Cosmetic Preservatives as Therapeutic Corneal and Scleral Tissue Cross-Linking Agents

PubMed Central

Babar, Natasha; Kim, MiJung; Cao, Kerry; Shimizu, Yukari; Kim, Su-Young; Takaoka, Anna; Trokel, Stephen L.; Paik, David C.

2015-01-01

Purpose. Previously, aliphatic β-nitroalcohols (BNAs) have been studied as a means to chemically induce tissue cross-linking (TXL) of cornea and sclera. There are a number of related and possibly more potent agents, known as formaldehyde releasers (FARs), that are in commercial use as preservatives in cosmetics and other personal care products. The present study was undertaken in order to screen such compounds for potential clinical utility as therapeutic TXL agents. Methods. A chemical registry of 62 FARs was created from a literature review and included characteristics relevant to TXL such as molecular weight, carcinogenicity/mutagenicity, toxicity, hydrophobicity, and commercial availability. From this registry, five compounds [diazolidinyl urea (DAU), imidazolidinyl urea (IMU), sodium hydroxymethylglycinate (SMG), DMDM hydantoin (DMDM), 5-Ethyl-3,7-dioxa-1-azabicyclo [3.3.0] octane (OCT)] were selected for efficacy screening using two independent systems, an ex vivo rabbit corneal cross-linking simulation setup and incubation of cut scleral tissue pieces. Treatments were conducted at pH 7.4 or 8.5 for 30 minutes. Efficacy was evaluated using thermal denaturation temperature (Tm), and cell toxicity was studied using the trypan blue exclusion method. Results. Cross-linking effects in the five selected FARs were pH and concentration dependent. Overall, the Tm shifts were in agreement with both cornea and sclera. By comparison with BNAs previously reported upon, the FARs identified in this study were significantly more potent but with similar or better cytotoxicity. Conclusions. The FARs, a class of compounds well known to the cosmetic industry, may have utility as therapeutic TXL agents. The compounds studied thus far show promise and will be further tested. PMID:25634979

21 CFR 177.1650 - Polysulfide polymer-polyepoxy resins.

Code of Federal Regulations, 2011 CFR

2011-04-01

...(2-chloroethyl) formal Bis(dichloropropyl) formal Cross-linking agent. Butyl alcohol Solvent. Carbon black (channel process) Chlorinated paraffins Cross-linking agent. Epoxidized linseed oil Epoxidized... monobutyl ether Solvent. Magnesium chloride Methyl isobutyl ketone Solvent. Naphthalene sulfonic acid...

Polymeric micelles with ionic cores containing biodegradable cross-links for delivery of chemotherapeutic agents.

PubMed

Kim, Jong Oh; Sahay, Gaurav; Kabanov, Alexander V; Bronich, Tatiana K

2010-04-12

Novel functional polymeric nanocarriers with ionic cores containing biodegradable cross-links were developed for delivery of chemotherapeutic agents. Block ionomer complexes (BIC) of poly(ethylene oxide)-b-poly(methacylic acid) (PEO-b-PMA) and divalent metal cations (Ca(2+)) were utilized as templates. Disulfide bonds were introduced into the ionic cores by using cystamine as a biodegradable cross-linker. The resulting cross-linked micelles with disulfide bonds represented soft, hydrogel-like nanospheres and demonstrated a time-dependent degradation in the conditions mimicking the intracellular reducing environment. The ionic character of the cores allowed to achieve a very high level of doxorubicin (DOX) loading (50% w/w) into the cross-linked micelles. DOX-loaded degradable cross-linked micelles exhibited more potent cytotoxicity against human A2780 ovarian carcinoma cells as compared to micellar formulations without disulfide linkages. These novel biodegradable cross-linked micelles are expected to be attractive candidates for delivery of anticancer drugs.

In-situ cross linking of polyvinyl alcohol. [application to battery separator films

NASA Technical Reports Server (NTRS)

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

1981-01-01

A method of producing a crosslinked polyvinyl alcohol structure, such as a battery separator membrane or electrode envelope is described. An aqueous solution of a film-forming polyvinyl alcohol is admixed with an aldehyde crosslinking agent a basic pH to inhibit crosslinking. The crosslinking agent, perferably a dialdehyde such as glutaraldehyde, is used in an amount of from about 1/2 to about 20% of the theoretical amount required to crosslink all of the hydroxyl groups of the polymer. The aqueous admixture is formed into a desired physical shape, such as by casting a sheet of the solution. The sheet is then dried to form a self-supporting film. Crosslinking is then effected by immersing the film in aqueous acid solution. The resultant product has excellent properties for use as a battery separator.

The Synergy of Double Cross-linking Agents on the Properties of Styrene Butadiene Rubber Foams

PubMed Central

Shao, Liang; Ji, Zhan-You; Ma, Jian-Zhong; Xue, Chao-Hua; Ma, Zhong-Lei; Zhang, Jing

2016-01-01

Sulfur (S) cross-linking styrene butadiene rubber (SBR) foams show high shrinkage due to the cure reversion, leading to reduced yield and increased processing cost. In this paper, double cross-linking system by S and dicumyl peroxide (DCP) was used to decrease the shrinkage of SBR foams. Most importantly, the synergy of double cross-linking agents was reported for the first time to our knowledge. The cell size and its distribution of SBR foams were investigated by FESEM images, which show the effect of DCP content on the cell structure of the SBR foams. The relationships between shrinkage and crystalline of SBR foams were analyzed by the synergy of double cross-linking agents, which were demonstrated by FTIR, Raman spectra, XRD, DSC and TGA. When the DCP content was 0.6 phr, the SBR foams exhibit excellent physical and mechanical properties such as low density (0.223 g/cm3), reduced shrinkage (2.25%) and compression set (10.96%), as well as elevated elongation at break (1.78 × 103%) and tear strength (54.63 N/mm). The results show that these properties are related to the double cross-linking system of SBR foams. Moreover, the double cross-linking SBR foams present high electromagnetic interference (EMI) shielding properties compared with the S cross-linking SBR foams. PMID:27841307

The Synergy of Double Cross-linking Agents on the Properties of Styrene Butadiene Rubber Foams

NASA Astrophysics Data System (ADS)

Shao, Liang; Ji, Zhan-You; Ma, Jian-Zhong; Xue, Chao-Hua; Ma, Zhong-Lei; Zhang, Jing

2016-11-01

Sulfur (S) cross-linking styrene butadiene rubber (SBR) foams show high shrinkage due to the cure reversion, leading to reduced yield and increased processing cost. In this paper, double cross-linking system by S and dicumyl peroxide (DCP) was used to decrease the shrinkage of SBR foams. Most importantly, the synergy of double cross-linking agents was reported for the first time to our knowledge. The cell size and its distribution of SBR foams were investigated by FESEM images, which show the effect of DCP content on the cell structure of the SBR foams. The relationships between shrinkage and crystalline of SBR foams were analyzed by the synergy of double cross-linking agents, which were demonstrated by FTIR, Raman spectra, XRD, DSC and TGA. When the DCP content was 0.6 phr, the SBR foams exhibit excellent physical and mechanical properties such as low density (0.223 g/cm3), reduced shrinkage (2.25%) and compression set (10.96%), as well as elevated elongation at break (1.78 × 103%) and tear strength (54.63 N/mm). The results show that these properties are related to the double cross-linking system of SBR foams. Moreover, the double cross-linking SBR foams present high electromagnetic interference (EMI) shielding properties compared with the S cross-linking SBR foams.

Nox Complex signal and MAPK cascade pathway are cross-linked and essential for pathogenicity and conidiation of mycoparasite Coniothyrium minitans

USDA-ARS?s Scientific Manuscript database

The NADPH oxidase complex of a sclerotial mycoparasite Coniothyrium minitans, an important biocontrol agent against crop diseases caused by Sclerotinia sclerotiorum, was identified and its functions involved in conidiation and mycoparasitism were studied. Gene knock-out and complementation experimen...

A novel crosslinking strategy for preparing poly(vinyl alcohol)-based proton-conducting membranes with high sulfonation

NASA Astrophysics Data System (ADS)

Tsai, Chun-En; Lin, Chi-Wen; Hwang, Bing-Joe

This study synthesizes poly(vinyl alcohol) (PVA)-based polymer electrolyte membranes by a two-step crosslinking process involving esterization and acetal ring formation reactions. This work also uses sulfosuccinic acid (SSA) as the first crosslinking agent to form an inter-crosslinked structure and a promoting sulfonating agent. Glutaraldehyde (GA) as the second crosslinking agent, reacts with the spare OH group of PVA and forms, not only a dense structure at the outer membrane surface, but also a hydrophobic protective layer. Compared with membranes prepared by a traditional one-step crosslinking process, membranes prepared by the two-step crosslinking process exhibit excellent dissolution resistance in water. The membranes become water-insoluble even at a molar ratio of SO 3H/PVA-OH as high as 0.45. Moreover, the synthesized membranes also exhibit high proton conductivities and high methanol permeability resistance. The current study measures highest proton conductivity of 5.3 × 10 -2 S cm -1 at room temperature from one of the synthesized membranes, higher than that of the Nafion ® membrane. Methanol permeability of the synthesized membranes measures about 1 × 10 -7 cm 2 S -1, about one order of magnitude lower than that of the Nafion ® membrane.

Multifunctional curing agents and their use in improving strength of composites containing carbon fibers embedded in a polymeric matrix

DOEpatents

Vautard, Frederic; Ozcan, Soydan

2017-04-11

A functionalized carbon fiber having covalently bound on its surface a sizing agent containing epoxy groups, at least some of which are engaged in covalent bonds with crosslinking molecules, wherein each of said crosslinking molecules possesses at least two epoxy-reactive groups and at least one free functional group reactive with functional groups of a polymer matrix in which the carbon fiber is to be incorporated, wherein at least a portion of said crosslinking molecules are engaged, via at least two of their epoxy-reactive groups, in crosslinking bonds between at least two epoxy groups of the sizing agent. Composites comprised of these functionalized carbon fibers embedded in a polymeric matrix are also described. Methods for producing the functionalized carbon fibers and composites thereof are also described.

Crosslinked polyethylene foams, via EB radiation

NASA Astrophysics Data System (ADS)

Cardoso, E. C. L.; Lugão, A. B.; Andrade E. Silva, L. G.

1998-06-01

Polyethylene foams, produced by radio-induced crosslinking, show a smooth and homogeneous surface, when compared to chemical crosslinking method using peroxide as crosslinking agent. This process fosters excellent adhesive and printability properties. Besides that, closed cells, intrinsic to theses foams, imparts opitmum mechanical, shocks and insulation resistance, indicating these foams to some markets segments as: automotive and transport; buoyancy, flotation and marine: building and insulation: packaging: domestic sports and leisure goods. We were in search of an ideal foam, by adding 5 to 15% of blowing agent in LDPE. A series of preliminary trials defined 203° C as the right blowing agent decomposition temperature. At a 22.7 kGy/dose ratio, the lowest dose for providing an efficient foam was 30 kGy, for a formulation comprising 10% of azodicarbonamide in LDPE, within a 10 minutes foaming time.

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

Cross-linking chemistry of squid beak.

PubMed

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

2010-12-03

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

Investigations of the Binding of [Pt2(DTBPA)Cl2](II) and [Pt2(TPXA)Cl2](II) to DNA via Various Cross-Linking Modes

PubMed Central

Yue, Hongwei; Yang, Bo; Wang, Yan; Chen, Guangju

2013-01-01

We have constructed models for a series of platinum-DNA adducts that represent the binding of two agents, [Pt2(DTBPA)Cl2](II) and [Pt2(TPXA)Cl2](II), to DNA via inter- and intra-strand cross-linking, and carried out molecular dynamics simulations and DNA conformational dynamics calculations. The effects of trans- and cis-configurations of the centers of these di-nuclear platinum agents, and of different bridging linkers, have been investigated on the conformational distortions of platinum-DNA adducts formed via inter- and intra-strand cross-links. The results demonstrate that the DNA conformational distortions for the various platinum-DNA adducts with differing cross-linking modes are greatly influenced by the difference between the platinum-platinum distance for the platinum agent and the platinum-bound N7–N7 distance for the DNA molecule, and by the flexibility of the bridging linkers in the platinum agent. However, the effects of trans/cis-configurations of the platinum-centers on the DNA conformational distortions in the platinum-DNA adducts depend on the inter- and intra-strand cross-linking modes. In addition, we discuss the relevance of DNA base motions, including opening, shift and roll, to the changes in the parameters of the DNA major and minor grooves caused by binding of the platinum agent. PMID:24077126

Interactions of collagen molecules in the presence of N-hydroxysuccinimide activated adipic acid (NHS-AA) as a crosslinking agent.

PubMed

Zhang, Min; Wu, Kun; Li, Guoying

2011-11-01

The effect of crosslinking agent on pepsin-soluble bovine collagen solution was examined using N-hydroxysuccinimide activated adipic acid (NHS-AA) as a crosslinker. Electrophoretic patterns indicated that crosslinks formed when NHS-AA was added. A higher polarity level deduced from the changes in the fluorescence emission spectrum of pyrene in the crosslinked collagen solution indicated that the formation of well-ordered aggregates was suppressed. The random aggregation of collagens was also observed by atomic force microscopy (AFM). Furthermore, the association of collagens into fibrils was influenced by crosslinking. Self-assembly was suppressed at 37°C; however, as temperature was increased to 39°C, a small amount of NHS-AA leaded to an improvement in the ability of self-aggregation. Although more random structure was brought about by crosslinking, self-aggregation might still be promoted as temperature was increased, accompanying by the thermal stability improvement of fibrils. Copyright © 2011 Elsevier B.V. All rights reserved.

Comparison of Hydrazone Heterobifunctional Crosslinking Agents for Reversible Conjugation of Thiol-Containing Chemistry

PubMed Central

Christie, R. James; Anderson, Diana J.; Grainger, David W.

2010-01-01

Reversible covalent conjugation chemistries that allow site- and condition-specific coupling and uncoupling reactions are attractive components in nanotechnologies, bioconjugation methods, imaging and drug delivery systems. Here, we compare three heterobifunctional crosslinkers, containing both thiol- and amine- reactive chemistry, to form pH-labile hydrazones with hydrazide derivatives of the known and often published water-soluble polymer, poly[N-(2-hydroxypropyl methacrylamide)] (pHPMA), while subsequently coupling thiol-containing molecules to the crosslinker via maleimide addition. Two novel crosslinkers were prepared from the popular heterobifunctional crosslinking agent, succinimidyl-4-(N-maleimidomethyl) cyclohexane-1-carboxylate (SMCC), modified to contain either terminal aldehyde groups (i.e., 1-(N-3-propanal)-4-(N-maleimidomethyl) cyclohexane carboxamide, PMCA) or methylketone groups (i.e., 1-(N-3-butanone)-4-(N-maleimidomethyl) cyclohexane carboxamide, BMCA). A third crosslinking agent was the commercially available N-4-acetylphenyl maleimide (APM). PMCA and BMCA exhibited excellent reactivity towards hydrazide-derivatized pHPMA with essentially complete hydrazone conjugation to polymer reactive sites, while APM coupled only ~ 60% of available reactive sites on the polymer despite a 3-fold molar excess relative to polymer hydrazide groups. All polymer hydrazone conjugates bearing these bifunctional agents were then further reacted with thiol-modified tetramethylrhodamine dye, confirming crosslinker maleimide reactivity after initial hydrazone polymer conjugation. Incubation of dye-labeled polymer conjugates in phosphate buffered saline at 37°C showed that hydrazone coupling resulting from APM exhibited the greatest difference in stability between pH 7.4 and 5.0, with hydrolysis and dye release increased at pH 5.0 over a 24hr incubation period. Polymer conjugates bearing hydrazones formed from crosslinker BMCA exhibited intermediate stability with hydrolysis much greater at pH 5.0 at early time points, but hydrolysis at pH 7.4 was significant after 5 hrs. Hydrazones formed with the PMCA crosslinker showed no difference in release rates at pH 7.4 and 5.0. PMID:20695431

Method of bonding functional surface materials to substrates and applications in microtechnology and anti-fouling

DOEpatents

Feng, Xiangdong; Liu, Jun; Liang, Liang

2001-01-01

A simple and effective method to bond a thin coating of poly(N-isopropylacylamide) (NIPAAm) on a glass surface by UV photopolymerization, and the use of such a coated surface in nano and micro technology applications. A silane coupling agent with a dithiocarbamate group is provided as a photosensitizer preferably, (N,N'-diethylamine) dithiocarbamoylpropyl-(trimethoxy) silane (DATMS). The thiocarbamate group of the sensitizer is then bonded to the glass surface by coupling the silane agent with the hydroxyl groups on the glass surface. The modified surface is then exposed to a solution of NIPAAm and a crosslinking agent which may be any organic molecule having an acrylamide group and at least two double bonds in its structure, such as N, N'-methylenebisacrylamide, and a polar solvent which may be any polar liquid which will dissolve the monomer and the crosslinking agent such as acetone, water, ethanol, or combinations thereof. By exposing the glass surface to a UV light, free radicals are generated in the thiocarbamate group which then bonds to the crosslinking agent and the NIPAAm. Upon bonding, the crosslinking agent and the NIPAAm polymerize to form a thin coating of PNIPAAm bonded to the glass. Depending upon the particular configuration of the glass, the properties of the PNIPAAm allow applications in micro and nano technology.

Method of bonding functional surface materials to substrates and applications in microtechnology and antifouling

DOEpatents

Feng, Xiangdong; Liu, Jun; Liang, Liang

1999-01-01

A simple and effective method to bond a thin coating of poly(N-isopropylacylamide) (NIPAAm) on a glass surface by UV photopolymerization, and the use of such a coated surface in nano and micro technology applications. A silane coupling agent with a dithiocarbamate group is provided as a photosensitizer, preferably, (N,N'-diethylamine)dithiocarbamoylpropyl-(trimethoxy)silane (DATMS). The thiocarbamate group of the sensitizer is then bonded to the glass surface by coupling the silane agent with the hydroxyl groups on the glass surface. The modified surface is then exposed to a solution of NIPAAm and a crosslinking agent which may be any organic molecule having an acrylamide group and at least two double bonds in its structure, such as N,N'-methylenebisacrylamide, and a polar solvent which may be any polar liquid which will dissolve the monomer and the crosslinking agent such as acetone, water, ethanol, or combinations thereof. By exposing the glass surface to a UV light, free radicals are generated in the thiocarbamate group which then bonds to the crosslinking agent and the NIPAAm. Upon bonding, the crosslinking agent and the NIPAAm polymerize to form a thin coating of PNIPAAm bonded to the glass. Depending upon the particular configuration of the glass, the properties of the PNIPAAm allow applications in micro and nano technology.

Diagnosis of Fanconi Anemia by Diepoxybutane Analysis

PubMed Central

Auerbach, Arleen D.

2015-01-01

Fanconi anemia (FA) is a genetically and phenotypically heterogeneous disorder characterized by congenital malformations, progressive bone marrow failure, and predisposition to cancer, particularly hematological malignancies and solid tumors of the head and neck. The main role of FA proteins is in the repair of DNA interstrand crosslinks (ICLs). FA results from pathogenic variants in at least 16 distinct genes, causing genomic instability. Although the highly variable phenotype makes accurate diagnosis on the basis of clinical manifestations difficult in some patients, diagnosis based on a profound sensitivity to DNA crosslinking agents can be used to identify the pre-anemia patient as well as patients with aplastic anemia or leukemia who may or may not have the physical stigmata associated with the syndrome. Diepoxybutane (DEB) analysis is the preferred test for FA because other agents have higher rates of false-positive and false-negative results. PMID:25827349

Development of toughened epoxy polymers for high performance composite and ablative applications

NASA Technical Reports Server (NTRS)

Allen, V. R.

1982-01-01

A survey of current procedures for the assessment of state of cure in epoxy polymers and for the evaluation of polymer toughness as related to nature of the crosslinking agent was made to facilitate a cause-effect study of the chemical modification of epoxy polymers. Various conformations of sample morphology were examined to identify testing variables and to establish optimum conditions for the selected physical test methods. Dynamic viscoelasticity testing was examined in conjunction with chemical analyses to allow observation of the extent of the curing reaction with size of the crosslinking agent the primary variable. Specifically the aims of the project were twofold: (1) to consider the experimental variables associated with development of "extent of cure" analysis, and (2) to assess methodology of fracture energy determination and to prescribe a meaningful and reproducible procedure. The following is separated into two categories for ease of presentation.

Deficiency of the Arabidopsis Helicase RTEL1 Triggers a SOG1-Dependent Replication Checkpoint in Response to DNA Cross-Links

PubMed Central

Hu, Zhubing; Cools, Toon; Kalhorzadeh, Pooneh; Heyman, Jefri; De Veylder, Lieven

2015-01-01

To maintain genome integrity, DNA replication is executed and regulated by a complex molecular network of numerous proteins, including helicases and cell cycle checkpoint regulators. Through a systematic screening for putative replication mutants, we identified an Arabidopsis thaliana homolog of human Regulator of Telomere Length 1 (RTEL1), which functions in DNA replication, DNA repair, and recombination. RTEL1 deficiency retards plant growth, a phenotype including a prolonged S-phase duration and decreased cell proliferation. Genetic analysis revealed that rtel1 mutant plants show activated cell cycle checkpoints, specific sensitivity to DNA cross-linking agents, and increased homologous recombination, but a lack of progressive shortening of telomeres, indicating that RTEL1 functions have only been partially conserved between mammals and plants. Surprisingly, RTEL1 deficiency induces tolerance to the deoxynucleotide-depleting drug hydroxyurea, which could be mimicked by DNA cross-linking agents. This resistance does not rely on the essential replication checkpoint regulator WEE1 but could be blocked by a mutation in the SOG1 transcription factor. Taken together, our data indicate that RTEL1 is required for DNA replication and that its deficiency activates a SOG1-dependent replication checkpoint. PMID:25595823

Combined use of chondroitinase-ABC, TGF-β1 and collagen crosslinking agent lysyl oxidase to engineer functional neotissues for fibrocartilage repair

PubMed Central

Makris, Eleftherios A.; MacBarb, Regina F.; Paschos, Nikolaos K.; Hu, Jerry C.; Athanasiou, Kyriacos A.

2014-01-01

Patients suffering from damaged or diseased fibrocartilages currently have no effective long-term treatment options. Despite their potential, engineered tissues suffer from inferior biomechanical integrity and an inability to integrate in vivo. The present study identifies a treatment regimen (including the biophysical agent chondroitinase-ABC, the biochemical agent TGF-β1, and the collagen crosslinking agent lysyl oxidase) to prime highly cellularized, scaffold-free neofibrocartilage implants, effecting continued improvement in vivo. We show these agents drive in vitro neofibrocartilage matrix maturation toward synergistically enhanced Young’s modulus and ultimate tensile strength values, which were increased 245% and 186%, respectively, over controls. Furthermore, an in vitro fibrocartilage defect model found this treatment regimen to significantly increase the integration tensile properties between treated neofibrocartilage and native tissue. Through translating this technology to an in vivo fibrocartilage defect model, our results indicate, for the first time, that a pre-treatment can prime neofibrocartilage for significantly enhanced integration potential in vivo, with interfacial tensile stiffness and strength increasing by 730% and 745%, respectively, compared to integration values achieved in vitro. Our results suggest that specifically targeting collagen assembly and organization is a powerful means to augment overall neotissue mechanics and integration potential toward improved clinical feasibility. PMID:24840619

Combined use of chondroitinase-ABC, TGF-β1, and collagen crosslinking agent lysyl oxidase to engineer functional neotissues for fibrocartilage repair.

PubMed

Makris, Eleftherios A; MacBarb, Regina F; Paschos, Nikolaos K; Hu, Jerry C; Athanasiou, Kyriacos A

2014-08-01

Patients suffering from damaged or diseased fibrocartilages currently have no effective long-term treatment options. Despite their potential, engineered tissues suffer from inferior biomechanical integrity and an inability to integrate in vivo. The present study identifies a treatment regimen (including the biophysical agent chondroitinase-ABC, the biochemical agent TGF-β1, and the collagen crosslinking agent lysyl oxidase) to prime highly cellularized, scaffold-free neofibrocartilage implants, effecting continued improvement in vivo. We show these agents drive in vitro neofibrocartilage matrix maturation toward synergistically enhanced Young's modulus and ultimate tensile strength values, which were increased 245% and 186%, respectively, over controls. Furthermore, an in vitro fibrocartilage defect model found this treatment regimen to significantly increase the integration tensile properties between treated neofibrocartilage and native tissue. Through translating this technology to an in vivo fibrocartilage defect model, our results indicate, for the first time, that a pre-treatment can prime neofibrocartilage for significantly enhanced integration potential in vivo, with interfacial tensile stiffness and strength increasing by 730% and 745%, respectively, compared to integration values achieved in vitro. Our results suggest that specifically targeting collagen assembly and organization is a powerful means to augment overall neotissue mechanics and integration potential toward improved clinical feasibility. Copyright © 2014 Elsevier Ltd. All rights reserved.

Analysis of the tertiary structure of the ribonuclease P ribozyme-substrate complex by site-specific photoaffinity crosslinking.

PubMed Central

Harris, M E; Kazantsev, A V; Chen, J L; Pace, N R

1997-01-01

Bacterial ribonuclease P (RNase P), an endonuclease involved in tRNA maturation, is a ribonucleoprotein containing a catalytic RNA. The secondary structure of this ribozyme is well-established, and a low-resolution model of the three-dimensional structure of the ribozyme-substrate complex has been proposed based on site-specific crosslinking and phylogenetic comparative data [Harris ME et al., 1994 EMBO J 13:3953-3963]. However, several substructures of that model were poorly constrained by the available data. In the present analysis, additional constraints between elements within the Escherichia coli RNase P RNA-pre-tRNA complex were determined by intra- and intermolecular crosslinking experiments. Circularly permuted RNase P RNAs were used to position an azidophenacyl photoactive crosslinking agent specifically at strategic sites within the ribozyme-substrate complex. Crosslink sites were mapped by primer extension and confirmed by analysis of the mobility of the crosslinked RNA lariats on denaturing acrylamide gels relative to circular and linear RNA standards. Crosslinked species generally retained significant catalytic activity, indicating that the results reflect the native ribozyme structure. The crosslinking results support the general configuration of the structure model and predicate new positions and orientations for helices that were previously poorly constrained by the data set. The expanded library of crosslinking constraints was used, together with secondary and tertiary structure identified by phylogenetic sequence comparisons, to refine significantly the model of RNase P RNA with bound substrate pre-tRNA. The crosslinking results and data from chemical-modification and mutational studies are discussed in the context of the current structural perspective on this ribozyme. PMID:9174092

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

PubMed

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

2005-10-07

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

Pervaporation separation of thiophene-heptane mixtures with polydimethylsiloxane (PDMS) membrane for desulfurization.

PubMed

Chen, Jian; Li, Jiding; Qi, Rongbin; Ye, Hong; Chen, Cuixian

2010-01-01

Cross-linked polydimethylsiloxane (PDMS)-polyetherimide (PEI) composite membranes were prepared, in which asymmetric microporous PEI membrane prepared with phase inversion method was acted as the microporous supporting layer in the flat-plate composite membrane. Membrane characterization was conducted by Fourier transform infrared and scanning electronic microscopy analysis. The composite membranes were employed in pervaporation separation of n-heptane-thiophene mixtures. Effect of amount of PDMS, cross-linking temperature, amount of cross-linking agent, and cross-linking time on the separation efficiency of n-heptane-thiophene mixtures was investigated experimentally. Experiment results demonstrated that 80-100 degrees degrees C of cross-linking temperature was more preferable for practical application, as the amount of cross-linking agent was up to 20 wt.%, and 25 wt.% of PDMS amount was more optimal as far as flux and sulfur enrichment factor were concerned. In addition, the swelling degree of and stableness of composite membrane during long-time operation were studied, which should be significant for practical application.

Genipin crosslinking reduced the immunogenicity of xenogeneic decellularized porcine whole-liver matrices through regulation of immune cell proliferation and polarization

NASA Astrophysics Data System (ADS)

Wang, Yujia; Bao, Ji; Wu, Xiujuan; Wu, Qiong; Li, Yi; Zhou, Yongjie; Li, Li; Bu, Hong

2016-04-01

Decellularized xenogeneic whole-liver matrices are plausible biomedical materials for the bioengineering of liver transplantation. A common method to reduce the inflammatory potential of xenogeneic matrices is crosslinking. Nevertheless, a comprehensive analysis of the immunogenic features of cross-linked decellularized tissue is still lacking. We aimed to reduce the immunogenicity of decellularized porcine whole-liver matrix through crosslinking with glutaraldehyde or genipin, a new natural agent, and investigated the mechanism of the immune-mediated responses. The histologic assessment of the host’s immune reaction activated in response to these scaffolds, as well as the M1/M2 phenotypic polarization profile of macrophages, was studied in vivo. The genipin-fixed scaffold elicited a predominantly M2 phenotype response, while the glutaraldehyde-fixed scaffold resulted in disrupted host tissue remodeling and a mixed macrophage polarization profile. The specific subsets of immune cells involved in the responses to the scaffolds were identified in vitro. Crosslinking alleviated the host response by reducing the proliferation of lymphocytes and their subsets, accompanied by a decreased release of both Th1 and Th2 cytokines. Therefore, we conclude that the natural genipin crosslinking could lower the immunogenic potential of xenogeneic decellularized whole-liver scaffolds.

The effect of electron beam irradiation on the mechanical properties of pineapple leaf fibre (PALF) reinforced high impact polystyrene (HIPS) composites

NASA Astrophysics Data System (ADS)

Siregar, J. P.; Sapuan, S. M.; Rahman, M. Z. A.; Zaman, H. M. D. K.

2010-05-01

The effects of electron beam irradiation on the mechanical properties of pineapple leaf fibre reinforced high impact polystyrene (HIPS) composites were studied. Two types of crosslinking agent that has been used in this study were trimethylolpropane triacrylate (TMPTA) and tripropylene gylcol diacrylate (TPGDA). A 50 wt.% of PALF was blended with HIPS and crosslinking agent using Brabender melt mixer at 165 °C. The composites were then irradiated using a 3 MeV electron beam accelerator with dosage of 0-100 kGy. The tensile strength, tensile modulus, flexural strength, flexural modulus, notched and unnotched impat and hardness of composites were measured and the effects of crosslinking agent were also compared.

Smart swelling biopolymer microparticles by a microfluidic approach: synthesis, in situ encapsulation and controlled release.

PubMed

Fang, Aiping; Cathala, Bernard

2011-01-01

This paper reports a microfluidic synthesis of biopolymer microparticles aiming at smart swelling. Monodisperse aqueous emulsion droplets comprising biopolymer and its cross-linking agent were formed in mineral oil and solidified in the winding microfluidic channels by in situ chaotic mixing, which resulted in internal chemical gelation for hydrogels. The achievement of pectin microparticles from in situ mixing pectin with its cross-linking agent, calcium ions, successfully demonstrates the reliability of this microfluidic synthesis approach. In order to achieve hydrogels with smart swelling, the following parameters and their impacts on the swelling behaviour, stability and morphology of microparticles were investigated: (1) the type of biopolymers (alginate or mixture of alginate and carboxymethylcellulose, A-CMC); (2) rapid mixing; (3) concentration and type of cross-linking agent. Superabsorbent microparticles were obtained from A-CMC mixture by using ferric chloride as an additional external cross-linking agent. The in situ encapsulation of a model protein, bovine serum albumin (BSA), was also carried out. As a potential protein drug-delivery system, the BSA release behaviours of the biopolymer particles were studied in simulated gastric and intestinal fluids. Compared with alginate and A-CMC microparticles cross-linked with calcium ions, A-CMC microparticles cross-linked with both calcium and ferric ions demonstrate a significantly delayed release. The controllable release profile, the facile encapsulation as well as their biocompatibility, biodegradability, mucoadhesiveness render this microfluidic approach promising in achieving biopolymer microparticles as protein drug carrier for site-specific release. Copyright © 2010 Elsevier B.V. All rights reserved.

Antimocrobial Polymer

DOEpatents

McDonald, William F.; Huang, Zhi-Heng; Wright, Stacy C.

2005-09-06

A polymeric composition having antimicrobial properties and a process for rendering the surface of a substrate antimicrobial are disclosed. The composition comprises a crosslinked chemical combination of (i) a polymer having amino group-containing side chains along a backbone forming the polymer, (ii) an antimicrobial agent selected from quaternary ammonium compounds, gentian violet compounds, substituted or unsubstituted phenols, biguanide compounds, iodine compounds, and mixtures thereof, and (iii) a crosslinking agent containing functional groups capable of reacting with the amino groups. In one embodiment, the polymer is a polyamide formed from a maleic anhydride or maleic acid ester monomer and alkylamines thereby producing a polyamide having amino substituted alkyl chains on one side of the polyamide backbone; the crosslinking agent is a phosphine having the general formula (A)3P wherein A is hydroxyalkyl; and the antimicrobial agent is chlorhexidine, dimethylchlorophenol, cetyl pyridinium chloride, gentian violet, triclosan, thymol, iodine, and mixtures thereof.

Antimicrobial Polymer

DOEpatents

McDonald, William F.; Wright, Stacy C.; Taylor, Andrew C.

2004-09-28

A polymeric composition having antimicrobial properties and a process for rendering the surface of a substrate antimicrobial are disclosed. The polymeric composition comprises a crosslinked chemical combination of (i) a polymer having amino group-containing side chains along a backbone forming the polymer, (ii) an antimicrobial agent selected from metals, metal alloys, metal salts, metal complexes and mixtures thereof, and (iii) a crosslinking agent containing functional groups capable of reacting with the amino groups. In one example embodiment, the polymer is a polyamide formed from a maleic anhydride or maleic acid ester monomer and alkylamines thereby producing a polyamide having amino substituted alkyl chains on one side of the polyamide backbone; the crosslinking agent is a phosphine having the general formula (A).sub.3 P wherein A is hydroxyalkyl; and the metallic antimicrobial agent is selected from chelated silver ions, silver metal, chelated copper ions, copper metal, chelated zinc ions, zinc metal and mixtures thereof.

Dithiothreitol (DTT) Acts as a Specific, UV-inducible Cross-linker in Elucidation of Protein–RNA Interactions*

PubMed Central

Zaman, Uzma; Richter, Florian M.; Hofele, Romina; Kramer, Katharina; Sachsenberg, Timo; Kohlbacher, Oliver; Lenz, Christof; Urlaub, Henning

2015-01-01

Protein–RNA cross-linking by UV irradiation at 254 nm wavelength has been established as an unbiased method to identify proteins in direct contact with RNA, and has been successfully applied to investigate the spatial arrangement of protein and RNA in large macromolecular assemblies, e.g. ribonucleoprotein-complex particles (RNPs). The mass spectrometric analysis of such peptide-RNA cross-links provides high resolution structural data to the point of mapping protein–RNA interactions to specific peptides or even amino acids. However, the approach suffers from the low yield of cross-linking products, which can be addressed by improving enrichment and analysis methods. In the present article, we introduce dithiothreitol (DTT) as a potent protein–RNA cross-linker. In order to evaluate the efficiency and specificity of DTT, we used two systems, a small synthetic peptide from smB protein incubated with U1 snRNA oligonucleotide and native ribonucleoprotein complexes from S. cerevisiae. Our results unambiguously show that DTT covalently participates in cysteine-uracil crosslinks, which is observable as a mass increment of 151.9966 Da (C4H8S2O2) upon mass spectrometric analysis. DTT presents advantages for cross-linking of cysteine containing regions of proteins. This is evidenced by comparison to experiments where (tris(2-carboxyethyl)phosphine) is used as reducing agent, and significantly less cross-links encompassing cysteine residues are found. We further propose insertion of DTT between the cysteine and uracil reactive sites as the most probable structure of the cross-linking products. PMID:26450613

Fanconi Anemia Proteins, DNA Interstrand Crosslink Repair Pathways, and Cancer Therapy

PubMed Central

Andreassen, Paul R.; Ren, Keqin

2016-01-01

DNA interstrand crosslinkers, a chemically diverse group of compounds which also induce alkylation of bases and DNA intrastrand crosslinks, are extensively utilized for cancer therapy. Understanding the cellular response to DNA damage induced by these agents is critical for more effective utilization of these compounds and for the identification of novel therapeutic targets. Importantly, the repair of DNA interstrand crosslinks (ICLs) involves many distinct DNA repair pathways, including nucleotide excision repair, translesion synthesis (TLS), and homologous recombination (HR). Additionally, proteins implicated in the pathophysiology of the multigenic disease Fanconi anemia (FA) have a role in the repair of ICLs that is not well understood. Cells from FA patients are hypersensitive to agents that induce ICLs, therefore FA proteins are potentially novel therapeutic targets. Here we will review current research directed at identifying FA genes and understanding the function of FA proteins in DNA damage responses. We will also examine interactions of FA proteins with other repair proteins and pathways, including signaling networks, which are potentially involved in ICL repair. Potential approaches to the modulation of FA protein function to enhance therapeutic outcome will be discussed. Also, mutation of many genes that encode proteins involved in ICL repair, including FA genes, increases susceptibility to cancer. A better understanding of these pathways is therefore critical for the design of individualized therapies tailored to the genetic profile of a particular malignancy. For this purpose, we will also review evidence for the association of mutation of FA genes with cancer in non-FA patients. PMID:19200054

Drilling fluid containing a copolymer filtration control agent

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

Enright, D.P.; Lucas, J.M.; Perricone, A.C.

1981-10-06

The invention relates to an aqueous drilling fluid composition, a filtration control agent for utilization in said aqueous drilling fluid, and a method of forming a filter cake on the wall of a well for the reduction of filtrate from said drilling fluid, by utilization of a copolymer of: (1) a (Meth) acrylamido alkyl sulfonic acid or alkali metal salt thereof; and (2) a (Meth) acrylamide or n-alkyl (Meth) acrylamide. The copolymer may be cross-linked with a quaternary ammonium salt cross-linking agent.

Drilling fluid containing a copolymer filtration control agent

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

Lucas, J. M.

1985-10-15

The invention relates to an aqueous drilling fluid composition, a filtration control agent for utilization in said aqueous drilling fluid, and a method of forming a filter cake on the wall of a well for the reduction of filtrate from said drilling fluid, by utilization of a copolymer of: a (meth) acrylamido alkyl sulfonic acid or alkali metal salt thereof; and N, N-dialkyl (meth) acrylamide. The copolymer may be cross-linked with N,N'-methylenebisacrylamide or other appropriate cross-linking agent.

Active oligonucleotides incorporating alkylating an agent as potential sequence- and base selective modifier of gene expression.

PubMed

Sasaki, S

2001-04-01

A number of cross-linking (alkylating) agents have been developed and incorporated into the oligonulceotides for sequence selective control of gene expression. Recently, potential application of such active oligonucleotides has been expanding from use for improvement of inhibition efficiency to new biotechnology that may enable chemical alteration of genetic information. These interests in active oligonucleotides have encouraged the generation of new cross-linking agents that exhibit high efficiency for application of either in vitro or in vivo. This mini review summarizes structures of alkylating agents, in particular, a new basic skeleton for cross-linking, a 2'-deoxyribose derivative of 2-amino-6-vinylpurine that has been recently developed by the author's group. The 2-amino-6-vinylpurine has been shown to form a complex with cytidine under acidic conditions, and brings the vinyl and the amino reactive groups into proximity to achieve efficient alkylation. A new strategy was designed so that the reactivity of 2-amino-6-vinylpurine can be induced from the corresponding phenylsulfoxide derivative within a duplex with the complementary strand. The validity of the new strategy has been proven by achievement of cytidine-selective cross-linking with remarkably efficiency.

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

NASA Astrophysics Data System (ADS)

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

2017-04-01

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

Electroactive hydrogel comprising poly(methyl 2-acetamido acrylate) for an artificial actuator

NASA Astrophysics Data System (ADS)

Ha, Eun-Ju; Kim, Bong-Soo; Park, Chun-ho; Lee, Jang-Oo; Paik, Hyun-jong

2013-08-01

A poly(methyl 2-acetamidoacrylic acrylate) (MAA) hydrogel was developed for use in an artificial actuator. The equilibrium swelling ratio of the MAA hydrogel was observed at different pH values with different concentrations of cross-linking agent; the hydrogel containing 2% cross-linking agent exhibited the maximum equilibrium swelling ratio at pH 10. The bending behavior of the MAA hydrogel under an electric field was measured in aqueous NaCl. The actuation response of the MAA hydrogel occurred via reversible bending behavior at 6 V. It was found that the MAA hydrogel features stable bending behavior over consecutive cycles in aqueous NaCl at different voltages depending on the cross-linking agent. Hence, the MAA hydrogel can be utilized as an artificial actuator using electrical stimulus.

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

PubMed

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

2017-01-22

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

Influence of different crosslinking treatments on the physical properties of collagen membranes.

PubMed

Charulatha, V; Rajaram, A

2003-02-01

The physical properties of collagen-based biomaterials are profoundly influenced by the method and extent of crosslinking. In this study, the influence of various crosslinking treatments on the physical properties of reconstituted collagen membranes was assessed. Five crosslinking agents viz., GTA, DMS, DTBP, a combination of DMS and GTA and acyl azide method were used to stabilize collagen matrices. Crosslinking density, swelling ratio, thermo-mechanical properties, stress-strain characteristics and resistance to collagenase digestion were determined to evaluate the physical properties of crosslinked matrices. GTA treatment induced the maximum number of crosslinks (13) while DMS treatment induced the minimum (7). Of the two diimidoesters (DMS and DTBP), DTBP was a more effective crosslinking agent due to the presence of disulphide bonds in the DTBP crosslinks. T(s) for DTBP and DMS crosslinked collagen were 80 degrees C and 70 degrees C, and their HIT values were 5.4 and 2.85MN/m(2), respectively. Low concentration of GTA (0.01%) increased the crosslinking density of an already crosslinked matrix (DMS treated matrix) from 7 to 12. Lowest fracture energy was observed for the acyl azide treated matrix (0.61MJ/m(3)) while the highest was observed for the GTA treated matrix (1.97MJ/m(3)). The tensile strength of GTA treated matrix was maximum (12.4MPa) and that of acyl azide treated matrix was minimum (7.2MPa). GTA, DTBP and acyl azide treated matrices were equally resistant to collagenase degradation with approximately 6% solubilization after 5h while the DMS treated was least stable with 52.4% solubilization after the same time period. The spatial orientation of amino acid side chain residues on collagen plays an important role in determining the crosslinking density and consequent physical properties of the collagen matrix.

Diffusion of ionic and non-ionic contrast agents in articular cartilage with increased cross-linking--contribution of steric and electrostatic effects.

PubMed

Kulmala, K A M; Karjalainen, H M; Kokkonen, H T; Tiitu, V; Kovanen, V; Lammi, M J; Jurvelin, J S; Korhonen, R K; Töyräs, J

2013-10-01

To investigate the effect of threose-induced collagen cross-linking on diffusion of ionic and non-ionic contrast agents in articular cartilage. Osteochondral plugs (Ø=6mm) were prepared from bovine patellae and divided into two groups according to the contrast agent to be used in contrast enhanced computed tomography (CECT) imaging: (I) anionic ioxaglate and (II) non-ionic iodixanol. The groups I and II contained 7 and 6 sample pairs, respectively. One of the paired samples served as a reference while the other was treated with threose to induce collagen cross-linking. The equilibrium partitioning of the contrast agents was imaged after 24h of immersion. Fixed charge density (FCD), water content, contents of proteoglycans, total collagen, hydroxylysyl pyridinoline (HP), lysyl pyridinoline (LP) and pentosidine (Pent) cross-links were determined as a reference. The equilibrium partitioning of ioxaglate (group I) was significantly (p=0.018) lower (-23.4%) in threose-treated than control samples while the equilibrium partitioning of iodixanol (group II) was unaffected by the threose-treatment. FCD in the middle and deep zones of the cartilage (p<0.05) and contents of Pent and LP (p=0.001) increased significantly due to the treatment. However, the proteoglycan concentration was not systematically altered after the treatment. Water content was significantly (-3.5%, p=0.007) lower after the treatment. Since non-ionic iodixanol showed no changes in partition after cross-linking, in contrast to anionic ioxaglate, we conclude that the cross-linking induced changes in charge distribution have greater effect on diffusion compared to the cross-linking induced changes in steric hindrance. Copyright © 2013 IPEM. Published by Elsevier Ltd. All rights reserved.

Deficiency of the Arabidopsis helicase RTEL1 triggers a SOG1-dependent replication checkpoint in response to DNA cross-links.

