating network and is prepared via blending IPAES and thermal treatment for direct methanol fuel cell (DMFC) applications. The synergistic effects of end-group crosslinking and ionic crosslinking improve chemical and thermal stability and mechanical properties. In addition, the DMFC performance of the DCM outperforms that of the end-group cross-linked SPAES and Nafion 212 due to its excellent fuel barrier property in spite of relatively low proton conductivity, which is derived from the content of the non-proton conducting IPAES copolymer. Consequently, the DCM has great potential as an electrolyte membrane for DMFC applications.

  7. Anionically cross linked homopolymer colloids applied in formation of platinum nanoparticles.

    PubMed

    Tangeysh, Behzad; Fryd, Michael; Hayik, Sara E N; Kamps, Amanda C; Wayland, Bradford B

    2012-03-01

    Diprotonic sulfuric and succinic acids react efficiently with the tertiary amine sites in polydimethylaminoethylmethacrylate (PDMAEMA) to produce polymer colloid nano-particles held together by dinegatively charged anions that cross link the partially protonated PDMAEMA homopolymer. This procedure is used to encorporate [PtCl(6)](2-) as a cross linker into the framework of well defined polymer network colloid particles that have dual roles as nanoreactors and a source of protective polymer coating. Reduction of the cross linking [PtCl(6)](2-) groups produces platinum metal nano-particles (1.12(.25)nm) that are relatively small and narrowly dispersed. Formation of colloid particles by reaction of diprotic acids with homopolymers that have proton accepting centers provides a convenient intentional route to incorporate a variety of homopolymers into self assembled polymer network materials for applications as nanoreactors and transport systems. PMID:22209582

  8. Metal-Free Preparation of Linear and Cross-Linked Polydicyclopentadiene.

    PubMed

    Goetz, Adam E; Boydston, Andrew J

    2015-06-24

    Metal-free ring-opening metathesis polymerization (ROMP) utilizes organic photoredox mediators as alternatives to traditional metal-based ROMP initiators to allow the preparation of polymers without residual metal contamination. Herein we report studies exploring the use of endo-dicyclopentadiene (DCPD), a common ROMP monomer, to form linear polyDCPD and copolymers with norbornene. Subsequent cross-linking of the linear polyDCPD using thiol-ene chemistry allows for a completely metal-free preparation of cross-linked polyDCPD. Furthermore, the examination of a number of structurally related monomers offers insights into mechanistic details of this polymerization and demonstrates new monomers that can be utilized for metal-free ROMP. PMID:26053254

  9. Cross-Linking Antisense Oligodeoxyribonucleotides with a Photoresponsive ?-Chloroaldehyde Moiety for RNA Point Mutations.

    PubMed

    Sugihara, Yuta; Nakata, Yuki; Yamayoshi, Asako; Murakami, Akira; Kobori, Akio

    2016-02-01

    Because point mutations in GTPase-coding genes have been reported to be responsible for the transformation of cells, anticancer reagents that react effectively and sequence selectively with target RNAs having a point mutation are highly desired. In this study, we developed novel photo-cross-linking oligodeoxyribonucleotides ((pro)PCA-ODNs) that had a caged ?-chloroaldehyde group conjugated to a 2-methylpropanediyl backbone ((pro)PCA) in the middle of the strand. A kinetic study of the deprotection reaction of (pro)PCA-ODN revealed that the bis(2-nitrobenzyl)acetal group was completely deprotected within 1 min. Photo-cross-linking studies of (pro)PCA-ODNs with complementary oligoribonucleotides (ORNs) revealed that (pro)PCA-ODNs reacts efficiently and selectively with the target ORNs that have an adenosine or cytidine residue at a frontal position of the (pro)PCA residue without adverse effects of bases adjacent to the mutation site. PMID:26788869

  10. Thermally Cross-Linked Anion Exchange Membranes from Solvent Processable Isoprene Containing Ionomers

    SciTech Connect

    Tsai, Tsung-Han; Ertem, S. Piril; Maes, Ashley M.; Seifert, Soenke; Herring, Andrew M; Coughlin, E. Bryan

    2015-01-28

    Random copolymers of isoprene and 4-vinylbenzyl chloride (VBCl) with varying compositions were synthesized via nitroxide-mediated polymerization. Subsequent quaternization afforded solvent processable and cross-linkable ionomers with a wide range of ion exchange capacities (IECs). Solution cast membranes were thermally cross-linked to form anion exchange membranes. Cross-linking was achieved by taking advantage of the unsaturations on the polyisoprene backbone, without added cross-linkers. A strong correlation was found between water uptake and ion conductivity of the membranes: conductivities of the membranes with IECs beyond a critical value were found to be constant related to their high water absorption. Environmentally controlled small-angle X-ray scattering experiments revealed a correlation between the average distance between ionic clusters and the ion conductivity, indicating that a well-connected network of ion clusters is necessary for efficient ion conduction and high ion conductivity.

  11. Chemistry for the analysis of proteinprotein interactions: Rapid and efficient cross-linking triggered by long wavelength light

    PubMed Central

    Fancy, David A.; Kodadek, Thomas

    1999-01-01

    Chemical cross-linking is a potentially useful technique for probing the architecture of multiprotein complexes. However, analyses using typical bifunctional cross-linkers often suffer from poor yields, and large-scale modification of nucleophilic side chains can result in artifactual results attributable to structural destabilization. We report here the de novo design and development of a type of protein cross-linking reaction that uses a photogenerated oxidant to mediate rapid and efficient cross-linking of associated proteins. The process involves brief photolysis of tris-bipyridylruthenium(II) dication with visible light in the presence of the electron acceptor ammonium persulfate and the proteins of interest. Very high yields of cross-linked products can be obtained with irradiation times of <1 second. This chemistry obviates many of the problems associated with standard cross-linking reagents. PMID:10339534

  12. The process of EDC-NHS cross-linking of reconstituted collagen fibres increases collagen fibrillar order and alignment

    SciTech Connect

    Shepherd, D. V. Shepherd, J. H.; Cameron, R. E.; Best, S. M.; Ghose, S.; Kew, S. J.

    2015-01-01

    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.

  13. The process of EDC-NHS Cross-linking of reconstituted collagen fibres increases collagen fibrillar order and alignment

    PubMed Central

    Shepherd, D.V.; Shepherd, J.H.; Ghose, S.; Kew, S.J.; Cameron, R.E.; Best, S.M.

    2014-01-01

    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 (AFM) 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. PMID:25506518

  14. Informatics Strategies for Large-Scale Novel Cross-Linking Analysis

    PubMed Central

    Anderson, Gordon A.; Tolic, Nikola; Tang, Xiaoting; Zheng, Chunxiang; Bruce, James E.

    2008-01-01

    The detection of protein interactions in biological systems represents a significant challenge for today’s technology. Chemical cross-linking provides the potential to impart new chemical bonds in a complex system that result in mass changes in a set of tryptic peptides detected by mass spectrometry. However, system complexity and cross-linking product heterogeneity have precluded widespread chemical cross-linking use for large-scale identification of protein–protein interactions. The development of mass spectrometry identifiable cross-linkers called protein interaction reporters (PIRs) has enabled on-cell chemical cross-linking experiments with product type differentiation. However, the complex datasets resultant from PIR experiments demand new informatics capabilities to allow interpretation. This manuscript details our efforts to develop such capabilities and describes the program X-links, which allows PIR product type differentiation. Furthermore, we also present the results from Monte Carlo simulation of PIR-type experiments to provide false discovery rate estimates for the PIR product type identification through observed precursor and released peptide masses. Our simulations also provide peptide identification calculations based on accurate masses and database complexity that can provide an estimation of false discovery rates for peptide identification. Overall, the calculations show a low rate of false discovery of PIR product types due to random mass matching of approximately 12% with 10 ppm mass measurement accuracy and spectral complexity resulting from 100 peptides. In addition, consideration of a reduced database resulting from stage 1 analysis of Shewanella oneidensis MR-1 containing 367 proteins resulted in a significant reduction of expected identification false discovery rate estimation compared to that from the entire Shewanella oneidensis MR-1 proteome. PMID:17676784