PubMed

Hu, Zhubing; Cools, Toon; Kalhorzadeh, Pooneh; Heyman, Jefri; De Veylder, Lieven

2015-01-01

To maintain genome integrity, DNA replication is executed and regulated by a complex molecular network of numerous proteins, including helicases and cell cycle checkpoint regulators. Through a systematic screening for putative replication mutants, we identified an Arabidopsis thaliana homolog of human Regulator of Telomere Length 1 (RTEL1), which functions in DNA replication, DNA repair, and recombination. RTEL1 deficiency retards plant growth, a phenotype including a prolonged S-phase duration and decreased cell proliferation. Genetic analysis revealed that rtel1 mutant plants show activated cell cycle checkpoints, specific sensitivity to DNA cross-linking agents, and increased homologous recombination, but a lack of progressive shortening of telomeres, indicating that RTEL1 functions have only been partially conserved between mammals and plants. Surprisingly, RTEL1 deficiency induces tolerance to the deoxynucleotide-depleting drug hydroxyurea, which could be mimicked by DNA cross-linking agents. This resistance does not rely on the essential replication checkpoint regulator WEE1 but could be blocked by a mutation in the SOG1 transcription factor. Taken together, our data indicate that RTEL1 is required for DNA replication and that its deficiency activates a SOG1-dependent replication checkpoint. © 2015 American Society of Plant Biologists. All rights reserved.

Novel magnetic cross-linked lipase aggregates for improving the resolution of (R, S)-2-octanol.

PubMed

Liu, Ying; Guo, Chen; Liu, Chun-Zhao

2015-03-01

Novel magnetic cross-linked lipase aggregates were fabricated by immobilizing the cross-linked lipase aggregates onto magnetic particles with a high number of -NH2 terminal groups using p-benzoquinone as the cross-linking agent. At the optimal fabrication conditions, 100% of immobilization efficiency and 139% of activity recovery of the magnetic cross-linked lipase aggregates were achieved. The magnetic cross-linked lipase aggregates were able to efficiently resolve (R, S)-2-octanol, and retained 100% activity and 100% enantioselectivity after 10 cycles of reuse, whereas the cross-linked lipase aggregates only retained about 50% activity and 70% enantioselectivity due to insufficient cross-linking. These results provide a great potential for industrial applications of the magnetic cross-linked lipase aggregates. © 2014 Wiley Periodicals, Inc.

Crosslinking of aromatic polyamides via pendant propargyl groups

NASA Technical Reports Server (NTRS)

St.clair, A. K.; St.clair, T. L.; Barrick, J. D.; Campbell, F. J.

1980-01-01

Methods for crosslinking N-methyl substituted aromatic polyamides were investigated in an effort to improve the applicability of these polymers as matrix resins for Kavlar trademark fiber composites. High molecular weight polymers were prepared from isophthaloyl dichloride and 4,4'- bis(methylamino)diphenylmethane with varying proportions of the N,N'bispropargyl diamine incorporated as a crosslinking agent. The propargylcontaining diamines were crosslinked thermally and characterized by infrared spectroscopy, differential scanning calorimetry, and thermogravimetric analysis. Attempts were also made to crosslink polyamide films by exposure to ultraviolet light, electron beam, and gamma radiation.

Synthesis of structurally diverse major groove DNA interstrand crosslinks using three different aldehyde precursors

PubMed Central

Mukherjee, Shivam; Guainazzi, Angelo; Schärer, Orlando D.

2014-01-01

DNA interstrand crosslinks (ICLs) are extremely cytotoxic lesions that block essential cellular processes, such as replication and transcription. Crosslinking agents are widely used in cancer chemotherapy and form an array of structurally diverse ICLs. Despite the clinical success of these agents, resistance of tumors to crosslinking agents, for example, through repair of these lesions by the cellular machinery remains a problem. We have previously reported the synthesis of site-specific ICLs mimicking those formed by nitrogen mustards to facilitate the studies of cellular responses to ICL formation. Here we extend these efforts and report the synthesis of structurally diverse major groove ICLs that induce severe, little or no distortion in the DNA. Our approach employs the incorporation of aldehyde precursors of different lengths into complementary strands and ICL formation using a double reductive amination with a variety of amines. Our studies provide insight into the structure and reactivity parameters of ICL formation by double reductive amination and yield a set of diverse ICLs that will be invaluable for exploring structure–activity relationships in ICL repair. PMID:24782532

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.

Gas-phase transfer of polymer cross-linking agents and by-products to solid oral pharmaceuticals.

PubMed

Maus, Russell G; Li, Min; Clement, Christopher M; Kinzer, Jeffery A

2007-11-05

In the pharmaceutical industry, solid oral compressed tablets (OCT) are frequently transported in bulk containers prior to packaging. While in this state, the product is generally protected from interaction with liquid and solid contaminants by physical barriers (e.g., polyethylene bags, drums, etc.). Vapor phase contamination, although generally less frequently observed, is possible. A specific example of the detection and identification of volatile by-products (acetophenone and 2-phenyl-2-propanol) of a common polymer cross-linking agent (dicumyl peroxide) is presented. The product tablets were compressed, placed into double polyethylene bags, and subsequently placed into a polyethylene drum for shipment overseas. To cushion the product during transit, a cross-linked polyethylene foam disk (designed to fit into the bottom of the drum) was placed below the bag of tablets. Initially, these contaminants were detected by HPLC with UV detection at the receiving laboratory, and assumed to be degradates of the active components of the product. Further analysis showed that neither the collected UV absorbance data nor the observed levels of the contaminants were consistent with known degradates of the product. Liquid extraction followed by GC-MS analysis of the product as well as the cross-linked foam disk exhibited measurable quantities of the contaminants in question. Vapor phase transfer of these cross-linking agent by-products, originating in the cross-linked foam pads, was determined to be the root cause for the presence of these compounds in the product.

Microorganism immobilization

DOEpatents

Compere, Alicia L.; Griffith, William L.

1981-01-01

Live metabolically active microorganisms are immobilized on a solid support by contacting particles of aggregate material with a water dispersible polyelectrolyte such as gelatin, crosslinking the polyelectrolyte by reacting it with a crosslinking agent such as glutaraldehyde to provide a crosslinked coating on the particles of aggregate material, contacting the coated particles with live microorganisms and incubating the microorganisms in contact with the crosslinked coating to provide a coating of metabolically active microorganisms. The immobilized microorganisms have continued growth and reproduction functions.

Reduction-responsive interlayer-crosslinked micelles prepared from star-shaped copolymer via click chemistry for drug controlled release

NASA Astrophysics Data System (ADS)

Dai, Yu; Wang, Hongquan; Zhang, Xiaojin

2017-12-01

To improve the stability of polymeric micelles, here we describe interlayer-crosslinked micelles prepared from star-shaped copolymer via click chemistry. The formation of interlayer-crosslinked micelles was investigated and confirmed by proton nuclear magnetic resonance, Fourier-transform infrared spectroscopy, and fluorescence spectroscopy. The morphology of un-crosslinked micelles and crosslinked micelles observed by transmission electron microscope is both uniform nano-sized spheres (approximately 20 nm). The crosslinking enhances the stability of polymeric micelles and improves the drug loading capacity of polymeric micelles. The interlayer-crosslinked micelles prepared from star-shaped copolymer and a crosslinker containing a disulfide bond are reduction-responsive and can release the drug quickly in the presence of the reducing agents such as glutathione (GSH).

DNA interstrand cross-link repair: understanding role of Fanconi anemia pathway and therapeutic implications.

PubMed

Shukla, Pallavi; Solanki, Avani; Ghosh, Kanjaksha; Vundinti, Babu Rao

2013-11-01

Interstrand cross-links (ICLs) are extremely toxic DNA lesions that prevent DNA double-helix separation due to the irreversible covalent linkage binding of some agents on DNA strands. Agents that induce these ICLs are thus widely used as chemotherapeutic drugs but may also lead to tumor growth. Fanconi anemia (FA) is a rare genetic disorder that leads to ICL sensitivity. This review provides update on current understanding of the role of FA proteins in repairing ICLs at various stages of cell cycle. We also discuss link between DNA cross-link genotoxicity caused by aldehydes in FA pathway. Besides this, we summarize various ICL agents that act as drugs to treat different types of tumors and highlight strategies for modulating ICL sensitivity for therapeutic interventions that may be helpful in controlling cancer and life-threatening disease, FA. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Adsorption of hexavalent chromium on modified corn stalk using different cross-linking agents

NASA Astrophysics Data System (ADS)

Chen, Suhong; Zhu, Yi; Han, Zhijun; Feng, Gao; Jia, Yuling; Fu, Kaifang; Yue, Qinyan

2017-12-01

In this study, four different types of adsorbents modified from corn stalk were synthesized after the reaction with epichlorohydrin, N,N-dimethylformamide, triethylamine and different cross-linking agents. The surface functional groups and thermal stability of modified corn stalk (MCSs) were characterized using FTIR and TG analysis, respectively. The feasibility of using MCSs to remove Cr(VI) were evaluated. Adsorption isotherms were determined and modeled with Langmuir, Freundlich and Temkin equations. The experimental results showed that MCS modified using diethylenetriamine (DETA) had the best modification effect, and the adsorption capacity of Cr(VI) reached as high as 227.27 mg/g at 323 K. Thermodynamic study showed that the Cr(VI) adsorption onto MCSs was endothermic processes. As a result, MCS by using DETA as cross-linking agent has good potential for the removal of Cr(VI) from aqueous solutions.

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

Applications of FT-IR spectroscopy to the studies of esterification and crosslinking of cellulose by polycarboxylic acids: Part II. The performance of the crosslinked cotton fabrics

NASA Astrophysics Data System (ADS)

Wei, Weishu; Yang, Charles Q.

1998-06-01

Durable press finishing processes are commonly used in the textile industry to produce wrinkle-free cotton fabrics and garments. A durable press finishing agent forms covalent bands with cellulosic hydroxyl groups, thus crosslinking the cellulose molecules. The crosslinking of cellulose increases wrinkle resistance of the treated cotton fabric and reduces fabric mechanical strength. Wrinkle recovery angle (WRA) and tensile strength are the two most important parameters used to evaluate the performance of the crosslinked cotton fabrics and garments. In this study, we investigated the correlation between WRA and tensile strength on one hand, and the amount of crosslinkages formed by the crosslinking agents including dimethyloldihydroxylethyleneurea (DMDHEU) and 1,2,3,4-butanetetracarboxylic acid (BTCA) determined by FT-IR spectroscopy on the other hand. Linear regression curves between the carbonyl band absorbance, and WRA and tensile strength of the treated cotton fabric were developed. The data indicated that FT-IR spectroscopy is a reliable technique for predicting the performance of durable press finished cotton fabrics, therefore can be used as a convenient instrumental method for quality control in the textile and garment industry.

Poly(iohexol) nanoparticles as contrast agents for in vivo X-ray computed tomography imaging.

PubMed

Yin, Qian; Yap, Felix Y; Yin, Lichen; Ma, Liang; Zhou, Qin; Dobrucki, Lawrence W; Fan, Timothy M; Gaba, Ron C; Cheng, Jianjun

2013-09-18

Biocompatible poly(iohexol) nanoparticles, prepared through cross-linking of iohexol and hexamethylene diisocyanate followed by coprecipitation of the resulting cross-linked polymer with mPEG-polylactide, were utilized as contrast agents for in vivo X-ray computed tomography (CT) imaging. Compared to conventional small-molecule contrast agents, poly(iohexol) nanoparticles exhibited substantially protracted retention within the tumor bed and a 36-fold increase in CT contrast 4 h post injection, which makes it possible to acquire CT images with improved diagnosis accuracy over a broad time frame without multiple administrations.

Structural interactions between retroviral Gag proteins examined by cysteine cross-linking.

PubMed Central

Hansen, M S; Barklis, E

1995-01-01

We have examined structural interactions between Gag proteins within Moloney murine leukemia virus (M-MuLV) particles by making use of the cysteine-specific cross-linking agents iodine and bis-maleimido hexane. Virion-associated wild-type M-MuLV Pr65Gag proteins in immature particles were intermolecularly cross-linked at cysteines to form Pr65Gag oligomers, from dimers to pentamers or hexamers. Following a systematic approach of cysteine-to-serine mutagenesis, we have shown that cross-linking of Pr65Gag occurred at cysteines of the nucleocapsid (NC) Cys-His motif, suggesting that the Cys-His motifs within virus particles are packed in close proximity. The M-MuLV Pr65Gag protein did not cross-link to the human immunodeficiency virus Pr55Gag protein when the two molecules were coexpressed, indicating either that they did not coassemble or that heterologous Gag proteins were not in close enough proximity to be cross-linked. Using an assembly-competent, protease-minus, cysteine-minus Pr65Gag protein as a template, novel cysteine residues were generated in the M-MuLV capsid domain major homology region (MHR). Cross-linking of proteins containing MHR cysteines showed above-background levels of Gag-Gag dimers but also identified a novel cellular factor, present in virions, that cross-linked to MHR residues. Although the NC cysteine mutation was compatible with M-MuLV particle assembly, deletions of the NC domain were not tolerated. These results suggest that the Cys-His motif is held in close proximity within immature M-MuLV particles by interactions between CA domains and/or non-Cys-His motif domains of the NC. PMID:7815493

Identifying chemicals of concern in hydraulic fracturing fluids used for oil production.

PubMed

Stringfellow, William T; Camarillo, Mary Kay; Domen, Jeremy K; Sandelin, Whitney L; Varadharajan, Charuleka; Jordan, Preston D; Reagan, Matthew T; Cooley, Heather; Heberger, Matthew G; Birkholzer, Jens T

2017-01-01

Chemical additives used for hydraulic fracturing and matrix acidizing of oil reservoirs were reviewed and priority chemicals of concern needing further environmental risk assessment, treatment demonstration, or evaluation of occupational hazards were identified. We evaluated chemical additives used for well stimulation in California, the third largest oil producing state in the USA, by the mass and frequency of use, as well as toxicity. The most frequently used chemical additives in oil development were gelling agents, cross-linkers, breakers, clay control agents, iron and scale control agents, corrosion inhibitors, biocides, and various impurities and product stabilizers used as part of commercial mixtures. Hydrochloric and hydrofluoric acids, used for matrix acidizing and other purposes, were reported infrequently. A large number and mass of solvents and surface active agents were used, including quaternary ammonia compounds (QACs) and nonionic surfactants. Acute toxicity was evaluated and many chemicals with low hazard to mammals were identified as potentially hazardous to aquatic environments. Based on an analysis of quantities used, toxicity, and lack of adequate hazard evaluation, QACs, biocides, and corrosion inhibitors were identified as priority chemicals of concern that deserve further investigation. Copyright © 2016 Elsevier Ltd. All rights reserved.

Impact of polymer modification on mechanical and viscoelastic properties.

DOT National Transportation Integrated Search

2015-10-01

This study was initiated with the aim of evaluating the relative impact of different cross-linking agents : on the rheological and morphological properties of polymer modified asphalt binders (PMAs). To : complete this objective, two cross-linking ag...

Induction and repair of DNA cross-links induced by sulfur mustard in the A-549 cell line followed by a comet assay.

PubMed

Jost, Petr; Svobodova, Hana; Stetina, Rudolf

2015-07-25

Sulfur mustard is a highly toxic chemical warfare agent with devastating impact on intoxicated tissues. DNA cross-links are probably the most toxic DNA lesions induced in the cell by sulfur mustard. The comet assay is a very sensitive method for measuring DNA damage. In the present study using the A-549 lung cell line, the comet assay protocol was optimized for indirect detection of DNA cross-links induced by sulfur mustard. The method is based on the additional treatment of the assayed cells containing cross-links with the chemical mutagen, styrene oxide. Alkali-labile adducts of styrene oxide cause DNA breaks leading to the formation of comets. A significant dose-dependent reduction of DNA migration of the comet's tail was found after exposing cells to sulfur mustard, indicative of the amount of sulfur mustard induced cross-links. The remarkable decrease of % tail DNA could be observed as early as 5min following exposure to sulfur mustard and the maximal effect was found after 30min, when DNA migration was reduced to the minimum. Sulfur mustard preincubated in culture medium without cells lost its ability to induce cross-links and had a half-life of about 15min. Pre-incubation longer than 30min does not lead to a significant increase in cross-links when applied to cells. However, the amount of cross-links is decreased during further incubation due to repair. The current modification of the comet assay provides a useful tool for detecting DNA cross-links induced by sulfur mustard and could be used for detection of other DNA cross-linking agents such as chemotherapeutic drugs. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Cross-linked enzyme aggregates of phenylalanine ammonia lyase: novel biocatalysts for synthesis of L-phenylalanine.

PubMed

Cui, Jian-Dong; Zhang, Si; Sun, Li-Mei

2012-06-01

Cross-linked enzyme aggregates of phenylalanine ammonia lyase (PAL-CLEAs) from Rhodotorula glutinis were prepared. The effects of the type of aggregating agent, its concentration, and that of cross-linking agent were studied. PAL-CLEAs production was most effective using ammonium sulfate (40 % saturation), followed by cross-linking for 1 h with 0.2 % (v/v) glutaraldehyde. Moreover, the storage and operational stability of the resulting PAL-CLEAs were also investigated. Compared to the free enzyme, the PAL-CLEAs exhibited the expected increased stability of the enzyme against various deactivating conditions such as pH, temperature, denaturants, and organic solvents and showed higher storage stability than its soluble counterpart. Additionally, the reusability of PAL-CLEAs with respect to the biotransformation of L-phenylalanine was evaluated. PAL-CLEAs could be recycled at least for 12 consecutive batch reactions without dramatic activity loss, which should dramatically increase the commercial potential of PAL for synthesis of L: -phenylalanine. To the best of our knowledge, this is the first report of immobilization of PAL as cross-linked enzyme aggregates.

Sulfolane-Cross-Polybenzimidazole Membrane For Gas Separation

DOEpatents

Young, Jennifer S.; Long, Gregory S.; Espinoza, Brent F.

2006-02-14

A cross-linked, supported polybenzimidazole membrane for gas separation is prepared by reacting polybenzimidazole (PBI) with the sulfone-containing crosslinking agent 3,4-dichloro-tetrahydro-thiophene-1,1-dioxide. The cross-linked reaction product exhibits enhanced gas permeability to hydrogen, carbon dioxide, nitrogen, and methane as compared to the unmodified analog, without significant loss of selectivity, at temperatures from about 20 degrees Celsius to about 400 degrees Celsius.

Reactive Oxygen Species (ROS) Inducible DNA Cross-Linking Agents and Their Effect on Cancer Cells and Normal Lymphocytes

PubMed Central

2015-01-01

Reducing host toxicity is one of the main challenges of cancer chemotherapy. Many tumor cells contain high levels of ROS that make them distinctively different from normal cells. We report a series of ROS-activated aromatic nitrogen mustards that selectively kill chronic lymphocytic leukemia (CLL) over normal lymphocytes. These agents showed powerful DNA cross-linking abilities when coupled with H2O2, one of the most common ROS in cancer cells, whereas little DNA cross-linking was detected without H2O2. Consistent with chemistry observation, in vitro cytotoxicity assay demonstrated that these agents induced 40–80% apoptosis in primary leukemic lymphocytes isolated from CLL patients but less than 25% cell death to normal lymphocytes from healthy donors. The IC50 for the most potent compound (2) was ∼5 μM in CLL cells, while the IC50 was not achieved in normal lymphocytes. Collectively, these data provide utility and selectivity of these agents that will inspire further and effective applications. PMID:24801734

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.

The rational design of recognitive polymeric networks for sensing applications

NASA Astrophysics Data System (ADS)

Noss, Kimberly Ryanne Dial

Testosterone recognitive networks were synthesized with varying feed crosslinking percentages and length of the bi-functional crosslinking agent to analyze the effect of changing structural parameters on template binding properties such as affinity, selectivity, capacity, and diffusional transport. The crosslinking percentage of the crosslinking monomer ethylene glycol dimethacrylate was varied from 50% to 90% and associated networks experienced a 2 fold increase in capacity and a 4 fold increase in affinity with the equilibrium association constants, Ka, ranging from 0.32 +/- 0.02 x 10 4 M-1 to 1.3 +/- 0.1 x 104 M -1, respectively. The higher concentration of crosslinking monomer increased the crosslinking points available for inter-chain stabilization creating an increased number of stable cavities for template association. However, by increasing the length of the crosslinking agent and increasing the feed crosslinking percentage from 77% crosslinked poly(methacrylic acid- co-ethylene glycol dimethacrylate) (poly(MAA-co-EGDMA)) to 50% crosslinked poly(methacrylic acid-co-poly(ethylene glycol)200 dimethacrylate) (poly(MAA-co-PEG200DMA)), the mesh size of the network increased resulting in an increased template diffusion coefficient from (2.83 +/- 0.06) x 109 cm2/s to (4.3 +/- 0.06) x 109 cm2/s, respectively, which is approximately a 40% faster template diffussional transport. A 77% crosslinked poly (MAA-co-PEG200DMA) recognitive network had an association constant of (0.20 +/- 0.05) x 104 M -1 and bound (0.72 +/- 0.04) x 10-2 mmol testosterone/g dry polymer, which was less by 6 and 3 fold, respectively, compared to a similarly crosslinked poly(MAA-co-EGDMA) recognitive network. Structural manipulation of the macromolecular architecture illustrates the programmability of recognitive networks for specific template binding parameters and diffusional transport, which may lead to enhanced imprinted sensor materials and successful integration onto sensor platforms.

Multi angle laser light scattering evaluation of field exposed thermoplastic photovoltaic encapsulant materials

DOE PAGES

Kempe, Michael D.; Miller, David C.; Wohlgemuth, John H.; ...

2016-01-08

As creep of polymeric materials is potentially a safety concern for photovoltaic modules, the potential for module creep has become a significant topic of discussion in the development of IEC 61730 and IEC 61215. To investigate the possibility of creep, modules were constructed, using several thermoplastic encapsulant materials, into thin-film mock modules and deployed in Mesa, Arizona. The materials examined included poly(ethylene)-co-vinyl acetate (EVA, including formulations both cross-linked and with no curing agent), polyethylene/polyoctene copolymer (PO), poly(dimethylsiloxane) (PDMS), polyvinyl butyral (PVB), and thermoplastic polyurethane (TPU). The absence of creep in this experiment is attributable to several factors of which themore » most notable one was the unexpected cross-linking of an EVA formulation without a cross-linking agent. It was also found that some materials experienced both chain scission and cross-linking reactions, sometimes with a significant dependence on location within a module. The TPU and EVA samples were found to degrade with cross-linking reactions dominating over chain scission. In contrast, the PO materials degraded with chain scission dominating over cross-linking reactions. Furthermore, we found no significant indications that viscous creep is likely to occur in fielded modules capable of passing the qualification tests, we note that one should consider how a polymer degrades, chain scission or cross-linking, in assessing the suitability of a thermoplastic polymer in terrestrial photovoltaic applications.« less

The Fanconi Anemia Pathway in Replication Stress and DNA Crosslink Repair

PubMed Central

Jones, Mathew JK.; Huang, Tony T.

2013-01-01

Interstand crosslinks (ICLs) are DNA lesions where the bases of opposing DNA strands are covalently linked, inhibiting critical cellular processes such as transcription and replication. Chemical agents that generate ICLs cause chromosomal abnormalities including breaks, deletions and rearrangements, making them highly genotoxic compounds. This toxicity has proven useful for chemotherapeutic treatment against a wide variety of cancer types. The majority of our understanding of ICL repair in humans has been uncovered thorough analysis of the rare genetic disorder Fanconi anemia, in which patients are extremely sensitive to crosslinking agents. Here, we discuss recent insights into ICL repair gained through new ICL repair assays and highlight the role of the Fanconi Anemia repair pathway during replication stress. PMID:22744751

Hydrothermal growth of cross-linked hyperbranched copper dendrites using copper oxalate complex

NASA Astrophysics Data System (ADS)

Truong, Quang Duc; Kakihana, Masato

2012-06-01

A facile and surfactant-free approach has been developed for the synthesis of cross-linked hyperbranched copper dendrites using copper oxalate complex as a precursor and oxalic acid as a reducing and structure-directing agent. The synthesized particles are composed of highly branched nanostructures with unusual cross-linked hierarchical networks. The formation of copper dendrites can be explained in view of both diffusion control and aggregation-based growth model accompanied by the chelation-assisted assembly. Oxalic acid was found to play dual roles as reducing and structure-directing agent based on the investigation results. The understanding on the crystal growth and the roles of oxalic acid provides clear insight into the formation mechanism of hyperbranched metal dendrites.

Effects of tetraamine crosslinking agents on the thermomechanical properties of PMR polyimide composites

NASA Technical Reports Server (NTRS)

Delvigs, P.

1976-01-01

The effects were investigated of partial substitution of tetraamine crosslinking agents for diamine reactants on the thermomechanical properties of PMR polyimide resins and graphite fiber-reinforced composites. The effect of tetraamine content on isothermal weight loss, glass transition, and softening temperatures of neat resin samples is discussed. Composites were fabricated using PMR methodology. Monomeric solution of various stoichiometric ratios was used to impregnate Hercules HTS graphite fiber. The mechanical property retention characteristics of the composites at 316 C (600 F) are described.

Study of Reactive Melt Processing Behavior of Externally Plasticized Cellulose Acetate in Presence of Isocyanate

PubMed Central

Erdmann, Rafael; Kabasci, Stephan; Kurek, Joanna; Zepnik, Stefan

2014-01-01

Two types of externally plasticized cellulose acetate (CA) were chemically modified using 4,4'-methylene diphenyl diisocyanate (MDI) as crosslinking agent. Crosslinking was performed in the molten state by means of melt mixing in an internal mixer. The viscoelastic properties of the non-crosslinked, externally plasticized CA show typical temperature dependence, similar to conventional thermoplastics. A strong increase in storage modulus is observed with increasing crosslink density indicating that the crosslinked compounds exhibit predominately elastic response. The complex viscosity also increases considerably with increasing crosslink density and does not reach the typical Newtonian plateau at low radial frequencies any more. The viscoelastic properties correlate well with the data recorded online during reactive melt processing in the internal mixer. In comparison to the non-crosslinked CA, the crosslinked compounds show higher glass transition temperature, higher VICAT softening temperatures, improved thermal stability and lower plasticizer evaporation at evaluated temperatures. PMID:28788273

Study of Reactive Melt Processing Behavior of Externally Plasticized Cellulose Acetate in Presence of Isocyanate.

PubMed

Erdmann, Rafael; Kabasci, Stephan; Kurek, Joanna; Zepnik, Stefan

2014-12-04

Two types of externally plasticized cellulose acetate (CA) were chemically modified using 4,4'-methylene diphenyl diisocyanate (MDI) as crosslinking agent. Crosslinking was performed in the molten state by means of melt mixing in an internal mixer. The viscoelastic properties of the non-crosslinked, externally plasticized CA show typical temperature dependence, similar to conventional thermoplastics. A strong increase in storage modulus is observed with increasing crosslink density indicating that the crosslinked compounds exhibit predominately elastic response. The complex viscosity also increases considerably with increasing crosslink density and does not reach the typical Newtonian plateau at low radial frequencies any more. The viscoelastic properties correlate well with the data recorded online during reactive melt processing in the internal mixer. In comparison to the non-crosslinked CA, the crosslinked compounds show higher glass transition temperature, higher VICAT softening temperatures, improved thermal stability and lower plasticizer evaporation at evaluated temperatures.

Characterization and thermal properties of polygenipin-crosslinked hide powders

USDA-ARS?s Scientific Manuscript database

Genipin is a naturally occurring iridoid compound, it is widely used as an ideal biological protein crosslinking agent due to its low toxicity compared to glutaraldehyde and formaldehyde. Under alkaline condition, genipin could undergo ring-opening polymerization via nucleophilic attack of hydroxyl ...

Modified in vivo comet assay detects the genotoxic potential of 14-hydroxycodeinone, an α,β-unsaturated ketone in oxycodone.

PubMed

Pant, Kamala; Roden, Nicholas; Zhang, Charles; Bruce, Shannon; Wood, Craig; Pendino, Kimberly

2015-12-01

14-Hydroxycodeinone (14-HC) is an α,β-unsaturated ketone impurity found in oxycodone drug substance and has a structural alert for genotoxicity. 14-HC was tested in a combined Modified and Standard Comet Assay to determine if the slight decrease in % Tail DNA noted in a previously conducted Standard Comet Assay with 14-HC could be magnified to clarify if the response was due to cross-linking activity. One limitation of the Standard Comet Assay is that DNA cross-links cannot be reliably detected. However, under certain modified testing conditions, DNA cross-links and chemical moieties that elicit such cross-links can be elucidated. One such modification involves the induction of additional breakages of DNA strands by gamma or X-ray irradiation. To determine if 14-HC is a DNA crosslinker in vivo, a Modified Comet Assay was conducted using X-ray irradiation as the modification to visualize crosslinking activity. In this assay, 14-HC was administered orally to mice up to 320 mg/kg/day. Results showed a statistically significant reduction in percent tail DNA in duodenal cells at 320 mg/kg/day, with a nonstatistically significant but dose-related reduction in percent tail DNA also observed at the mid dose of 160 mg/kg/day. Similar decreases were not observed in cells from the liver or stomach, and no increases in percent tail DNA were noted for any tissue in the concomitantly conducted Standard Comet Assay. Taken together, 14-HC was identified as a cross-linking agent in the duodenum in the Modified Comet Assay. © 2015 Wiley Periodicals, Inc.

Enhanced drug encapsulation and extended release profiles of calcium-alginate nanoparticles by using tannic acid as a bridging cross-linking agent.

PubMed

Abulateefeh, Samer R; Taha, Mutasem O

2015-01-01

Calcium alginate nanoparticles (NPs) suffer from sub-optimal stability in bio-relevant media leading to low drug encapsulation efficiency and uncontrolled release profiles. To sort out these drawbacks, a novel approach is proposed herein based on introducing tannic acid into these NPs to act as a bridging cross-linking aid agent. Calcium-alginate NPs were prepared by the ionotropic gelation method and loaded with diltiazem hydrochloride as a model drug. These NPs were characterized in terms of particle size, zeta potential, and morphology, and results were explained in accordance with Fourier-transform infrared (FTIR) spectroscopy and differential scanning calorimetry (DSC). The incorporation of tannic acid led to more than four folds increase in drug encapsulation efficiency (i.e. from 15.3% to 69.5%) and reduced burst drug release from 44% to around 10% within the first 30 min. These findings suggest the possibility of improving the properties of Ca-alginate NPs by incorporating cross-linking aid agents under mild conditions.

Aryl sulfonate based anticancer alkylating agents.

PubMed

Sheikh, Hamdullah Khadim; Arshad, Tanzila; Kanwal, Ghazala

2018-05-01

This research work revolves around synthesis of antineoplastic alkylating sulfonate esters with dual alkylating sites for crosslinking of the DNA strands. These molecules were evaluated as potential antineoplastic cross linking alkylating agents by reaction with the nucleoside of Guanine DNA nucleobase at both ends of the synthesized molecule. Synthesis of the alkylating molecules and the crosslinking with the guanosine nucleoside was monitored by MALDITOF mass spectroscopy. The synthesized molecule's crosslinking or adduct forming rate with the nucleoside was compared with that of 1,4 butane disulfonate (busulfan), in form of time taken for the appearance of [M+H] + . It was found that aryl sulfonate leaving group was causing higher rate of nucleophilic attack by the Lewis basic site of the nucleobase. Furthermore, the rate was also found to be a function of electron withdrawing or donating nature of the substituent on the aryl ring. Compound with strong electron withdrawing substituent on the para position of the ring reacted fastest. Hence, new alkylating agents were synthesized with optimized or desired reactivity.

Nano-hydrogels of methoxy polyethylene glycol-grafted branched polyethyleneimine via biodegradable cross-linking of Zn2+-ionomer micelle template

NASA Astrophysics Data System (ADS)

Abolmaali, Samira Sadat; Tamaddon, Ali Mohammad; Dinarvand, Rasoul

2013-12-01

Soft polymeric nanomaterials were synthesized by the template-assisted method involving self-association of methoxy polyethylene glycol- g-branched polyethyleneimine (mPEG- g-branched PEI) ionomer by transition metal ions such as Zn2+ followed by chemical cross-linking of the polyamine core by dithiopropionic acid. The formation of donor-acceptor complexes of Zn2+ and PEI ionomer was characterized by FT-IR spectroscopy and potentiometric titration. Turbidimetry was performed to study the solution property of the complexes which depended on pH, relative weight fraction of mPEG, and the molar ratio of Zn2+. The cross-linking reaction was studied by TNBS assay, 1H-NMR, and size exclusion chromatography. Upon removal of Zn2+ from cl-mPEG- g-branched PEI/Zn2+ at pH 3 by dialysis, the resulting cross-linked self-assembly represented a uniform, stable, and less positively charged hydrogel-like nanosphere with an intensity-averaged size ranging from 150 to 250 nm as determined by a Zetasizer. Atomic forced microscopy imaging was performed in intermittent contact mode in air that revealed discrete and oval-to-spherically shaped particles with average sizes ranging from 40 to 50 nm depending on the degree of cross-linking. This functional nanocarrier is expected to exhibit some key features such as active encapsulation of negatively charged hydrophilic agents in the swollen core of polyamine network and a hydrophilic mPEG shell which provides an increased solubility and passive targeting of active pharmaceutical agents to impaired tissues. The nano-hydrogels especially at 12 % degrees of cross-link demonstrated excellent biocompatibility determined by different experiments such as albumin aggregation, erythrocyte aggregation, hemolysis, and MTT cytotoxicity assay. Moreover, biodegradability of the cross-links as shown by the Ellman assay can offer a time-dependent degradation and redox-stimulated release of active agents.

1,3,5-Triazine-2,4,6-tribenzaldehyde derivative as a new crosslinking agent for synthesis of pH-thermo dual responsive chitosan hydrogels and their nanocomposites: Swelling properties and drug release behavior.

PubMed

Karimi, Ali Reza; Tarighatjoo, Mahsa; Nikravesh, Golara

2017-12-01

In this work, 1,3,5-triazine-2,4,6-tribenzaldehyde was synthesized and chosen as the cross-linking agent for preparation of novel thermo- and pH-responsive hydrogels based on chitosan. The cross-linking proceeds through formation of imine bond by reaction of amino groups of chitosan with aldehyde groups of the cross-linker. The various amounts (6, 10, 14% w/w) of the cross-linker were used with respect to chitosan to produce three 1,3,5-triazine-2,4,6-tribenzaldehyde cross-linked chitosans. Then, their hydrogel nanocomposites were prepared by crosslinking of chitosan with 1,3,5-triazine-2,4,6-tribenzaldehyde in the presence of 0.1% and 0.3% (w/w) multi-walled carbon nanotubes (MWCNTs). The structure and properties of the hydrogels and their nanocomposites were characterized by FT-IR, 1 H NMR and scanning electron microscopy (SEM). The swelling behavior of prepared hydrogels and their nanocomposites at different pHs and temperatures was investigated. The results showed that they exhibit a pH and temperature-responsive swelling ratio. The swelling behavior of the prepared chitosan hydrogels was strongly dependent on the amounts of cross-linker and MWCNTs. In vitro controlled release behavior of metronidazole model drug was studied with prepared hydrogels and nanocomposite hydrogels. The pH, temperature and wt% of MWCNTs were found to strongly influence the drug release behavior of the hydrogels. Copyright © 2017 Elsevier B.V. All rights reserved.

Poly(vinyl alcohol)-based film potentially suitable for antimicrobial packaging applications.

PubMed

Musetti, Alessandro; Paderni, Katia; Fabbri, Paola; Pulvirenti, Andrea; Al-Moghazy, Marwa; Fava, Patrizia

2014-04-01

This work aimed at developing a thin and water-resistant food-grade poly(vinyl alcohol) (PVOH)-based matrix able to swell when in contact with high moisture content food products without rupturing to release antimicrobial agents onto the food surface. This film was prepared by blending PVOH and 7.20% (wt/wt of PVOH) of poly(ethylene glycol) (PEG) with citric acid as crosslinking agent. The film-forming solution was then casted onto a flat surface and the obtained film was 60 μm in thickness and showed a good transparency (close to T = 100%) in the visible region (400 to 700 nm). After immersion in water for 72 h at room temperature, the crosslinked matrix loses only 19.2% of its original weight (the percentage includes the amount of unreacted crosslinking agent, antimicrobial in itself). Water content, degree of swelling, and crosslinking density of the film prove that the presence of PEG diminishes the hydrophilic behavior of the material. Also the mechanical properties of the wet and dry film were assessed. Alongside this, 2.5% (wt/wt of dry film) of grapefruit seed extract (GSE), an antimicrobial agent, was added to the film-forming solution just before casting and the ability of the plastic matrix to release the additive was then evaluated in vitro against 2 GSE-susceptible microorganisms, Salmonella enteritidis and Listeria innocua. The results indicate that the developed matrix may be a promising food-grade material for the incorporation of active substances. © 2014 Institute of Food Technologists®

In vivo therapeutic responses contingent on Fanconi anemia/BRCA2 status of the tumor.

PubMed

van der Heijden, Michiel S; Brody, Jonathan R; Dezentje, David A; Gallmeier, Eike; Cunningham, Steven C; Swartz, Michael J; DeMarzo, Angelo M; Offerhaus, G Johan A; Isacoff, William H; Hruban, Ralph H; Kern, Scott E

2005-10-15

BRCA2, FANCC, and FANCG gene mutations are present in a subset of pancreatic cancer. Defects in these genes could lead to hypersensitivity to interstrand cross-linkers in vivo and a more optimal treatment of pancreatic cancer patients based on the genetic profile of the tumor. Two retrovirally complemented pancreatic cancer cell lines having defects in the Fanconi anemia pathway, PL11 (FANCC-mutated) and Hs766T (FANCG-mutated), as well as several parental pancreatic cancer cell lines with or without mutations in the Fanconi anemia/BRCA2 pathway, were assayed for in vitro and in vivo sensitivities to various chemotherapeutic agents. A distinct dichotomy of drug responses was observed. Fanconi anemia-defective cancer cells were hypersensitive to the cross-linking agents mitomycin C (MMC), cisplatin, chlorambucil, and melphalan but not to 5-fluorouracil, gemcitabine, doxorubicin, etoposide, vinblastine, or paclitaxel. Hypersensitivity to cross-linking agents was confirmed in vivo; FANCC-deficient xenografts of PL11 and BRCA2-deficient xenografts of CAPAN1 regressed on treatment with two different regimens of MMC whereas Fanconi anemia-proficient xenografts did not. The MMC response comprised cell cycle arrest, apoptosis, and necrosis. Xenografts of PL11 also regressed after a single dose of cyclophosphamide whereas xenografts of genetically complemented PL11(FANCC) did not. MMC or other cross-linking agents as a clinical therapy for pancreatic cancer patients with tumors harboring defects in the Fanconi anemia/BRCA2 pathway should be specifically investigated.

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

NASA Astrophysics Data System (ADS)

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

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

Data of thermal degradation and dynamic mechanical properties of starch–glycerol based films with citric acid as crosslinking agent

PubMed Central

González Seligra, Paula; Medina Jaramillo, Carolina; Famá, Lucía; Goyanes, Silvia

2016-01-01

Interest in biodegradable edible films as packaging or coating has increased because their beneficial effects on foods. In particular, food products are highly dependents on thermal stability, integrity and transition process temperatures of the packaging. The present work describes a complete data of the thermal degradation and dynamic mechanical properties of starch–glycerol based films with citric acid (CA) as crosslinking agent described in the article titled: “Biodegradable and non-retrogradable eco-films based on starch–glycerol with citric acid as crosslinking agent” González Seligra et al. (2016) [1]. Data describes thermogravimetric and dynamical mechanical experiences and provides the figures of weight loss and loss tangent of the films as a function of the temperature. PMID:27158645

Potent antitumor bifunctional DNA alkylating agents, synthesis and biological activities of 3a-aza-cyclopenta[a]indenes.

PubMed

Kakadiya, Rajesh; Dong, Huajin; Lee, Pei-Chih; Kapuriya, Naval; Zhang, Xiuguo; Chou, Ting-Chao; Lee, Te-Chang; Kapuriya, Kalpana; Shah, Anamik; Su, Tsann-Long

2009-08-01

A series of bifunctional DNA interstrand cross-linking agents, bis(hydroxymethyl)- and bis(carbamates)-8H-3a-azacyclopenta[a]indene-1-yl derivatives were synthesized for antitumor evaluation. The preliminary antitumor studies revealed that these agents exhibited potent cytotoxicity in vitro and antitumor therapeutic efficacy against human tumor xenografts in vivo. Furthermore, these derivatives have little or no cross-resistance to either Taxol or Vinblastine. Remarkably, complete tumor remission in nude mice bearing human breast carcinoma MX-1 xenograft by 13a,b and 14g,h and significant suppression against prostate adenocarcinoma PC3 xenograft by 13b were achieved at the maximum tolerable dose with relatively low toxicity. In addition, these agents induce DNA interstrand cross-linking and substantial G2/M phase arrest in human non-small lung carcinoma H1299 cells. The current studies suggested that these agents are promising candidates for preclinical studies.

Inability of keratinocytes lacking their specific transglutaminase to form cross-linked envelopes: absence of envelopes as a simple diagnostic test for lamellar ichthyosis.

PubMed

Jeon, S; Djian, P; Green, H

1998-01-20

Epidermal keratinocytes, late in their terminal differentiation, form cross-linked envelopes resistant to ionic detergent and reducing agent. Because the cross-linking process is catalyzed by the keratinocyte transglutaminase, the absence of active transglutaminase should result in failure of the keratinocyte to form a cross-linked envelope. Three keratinocyte strains bearing mutations in the keratinocyte transglutaminase were examined: two contained no detectable transglutaminase mRNA and none contained active enzyme. All three were unable to form cross-linked envelopes, either spontaneously in stratified cultures or upon induction with Ca2+. Although stratum corneum of normal humans and scales from patients with different ichthyotic diseases contain cross-linked envelopes, those from patients with transglutaminase-negative lamellar ichthyosis do not. Therefore, the disease due to the absence of transglutaminase may be readily distinguished from other ichthyotic disease by a simple test for cross-linked envelopes.

Photocontrolled Cargo Release from Dual Cross-Linked Polymer Particles.

PubMed

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

2016-03-09

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

γ-H2AX formation in response to interstrand crosslinks requires XPF in human cells

PubMed Central

Mogi, Seiki; Oh, Dennis H.

2009-01-01

To further define the molecular mechanisms involved in processing interstrand crosslinks, we monitored the formation of phosphorylated histone H2AX (γ-H2AX), which is generated in chromatin near double strand break sites, following DNA damage in normal and repair-deficient human cells. Following treatment with a psoralen derivative and ultraviolet A radiation doses that produce significant numbers of crosslinks, γ-H2AX levels in nucleotide excision repair-deficient XP-A fibroblasts (XP12RO-SV) increased to levels that were twice those observed in normal control GM637 fibroblasts. A partial XPA revertant cell line (XP129) that is proficient in crosslink removal, exhibited reduced γ-H2AX levels that were intermediate between those of GM637 and XP-A cells. XP-F fibroblasts (XP2YO-SV and XP3YO) that are also repair-deficient exhibited γ-H2AX levels below even control fibroblasts following treatment with psoralen and ultraviolet A radiation. Similarly, another crosslinking agent, mitomycin C, did not induce γ-H2AX in XP-F cells, although it did induce equivalent levels of γ-H2AX in XPA and control GM637 cells. Ectopic expression of XPF in XP-F fibroblasts restored γ-H2AX induction following treatment with crosslinking agents. Angelicin, a furocoumarin which forms only monoadducts and not crosslinks following ultraviolet A radiation, as well as ultraviolet C radiation, resulted only in weak induction of γ-H2AX in all cells, suggesting that the double strand breaks observed with psoralen and ultraviolet A treatment result preferentially following crosslink formation. These results indicate that XPF is required to form γ-H2AX and likely double strand breaks in response to interstrand crosslinks in human cells. Furthermore, XPA may be important to allow psoralen interstrand crosslinks to be processed without forming a double strand break intermediate. PMID:16678501

Hypoxia-Activated Alkylating Agents in BRCA1-Mutant Ovarian Serous Carcinoma.

PubMed

Conroy, Michael; Borad, Mitesh J; Bryce, Alan H

2017-07-26

Breast cancer 1 antigen (BRCA 1) and breast cancer 2 antigen (BRCA2) genes play a significant role in deoxyribonucleic acid (DNA) repair by means of interstrand crosslink repair, and deleterious germline mutations of these are responsible for most hereditary breast and ovarian cancers. Therapeutic strategies which specifically target interstrand crosslink repair can therefore be helpful in patients with harmful mutations. We describe two patients with advanced ovarian cancer and deleterious BRCA1 mutations who were treated with TH-302, a hypoxia-activated alkylating agent.

Synthesis and Characterization of Ionically Crosslinked Elastomers

DTIC Science & Technology

2016-01-01

ionomer with a anionic monomer, and crosslinking by copolymerization with a  hydrophobic  monomer.  While these materials displayed signatures of...Therefore, these RAFT agents should have utility in the  synthesis of  super ‐soft and/or higher damping elastomers.  2. An ionic, divinyl crosslinking

Novel in-situ nanocomposites of phthalonitrile end-capped polyarylene ether nitrile/copper phthalocyanine

NASA Astrophysics Data System (ADS)

Tong, Lifen; Wei, Renbo; Liu, Xiaobo

2017-01-01

A novel phthalonitrile end-capped polyarylene ether nitrile (PEN-Ph)/copper phthalocyanine (CuPc) nanocomposites which possesses crosslinking reaction combined with crystallization behaviour were prepared successfully through in-situ reaction and hot-compression. In the presence of copper ion, CuPc were formed through crosslinking reaction among the phthalonitrile at the end of the PEN-Ph main chain and 1, 3, 5-Tri-(3, 4-dicyanophenoxy) benzene (TPh). Besides, the formed CuPc can play the role of nucleating agent to improve the crystallinity of the polymers. The influence of the crosslinking reaction and crystallization behaviour were investigated. The results show that the crystallization and crosslinking coexist in the system at the same time. Scanning electron microscope (SEM) images show that the crystals of the PEN-Ph grow after the hot-compressing procedure. Moreover, the glass transition temperature (Tg) increases while the crystallinity declines slightly with the low amount of copper ions. The increase of Tg is mainly caused by the crosslinking reaction, indicating that the copper can be used as a crosslinking agent in this system. Due to formation of the CuPc and the crystallization behaviour, the dielectric constant increased as expected from 3.2 to 4.9 while the dielectric loss decreased. Therefore, the PEN-Ph/CuPc in-situ nanocomposites will have a good prospect for application in electronic field.

Preparation of Thermo-Responsive and Cross-Linked Fluorinated Nanoparticles via RAFT-Mediated Aqueous Polymerization in Nanoreactors.

PubMed

Ma, Jiachen; Zhang, Luqing; Geng, Bing; Azhar, Umair; Xu, Anhou; Zhang, Shuxiang

2017-01-25

In this work, a thermo-responsive and cross-linked fluoropolymer poly(2,2,2-Trifluoroethyl) methacrylate (PTFEMA) was successfully prepared by reversible addition-fragmentation chain transfer (RAFT) mediated aqueous polymerization with a thermo-responsive diblock poly(dimethylacrylamide- b - N -isopropylacrylamide) (PDMA- b -PNIPAM) that performed a dual function as both a nanoreactor and macro-RAFT agent. The cross-linked polymer particles proved to be in a spherical-like structure of about 50 nm in diameter and with a relatively narrow particle size distribution. ¹H-NMR and 19 F-NMR spectra showed that thermo-responsive diblock P(DMA- b -NIPAM) and cross-linked PTFEMA particles were successfully synthesized. Influence of the amount of ammonium persulfate (APS), the molar ratio of monomers to RAFT agent, influence of the amount of cross-linker on aqueous polymerization and thermo-responsive characterization of the particles are investigated. Monomer conversion increased from 44% to 94% with increasing the molar ratio of APS and P(DMA- b -NIPAM) from 1:9 to1:3. As the reaction proceeded, the particle size increased from 29 to 49 nm due to the consumption of TFEMA monomer. The size of cross-linked nanoparticles sharply decreased from 50.3 to 40.5 nm over the temperature range 14-44 °C, suggesting good temperature sensitivity for these nanoparticles.

Crosslinking Protein Glutathionylation Mediated by O2-Arylated Bis-Diazeniumdiolate “Double JS-K”

PubMed Central

Holland, Ryan J.; Maciag, Anna E.; Kumar, Varun; Shi, Lei; Saavedra, Joseph E.; Prud’homme, Robert K.; Chakrapani, Harinath; Keefer, Larry K.

2012-01-01

Attachment of glutathione (GSH) to cysteine residues in proteins (S-glutathionylation) is a reversible post-translational modification that can profoundly alter protein structure and function. Often serving in a protective role, e.g., by temporarily saving protein thiols from irreversible oxidation and inactivation, glutathionylation can be identified and semi-quantitatively assessed using anti-GSH antibodies, thought to be specific for recognition of the S-glutathionylation modification. Here we describe an alternate mechanism of protein glutathionylation in which the sulfur atoms of the GSH and the protein’s thiol group are covalently bound via a crosslinking agent, rather than through a disulfide bond. This form of thiol crosslinking has been shown to occur and confirmed by mass spectrometry at the solution chemistry level, as well as in experiments documenting the potent antiproliferative activity of the bis-diazeniumdiolate Double JS-K in H1703 cells in vitro and in vivo. The modification is recognized by the anti-GSH antibody as if it were authentic S-glutathionylation, requiring mass spectrometry to distinguish between them. PMID:23106594

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.

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

PubMed

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

2010-03-08

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

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

PubMed

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

1987-08-15

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

Resistance of uveal melanoma to the interstrand cross-linking agent mitomycin C is associated with reduced expression of CYP450R

PubMed Central

Gravells, P; Hoh, L; Canovas, D; Rennie, I G; Sisley, K; Bryant, H E

2011-01-01

Background: Uveal melanoma (UM) is the most common primary intraocular tumour of adults, frequently metastasising to the liver. Hepatic metastases are difficult to treat and are mainly unresponsive to chemotherapy. To investigate why UM are so chemo-resistant we explored the effect of interstrand cross-linking agents mitomycin C (MMC) and cisplatin in comparison with hydroxyurea (HU). Methods: Sensitivity to MMC, cisplatin and HU was tested in established UM cell lines using clonogenic assays. The response of UM to MMC was confirmed in MTT assays using short-term cultures of primary UM. The expression of cytochrome P450 reductase (CYP450R) was analysed by western blotting, and DNA cross-linking was assessed using COMET analysis supported by γ-H2AX foci formation. Results: Both established cell lines and primary cultures of UM were resistant to the cross-linking agent MMC (in each case P<0.001 in Student's t-test compared with controls). In two established UM cell lines, DNA cross-link damage was not induced by MMC (in both cases P<0.05 in Students's t-test compared with damage induced in controls). In all, 6 out of 6 UMs tested displayed reduced expression of the metabolising enzyme CYP450R and transient expression of CYP450R increased MMC sensitivity of UM. Conclusion: We suggest that reduced expression of CYP450R is responsible for MMC resistance of UM, through a lack of bioactivation, which can be reversed by complementing UM cell lines with CYP450R. PMID:21386838

Nucleotide excision repair is a potential therapeutic target in multiple myeloma

PubMed Central

Szalat, R; Samur, M K; Fulciniti, M; Lopez, M; Nanjappa, P; Cleynen, A; Wen, K; Kumar, S; Perini, T; Calkins, A S; Reznichenko, E; Chauhan, D; Tai, Y-T; Shammas, M A; Anderson, K C; Fermand, J-P; Arnulf, B; Avet-Loiseau, H; Lazaro, J-B; Munshi, N C

2018-01-01

Despite the development of novel drugs, alkylating agents remain an important component of therapy in multiple myeloma (MM). DNA repair processes contribute towards sensitivity to alkylating agents and therefore we here evaluate the role of nucleotide excision repair (NER), which is involved in the removal of bulky adducts and DNA crosslinks in MM. We first evaluated NER activity using a novel functional assay and observed a heterogeneous NER efficiency in MM cell lines and patient samples. Using next-generation sequencing data, we identified that expression of the canonical NER gene, excision repair cross-complementation group 3 (ERCC3), significantly impacted the outcome in newly diagnosed MM patients treated with alkylating agents. Next, using small RNA interference, stable knockdown and overexpression, and small-molecule inhibitors targeting xeroderma pigmentosum complementation group B (XPB), the DNA helicase encoded by ERCC3, we demonstrate that NER inhibition significantly increases sensitivity and overcomes resistance to alkylating agents in MM. Moreover, inhibiting XPB leads to the dual inhibition of NER and transcription and is particularly efficient in myeloma cells. Altogether, we show that NER impacts alkylating agents sensitivity in myeloma cells and identify ERCC3 as a potential therapeutic target in MM. PMID:28588253

Nanocomposite scaffolds with tunable mechanical and degradation capabilities: co-delivery of bioactive agents for bone tissue engineering.

PubMed

Cattalini, Juan P; Roether, Judith; Hoppe, Alexander; Pishbin, Fatemeh; Haro Durand, Luis; Gorustovich, Alejandro; Boccaccini, Aldo R; Lucangioli, Silvia; Mouriño, Viviana

2016-10-21

Novel multifunctional nanocomposite scaffolds made of nanobioactive glass and alginate crosslinked with therapeutic ions such as calcium and copper were developed for delivering therapeutic agents, in a highly controlled and sustainable manner, for bone tissue engineering. Alendronate, a well-known antiresorptive agent, was formulated into microspheres under optimized conditions and effectively loaded within the novel multifunctional scaffolds with a high encapsulation percentage. The size of the cation used for the alginate crosslinking impacted directly on porosity and viscoelastic properties, and thus, on the degradation rate and the release profile of copper, calcium and alendronate. According to this, even though highly porous structures were created with suitable pore sizes for cell ingrowth and vascularization in both cases, copper-crosslinked scaffolds showed higher values of porosity, elastic modulus, degradation rate and the amount of copper and alendronate released, when compared with calcium-crosslinked scaffolds. In addition, in all cases, the scaffolds showed bioactivity and mechanical properties close to the endogenous trabecular bone tissue in terms of viscoelasticity. Furthermore, the scaffolds showed osteogenic and angiogenic properties on bone and endothelial cells, respectively, and the extracts of the biomaterials used promoted the formation of blood vessels in an ex vivo model. These new bioactive nanocomposite scaffolds represent an exciting new class of therapeutic cell delivery carrier with tunable mechanical and degradation properties; potentially useful in the controlled and sustainable delivery of therapeutic agents with active roles in bone formation and angiogenesis, as well as in the support of cell proliferation and osteogenesis for bone tissue engineering.

Influence of Glutathione and Glutathione S-transferases on DNA Interstrand Cross-Link Formation by 1,2-Bis(methylsulfonyl)-1-(2-chloroethyl)hydrazine, the Active Anticancer Moiety Generated by Laromustine

PubMed Central

2015-01-01

Prodrugs of 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)hydrazine (90CE) are promising anticancer agents. The 90CE moiety is a readily latentiated, short-lived (t1/2 ∼ 30 s) chloroethylating agent that can generate high yields of oxophilic electrophiles responsible for the chloroethylation of the O-6 position of guanine in DNA. These guanine O-6 alkylations are believed to be responsible for the therapeutic effects of 90CE and its prodrugs. Thus, 90CE demonstrates high selectivity toward tumors with diminished levels of O6-alkylguanine-DNA alkyltransferase (MGMT), the resistance protein responsible for O6-alkylguanine repair. The formation of O6-(2-chloroethyl)guanine lesions ultimately leads to the generation of highly cytotoxic 1-(N3-cytosinyl),-2-(N1-guaninyl)ethane DNA interstrand cross-links via N1,O6-ethanoguanine intermediates. The anticancer activity arising from this sequence of reactions is thus identical to this component of the anticancer activity of the clinically used chloroethylnitrosoureas. Herein, we evaluate the ability of glutathione (GSH) and other low molecular weight thiols, as well as GSH coupled with various glutathione S-transferase enzymes (GSTs) to attenuate the final yields of cross-links generated by 90CE when added prior to or immediately following the initial chloroethylation step to determine the major point(s) of interaction. In contrast to studies utilizing BCNU as a chloroethylating agent by others, GSH (or GSH/GST) did not appreciably quench DNA interstrand cross-link precursors. While thiols alone offered little protection at either alkylation step, the GSH/GST couple was able to diminish the initial yields of cross-link precursors. 90CE exhibited a very different GST isoenzyme susceptibility to that reported for BCNU, this could have important implications in the relative resistance of tumor cells to these agents. The protection afforded by GSH/GST was compared to that produced by MGMT. PMID:25012050

Influence of glutathione and glutathione S-transferases on DNA interstrand cross-link formation by 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)hydrazine, the active anticancer moiety generated by laromustine.

PubMed

Penketh, Philip G; Patridge, Eric; Shyam, Krishnamurthy; Baumann, Raymond P; Zhu, Rui; Ishiguro, Kimiko; Sartorelli, Alan C

2014-08-18

Prodrugs of 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)hydrazine (90CE) are promising anticancer agents. The 90CE moiety is a readily latentiated, short-lived (t1/2 ∼ 30 s) chloroethylating agent that can generate high yields of oxophilic electrophiles responsible for the chloroethylation of the O-6 position of guanine in DNA. These guanine O-6 alkylations are believed to be responsible for the therapeutic effects of 90CE and its prodrugs. Thus, 90CE demonstrates high selectivity toward tumors with diminished levels of O(6)-alkylguanine-DNA alkyltransferase (MGMT), the resistance protein responsible for O(6)-alkylguanine repair. The formation of O(6)-(2-chloroethyl)guanine lesions ultimately leads to the generation of highly cytotoxic 1-(N(3)-cytosinyl),-2-(N(1)-guaninyl)ethane DNA interstrand cross-links via N(1),O(6)-ethanoguanine intermediates. The anticancer activity arising from this sequence of reactions is thus identical to this component of the anticancer activity of the clinically used chloroethylnitrosoureas. Herein, we evaluate the ability of glutathione (GSH) and other low molecular weight thiols, as well as GSH coupled with various glutathione S-transferase enzymes (GSTs) to attenuate the final yields of cross-links generated by 90CE when added prior to or immediately following the initial chloroethylation step to determine the major point(s) of interaction. In contrast to studies utilizing BCNU as a chloroethylating agent by others, GSH (or GSH/GST) did not appreciably quench DNA interstrand cross-link precursors. While thiols alone offered little protection at either alkylation step, the GSH/GST couple was able to diminish the initial yields of cross-link precursors. 90CE exhibited a very different GST isoenzyme susceptibility to that reported for BCNU, this could have important implications in the relative resistance of tumor cells to these agents. The protection afforded by GSH/GST was compared to that produced by MGMT.

Crosslinked, porous, polyacrylate beads

NASA Technical Reports Server (NTRS)

Rembaum, Alan (Inventor); Yen, Shiao-Ping Siao (Inventor); Dreyer, William J. (Inventor)

1976-01-01

Uniformly-shaped, porous, round beads are prepared by the co-polymerization of an acrylic monomer and a cross-linking agent in the presence of 0.05 to 5% by weight of an aqueous soluble polymer such as polyethylene oxide. Cross-linking proceeds at high temperature above about 50.degree.C or at a lower temperature with irradiation. Beads of even shape and even size distribution of less than 2 micron diameter are formed. The beads will find use as adsorbents in chromatography and as markers for studies of cell surface receptors.

Crosslinked, porous, polyacrylate beads

NASA Technical Reports Server (NTRS)

Rembaum, Alan (Inventor); Yen, Shiao-Ping S. (Inventor); Dreyer, William J. (Inventor)

1977-01-01

Uniformly-shaped, porous, round beads are prepared by the co-polymerization of an acrylic monomer and a cross-linking agent in the presence of 0.05 to 5% by weight of an aqueous soluble polymer such as polyethylene oxide. Cross-linking proceeds at high temperature above about 50.degree. C or at a lower temperature with irradiation. Beads of even shape and even size distribution of less than 2 micron diameter are formed. The beads will find use as adsorbents in chromatography and as markers for studies of cell surface receptors.

In situ self cross-linking of polyvinyl alcohol battery separators

NASA Technical Reports Server (NTRS)

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

1979-01-01

A battery separator was produced from a polyvinyl alcohol sheet structure which was subjected to an in situ, self crosslinking process by selective oxidation of the 1,2 diol units present in the polyvinyl alcohol sheet structure. The 1,2 diol units were cleaved to form aldehyde end groups which subsequently crosslink through acetalization of the 1,3 diol units of the polyvinyl alcohol. Selective oxidation was achieved using a solution of a suitable oxidizing agent such as periodic acid or lead tetraacetate.

The Use of Polymerized Genipin for the Stabilization of the Collagen Structure of Animal Hides

USDA-ARS?s Scientific Manuscript database

Animal hides are the major byproduct of meat industry and the collagen fibers is the main constituent. Crosslinkers play a key role in stabilizing the collagen structure for useful applications. Genipin is widely used as an ideal biological protein crosslinking agent due to its low toxicity compare...

Process for Preparing Microcapsules Having Gelatin Walls Crosslinked with Quinone.

DTIC Science & Technology

A process for conveniently producing microcapsules containing a gelatin wall crosslinked with quinone and a core of an active compound such as a...provides microcapsules of excellent strength, storage stability, and resistance to aqueous exposure, such that the rate of release of the fouling reducing agent can be controlled with precision. jg

Consequences of placing an intramolecular crosslink in myosin S1

PubMed Central

Konno, Kunihiko; Ue, Kathleen; Khoroshev, Mikhail; Martinez, Hugo; Ray, Bruce; Morales, Manuel F.

2000-01-01

This paper describes the placement of a crosslinking agent (dibromobimane) between two thiols (Cys-522 and Cys-707) of a fragment, “S1,” of the motor protein, myosin. It turns out that fastening the first anchor of the crosslinker is easy and rapid, but fastening the second anchor (Cys-522) is very temperature dependent, taking 30 min at room temperature but about a week on ice. Moreover, crystallography taken at 4°C would seem to predict that the linkage is impossible, because the span of the crosslinking agent is much less than the interthiol distance. The simplest resolution of this seeming paradox is that structural fluctuations of the protein render the linkage increasingly likely as the temperature increases. Also, measurements of the affinity of MgADP for the protein, as well as the magnetic resonance of the P-atoms of the ADP once emplaced, suggest that binding the first reagent anchor to Cys-707 initiates an influence that travels to the rather distant ADP-binding site, and it is speculated what this “path of influence” might be. PMID:10677484

Click chemistry improved wet adhesion strength of mussel-inspired citrate-based antimicrobial bioadhesives

PubMed Central

Guo, Jinshan; Kim, Gloria B.; Shan, Dingying; Kim, Jimin P.; Hu, Jianqing; Wang, Wei; Hamad, Fawzi G.; Qian, Guoying; Rizk, Elias B.; Yang, Jian

2016-01-01

For the first time, a convenient copper-catalyzed azide-alkyne cycloaddition (CuAAC, click chemistry) was successfully introduced into injectable citrate-based mussel-inspired bioadhesives (iCMBAs, iCs) to improve both cohesive and wet adhesive strengths and elongate the degradation time, providing numerous advantages in surgical applications. The major challenge to developing such an adhesive was the mutual inhibition effect between the oxidant used for crosslinking catechol groups and the Cu(II) reductant used for CuAAC, which was successfully minimized by adding a biocompatible buffering agent typically used in cell culture, 4-(2-hydroxyethyl) -1-piperazineethanesulfonic acid (HEPES), as a copper chelating agent. Among the investigated formulations, the highest adhesion strength achieved (223.11 ± 15.94 kPa) was around 13 times higher than that of a commercially available fibrin glue (15.4 ± 2.8 kPa). In addition, dual-crosslinked (i.e. click crosslinking and mussel-inspired crosslinking) iCMBAs still preserved considerable antibacterial and antifungal capabilities that are beneficial for the bioadhesives used as hemostatic adhesives or sealants for wound management. PMID:27770631

Optical and mechanical behaviors of glassy silicone networks derived from linear siloxane precursors

NASA Astrophysics Data System (ADS)

Jang, Heejun; Seo, Wooram; Kim, Hyungsun; Lee, Yoonjoo; Kim, Younghee

2016-01-01

Silicon-based inorganic polymers are promising materials as matrix materials for glass fiber composites because of their good process ability, transparency, and thermal property. In this study, for utilization as a matrix precursor for a glass-fiber-reinforced composite, glassy silicone networks were prepared via hydrosilylation of linear/pendant Si-H polysiloxanes and the C=C bonds of viny-lterminated linear/cyclic polysiloxanes. 13C nuclear magnetic resonance spectroscopy was used to determine the structure of the cross-linked states, and a thermal analysis was performed. To assess the mechanical properties of the glassy silicone networks, we performed nanoindentation and 4-point bending tests. Cross-linked networks derived from siloxane polymers are thermally and optically more stable at high temperatures. Different cross-linking agents led to final networks with different properties due to differences in the molecular weights and structures. After stepped postcuring, the Young's modulus and the hardness of the glassy silicone networks increased; however, the brittleness also increased. The characteristics of the cross-linking agent played an important role in the functional glassy silicone networks.

Improving Properties of Arrowroot Starch (Maranta arundinacea)/PVA Blend Films by Using Citric Acid as Cross-linking Agent

NASA Astrophysics Data System (ADS)

Sholichah, Enny; Purwono, Bambang; Nugroho, Pramono

2017-12-01

This research studied the effect of PVA as organic polymer and citric acid as crosslinker agent in the arrowroot starch/PVA blend films. The properties of films were investigated by water uptake, water vapor permeability, mechanical properties, thermal stability, spectra of FTIR and XRD patterns. PVA used in this research influenced the film properties at the highest concentration. The cross-linkingsinter or intra molecules of arrowroot and PVA were developed as ester bonds which are formed from the reaction of hydroxyl groups consisting of starch and PVA with citric acid. The ester bond was confirmed by FTIR spectra. The increase of the amount of citric acid affected significantly on physical, chemical and mechanical properties, water uptake, WVP and crystallinity. Water barrier level was reduced by decreasing of water uptake and WVP succeeded significantly with increased crosslinking. Cross-linking impact the thermal stability of the films. The elasticity of the films also increases the production of citric acid as a plasticizer in the making of the films as a food packaging material.

Macromolecular crowding-assisted fabrication of liquid-crystalline imprinted polymers.

PubMed

Zhang, Chen; Zhang, Jing; Huang, Yan-Ping; Liu, Zhao-Sheng

2015-04-01

A macromolecular crowding-assisted liquid-crystalline molecularly imprinted monolith (LC-MIM) was prepared successfully for the first time. The imprinted stationary phase was synthesized with polymethyl methacrylate (PMMA) or polystyrene (PS) as the crowding agent, 4-cyanophenyl dicyclohexyl propylene (CPCE) as the liquid-crystal monomer, and hydroquinidine as the pseudo-template for the chiral separation of cinchona alkaloids in HPLC. A low level of cross-linker (26%) has been found to be sufficient to achieve molecular recognition on the crowding-assisted LC-MIM due to the physical cross-linking of mesogenic groups in place of chemical cross-linking, and baseline separation of quinidine and quinine could be achieved with good resolution (R(s) = 2.96), selectivity factor (α = 2.16), and column efficiency (N = 2650 plates/m). In contrast, the LC-MIM prepared without crowding agents displayed the smallest diastereoselectivity (α = 1.90), while the crowding-assisted MIM with high level of cross-linker (80%) obtained the greatest selectivity factor (α = 7.65), but the lowest column efficiency (N = 177 plates/m).

Genipin crosslinker releasing sutures for improving the mechanical/repair strength of damaged connective tissue.

PubMed

Sundararaj, Sharath; Slusarewicz, Paul; Brown, Matt; Hedman, Thomas

2017-11-01

The most common mode of surgical repair of ruptured tendons and ligaments involves the use of sutures for reattachment. However, there is a high incidence of rerupture and repair failure due to pulling out of the suture material from the damaged connective tissue. The main goal of this research was to achieve a localized delivery of crosslinking agent genipin (GP) from rapid-release biodegradable coatings on sutures, for strengthening the repair of ruptured connective tissue. Our hypothesis is that GP released from the suture coating will lead to exogenous crosslinking of native connective tissue resulting in beneficial effects on clinically relevant mechanical parameters such as tear resistance, tissue strength, and energy required to rupture the tissue (toughness). Sutures were successfully coated with a biodegradable polymer layer loaded with the crosslinking agent genipin, without compromising the mechanical properties of the suture. The rapid-release of genipin was achieved under both in vitro and ex vivo conditions. Exogenous crosslinking using these genipin releasing sutures was demonstrated using equine tendons. The tendons treated with genipin releasing sutures showed significant improvement in failure load, energy required for pull-out failure, and stiffness. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 2199-2205, 2017. © 2016 Wiley Periodicals, Inc.

Photoinduced DNA Interstrand Cross-Link Formation by Naphthalene Boronates via a Carbocation.

PubMed

Wang, Yibin; Lin, Zechao; Fan, Heli; Peng, Xiaohua

2016-07-18

Most photoinduced DNA cross-link formation by a bifunctional aryl derivative is through a bisquinone methide. DNA cross-linking via a bisarylcarbocation remains a less explored area. We designed and synthesized a series of naphthalene boronates that produce DNA interstrand cross-links via a carbocation upon UV irradiation. A free radical was generated from the naphthalene boronates with 350 nm irradiation and further converted to a carbocation by electron transfer. The activation mechanism was determined using the orthogonal traps, 2,2,6,6-tetramethylpiperidin-1-oxyl (TEMPO) and methoxyamine that react with either the free radical or the carbocation but not both. This represents a novel example of photoinduced DNA cross-link formation via carbocations generated from a bisaryl derivative. This work provides information useful for the design of novel photoactivated DNA cross-linking agents. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Modulation of biomechanical properties of hyaluronic acid hydrogels by crosslinking agents.

PubMed

Choi, Sung Chul; Yoo, Mi Ae; Lee, Su Yeon; Lee, Hyun Ji; Son, Dong Hoon; Jung, Jessica; Noh, Insup; Kim, Chan-Wha

2015-09-01

Modulation of both mechanical properties and biocompatibilities of hyaluronic acid (HA) hydrogels is very importance for their applications in biomaterials. Pure HA solution was converted into a hydrogel by using butanediol diglycidyl ether (BDDE) as a crosslinking agent. Mechanical properties of the HA hydrogels have been evaluated by adding up different amount of BDDEs. While the mechanical properties of the obtained HA hydrogels were evaluated by measuring their crosslinking degrees, elastic modulus and viscosity, their in vitro biocompatibilities were done by measuring the degrees of anti-inflammatory reactions, cell viabilities and cytotoxicity. The degrees of anti-inflammatory reactions were determined by measuring the amount of nitric oxides (NOs) released from lipopolysaccharide(LPS)(+)-induced macrophages; cell viability was evaluated by observing differences in the behaviors of fibroblasts covered with the HA hydrogels, compared with those covered with the films of Teflon and Latex. Cytotoxicity of the HA hydrogels was also evaluated by measuring the degrees of viability of the cells exposed on the extracts of the HA hydrogels over those of Teflon, Latex and pure HA solutions by the assays of thiazoly blue tetrazolium bromide (MTT), neutral reds, and bromodeoxyuridine (BrdU). The results showed that employment of BDDEs beyond critical amounts showed lower biocompatibility of the crosslinked HA hydrogels but higher crosslinking degrees and mechanical properties, indicating the importance of controlling the HA concentrations, BDDE amounts and their reaction times for the synthesis of the crosslinked HA hydrogels for their clinical applications as biomaterials. © 2015 Wiley Periodicals, Inc.

Inability of keratinocytes lacking their specific transglutaminase to form cross-linked envelopes: Absence of envelopes as a simple diagnostic test for lamellar ichthyosis

PubMed Central

Jeon, Saewha; Djian, Philippe; Green, Howard

1998-01-01

Epidermal keratinocytes, late in their terminal differentiation, form cross-linked envelopes resistant to ionic detergent and reducing agent. Because the cross-linking process is catalyzed by the keratinocyte transglutaminase, the absence of active transglutaminase should result in failure of the keratinocyte to form a cross-linked envelope. Three keratinocyte strains bearing mutations in the keratinocyte transglutaminase were examined: two contained no detectable transglutaminase mRNA and none contained active enzyme. All three were unable to form cross-linked envelopes, either spontaneously in stratified cultures or upon induction with Ca2+. Although stratum corneum of normal humans and scales from patients with different ichthyotic diseases contain cross-linked envelopes, those from patients with transglutaminase-negative lamellar ichthyosis do not. Therefore, the disease due to the absence of transglutaminase may be readily distinguished from other ichthyotic diseases by a simple test for cross-linked envelopes. PMID:9435253

The role of ADP-ribosylation in regulating DNA interstrand crosslink repair

PubMed Central

Gunn, Alasdair R.; Banos-Pinero, Benito; Paschke, Peggy; Sanchez-Pulido, Luis; Ariza, Antonio; Day, Joseph; Emrich, Mehera; Leys, David; Ponting, Chris P.

2016-01-01

ABSTRACT ADP-ribosylation by ADP-ribosyltransferases (ARTs) has a well-established role in DNA strand break repair by promoting enrichment of repair factors at damage sites through ADP-ribose interaction domains. Here, we exploit the simple eukaryote Dictyostelium to uncover a role for ADP-ribosylation in regulating DNA interstrand crosslink repair and redundancy of this pathway with non-homologous end-joining (NHEJ). In silico searches were used to identify a protein that contains a permutated macrodomain (which we call aprataxin/APLF-and-PNKP-like protein; APL). Structural analysis reveals that this permutated macrodomain retains features associated with ADP-ribose interactions and that APL is capable of binding poly(ADP-ribose) through this macrodomain. APL is enriched in chromatin in response to cisplatin treatment, an agent that induces DNA interstrand crosslinks (ICLs). This is dependent on the macrodomain of APL and the ART Adprt2, indicating a role for ADP-ribosylation in the cellular response to cisplatin. Although adprt2− cells are sensitive to cisplatin, ADP-ribosylation is evident in these cells owing to redundant signalling by the double-strand break (DSB)-responsive ART Adprt1a, promoting NHEJ-mediated repair. These data implicate ADP-ribosylation in DNA ICL repair and identify that NHEJ can function to resolve this form of DNA damage in the absence of Adprt2. PMID:27587838

Absence of cross-resistance between two alkylating agents: BCNU vs bifunctional galactitol.

PubMed

Institóris, E; Szikla, K; Otvös, L; Gál, F

1989-01-01

Dianhydrogalactitol (DAG) increased the life span of both BCNU-sensitive and -resistant L1210 tumor-bearing mice. However, the BCNU-resistant strain showed slightly lower sensitivity against DAG, which could be overcome by an increase in drug dose of ca. 20%. The somewhat lower sensitivity was proportional to a slightly reduced DNA cross-linking formation induced by DAG in BCNU-resistant cells. The amount of DNA cross-links was determined by measurement of the 1,6-di(guaninyl)-galactitol content of DNA. The slight reduction in cross-links is not attributable to DNA repair but rather to other factors that seem to prevent the formation of DNA-drug adducts. The absence of cross-resistance is explained by different kinds of DNA damage caused by the two alkylating agents and the presumably different defense mechanisms developed by cells against these lesions.

Preparation and characterization of safe microparticles based on xylan.

PubMed

Cartaxo da Costa Urtiga, Silvana; Aquino Azevedo de Lucena Gabi, Camilla; Rodrigues de Araújo Eleamen, Giovanna; Santos Souza, Bartolomeu; Pessôa, Hilzeth de Luna Freire; Marcelino, Henrique Rodrigues; Afonso de Moura Mendonça, Elisângela; Egito, Eryvaldo Sócrates Tabosa do; Oliveira, Elquio Eleamen

2017-10-01

This work describes the preparation and evaluation of safe xylan-based microparticles prepared by cross-linking polymerization using sodium trimetaphosphate. The resulting microparticles were evaluated for morphology, particle size, polymer-cross-link agent interaction, and in vitro toxicity. The microparticles showed narrow monodisperse size distributions with their mean sizes being between 3.5 and 12.5 µm in dried state. FT-IR analyzes confirmed the interaction between sodium trimetaphosphate and xylan during the cross-linking process with formation of phosphate ester bonds. Additionally, the X-ray diffraction patterns and FT-IR analyzes suggested that little or no cross-linking agent remained inside the microparticles. Furthermore, the in-vitro studies using Artemia salina and human erythrocytes revealed that the microparticles are not toxic. Therefore, the overall results suggest that these xylan microparticles can be used as a platform for new drug delivery system.

Hydrogels in a historical perspective: from simple networks to smart materials.

PubMed

Buwalda, Sytze J; Boere, Kristel W M; Dijkstra, Pieter J; Feijen, Jan; Vermonden, Tina; Hennink, Wim E

2014-09-28

Over the past decades, significant progress has been made in the field of hydrogels as functional biomaterials. Biomedical application of hydrogels was initially hindered by the toxicity of crosslinking agents and limitations of hydrogel formation under physiological conditions. Emerging knowledge in polymer chemistry and increased understanding of biological processes resulted in the design of versatile materials and minimally invasive therapies. Hydrogel matrices comprise a wide range of natural and synthetic polymers held together by a variety of physical or chemical crosslinks. With their capacity to embed pharmaceutical agents in their hydrophilic crosslinked network, hydrogels form promising materials for controlled drug release and tissue engineering. Despite all their beneficial properties, there are still several challenges to overcome for clinical translation. In this review, we provide a historical overview of the developments in hydrogel research from simple networks to smart materials. Copyright © 2014 Elsevier B.V. All rights reserved.

Macromolecular cross-linked enzyme aggregates (M-CLEAs) of α-amylase.

PubMed

Nadar, Shamraja S; Muley, Abhijeet B; Ladole, Mayur R; Joshi, Pranoti U

2016-03-01

Macromolecular cross-linked enzyme aggregates (M-CLEAs) of α-amylase were prepared by precipitation and subsequent cross-linking. The non-toxic, biodegradable, biocompatible, renewable polysaccharide based macromolecular cross-linkers viz. agar, chitosan, dextran, and gum arabic were used as a substitute for traditional glutaraldehyde to augment activity recovery toward macromolecular substrate. Macromolecular cross-linkers were prepared by periodate mediated controlled oxidation of polysaccharides. The effects of precipitating agent, concentration and different cross-linkers on activity recovery of α-amylase CLEAs were investigated. α-Amylase aggregated with ammonium sulphate and cross-linked by dextran showed 91% activity recovery, whereas glutaraldehyde CLEAs (G-CLEAs) exhibited 42% activity recovery. M-CLEAs exhibited higher thermal stability in correlation with α-amylase and G-CLEAs. Moreover, dextran and chitosan M-CLEAs showed same affinity for starch hydrolysis as of free α-amylase. The changes in secondary structures revealed the enhancements in structural and conformational rigidity attributed by cross-linkers. Finally, after five consecutive cycles dextran M-CLEAs retained 1.25 times higher initial activity than G-CLEAs. Copyright © 2015 Elsevier B.V. All rights reserved.

Crosslinking to enhance colloidal stability and redispersity of layered double hydroxide nanoparticles.

PubMed

Zuo, Huali; Gu, Zi; Cooper, Helen; Xu, Zhi Ping

2015-12-01

This article introduces a strategy for stabilizing and redispersing layered double hydroxide (LDH) nanoparticles by crosslinking bovine serum albumin (BSA) coated onto the surface. The strategy involves optimization of the amount of the crosslinking agent glutaraldehyde (GTA) to achieve minimal aggregation and ready redispersion. LDH nanoparticles were prepared by co-precipitation and hydrothermal treatment, with subsequent BSA coating at the BSA/LDH mass ratio of 5:2. BSA coated onto LDH nanoparticles was crosslinked with different amounts of GTA. Aggregation studies using dilution assays, dynamic light scattering, and zeta potential analysis indicated that severe aggregation at lower LDH nanoparticle concentrations can be prevented by proper crosslinking of BSA with GTA. The GTA-crosslinked BSA-coated nanoparticles showed excellent redispersity compared to the non-crosslinked nanoparticles. In vitro cytotoxicity and cell uptake were found to be minimally affected by GTA-crosslinking. The new strategy therefore provides a much more effective method for the prevention of LDH nanoparticle aggregation and improved LDH nanoparticle redispersion for use in a wide variety of bio-applications in vitro and in vivo. Copyright © 2015 Elsevier Inc. All rights reserved.

Repair of DNA-polypeptide crosslinks by human excision nuclease

NASA Astrophysics Data System (ADS)

Reardon, Joyce T.; Sancar, Aziz

2006-03-01

DNA-protein crosslinks are relatively common DNA lesions that form during the physiological processing of DNA by replication and recombination proteins, by side reactions of base excision repair enzymes, and by cellular exposure to bifunctional DNA-damaging agents such as platinum compounds. The mechanism by which pathological DNA-protein crosslinks are repaired in humans is not known. In this study, we investigated the mechanism of recognition and repair of protein-DNA and oligopeptide-DNA crosslinks by the human excision nuclease. Under our assay conditions, the human nucleotide excision repair system did not remove a 16-kDa protein crosslinked to DNA at a detectable level. However, 4- and 12-aa-long oligopeptides crosslinked to the DNA backbone were recognized by some of the damage recognition factors of the human excision nuclease with moderate selectivity and were excised from DNA at relatively efficient rates. Our data suggest that, if coupled with proteolytic degradation of the crosslinked protein, the human excision nuclease may be the major enzyme system for eliminating protein-DNA crosslinks from the genome. damage recognition | nucleotide excision repair

Adhesion strength of norbornene-based self-healing agents to an amine-cured epoxy

NASA Astrophysics Data System (ADS)

Huang, Guang Chun; Lee, Jong Keun; Kessler, Michael R.; Yoon, Sungho

2009-07-01

Self-healing is triggered by crack propagation through embedded microcapsules in an epoxy matrix, which then release the liquid healing agent into the crack plane. Subsequent exposure of the healing agent to the chemical catalyst initiates ring-opening metathesis polymerization (ROMP) and bonding of the crack faces. In order to improve self-healing functionality, it is necessary to enhance adhesion of polymerized healing agent within the crack to the matrix resin. In this study, shear bond strength between different norbornene-based healing agents and an amine-cured epoxy resin was evaluated using the single lap shear test method (ASTM D3163, modified). The healing agents tested include endodicyclopentadiene (endo-DCPD), 5-ethylidene-2-norbornene (ENB) and DCPD/ENB blends. 5-Norbornene-2-methanol (NBM) was used as an adhesion promoter, containing hydroxyl groups to form hydrogen bonds with the amine-cured epoxy. A custom synthesized norbornene-based crosslinking agent was also added to improve adhesion for ENB by increasing the crosslinking density of the adhesive after ROMP. The healing agents were polymerized with varying loadings of the 1st generation Grubbs' catalyst at different reaction times and temperatures.

Mechanical Properties and Morphologies of Carboxyl-Terminated Butadiene Acrylonitrile Liquid Rubber/Epoxy Blends Compatibilized by Pre-Crosslinking.

PubMed

Xu, Shiai; Song, Xiaoxue; Cai, Yangben

2016-07-29

In order to enhance the compatibilization and interfacial adhesion between epoxy and liquid carboxyl-terminated butadiene acrylonitrile (CTBN) rubber, an initiator was introduced into the mixture and heated to initiate the cross-linking reaction of CTBN. After the addition of curing agents, the CTBN/epoxy blends with a localized interpenetrating network structure were prepared. The mechanical properties and morphologies of pre-crosslinked and non-crosslinked CTBN/epoxy blends were investigated. The results show that the tensile strength, elongation at break and impact strength of pre-crosslinked CTBN/epoxy blends are significantly higher than those of non-crosslinked CTBN/epoxy blends, which is primarily due to the enhanced interfacial strength caused by the chemical bond between the two phases and the localized interpenetrating network structure. Both pre-crosslinked and non-crosslinked CTBN/epoxy blends show a bimodal distribution of micron- and nano-sized rubber particles. However, pre-crosslinked CTBN/epoxy blends have smaller micron-sized rubber particles and larger nano-sized rubber particles than non-crosslinked CTBN/epoxy blends. The dynamic mechanical analysis shows that the storage modulus of pre-crosslinked CTBN/epoxy blends is higher than that of non-crosslinked CTBN/epoxy blends. The glass transition temperature of the CTBN phase in pre-crosslinked CTBN/epoxy blends increases slightly compared with the CTBN/epoxy system. The pre-crosslinking of rubber is a promising method for compatibilization and controlling the morphology of rubber-modified epoxy materials.

Mechanical Properties and Morphologies of Carboxyl-Terminated Butadiene Acrylonitrile Liquid Rubber/Epoxy Blends Compatibilized by Pre-Crosslinking

PubMed Central

Xu, Shiai; Song, Xiaoxue; Cai, Yangben

2016-01-01

In order to enhance the compatibilization and interfacial adhesion between epoxy and liquid carboxyl-terminated butadiene acrylonitrile (CTBN) rubber, an initiator was introduced into the mixture and heated to initiate the cross-linking reaction of CTBN. After the addition of curing agents, the CTBN/epoxy blends with a localized interpenetrating network structure were prepared. The mechanical properties and morphologies of pre-crosslinked and non-crosslinked CTBN/epoxy blends were investigated. The results show that the tensile strength, elongation at break and impact strength of pre-crosslinked CTBN/epoxy blends are significantly higher than those of non-crosslinked CTBN/epoxy blends, which is primarily due to the enhanced interfacial strength caused by the chemical bond between the two phases and the localized interpenetrating network structure. Both pre-crosslinked and non-crosslinked CTBN/epoxy blends show a bimodal distribution of micron- and nano-sized rubber particles. However, pre-crosslinked CTBN/epoxy blends have smaller micron-sized rubber particles and larger nano-sized rubber particles than non-crosslinked CTBN/epoxy blends. The dynamic mechanical analysis shows that the storage modulus of pre-crosslinked CTBN/epoxy blends is higher than that of non-crosslinked CTBN/epoxy blends. The glass transition temperature of the CTBN phase in pre-crosslinked CTBN/epoxy blends increases slightly compared with the CTBN/epoxy system. The pre-crosslinking of rubber is a promising method for compatibilization and controlling the morphology of rubber-modified epoxy materials. PMID:28773762

Effect of the cross-linking agent on performances of NaCS-CS/WSC microcapsules.

PubMed

Wu, Qing-Xi; Xu, Xin; Wang, Zu-Li; Yao, Shan-Jing; Tong, Wang-Yu; Chen, Yan

2016-11-01

Based on the properties of oppositely charged natural polysaccharides, the polyelectrolyte complexes (PECs) prepared with chitosan-related polycationic polyelectrolytes and cellulose-related polyanionic polyelectrolytes have been widely concerned for their potential applications as micro-drug-carriers for colon. However, the poor mechanical property of the PECs becomes the obstacle encountered in practical applications. This study investigated the effect of the cross-linking agent (sodium polyphosphate, PPS) on the performances of sodium cellulose sulfate -chitosan/water soluble chitosan (NaCS-CS/WSC) microcapsules. The results revealed that PPS could penetrate through the PEC film and form tighter interior structures compared with the microcapsules without the addition of cross-linking agent. The NaCS-CS microcapsules and NaCS-WSC microcapsules with or without PPS had distinct microstructures, which could be ascribed to the different physicochemical properties of CS and WSC. During the formation process, CS can be dissolved in water under acidic conditions, while WSC can be directly dissolved and protonated in acid-free aqueous providing NH3(+) groups quickly, which resulted in the microstructure's difference. Further analysis showed the NaCS-CS-PPS microcapsules and NaCS-WSC-PPS microcapsules had lower swelling ratios due to their tighter interior microstructures that formed. The cross-linking agent had important effect on the total mass of PECs that produced; moreover, the decline of zeta potential of NaCS-CS-PPS microcapsules was lower than that of NaCS-CS microcapsules, similar trend was found in the NaCS-WSC-PPS microcapsules compared with NaCS-WSC microcapsules, indicating the PPS participated in the interactions and played a role in the microcapsules' formation process. Copyright © 2016 Elsevier B.V. All rights reserved.

Efficacy of new natural biomodification agents from Anacardiaceae extracts on dentin collagen cross-linking.

PubMed

Moreira, M A; Souza, N O; Sousa, R S; Freitas, D Q; Lemos, M V; De Paula, D M; Maia, F J N; Lomonaco, D; Mazzetto, S E; Feitosa, V P

2017-10-01

Several polyphenols from renewable sources were surveyed for dentin biomodification. However, phenols from cashew nut shell liquid (CNSL, Anacardium occidentale) and from Aroeira (Myracrodruon urundeuva) extract have never been evaluated. The present investigation aimed to compare the dentin collagen crosslinking (biomodification) effectiveness of polyphenols from Aroeira stem bark extract, proanthocyanidins (PACs) from grape-seed extract (Vitis vinifera), cardol and cardanol from CNSL after clinically relevant treatment for one minute. Three-point bending test was used to obtain the elastic modulus of fully demineralized dentin beams before and after biomodification, whilst color change and mass variation were evaluated after four weeks water biodegradation. Color aspect was assessed by optical images after biodegradation whereas collagen cross-linking was investigated by micro-Raman spectroscopy. Statistical analysis was performed with repeated-measures two way ANOVA and Tukey's test (p<0.05). The increase in elastic modulus after biomodification was in the order cardol>cardanol>aroeira=PACs with cardol solution achieving mean 338.2% increase. The mass increase after biomodification followed the same order aforementioned. Nevertheless, after four weeks aging, more hydrophobic agent (cardanol) induced the highest resistance against water biodegradation. Aroeira and cardol attained intermediate outcomes whereas PACs provided the lower resistance. Tannin-based agents (Aroeira and PACs) stained the specimens in dark brown color. No color alteration was observed with cardol and cardanol treatments. All four agents achieved crosslinking in micro-Raman after one minute application. In conclusion, major components of CNSL yield overall best dentin biomodification outcomes when applied for one minute without staining the dentin collagen. Copyright © 2017 The Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

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.