  15. Thin-Walled Cross-Linked Acetabular Liners Need Not Exhibit Reduced Locking Strength.

    PubMed

    Murtha, Andrew S; Roy, Marcel E; Whiteside, Leo A; Tilden, David S; Schmitt, Krystal L

    2015-08-01

    Use of larger diameter femoral heads has emerged as a promising strategy to reduce the risk of dislocation after total hip arthroplasty, but thinning the walls of cross-linked ultra-high-molecular-weight polyethylene (UHMWPE) acetabular liners to accommodate these larger heads may compromise the locking mechanism of the liner. The purpose of this study was to test the mechanical integrity of the locking mechanism in cross-linked and re-melted UHMWPE acetabular components with reduced wall thickness. The locking mechanism of cross-linked (100 kGy/re-melted) acetabular liners in sizes 50/28, 50/36, and 52/36 mm of 1 design was evaluated by lever-out tests and torsion tests. Torsion tests were performed at 2 angles to isolate the liner's locking tabs independent of the contribution of its central post. Lever-out testing demonstrated nominally reduced failure strength in 50/36-mm liners (13.3 N m) compared with 50/28-mm liners (12.3 N m; P=.0502), whereas the lever-out strength of 52/36-mm liners was 12.20.94 N m. Failure torques were similar between 50/28- and 50/36-mm liners at 45 and 90, but the failure torque of size 52/36-mm liners was significantly higher at each angle. The use of larger diameter femoral heads does not compromise the locking mechanism of thinned MicroSeal (Signal Medical Corp, Marysville, Michigan) acetabular liners. Use of a cross-linked UHMWPE acetabular liner, with a locking mechanism that is not compromised when the liner is thinned to a thickness of at least 2.86 mm, appears to be a biomechanically sound construct when articulated with large diameter femoral heads. PMID:26270761

  16. Generation of Guanine Amino Acid Cross-Links by a Free Radical Combination Mechanism

    PubMed Central

    Uvaydov, Yuriy; Geacintov, Nicholas E.

    2014-01-01

    A direct method has been developed for the in vitro synthesis of stable DNA protein cross-links (DPCs) between guanine and amino acids (lysine, arginine). This method employs the combination of guanine neutral radicals, G(-H), and side-chain C-centered amino acid radicals. The latter were generated indirectly after first causing the selective photoionization of 2-aminopurine (2AP) embedded in the oligonucleotide, 5-d(CC[2AP]TCGCTACC) by intense nanosecond 308 nm excimer laser pulses. The 2AP radical cation deprotonates rapidly to form the 2AP(-H) neutral radical which, in turn, oxidizes the nearby guanine to form the neutral guanine G(-H) radical, as described previously (Shafirovich et al. J. Phys. Chem. B, 2001, 105, 8431). In parallel, the hydrated electrons generated by the photoionization of 2AP, are scavenged by nitrous oxide to generate hydroxyl radicals. In the presence of a large excess of the amino acids, the hydroxyl radicals oxidize the latter to produce C-centered amino acid radicals that combine with the G(-H) radicals to form the guanine-amino acid cross-linked oligonucleotide product. Analogous products were generated by photoionizing the free nucleoside, 2,3,5-tri-O-acetylguanoise, (tri-O-Ac-Guo) using intense nanosecond 266 nm Nd: Yag laser pulse irradiation. The guanine amino acid cross-links thus produced either site-specifically positioned in oligonucleotides, or in the free nucleoside tri-O-Ac-Guo were isolated by HPLC methods and identified by high resolution LC-TOF/MS and LC-MS/MS methods. The possibility is discussed that analogous guanine-amino acid cross-linked products could be formed in vivo under single hit radical generation mechanisms during oxidative stress. PMID:24810398