Studies of the permeation properties of glomerular basement membrane: cross-linking renders glomerular basement membrane permeable to protein.

PubMed

Walton, H A; Byrne, J; Robinson, G B

1992-03-20

Cross-linking glomerular basement membrane (GBM) has been shown to render it more permeable to protein. Isolated pig GBM was cross-linked with dimethylmalonimidate which reacts selectively with lysine epsilon-NH2 groups or with glutaraldehyde, a less selective cross-linking agent. Studies of the ultrafiltration properties of these materials in vitro using cytochrome c, myoglobin, bovine serum albumin and immunoglobulin showed that cross-linking had markedly increased solvent and protein fluxes as compared with native membranes particularly at higher pressures. Filtration studies with serum demonstrated that the cross-linked membranes were more permeable to serum proteins. Thickness measurements under pressure indicated that cross-linked membrane was less compressed than native membrane as pressure was increased. Pore theory did not provide a suitable model for analysis of the results, but analysis of the results using the fibre-matrix hypothesis indicated that cross-linking had the effect of bundling together the fibres (type IV collagen) in the GBM matrix. The effect of cross-linking on filtration could be explained by a combination of contraction of the membrane, fibre bundling and increased rigidity compared with native membrane. Cross-linking of GBM might lead to long-term damage of the glomerular capillary wall in nephritis, so promoting proteinuria.

Targeted Delivery of Ubiquitin-Conjugated BH3 Peptide-Based Mcl-1 Inhibitors into Cancer Cells

PubMed Central

2015-01-01

BH3 peptides are key mediators of apoptosis and have served as the lead structures for the development of anticancer therapeutics. Previously, we reported the application of a simple cysteine-based side chain cross-linking chemistry to NoxaBH3 peptides that led to the generation of the cross-linked NoxaBH3 peptides with increased cell permeability and higher inhibitory activity against Mcl-1 (Muppidi, A., Doi, K., Edwardraja, S., Drake, E. J., Gulick, A. M., Wang, H.-G., Lin, Q. (2012) J. Am. Chem. Soc.134, 1473422920569). To deliver cross-linked NoxaBH3 peptides selectively into cancer cells for enhanced efficacy and reduced systemic toxicity, here we report the conjugation of the NoxaBH3 peptides with the extracellular ubiquitin, a recently identified endogenous ligand for CXCR4, a chemokine receptor overexpressed in cancer cells. The resulting ubiquitin-NoxaBH3 peptide conjugates showed increased inhibitory activity against Mcl-1 and selective killing of the CXCR4-expressing cancer cells. The successful delivery of the NoxaBH3 peptides by ubiquitin into cancer cells suggests that the ubiquitin/CXCR4 axis may serve as a general route for the targeted delivery of anticancer agents. PMID:24410055

Comparative investigation of thermal and mechanical properties of cross-linked epoxy polymers with different curing agents by molecular dynamics simulation.

PubMed

Jeyranpour, F; Alahyarizadeh, Gh; Arab, B

2015-11-01

Molecular dynamics (MD) simulations were carried out to predict the thermal and mechanical properties of the cross-linked epoxy system composed of DGEBA resin and the curing agent TETA. To investigate the effects of curing agents, a comprehensive and comparative study was also performed on the thermal and mechanical properties of DGEBA/TETA and DGEBA/DETDA epoxy systems such as density, glass transition temperature (Tg), coefficient of thermal expansion (CTE) and elastic properties of different cross-linking densities and different temperatures. The results indicated that the glass transition temperature of DGEBA/TETA system calculated through density-temperature data, ∼ 385-395 °K, for the epoxy system with the cross-linking density of 62.5% has a better agreement with the experimental value (Tg, ∼ 400 °K) in comparison to the value calculated through the variation of cell volume in terms of temperature, 430-440 °K. They also indicated that CTE related parameters and elastic properties including Young, Bulk, and shear's moduli, and Poisson's ratio have a relative agreement with the experimental results. Comparison between the thermal and mechanical properties of epoxy systems of DGEBA/TETA and DGEBA/DETDA showed that the DGEBA/DETDA has a higher Tg in all cross linking densities than that of DGEBA/TETA, while higher mechanical properties was observed in the case of DGEBA/TETA in almost all cross linking densities. Copyright © 2015 Elsevier Inc. All rights reserved.

Synthesis and biological activity of mustard derivatives of thymine.

PubMed

Hadj-Bouazza, Amel; Teste, Karine; Colombeau, Ludovic; Chaleix, Vincent; Zerrouki, Rachida; Kraemer, Michel; Sainte Catherine, Odile

2008-05-01

The synthesis and biological activity of a novel DNA cross-linking antitumor agent is presented. The new alkylating agent significantly inhibited cell proliferation, migration and invasion as tested in vitro on the A431 vulvar epidermal carcinoma cell line.

CuC1 thermochemical cycle for hydrogen production

DOEpatents

Fan, Qinbai [Chicago, IL; Liu, Renxuan [Chicago, IL

2012-01-03

An electrochemical cell for producing copper having a dense graphite anode electrode and a dense graphite cathode electrode disposed in a CuCl solution. An anion exchange membrane made of poly(ethylene vinyl alcohol) and polyethylenimine cross-linked with a cross-linking agent selected from the group consisting of acetone, formaldehyde, glyoxal, glutaraldehyde, and mixtures thereof is disposed between the two electrodes.

Cross-linking of bovine and caprine caseins by microbial transglutaminase and their use as microencapsulating agents for n-3 fatty acids

USDA-ARS?s Scientific Manuscript database

Bovine and caprine caseins were cross-linked with microbial transglutaminase (mTG). The mTG-cross-linked bovine or caprine casein dispersion, mixed with 14.5% maltodextrin (DE = 40), was used to prepare emulsions with 10.5% algae oil. Oxidative stability of emulsions was evaluated by peroxide valu...

Cross-linkable liposomes stabilize a magnetic resonance contrast-enhancing polymeric fastener.

PubMed

Smith, Cartney E; Kong, Hyunjoon

2014-04-08

Liposomes are commonly used to deliver drugs and contrast agents to their target site in a controlled manner. One of the greatest obstacles in the performance of such delivery vehicles is their stability in the presence of serum. Here, we demonstrate a method to stabilize a class of liposomes that load gadolinium, a magnetic resonance (MR) contrast agent, as a model cargo on their surfaces. We hypothesized that the sequential adsorption of a gadolinium-binding chitosan fastener on the liposome surface followed by covalent cross-linking of the lipid bilayer would provide enhanced stability and improved MR signal in the presence of human serum. To investigate this hypothesis, liposomes composed of diyne-containing lipids were assembled and functionalized via chitosan conjugated with a hydrophobic anchor and diethylenetriaminepentaacetic acid (DTPA). This postadsorption cross-linking strategy served to stabilize the thermodynamically favorable association between liposome and polymeric fastener. Furthermore, the chitosan-coated, cross-linked liposomes proved more effective as delivery vehicles of gadolinium than uncross-linked liposomes due to the reduced liposome degradation and chitosan desorption. Overall, this study demonstrates a useful method to stabilize a broad class of particles used for systemic delivery of various molecular payloads.

Cross-Linkable Liposomes Stabilize a Magnetic Resonance Contrast-Enhancing Polymeric Fastener

PubMed Central

2015-01-01

Liposomes are commonly used to deliver drugs and contrast agents to their target site in a controlled manner. One of the greatest obstacles in the performance of such delivery vehicles is their stability in the presence of serum. Here, we demonstrate a method to stabilize a class of liposomes that load gadolinium, a magnetic resonance (MR) contrast agent, as a model cargo on their surfaces. We hypothesized that the sequential adsorption of a gadolinium-binding chitosan fastener on the liposome surface followed by covalent cross-linking of the lipid bilayer would provide enhanced stability and improved MR signal in the presence of human serum. To investigate this hypothesis, liposomes composed of diyne-containing lipids were assembled and functionalized via chitosan conjugated with a hydrophobic anchor and diethylenetriaminepentaacetic acid (DTPA). This postadsorption cross-linking strategy served to stabilize the thermodynamically favorable association between liposome and polymeric fastener. Furthermore, the chitosan-coated, cross-linked liposomes proved more effective as delivery vehicles of gadolinium than uncross-linked liposomes due to the reduced liposome degradation and chitosan desorption. Overall, this study demonstrates a useful method to stabilize a broad class of particles used for systemic delivery of various molecular payloads. PMID:24635565

Tunable Elastomers with an Antithrombotic Component for Cardiovascular Applications.

PubMed

Stahl, Alexander M; Yang, Yunzhi Peter

2018-05-31

This study reports the development of a novel family of biodegradable polyurethanes for use as tissue engineered cardiovascular scaffolds or blood-contacting medical devices. Covalent incorporation of the antiplatelet agent dipyridamole into biodegradable polycaprolactone-based polyurethanes yields biocompatible materials with improved thromboresistance and tunable mechanical strength and elasticity. Altering the ratio of the dipyridamole to the diisocyanate linking unit and the polycaprolactone macromer enables control over both the drug content and the polymer cross-link density. Covalent cross-linking in the materials achieves significant elasticity and a tunable range of elastic moduli similar to that of native cardiovascular tissues. Interestingly, the cross-link density of the polyurethanes is inversely related to the elastic modulus, an effect attributed to decreasing crystallinity in the more cross-linked polymers. In vitro characterization shows that the antiplatelet agent is homogeneously distributed in the materials and is released slowly throughout the polymer degradation process. The drug-containing polyurethanes support endothelial cell and vascular smooth muscle cell proliferation, while demonstrating reduced levels of platelet adhesion and activation, supporting their candidacy as promising substrates for cardiovascular tissue engineering. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

DSC investigation of bovine hide collagen at varying degrees of crosslinking and humidities.

PubMed

Schroepfer, Michaela; Meyer, Michael

2017-10-01

Bovine hide collagen (nonCLC; non-CrossLinked Collagen) was analysed by differential scanning calorimetry (DSC) at different hydration degrees and compared with hide collagen samples crosslinked with glutaraldehyde (CLC-GA) and chromium(III) ions (CLC-Cr), respectively. Crosslinking and drying were confirmed to increase the denaturation temperature. Different regions were assigned, that reflect the variation of the influence of water on the denaturation temperature. Furthermore, at moderate hydration degrees, the enthalpies of non-crosslinked collagen increase compared to the fully hydrated state. This reflects a glue-like action of water in the range of 25% hydration. Crosslinking of bovine hide collagen decreases the enthalpy by 25% in the fully hydrated state, even at very low levels of crosslinking This can be explained by intensive effects of the crosslinking agent on the hydration network of the collagen molecules, assuming that the enthalpies are principally a result of hydrogen bonding. At very low water contents DSC peaks of CLC-Cr completely disappear. This could be explained by competition between hydroxosulfochromate(III) complexes and collagen for water. Copyright © 2017 Elsevier B.V. All rights reserved.

Crosslinkable coatings from phosphorylcholine-based polymers.

PubMed

Lewis, A L; Cumming, Z L; Goreish, H H; Kirkwood, L C; Tolhurst, L A; Stratford, P W

2001-01-01

2-Methacryloyloxyethyl phosphorylcholine (MPC) was synthesised and then used in the preparation of crosslinked polymer membranes with lauryl methacrylate, hydroxypropyl methacrylate and trimethoxysilylpropyl methacrylate (crosslinker) comonomers. Some physical aspects of the membrane properties were evaluated in order to establish the basis for the synthesis of a series of post-crosslinkable polymers. These materials were made by copolymerisation of the constituent monomers via a free radical method, and characterised using NMR, FT-IR, viscometry and elemental analysis. The optimum crosslink density and conditions required for curing coatings of these polymers were investigated using atomic force microscopy (AFM) and showed the inclusion of 5 mol% silyl crosslinking agent to be ideal. A nanoindentation technique was employed to determine if the coating developed elasticity upon crosslinking. The biological properties of the coatings were evaluated using a variety of protein adsorption assays and blood contacting experiments, and an enzyme immunoassay was developed to detect E. coli in order to assess the level of bacterial adhesion to these biomaterials. Polymers of this type were shown to be very useful as coating materials for improving the biocompatibility of, or reducing the levels of adherent bacteria to medical devices.

Recruitment of Fanconi Anemia and Breast Cancer Proteins to DNA Damage Sites is differentially Governed by Replication

PubMed Central

Shen, Xi; Do, Huong; Li, Yongjian; Chung, Woo-Hyun; Tomasz, Maria; de Winter, Johan P.; Xia, Bing; Elledge, Stephen J.; Wang, Weidong; Li, Lei

2009-01-01

Summary Fanconi anemia (FA) is characterized by cellular hypersensivity to DNA crosslinking agents, but how the Fanconi pathway protects cells from DNA crosslinks and whether FA proteins act directly on crosslinks remains unclear. We developed a chromatin-IP-based strategy termed eChIP and detected association of multiple FA proteins with DNA crosslinks in vivo. Inter-dependence analyses revealed that crosslink-specific enrichment of various FA proteins is controlled by distinct mechanisms. BRCA-related FA proteins (BRCA2, FANCJ/BACH1, and FANCN/PALB2), but not FA core and I/D2 complexes, require replication for their crosslink association. FANCD2, but not FANCJ and FANCN, requires the FA core complex for its recruitment. FA core complex requires nucleotide excision repair proteins XPA and XPC for its association. Consistent with the distinct recruitment mechanism, recombination-independent crosslink repair was inversely affected in cells deficient of FANC-core versus BRCA-related FA proteins. Thus, FA proteins participate in distinct DNA damage response mechanisms governed by DNA replication status. PMID:19748364

Mechanical Strength Improvements of Carbon Nanotube Threads through Epoxy Cross-Linking

PubMed Central

Yu, Qingyue; Alvarez, Noe T.; Miller, Peter; Malik, Rachit; Haase, Mark R.; Schulz, Mark; Shanov, Vesselin; Zhu, Xinbao

2016-01-01

Individual Carbon Nanotubes (CNTs) have a great mechanical strength that needs to be transferred into macroscopic fiber assemblies. One approach to improve the mechanical strength of the CNT assemblies is by creating covalent bonding among their individual CNT building blocks. Chemical cross-linking of multiwall CNTs (MWCNTs) within the fiber has significantly improved the strength of MWCNT thread. Results reported in this work show that the cross-linked thread had a tensile strength six times greater than the strength of its control counterpart, a pristine MWCNT thread (1192 MPa and 194 MPa, respectively). Additionally, electrical conductivity changes were observed, revealing 2123.40 S·cm−1 for cross-linked thread, and 3984.26 S·cm−1 for pristine CNT thread. Characterization suggests that the obtained high tensile strength is due to the cross-linking reaction of amine groups from ethylenediamine plasma-functionalized CNT with the epoxy groups of the cross-linking agent, 4,4-methylenebis(N,N-diglycidylaniline). PMID:28787868

Preparation and properties of adjacency crosslinked polyurethane-urea elastomers

NASA Astrophysics Data System (ADS)

Wu, Yuan; Cao, Yu-Yang; Wu, Shou-Peng; Li, Zai-Feng

2012-12-01

Adjacency crosslinked polyurethane-urea (PUU) elastomers with different crosslinking density were prepared by using hydroxyl-terminated liquid butadiene-nitrile (HTBN), toluene diisocyanate (TDI) and chain extender 3,5-dimethyl thio-toluene diamine (DMTDA) as raw materials, dicumyl peroxide (DCP) as initiator, and N,N'-m-phenylene dimaleimide (HVA-2) as the crosslinking agent. The influences of the crosslinking density and temperature on the structure and properties of such elastomers were investigated. The crosslinking density of PUU elastomer was tested by the NMR method. It is found that when the content of HVA-2 is 1.5%, the mechanical properties of polyurethane elastomer achieve optimal performance. By testing thermal performance of PUU, compared with linear PUU, the thermal stability of the elastomers has a marked improvement. With the addition of HVA-2, the loss factor tan δ decreases. FT-IR spectral studies of PUU elastomer at various temperatures were performed. From this study, heat-resistance polyurethane could be prepared, and the properties of PUU at high temperature could be improved obviously.

Supercritical CO2 foaming of radiation crosslinked polypropylene/high-density polyethylene blend: Cell structure and tensile property

NASA Astrophysics Data System (ADS)

Yang, Chenguang; Xing, Zhe; Zhang, Mingxing; Zhao, Quan; Wang, Mouhua; Wu, Guozhong

2017-12-01

A blend of isotactic polypropylene (PP) with high-density polyethylene (HDPE) in different PP/HDPE ratios was irradiated by γ-ray to induce cross-linking and then foamed using supercritical carbon dioxide (scCO2) as a blowing agent. Radiation effect on the melting point and crystallinity were analyzed in detail. The average cell diameter and cell density were compared for PP/HDPE foams prepared under different conditions. The optimum absorbed dose for the scCO2 foaming of PP/HDPE in terms of foaming ability and cell structure was 20 kGy. Tensile measurements showed that the elongation at break and tensile strength at break of the crosslinked PP/HDPE foams were higher than the non-crosslinked ones. Of particular interest was the increase in the foaming temperature window from 4 ℃ for pristine PP to 8-12 ℃ for the radiation crosslinked PP/HDPE blends. This implies much easier handling of scCO2 foaming of crosslinked PP with the addition of HDPE.

Crosslinking of surface antibodies and Fc sub. gamma. receptors: Theory and application

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

Wofsy, C.; Goldstein, B.

1991-03-15

In an immune response, the crosslinking of surface immunoglobulin (sIg) on B cells by multiply-bound ligand activates a range of cell responses, culminating in the production of antibody-secreting cells. However, when the crosslinking agent is itself an antibody, B cell activation is inhibited. Solution antibody (IgG) can bind simultaneously to sIg and to another cell surface receptor, Fc{sub {gamma}}R, co-crosslinking' the distinct receptors. Experiments point to co-crosslinking as the inhibitory signal. It is not clear how co-crosslinking inhibits B cell stimulation. The authors construct and analyze a mathematical model aimed at clarifying the nature and mechanisms of action of themore » separate cell signals controlling B cell responses to antibodies. Basophils and mast cells respond to the crosslinking of cell surface antibody by releasing histamine. Like B cells, basophils also express FC{sub {gamma}}R. They use their model to analyze new data on the effect of antibody-induced co-crosslinking of the two types of receptor on this family of cells. Predictions of the model indicate that an observed difference between the response patterns induced by antibodies and by antibody fragments that cannot bind to FC{sub {gamma}}R can be explained if co-crosslinking is neither inhibitory nor stimulatory in this system.« less

Ionic Hydrogel Based on Chitosan Cross-Linked with 6-Phosphogluconic Trisodium Salt as a Drug Delivery System.

PubMed

Martínez-Martínez, Mayte; Rodríguez-Berna, Guillermo; Gonzalez-Alvarez, Isabel; Hernández, Ma Jesús; Corma, Avelino; Bermejo, Marival; Merino, Virginia; Gonzalez-Alvarez, Marta

2018-04-09

In this work, 6-phosphogluconic trisodium salt (6-PG - Na + ) is introduced as a new aqueous and nontoxic cross-linking agent to obtain ionic hydrogels. Here, it is shown the formation of hydrogels based on chitosan cross-linked with 6-PG - Na + . This formulation is obtained by ionic interaction of cationic groups of polymer with anionic groups of the cross-linker. These hydrogels are nontoxic, do not cause dermal irritation, are easy to extend, and have an adequate adhesion force to be applied as polymeric film over the skin. This formulation exhibits a first order release kinetic and can be applied as drug vehicle for topical administration or as wound dressing for wound healing. The primary goal of this communication is to report the identification and utility of 6-phosphogluconic trisodium salt (6-PG - Na + ) as a nontoxic cross-linker applicable for cationic polymers.

A New Class of Antibody-Drug Conjugates with Potent DNA Alkylating Activity.

PubMed

Miller, Michael L; Fishkin, Nathan E; Li, Wei; Whiteman, Kathleen R; Kovtun, Yelena; Reid, Emily E; Archer, Katie E; Maloney, Erin K; Audette, Charlene A; Mayo, Michele F; Wilhelm, Alan; Modafferi, Holly A; Singh, Rajeeva; Pinkas, Jan; Goldmacher, Victor; Lambert, John M; Chari, Ravi V J

2016-08-01

The promise of tumor-selective delivery of cytotoxic agents in the form of antibody-drug conjugates (ADC) has now been realized, evidenced by the approval of two ADCs, both of which incorporate highly cytotoxic tubulin-interacting agents, for cancer therapy. An ongoing challenge remains in identifying potent agents with alternative mechanisms of cell killing that can provide ADCs with high therapeutic indices and favorable tolerability. Here, we describe the development of a new class of potent DNA alkylating agents that meets these objectives. Through chemical design, we changed the mechanism of action of our novel DNA cross-linking agent to a monofunctional DNA alkylator. This modification, coupled with linker optimization, generated ADCs that were well tolerated in mice and demonstrated robust antitumor activity in multiple tumor models at doses 1.5% to 3.5% of maximally tolerated levels. These properties underscore the considerable potential of these purpose-created, unique DNA-interacting conjugates for broadening the clinical application of ADC technology. Mol Cancer Ther; 15(8); 1870-8. ©2016 AACR. ©2016 American Association for Cancer Research.

Microgel particles for the delivery of bioactive materials

DOEpatents

Frechet, Jean M. J.; Murthy Niren

2010-03-23

Novel microgels, microparticles and related polymeric materials capable of delivering bioactive materials to cells for use as vaccines or therapeutic agents. The materials are made using a crosslinker molecule that contains a linkage cleavable under mild acidic conditions. The crosslinker molecule is exemplified by a bisacryloyl acetal crosslinker. The new materials have the common characteristic of being able to degrade by acid hydrolysis under conditions commonly found within the endosomal or lysosomal compartments of cells thereby releasing their payload within the cell. The materials can also be used for the delivery of therapeutics to the acidic regions of tumors and sites of inflammation.

Microgel particles for the delivery of bioactive materials

DOEpatents

Frechet, Jean M.; Murthy, Niren

2006-06-06

Novel microgels, microparticles and related polymeric materials capable of delivering bioactive materials to cells for use as vaccines or therapeutic agents. The materials are made using a crosslinker molecule that contains a linkage cleavable under mild acidic conditions. The crosslinker molecule is exemplified by a bisacryloyl acetal crosslinker. The new materials have the common characteristic of being able to degrade by acid hydrolysis under conditions commonly found within the endosomal or lysosomal compartments of cells thereby releasing their payload within the cell. The materials can also be used for the delivery of therapeutics to the acidic regions of tumors and sites of inflammation.

Mixed-Isotope Labeling with LC-IMS-MS for Characterization of Protein–Protein Interactions by Chemical Cross-Linking

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

Merkley, Eric D.; Baker, Erin S.; Crowell, Kevin L.

2013-02-20

Chemical cross-linking of proteins followed by proteolysis and mass spectrometric analysis of the resulting cross-linked peptides can provide insights into protein structure and protein-protein interactions. However, cross-linked peptides are by necessity of low stoichometry and have different physicochemical properties than linear peptides, routine unambiguous identification of the cross-linked peptides has remained difficult. To address this challenge, we demonstrated the use of liquid chromatography and ion mobility separations coupled with mass spectrometry in combination with a heavy-isotope labeling method. The combination of mixed-isotope cross-linking and ion mobility provided unique and easily interpretable spectral multiplet features for the intermolecular cross-linked peptides. Applicationmore » of the method to two different homodimeric proteins - SrfN, a virulence factor from Salmonella Typhimurium and SO_2176, a protein of unknown function from Shewanella oneidensis- revealed several cross-linked peptides from both proteins that were identified with a low false discovery rate (estimated using a decoy approach). A greater number of cross-linked peptides were identified using ion mobility drift time information in the analysis than when the data were summed across the drift time dimension before analysis. The identified cross-linked peptides migrated more quickly in the ion mobility drift tube than the unmodified peptides.« less

Inhibition of the Nedd8 system sensitizes cells to DNA Inter-strand crosslinking agents

PubMed Central

Kee, Younghoon; Huang, Min; Chang, Sophia; Moreau, Lisa A.; Park, Eunmi; Smith, Peter G.; D’Andrea, Alan D.

2012-01-01

The Fanconi Anemia (FA) pathway is required for repair of DNA interstrand crosslinks (ICLs). FA pathway-deficient cells are hypersensitive to DNA ICL-inducing drugs such as Cisplatin. Conversely, hyperactivation of the FA pathway is a mechanism that may underlie cellular resistance to DNA ICL agents. Modulating FANCD2 monoubiquitination, a key step in the FA pathway, may be an effective therapeutic approach to conferring cellular sensitivity to ICL agents. Here, we show that inhibition of the Nedd8 conjugation system increases cellular sensitivity to DNA ICL-inducing agents. Mechanistically, the Nedd8 inhibition, either by siRNA-mediated knockdown of Nedd8 conjugating enzymes or treatment with a Nedd8 activating enzyme inhibitor MLN4924, suppressed DNA damage-induced FANCD2 monoubiquitination and CHK1 phosphorylation. Our data indicate that inhibition of the FA pathway is largely responsible for the heightened cellular sensitivity to DNA ICLs upon Nedd8 inhibition. These results suggest that a combination of Nedd8 inhibition with ICL-inducing agents may be an effective strategy for sensitizing a subset of drug-resistant cancer cells. PMID:22219386

Genipin-crosslinked silk fibroin/hydroxybutyl chitosan nanofibrous scaffolds for tissue-engineering application.

PubMed

Zhang, Kuihua; Qian, Yongfang; Wang, Hongsheng; Fan, Linpeng; Huang, Chen; Yin, Anlin; Mo, Xiumei

2010-12-01

To improve water-resistant ability and mechanical properties of silk fibroin (SF)/hydroxybutyl chitosan (HBC) nanofibrous scaffolds for tissue-engineering applications, genipin, glutaraldehyde (GTA), and ethanol were used to crosslink electrospun nanofibers, respectively. The mechanical properties of nanofibrous scaffolds were obviously improved after 24 h of crosslinking with genipin and were superior to those crosslinked with GTA and ethanol for 24 h. SEM indicated that crosslinked nanofibers with genipin and GTA vapor had good water-resistant ability. Characterization of the microstructure (porosity and pore structure) demonstrated crosslinked nanofibrous scaffolds with genipin and GTA vapor had lager porosities and mean diameters than those with ethanol. Characterization of FTIR-ATR and (13)C NMR clarified both genipin and GTA acted as crosslinking agents for SF and HBC. Furthermore, genipin could induce SF conformation from random coil or α-helix to β-sheet. Although GTA could also successfully crosslink SF/HBC nanofibrous scaffolds, in long run, genipin maybe a better method due to lower cytotoxicity than GTA. Cell viability studies and wound-healing test in rats clarified that the genipin-crosslinked SF/HBC nanofibrous scaffolds had a good biocompatibility both in vitro and in vivo. These results suggested that genipin-crosslinked SF/HBC nanofibrous scaffolds might be potential candidates for wound dressing and tissue-engineering scaffolds. © 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010.

Chitosan membranes for tissue engineering: comparison of different crosslinkers.

PubMed

Ruini, F; Tonda-Turo, C; Chiono, V; Ciardelli, G

2015-11-03

Chitosan (CS), a derivative of the naturally occurring biopolymer chitin, is an attractive material for biomedical applications thanks to its biocompatibility, biodegradability, antibacterial properties and ability to enhance cell adhesion and growth compared to other biopolymers. However, the physical and mechanical stability of CS based materials in aqueous solutions is limited and crosslinking agents are required to increase CS performances in a biological environment. In this work, the effect of three highly-biocompatible crosslinkers as genipin (GP), γ-glycidoxypropyltrimethoxysilane (GPTMS), dibasic sodium phosphate (DSP) and a combination of GPTMS and DSP (GPTMS_DSP) on CS physicochemical, thermal, morphological, mechanical properties, swelling and degradation behavior was investigated. Infrared spectroscopy and thermogravimetric analyses confirmed the chemical reaction between CS and the different crosslinkers. CS wettability was enhanced when CS was DSP ionically crosslinked showing contact angle values of about 65° and exhibiting a higher swelling behavior compared to covalently crosslinked films. Moreover, all the crosslinking methods analyzed improved the stability of CS in aqueous media, showed model molecule permeation in time and increased the mechanical properties when compared with non-crosslinked films. The possibility to tailor the final properties of CS scaffolds through crosslinking is a key strategy in applying CS in different biomedical and tissue engineering applications. The obtained results reveal that the optimization of the crosslinking mechanism provides CS membrane properties required in different biomedical applications.

Nanocelluloses from jute fibers and their nanocomposites with natural rubber: Preparation and characterization.

PubMed

Thomas, Martin George; Abraham, Eldho; Jyotishkumar, P; Maria, Hanna J; Pothen, Laly A; Thomas, Sabu

2015-11-01

Nanocellulose fibers having an average diameter of 50nm were isolated from raw jute fibers by steam explosion process. The isolation of nanocellulose from jute fibers by this extraction process is proved by SEM, XRD, FTIR, birefringence and TEM characterizations. This nanocellulose was used as the reinforcing agent in natural rubber (NR) latex along with crosslinking agents to prepare crosslinked nanocomposite films. The effects of nanocellulose loading on the morphology and mechanics of the nanocomposites have been carefully analyzed. Significant improvements in the Young's modulus and tensile strength of the nanocomposite were observed because of the reinforcing ability of the nanocellulose in the rubber matrix. A mechanism is suggested for the formation of the Zn-cellulose complex. The three-dimensional network of cellulose nanofibers (cellulose/cellulose network and Zn/cellulose network) in the NR matrix plays a major role in improving the properties of the crosslinked nanocomposites. Copyright © 2015 Elsevier B.V. All rights reserved.

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

Park, Jongmin; Saba, Stacey A.; Hillmyer, Marc A.

We report on the phase separation behaviors of polymerization mixtures containing a polylactide macro-chain transfer agent (PLA-CTA), styrene, divinylbenzene, hydroxyl-terminated PLA (PLA-OH), and a molecular chain transfer agent which enable the ability to tune the pore size of a cross-linked polymer monolith in a facile manner. Cross-linked monoliths were produced from the mixtures via reversible addition-fragmentation chain transfer (RAFT) polymerization and converted into cross-linked porous polymers by selective removal of PLA while retaining the parent morphology. We demonstrate that pore sizes are tunable over a wide range of length scales from the meso- to macroporous regimes by adjusting the ratiomore » of PLA-CTA to PLA-OH in the reaction mixture which causes the phase separation mechanism to change from polymerization-induced microphase separation to polymerization-induced phase separation. The possibility of increasing porosity and inducing simultaneous micro- and macrophase separation was also realized by adjustments in the molar mass of PLA which enabled the synthesis of hierarchically meso- and macroporous polymers.« less

The use of molecular dynamics simulations to evaluate the DNA sequence-selectivity of G-A cross-linking PBD-duocarmycin dimers.

PubMed

Jackson, Paul J M; Rahman, Khondaker M; Thurston, David E

2017-01-01

The pyrrolobenzodiazepine (PBD) and duocarmycin families are DNA-interactive agents that covalently bond to guanine (G) and adenine (A) bases, respectively, and that have been joined together to create synthetic dimers capable of cross-linking G-G, A-A, and G-A bases. Three G-A alkylating dimers have been reported in publications to date, with defined DNA-binding sites proposed for two of them. In this study we have used molecular dynamics simulations to elucidate preferred DNA-binding sites for the three published molecular types. For the PBD-CPI dimer UTA-6026 (1), our simulations correctly predicted its favoured binding site (i.e., 5'-C(G)AATTA-3') as identified by DNA cleavage studies. However, for the PBD-CI molecule ('Compound 11', 3), we were unable to reconcile the results of our simulations with the reported preferred cross-linking sequence (5'-ATTTTCC(G)-3'). We found that the molecule is too short to span the five base pairs between the A and G bases as claimed, but should target instead a sequence such as 5'-ATTTC(G)-3' with two less base pairs between the reacting G and A residues. Our simulation results for this hybrid dimer are also in accord with the very low interstrand cross-linking and in vitro cytotoxicity activities reported for it. Although a preferred cross-linking sequence was not reported for the third hybrid dimer ('27eS', 2), our simulations predict that it should span two base pairs between covalently reacting G and A bases (e.g., 5'-GTAT(A)-3'). Copyright © 2016. Published by Elsevier Ltd.

Field Testing of Thermoplastic Encapsulants in High-Temperature Installations

DOE PAGES

Kempe, Michael D.; Miller, David C.; Wohlgemuth, John H.; ...

2015-11-01

Recently there has been increased interest in using thermoplastic encapsulant materials in photovoltaic modules, but concerns have been raised about whether these would be mechanically stable at high temperatures in the field. This has become a significant topic of discussion in the development of IEC 61730 and IEC 61215. We constructed eight pairs of crystalline-silicon modules and eight pairs of glass/encapsulation/glass thin-film mock modules using different encapsulant materials, of which only two were formulated to chemically crosslink. One module set was exposed outdoors with thermal insulation on the back side in Mesa, Arizona, in the summer (hot-dry), and an identicalmore » module set was exposed in environmental chambers. High-precision creep measurements (±20 μm) and electrical performance measurements indicate that despite many of these polymeric materials operating in the melt or rubbery state during outdoor deployment, no significant creep was seen because of their high viscosity, lower operating temperature at the edges, and/or the formation of chemical crosslinks in many of the encapsulants with age despite the absence of a crosslinking agent. Only an ethylene-vinyl acetate (EVA) encapsulant formulated without a peroxide crosslinking agent crept significantly. When the crystalline-silicon modules, the physical restraint of the backsheet reduced creep further and was not detectable even for the EVA without peroxide. Because of the propensity of some polymeric materials to crosslink as they age, typical thermoplastic encapsulants would be unlikely to result in creep in the vast majority of installations.« less

Preparation of crosslinked enzyme aggregates (CLEAs) of acid urease with urethanase activity and their application.

PubMed

Zhang, Qian; Zha, Xiaohong; Zhou, Nandi; Tian, Yaping

2016-04-01

An acid urease from Providencia rettgeri JN-B815 was purified via ultrasonication, ethanol precipitation, and DEAE ion-exchange column chromatography. It was found that the enzyme exhibits not only urease activity, but also urethanase activity, which made it possible to reduce EC already existed or would produce and its precursor urea at the same time. Then, crosslinked enzyme aggregates of P. rettgeri urease (PRU-CLEAs) were prepared using genipin as crosslinking agent. The purification process of acid urease, the effects of genipin concentration, and crosslinking time on PRU-CLEAs activity were investigated. The crosslinking was performed at pH 4.5 for 2.5 h, using 0.3% genipin as crosslinking agent, and 0.3 g · L(-1) bovine serum albumin as protein feeder. Using the obtained PRU-CLEAs, the removal rate of urea was up to 9.31 mg · L(-1) · h(-1). The removal rate of urea was still up to 7.56 mg · L(-1) · h(-1) after PRU-CLEAs was re-used for 6 times. When PRU-CLEAs were applied in a batch stirred and membrane reactor, the removal rate of urea in rice wine reached 5.16 mg · L(-1) · h(-1) and the removal rate of EC was 9.21 μg · L(-1) · h(-1). Furthermore, the treatment with PRU-CLEAs revealed no significant change of volatile flavor substances in Chinese rice wine. Thus PRU-CLEAs have great potential in the elimination of EC in Chinese rice wine. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Biallelic inactivation of REV7 is associated with Fanconi anemia.

PubMed

Bluteau, Dominique; Masliah-Planchon, Julien; Clairmont, Connor; Rousseau, Alix; Ceccaldi, Raphael; Dubois d'Enghien, Catherine; Bluteau, Olivier; Cuccuini, Wendy; Gachet, Stéphanie; Peffault de Latour, Régis; Leblanc, Thierry; Socié, Gérard; Baruchel, André; Stoppa-Lyonnet, Dominique; D'Andrea, Alan D; Soulier, Jean

2016-09-01

Fanconi anemia (FA) is a recessive genetic disease characterized by congenital abnormalities, chromosome instability, progressive bone marrow failure (BMF), and a strong predisposition to cancer. Twenty FA genes have been identified, and the FANC proteins they encode cooperate in a common pathway that regulates DNA crosslink repair and replication fork stability. We identified a child with severe BMF who harbored biallelic inactivating mutations of the translesion DNA synthesis (TLS) gene REV7 (also known as MAD2L2), which encodes the mutant REV7 protein REV7-V85E. Patient-derived cells demonstrated an extended FA phenotype, which included increased chromosome breaks and G2/M accumulation upon exposure to DNA crosslinking agents, γH2AX and 53BP1 foci accumulation, and enhanced p53/p21 activation relative to cells derived from healthy patients. Expression of WT REV7 restored normal cellular and functional phenotypes in the patient's cells, and CRISPR/Cas9 inactivation of REV7 in a non-FA human cell line produced an FA phenotype. Finally, silencing Rev7 in primary hematopoietic cells impaired progenitor function, suggesting that the DNA repair defect underlies the development of BMF in FA. Taken together, our genetic and functional analyses identified REV7 as a previously undescribed FA gene, which we term FANCV.

Biallelic inactivation of REV7 is associated with Fanconi anemia

PubMed Central

Masliah-Planchon, Julien; Clairmont, Connor; Rousseau, Alix; Ceccaldi, Raphael; Dubois d’Enghien, Catherine; Bluteau, Olivier; Cuccuini, Wendy; Gachet, Stéphanie; Peffault de Latour, Régis; Leblanc, Thierry; Socié, Gérard; Baruchel, André; Stoppa-Lyonnet, Dominique; D’Andrea, Alan D.

2016-01-01

Fanconi anemia (FA) is a recessive genetic disease characterized by congenital abnormalities, chromosome instability, progressive bone marrow failure (BMF), and a strong predisposition to cancer. Twenty FA genes have been identified, and the FANC proteins they encode cooperate in a common pathway that regulates DNA crosslink repair and replication fork stability. We identified a child with severe BMF who harbored biallelic inactivating mutations of the translesion DNA synthesis (TLS) gene REV7 (also known as MAD2L2), which encodes the mutant REV7 protein REV7-V85E. Patient-derived cells demonstrated an extended FA phenotype, which included increased chromosome breaks and G2/M accumulation upon exposure to DNA crosslinking agents, γH2AX and 53BP1 foci accumulation, and enhanced p53/p21 activation relative to cells derived from healthy patients. Expression of WT REV7 restored normal cellular and functional phenotypes in the patient’s cells, and CRISPR/Cas9 inactivation of REV7 in a non-FA human cell line produced an FA phenotype. Finally, silencing Rev7 in primary hematopoietic cells impaired progenitor function, suggesting that the DNA repair defect underlies the development of BMF in FA. Taken together, our genetic and functional analyses identified REV7 as a previously undescribed FA gene, which we term FANCV. PMID:27500492

EVIDENCE FOR BASE EXCISION REPAIR PROCESSING OF DNA INTERSTRAND CROSSLINKS

PubMed Central

Kothandapani, Anbarasi; Patrick, Steve M

2013-01-01

Many bifunctional alkylating agents and anticancer drugs exert their cytotoxicity by producing cross links between the two complementary strands of DNA, termed interstrand crosslinks (ICLs). This blocks the strand separating processes during DNA replication and transcription, which can lead to cell cycle arrest and apoptosis. Cells use multiple DNA repair systems to eliminate the ICLs. Concerted action of repair proteins involved in Nucleotide Excision Repair and Homologous Recombination pathways are suggested to play a key role in the ICL repair. However, recent studies indicate a possible role for Base Excision Repair (BER) in mediating the cytotoxicity of ICL inducing agents in mammalian cells. Elucidating the mechanism of BER mediated modulation of ICL repair would help in understanding the recognition and removal of ICLs and aid in the development of potential therapeutic agents. In this review, the influence of BER proteins on ICL DNA repair and the possible mechanisms of action are discussed. PMID:23219605

Synthesis and antitumor activity evaluation of a novel combi-nitrosourea prodrug: Designed to release a DNA cross-linking agent and an inhibitor of O(6)-alkylguanine-DNA alkyltransferase.

PubMed

Sun, Guohui; Zhang, Na; Zhao, Lijiao; Fan, Tengjiao; Zhang, Shufen; Zhong, Rugang

2016-05-01

The drug resistance of CENUs induced by O(6)-alkylguanine-DNA alkyltransferase (AGT), which repairs the O(6)-alkylated guanine and subsequently inhibits the formation of dG-dC cross-links, hinders the application of CENU chemotherapies. Therefore, the discovery of CENU analogs with AGT inhibiting activity is a promising approach leading to novel CENU chemotherapies with high therapeutic index. In this study, a new combi-nitrosourea prodrug 3-(3-(((2-amino-9H-purin-6-yl)oxy)methyl)benzyl)-1-(2-chloroethyl)-1-nitrosourea (6), designed to release a DNA cross-linking agent and an inhibitor of AGT, was synthesized and evaluated for its antitumor activity and ability to induce DNA interstrand cross-links (ICLs). The results indicated that 6 exhibited higher cytotoxicity against mer(+) glioma cells compared with ACNU, BCNU, and their respective combinations with O(6)-benzylguanine (O(6)-BG). Quantifications of dG-dC cross-links induced by 6 were performed using HPLC-ESI-MS/MS. Higher levels of dG-dC cross-link were observed in 6-treated human glioma SF763 cells (mer(+)), whereas lower levels of dG-dC cross-link were observed in 6-treated calf thymus DNA, when compared with the groups treated with BCNU and ACNU. The results suggested that the superiority of 6 might result from the AGT inhibitory moiety, which specifically functions in cells with AGT activity. Molecular docking studies indicated that five hydrogen bonds were formed between the O(6)-BG analogs released from 6 and the five residues in the active pocket of AGT, which provided a reasonable explanation for the higher AGT-inhibitory activity of 6 than O(6)-BG. Copyright © 2016 Elsevier Ltd. All rights reserved.

Protein glycation, diabetes, and aging.

PubMed

Ulrich, P; Cerami, A

2001-01-01

Biological amines react with reducing sugars to form a complex family of rearranged and dehydrated covalent adducts that are often yellow-brown and/or fluorescent and include many cross-linked structures. Food chemists have long studied this process as a source of flavor, color, and texture changes in cooked, processed, and stored foods. During the 1970s and 1980s, it was realized that this process, called the Maillard reaction or advanced glycation, also occurs slowly in vivo. Advanced glycation endproducts (AGEs) that form are implicated, causing the complications of diabetes and aging, primarily via adventitious and crosslinking of proteins. Long-lived proteins such as structural collagen and lens crystallins particularly are implicated as pathogenic targets of AGE processes. AGE formation in vascular wall collagen appears to be an especially deleterious event, causing crosslinking of collagen molecules to each other and to circulating proteins. This leads to plaque formation, basement membrane thickening, and loss of vascular elasticity. The chemistry of these later-stage, glycation-derived crosslinks is still incompletely understood but, based on the hypothesis that AGE formation involves reactive carbonyl groups, the authors introduced the carbonyl reagent aminoguanidine hydrochloride as an inhibitor of AGE formation in vivo in the mid 1980s. Subsequent studies by many researchers have shown the effectiveness of aminoguanidine in slowing or preventing a wide range of complications of diabetes and aging in animals and, recently, in humans. Since, the authors have developed a new class of agents, exemplified by 4,5-dimethyl-3-phenacylthiazolium chloride (DPTC), which can chemically break already-formed AGE protein-protein crosslinks. These agents are based on a new theory of AGE crosslinking that postulates that alpha-dicarbonyl structures are present in AGE protein-protein crosslinks. In studies in aged animals, DPTC has been shown to be capable of reverting indices of vascular compliance to levels seen in younger animals. Human clinical trials are underway.

Cross-linked polyvinyl pyridine coated glass particle catalyst support and aqueous composition or polyvinyl pyridine adducted microspheres

NASA Technical Reports Server (NTRS)

Rembaum, Alan (Inventor); Gupta, Amitava (Inventor); Volksen, Willi (Inventor)

1981-01-01

Microspheres are produced by cobalt gamma radiation initiated polymerization of a dilute aqueous vinyl pyridine solution. Addition of cross-linking agent provides higher surface area beads. Addition of monomers such as hydroxyethylmethacrylate acrylamide or methacrylamide increases hydrophilic properties and surface area of the beads. High surface area catalytic supports are formed in the presence of controlled pore glass substrate.

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

PubMed

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

2016-10-18

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

Accelerated healing of full-thickness wounds by genipin-crosslinked silk sericin/PVA scaffolds.

PubMed

Aramwit, Pornanong; Siritienthong, Tippawan; Srichana, Teerapol; Ratanavaraporn, Juthamas

2013-01-01

Silk sericin has recently been studied for its advantageous biological properties, including its ability to promote wound healing. This study developed a delivery system to accelerate the healing of full-thickness wounds. Three-dimensional scaffolds were fabricated from poly(vinyl alcohol) (PVA), glycerin (as a plasticizer) and genipin (as a crosslinking agent), with or without sericin. The physical and biological properties of the genipin-crosslinked sericin/PVA scaffolds were investigated and compared with those of scaffolds without sericin. The genipin-crosslinked sericin/PVA scaffolds exhibited a higher compressive modulus and greater swelling in water than the scaffolds without sericin. Sericin also exhibited controlled release from the scaffolds. The genipin-crosslinked sericin/PVA scaffolds promoted the attachment and proliferation of L929 mouse fibroblasts. After application to full-thickness rat wounds, the wounds treated with genipin-crosslinked sericin/PVA scaffolds showed a significantly greater reduction in wound size, collagen formation and epithelialization compared with the control scaffolds without sericin but lower numbers of macrophages and multinucleated giant cells. These results indicate that the delivery of sericin from the novel genipin-crosslinked scaffolds efficiently healed the wound. Therefore, these genipin-crosslinked sericin/PVA scaffolds represent a promising candidate for the accelerated healing of full-thickness wounds. Copyright © 2013 S. Karger AG, Basel.

Improving the performance of transglutaminase-crosslinked microparticles for enteric delivery.

PubMed

Tello, Fernando; Prata, Ana S; Rodrigues, Rodney A F; Sartoratto, Adilson; Grosso, Carlos R F

2016-10-01

Various agents for cross-linking have been investigated for stabilizing and controlling the barrier properties of microparticles for enteric applications. Transglutaminase, in addition to being commercially available for human consumption, presents inferior cross-linking action compared to glutaraldehyde. In this study, the intensity of this enzymatic cross-linking was investigated in microparticles obtained by complex coacervation between gelatin and gum Arabic. The effectiveness of cross-linking in these microparticles was evaluated based on swelling, release of a model substance (parika oleoresin: colored and hydrophobic) and gastrointestinal assays. The cross-linked microparticles remained intact under gastric conditions, whereas the uncross-linked microparticles have been dissolved. However, all of the microparticles have been dissolved under intestinal conditions. The amount of oily core that was released decreased as the amount of transglutaminase increased. For the most efficient microparticles (50U/g of protein), the performance was improved by increasing the pH of cross-linking from 4.0 to 6.0, resulting in a release of 17.1% rather than 32.3% of the core material. These results were considerably closer to the 10.3% of core material released by glutaraldehyde-cross-linked microparticles (1mM/g of protein). Copyright © 2016 Elsevier Ltd. All rights reserved.

Interstrand cross-linking of DNA by 1,3-bis(2-chloroethyl)-1-nitrosourea and other 1-(2-haloethyl)-1-nitrosoureas.

PubMed

Kohn, K W

1977-05-01

Bifunctional alkylating agents are known to cross-link DNA by simultaneously alkylating two guanine residues located on opposite strands. Despite this apparent requirement for bifunctionality, 1-(2-chloroethyl)-1-nitrosoureas bearing a single alkylating function were found to cross-link DNA in vitro. Cross-linking was demonstrated by showing inhibition of alkali-induced strand separation. Extensive cross-linking was observed in DNA treated with 1-(2-chloroethyl)-1-nitrosourea, 1,3-bis-(2-chloroethyl)-1-nitrosourea, and 1-(2-chloroethyl(-3-cyclohexyl-1-nitrosourea. The reaction occurs in two steps, an intital binding followed by a second step which can proceed after removal of unbound drug. It is suggested that the first step is chloroethylation of a nucleophilic site on one strand and that the second step involves displacement of Cl- by a nucleophilic site on the opposite strand, resulting in an ethyl bridge between the strands. Consistent with this possibility, 1-(2-fluoroethyl)-3-cyclohexyl-1-nitrosourea produced much less cross-linking, as expected from the known low activity of F-, compared with Cl-, as leaving group. 1-Methyl-1-nitrosourea, which is known to depurinate DNA, produced no detectable cross-linking.

Increasing Fire Safety of Epoxies

NASA Technical Reports Server (NTRS)

Kourtides, D. A.; Mikroyannidis, J. A.

1985-01-01

Epoxy with increased resistance to fire made by reacting any of three commercial epoxide monomers with curing agent consisting of mixture of isomers called "DCEPD". Curing agent incorporates phosphorus and chlorine directly into crosslinking part of polymer. DCEPD produced by nitrating precursor phosphonylmethyl benzene, then reducing resulting isomeric mixture of dinitro compounds.

Diepoxybutane Interstrand Cross-Links Induce DNA Bending

PubMed Central

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

2011-01-01

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

Assessment of the characteristics and biocompatibility of gelatin sponge scaffolds prepared by various crosslinking methods.

PubMed

Yang, Gang; Xiao, Zhenghua; Long, Haiyan; Ma, Kunlong; Zhang, Junpeng; Ren, Xiaomei; Zhang, Jiang

2018-01-25

This comparative study aims to identify a biocompatible and effective crosslinker for preparing gelatin sponges. Glutaraldehyde (GTA), genipin (GP), 1-ethyl-3-(3-dimethyl aminopropyl)carbodiimide (EDC), and microbial transglutaminase (mTG) were used as crosslinking agents. The physical properties of the prepared samples were characterized, and material degradation was studied in vitro with various proteases and in vivo through subcutaneous implantation of the sponges in rats. Adipose-derived stromal stem cells (ADSCs) were cultured and inoculated onto the scaffolds to compare the cellular biocompatibility of the sponges. Cellular seeding efficiency and digestion time of the sponges were also evaluated. Cellular viability and proliferation in scaffolds were analyzed by fluorescence staining and MTT assay. All the samples exhibited high porosity, good swelling ratio, and hydrolysis properties; however, material strength, hydrolysis, and enzymolytic properties varied among the samples. GTA-sponge and GP-sponge possessed high compressive moduli, and EDC-sponge exhibited fast degradation performance. GTA and GP sponge implants exerted strong in vivo rejections, and the former showed poor cell growth. mTG-sponge exhibited the optimal comprehensive performance, with good porosity, compressive modulus, anti-degradation ability, and good biocompatibility. Hence, mTG-sponge can be used as a scaffold material for tissue engineering applications.

Mechanism and Regulation of DNA-Protein Crosslink Repair by the DNA-Dependent Metalloprotease SPRTN

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

Stingele, Julian; Bellelli, Roberto; Alte, Ferdinand

Covalent DNA-protein crosslinks (DPCs) are toxic DNA lesions that interfere with essential chromatin transactions, such as replication and transcription. Little was known about DPC-specific repair mechanisms until the recent identification of a DPC-processing protease in yeast. The existence of a DPC protease in higher eukaryotes is inferred from data in Xenopus laevis egg extracts, but its identity remains elusive. Here we identify the metalloprotease SPRTN as the DPC protease acting in metazoans. Loss of SPRTN results in failure to repair DPCs and hypersensitivity to DPC-inducing agents. SPRTN accomplishes DPC processing through a unique DNA-induced protease activity, which is controlled bymore » several sophisticated regulatory mechanisms. Cellular, biochemical, and structural studies define a DNA switch triggering its protease activity, a ubiquitin switch controlling SPRTN chromatin accessibility, and regulatory autocatalytic cleavage. Our data also provide a molecular explanation on how SPRTN deficiency causes the premature aging and cancer predisposition disorder Ruijs-Aalfs syndrome.« less

Mechanism and Regulation of DNA-Protein Crosslink Repair by the DNA-Dependent Metalloprotease SPRTN

DOE PAGES

Stingele, Julian; Bellelli, Roberto; Alte, Ferdinand; ...

2016-10-27

Covalent DNA-protein crosslinks (DPCs) are toxic DNA lesions that interfere with essential chromatin transactions, such as replication and transcription. Little was known about DPC-specific repair mechanisms until the recent identification of a DPC-processing protease in yeast. The existence of a DPC protease in higher eukaryotes is inferred from data in Xenopus laevis egg extracts, but its identity remains elusive. Here we identify the metalloprotease SPRTN as the DPC protease acting in metazoans. Loss of SPRTN results in failure to repair DPCs and hypersensitivity to DPC-inducing agents. SPRTN accomplishes DPC processing through a unique DNA-induced protease activity, which is controlled bymore » several sophisticated regulatory mechanisms. Cellular, biochemical, and structural studies define a DNA switch triggering its protease activity, a ubiquitin switch controlling SPRTN chromatin accessibility, and regulatory autocatalytic cleavage. Our data also provide a molecular explanation on how SPRTN deficiency causes the premature aging and cancer predisposition disorder Ruijs-Aalfs syndrome.« less

Mammals divert endogenous genotoxic formaldehyde into one-carbon metabolism.

PubMed

Burgos-Barragan, Guillermo; Wit, Niek; Meiser, Johannes; Dingler, Felix A; Pietzke, Matthias; Mulderrig, Lee; Pontel, Lucas B; Rosado, Ivan V; Brewer, Thomas F; Cordell, Rebecca L; Monks, Paul S; Chang, Christopher J; Vazquez, Alexei; Patel, Ketan J

2017-08-31

The folate-driven one-carbon (1C) cycle is a fundamental metabolic hub in cells that enables the synthesis of nucleotides and amino acids and epigenetic modifications. This cycle might also release formaldehyde, a potent protein and DNA crosslinking agent that organisms produce in substantial quantities. Here we show that supplementation with tetrahydrofolate, the essential cofactor of this cycle, and other oxidation-prone folate derivatives kills human, mouse and chicken cells that cannot detoxify formaldehyde or that lack DNA crosslink repair. Notably, formaldehyde is generated from oxidative decomposition of the folate backbone. Furthermore, we find that formaldehyde detoxification in human cells generates formate, and thereby promotes nucleotide synthesis. This supply of 1C units is sufficient to sustain the growth of cells that are unable to use serine, which is the predominant source of 1C units. These findings identify an unexpected source of formaldehyde and, more generally, indicate that the detoxification of this ubiquitous endogenous genotoxin creates a benign 1C unit that can sustain essential metabolism.

Computational investigation of kinetics of cross-linking reactions in proteins: importance in structure prediction.

PubMed

Bandyopadhyay, Pradipta; Kuntz, Irwin D

2009-01-01

The determination of protein structure using distance constraints is a new and promising field of study. One implementation involves attaching residues of a protein using a cross-linking agent, followed by protease digestion, analysis of the resulting peptides by mass spectroscopy, and finally sequence threading to detect the protein folds. In the present work, we carry out computational modeling of the kinetics of cross-linking reactions in proteins using the master equation approach. The rate constants of the cross-linking reactions are estimated using the pKas and the solvent-accessible surface areas of the residues involved. This model is tested with fibroblast growth factor (FGF) and cytochrome C. It is consistent with the initial experimental rate data for individual lysine residues for cytochrome C. Our model captures all observed cross-links for FGF and almost 90% of the observed cross-links for cytochrome C, although it also predicts cross-links that were not observed experimentally (false positives). However, the analysis of the false positive results is complicated by the fact that experimental detection of cross-links can be difficult and may depend on specific experimental conditions such as pH, ionic strength. Receiver operator characteristic plots showed that our model does a good job in predicting the observed cross-links. Molecular dynamics simulations showed that for cytochrome C, in general, the two lysines come closer for the observed cross-links as compared to the false positive ones. For FGF, no such clear pattern exists. The kinetic model and MD simulation can be used to study proposed cross-linking protocols.

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

PubMed

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

2017-01-27

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

Bifunctional alkylating agent-mediated MGMT-DNA cross-linking and its proteolytic cleavage in 16HBE cells

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

Cheng, Jin; Ye, Feng; Dan, Guorong

Nitrogen mustard (NM), a bifunctional alkylating agent (BAA), contains two alkyl arms and can act as a cross-linking bridge between DNA and protein to form a DNA-protein cross-link (DPC). O{sup 6}-methylguanine–DNA methyltransferase (MGMT), a DNA repair enzyme for alkyl adducts removal, is found to enhance cell sensitivity to BAAs and to promote damage, possibly due to its stable covalent cross-linking with DNA mediated by BAAs. To investigate MGMT-DNA cross-link (mDPC) formation and its possible dual roles in NM exposure, human bronchial epithelial cell line 16HBE was subjected to different concentrations of HN2, a kind of NM, and we found mDPCmore » was induced by HN2 in a concentration-dependent manner, but the mRNA and total protein of MGMT were suppressed. As early as 1 h after HN2 treatment, high mDPC was achieved and the level maintained for up to 24 h. Quick total DPC (tDPC) and γ-H2AX accumulation were observed. To evaluate the effect of newly predicted protease DVC1 on DPC cleavage, we applied siRNA of MGMT and DVC1, MG132 (proteasome inhibitor), and NMS-873 (p97 inhibitor) and found that proteolysis plays a role. DVC1 was proven to be more important in the cleavage of mDPC than tDPC in a p97-dependent manner. HN2 exposure induced DVC1 upregulation, which was at least partially contributed to MGMT cleavage by proteolysis because HN2-induced mDPC level and DNA damage was closely related with DVC1 expression. Homologous recombination (HR) was also activated. Our findings demonstrated that MGMT might turn into a DNA damage promoter by forming DPC when exposed to HN2. Proteolysis, especially DVC1, plays a crucial role in mDPC repair. - Highlights: • Nitrogen mustard-induced MGMT-DNA cross-linking was detected in a living cell. • Concentration- and time-dependent manners of MGMT-DNA cross-linking were revealed. • Proteolysis played an important role in protein (MGMT)-DNA cross-linking repair. • DVC1 acts as a proteolytic enzyme in cross-linking repair in a p97-dependent manner.« less

Polymeric foams from cross-linkable poly-N-ary lenebenzimidazoles

NASA Technical Reports Server (NTRS)

Harrison, E. S.; Delano, C. B.; Riccitello, S. R. (Inventor)

1978-01-01

Foamed cross-linked poly-N-arylenebinzimidazoles are prepared by mixing an organic tetraamine and an ortho substituted aromatic dicarboxylic acid anhydride in the presence of a blowing agent, and then heating the prepolymer to a temperature sufficient to complete polymerization and foaming of the reactants. In another embodiment of the process, the reactants are heated to form a prepolymer. The prepolymer is then cured at higher temperatures to complete foaming and polymerization.

From nano- to macro-engineering of oxide-encapsulated-nanoparticles for harsh reactions: one-step organization via cross-linking molecules.

PubMed

Zhang, Qiaofei; Zhao, Guofeng; Zhang, Zhiqiang; Han, Lupeng; Fan, Songyu; Chai, Ruijuan; Li, Yakun; Liu, Ye; Huang, Jun; Lu, Yong

2016-09-29

A strategy of "macro-micro-nano" organization is reported for embedding oxide-encapsulated-nanoparticles onto monolithic substrates in one-step with the aid of molecularly defined cross-linking agents. Such catalysts, with enhanced heat/mass transfer and high permeability, are qualified for several harsh reaction processes such as CH 4 /VOC abatement, gas-phase hydrogenation of dimethyl oxalate and oxidative dehydrogenation of ethane.

Structural analysis of photosystem I polypeptides using chemical crosslinking

NASA Technical Reports Server (NTRS)

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

1994-01-01

Thylakoid membranes, obtained from leaves of 14 d soybean (Glycine max L. cv. Williams) plants, were treated with the chemical crosslinkers glutaraldehyde or 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) to investigate the structural organization of photosystem I. Polypeptides were resolved using lithium dodecyl sulfate polyacrylamide gel electrophoresis, and were identified by western blot analysis using a library of polyclonal antibodies specific for photosystem I subunits. An electrophoretic examination of crosslinked thylakoids revealed numerous crosslinked products, using either glutaraldehyde or EDC. However, only a few of these could be identified by western blot analysis using subunit-specific polyclonal antibodies. Several glutaraldehyde dependent crosslinked species were identified. A single band was identified minimally composed of PsaC and PsaD, documenting the close interaction between these two subunits. The most interesting aspect of these studies was a crosslinked species composed of the PsaB subunit observed following EDC treatment of thylakoids. This is either an internally crosslinked species, which will provide structural information concerning the topology of the complex PsaB protein, a linkage with a polypeptide for which we do not yet have an immunological probe, or a masking of epitopes by the EDC linkage at critical locations in the peptide which is linked to PsaB.

Atmospheric Pressure Non-Equilibrium Plasma as a Green Tool to Crosslink Gelatin Nanofibers

NASA Astrophysics Data System (ADS)

Liguori, Anna; Bigi, Adriana; Colombo, Vittorio; Focarete, Maria Letizia; Gherardi, Matteo; Gualandi, Chiara; Oleari, Maria Chiara; Panzavolta, Silvia

2016-12-01

Electrospun gelatin nanofibers attract great interest as a natural biomaterial for cartilage and tendon repair despite their high solubility in aqueous solution, which makes them also difficult to crosslink by means of chemical agents. In this work, we explore the efficiency of non-equilibrium atmospheric pressure plasma in stabilizing gelatin nanofibers. We demonstrate that plasma represents an innovative, easy and environmentally friendly approach to successfully crosslink gelatin electrospun mats directly in the solid state. Plasma treated gelatin mats display increased structural stability and excellent retention of fibrous morphology after immersion in aqueous solution. This method can be successfully applied to induce crosslinking both in pure gelatin and genipin-containing gelatin electrospun nanofibers, the latter requiring an even shorter plasma exposure time. A complete characterization of the crosslinked nanofibres, including mechanical properties, morphological observations, stability in physiological solution and structural modifications, has been carried out in order to get insights on the occurring reactions triggered by plasma.

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.

Calcium gluconate as cross-linker improves survival and shelf life of encapsulated and dried Metarhizium brunneum and Saccharomyces cerevisiae for the application as biological control agents.

PubMed

Humbert, Pascal; Przyklenk, Michael; Vemmer, Marina; Patel, Anant V

2017-02-01

Calcium chloride (CC) is the most common cross-linker for the encapsulation of biocontrol microorganisms in alginate beads. The aim of this study was to evaluate if calcium gluconate (CG) can replace CC as cross-linker and at the same time improve viability after drying and rehydration, hygroscopic properties, shelf life and nutrient supply. Hence, the biocontrol fungi Metarhizium brunneum and Saccharomyces cerevisiae were encapsulated in Ca-alginate beads supplemented with starch. Beads were dried and maximum survival was found in beads cross-linked with CG. Beads prepared with CG showed lower hygroscopic properties, but a higher shelf life for encapsulated fungi. Moreover, we demonstrated that gluconate has a nutritive effect on encapsulated fungi, leading to increased mycelium growth of M. brunneum and to enhanced CO 2 release from beads containing Saccharomyces cerevisiae. The application of CG as cross-linker will pave the way towards increasing drying survival and shelf life of various, especially drying-sensitive microbes.

Dual Sulfide-Disulfide Crosslinked Networks with Rapid and Room Temperature Self-Healability.

PubMed

An, So Young; Noh, Seung Man; Nam, Joon Hyun; Oh, Jung Kwon

2015-07-01

Polymer-based crosslinked networks with intrinsic self-repairing ability have emerged due to their built-in ability to repair physical damages. Here, novel dual sulfide-disulfide crosslinked networks (s-ssPxNs) are reported exhibiting rapid and room temperature self-healability within seconds to minutes, with no extra healing agents and no change under any environmental conditions. The method to synthesize these self-healable networks utilizes a combination of well-known crosslinking chemistry: photoinduced thiol-ene click-type radical addition, generating lightly sulfide-crosslinked polysulfide-based networks with excess thiols, and their oxidation, creating dynamic disulfide crosslinkages to yield the dual s-ssPxNs. The resulting s-ssPxN networks show rapid self-healing within 30 s to 30 min at room temperature, as well as self-healing elasticity with reversible viscoelastic properties. These results, combined with tunable self-healing kinetics, demonstrate the versatility of the method as a new means to synthesize smart multifunctional polymeric materials. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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

PubMed

Bragg, P D; Hou, C

1980-05-01

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

[Fanconi anemia: genes and function(s) revisited].

PubMed

Papadopoulo, Dora; Moustacchi, Ethel

2005-01-01

Fanconi anemia (FA), a rare inherited disorder, exhibits a complex phenotype including progressive bone marrow failure, congenital malformations and increased risk of cancers, mainly acute myeloid leukaemia. At the cellular level, FA is characterized by hypersensitivity to DNA cross-linking agents and by high frequencies of induced chromosomal aberrations, a property used for diagnosis. FA results from mutations in one of the eleven FANC (FANCA to FANCJ) genes. Nine of them have been identified. In addition, FANCD1 gene has been shown to be identical to BRCA2, one of the two breast cancer susceptibility genes. Seven of the FANC proteins form a complex, which exists in four different forms depending of its subcellular localisation. Four FANC proteins (D1(BRCA2), D2, I and J) are not associated to the complex. The presence of the nuclear form of the FA core complex is necessary for the mono-ubiquitinylation of FANCD2 protein, a modification required for its re-localization to nuclear foci, likely to be sites of DNA repair. A clue towards understanding the molecular function of the FANC genes comes from the recently identified connection of FANC to the BRCA1, ATM, NBS1 and ATR genes. Two of the FANC proteins (A and D2) directly interact with BRCA1, which in turn interacts with the MRE11/RAD50/NBS1 complex, which is one of the key components in the mechanisms involved in the cellular response to DNA double strand breaks (DSB). Moreover, ATM, a protein kinase that plays a central role in the network of DSB signalling, phosphorylates in vitro and in vivo FANCD2 in response to ionising radiations. Moreover, the NBS1 protein and the monoubiquitinated form of FANCD2 seem to act together in response to DNA crosslinking agents. Taken together with the previously reported impaired DSB and DNA interstrand crosslinks repair in FA cells, the connection of FANC genes to the ATM, ATR, NBS1 and BRCA1 links the FANC genes function to the finely orchestrated network involved in the sensing, signalling and repair of DNA replication-blocking lesions.

A method for the production of weakly acidic cation exchange resins

NASA Astrophysics Data System (ADS)

Heller, H.; Werner, F.; Mitschker, A.; Diehl, H. V.; Schaefer, A.

1991-12-01

The invention relates to a nonpolluting method for the production of weakly acidic cation exchange resins by saponification of cross-linked acrylonitrile bead polymers, with an alkaline saponification agent at elevated temperature, according to which method the bead polymer and alkaline saponification agent are jointly added only at elevated temperature.

Photo-triggered fluorescent theranostic prodrugs as DNA alkylating agents for mechlorethamine release and spatiotemporal monitoring.

PubMed

Cao, Yanting; Pan, Rong; Xuan, Weimin; Wei, Yongyi; Liu, Kejian; Zhou, Jiahong; Wang, Wei

2015-06-28

We describe a new theranostic strategy for selective delivery and spatiotemporal monitoring of mechlorethamine, a DNA alkylating agent. A photo-responsive prodrug is designed and composed of a photolabile o-nitrophenylethyl group, a DNA alkylating mechlorethamine drug and a coumarin fluorophore. Masking of the "N" in mechlorethamine in a positively charged state in the prodrug renders it inactive, non-toxic, selective and non-fluorescent. Indeed, the stable prodrug shows negligible cytotoxicity towards normal cells with and without UV activation and is completely non-fluorescent. However, upon photo-irradiation, the active mechlorethamine is released and induces efficient DNA cross-links, accompanied by a strong fluorescence enhancement (152 fold). Furthermore, DNA cross-linking activity from the release can be transformed into anticancer activity observed in in vitro studies of tumor cells. Importantly, the drug release progress and the movement can be conveniently monitored by fluorescence spectroscopy. The mechanistic study proves that the DNA cross-linking activity is mainly due to the release of DNA alkylating mechlorethamine. Altogether, the studies show the power of the theranostic strategy for efficient therapy in cancer treatment.

Enantiomers Recognition of Propranolol Based on Organic-Inorganic Hybrid Open-Tubular MIPs-CEC Column Using 3-(Trimethoxysilyl)Propyl Methacrylate as a Cross-Linking Monomer.

PubMed

Chen, Guo-Ning; Li, Ning; Luo, Tian; Dong, Yu-Ming

2017-04-01

In this study, 3-(trimethoxysilyl)propyl methacrylate (γ-MPS), a bifunctional group compound, was used as a single cross-linking agent to prepare molecular imprinted inorganic-organic hybrid polymers by in situ polymerization for open-tubular capillary electro chromatography (CEC) column. The optimal preparation conditions were: the ratio between template molecule and functional monomer was 1:4; the volume proportion of porogen toluene and methanol was 1:1 and the volume of cross-linking agent γ-MPS was 69 μL. The optimal separation conditions were separation voltage of 15 kV; detection wavelength at 215 nm and background electrolyte composed of 70% acetonitrile/20 mmol/L boric acid salt (pH 6.9). Under the optimized conditions, the propranolol enantiomers can be separated well by CEC. The method is simple and fast, it can be a potentially useful approach for propranolol enantiomers separation. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

21 CFR 177.1211 - Cross-linked polyacrylate copolymers.

Code of Federal Regulations, 2011 CFR

2011-04-01

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

21 CFR 177.1211 - Cross-linked polyacrylate copolymers.

Code of Federal Regulations, 2010 CFR

2010-04-01

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

Cellulose-silica/gold nanomaterials for electronic applications.

PubMed

Kim, Gwang-Hoon; Ramesh, Sivalingam; Kim, Joo-Hyung; Jung, Dongsoo; Kim, Heung Soo

2014-10-01

Cellulose and one dimensional nano-material composite has been investigated for various industrial applications due to their optical, mechanical and electrical properties. In present investigation, cellulose/silica and silica-gold hybrid biomaterials were prepared by sol-gel covalent cross-linking process. The tetraethoxysiliane (TEOS) and gold precursors and γ-aminopropyltriethoxysilane (γ-APTES) as coupling agent were used for sol-gel cross-linking process. The chemical and morphological properties of cellulose/silica and cellulose/silica-gold nano-materials via covalent cross-linking hybrids were confirmed by FTIR, XRD, SEM, and TEM analysis. In the sol-gel process, the inorganic particles were dispersed in the cellulose host matrix at the nanometer scale, bonding to the cellulose through the covalent bonds.

Durable low surface-energy surfaces

NASA Technical Reports Server (NTRS)

Willis, Paul B. (Inventor); McElroy, Paul M. (Inventor); Hickey, Gregory H. (Inventor)

1993-01-01

A formulation for forming a low surface-energy surface on a substrate having (i) a fluoroalkyl silane having a low surface energy part, (ii) a liquid crystal silane operable for enhancing the orientation of the molecules of the fluoroalkyl silane and for crosslinking with the fluoroalkyl silane, and, (iii) a transport medium for applying the fluoroalkyl silane and the liquid crystal silane to the surface of a substrate. In one embodiment the formulation can includes a crosslinking agent for crosslinking the fluoroalkyl silane. In another embodiment the formulation has a condensation catalyst for enhancing chemical bonding of the fluoroalkyl silane to the substrate. The transport medium can be an alcohol such as methanol or ethanol.

FAN1 acts with FANCI-FANCD2 to promote DNA interstrand cross-link repair.

PubMed

Liu, Ting; Ghosal, Gargi; Yuan, Jingsong; Chen, Junjie; Huang, Jun

2010-08-06

Fanconi anemia (FA) is caused by mutations in 13 Fanc genes and renders cells hypersensitive to DNA interstrand cross-linking (ICL) agents. A central event in the FA pathway is mono-ubiquitylation of the FANCI-FANCD2 (ID) protein complex. Here, we characterize a previously unrecognized nuclease, Fanconi anemia-associated nuclease 1 (FAN1), that promotes ICL repair in a manner strictly dependent on its ability to accumulate at or near sites of DNA damage and that relies on mono-ubiquitylation of the ID complex. Thus, the mono-ubiquitylated ID complex recruits the downstream repair protein FAN1 and facilitates the repair of DNA interstrand cross-links.

Application of conjugated heparin-albumin microparticles with laser-balloon angioplasty: a potential method for reducing adverse biologic reactivity after angioplasty

NASA Astrophysics Data System (ADS)

Kundu, Sourav K.; McMath, Linda P.; Zaidan, Jonathan T.; Spears, J. Richard

1991-05-01

Laser-balloon angioplasty (LBA) may potentially be used for local application of pharmacologically active agents which will reduce thrombogenic and proliferative responses after the angioplasty. In this study, the feasibility of applying covalently conjugated heparin- albumin microparticles onto arterial luminal surface was demonstrated. The covalent linkages were formed by reaction with 1-ethyl-3-dimethyl-aminopropyl-carbodiimide (EDC), and the resultant conjugates were used for preparation of microparticles by employing standard emulsification and heat-crosslinking techniques. The heparin release rate from the microparticles was found to be dependent upon the degree of crosslinking. When a thin coagulum of a suspension of microparticles was formed with heat on a glass surface, the treated surface demonstrated resistance to clot formation in contact with non-anticoagulated blood. A suspension of the microparticles applied during laser-balloon angioplasty onto the luminal surface of dog carotid and femoral arteries showed persistence for up to one week without thrombus formation or occlusion of the vessel. Since the rate of biodegradation is primarily dictated by the extent of crosslinking, an optimal degree of thermal denaturation will permit longer persistence of the carrier while allowing adequate release of the entrapped pharmacologic agent. A variety of antithrombotic and antiinflammatory agents are being considered as candidate bioprotective materials for local application after angioplasty.

Synthesis and evaluation of acryloylated starch-g-poly (Acrylamide/Vinylmethacrylate/1-Vinyl-2-pyrrolidone) crosslinked terpolymer functionalized by dimethylphenylvinylsilane derivative as a novel polymer-flooding agent.

PubMed

El-Hoshoudy, A N; Desouky, S M

2018-05-16

Starch is a natural biopolymer that subjected to various chemical modifications through different industrial applications. Molecular structure of starch allow its grafting with different vinyl monomers in the presence of crosslinking agents to synthesize cross-linked hydrogels, which used in enhanced oil recovery (EOR) applications, water shutoff and drag reduction. Application of native starch in the field of petroleum reservoirs as a flooding agent suffer from some limitations concerned with microbial degradation, thermal and salinity resistance under harsh petroleum reservoir conditions. In the current research, we stated the synthesis of acryloylated starch then it's grafting with poly (Acrylamide/Vinylmethacrylate/1-Vinyl-2-pyrrolidone) terpolymer in presence of dimethylphenylvinylsilane through emulsification polymerization. Characterization and structure determination occur by different spectroscopic techniques as stated throughout the manuscript. Rheological and solution properties carried out as a function of shear rate, salinity and temperature at simulated reservoir conditions. Flooding tests carried out through linear-dimensional sandstone model at simulated reservoir conditions, and recovered oil amount calculated on volumetric basis. The obtained results indicate that the prepared starch-g-terpolymer can tolerate to severe flooding conditions of high temperature and salinity, moreover it can increase recovery factor up to 49% of residual oil saturation so considered as a promised EOR candidate. Copyright © 2017 Elsevier B.V. All rights reserved.

Crosslinked anion exchange membranes prepared from poly(phenylene oxide) (PPO) for non-aqueous redox flow batteries

NASA Astrophysics Data System (ADS)

Li, Yun; Sniekers, Jeroen; Malaquias, João C.; Van Goethem, Cedric; Binnemans, Koen; Fransaer, Jan; Vankelecom, Ivo F. J.

2018-02-01

A stable and eco-friendly anion-exchange membrane (AEM) was prepared and applied in a non-aqueous all-copper redox flow battery (RFB). The AEM was prepared via a simple procedure, leading to a cross-linked structure containing quaternary ammonium groups without involvement of harmful trimethylamine. A network was thus constructed which ensured both ion transport and solvent resistance. The ion exchange capacity (IEC) of the membrane was tuned from 0.49 to 1.03 meq g-1 by varying the content of the 4, 4‧-bipyridine crosslinking agent. The membrane showed a good anion conductivity and retention of copper ions. As a proof of principle, a RFB single cell with this crosslinked membrane yielded a coulombic efficiency of 89%, a voltage efficiency of 61% and an energy efficiency of 54% at 7.5 mA cm-2.

Effects of Chemical Cross-linkers on Caries-affected Dentin Bonding

PubMed Central

Macedo, G.V.; Yamauchi, M.; Bedran-Russo, A.K.

2009-01-01

The achievement of a strong and stable bond between composite resin and dentin remains a challenge in restorative dentistry. Over the past two decades, dental materials have been substantially improved, with better handling and bonding characteristics. However, little attention has been paid to the contribution of collagen structure/stability to bond strength. We hypothesized that the induction of cross-linking in dentin collagen improves dentin collagen stability and bond strength. This study investigated the effects of glutaraldehyde- and grape seed extract-induced cross-linking on the dentin bond strengths of sound and caries-affected dentin, and on the stability of dentin collagen. Our results demonstrated that the application of chemical cross-linking agents to etched dentin prior to bonding procedures significantly enhanced the dentin bond strengths of caries-affected and sound dentin. Glutaraldehyde and grape seed extract significantly increased dentin collagen stability in sound and caries-affected dentin, likely via distinct mechanisms. PMID:19892915

Polyvinyl alcohol cross-linked with two aldehydes

NASA Technical Reports Server (NTRS)

Sheibley, D. W.; Rieker, L. L.; Hsu, L. C.; Manzo, M. A. (Inventor)

1982-01-01

A film forming polyvinyl alcohol resin is admixed, in aqueous solution, with a dialdehyde crosslinking agent which is capable of crosslinking the polyvinyl alcohol resin and a water soluble acid aldehyde containing a reactive aldehyde group capable of reacting with hydroxyl groups in the polyvinyl alcohol resin and an ionizable acid hydrogen atom. The dialdehyde is present in an amount sufficient to react with from 1 to 20% by weight of the theoretical amount required to react with all of the hydroxyl groups of the polyvinyl alcohol. The amount of acid aldehyde is from 1 to 50% by weight, same basis, and is sufficient to reduce the pH of the aqueous admixture to 5 or less. The admixture is then formed into a desired physical shape, such as by casting a sheet or film, and the shaped material is then heated to simultaneously dry and crosslink the article.

Influence of crosslinking on the mechanical behavior of 3D printed alginate scaffolds: Experimental and numerical approaches.

PubMed

Naghieh, Saman; Karamooz-Ravari, Mohammad Reza; Sarker, M D; Karki, Eva; Chen, Xiongbiao

2018-04-01

Tissue scaffolds fabricated by three-dimensional (3D) bioprinting are attracting considerable attention for tissue engineering applications. Because the mechanical properties of hydrogel scaffolds should match the damaged tissue, changing various parameters during 3D bioprinting has been studied to manipulate the mechanical behavior of the resulting scaffolds. Crosslinking scaffolds using a cation solution (such as CaCl 2 ) is also important for regulating the mechanical properties, but has not been well documented in the literature. Here, the effect of varied crosslinking agent volume and crosslinking time on the mechanical behavior of 3D bioplotted alginate scaffolds was evaluated using both experimental and numerical methods. Compression tests were used to measure the elastic modulus of each scaffold, then a finite element model was developed and a power model used to predict scaffold mechanical behavior. Results showed that crosslinking time and volume of crosslinker both play a decisive role in modulating the mechanical properties of 3D bioplotted scaffolds. Because mechanical properties of scaffolds can affect cell response, the findings of this study can be implemented to modulate the elastic modulus of scaffolds according to the intended application. Copyright © 2018 Elsevier Ltd. All rights reserved.

Use of 2,5-dimethyl-2,5-hexane diamine as a curing agent for epoxy resins. [Patent application

DOEpatents

Rinde, J.A.; Newey, H.A.

Primary diamines are prepared for use as a curing agent for epoxy resins. These curing agents can be used to form epoxy resin mixtures useful in filament winding and preimpregnated fiber molding and in formulating film adhesives, powder coatings and molding powders. The epoxy mixtures form for such uses a room temperature non-reacting, intermediate stable state which has a latent cross-linking capability.

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

PubMed

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

2015-11-01

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

Improving performance of nanoscale ultrasound contrast agents using N,N-diethylacrylamide stabilization.

PubMed

Perera, Reshani H; Wu, Hanping; Peiris, Pubudu; Hernandez, Christopher; Burke, Alan; Zhang, Helen; Exner, Agata A

2017-01-01

The design of nanoscale yet highly echogenic agents for imaging outside of the vasculature and for ultrasound-mediated drug delivery remains a formidable challenge. We have previously reported on formulation of echogenic perfluoropropane gas nanobubbles stabilized by a lipid-pluronic surfactant shell. In the current work we describe the development of a new generation of these nanoparticles which consist of perfluoropropane gas stabilized by a surfactant and lipid membrane and a crosslinked network of N,N-diethylacrylamide. The resulting crosslinked nanobubbles (CL-PEG-NB) were 95.2±25.2nm in diameter and showed significant improvement in stability and retention of echogenic signal over 24h. In vivo analysis via ultrasound and fluorescence mediated tomography showed greater tumor extravasation and accumulation with CL-PEG-NB compared to microbubbles. Together these results demonstrate the capabilities and advantages of a new, more stable, nanometer-scale ultrasound contrast agent that can be utilized in future work for diagnostic scans and molecular imaging. Copyright © 2016 Elsevier Inc. All rights reserved.

Covalent immobilization of lipase onto chitosan-mesoporous silica hybrid nanomaterials by carboxyl functionalized ionic liquids as the coupling agent.

PubMed

Xiang, Xinran; Suo, Hongbo; Xu, Chao; Hu, Yi

2018-05-01

Chitosan-mesoporous silica SBA-15 hybrid nanomaterials (CTS-SBA-15) were synthesized by means of carboxyl functionalized ionic liquids as the coupling agent. The as-prepared CTS-SBA-15 support was characterized by TEM, FTIR, TG and nitrogen adsorption-desorption techniques. Porcine pancreas lipase (PPL) was then bound to the hybrid nanomaterials by using the cross-linking reagent glutaraldehyde (GA). Further, the parameters like cross-linking concentration, time and ratio of supports to enzyme were optimized. The property of immobilized lipase were tested in detail by enzyme activity assays. The results indicated that the hybrid nanomaterials could form three-dimensional (3D) structure with homogeneous mesoporous structures and immobilized PPL revealed excellent enzymatic performance. Copyright © 2018 Elsevier B.V. All rights reserved.

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.

Effect of Instrument Lubricants on the Surface Degree of Conversion and Crosslinking Density of Nanocomposites.

PubMed

de Paula, Felipe Costa; Valentin, Regis de Souza; Borges, Boniek Castillo Dutra; Medeiros, Maria Cristina Dos Santos; de Oliveira, Raiza Freitas; da Silva, Ademir Oliveira

2016-01-01

The surface degree of conversion and crosslink density of composites should not be affected by the use of instrument lubricants in order to provide long-lasting tooth restorations. This study aimed to analyze the effect of instrument lubricants on the degree of conversion and crosslink density of nanocomposites. Samples (N = 10) were fabricated according to the composites (Filtek Z350 XT, 3M ESPE, St. Paul, MN, USA; and IPS Empress Direct, Ivoclar Vivadent AG, Schaan, Liechtenstein and lubricants used (Adper Single Bond 2 and Scotchbond Multi-Purpose bonding agent adhesive systems, 3M ESPE; 70% ethanol, absolute ethanol, and no lubricant). Single composite increments were inserted into a Teflon mold using the same dental instrument. The composite surface was then modeled using a brush wiped with each adhesive system and a spatula wiped with each ethanol. The control group was fabricated with no additional modeling. The surface degree of conversion and crosslink density were measured by Fourier transform infrared spectroscopy and the hardness decrease test, respectively. Data were analyzed using two-way analysis of variance and the Tukey's test (p < 0.05). Filtek Z350 XT showed statistically similar degree of conversion regardless of the lubricant used, whereas the use of adhesive systems and 70% ethanol decreased the degree of conversion for IPS Empress Direct. Only Scotchbond Multi-Purpose bonding agent decreased crosslink density for Filtek Z350 XT, whereas both adhesive systems decreased crosslink density for IPS Empress Direct. Filtek Z350 XT appeared to be less sensitive to the effects of lubricants, and absolute ethanol did not affect the degree of conversion and crosslink density of the nanocomposites tested. Although the use of lubricants may be recommended to minimize the stickiness of dental instruments and composite resin, dentists should choose materials that do not have a negative effect on the surface properties of composites. Only the use of absolute ethanol safely maintains the surface integrity of nanocomposites in comparison with adhesive system and 70% ethanol. © 2015 Wiley Periodicals, Inc.

Spontaneous translocation of antitumor oxaliplatin, its enantiomeric analogue, and Cisplatin from one strand to another in double-helical DNA.

PubMed

Malina, Jaroslav; Natile, Giovanni; Brabec, Viktor

2013-09-02

Oxaliplatin and cisplatin belong to the class of platinum-based anticancer agents. Formation of DNA adducts by these complexes and the consequences for its structure and function, is the mechanistic paradigm by which these drugs exert their antitumor activity. We show that employing short oligonucleotide duplexes containing single, site-specific 1,3-intrastrand cross-links of oxaliplatin, its enantiomeric analogue, or cisplatin and by using gel electrophoresis that under physiological conditions the coordination bonds between platinum and the N7 position of guanine residues involved in the cross-links of the Pt(II) complexes can be cleaved. This cleavage may lead to linkage isomerization reactions between these metallodrugs and double-helical DNA. For instance, approximately 25 % 1,3-intrastrand cross-links of the platinum complexes isomerized after 192 h (at 310 K in 200 mM NaClO4). Differential scanning calorimetry of duplexes containing single, site-specific cross-links of oxaliplatin, its enantiomeric analogue, and cisplatin reveals that one of the driving forces that leads to the lability of DNA cross-links of these metallodrugs is a difference between the thermodynamic destabilization induced by the cross-link and by the adduct into which it could isomerize. The rearrangements may proceed in the way that cross-links originally formed in one strand of the DNA can spontaneously translocate from one DNA strand to its complementary counterpart, which may evoke walking of the platinum complex on DNA molecule. In addition, the differences in the kinetics of the rearrangement reactions and the thermodynamic destabilization of DNA observed for adducts of oxaliplatin and its enantiomeric analogue confirm that the chirality at the carrier 1,2-diaminocyclohexane ligand can considerably affect structural and other physical properties of DNA adducts and consequently their biological effects. In aggregate, interesting generalization of the results described in this work might be that the migration of oxaliplatin, its enantiomeric analogue, or cisplatin from one strand to another in double-helical DNA controlled by energetic signatures of these agents might contribute to a better understanding of their cytotoxic and mutagenic potential. Copyright © 2013 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim.

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

NASA Astrophysics Data System (ADS)

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

2018-02-01

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

Microfluidic production of bioactive fibrin micro-beads embedded in crosslinked collagen used as an injectable bulking agent for urinary incontinence treatment.

PubMed

Vardar, E; Larsson, H M; Allazetta, S; Engelhardt, E M; Pinnagoda, K; Vythilingam, G; Hubbell, J A; Lutolf, M P; Frey, P

2018-02-01

Endoscopic injection of bulking agents has been widely used to treat urinary incontinence, often due to urethral sphincter complex insufficiency. The aim of the study was to develop a novel injectable bioactive collagen-fibrin bulking agent restoring long-term continence by functional muscle tissue regeneration. Fibrin micro-beads were engineered using a droplet microfluidic system. They had an average diameter of 140 μm and recombinant fibrin-binding insulin-like growth factor-1 (α 2 PI 1-8 -MMP-IGF-1) was covalently conjugated to the beads. A plasmin fibrin degradation assay showed that 72.5% of the initial amount of α 2 PI 1-8 -MMP-IGF-1 loaded into the micro-beads was retained within the fibrin micro-beads. In vitro, the growth factor modified fibrin micro-beads enhanced cell attachment and the migration of human urinary tract smooth muscle cells, however, no change of the cellular metabolic activity was seen. These bioactive micro-beads were mixed with genipin-crosslinked homogenized collagen, acting as a carrier. The collagen concentration, the degree of crosslinking, and the mechanical behavior of this bioactive collagen-fibrin injectable were comparable to reference samples. This novel injectable showed no burst release of the growth factor, had a positive effect on cell behavior and may therefore induce smooth muscle regeneration in vivo, necessary for the functional treatment of stress and other urinary incontinences. Urinary incontinence is involuntary urine leakage, resulting from a deficient function of the sphincter muscle complex. Yet there is no functional cure for this devastating condition using current treatment options. Applied physical and surgical therapies have limited success. In this study, a novel bioactive injectable bulking agent, triggering new muscle regeneration at the injection site, has been evaluated. This injectable consists of cross-linked collagen and fibrin micro-beads, functionalized with bound insulin-like growth factor-1 (α 2 PI 1-8 -MMP-IGF-1). These bioactive fibrin micro-beads induced human smooth muscle cell migration in vitro. Thus, this injectable bulking agent is apt to be a good candidate for regeneration of urethral sphincter muscle, ensuring a long-lasting treatment for urinary incontinence. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Treatment with eldecalcitol positively affects mineralization, microdamage, and collagen crosslinks in primate bone.