  17. Volume phase transitions in surface-tethered, photo-cross-linked poly(N-isopropylacrylamide) networks

    NASA Astrophysics Data System (ADS)

    Vidyasagar, Ajay Kumar

    The overall thrust of this dissertation is to gain a comprehensive understanding over the factors that govern the performance and behavior of ultra-thin, cross-linked polymer films. Poly(NIPAAm) was used as a model polymer to study volume phase transition in surface tethered networks. Poly(NIPAAm) undergoes a reversible phase transition at approximately 32C between a swollen hydrophilic random coil to a collapsed hydrophobic globule state, thought to be caused by increased hydrophobic attractions between the isopropyl groups at elevated temperatures. We present a simple photochemical technique for fabricating structured polymer networks, enabling the construction of responsive surfaces with unique properties. The approach is based on the photo-cross-linking of copolymers synthesized from N-isopropylacrylamide and methacroyloxybenzophenone (MaBP). In order correlate layer swelling to the MaBP content, we have studied the swelling behavior of such layers in contact with aqueous solutions with neutron reflection. The cross-linked networks provide a three-dimensional scaffold to host a variety of functionalities. These networks serve as a platform which can be used to amplify small local perturbations induced by various stimuli like temperature, pH, solvent, ionic strength and peptide modified hydrogels to bring about a macroscopic change. Neutron reflection experiments have shown that the volume-phase transition of a surface-tethered, cross-linked poly(NIPAAm) network coincided with the two-phase region of uncross-linked poly(NIPAAm) in solution. Parallel measurements with ATR-FTIR investigating the effect of temperature, pH and salts suggest that the discontinuous transition is the result of cooperative dehydration of the isopropyl groups, with water remaining confined between amide groups in the collapsed state as weakly hydrogen bonded bridges. Hybrid polymers with specific peptide sequences have shown specific response to external cues such as pH and metal ions exhibiting unique phase behavior.

  18. Fabrication and properties of irradiation-cross-linked poly(vinyl alcohol)/clay aerogel composites.

    PubMed

    Chen, Hong-Bing; Liu, Bo; Huang, Wei; Wang, Jun-Sheng; Zeng, Guang; Wu, Wen-Hao; Schiraldi, David A

    2014-09-24

    Poly(vinyl alcohol) (PVOH)/clay aerogel composites were fabricated by an environmentally friendly freeze-drying of the aqueous precursor suspensions, followed by cross-linking induced by gamma irradiation without chemical additives. The influences of cross-linking conditions, i.e., absorbed dose and polymer loading as well as density on the aerogel structure and properties, were investigated. The absorbed dose of 30 kGy was found to be the optimum dose for fabricating strong PVOH composites; the compressive modulus of an aerogel prepared from an aqueous suspension containing 2 wt % PVOH/8 wt % clay increased 10-fold, and that containing 1 wt % PVOH/9 wt % clay increased 12 times upon cross-linking with a dose of 30 kGy. Increasing the solids concentration led to an increase in the mechanical strength, in accordance with the changes in microstructure from layered structure to network structure. The increase of absorbed dose also led to decreased porous size of the network structure. Cross-linking and the increase of the PVOH lead to decreased thermal stability. The strengthened PVOH/clay aerogels possess very low flammability, as measured by cone calorimetry, with heat, smoke, and volatile products release value decreasing as increasing clay content. The mechanism of flame retardation in these materials was investigated with weight loss, FTIR, WAXD, and SEM of the burned residues. The proposed mechanism is that with decreasing fuel content (increasing clay content), increased heat and mass transport barriers are developed; simultaneously low levels of thermal conductivity are maintained during the burning. PMID:25164075