PubMed

Saito, Mitsuru; Grynpas, Marc D; Burr, David B; Allen, Matthew R; Smith, Susan Y; Doyle, Nancy; Amizuka, Norio; Hasegawa, Tomoka; Kida, Yoshikuni; Marumo, Keishi; Saito, Hitoshi

2015-04-01

Eldecalcitol (ELD), an active form of vitamin D analog approved for the treatment of osteoporosis in Japan, increases lumbar spine bone mineral density (BMD), suppresses bone turnover markers, and reduces fracture risk in patients with osteoporosis. We have previously reported that treatment with ELD for 6 months improved the mechanical properties of the lumbar spine in ovariectomized (OVX) cynomolgus monkeys. ELD treatment increased lumbar BMD, suppressed bone turnover markers, and reduced histomorphometric parameters of both bone formation and resorption in vertebral trabecular bone. In this study, we elucidated the effects of ELD on bone quality (namely, mineralization, microarchitecture, microdamage, and bone collagen crosslinks) in OVX cynomolgus monkeys in comparison with OVX-vehicle control monkeys. Density fractionation of bone powder prepared from lumbar vertebrae revealed that ELD treatment shifted the distribution profile of bone mineralization to a higher density, and backscattered electron microscopic imaging showed improved trabecular bone connectivity in the ELD-treated groups. Higher doses of ELD more significantly reduced the amount of microdamage compared to OVX-vehicle controls. The fractionated bone powder samples were divided according to their density, and analyzed for collagen crosslinks. Enzymatic crosslinks were higher in both the high-density (≥2.0 mg/mL) and low-density (<2.0 mg/mL) fractions from the ELD-treated groups than in the corresponding fractions in the OVX-vehicle control groups. On the other hand, non-enzymatic crosslinks were lower in both the high- and low-density fractions. These observations indicated that ELD treatment stimulated the enzymatic reaction of collagen crosslinks and bone mineralization, but prevented non-enzymatic reaction of collagen crosslinks and accumulation of bone microdamage. Bone anti-resorptive agents such as bisphosphonates slow down bone remodeling so that bone mineralization, bone microdamage, and non-enzymatic collagen crosslinks all increase. Bone anabolic agents such as parathyroid hormone decrease bone mineralization and bone microdamage by stimulating bone remodeling. ELD did not fit into either category. Histological analysis indicated that the ELD treatment strongly suppressed bone resorption by reducing the number of osteoclasts, while also stimulating focal bone formation without prior bone resorption (bone minimodeling). These bidirectional activities of ELD may account for its unique effects on bone quality. Copyright © 2014. Published by Elsevier Inc.

Small, porous polyacrylate beads

NASA Technical Reports Server (NTRS)

Yen, Shiao-Ping Siao (Inventor); Rembaum, Alan (Inventor); Dreyer, William J. (Inventor)

1976-01-01

Uniformly-shaped, porous, round beads are prepared by the co-polymerization of an acrylic monomer and a cross-linking agent in the presence of 0.05 to 5% by weight of an aqueous soluble polymer such as polyethylene oxide. Cross-linking proceeds at high temperature above about 50.degree.C or at a lower temperature with irradiation. Beads of even shape and even size distribution of less than 2 micron diameter are formed. The beads will find use as adsorbents in chromatography and as markers for studies of cell surface receptors.

3D freeform printing of silk fibroin.

PubMed

Rodriguez, Maria J; Dixon, Thomas A; Cohen, Eliad; Huang, Wenwen; Omenetto, Fiorenzo G; Kaplan, David L

2018-04-15

Freeform fabrication has emerged as a key direction in printing biologically-relevant materials and structures. With this emerging technology, complex structures with microscale resolution can be created in arbitrary geometries and without the limitations found in traditional bottom-up or top-down additive manufacturing methods. Recent advances in freeform printing have used the physical properties of microparticle-based granular gels as a medium for the submerged extrusion of bioinks. However, most of these techniques require post-processing or crosslinking for the removal of the printed structures (Miller et al., 2015; Jin et al., 2016) [1,2]. In this communication, we introduce a novel method for the one-step gelation of silk fibroin within a suspension of synthetic nanoclay (Laponite) and polyethylene glycol (PEG). Silk fibroin has been used as a biopolymer for bioprinting in several contexts, but chemical or enzymatic additives or bulking agents are needed to stabilize 3D structures. Our method requires no post-processing of printed structures and allows for in situ physical crosslinking of pure aqueous silk fibroin into arbitrary geometries produced through freeform 3D printing. 3D bioprinting has emerged as a technology that can produce biologically relevant structures in defined geometries with microscale resolution. Techniques for fabrication of free-standing structures by printing into granular gel media has been demonstrated previously, however, these methods require crosslinking agents and post-processing steps on printed structures. Our method utilizes one-step gelation of silk fibroin within a suspension of synthetic nanoclay (Laponite), with no need for additional crosslinking compounds or post processing of the material. This new method allows for in situ physical crosslinking of pure aqueous silk fibroin into defined geometries produced through freeform 3D printing. Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Preparation and Optimisation of Cross-Linked Enzyme Aggregates Using Native Isolate White Rot Fungi Trametes versicolor and Fomes fomentarius for the Decolourisation of Synthetic Dyes

PubMed Central

Vršanská, Martina; Voběrková, Stanislava; Jiménez Jiménez, Ana María; Strmiska, Vladislav; Adam, Vojtěch

2017-01-01

The key to obtaining an optimum performance of an enzyme is often a question of devising a suitable enzyme and optimisation of conditions for its immobilization. In this study, laccases from the native isolates of white rot fungi Fomes fomentarius and/or Trametes versicolor, obtained from Czech forests, were used. From these, cross-linked enzyme aggregates (CLEA) were prepared and characterised when the experimental conditions were optimized. Based on the optimization steps, saturated ammonium sulphate solution (75 wt.%) was used as the precipitating agent, and different concentrations of glutaraldehyde as a cross-linking agent were investigated. CLEA aggregates formed under the optimal conditions showed higher catalytic efficiency and stabilities (thermal, pH, and storage, against denaturation) as well as high reusability compared to free laccase for both fungal strains. The best concentration of glutaraldehyde seemed to be 50 mM and higher efficiency of cross-linking was observed at a low temperature 4 °C. An insignificant increase in optimum pH for CLEA laccases with respect to free laccases for both fungi was observed. The results show that the optimum temperature for both free laccase and CLEA laccase was 35 °C for T. versicolor and 30 °C for F. fomentarius. The CLEAs retained 80% of their initial activity for Trametes and 74% for Fomes after 70 days of cultivation. Prepared cross-linked enzyme aggregates were also investigated for their decolourisation activity on malachite green, bromothymol blue, and methyl red dyes. Immobilised CLEA laccase from Trametes versicolor showed 95% decolourisation potential and CLEA from Fomes fomentarius demonstrated 90% decolourisation efficiency within 10 h for all dyes used. These results suggest that these CLEAs have promising potential in dye decolourisation. PMID:29295505

BLM and the FANC proteins collaborate in a common pathway in response to stalled replication forks

PubMed Central

Pichierri, Pietro; Franchitto, Annapaola; Rosselli, Filippo

2004-01-01

Fanconi anaemia (FA) and Bloom syndrome (BS) are autosomal recessive diseases characterised by chromosome fragility and cancer proneness. Here, we report that BLM and the FA pathway are activated in response to both crosslinked DNA and replication fork stall. We provide evidence that BLM and FANCD2 colocalise and co-immunoprecipitate following treatment with either DNA crosslinkers or agents inducing replication arrest. We also find that the FA core complex is necessary for BLM phosphorylation and assembly in nuclear foci in response to crosslinked DNA. Moreover, we show that knock-down of the MRE11 complex, whose function is also under the control of the FA core complex, enhances cellular and chromosomal sensitivity to DNA interstrand crosslinks in BS cells. These findings suggest the existence of a functional link between BLM and the FA pathway and that BLM and the MRE11 complex are in two separated branches of a pathway resulting in S-phase checkpoint activation, chromosome integrity and cell survival in response to crosslinked DNA. PMID:15257300

Shell crosslinked nanoparticles carrying silver antimicrobials as therapeutics†

PubMed Central

Li, Yali; Hindi, Khadijah; Watts, Kristin M.; Taylor, Jane B.; Zhang, Ke; Li, Zicheng

2010-01-01

Amphiphilic polymer nanoparticles loaded with silver cations or/and N-heterocyclic carbene–silver complexes were assessed as antimicrobial agents against Gram-negative pathogens Escherichia coli and Pseudomonas aeruginosa. PMID:20024313

In-Culture Cross-Linking of Bacterial Cells Reveals Large-Scale Dynamic Protein-Protein Interactions at the Peptide Level.

PubMed

de Jong, Luitzen; de Koning, Edward A; Roseboom, Winfried; Buncherd, Hansuk; Wanner, Martin J; Dapic, Irena; Jansen, Petra J; van Maarseveen, Jan H; Corthals, Garry L; Lewis, Peter J; Hamoen, Leendert W; de Koster, Chris G

2017-07-07

Identification of dynamic protein-protein interactions at the peptide level on a proteomic scale is a challenging approach that is still in its infancy. We have developed a system to cross-link cells directly in culture with the special lysine cross-linker bis(succinimidyl)-3-azidomethyl-glutarate (BAMG). We used the Gram-positive model bacterium Bacillus subtilis as an exemplar system. Within 5 min extensive intracellular cross-linking was detected, while intracellular cross-linking in a Gram-negative species, Escherichia coli, was still undetectable after 30 min, in agreement with the low permeability in this organism for lipophilic compounds like BAMG. We were able to identify 82 unique interprotein cross-linked peptides with <1% false discovery rate by mass spectrometry and genome-wide database searching. Nearly 60% of the interprotein cross-links occur in assemblies involved in transcription and translation. Several of these interactions are new, and we identified a binding site between the δ and β' subunit of RNA polymerase close to the downstream DNA channel, providing a clue into how δ might regulate promoter selectivity and promote RNA polymerase recycling. Our methodology opens new avenues to investigate the functional dynamic organization of complex protein assemblies involved in bacterial growth. Data are available via ProteomeXchange with identifier PXD006287.

Oligo(ethylene glycol)-sidechain microgels prepared in absence of cross-linking agent: Polymerization, characterization and variation of particle deformability.

PubMed

Welsch, Nicole; Lyon, L Andrew

2017-01-01

We present a systematic study of self-cross-linked microgels formed by precipitation polymerization of oligo ethylene glycol methacrylates. The cross-linking density of these microgels and, thus, the network flexibility can be easily tuned through the modulation of the reaction temperature during polymerization. Microgels prepared in absence of any difunctional monomer, i.e. cross-linker, show enhanced deformability and particle spreading on solid surfaces as compared to microgels cross-linked with varying amounts of poly(ethylene glycol diacrylate) (PEG-DA) in addition to self-crosslinking. Particles prepared at low reaction temperatures exhibit the highest degree of spreading due to the lightly cross-linked and flexible polymer network. Moreover, AFM force spectroscopy studies suggest that cross-linker-free microgels constitute of a more homogeneous polymer network than PEG-DA cross-linked particles and have elastic moduli at the particle apex that are ~5 times smaller than the moduli of 5 mol-% PEG-DA cross-linked microgels. Resistive pulse sensing experiments demonstrate that microgels prepared at 75 and 80°C without PEG-DA are able to deform significantly to pass through nanopores that are smaller than the microgel size. Additionally, we found that polymer network flexibility of microgels is a useful tool to control the formation of particle dewetting patterns. This offers a promising new avenue for build-up of 2D self-assembled particle structures with patterned chemical and mechanical properties.

Oligo(ethylene glycol)-sidechain microgels prepared in absence of cross-linking agent: Polymerization, characterization and variation of particle deformability

PubMed Central

Lyon, L. Andrew

2017-01-01

We present a systematic study of self-cross-linked microgels formed by precipitation polymerization of oligo ethylene glycol methacrylates. The cross-linking density of these microgels and, thus, the network flexibility can be easily tuned through the modulation of the reaction temperature during polymerization. Microgels prepared in absence of any difunctional monomer, i.e. cross-linker, show enhanced deformability and particle spreading on solid surfaces as compared to microgels cross-linked with varying amounts of poly(ethylene glycol diacrylate) (PEG-DA) in addition to self-crosslinking. Particles prepared at low reaction temperatures exhibit the highest degree of spreading due to the lightly cross-linked and flexible polymer network. Moreover, AFM force spectroscopy studies suggest that cross-linker-free microgels constitute of a more homogeneous polymer network than PEG-DA cross-linked particles and have elastic moduli at the particle apex that are ~5 times smaller than the moduli of 5 mol-% PEG-DA cross-linked microgels. Resistive pulse sensing experiments demonstrate that microgels prepared at 75 and 80°C without PEG-DA are able to deform significantly to pass through nanopores that are smaller than the microgel size. Additionally, we found that polymer network flexibility of microgels is a useful tool to control the formation of particle dewetting patterns. This offers a promising new avenue for build-up of 2D self-assembled particle structures with patterned chemical and mechanical properties. PMID:28719648

Ubiquitylation and the Fanconi Anemia Pathway

PubMed Central

Garner, Elizabeth; Smogorzewska, Agata

2012-01-01

The Fanconi anemia (FA) pathway maintains genome stability through co-ordination of DNA repair of interstrand crosslinks (ICLs). Disruption of the FA pathway yields hypersensitivity to interstrand crosslinking agents, bone marrow failure and cancer predisposition. Early steps in DNA damage dependent activation of the pathway are governed by monoubiquitylation of FANCD2 and FANCI by the intrinsic FA E3 ubiquitin ligase, FANCL. Downstream FA pathway components and associated factors such as FAN1 and SLX4 exhibit ubiquitin-binding motifs that are important for their DNA repair function, underscoring the importance of ubiquitylation in FA pathway mediated repair. Importantly, ubiquitylation provides the foundations for cross-talk between repair pathways, which in concert with the FA pathway, resolve interstrand crosslink damage and maintain genomic stability. PMID:21605559

Cross-Linked Micellar Spherical Nucleic Acids from Thermoresponsive Templates

PubMed Central

2017-01-01

A one-pot synthesis of micellar spherical nucleic acid (SNA) nanostructures using Pluronic F127 as a thermoresponsive template is reported. These novel constructs are synthesized in a chemically straightforward process that involves intercalation of the lipid tails of DNA amphiphiles (CpG motifs for TLR-9 stimulation) into the hydrophobic regions of Pluronic F127 micelles, followed by chemical cross-linking and subsequent removal of non-cross-linked structures. The dense nucleic acid shell of the resulting cross-linked micellar SNA enhances their stability in physiological media and facilitates their rapid cellular internalization, making them effective TLR-9 immunomodulatory agents. These constructs underscore the potential of SNAs in regulating immune response and address the relative lack of stability of noncovalent constructs. PMID:28207251

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

PubMed Central

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

2014-01-01

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

Innovative Ingredients and Emerging Technologies for Controlling Ice Recrystallization, Texture, and Structure Stability in Frozen Dairy Desserts: A Review.

PubMed

Soukoulis, Christos; Fisk, Ian

2016-11-17

Over the past decade, ice cream manufacturers have developed a strong understanding of the functionality of key ingredients and processing, developing effective explanations for the link between structure forming agents, stability mechanisms, and perceived quality. Increasing demand for products perceived as healthier/more natural with minimal processing has identified a number of new tools to improve quality and storage stability of frozen dairy desserts. Ingredients such as dietary fiber, polysaccharides, prebiotics, alternate sweeteners, fat sources rich in unsaturated fatty acids and ice strucsturing proteins (ISP) have been successfully applied as cryoprotective, texturizing, and structuring agents. Emerging minimal processing technologies including hydrostatic pressure processing, ultrasonic or high pressure assisted freezing, low temperature extrusion and enzymatically induced biopolymers crosslinking have been evaluated for their ability to improve colloidal stability, texture and sensory quality. It is therefore timely for a comprehensive review.

Collision-induced dissociative chemical cross-linking reagents and methodology: Applications to protein structural characterization using tandem mass spectrometry analysis.

PubMed

Soderblom, Erik J; Goshe, Michael B

2006-12-01

Chemical cross-linking combined with mass spectrometry is a viable approach to study the low-resolution structure of protein and protein complexes. However, unambiguous identification of the residues involved in a cross-link remains analytically challenging. To enable a more effective analysis across various MS platforms, we have developed a novel set of collision-induced dissociative cross-linking reagents and methodology for chemical cross-linking experiments using tandem mass spectrometry (CID-CXL-MS/MS). These reagents incorporate a single gas-phase cleavable bond within their linker region that can be selectively fragmented within the in-source region of the mass spectrometer, enabling independent MS/MS analysis for each peptide. Initial design concepts were characterized using a synthesized cross-linked peptide complex. Following verification and subsequent optimization of cross-linked peptide complex dissociation, our reagents were applied to homodimeric glutathione S-transferase and monomeric bovine serum albumin. Cross-linked residues identified by our CID-CXL-MS/MS method were in agreement with published crystal structures and previous cross-linking studies using conventional approaches. Common LC/MS/MS acquisition approaches such as data-dependent acquisition experiments using ion trap mass spectrometers and product ion spectral analysis using SEQUEST were shown to be compatible with our CID-CXL-MS/MS reagents, obviating the requirement for high resolution and high mass accuracy measurements to identify both intra- and interpeptide cross-links.

The effect of pH on hydrolysis, cross-linking and barrier properties of starch barriers containing citric acid.

PubMed

Olsson, Erik; Menzel, Carolin; Johansson, Caisa; Andersson, Roger; Koch, Kristine; Järnström, Lars

2013-11-06

Citric acid cross-linking of starch for e.g. food packaging applications has been intensely studied during the last decade as a method of producing water-insensitive renewable barrier coatings. We managed to improve a starch formulation containing citric acid as cross-linking agent for industrial paper coating applications by adjusting the pH of the starch solution. The described starch formulations exhibited both cross-linking of starch by citric acid as well as satisfactory barrier properties, e.g. fairly low OTR values at 50% RH that are comparable with EVOH. Furthermore, it has been shown that barrier properties of coated papers with different solution pH were correlated to molecular changes in starch showing both hydrolysis and cross-linking of starch molecules in the presence of citric acid. Hydrolysis was shown to be almost completely hindered at solution pH≥4 at curing temperatures≤105 °C and at pH≥5 at curing temperatures≤150 °C, whereas cross-linking still occurred to some extent at pH≤6.5 and drying temperatures as low as 70 °C. Coated papers showed a minimum in water vapor transmission rate at pH 4 of the starch coating solution, corresponding to the point where hydrolysis was effectively hindered but where a significant degree of cross-linking still occurred. Copyright © 2013 Elsevier Ltd. All rights reserved.

Nanogels from Metal-Chelating Crosslinkers as Versatile Platforms Applied to Copper-64 PET Imaging of Tumors and Metastases

DOE PAGES

Lux, Jacques; White, Alexander G.; Chan, Minnie; ...

2015-01-01

Metals are essential in medicine for both therapy and diagnosis. We recently created the first metal-chelating nanogel imaging agent, which employed versatile, reproducible chemistry that maximizes chelation stability. Here we demonstrate that our metal chelating crosslinked nanogel technology is a powerful platform by incorporating 64Cu to obtain PET radiotracers. Polyacrylamide-based nanogels were crosslinked with three different polydentate ligands (DTPA, DOTA, NOTA). NOTA-based nanogels stably retained 64Cu in mouse serum and accumulated in tumors in vivo as detected by PET/CT imaging. Measurement of radioactivity in major organs ex vivo confirmed this pattern, revealing a high accumulation (12.3% ID/g and 16.6% ID/g)more » in tumors at 24 and 48 h following administration, with lower accumulation in the liver (8.5% ID/g at 24 h) and spleen (5.5% ID/g). Nanogels accumulated even more efficiently in metastases (29.9% and 30.4% ID/g at 24 and 48 h). These metal-chelating nanogels hold great promise for future application as bimodal PET/MRI agents; chelation of β-emitting radionuclides could enable radiation therapy.« less

Fanconi anemia: a disorder defective in the DNA damage response.

PubMed

Kitao, Hiroyuki; Takata, Minoru

2011-04-01

Fanconi anemia (FA) is a cancer predisposition disorder characterized by progressive bone marrow failure, congenital developmental defects, chromosomal abnormalities, and cellular hypersensitivity to DNA interstrand crosslink (ICL) agents. So far mutations in 14 FANC genes were identified in FA or FA-like patients. These gene products constitute a common ubiquitin-phosphorylation network called the "FA pathway" and cooperate with other proteins involved in DNA repair and cell cycle control to repair ICL lesions and to maintain genome stability. In this review, we summarize recent exciting discoveries that have expanded our view of the molecular mechanisms operating in DNA repair and DNA damage signaling.

Application of a fast sorting algorithm to the assignment of mass spectrometric cross-linking data.

PubMed

Petrotchenko, Evgeniy V; Borchers, Christoph H

2014-09-01

Cross-linking combined with MS involves enzymatic digestion of cross-linked proteins and identifying cross-linked peptides. Assignment of cross-linked peptide masses requires a search of all possible binary combinations of peptides from the cross-linked proteins' sequences, which becomes impractical with increasing complexity of the protein system and/or if digestion enzyme specificity is relaxed. Here, we describe the application of a fast sorting algorithm to search large sequence databases for cross-linked peptide assignments based on mass. This same algorithm has been used previously for assigning disulfide-bridged peptides (Choi et al., ), but has not previously been applied to cross-linking studies. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Boron Nitride Nanotubes and Nanoplatelets as Reinforcing Agents of Polymeric Matrices for Bone Tissue Engineering

PubMed Central

Farshid, Behzad; Lalwani, Gaurav; Mohammadi, Meisam Shir; Simonsen, John; Sitharaman, Balaji

2016-01-01

This study investigates the mechanical properties and in vitro cytotoxicity of one- and two-dimensional boron nitride nanomaterials-reinforced biodegradable polymeric nanocomposites. Poly(propylene fumarate) (PPF) nanocomposites were fabricated using crosslinking agent N-vinyl pyrrolidone (NVP) and inorganic nanomaterials: boron nitride nanotubes (BNNTs) and boron nitride nanoplatelets (BNNPs) dispersed at 0.2 wt.% in the polymeric matrix. The incorporation of BNNPs and BNNTs resulted in a ~38% and ~15% increase in compressive (young's) modulus, and ~31% and ~6% increase in compressive yield strength compared to PPF control, respectively. The nanocomposites showed a time-dependent increased protein adsorption for only collagen-I protein. The cytotoxicity evaluation of aqueous BNNT and BNNP dispersions (at 1-100 μg/mL concentrations) using a representative murine MC3T3 preosteoblast cell line showed cytocompatibility of BNNTs and BNNPs (~73-99% viability). The cytotoxicity evaluation of media extracts of nanocomposites prior to crosslinking, after crosslinking and upon degradation (using 1X-100X dilutions) showed dose-dependent cytotoxicity responses. Crosslinked nanocomposites showed excellent (~79-100%) cell viability, cellular attachment (~57-67%), and spreading similar to cells grown on the surface of tissue culture polystyrene (TCPS) control. The media extracts of degradation products showed a dose-dependent cytotoxicity. The favorable cytocompatibility results in combination with improved mechanical properties of BNNT and BNNP nanocomposites opens new avenues for further in vitro and in vivo safety and efficacy studies for their bone tissue engineering applications. PMID:26526153

Mechanical characterization of proanthocyanidin-dentin matrix interaction

PubMed Central

Castellan, Carina Strano; Pereira, Patricia Nobrega; Grande, Rosa Helena Miranda; Bedran-Russo, Ana Karina

2010-01-01

Objectives To characterize the properties of dentin matrix treated with two proanthocyanidin rich cross-linking agents and their effect on dentin bonded interfaces. Methods Sound human molars were cut into 0.5 mm thick dentin slabs, demineralized and either treated with one of two cross-linking agents (grape seed - GSE and cocoa seed - COE extracts) or left untreated. The modulus of elasticity of demineralized dentin was assessed after 10 or 60 min and the swelling ratio after 60 min treatment. Bacterial collagenase was also used to assess resistance to enzymatic degradation of samples subjected to ultimate tensile strength. The effect of GSE or COE on the resin-dentin bond strength was evaluated after 10 or 60 min of exposure time. Data were statistically analyzed at a 95% confidence interval. Results Both cross-linkers increased the elastic modulus of demineralized dentin as exposure time increased. Swelling ratio was lower for treated samples when compared to control groups. No statistically significant changes to the UTS indicate that collagenase had no effect on dentin matrix treated with either GSE or COE. Dentin-resin bonds significantly increased following treatment with GSE regardless of the application time or adhesive system used. Significance Increased mechanical properties and stability of dentin matrix can be achieved by the use of PA-rich collagen cross-linkers most likely due to the formation of a PA-collagen complex. The short term dentin-resin bonds can be improved after 10 minutes dentin treatment. PMID:20650510

Chitosan-bound pyridinedicarboxylate Ni(II) and Fe(III) complex biopolymer films as waste water decyanidation agents.

PubMed

Adewuyi, Sheriff; Jacob, Julianah Modupe; Olaleye, Oluwatoyin Omolola; Abdulraheem, Taofiq Olanrewaju; Tayo, Jubril Ayopo; Oladoyinbo, Fatai Oladipupo

2016-10-20

Chitosan is a biopolymer with immense structural advantage for chemical and mechanical modifications to generate novel properties, functions and applications. This work depicts new pyridinedicarboxylicacid (PDC) crosslinked chitosan-metal ion films as veritable material for cyanide ion removal from aqueous solution. The PDC-crosslinked chitosan-metal films (PDC-Chit-Ni(II) and PDC-Chit-Fe(III)) were formed by complexing PDC-crosslinked chitosan film with anhydrous nickel(II) and iron(III) chloride salts respectively. The PDC-Chit and its metal films were characterized employing various analytical and spectroscopic techniques. The FT-IR, UV-vis and the XRD results confirm the presence of the metal ions in the metal coordinated PDC-crosslinked chitosan film. The surface morphological difference of PDC-Chit-Ni(II) film before and after decyanidation was explored with scanning electron microscopy. Furthermore, the quantitative amount of nickel(II) and iron(III) present in the complex were determined using Atomic Absorption Spectrophotometer as 32.3 and 37.2μg/g respectively which portends the biopolymer film as a good complexing agent. Removal of cyanide from aqueous solution with PDC-Chit, PDC-Chit-Ni(II) and PDC-Chit-Fe(III) films was studied with batch equilibrium experiments. At equilibrium, decyanidation capacity (DC) followed the order PDC-Chit-Ni (II)≈PDC-Chit-Fe(III)>PDC-Chit. PDC-Chit-Ni(II) film gave 100% CN(-) removal within 40min decyanidation owing to favorable coordination geometry. Copyright © 2016 Elsevier Ltd. All rights reserved.

Inert Reassessment Document for Trimethylolpropane - CAS No. 77-99-6

EPA Pesticide Factsheets

Trimethylolpropane is used in a large variety of commercial applications, including in the manufacturer of varnishes, resins; polyesters for polyurethane foams, textile finishes, plastictzers, and cross-linking agents for spandex fibers.

Investigation of FANCA gene in Fanconi anaemia patients in Iran

PubMed Central

Saffar Moghadam, Ali Akbar; Mahjoubi, Frouzandeh; Reisi, Nahid; Vosough, Parvaneh

2016-01-01

Background & objectives: Fanconi anaemia (FA) is a syndrome with a predisposition to bone marrow failure, congenital anomalies and malignancies. It is characterized by cellular hypersensitivity to cross-linking agents such as mitomycin C (MMC). In the present study, a new approach was selected to investigate FANCA (Fanconi anaemia complementation group A) gene in patients clinically diagnosed with cellular hypersensitivity to DNA cross-linking agent MMC. Methods: Chromosomal breakage analysis was performed to prove the diagnosis of Fanconi anaemia in 318 families. Of these, 70 families had a positive result. Forty families agreed to molecular genetic testing. In total, there were 27 patients with unknown complementary types. Genomic DNA was extracted and total RNA was isolated from fresh whole blood of the patients. The first-strand cDNA was synthesized and the cDNA of each patient was then tested with 21 pairs of overlapping primers. High resolution melting curve analysis was used to screen FANCA, and LinReg software version 1.7 was utilized for analysis of expression. Results: In total, six sequence alterations were identified, which included two stop codons, two frames-shift mutations, one large deletion and one amino acid exchange. FANCA expression was downregulated in patients who had sequence alterations. Interpretation & conclusions: The results of the present study show that high resolution melting (HRM) curve analysis may be useful in the detection of sequence alteration. It is simpler and more costeffective than the multiplex ligation-dependent probe amplification (MLPA) procedure. PMID:27121516

Investigation of FANCA gene in Fanconi anaemia patients in Iran.

PubMed

Moghadam, Ali Akbar Saffar; Mahjoubi, Frouzandeh; Reisi, Nahid; Vosough, Parvaneh

2016-02-01

Fanconi anaemia (FA) is a syndrome with a predisposition to bone marrow failure, congenital anomalies and malignancies. It is characterized by cellular hypersensitivity to cross-linking agents such as mitomycin C (MMC). In the present study, a new approach was selected to investigate FANCA (Fanconi anaemia complementation group A) gene in patients clinically diagnosed with cellular hypersensitivity to DNA cross-linking agent MMC. Chromosomal breakage analysis was performed to prove the diagnosis of Fanconi anaemia in 318 families. Of these, 70 families had a positive result. Forty families agreed to molecular genetic testing. In total, there were 27 patients with unknown complementary types. Genomic DNA was extracted and total RNA was isolated from fresh whole blood of the patients. The first-strand cDNA was synthesized and the cDNA of each patient was then tested with 21 pairs of overlapping primers. High resolution melting curve analysis was used to screen FANCA, and LinReg software version 1.7 was utilized for analysis of expression. In total, six sequence alterations were identified, which included two stop codons, two frames-shift mutations, one large deletion and one amino acid exchange. FANCA expression was downregulated in patients who had sequence alterations. The results of the present study show that high resolution melting (HRM) curve analysis may be useful in the detection of sequence alteration. It is simpler and more cost-effective than the multiplex ligation-dependent probe amplification (MLPA) procedure.

Development of Microbubble Contrast Agents with Biochemical Recognition and Tunable Acoustic Response

NASA Astrophysics Data System (ADS)

Nakatsuka, Matthew Allan Masao

Microbubbles, consisting of gas-filled cores encapsulated within phospholipid or polymer shells, are the most widely used ultrasound contrast agents in the world. Because of their acoustic impedance mismatch with surrounding tissues and compressible gaseous interiors, they have high echogenicities that allow for efficient backscatter of ultrasound. They can also generate unique harmonic frequencies when insonated near their resonance frequency, depending on physical microbubble properties such as the stiffness and thickness of the encapsulating shell. Microbubbles are used to detect a number of cardiovascular diseases, but current methodologies lack the ability to detect and distinguish small, rapidly growing abnormalities that do not produce visible blockage or slowing of blood flow. This work describes the development, formulation, and validation of microbubbles with various polymer shell architectures designed to modulate their acoustic ability. We demonstrate that the addition of a thick disulfide crosslinked, poly(acrylic acid) encapsulating shell increases a bubble's resistance to cavitation and changes its resonance frequency. Modification of this shell architecture to use hybridized DNA strands to form crosslinks between the polymer chains allows for tuning of the bubble acoustic response. When the DNA crosslinks are in place, shell stiffness is increased so the bubbles do not oscillate and acoustic signal is muted. Subsequently, when these DNA strands are displaced, partial acoustic activity is restored. By using aptamer sequences with a specific affinity towards the biomolecule thrombin as the DNA crosslinking strand, this acoustic "ON/OFF" behavior can be specifically tailored towards the presence of a specific biomarker, and produces a change in acoustic signal at concentrations of thrombin consistent with acute deep venous thrombosis. Incorporation of the emulsifying agent poly(ethylene glycol) into the encapsulating shell improves microbubble yield and circulation half-life substantially, allowing for the in vivo detection of a blood clot in a rabbit model. The results presented here show a unique marriage of highly specific molecular imaging and sonography that could be tailored towards a wide variety of cardiovascular abnormalities.

Role of solvent-mediated carbodiimide cross-linking in fabrication of electrospun gelatin nanofibrous membranes as ophthalmic biomaterials.

PubMed

Chou, Shih-Feng; Luo, Li-Jyuan; Lai, Jui-Yang; Ma, David Hui-Kang

2017-02-01

Due to their ability to mimic the structure of extracellular matrix, electrospun gelatin nanofibers are promising cell scaffolding materials for tissue engineering applications. However, the hydrophilic gelatin molecules usually need stabilization before use in aqueous physiological environment. Considering that biomaterials cross-linked via film immersion technique may have a more homogeneous cross-linked structure than vapor phase cross-linking, this work aims to investigate the chemical modification of electrospun gelatin nanofibrous membranes by liquid phase carbodiimide in the presence of ethanol/water co-solvents with varying ethanol concentrations ranging from 80 to 99.5vol%. The results of characterization showed that increasing water content in the binary reaction solvent system increases the extent of cross-linking of gelatin nanofibers, but simultaneously promotes the effect of biopolymer swelling and distortion in fiber mat structure. As compared to non-cross-linked counterparts, carbodiimide treated gelatin nanofibrous mats exhibited better thermal and biological stability where the shrinkage temperature and resistance to enzymatic degradation varied in response to ethanol/water solvent composition-mediated generation of cross-links. Irrespective of their cross-linking density, all studied membrane samples did not induce any responses in ocular epithelial cell cultures derived from cornea, lens, and retina. Unlike many other cross-linking agents and/or methods (e.g., excessive vapor phase cross-linking) that may pose a risk of toxicity, our study demonstrated that these nanofibrous materials are well tolerated by anterior segment tissues. These findings also indicate the safety of using ethanol/water co-solvents for chemical cross-linking of gelatin to engineer nanofibrous materials with negligible biological effects. In summary, the present results suggest the importance of solvent-mediated carbodiimide cross-linking in modulating structure-property relationship without compromising in vitro and in vivo biocompatibility of electrospun gelatin nanofibers for future ophthalmic applications. Copyright © 2016 Elsevier B.V. All rights reserved.

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

PubMed

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

2017-11-01

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

Water soluble folate-chitosan nanogels crosslinked by genipin.

PubMed

Pujana, Maite Arteche; Pérez-Álvarez, Leyre; Iturbe, L Carlos Cesteros; Katime, Issa

2014-01-30

Folate-chitosan conjugates were prepared by a concurrent functionalization and crosslinking reaction with the natural crosslinker genipin. Genipin molecule was employed simultaneously as crosslinker agent and spacer molecule in order to allow the functionalization with folic acid for active tumor targeting. The reaction was carried out in reverse microemulsion which provided colloidal size and monodisperse particle size distribution. The water solubility of the obtained folate-genipin-chitosan nanogels was studied as function of the pH of the medium and all nanoparticles were totally dispersible at physiological pH. The enzymatic degradability of the nanogels in a lysozyme solution was evaluated at acidic and physiological pH. QELS analyses of the swelling behavior of the nanogels with the pH did not show a clear pH-sensitivity. However, the study on the loading and release capacity of 5-fluorouracil revealed an interesting pH-responsive behavior of the nanogels that makes them promising as nanodevices for targeted anticancer drug delivery. Copyright © 2013 Elsevier Ltd. All rights reserved.

Ion Exchange Equilibrium and Kinetic Properties of Polyacrylate Films and Applications to Chemical Analysis and Environmental Decontamination

NASA Technical Reports Server (NTRS)

Tanner, Stephen P.

1997-01-01

One of the goals of the original proposal was to study how cross-linking affects the properties of an ion exchange material(IEM) developed at Lewis Research Center. However, prior to the start of this work, other workers at LERC investigated the effect of cross-linking on the properties of this material. Other than variation in the ion exchange capacity, the chemical characteristics were shown to be independent of the cross-linking agent, and the degree of cross-linking. New physical forms of the film were developed (film, supported film, various sizes of beads, and powder). All showed similar properties with respect to ion exchange equilibria but the kinetics of ion exchange depended on the surface area per unit mass; the powder form of the IEM exchanging much more rapidly than the other forms. The research performed under this grant was directed towards the application of the IEM to the analysis of metal ions at environmental concentrations.

In vitro and in vivo evaluation of novel interpenetrated polymer network microparticles containing repaglinide.

PubMed

Kulkarni, Raghavendra V; Patel, Foram S; Nanjappaiah, H M; Naikawadi, Akram A

2014-08-01

Interpenetrated polymer network (IPN) microparticles of sterculia gum and sodium alginate loaded with repaglinide were developed by ionic gelation and emulsion crosslinking method. The drug entrapment efficiency was as high as 91%. FTIR and TG analyses confirmed the crosslinking and IPN formation. Microparticles have demonstrated the drug release up to 24h depending upon type of crosslinking agents; the glutaraldehyde treatment of ionically crosslinked microparticles has resulted in decreased drug release rate. The in-vivo anti-diabetic activity performed on streptozotocin induced diabetic rats indicated that the pristine repaglinide has shown maximum percentage reduction of elevated blood glucose within 3h and then the percentage reduction in blood glucose was decreased. In the case of rats treated with KA8 IPN microparticles, percentage reduction of elevated glucose was slow as compared to pristine drug within 3h, but it was gradually increased to 81.27% up to 24h. Copyright © 2014 Elsevier B.V. All rights reserved.

Gamma-irradiated cross-linked LDPE foams: Characteristics and properties

NASA Astrophysics Data System (ADS)

Cardoso, E. C. L.; Scagliusi, S. R.; Parra, D. F.; Lugão, A. B.

2013-03-01

Foamed polymers are future materials, as they are increasingly considered "green materials" due to their interesting properties at very low consumption of raw materials. They can be used to improve appearance of insulation structures, thermal and acoustic insulation, core materials for sandwich panels, fabrication of furniture and flotation materials or to reduce costs involving materials. Low-density polyethylene is widely used because of its excellent properties, such as softness, elasticity, processibility and insulation. In general, cross-linking is often applied to improve the thermal and mechanical properties of polyethylene products, due to the formation of a three-dimensional network. In particular for the production of PE foams, cross-linking is applied prior the expansion to control bubble formation, cell characteristics and final properties of the foam. However, the usual production process of PE foams is a process in which a gaseous blowing agent is injected into a melted thermoplastic polymer, under pressure, to form a solution between blowing agent and melted polymer. An extrusion system is provided for foaming the polymer, supplied to an extruder and moving through a rotating screw. The pressure must be high enough to keep the gas blowing agent (or foaming agent) in the solution with the melt. The foaming agent is then diffused and dissolved in the molten material to form a single-phase solution. In the present work carbon dioxide was used as the bowing agent, a chemically stable and non-toxic gas, with good diffusion coefficient; gas pressure used varied within a 20-40 bar range. Some requirements for physical foaming are required, as low friction heat generation, homogeneous melt temperature distribution, melt temperature at die exit just above crystallization temperature (die) and high melt strength during expansion. This work studied foams properties gamma-irradiated within 0, 10, 15, 20, 25, and 30 kGy, from a LDPE exhibiting 2.6 g/10 min Melt Index. Accomplished tests: DSC, gel-fraction, swelling ratio in various solvents, rheological measurements, infra-red spectroscopy and melt strength. It was verified that within a given radiation dose range; the material exhibited an optimization in viscoelastic properties, providing the desired melt strength range for obtaining foams.

Study on Movable gel Profiling/Flooding System Optimization at Boundary Temperature

NASA Astrophysics Data System (ADS)

Gao, Shanshan; Xie, Gang; Zhang, Tiantian; Wang, Zhiqiang; Jiang, Peijun; Wei, Junjie; Gu, Yi; Lei, Xiaoyang; Guo, Suzhen; Lei, Shi

2017-12-01

65-75°C is the boundary temperature of phenol-formaldehyde resin and organic chrome crosslink with HPAM to generate movable gel. Generally speaking, when the reservoir temperature is below 65-75°C, the crosslinking agent is Organic chrome. Phenol-formaldehyde resin is used when the temperature is above 65-75°C. In this paper the gelation properties of phenol-formaldehyde resin and organic chrome were compared at 70°C. The experiment results showed that the crosslinking time of phenol-formaldehyde resin gel was longer and more conducive to field injection. In addition the phenol-formaldehyde resin gel had greater viscosity, adhesion and shearing resistance compared with the organic chrome gel. So the phenol-formaldehyde resin was chosen for further optimization. The crosslinking time was shortened and the gel viscosity increased with the increase of the polymer concentration. As the polymer crosslinker proportion increased The crosslinking time was shortened and the gel viscosity increased first and then decreased. Reinforcer NC and stabilizer WG were added to improve the temperature and salt tolerance of the gel. The gel formula suitable for the boundary temperature was obtained. The optimum polymer concentration is 1200mg/L, the polymer crosslinker proportion is 1:1.1, the best reinforcer concentration is 400mg/L and the concentration of the stabilizer is 150mg/L. The crosslinking time is 31h, the gel viscosity is above 2100mPa·s. The gel did not shrink and no water separation was observed at 70°C for 150 days. The viscosity retention rate was more than 70%.

The Study of the Interaction between Silica Filler and Silicone Rubber

NASA Astrophysics Data System (ADS)

Liu, Jiesheng; Gong, Xiaoqiang; Zhang, Rongtang

2018-01-01

The interaction between silica filler and silicone rubber was studied by swelling ratio, Kraus curve and crosslinking density test. The results showed that lower values of Vro/Vrf and swelling ratio in modified filler system suggests good filler-matrix interactions. The composites with silane coupling agents show higher crosslink-density compared that of untreated ones. In the light of the above statement, it can be concluded that modification of filler is the crucial factor in creating a good interaction between the filler and silicone rubber.

Fractionation and reconstitution experiments provide insight into the role of starch gelatinization and pasting properties in pasta quality.

PubMed

Delcour, J A; Vansteelandt, J; Hythier, M; Abécassis, J

2000-09-01

Commercial durum wheat semolina was fractionated into protein, starch, water-extractable, and sludge fractions. The starch fraction was hydroxypropylated, annealed, or cross-linked to change its gelatinization and pasting properties. Spaghettis were made by reconstitution of the fractions, and their quality was assessed. Hydroxypropylated starches were detrimental for cooked pasta quality. Cross-linked starches made the reconstituted pasta firmer and even brittle when the degree of cross-linking was too high. These results indicate that starch properties play a role in pasta quality, although the gluten remains very important as an ultrastructure agent. It was concluded that, given a certain gluten ultrastructure, starch water uptake and gel properties and/or its interference with or breakdown of the continuous gluten network during cooking determine pasta quality.

Near-Infrared-Light-Responsive Magnetic DNA Microgels for Photon- and Magneto-Manipulated Cancer Therapy.

PubMed

Wang, Yitong; Wang, Ling; Yan, Miaomiao; Dong, Shuli; Hao, Jingcheng

2017-08-30

Functional DNA molecules have been introduced into polymer-based nanocarrier systems to incorporate chemotherapy drugs for cancer therapy. Here is the first report of dual-responsive microgels composed of a core of Au nanorods and a shell of magnetic ionic liquid and DNA moieties in the cross-linking network simultaneously, as effective drug delivery vectors. TEM images indicated a magnetic polymer shell has an analogous "doughnut" shape which loosely surround the AuNRs core. When irradiated with a near-infrared-light (near-IR) laser, Au nanorods are the motors which convert the light to heat, leading to the release of the encapsulated payloads with high controllability. DNA acts not only as a cross-linker agent, but also as a gatekeeper to regulate the release of drugs. The internalization study and MTT assay confirm that these core-shell DNA microgels are excellent candidates which can enhance the cytotoxicity of cancer cells controlled by near-IR laser and shield the high toxicity of chemotherapeutic agents to improve the killing efficacy of chemotherapeutic agents efficiently in due course.

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

PubMed

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

2017-02-03

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

In-Silico Design, Synthesis and Evaluation of a Nanostructured Hydrogel as a Dimethoate Removal Agent.

PubMed

Avila-Salas, Fabian; Marican, Adolfo; Villaseñor, Jorge; Arenas-Salinas, Mauricio; Argandoña, Yerko; Caballero, Julio; Durán-Lara, Esteban F

2018-01-04

This study describes the in-silico design, synthesis, and evaluation of a cross-linked PVA hydrogel (CLPH) for the absorption of organophosphorus pesticide dimethoate from aqueous solutions. The crosslinking effectiveness of 14 dicarboxilic acids was evaluated through in-silico studies using semiempirical quantum mechanical calculations. According to the theoretical studies, the nanopore of PVA cross-linked with malic acid (CLPH-MA) showed the best interaction energy with dimethoate. Later, using all-atom molecular dynamics simulations, three hydrogels with different proportions of PVA:MA (10:2, 10:4, and 10:6) were used to evaluate their interactions with dimethoate. These results showed that the suitable crosslinking degree for improving the affinity for the pesticide was with 20% ( W %) of the cross-linker. In the experimental absorption study, the synthesized CLPH-MA20 recovered 100% of dimethoate from aqueous solutions. Therefore, the theoretical data were correlated with the experimental studies. Surface morphology of CLPH-MA20 by Scanning Electron Microscopy (SEM) was analyzed. In conclusion, the ability of CLPH-MA20 to remove dimethoate could be used as a technological alternative for the treatment of contaminated water.

In-Silico Design, Synthesis and Evaluation of a Nanostructured Hydrogel as a Dimethoate Removal Agent

PubMed Central

Avila-Salas, Fabian; Marican, Adolfo; Villaseñor, Jorge; Argandoña, Yerko

2018-01-01

This study describes the in-silico design, synthesis, and evaluation of a cross-linked PVA hydrogel (CLPH) for the absorption of organophosphorus pesticide dimethoate from aqueous solutions. The crosslinking effectiveness of 14 dicarboxilic acids was evaluated through in-silico studies using semiempirical quantum mechanical calculations. According to the theoretical studies, the nanopore of PVA cross-linked with malic acid (CLPH-MA) showed the best interaction energy with dimethoate. Later, using all-atom molecular dynamics simulations, three hydrogels with different proportions of PVA:MA (10:2, 10:4, and 10:6) were used to evaluate their interactions with dimethoate. These results showed that the suitable crosslinking degree for improving the affinity for the pesticide was with 20% (W%) of the cross-linker. In the experimental absorption study, the synthesized CLPH-MA20 recovered 100% of dimethoate from aqueous solutions. Therefore, the theoretical data were correlated with the experimental studies. Surface morphology of CLPH-MA20 by Scanning Electron Microscopy (SEM) was analyzed. In conclusion, the ability of CLPH-MA20 to remove dimethoate could be used as a technological alternative for the treatment of contaminated water. PMID:29300312

Reprocessing and Recycling of Highly Cross-Linked Ion-Conducting Networks through Transalkylation Exchanges of C-N Bonds.

PubMed

Obadia, Mona M; Mudraboyina, Bhanu P; Serghei, Anatoli; Montarnal, Damien; Drockenmuller, Eric

2015-05-13

Exploiting exchangeable covalent bonds as dynamic cross-links recently afforded a new class of polymer materials coined as vitrimers. These permanent networks are insoluble and infusible, but the network topology can be reshuffled at high temperatures, thus enabling glasslike plastic deformation and reprocessing without depolymerization. We disclose herein the development of functional and high-value ion-conducting vitrimers that take inspiration from poly(ionic liquid)s. Tunable networks with high ionic content are obtained by the solvent- and catalyst-free polyaddition of an α-azide-ω-alkyne monomer and simultaneous alkylation of the resulting poly(1,2,3-triazole)s with a series of difunctional cross-linking agents. Temperature-induced transalkylation exchanges of C-N bonds between 1,2,3-triazolium cross-links and halide-functionalized dangling chains enable recycling and reprocessing of these highly cross-linked permanent networks. They can also be recycled by depolymerization with specific solvents able to displace the transalkylation equilibrium, and they display a great potential for applications that require solid electrolytes with excellent mechanical performances and facile processing such as supercapacitors, batteries, fuel cells, and separation membranes.

Chemically crosslinked protein dimers: stability and denaturation effects.

PubMed Central

Byrne, M. P.; Stites, W. E.

1995-01-01

Nine single substitution cysteine mutants of staphylococcal nuclease (nuclease) were preferentially crosslinked at the introduced cysteine residues using three different bifunctional crosslinking reagents; 1,6-bismaleimidohexane (BMH), 1,3-dibromo-2-propanol (DBP), and the chemical warfare agent, mustard gas (bis(2-chloroethyl)sulfide; mustard). BMH and mustard gas are highly specific reagents for cysteine residues, whereas DBP is not as specific. Guanidine hydrochloride (GuHCl) denaturations of the resulting dimeric proteins exhibited biphasic unfolding behavior that did not fit the two-state model of unfolding. The monofunctional reagent, epsilon-maleimidocaproic acid (MCA), was used as a control for the effects of alkylation. Proteins modified with MCA unfolded normally, showing that this unusual unfolding behavior is due to crosslinking. The data obtained from these crosslinked dimers was fitted to a three-state thermodynamic model of two successive transitions in which the individual subunits cooperatively unfold. These two unfolding transitions were very different from the unfolding of the monomeric protein. These differences in unfolding behavior can be attributed in large part to changes in the denatured state. In addition to GuHCl titrations, the crosslinked dimers were also thermally unfolded. In contrast to the GuHCl denaturations, analysis of this data fit a two-state model well, but with greatly elevated van't Hoff enthalpies in many cases. However, clear correlations between the thermal and GuHCl denaturations exist, and the differences in thermal unfolding can be rationalized by postulating interactions of the denatured crosslinked proteins. PMID:8580845

Walking of antitumor bifunctional trinuclear PtII complex on double-helical DNA

PubMed Central

Malina, Jaroslav; Kasparkova, Jana; Farrell, Nicholas P.; Brabec, Viktor

2011-01-01

The trinuclear BBR3464 ([{trans-PtCl(NH3)2}2µ-(trans-Pt(NH3)2(H2N(CH2)6NH2)2)]4+) belongs to the polynuclear class of platinum-based anticancer agents. DNA adducts of this complex differ significantly in structure and type from those of clinically used mononuclear platinum complexes, especially, long-range (Pt, Pt) intrastrand and interstrand cross-links are formed in both 5′–5′ and 3′–3′ orientations. We show employing short oligonucleotide duplexes containing single, site-specific cross-links of BBR3464 and gel electrophoresis that in contrast to major DNA adducts of clinically used platinum complexes, under physiological conditions the coordination bonds between platinum and N7 of G residues involved in the cross-links of BBR3464 can be cleaved. This cleavage may lead to the linkage isomerization reactions between this metallodrug and double-helical DNA. Differential scanning calorimetry of duplexes containing single, site-specific cross-links of BBR3464 reveals that one of the driving forces that leads to the lability of DNA cross-links of this metallodrug is a difference between the thermodynamic destabilization induced by the cross-link and by the adduct into which it could isomerize. The rearrangements may proceed in the way that cross-links originally formed in one strand of DNA can spontaneously translocate from one DNA strand to its complementary counterpart, which may evoke walking of the platinum complex on DNA molecule. PMID:20833634

Unique Dynamic Properties of DNA Duplexes Containing Interstrand Crosslinks†

PubMed Central

Friedman, Joshua I.; Jiang, Yu Lin; Miller, Paul S.; Stivers, James T.

2010-01-01

Bifunctional DNA alkylating agents form a diverse assortment of covalent DNA interstrand crosslinked (ICL) structures that are potent cytotoxins. Since it is implausible that cells could possess distinct DNA repair systems for each individual ICL, it is believed that common structural and dynamic features of ICL damage are recognized, rather than specific structural characteristics of each cross-linking agent. Investigation of the structural and dynamic properties of ICLs that might be important for recognition has been complicated by heterogeneous incorporation of these lesions into DNA. To address this problem we have synthesized and characterized several homogenous ICL-DNAs containing site–specific staggered N4-cytosine-ethyl-N4-cytosine crosslinks. Staggered crosslinks were introduced in two ways: in a manner that preserves the overall structure of B-form duplex DNA, and in a manner that highly distorts the DNA structure, with the goal of understanding how structural and dynamic properties of diverse ICL duplexes might flag these sites for repair. Measurements of base pair opening dynamics in the B-form ICL duplex by 1H NMR linewidth or imino proton solvent exchange showed that the guanine base opposite to the crosslinked cytosine opened at least an order of magnitude more slowly than when in a control matched normal duplex. To a lesser degree, the B-form ICL also induced a decrease in base pair opening dynamics that extended from the site of the crosslink to adjacent base pairs. In contrast, the non-B-form ICL showed extensive conformational dynamics at the site of the cross link, which extended over the entire DNA sequence. Since DNA duplexes containing the B-form and non-B-form ICL crosslinks have both been shown to be incised when incubated in mammalian whole cell extracts, while a matched normal duplex is not, we conclude that intrinsic DNA dynamics is not a requirement for specific damage incision of these ICLs. Instead, we propose a general model where destabilized ICL-duplexes serve to energetically facilitate binding of DNA repair factors that must induce bubbles or other distortions in the duplex. However, the essential requirement for incision is an immobile Y-junction where the repair factors are stably bound at the site of the ICL, and the two DNA strands are unpaired. PMID:21174443

Self-assembled high-strength hydroxyapatite/graphene oxide/chitosan composite hydrogel for bone tissue engineering.

PubMed

Yu, Peng; Bao, Rui-Ying; Shi, Xiao-Jun; Yang, Wei; Yang, Ming-Bo

2017-01-02

Graphene hydrogel has shown greatly potentials in bone tissue engineering recently, but it is relatively weak in the practical use. Here we report a facile method to synthesize high strength composite graphene hydrogel. Graphene oxide (GO), hydroxyapatite (HA) nanoparticles (NPs) and chitosan (CS) self-assemble into a 3-dimensional hydrogel with the assistance of crosslinking agent genipin (GNP) for CS and reducing agent sodium ascorbate (NaVC) for GO simultaneously. The dense and oriented microstructure of the resulted composite gel endows it with high mechanical strength, high fixing capacity of HA and high porosity. These properties together with the good biocompatibility make the ternary composite gel a promising material for bone tissue engineering. Such a simultaneous crosslinking and reduction strategy can also be applied to produce a variety of 3D graphene-polymer based nanocomposites for biomaterials, energy storage materials and adsorbent materials. Copyright © 2016 Elsevier Ltd. All rights reserved.

Final Report

NASA Technical Reports Server (NTRS)

1995-01-01

This cooperative agreement explored the novel polymerization of vinylsilane, alone and in combination with other alkenylsilanes, alkylsilanes, and/or crosslinking agents, using dimethyltitanocene as homogeneous catalyst. The reactions were found to be initiated photochemically under mild conditions, and no hydrogen gas was evolved when alkenylsilanes were polymerized. The polymers were found to have predominately a carbosilane-type backbone containing -SiH2-CH2-CH2-Si and -SiH2-CH(CH3)-Si type linkages. The mechanism of polymerization was found to be step-growth. Despite the relatively low molecular weight of the polymer (M(sub n) - 500 and M(sub W) - 1500), pyrolysis resulted in conversion to C-rich SiC ceramic in high char yields (-60%). Copolymerization with methylsilane resulted in higher chars and more crystalline polymer. Addition of crosslinking agents such as polybutadiene or methyltrivinylsilane increased the viscosity of the polymer produced and enabled application as coatings to fiber toes.

Mapping contacts between gRNA and mRNA in trypanosome RNA editing.

PubMed

Leung, S S; Koslowsky, D J

1999-02-01

All guide RNAs (gRNAs) identified to date have defined 5' anchor sequences, guiding sequences and a non-encoded 3' uridylate tail. The 5' anchor is required for in vitro editing and is thought to be responsible for selection and binding to the pre-edited mRNA. Little is known, however, about how the gRNAs are used to direct RNA editing. Utilizing the photo-reactive crosslinking agent, azidophenacyl (APA), attached to the 5'- or 3'-terminus of the gRNA, we have begun to map the structural relationships between the different defined regions of the gRNA with the pre-edited mRNA. Analyses of crosslinked conjugates produced with a 5'-terminal APA group confirm that the anchor of the gRNA is correctly positioning the interacting molecules. 3' Crosslinks (X-linker placed at the 3'-end of a U10tail) have also been mapped for three different gRNA/mRNA pairs. In all cases, analyses indicate that the U-tail can interact with a range of nucleotides located upstream of the first edited site. It appears that the U-tail prefers purine-rich sites, close to the first few editing sites. These results suggest that the U-tail may act in concert with the anchor to melt out secondary structure in the mRNA in the immediate editing domain, possibly increasing the accessibility of the editing complex to the proper editing sites.

Boron nitride nanotubes and nanoplatelets as reinforcing agents of polymeric matrices for bone tissue engineering.

PubMed

Farshid, Behzad; Lalwani, Gaurav; Shir Mohammadi, Meisam; Simonsen, John; Sitharaman, Balaji

2017-02-01

This study investigates the mechanical properties and in vitro cytotoxicity of one- and two-dimensional boron nitride nanomaterials-reinforced biodegradable polymeric nanocomposites. Poly(propylene fumarate) (PPF) nanocomposites were fabricated using crosslinking agent N-vinyl pyrrolidone and inorganic nanomaterials: boron nitride nanotubes (BNNTs) and boron nitride nanoplatelets (BNNPs) dispersed at 0.2 wt % in the polymeric matrix. The incorporation of BNNPs and BNNTs resulted in a ∼38 and ∼15% increase in compressive (Young's) modulus, and ∼31 and ∼6% increase in compressive yield strength compared to PPF control, respectively. The nanocomposites showed a time-dependent increased protein adsorption for collagen I protein. The cytotoxicity evaluation of aqueous BNNT and BNNP dispersions (at 1-100 μg/mL concentrations) using murine MC3T3 preosteoblast cells showed ∼73-99% viability. The cytotoxicity evaluation of media extracts of nanocomposites before crosslinking, after crosslinking, and upon degradation (using 1×-100× dilutions) showed dose-dependent cytotoxicity responses. Crosslinked nanocomposites showed excellent (∼79-100%) cell viability, cellular attachment (∼57-67%), and spreading similar to cells grown on the surface of tissue culture polystyrene control. The media extracts of degradation products showed a dose-dependent cytotoxicity. The favorable cytocompatibility results in combination with improved mechanical properties of BNNT and BNNP nanocomposites opens new avenues for further in vitro and in vivo safety and efficacy studies towards bone tissue engineering applications. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 406-419, 2017. © 2015 Wiley Periodicals, Inc.

Biohydrogels with magnetic nanoparticles as crosslinker: characteristics and potential use for controlled antitumor drug-delivery.

PubMed

Barbucci, Rolando; Giani, Gabriele; Fedi, Serena; Bottari, Severino; Casolaro, Mario

2012-12-01

Hybrid magnetic hydrogels are of interest for applications in biomedical science as controlled drug-delivery systems. We have developed a strategy to obtain novel hybrid hydrogels with magnetic nanoparticles (NPs) of CoFe(2)O(3) and Fe(3)O(4) as crosslinker agents of carboxymethylcellulose (CMC) or hyaluronic acid (HYAL) polymers and we have tested these systems for controlled doxorubicin release. The magnetic NPs are functionalized with (3-aminopropyl)trimethoxysilane (APTMS) in order to introduce amino groups on the surface. The amino coating is determined and quantified by standard Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy methods, and by cyclic voltammetry, a novel approach that permits us to look at the solution properties of the functionalized NPs. The gel formation involves the creation of an amide bond between the carboxylic groups of CMC or HYAL and the amine groups of functionalized NPs, which work as crosslinking agents of the polymer chains. The hybrid hydrogels are chemically and morphologically characterized. The rheological and the water uptake properties of the hydrogels are also investigated. Under the application of an alternating magnetic field, the CMC-HYAL hybrid hydrogel previously loaded with doxorubicin shows a drug release greater than that showed by the CMC-HYAL hydrogel crosslinked with 1,3-diaminopropane. In conclusion, the presence of magnetic NPs makes the synthesized hybrid hydrogels suitable for application as a drug-delivery system by means of alternating magnetic fields. Copyright © 2012 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Evaluation of different crosslinking agents on hybrid biomimetic collagen-hydroxyapatite composites for regenerative medicine.

PubMed

Krishnakumar, Gopal Shankar; Gostynska, Natalia; Dapporto, Massimiliano; Campodoni, Elisabetta; Montesi, Monica; Panseri, Silvia; Tampieri, Anna; Kon, Elizaveta; Marcacci, Maurilio; Sprio, Simone; Sandri, Monica

2018-01-01

This study focuses on the development of novel bone-like scaffolds by bio-inspired, pH-driven, mineralization of type I collagen matrix with magnesium-doped hydroxyapatite nanophase (MgHA/Coll). To this aim, this study evaluates the altered modifications in the obtained composite due to different crosslinkers such as dehydrothermal treatment (DHT), 1,4-butanediol diglycidyl ether (BDDGE) and ribose in terms of morphological, physical-chemical and biological properties. The physical-chemical properties of the composites evaluated by XRD, FTIR, ICP and TGA demonstrated that the chemical mimesis of bone was effectively achieved using the in-lab biomineralization process. Furthermore, the presence of various crosslinkers greatly promoted beneficial enzymatic resistivity and swelling ability. The morphological results revealed highly porous and fibrous micro-architecture with total porosity above 85% with anisotropic pore size within the range of 50-200μm in all the analysed composites. The mechanical behaviour in response to compressive forces demonstrated enhanced compressive modulus in all crosslinked composites, suggesting that mechanical behaviour is largely dependent on the type of crosslinker used. The biomimetic compositional and morphological features of the composites elicited strong cell-material interaction. Therefore, the results showed that by activating specific crosslinking mechanisms, hybrid composites can be designed and tailored to develop tissue-specific biomimetic biomaterials for hard tissue engineering. Copyright © 2017 Elsevier B.V. All rights reserved.

Cross-linked polyimides for integrated optics

NASA Astrophysics Data System (ADS)

Singer, Kenneth D.; Kowalczyk, Tony C.; Nguyen, Hung D.; Beuhler, Allyson J.; Wargowski, David A.

1997-01-01

We have investigated a promising class of polyimide materials for both passive and active electro-optic devices, namely crosslinkable polyimides. These fluorinated polyimides are soluble in the imidized form and are both thermally and photo-crosslinkable leading to easy processability into waveguide structures and the possibility of stable electro-optic properties. We have fabricated channel and slab waveguides and investigated the mechanism of optical propagation loss using photothermal deflection spectroscopy and waveguide loss spectroscopy, and found the losses to arise from residual absorption due to the formation of charge transfer states. The absorption is inhibited by fluorination leading to propagation losses as low as 0.3 dB/cm in the near infrared. Because of the ability to photocrosslink, channel waveguides are fabricated using a simple wet-etch process. Channel waveguides so formed are observed to have no excess loss over slab structures. Solubility followed by thermal cross-linking allows the formation of multilayer structures. We have produced electro-optic polymers by doping with the nonlinear optical chromophores, DCM and DADC; and a process of concurrent poling and thermal crosslinking. Multilayer structures have been investigated and poling fields optimized in the active layer by doping the cladding with an anti-static agent. The high glass-transition temperature and cross-linking leads to very stable electro-optic properties. We are currently building electro-optic modulators based on these materials. Progress and results in this area also are reported.

Optimization of the fabrication of novel stealth PLA-based nanoparticles by dispersion polymerization using D-optimal mixture design

PubMed Central

Adesina, Simeon K.; Wight, Scott A.; Akala, Emmanuel O.

2015-01-01

Purpose Nanoparticle size is important in drug delivery. Clearance of nanoparticles by cells of the reticuloendothelial system has been reported to increase with increase in particle size. Further, nanoparticles should be small enough to avoid lung or spleen filtering effects. Endocytosis and accumulation in tumor tissue by the enhanced permeability and retention effect are also processes that are influenced by particle size. We present the results of studies designed to optimize crosslinked biodegradable stealth polymeric nanoparticles fabricated by dispersion polymerization. Methods Nanoparticles were fabricated using different amounts of macromonomer, initiators, crosslinking agent and stabilizer in a dioxane/DMSO/water solvent system. Confirmation of nanoparticle formation was by scanning electron microscopy (SEM). Particle size was measured by dynamic light scattering (DLS). D-optimal mixture statistical experimental design was used for the experimental runs, followed by model generation (Scheffe polynomial) and optimization with the aid of a computer software. Model verification was done by comparing particle size data of some suggested solutions to the predicted particle sizes. Results and Conclusion Data showed that average particle sizes follow the same trend as predicted by the model. Negative terms in the model corresponding to the crosslinking agent and stabilizer indicate the important factors for minimizing particle size. PMID:24059281

Optimization of the fabrication of novel stealth PLA-based nanoparticles by dispersion polymerization using D-optimal mixture design.

PubMed

Adesina, Simeon K; Wight, Scott A; Akala, Emmanuel O

2014-11-01

Nanoparticle size is important in drug delivery. Clearance of nanoparticles by cells of the reticuloendothelial system has been reported to increase with increase in particle size. Further, nanoparticles should be small enough to avoid lung or spleen filtering effects. Endocytosis and accumulation in tumor tissue by the enhanced permeability and retention effect are also processes that are influenced by particle size. We present the results of studies designed to optimize cross-linked biodegradable stealth polymeric nanoparticles fabricated by dispersion polymerization. Nanoparticles were fabricated using different amounts of macromonomer, initiators, crosslinking agent and stabilizer in a dioxane/DMSO/water solvent system. Confirmation of nanoparticle formation was by scanning electron microscopy (SEM). Particle size was measured by dynamic light scattering (DLS). D-optimal mixture statistical experimental design was used for the experimental runs, followed by model generation (Scheffe polynomial) and optimization with the aid of a computer software. Model verification was done by comparing particle size data of some suggested solutions to the predicted particle sizes. Data showed that average particle sizes follow the same trend as predicted by the model. Negative terms in the model corresponding to the cross-linking agent and stabilizer indicate the important factors for minimizing particle size.

Navy LOVA propellant development

NASA Technical Reports Server (NTRS)

Vreatt, W. H.; Mitchell, S. E.

1980-01-01

The progress realized on evaluation of inert binder, nitramine formulations is considered with respect to their development for use as low vulnerability ammunition (LOVA) propellants. Burning rate, plasticizers, crosslink agents, physical property and vulnerability studies are discussed and some preliminary conclusions presented.

Compositional effects on the chemorheological properties and forming behavior of aqueous alumina-poly(vinyl alcohol) gelcasting suspensions

NASA Astrophysics Data System (ADS)

Morissette, Sherry L.

A new gelcasting system based on aqueous, alumina-poly(vinyl alcohol) (PVA) suspensions cross-linked by an organotitanate coupling agent has been developed. Both the chemorheological properties and forming behavior of this system exhibited a strong compositional dependence. A sol- gel phase diagram was established, which yielded the critical titanium concentration [Ti] c required for gelation at a given PVA volume fraction, as well as the minimum PVA volume fraction ( fminPVA = 0.0245) and titanium PVA concentration ([Ti]min = 9.984 x 10--4 g Ti/ml) below which gelation was not observed irrespective of solution composition. The gelation time of suspensions of constant PVA volume fraction ( fsolnPVA ) decreased with increasing cross-linking agent concentration, PVA temperature, and solids volume fraction. The steady-state viscosity and elastic modulus of polymer solutions ( fsolnPVA = 0.05) of varying [Ti] were well described by the PVA percolation model, giving scaling exponents of 0.84 and 1.79, respectively. The steady-state elastic modulus of gel casting suspensions, which provides a measure of their handling strength in the as-gelled state, increased with increasing solids volume fraction. Gelcasting suspensions were used as feedstock for solid free-form fabrication (SFF) of ceramic components. The influence of processing conditions (e.g., tip diameter, mixing rate, table speed, etc.) and suspension rheology on deposition behavior was investigated. Continuous printablity was achieved for tip diameters ranging from dt = 0.254 -- 1.370 mm for all mixing rates (Rmix 5 -- 300 rpm) and suspension compositions (i.e., fAl2O3 = 0.45, φPVA = 0.275, [Ti] 0 -- 6.30 x 10--3 g Ti/ml) probed, where the minimum tip diameter for continuous printing was 0.203 mm. Printed lines were uniform with good edge definition. Line dimensions were independent of mixing rate for the given process conditions. The as-cast alumina volume fraction ( fAl2O3 ) depended on casting conditions and cross-linking agent concentration, where fAl2O3 decreased with increasing tip diameter and increased with increasing cross-linking agent concentration. Free-fomied Al2O3 components exhibited uniform particle packing and had minimal macro-defects (e.g., slumping or stair casing) and no detectable micro-defects (e.g., bubbles or cracking).

[Molecular basis of Fanconi's anemia].

PubMed

Digweed, M

1999-01-01

Fanconi anaemia (FA) is an autosomal recessive genetic disorder characterised clinically by progressive bone marrow failure, skeletal deformities and a predisposition to neoplasia. Patient cells manifest an extreme chromosomal instability and hypersensitivity to polyfunctional alkylating agents. It is assumed that the basic defect is related to the repair of DNA damage, in particular that of so-called DNA crosslinks. Currently there are eight complementation groups in FA (FA-A-FA-H) which indicates that at least eight independent genes can lead to FA. Three of these genes have been identified: FANCA, FANCC and FANCG. In this review, the molecular biology and genetics of FA are presented and possible functions of the FANC proteins are discussed.

A new injectable biphasic hydrogel based on partially hydrolyzed polyacrylamide and nano hydroxyapatite, crosslinked with chromium acetate, as scaffold for cartilage regeneration

NASA Astrophysics Data System (ADS)

Koushki, N.; Tavassoli, H.; Katbab, A. A.; Katbab, P.; Bonakdar, S.

2015-05-01

Polymer scaffolds are applied in the field of tissue engineering as three dimensional structures to organize cells and present stimuli to direct generation of a desired damaged tissue. In situ gelling scaffolds have attracted great attentions, as they are structurally similar to the extra cellular matrix (ECM). In the present work, attempts have been made to design and fabricate a new injectable and crosslinkable biphasic hydrogel based on partially hydrolyzed polyacrylamide (HPAM), chromium acetate as crosslink agent and nanocrystalline hydroxyapatite (nHAp) as reinforcing and bioactive agent for repair and regeneration of damaged cartilage. The distinct characteristic of HPAM is the presence of carboxylate anion groups on its backbone which allows to engineer the structure of the hydrogel for the desired bioactivity with appropriate cells differentiation towards both soft and hard (bone) tissues. The synthesized hydrogel exhibited bifunctional behavior which was derived by its biphasic structure in which one phase was loaded with nano hydroxyapatite to provide integration capability by subchondral bones and fix the hydrogel at cartilage defect without a need for suturing. The other phase differentiates the rabbit adipogenic mesenchymal stem cells (MSCs) towards soft tissue. Rheomechanical spectrometry (RMS) was employed to study the kinetic of the gelation including induction time and rate, as well as to measure the ultimate elastic modulus of the optimum crosslinked hydrogel. Surface tension measurement was also performed to tailor the surface characteristics of the gels. In vitro culturing of the cells inside the crosslinked hydrogel revealed high viability and high differentiation of the encapsulated rabbit stem cells, providing that the chromium acetate level was kept below 0.2 wt%. Based on the obtained results, the designed and fabricated biphasic hydrogel exhibited high potential as carrier for the stem cells for cartilage tissue engineering application with excellent injectability.

The use of cross-linked chitosan beads for nutrients (nitrate and orthophosphate) removal from a mixture of P-PO4, N-NO2 and N-NO3.

PubMed

Jóźwiak, Tomasz; Filipkowska, Urszula; Szymczyk, Paula; Kuczajowska-Zadrożna, Małgorzata; Mielcarek, Artur

2017-11-01

A hydrogel chitosan sorbent ionically cross-linked with sodium citrate and covalently cross-linked with epichlorohydrin was used to remove nutrients from an equimolar mixture of P-PO 4 , N-NO 2 and N-NO 3 . The scope of the study included, among other things, determination of the influence of pH on nutrient sorption effectiveness, nutrient sorption kinetics as well as determination of the maximum sorption capacity of cross-linked chitosan sorbents regarding P-PO 4 (H 2 PO 4 - , HPO 4 2- ), N-NO 2 (HNO 2 , NO 2 - ), and N-NO 3 (NO 3 - ). The effect of the type of the cross-linking agent on the affinity of the modified chitosan to each nutrient was studied as well. The kinetics of nutrient sorption on the tested chitosan sorbents was best described with the pseudo-second order model. The model of intramolecular diffusion showed that P-PO 4 , N-NO 2 and N-NO3 sorption on cross-linked hydrogel chitosan beads proceeded in two phases. The best sorbent of nutrients turned out to be chitosan cross-linked covalently with epichlorohydrin; with P-PO 4 , N-NO 2 and N-NO 3 sorption capacity reaching: 1.23, 0.94 and 0.76mmol/g, respectively (total of 2.92mmol/g). For comparison, the sorption capacity of chitosan cross-linked ionically with sodium citrate was: 0.43, 0.39 and 0.39mmol/g for P-PO 4 , N-NO 2 and N-NO 3 , respectively (total of 1.21mmol/g). Copyright © 2017 Elsevier B.V. All rights reserved.

Identification of MS-Cleavable and Non-Cleavable Chemically Crosslinked Peptides with MetaMorpheus.

PubMed

Lu, Lei; Millikin, Robert J; Solntsev, Stefan K; Rolfs, Zach; Scalf, Mark; Shortreed, Michael R; Smith, Lloyd M

2018-05-25

Protein chemical crosslinking combined with mass spectrometry has become an important technique for the analysis of protein structure and protein-protein interactions. A variety of crosslinkers are well developed, but reliable, rapid, and user-friendly tools for large-scale analysis of crosslinked proteins are still in need. Here we report MetaMorpheusXL, a new search module within the MetaMorpheus software suite that identifies both MS-cleavable and non-cleavable crosslinked peptides in MS data. MetaMorpheusXL identifies MS-cleavable crosslinked peptides with an ion-indexing algorithm, which enables an efficient large database search. The identification does not require the presence of signature fragment ions, an advantage compared to similar programs such as XlinkX. One complication associated with the need for signature ions from cleavable crosslinkers such as DSSO (disuccinimidyl sulfoxide) is the requirement for multiple fragmentation types and energy combinations, which is not necessary for MetaMorpheusXL. The ability to perform proteome-wide analysis is another advantage of MetaMorpheusXl compared to such programs as MeroX and DXMSMS. MetaMorpheusXL is also faster than other currently available MS-cleavable crosslink search software programs. It is imbedded in MetaMorpheus, an open-source and freely available software suite that provides a reliable, fast, user-friendly graphical user interface that is readily accessible to researchers.

Sequestration of GPI-anchored proteins in caveolae triggered by cross-linking.

PubMed

Mayor, S; Rothberg, K G; Maxfield, F R

1994-06-24

Glycosyl-phosphatidylinositol (GPI)-anchored proteins have been reported to reside in clusters collected over small membrane invaginations called caveolae. The detection of different GPI-anchored proteins with fluorescently labeled monoclonal antibodies showed that these proteins are not constitutively concentrated in caveolae; they enter these structures independently after cross-linking with polyclonal secondary antibodies. Analysis of the cell surface distribution of the GPI-anchored folate receptor by electron microscopy confirms these observations. Thus, multimerization of GPI-anchored proteins regulates their sequestration in caveolae, but in the absence of agents that promote clustering they are diffusely distributed over the plasma membrane.

Two DD-carboxypeptidases from Mycobacterium smegmatis affect cell surface properties through regulation of peptidoglycan cross-linking and glycopeptidolipids.

PubMed

Pandey, Satya Deo; Pal, Shilpa; Kumar N, Ganesh; Bansal, Ankita; Mallick, Sathi; Ghosh, Anindya S

2018-05-07

During the peptidoglycan (PG) maturation of mycobacteria, the glycan strands are interlinked by both 3-3 (between two meso-DAP) and 4-3 cross-links (between D-ala and meso-DAP), though there is a predominance (60-80%) of 3-3 cross-links. The DD-CPases act on pentapeptides to generate tetrapeptides that are used by LD-transpeptidases as substrates to form 3-3 cross-links. Therefore, DD-CPases play a crucial role in mycobacterial PG cross-link formation. However, the physiology of DD-CPases in mycobacteria is relatively unexplored. Here, we deleted two DD-CPase genes, msmeg_2433 , and msmeg_2432 , both individually and in combination, from Mycobacterium smegmatis mc 2 155. Though the single DD-CPase deletions had no significant impact on the mycobacterial physiology, many interesting functional alterations were observed in the double deletion mutant, viz. , a predominance in PG cross-link formation was shifted from 3-3 cross-links to 4-3, cell surface glycopeptidolipid (GPL) expression was reduced and susceptibility towards β-lactams and anti-tubercular agents was enhanced. Moreover, the existence of the double mutant within murine macrophages was better as compared to the parent. Interestingly, the complementation with any one of the DD-CPase genes could restore the wild-type phenotype. In a nutshell, we infer that the altered ratio of 4-3: 3-3 PG cross-links might have influenced the expression of surface GPLs, colony morphology, biofilm formation,, drug susceptibility and subsistence of the cells within macrophages. Importance The glycan strands in mycobacterial peptidoglycan (PG) are interlinked by both 3-3 and 4-3 cross-links. The DD-CPases generate tetrapeptides by acting on the pentapeptides, and LD-transpeptidases use tetrapeptides as substrates to form 3-3 cross-links. Here, we showed that simultaneous deletions of two DD-CPases alter the nature of PG cross-linking from 3-3 cross-links to 4-3 cross-links. The deletions subsequently decrease the expression of Glycopeptidolipids (a significant surface lipid present in many non-tuberculous mycobacteria including Mycobacterium smegmatis ) and affect other physiological parameters like cell morphology, growth rate, biofilm formation, antibiotic susceptibility and existence within murine macrophages. Thus, unraveling the physiology of DD-CPases might help us design anti-mycobacterial therapeutics in future. Copyright © 2018 American Society for Microbiology.

Synthesis and characterization of drug loaded albumin mesospheres for intratumoral chemotherapy

NASA Astrophysics Data System (ADS)

Freeman, Shema Taian

Conventional chemotherapy is problematic due to toxic complications. Intratumoral (IT) drug delivery, offers a new, less toxic, potentially more effective treatment concept. The objectives of this research encompassed (1) an investigation of the synthesis of BSA mesospheres (MS) employing genipin (GEN) as a novel crosslinking agent, (2) comparison with glutaraldehyde (GTA) crosslinked mesosphere, (3) a study of process parameters to define conditions for the synthesis of 1-10microm drug loaded mesospheres, and (4) investigation of the drug delivery properties of such mesospheres for IT chemotherapy. Smooth, spherical BSA-MS, crosslinked with glutaraldehyde and genipin, were prepared in a dry particle size range of 1microm to 10microm. It was shown that increasing dispersion stirring rate, crosslinking time and GEN/BSA ratio led to a decrease in particle size and a narrower particle distribution. It was also shown that increasing crosslinking time, GEN/BSA ratio, BSA concentrations, GEN concentration slowed enzymatic degradation. Post-loading and in situ drug loading methods were studied for the incorporation of cyclophosphamide and cisplatin into mesospheres. Maximum post loading of cisplatin was 3.2% (w/w) and 2.6% (w/w) with GEN and with GTA crosslinking. For cyclophosphamide 8.2% (w/w) and 7.1% (w/w) loading was achieved with GEN and GTA respectively. In situ drug loaded MS genipin and glutaraldehyde crosslinked mesospheres were also synthesized with 1.8% (w/w) cisplatin (using GEN) and 1.2% (w/w) (using GTA). Maximum loading of 13.3% (w/w) was achieved for cyclophosphamide in genipin crosslinked mesospheres. The cytotoxicity of in situ loaded genipin and glutaraldehyde crosslinked cisplatin mesospheres was evaluated using a murine Lewis lung model. Both genipin and glutaraldehyde crosslinked BSA-cisplatin mesospheres proved to be cytotoxic during a 48 hour test. Ultimately a standard set of processing parameters (BSA concentration, CAB concentration, GEN concentration, GEN/BSA ratio, stabilization stirring rate and crosslinking time) were defined to produce both GEN and GTA crosslinked cisplatin and cyclophosphamide BSA mesospheres. In vitro analysis confirmed the utility of mesosphere bound drug. In several related studies, (1) IT delivered dispersions of mitoxantrone loaded albumin microspheres were shown to afford an effective treatment, with significantly prolonging animal survival and (2) genipin and gadolinium crosslinked MS were prepared from HA and BSA/HA.

Identification of cross-linked amino acids in the protein pair HmaL23-HmaL29 from the 50S ribosomal subunit of the archaebacterium Haloarcula marismortui.

PubMed

Bergmann, U; Wittmann-Liebold, B

1993-03-23

50S ribosomal subunits from the extreme halophilic archaebacterium Haloarcula marismortui were treated with the homobifunctional protein-protein cross-linking reagents diepoxybutane (4 A) and dithiobis(succinimidyl propionate) (12 A). The dominant product with both cross-linking reagents was identified on the protein level as HmaL23-HmaL29, which is homologous to the protein pair L23-L29 from Escherichia coli [Walleczek, J., Martin, T., Redl, B., Stöffler-Meilicke, M., & Stöffler, G. (1989) Biochemistry 28, 4099-4105] and from Bacillus stearothermophilus [Brockmöller, J., & Kamp, R. M. (1986) Biol. Chem. Hoppe-Seyler 367, 925-935]. To reveal the exact cross-linking site in HmaL23-HmaL29, the cross-linked complex was purified on a preparative scale by conventional and high-performance liquid chromatography. After endoproteolytic fragmentation of the protein pair, the amino acids engaged in cross-link formation were unambiguously identified by N-terminal sequence analysis and mass spectrometry of the cross-linked peptides. The cross-link is formed between lysine-57 in the C-terminal region of HmaL29 and the alpha-amino group of the N-terminal serine in protein HmaL23, irrespective of the cross-linking reagent. This result demonstrates that the N-terminal region of protein HmaL23 and the C-terminal domain of HmaL29 are highly flexible so that the distance between the two polypeptide chains can vary by at least 8 A. Comparison of our cross-linking results with those obtained with B. stearothermophilus revealed that the fine structure within this ribosomal domain is at least partially conserved.

Hekate: Software Suite for the Mass Spectrometric Analysis and Three-Dimensional Visualization of Cross-Linked Protein Samples

PubMed Central

2013-01-01

Chemical cross-linking of proteins combined with mass spectrometry provides an attractive and novel method for the analysis of native protein structures and protein complexes. Analysis of the data however is complex. Only a small number of cross-linked peptides are produced during sample preparation and must be identified against a background of more abundant native peptides. To facilitate the search and identification of cross-linked peptides, we have developed a novel software suite, named Hekate. Hekate is a suite of tools that address the challenges involved in analyzing protein cross-linking experiments when combined with mass spectrometry. The software is an integrated pipeline for the automation of the data analysis workflow and provides a novel scoring system based on principles of linear peptide analysis. In addition, it provides a tool for the visualization of identified cross-links using three-dimensional models, which is particularly useful when combining chemical cross-linking with other structural techniques. Hekate was validated by the comparative analysis of cytochrome c (bovine heart) against previously reported data.1 Further validation was carried out on known structural elements of DNA polymerase III, the catalytic α-subunit of the Escherichia coli DNA replisome along with new insight into the previously uncharacterized C-terminal domain of the protein. PMID:24010795

BLM promotes the activation of Fanconi Anemia signaling pathway.

PubMed

Panneerselvam, Jayabal; Wang, Hong; Zhang, Jun; Che, Raymond; Yu, Herbert; Fei, Peiwen

2016-05-31

Mutations in the human RecQ helicase, BLM, causes Bloom Syndrome, which is a rare autosomal recessive disorder and characterized by genomic instability and an increased risk of cancer. Fanconi Anemia (FA), resulting from mutations in any of the 19 known FA genes and those yet to be known, is also characterized by chromosomal instability and a high incidence of cancer. BLM helicase and FA proteins, therefore, may work in a common tumor-suppressor signaling pathway. To date, it remains largely unclear as to how BLM and FA proteins work concurrently in the maintenance of genome stability. Here we report that BLM is involved in the early activation of FA group D2 protein (FANCD2). We found that FANCD2 activation is substantially delayed and attenuated in crosslinking agent-treated cells harboring deficient Blm compared to similarly treated control cells with sufficient BLM. We also identified that the domain VI of BLM plays an essential role in promoting FANCD2 activation in cells treated with DNA crosslinking agents, especially ultraviolet B. The similar biological effects performed by ΔVI-BLM and inactivated FANCD2 further confirm the relationship between BLM and FANCD2. Mutations within the domain VI of BLM detected in human cancer samples demonstrate the functional importance of this domain, suggesting human tumorigenicity resulting from mtBLM may be at least partly attributed to mitigated FANCD2 activation. Collectively, our data show a previously unknown regulatory liaison in advancing our understanding of how the cancer susceptibility gene products act in concert to maintain genome stability.

Structure-based non-canonical amino acid design to covalently crosslink an antibody–antigen complex

PubMed Central

Xu, Jianqing; Tack, Drew; Hughes, Randall A.; Ellington, Andrew D.; Gray, Jeffrey J.

2014-01-01

Engineering antibodies to utilize non-canonical amino acids (NCAA) should greatly expand the utility of an already important biological reagent. In particular, introducing crosslinking reagents into antibody complementarity determining regions (CDRs) should provide a means to covalently crosslink residues at the antibody–antigen interface. Unfortunately, finding the optimum position for crosslinking two proteins is often a matter of iterative guessing, even when the interface is known in atomic detail. Computer-aided antibody design can potentially greatly restrict the number of variants that must be explored in order to identify successful crosslinking sites. We have therefore used Rosetta to guide the introduction of an oxidizable crosslinking NCAA, l-3,4-dihydroxyphenylalanine (l-DOPA), into the CDRs of the anti-protective antigen scFv antibody M18, and have measured crosslinking to its cognate antigen, domain 4 of the anthrax protective antigen. Computed crosslinking distance, solvent accessibility, and interface energetics were three factors considered that could impact the efficiency of l-DOPA-mediated crosslinking. In the end, 10 variants were synthesized, and crosslinking efficiencies were generally 10% or higher, with the best variant crosslinking to 52% of the available antigen. The results suggest that computational analysis can be used in a pipeline for engineering crosslinking antibodies. The rules learned from l-DOPA crosslinking of antibodies may also be generalizable to the formation of other crosslinked interfaces and complexes. PMID:23680795

A new glycation product ‘norpronyl-lysine,’ and direct characterization of cross linking and other glycation adducts: NMR of model compounds and collagen

PubMed Central

Bullock, Peter T. B.; Reid, David G.; Ying Chow, W.; Lau, Wendy P. W.; Duer, Melinda J.

2014-01-01

NMR is ideal for characterizing non-enzymatic protein glycation, including AGEs (advanced glycation endproducts) underlying tissue pathologies in diabetes and ageing. Ribose, R5P (ribose-5-phosphate) and ADPR (ADP-ribose), could be significant and underinvestigated biological glycating agents especially in chronic inflammation. Using [U-13C]ribose we have identified a novel glycoxidation adduct, 5-deoxy-5-desmethylpronyl-lysine, ‘norpronyl-lysine’, as well as numerous free ketones, acids and amino group reaction products. Glycation by R5P and ADPR proceeds rapidly with R5P generating a brown precipitate with PLL (poly-L-lysine) within hours. ssNMR (solid-state NMR) 13C–13C COSY identifies several crosslinking adducts such as the newly identified norpronyl-lysine, in situ, from the glycating reaction of 13C5-ribose with collagen. The same adducts are also identifiable after reaction of collagen with R5P. We also demonstrate for the first time bio-amine (spermidine, N-acetyl lysine, PLL) catalysed ribose 2-epimerization to arabinose at physiological pH. This work raises the prospect of advancing understanding of the mechanisms and consequences of glycation in actual tissues, in vitro or even ex vivo, using NMR isotope-labelled glycating agents, without analyses requiring chemical or enzymatic degradations, or prior assumptions about glycation products. PMID:27919030

Caffeine-catalyzed gels.

PubMed

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

2018-07-01

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

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

PubMed

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

2014-10-01

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

Synthesis and evaluation of thermo-rheological behaviour and ionotropic crosslinking of new gellan gum-alkyl derivatives.

PubMed

Agnello, Stefano; Palumbo, Fabio Salvatore; Pitarresi, Giovanna; Fiorica, Calogero; Giammona, Gaetano

2018-04-01

This paper reports the synthesis and the physicochemical characterization of two series of gellan gum (GG) derivatives functionalized with alkyl chains with different number of carbon, from 8 to 18. In particular, low molecular weight gellan gum samples with 52.6 or 96.7 kDa, respectively, were functionalized with octylamine (C 8 ), dodecylamine (C 12 ) and octadecylamine (C 18 ) by using bis(4-nitrophenyl) carbonate (4-NPBC) as a coupling agent. Thermo-rheological and ionotropic crosslinking properties of these gellan gum-alkyl derivatives were evaluated and related to the degree of derivatization in alkyl chains. Results suggested as length and degree of derivatization differently influenced coil-to-helix gelation mechanism of GG derivatives, ionotropic crosslinking, and strength of crosslinked hydrogels obtained in CaCl 2 0.102 M and NaCl 0.15 M. Statement of hypothesis: The insertion of alkyl chains on the gellan gum backbone interferes with coil-to-helix transition mechanism and allows the production of hydrophobically assembled hydrogels. Copyright © 2018 Elsevier Ltd. All rights reserved.