  19. Tyrosyl radical generated by myeloperoxidase catalyzes the oxidative cross-linking of proteins.

    PubMed Central

    Heinecke, J W; Li, W; Francis, G A; Goldstein, J A

    1993-01-01

    Phagocytes generate H2O2 for use by a secreted heme enzyme, myeloperoxidase, to kill invading bacteria, viruses, and fungi. We have explored the possibility that myeloperoxidase might also convert L-tyrosine to a radical catalyst that cross-links proteins. Protein-bound tyrosyl residues exposed to myeloperoxidase, H2O2, and L-tyrosine were oxidized to o,o'-dityrosine, a stable product of the tyrosyl radical. The cross-linking reaction required L-tyrosine but was independent of halide and free transition metal ions; the heme poisons azide and aminotriazole were inhibitory. Activated neutrophils likewise converted polypeptide tyrosines to dityrosine. The pathway for oxidation of peptide tyrosyl residues was dependent upon L-tyrosine and was inhibited by heme poisons and catalase. Dityrosine synthesis was little affected by plasma concentrations of Cl- and amino acids, suggesting that the reaction pathway might be physiologically relevant. The requirement for free L-tyrosine and H2O2 for dityrosine formation and the inhibition by heme poisons support the hypothesis that myeloperoxidase catalyzes the cross-linking of proteins by a peroxidative mechanism involving tyrosyl radical. In striking contrast to the pathways generally used to study protein oxidation in vitro, the reaction does not require free metal ions. We speculate that protein dityrosine cross-linking by myeloperoxidase may play a role in bacterial killing or injuring normal tissue. The intense fluorescence and stability of biphenolic compounds may allow dityrosine to act as a marker for proteins oxidatively damaged by myeloperoxidase in phagocyte-rich inflammatory lesions. Images PMID:8390491

  20. Covalent DNA-Protein Cross-Linking by Phosphoramide Mustard and Nornitrogen Mustard in Human Cells.

    PubMed

    Groehler, Arnold; Villalta, Peter W; Campbell, Colin; Tretyakova, Natalia

    2016-02-15

    N,N-Bis-(2-chloroethyl)-phosphorodiamidic acid (phosphoramide mustard, PM) and N,N-bis-(2-chloroethyl)-amine (nornitrogen mustard, NOR) are the two biologically active metabolites of cyclophosphamide, a DNA alkylating drug commonly used to treat lymphomas, breast cancer, certain brain cancers, and autoimmune diseases. PM and NOR are reactive bis-electrophiles capable of cross-linking cellular biomolecules to form covalent DNA-DNA and DNA-protein cross-links (DPCs). In the present work, a mass spectrometry-based proteomics approach was employed to characterize PM- and NOR-mediated DNA-protein cross-linking in human cells. Following treatment of human fibrosarcoma cells (HT1080) with cytotoxic concentrations of PM, over 130 proteins were found to be covalently trapped to DNA, including those involved in transcriptional regulation, RNA splicing/processing, chromatin organization, and protein transport. HPLC-ESI(+)-MS/MS analysis of proteolytic digests of DPC-containing DNA from NOR-treated cells revealed a concentration-dependent formation of N-[2-[cysteinyl]ethyl]-N-[2-(guan-7-yl)ethyl]amine (Cys-NOR-N7G) conjugates, confirming that it cross-links cysteine thiols of proteins to the N7 position of guanines in DNA. Cys-NOR-N7G adduct numbers were higher in NER-deficient xeroderma pigmentosum cells (XPA) as compared with repair proficient cells. Furthermore, both XPA and FANCD2 deficient cells were sensitized to PM treatment as compared to that of wild type cells, suggesting that Fanconi anemia and nucleotide excision repair pathways are involved in the removal of cyclophosphamide-induced DNA damage. PMID:26692166