Preparation and physico-chemical properties of hydrogels from carboxymethyl cassava starch crosslinked with citric acid

NASA Astrophysics Data System (ADS)

Boonkham, Sasikan; Sangseethong, Kunruedee; Chatakanon, Pathama; Niamnuy, Chalida; Nakasaki, Kiyohiko; Sriroth, Klanarong

2014-06-01

Recently, environmentally friendly hydrogels prepared from renewable bio-based resources have drawn significant attention from both industrial and academic sectors. In this study, chemically crosslinked hydrogels have been developed from cassava starch which is a bio-based polymer using a non-toxic citric acid as a crosslinking agent. Cassava starch was first modified by carboxymethylation to improve its water absorbency property. The carboxymethyl cassava starch (CMCS) obtained was then crosslinked with citric acid at different concentrations and reaction times. The gel fraction of hydrogels increased progressively with increasing citric acid concentration. Free swelling capacity of hydrogels in de-ionized water, saline solution and buffers at various pHs as well as absorption under load were investigated. The results revealed that swelling behavior and mechanical characteristic of hydrogels depended on the citric acid concentration used in reaction. Increasing citric acid concentration resulted in hydrogels with stronger network but lower swelling and absorption capacity. The cassava starch hydrogels developed were sensitive to ionic strength and pH of surrounding medium, showing much reduced swelling capacity in saline salt solution and acidic buffers.

Rapid, High Affinity Binding by a Fluorescein Templated Copolymer Combining Covalent, Hydrophobic, and Acid–Base Noncovalent Crosslinks

PubMed Central

Timberman, Anthony; Yang, Rongfang; Papantones, Alex; Seitz, W. Rudolf

2018-01-01

A new type of biomimetic templated copolymer has been prepared by reverse addition fragmentation chain transfer polymerization (RAFT) in dioxane. The initial formulation includes the template fluorescein, N-isopropylacrylamide (NIPAM, 84 mol %), methacrylic acid (MAA, 5-mol %), 4-vinylpyridine (4-VP, 9 mmol %), and N,N′-methylenebis(acrylamide) (MBA, 2 mol %). PolyNIPAM is a thermosensitive polymer that comes out of aqueous solution above its lower critical solution temperature forming hydrophobic ‘crosslinks’. MAA and 4-VP interact in dioxane forming acid–base crosslinks. The excess 4-VP serves as a recognition monomer organizing around the template fluorescein to form a binding site that is held in place by the noncovalent and covalent crosslinks. The MBA is a covalent crosslinker. The RAFT agent in the resulting copolylmer was reduced to a thiol and attached to gold nanoparticles. The gold nanoparticle bound copolymer binds fluorescein completely in less than two seconds with an affinity constant greater than 108 M−1. A reference copolymer prepared with the same monomers by the same procedure binds fluorescein much more weakly. PMID:29693601

Electrically responsive materials based on polycarbazole/sodium alginate hydrogel blend for soft and flexible actuator application.

PubMed

Sangwan, Watchara; Petcharoen, Karat; Paradee, Nophawan; Lerdwijitjarud, Wanchai; Sirivat, Anuvat

2016-10-20

The electromechanical properties, namely the storage modulus sensitivity and bending, of sodium alginate (SA) hydrogels and polycarbazole/sodium alginate (PCB/SA) hydrogel blends under applied electric field was investigated. The electromechanical properties of the pristine SA were studied under effects of crosslinking types and SA molecular weights, whereas the PCB/SA hydrogel blends were studied under the effect of PCB concentrations. The storage modulus sensitivity and bending of the pristine SA as crosslinked by the ionic crosslinking agent were found to be higher than those of the covalent crosslinking. The storage modulus sensitivity and deflection of the SA increased monotonically with increasing molecular weight. The highest electromechanical response of the PCB/SA hydrogel blends was obtained from the blend with 0.10% v/v PCB as it provided surprisingly the highest ever storage modulus sensitivity, (G'-G'0)/G'0 where G'0 and G' are the storage modulus without and with applied electric field, respectively, at 18.5 under applied electric field strength of 800V/mm. Copyright © 2016 Elsevier Ltd. All rights reserved.

Deficiency of UBE2T, the E2 Ubiquitin Ligase Necessary for FANCD2 and FANCI Ubiquitination, Causes FA-T Subtype of Fanconi Anemia.

PubMed

Rickman, Kimberly A; Lach, Francis P; Abhyankar, Avinash; Donovan, Frank X; Sanborn, Erica M; Kennedy, Jennifer A; Sougnez, Carrie; Gabriel, Stacey B; Elemento, Olivier; Chandrasekharappa, Settara C; Schindler, Detlev; Auerbach, Arleen D; Smogorzewska, Agata

2015-07-07

Fanconi anemia (FA) is a rare bone marrow failure and cancer predisposition syndrome resulting from pathogenic mutations in genes encoding proteins participating in the repair of DNA interstrand crosslinks (ICLs). Mutations in 17 genes (FANCA-FANCS) have been identified in FA patients, defining 17 complementation groups. Here, we describe an individual presenting with typical FA features who is deficient for the ubiquitin-conjugating enzyme (E2), UBE2T. UBE2T is known to interact with FANCL, the E3 ubiquitin-ligase component of the multiprotein FA core complex, and is necessary for the monoubiquitination of FANCD2 and FANCI. Proband fibroblasts do not display FANCD2 and FANCI monoubiquitination, do not form FANCD2 foci following treatment with mitomycin C, and are hypersensitive to crosslinking agents. These cellular defects are complemented by expression of wild-type UBE2T, demonstrating that deficiency of the protein UBE2T can lead to Fanconi anemia. UBE2T gene gains an alias of FANCT. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Immobilized laccase mediated dye decolorization and transformation pathway of azo dye acid red 27.

PubMed

Chhabra, Meenu; Mishra, Saroj; Sreekrishnan, Trichur Ramaswamy

2015-01-01

Laccases have good potential as bioremediating agents and can be used continuously in the immobilized form like many other enzymes. In the present study, laccase from Cyathus bulleri was immobilized by entrapment in Poly Vinyl Alcohol (PVA) beads cross-linked with either nitrate or boric acid. Immobilized laccase was used for dye decolorization in both batch and continuous mode employing a packed bed column. The products of degradation of dye Acid Red 27 were identified by LC MS/MS analysis. The method led to very effective (90%) laccase immobilization and also imparted significant stability to the enzyme (more than 70% after 5 months of storage at 4°C). In batch decolorization, 90-95% decolorization was achieved of the simulated dye effluent for up to 10-20 cycles. Continuous decolorization in a packed bed bioreactor led to nearly 90% decolorization for up to 5 days. The immobilized laccase was also effective in decolorization and degradation of Acid Red 27 in the presence of a mediator. Four products of degradation were identified by LC-MS/MS analysis. The immobilized laccase in PVA-nitrate was concluded to be an effective agent in treatment of textile dye effluents.

Degradable Hydrogels and Nanogels for the Delivery of Cells and Therapeutics

NASA Astrophysics Data System (ADS)

Boehnke, Natalie

Degradable polymeric materials such as hydrogels are extensively utilized as delivery vehicles due to their biocompatibility and tunable properties. Encapsulating therapeutic agents inside hydrogels stabilizes the cargo by preventing degradation, extending circulation time, and also allows for targeted release and delivery. Due to their small size and tunable properties, nano-scale hydrogels, or nanogels, are frequently utilized to deliver therapeutics to areas difficult to reach, such as tumors and the cytoplasm, through traditional means. To control hydro- and nanogel function, degradable cross-links can be installed, allowing for cargo release in response to specific stimuli, such as hydrolysis or reduction. This dissertation offers three degradable strategies that can be applied to synthesize hydrogels and nanogels for the stabilization and release of therapeutic cargo. In the first example, mixed imine cross-linking chemistry was applied to synthesize poly(ethylene glycol) (PEG)-based hydrogels with tunable degradability to encapsulate and deliver cells. Time to degradation of the gels could be controlled from 24 hours to more than 7 days by varying the hydrazone structure and the ratio of hydrazone and oxime cross-links. Encapsulated cells exhibited high viability up to at least 7 days, suggesting this system may be useful for cell delivery applications. In the second example, disulfide cross-links were utilized to form redox-responsive nanogels comprised of trehalose copolymers. The synthesis of a methacrylate trehalose monomer (TrMA) was optimized, improving the overall yield from 14% to 42%. TrMA was subsequently copolymerized with pyridyl disulfide ethyl methacrylate (PDSMA) using free radical polymerization conditions to form copolymers with two monomer ratios (1:1 and 2:1) which were cross-linked with 1 kDa PEG-dithiol via disulfide exchange to form uniform nanogels approximately 9 nm in diameter. The addition of a cross-linker eliminated the need to add reducing agent to facilitate cross-linking and nanogel formation, making this approach ideal for the encapsulation of sensitive therapeutic agents. Next, PDSMA-co-TrMA nanogels were utilized to encapsulate, stabilize, and release glucagon, an unstable peptide hormone used to treat hypoglycemia. The amines on glucagon were modified with thiol groups while retaining their positive charges for reversible conjugation and cross-linking. Glucagon-nanogel conjugates were synthesized with >80% conjugation yield, and the reversible disulfide linkage between peptide and polymer allowed for efficient cargo release under mild reducing conditions. The nanogels stabilized glucagon against aggregation in solution up to five days as well as solubilized the peptide at neutral pH. In vitro bioactivity of the modified peptide was found to be comparable to native glucagon, suggesting this may be a promising formulation strategy for further in vivo study. Finally, a series of dual-enzyme responsive peptides was synthesized by masking the epsilon-amine of lysine with protease substrates. After unmasking the amine by enzymatic cleavage, a second enzyme was able to cleave at the C terminus of lysine, which was monitored colorimetrically. Three different dual-enzyme responsive peptides were prepared, (AcAAF)K-pNA, (AcFG)K-pNA, and (AcDEVD)K-pNA, for chymotrypsin, papain, and caspase 3 sensitivity, respectively, followed by trypsin sensitivity after cleavage by the first enzyme. This modular peptide design could be useful for selective drug delivery, studies on dual enzyme activity, as well as for diagnostic enzyme screening.

A new class of dual responsive self-healable hydrogels based on a core crosslinked ionic block copolymer micelle prepared via RAFT polymerization and Diels-Alder "click" chemistry.

PubMed

Banerjee, Sovan Lal; Singha, Nikhil K

2017-12-06

Amphiphilic diblock copolymers of poly(furfuryl methacrylate) (PFMA) with cationic poly(2-(methacryloyloxy)ethyltrimethyl ammonium chloride) (PFMA-b-PMTAC) and anionic poly(sodium 4-vinylbenzenesulfonate) (PFMA-b-PSS) were prepared via reversible addition fragmentation chain-transfer (RAFT) polymerization by using PFMA as a macro-RAFT agent. The formation of the block copolymer was confirmed by FTIR and 1 H NMR analyses. In water, the amphiphilic diblock copolymers, (PFMA-b-PMTAC) and (PFMA-b-PSS), formed micelles with PFMA in the core and the rest of the hydrophilic polymers like PMTAC and PSS in the corona. The PFMA core was crosslinked by using Diels-Alder (DA) "Click" chemistry in water at 60 °C where bismaleimide acted as a crosslinker. Afterwards, both the core crosslinked micelles were mixed at an almost equal charge ratio which was determined by zeta potential analysis to prepare the self-assembled hydrogel. The de-crosslinking of the hydrophobic PFMA core in the self-assembled hydrogel via rDA reaction took place at 165 °C as determined from DSC analysis. This hydrogel showed self-healing behavior using ionic interaction (in the presence of water) and DA chemistry (in the presence of heat).

Laponite crosslinked starch/polyvinyl alcohol hydrogels by freezing/thawing process and studying their cadmium ion absorption.

PubMed

Yu, Chen; Tang, Xiaozhi; Liu, Shaowei; Yang, Yuling; Shen, Xinchun; Gao, Chengcheng

2018-05-22

In this study, Laponite RD (LRD) cross-linked hydrogels consisting of starch, polyvinyl alcohol (PVA) were prepared by freezing/thawing process and the influence of LRD content on structure and properties of hydrogels was investigated. FTIR showed a new structure of hydrogen bonding might result from cross-linking reactions between LRD and polymers. X-ray diffraction (XRD) analysis showed that high degree of exfoliation of LRD clay layers had occurred during the preparation of hydrogels. The synergistic effect of physical cross-linking by freeze/thaw cycles and by LRD led to more porous, uniform and stable network, which was shown in SEM images. The melting temperature decreased and thermal stability got improved with the increase of LRD content. Reswelling ratios of hydrogels had the highest value when LRD content was 10%. Additionally, cadmium ion absorption capacity of the hydrogel was studied and the results showed that increasing the concentration of LRD increased absorption ratio and amount of Cd 2+ ion in the solution. In a word, LRD could be used as a physical crosslinker and reinforced agent for starch-PVA based hydrogels and the formed hydrogels could be used as novel type and high capacity absorbent materials in heavy metal removing processes. Copyright © 2018. Published by Elsevier B.V.

Biohybrid Fibro-Porous Vascular Scaffolds: Effect of Crosslinking on Properties

PubMed Central

Nozik, Danna; Patel, Harsh; Singh, Raj K.; Vohra, Yogesh K.

2015-01-01

Tubular grafts were fabricated from blends of polycaprolactone (PCL) and poly(glycolide -co-caprolactone) (PGC) polymers and coated with an extracellular matrix containing collagens, laminin, and proteoglycans, but not growth factors (HuBiogel™). Multifunctional scaffolds from polymer blends and membrane proteins provide the necessary biomechanics and biological functions for tissue regeneration. Two crosslinking agents, a natural crosslinker namely genipin (Gp) and a carbodiimide reagent namely 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC), were used for further stabilizing the protein matrix and the effect of crosslinking was evaluated for structural, morphological, mechanical properties using SEM, DSC and DMA. SEM images and fiber diameter distribution showed fiber-size between 0.2 µm to 1 µm with the majority of fiber diameters being under 500 nm, indicating upper range of protein fiber-sizes (for example, collagen fibers in extracellular matrix are in 50 to 500 nm diameter range). HB coating did not affect the mechanical properties, but increased its hydrophilicity of the graft. Overall data showed that PCL/PGC blends with 3:1 mass ratio exhibited mechanical properties comparable to those of human native arteries (tensile strength of 1–2 MPa and Young’s modulus of <10 MPa). Additionally, the effect of crosslinking on coating stability was investigated to assure the retention of proteins on scaffold for effective cell-matrix interactions. PMID:26082566

Biohybrid Fibro-Porous Vascular Scaffolds: Effect of Crosslinking on Properties.

PubMed

Thomas, Vinoy; Nozik, Danna; Patel, Harsh; Singh, Raj K; Vohra, Yogesh K

Tubular grafts were fabricated from blends of polycaprolactone (PCL) and poly(glycolide -co-caprolactone) (PGC) polymers and coated with an extracellular matrix containing collagens, laminin, and proteoglycans, but not growth factors (HuBiogel™). Multifunctional scaffolds from polymer blends and membrane proteins provide the necessary biomechanics and biological functions for tissue regeneration. Two crosslinking agents, a natural crosslinker namely genipin (Gp) and a carbodiimide reagent namely 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC), were used for further stabilizing the protein matrix and the effect of crosslinking was evaluated for structural, morphological, mechanical properties using SEM, DSC and DMA. SEM images and fiber diameter distribution showed fiber-size between 0.2 µm to 1 µm with the majority of fiber diameters being under 500 nm, indicating upper range of protein fiber-sizes (for example, collagen fibers in extracellular matrix are in 50 to 500 nm diameter range). HB coating did not affect the mechanical properties, but increased its hydrophilicity of the graft. Overall data showed that PCL/PGC blends with 3:1 mass ratio exhibited mechanical properties comparable to those of human native arteries (tensile strength of 1-2 MPa and Young's modulus of <10 MPa). Additionally, the effect of crosslinking on coating stability was investigated to assure the retention of proteins on scaffold for effective cell-matrix interactions.

The mechanism of collagen cross-linking in diabetes: a puzzle nearing resolution.

PubMed

Monnier, V M; Glomb, M; Elgawish, A; Sell, D R

1996-07-01

Considerable interest has been focused in recent years on the mechanism of collagen cross-linking by high glucose in vitro and in vivo. Experiments in both diabetic humans and in animals have shown that over time collagen becomes less soluble, less digestible by collagenase, more stable to heat-induced denaturation, and more glycated. In addition, collagen becomes more modified by advanced products of the Maillard reaction, i.e., immunoreactive advanced glycation end products and the glycoxidation markers carboxymethyllysine and pentosidine. Mechanistic studies have shown that collagen cross-linking in vitro can be uncoupled from glycation by the use of antioxidants and chelating agents. Experiments in the authors' laboratory revealed that approximately 50% of carboxymethyllysine formed in vitro originates from pathways other than oxidation of Amadori products, i.e., most likely the oxidation of Schiff base-linked glucose. In addition, the increase in thermal stability of rat tail tendons exposed to high glucose in vitro or in vivo was found to strongly depend on H2O2 formation. The final missing piece of the puzzle is that of the structure of the major cross-link. We speculate that it is a nonfluorescent nonultraviolet active cross-link between two lysine residues, which includes a fragmentation product of glucose linked in a nonreducible bond labile to both strong acids and bases.

Effect of curcumin caged silver nanoparticle on collagen stabilization for biomedical applications.

PubMed

Srivatsan, Kunnavakkam Vinjimur; Duraipandy, N; Begum, Shajitha; Lakra, Rachita; Ramamurthy, Usha; Korrapati, Purna Sai; Kiran, Manikantan Syamala

2015-04-01

The current study aims at understanding the influence of curcumin caged silver nanoparticle (CCSNP) on stability of collagen. The results indicated that curcumin caged silver nanoparticles efficiently stabilize collagen, indicated by enhanced tensile strength, fibril formation and viscosity. The tensile strength of curcumin caged silver nanoparticle cross-linked collagen and elongation at break was also found to be higher than glutaraldehyde cross-linked collagen. The physicochemical characteristics of curcumin caged nanoparticle cross-linked collagen exhibited enhanced strength. The thermal properties were also good with both thermal degradation temperature and hydrothermal stability higher than native collagen. CD analysis showed no structural disparity in spite of superior physicochemical properties suggesting the significance of curcumin caged nanoparticle mediated cross-linking. The additional enhancement in the stabilization of collagen could be attributed to multiple sites for interaction with collagen molecule provided by curcumin caged silver nanoparticles. The results of cell proliferation and anti-microbial activity assays indicated that curcumin caged silver nanoparticles promoted cell proliferation and inhibited microbial growth making it an excellent biomaterial for wound dressing application. The study opens scope for nano-biotechnological strategies for the development of alternate non-toxic cross-linking agents facilitating multiple site interaction thereby improving therapeutic values to the collagen for biomedical application. Copyright © 2015 Elsevier B.V. All rights reserved.

[PREPARATION AND BIOCOMPATIBILITY OF IN SITU CROSSLINKING HYALURONIC ACID HYDROGEL].

PubMed

Liang, Jiabi; Li, Jun; Wang, Ting; Liang, Yuhong; Zou, Xuenong; Zhou, Guangqian; Zhou, Zhiyu

2016-06-08

To fabricate in situ crosslinking hyaluronic acid hydrogel and evaluate its biocompatibility in vitro. The acrylic acid chloride and polyethylene glycol were added to prepare crosslinking agent polyethylene glycol acrylate (PEGDA), and the molecular structure of PEGDA was analyzed by Flourier transformation infrared spectroscopy and 1H nuclear magnetic resonance spectroscopy. Hyaluronic acid hydrogel was chemically modified to prepare hyaluronic acid thiolation (HA-SH). And the degree of HA-SH was analyzed qualitatively and quantitatively by Ellman method. HA-SH solution in concentrations ( W/V ) of 0.5%, 1.0%, and 1.5% and PEGDA solution in concentrations ( W/V ) of 2%, 4%, and 6% were prepared with PBS. The two solutions were mixed in different ratios, and in situ crosslinking hyaluronic acid hydrogel was obtained; the crosslinking time was recorded. The cellular toxicity of in situ crosslinking hyaluronic acid hydrogel (1.5% HA-SH and 4% PEGDA mixed) was tested by L929 cells. Meanwhile, the biocompatibility of hydrogel was tested by co-cultured with human bone mesenchymal stem cells (hBMSCs). Flourier transformation infrared spectroscopy showed that most hydroxyl groups were replaced by acrylate groups; 1H nuclear magnetic resonance spectroscopy showed 3 characteristic peaks of hydrogen representing acrylate and olefinic bond at 5-7 ppm. The thiolation yield of HA-SH was 65.4%. In situ crosslinking time of hyaluronic acid hydrogel was 2 to 70 minutes in the PEGDA concentrations of 2%-6% and HA-SH concentrations of 0.5%-1.5%. The hyaluronic acid hydrogel appeared to be transparent. The toxicity grade of leaching solution of hydrogel was grade 1. hBMSCs grew well and distributed evenly in hydrogel with a very high viability. In situ crosslinking hyaluronic acid hydrogel has low cytotoxicity, good biocompatibility, and controllable crosslinking time, so it could be used as a potential tissue engineered scaffold or repairing material for tissue regeneration.

The Impact of HA Oligomer Content on Physical, Mechanical, and Biologic Properties of Divinyl Sulfone-Crosslinked HA Hydrogels

PubMed Central

Ibrahim, Samir; Kang, Qian K; Ramamurthi, Anand

2009-01-01

In recent studies, we showed that exogenous hyaluronic acid oligomers (HA-o) stimulate functional endothelialization, though native long-chain HA is more bioinert and possibly more biocompatible. Thus, in this study, hydrogels containing high molecular weight (HMW) HA (1×106 Da) and HA oligomer mixtures (HA-o: 0.75–10 kDa) were created by crosslinking with divinyl sulfone (DVS). The incorporation of HA oligomers was found to compromise the physical and mechanical properties of the gels (rheology, apparent crosslinking density, swelling ratio, degradation) and to very mildly enhance inflammatory cell recruitment in vivo; increasing the DVS crosslinker content within the gels in general, had the opposite effect, though the relatively high concentration of DVS within these gels (necessary to create a solid gel) also stimulated a mild sub-cutaneous inflammatory response in vivo and VCAM-1 expression by ECs cultured atop; ICAM-expression levels remained very low irrespective extent of DVS crosslinking or HA-o content. The greatest EC attachment and proliferation (MTT assay) was observed on gels that contained the highest amount of HA-o. The study shows that the beneficial EC response to HA oligomers and biocompatibility of HA is mostly unaltered by their chemical derivatization and crosslinking into a hydrogel. However, the study also demonstrates that the relatively high concentrations of DVS, necessary to create solid gels, compromises their biocompatibility. Moreover, the poor mechanics of even these heavily crosslinked gels, in the context of vascular implantation, necessitates the investigation of other, more appropriate crosslinking agents. Alternately, the outcomes of this study may be used to guide an approach based on chemical immobilization and controlled surface-presentation of both bioactive HA oligomers and more biocompatible HMW HAon synthetic or tissue engineered grafts already in use, without the use of a crosslinker, so that improved, predictable, and functional endothelialization can be achieved, and the need to create a mechanically compliant biomaterial for standalone use, circumvented. PMID:20186732

Sensitization of melanoma cells to alkylating agent-induced DNA damage and cell death via orchestrating oxidative stress and IKKβ inhibition.

PubMed

Tse, Anfernee Kai-Wing; Chen, Ying-Jie; Fu, Xiu-Qiong; Su, Tao; Li, Ting; Guo, Hui; Zhu, Pei-Li; Kwan, Hiu-Yee; Cheng, Brian Chi-Yan; Cao, Hui-Hui; Lee, Sally Kin-Wah; Fong, Wang-Fun; Yu, Zhi-Ling

2017-04-01

Nitrosourea represents one of the most active classes of chemotherapeutic alkylating agents for metastatic melanoma. Treatment with nitrosoureas caused severe systemic side effects which hamper its clinical use. Here, we provide pharmacological evidence that reactive oxygen species (ROS) induction and IKKβ inhibition cooperatively enhance nitrosourea-induced cytotoxicity in melanoma cells. We identified SC-514 as a ROS-inducing IKKβ inhibitor which enhanced the function of nitrosoureas. Elevated ROS level results in increased DNA crosslink efficiency triggered by nitrosoureas and IKKβ inhibition enhances DNA damage signals and sensitizes nitrosourea-induced cell death. Using xenograft mouse model, we confirm that ROS-inducing IKKβ inhibitor cooperates with nitrosourea to reduce tumor size and malignancy in vivo. Taken together, our results illustrate a new direction in nitrosourea treatment, and reveal that the combination of ROS-inducing IKKβ inhibitors with nitrosoureas can be potentially exploited for melanoma therapy. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

3′-Phosphoadenosine 5′-phosphosulfate synthase 1 (PAPSS1) knockdown sensitizes non-small cell lung cancer cells to DNA damaging agents

PubMed Central

Leung, Ada W. Y.; Dragowska, Wieslawa H.; Ricaurte, Daniel; Kwok, Brian; Mathew, Veena; Roosendaal, Jeroen; Ahluwalia, Amith; Warburton, Corinna; Laskin, Janessa J.; Stirling, Peter C.; Qadir, Mohammed A.; Bally, Marcel B.

2015-01-01

Standard treatment for advanced non-small cell lung cancer (NSCLC) with no known driver mutation is platinum-based chemotherapy, which has a response rate of only 30–33%. Through an siRNA screen, 3′-phosphoadenosine 5′-phosphosulfate (PAPS) synthase 1 (PAPSS1), an enzyme that synthesizes the biologically active form of sulfate PAPS, was identified as a novel platinum-sensitizing target in NSCLC cells. PAPSS1 knockdown in combination with low-dose (IC10) cisplatin reduces clonogenicity of NSCLC cells by 98.7% (p < 0.001), increases DNA damage, and induces G1/S phase cell cycle arrest and apoptosis. PAPSS1 silencing also sensitized NSCLC cells to other DNA crosslinking agents, radiation, and topoisomerase I inhibitors, but not topoisomerase II inhibitors. Chemo-sensitization was not observed in normal epithelial cells. Knocking out the PAPSS1 homolog did not sensitize yeast to cisplatin, suggesting that sulfate bioavailability for amino acid synthesis is not the cause of sensitization to DNA damaging agents. Rather, sensitization may be due to sulfation reactions involved in blocking the action of DNA damaging agents, facilitating DNA repair, promoting cancer cell survival under therapeutic stress or reducing the bioavailability of DNA damaging agents. Our study demonstrates for the first time that PAPSS1 could be targeted to improve the activity of multiple anticancer agents used to treat NSCLC. PMID:26220590

Multiple binding modes of substrate to the catalytic RNA subunit of RNase P from Escherichia coli.

PubMed Central

Pomeranz Krummel, D A; Altman, S

1999-01-01

M1 RNA that contained 4'-thiouridine was photochemically cross-linked to different substrates and to a product of the reaction it governs. The locations of the cross-links in these photochemically induced complexes were identified. The cross-links indicated that different substrates share some contacts but have distinct binding modes to M1 RNA. The binding of some substrates also results in a substrate-dependent conformational change in the enzymatic RNA, as evidenced by the appearance of an M1 RNA intramolecular cross-link. The identification of the cross-links between M1 RNA and product indicate that they are shared with only one of the three cross-linked E-S complexes that were identified, an indication of noncompetitive inhibition by the product. We also examined whether the cross-linked complexes between M1 RNA and substrate(s) or product are altered in the presence of the enzyme's protein cofactor (C5 protein) and in the presence of different concentrations of divalent metal ions. C5 protein enhanced the yield of certain M1 RNA-substrate cross-linked complexes for both wild-type M1 RNA and a deletion mutant of M1 RNA (delta[273-281]), but not for the M1 RNA-product complex. High concentrations of Mg2+ increased the yield of all M1 RNA-substrate complexes but not the M1 RNA-product complex. PMID:10445877

Redox-responsive solid lipid microparticles composed of octadecyl acrylate and allyl disulfide.

PubMed

Kim, Tae Hoon; Kim, Jin-Chul

2018-04-01

Redox-responsive solid lipid microparticles were prepared by an emulsification photo-polymerization method. Octadecyl acrylate (ODA) and a cross-linker (i.e. allyl disulfide (ADS) and octadiene (ODE)) were dissolved in dichloromethane, it was emulsified in poly(vinyl alcohol) solution, and the resulting O/W emulsion was irradiated with UV light. On the scanning electron microscope micrographs, the microparticles were sphere-like and they were not markedly different from the oil droplets in size. Using the atomic compositions analyzed by energy dispersive X-ray spectroscopy, the ODA to cross-linker molar ratio of ODA/ADS microparticles and ODA/ODE ones were calculated to be 1:0.13 and 1:0.15, respectively. In the FT-IR spectra of the microparticles, the signal of the vinyl group was hardly detected, implying that the monomer and the cross-linkers participated in the photo-polymerization. In differential scanning calorimetry study, ODA/ADS microparticles and ODA/ODE ones exhibited their endothermic peaks around 42.9 and 41.3 °C, respectively, possibly due to the melting of polymeric ODA. Dithiothreitol (DTT, a reducing agent) concentration had little effect on the release degree of dye loaded in ODA/ODE microparticles. Whereas, DTT concentration had a significant effect on the release degree of dye loaded in ODA/ADS microparticles. The release degree at 26 °C was weakly affected by DTT concentration. When the temperature was 37 °C, DTT concentration had a strong effect on the release degree. The disulfide cross-linker (i.e. ADS) can be broken to thiol compounds by the reducing agent, resulting in an increase in the release degree.

Evaluation of Gentamicin and Lidocaine Release Profile from Gum Acacia-crosslinked-poly(2-hydroxyethylmethacrylate)-carbopol Based Hydrogels.

PubMed

Singh, Baljit; Dhiman, Abhishek

2017-01-01

No doubt, the prevention of infection is an indispensable aspect of the wound management, but, simultaneous wound pain relief is also required. Therefore, herein this article, incorporation of antibiotic agent 'gentamicin' and pain relieving agent 'lidocaine' into hydrogel wound dressings, prepared by using acacia gum, carbopol and poly(2-hydroxyethylmethacrylate) polymers, has been carried out. The hydrogels were evaluated as a drug carrier for model drugs gentamicin and lidocaine. Synthesis of hydrogel wound dressing was carried out by free radical polymerization technique. The drug loading was carried out by swelling equilibrium method and gel strength of hydrogels was measured by a texture analyzer. Porous microstructure of the hydrogel was observed in cryo-SEM images. The hydrogel showed mesh size 37.29 nm, cross-link density 2.19× 10-5 mol/cm3, molecular weight between two cross-links 60.25× 10-3 g/mol and gel strength 0.625±0.112 N in simulated wound fluid. It is concluded that the pH of swelling medium has influenced the network structure of hydrogel i.e., molecular weight of the polymer chain between two neighboring cross links, crosslink density and the corresponding mesh size. A good correlation was established between gel strength and network parameters. Cryo-SEM images showed porous morphology of hydrogels. These hydrogels were found to be biodegradable and antimicrobial in nature. Drug release occurred through Fickian diffusion mechanism and release profile was best fitted in first order model. Overall it is concluded that modification in GA has led to formation of a porous hydrogels for wound dressing applications. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Deformable microparticles with multiple functions for drug delivery and device testing

NASA Astrophysics Data System (ADS)

Thula, Taili T.

Since the HIV epidemic of the 1990s, researchers have attempted to develop a red blood cell analog. Even though some of these substitutes are now in Phase III of clinical trials, their use is limited by side effects and short half-life in the human body. As a result, there is still a need for an effective erythrocyte analog with minimum immunogenic and side effects, so that it can be used for multiple applications. Finding new approaches to develop more efficient blood substitutes will not only bring valuable advances in the clinical approach, but also in the area of in vitro testing of medical devices. We examined the feasibility of creating a deformable multi-functional, biodegradable, biocompatible particle for applications in drug delivery and device testing. As a preliminary evaluation, we synthesized different types of microcapsules using natural and synthetic polymers, various cross-linking agents, and diverse manufacturing techniques. After fully characterizing of each system, we determined the most promising red blood cell analog in terms of deformability, stability and toxicity. We also examined the encapsulation and release of bovine serum albumin (BSA) within these deformable particles. After removal of cross-linkers, zinc- and copper-alginate microparticles surrounded by multiple polyelectrolyte layers of chitosan oligosaccharide and alginate were deformable and remained stable under physiological pressures applied by the micropipette technique. In addition, multiple coatings decreased toxicity of heavy-metal crosslinked particles. BSA encapsulation and release from chitosan-alginate microspheres were contingent on the crosslinker and number of polyelectrolyte coatings, respectively. Further rheological studies are needed to determine how closely these particles simulate the behavior of erythrocytes. Also, studies on the encapsulation and release of different proteins, including hemoglobin, are needed to establish the desired controlled release of bioactive agents for the proposed delivery system.

Two Dimensional Polymer That Generates Nitric Oxide.

DOEpatents

McDonald, William F.; Koren, Amy B.

2005-10-04

A polymeric composition that generates nitric oxide and a process for rendering the surface of a substrate nonthrombogenic by applying a coating of the polymeric composition to the substrate are disclosed. The composition comprises: (1) a crosslinked chemical combination of (i) a polymer having amino group-containing side chains along a backbone forming the polymer, and (ii) a crosslinking agent containing functional groups capable of reacting with the amino groups; and (2) a plurality of nitric oxide generating functional groups associated with the crosslinked chemical combination. Once exposed to a physiological environment, the coating generates nitric oxide thereby inhibiting platelet aggregation. In one embodiment, the nitric oxide generating functional groups are provided by a nitrated compound (e.g., nitrocellulose) imbedded in the polymeric composition. In another embodiment, the nitric oxide generating functional groups comprise N2O2- groups covalently bonded to amino groups on the polymer.

Synthesis of Photocrosslinkable and Amine Containing Multifunctional Nanoparticles via Polymerization-Induced Self-Assembly.

PubMed

Huang, Jianbing; Li, Decai; Liang, Hui; Lu, Jiang

2017-08-01

Photo-crosslinkable and amine-containing block copolymer nanoparticles are synthesized via reversible addition-fragmentation chain transfer (RAFT) polymerization-induced self-assembly of a multifunctional core-forming monomer, 2-((3-(4-(diethylamino)phenyl)acryloyl)oxy)ethyl methacrylate (DEMA), using poly(2-hydroxypropyl methacrylate) macromolecular chain transfer agent as a steric stabilizer in methanol at 65 °C. By tuning the chain length of PDEMA, a range of nanoparticle morphologies (sphere, worm, and vesicle) can be obtained. Since cinnamate groups can easily undergo a [2 + 2] cycloaddition of the carbon-carbon double bonds upon UV irradiation, the as-prepared block copolymer nanoparticles are readily stabilized by photo-crosslinking to produce anisotropic nanoparticles. The crosslinked block copolymer nanoparticles can be used as templates for in situ formation polymer/gold hybrid nanoparticles. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Characterization of the Deoxyguanosine–Lysine Cross-Link of Methylglyoxal

PubMed Central

2015-01-01

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

Cytotoxicity of silica-glass fiber reinforced composites.

PubMed

Meriç, Gökçe; Dahl, Jon E; Ruyter, I Eystein

2008-09-01

Silica-glass fiber reinforced polymers can be used for many kinds of dental applications. The fiber reinforcement enhances the mechanical properties of the polymers, and they have good esthetic attributes. There is good initial bonding of glass fibers to polymers via an interface made from silane coupling agents. The aim of this in vitro study was to determine the cytotoxicity of two polymers reinforced with two differently sized silica-glass fibers before and after thermal cycling. Cytotoxicity of the polymers without fibers was also evaluated. Two different resin mixtures (A and B) were prepared from poly(vinyl chloridecovinylacetate) powder and poly(methyl methacrylate) (PMMA) dissolved in methyl methacrylate and mixed with different cross-linking agents. The resin A contained the cross-linking agents ethylene glycol dimethacrylate and 1,4-butanediol dimethacrylate, and for resin B diethylene glycol dimethacrylate was used. Woven silica-glass fibers were used for reinforcement. The fibers were sized with either linear poly(butyl methacrylate)-sizing or cross-linking PMMA-sizing. Cytotoxicity was evaluated by filter diffusion test (ISO 7405:1997) of newly made and thermocycled test specimens. Extracts were prepared according to ISO 10993-12 from newly made and from thermocycled specimens and tested by the MTT assay. The results from the experiments were statistically analyzed by one-way ANOVA and Tukey's test (rho<0.05). The filter diffusion test disclosed no change in staining intensity at the cell-test sample contact area indicating non-cytotoxicity in all experimental groups. Cell viability assessed by MTT assay was more than 90% in all experimental groups. All are non-cytotoxic. It can be concluded that correctly processed heat polymerized silica-glass fiber reinforced polymers induced no cytotoxicity and that thermocycling did not alter this property.

Elevated levels of STAT1 in Fanconi anemia group A lymphoblasts correlate with the cells’ sensitivity to DNA interstrand crosslinking drugs

PubMed Central

Prieto-Remón, Inés; Sánchez-Carrera, Dámaso; López-Duarte, Mónica; Richard, Carlos; Pipaón, Carlos

2013-01-01

Progressive bone marrow failure starting in the first decade of life is one of the main characteristics of Fanconi anemia. Along with the bone marrow failure, this pathology is characterized by congenital malformations, endocrine dysfunction and an extraordinary predisposition to develop cancer. The fact that hematopoietic progenitor cells from subjects with Fanconi anemia are sensitive to both DNA-interstrand crosslinking agents and inflammatory cytokines, which are aberrantly overproduced in these patients, has led to different explanations for the causes of the bone marrow failure. We analyzed STAT1 expression in lymphoblastoid cell lines derived from patients with Fanconi anemia group A and correlated this with aspects of the Fanconi anemia phenotype such as sensitivity to genotoxic agents or to inhibitory cytokines. We provide evidence of overexpression of STAT1 in FANCA-deficient cells which has both transcriptional and post-translational components, and is related to the constitutive activation of ERK in Fanconi anemia group A cells, since it can be reverted by treatment with U0126. STAT1 phosphorylation was not defective in the lymphoblasts, so these cells accumulated higher levels of active STAT1 in response to interferon gamma, probably in relation to their greater sensitivity to this cytokine. On the other hand, inhibition of STAT1 by genetic or chemical means reverted the hypersensitivity of Fanconi anemia group A lymphoblasts to DNA interstrand crosslinking agents. Our data provide an explanation for the mixed sensitivity of Fanconi anemia group A cells to both genotoxic stress and inflammatory cytokines and indicate new targets for the treatment of bone marrow failure in these patients. PMID:23585528

Cytokine overproduction and crosslinker hypersensitivity are unlinked in Fanconi anemia macrophages.

PubMed

Garbati, Michael R; Hays, Laura E; Rathbun, R Keaney; Jillette, Nathaniel; Chin, Kathy; Al-Dhalimy, Muhsen; Agarwal, Anupriya; Newell, Amy E Hanlon; Olson, Susan B; Bagby, Grover C

2016-03-01

The Fanconi anemia proteins participate in a canonical pathway that repairs cross-linking agent-induced DNA damage. Cells with inactivated Fanconi anemia genes are universally hypersensitive to such agents. Fanconi anemia-deficient hematopoietic stem cells are also hypersensitive to inflammatory cytokines, and, as importantly, Fanconi anemia macrophages overproduce such cytokines in response to TLR4 and TLR7/8 agonists. We questioned whether TLR-induced DNA damage is the primary cause of aberrantly regulated cytokine production in Fanconi anemia macrophages by quantifying TLR agonist-induced TNF-α production, DNA strand breaks, crosslinker-induced chromosomal breakage, and Fanconi anemia core complex function in Fanconi anemia complementation group C-deficient human and murine macrophages. Although both M1 and M2 polarized Fanconi anemia cells were predictably hypersensitive to mitomycin C, only M1 macrophages overproduced TNF-α in response to TLR-activating signals. DNA damaging agents alone did not induce TNF-α production in the absence of TLR agonists in wild-type or Fanconi anemia macrophages, and mitomycin C did not enhance TLR responses in either normal or Fanconi anemia cells. TLR4 and TLR7/8 activation induced cytokine overproduction in Fanconi anemia macrophages. Also, although TLR4 activation was associated with induced double strand breaks, TLR7/8 activation was not. That DNA strand breaks and chromosome breaks are neither necessary nor sufficient to account for the overproduction of inflammatory cytokines by Fanconi anemia cells suggests that noncanonical anti-inflammatory functions of Fanconi anemia complementation group C contribute to the aberrant macrophage phenotype and suggests that suppression of macrophage/TLR hyperreactivity might prevent cytokine-induced stem cell attrition in Fanconi anemia. © Society for Leukocyte Biology.

Use of 2,5-dimethyl-2,5-hexane diamine as a curing agent for epoxy resins

DOEpatents

Rinde, J.A.; Newey, H.A.

1981-02-24

Primary diamines are disclosed of the formula shown in a diagram wherein R is a straight chain saturated hydrocarbon of 2 to 4 carbons, a disubstituted benzene ring, or disubstituted dibenzomethane for use as a curing agent for epoxy resins. These curing agents can be used to form epoxy resin mixtures useful in filament winding and pre-impregnated fiber molding and in formulating film adhesives, powder coatings and molding powders. The epoxy mixtures form for such uses as room temperature non-reacting, intermediate stable state which has a latent cross-linking capability.

Use of 2,5-dimethyl-2,5-hexane diamine as a curing agent for epoxy resins

DOEpatents

Rinde, James A. [Livermore, CA; Newey, Herbert A. [Lafayette, CA

1981-02-24

Primary diamines of the formula ##STR1## wherein R is a straight chain saturated hydrocarbon of 2 to 4 carbons, a disubstituted benzene ring, or disubstituted dibenzo methane for use as a curing agent for epoxy resins. These curing agents can be used to form epoxy resin mixtures useful in filament winding and pre-impregnated fiber molding and in formulating film adhesives, powder coatings and molding powders. The epoxy mixtures form for such uses as room temperature non-reacting, intermediate stable state which has a latent cross-linking capability.

Aptamer-crosslinked microbubbles: smart contrast agents for thrombin-activated ultrasound imaging.

PubMed

Nakatsuka, Matthew A; Mattrey, Robert F; Esener, Sadik C; Cha, Jennifer N; Goodwin, Andrew P

2012-11-27

Thrombosis, or malignant blood clotting, is associated with numerous cardiovascular diseases and cancers. A microbubble contrast agent is presented that produces ultrasound harmonic signal only when exposed to elevated thrombin levels. Initially silent microbubbles are activated in the presence of both thrombin-spiked and freshly clotting blood in three minutes with detection limits of 20 nM thrombin and 2 aM microbubbles. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Kinetics of micronucleus induction and cytotoxicity caused by distinct antineoplastics and alkylating agents in vivo.

PubMed

Morales-Ramírez, Pedro; Vallarino-Kelly, Teresita; Cruz-Vallejo, Virginia

2014-01-30

This mini-review aims to compare the differences in the kinetics of the induction of micronucleated polychromatic erythrocytes (MN-PCE) and cytotoxicity by distinct antineoplastic and genotoxic agents in murine peripheral blood in vivo and to correlate these kinetics with the underlying processes. Comparisons were carried out using our previously obtained data with nominal doses causing similar levels of cytotoxicity, as measured in terms reduction of PCE. The aneuploidogens caused the most rapid induction of MN-PCEs and had the highest rates of cytotoxicity and genotoxicity. The promutagens cyclophosphamide and dimethylnitrosamine showed the most delayed responses and had the lowest genotoxic and cytotoxic efficiencies. DNA crosslinking agents had a similar delay of 4-5 h, greater than those of aneuploidogens, but differed in their cytotoxic and genotoxic efficiencies. Methylnitrosourea and 5-aza-cytidine caused greater delays than crosslinking agents. These delays can be due to the methylnitrosourea-mediated induction of formation of mono alkyl adducts which are interpreted as mismatches during DNA duplication, whereas 5-aza-cytidine requires incorporation into the DNA to induce breakage. This review allows us to conclude that the requirement for metabolic activation and the mechanisms of DNA breakage and of micronucleus induction are the main factors that affect the time of maximal MN-PCE induction. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

A Study into the Collision-induced Dissociation (CID) Behavior of Cross-Linked Peptides*

PubMed Central

Giese, Sven H.; Fischer, Lutz; Rappsilber, Juri

2016-01-01

Cross-linking/mass spectrometry resolves protein–protein interactions or protein folds by help of distance constraints. Cross-linkers with specific properties such as isotope-labeled or collision-induced dissociation (CID)-cleavable cross-linkers are in frequent use to simplify the identification of cross-linked peptides. Here, we analyzed the mass spectrometric behavior of 910 unique cross-linked peptides in high-resolution MS1 and MS2 from published data and validate the observation by a ninefold larger set from currently unpublished data to explore if detailed understanding of their fragmentation behavior would allow computational delivery of information that otherwise would be obtained via isotope labels or CID cleavage of cross-linkers. Isotope-labeled cross-linkers reveal cross-linked and linear fragments in fragmentation spectra. We show that fragment mass and charge alone provide this information, alleviating the need for isotope-labeling for this purpose. Isotope-labeled cross-linkers also indicate cross-linker-containing, albeit not specifically cross-linked, peptides in MS1. We observed that acquisition can be guided to better than twofold enrich cross-linked peptides with minimal losses based on peptide mass and charge alone. By help of CID-cleavable cross-linkers, individual spectra with only linear fragments can be recorded for each peptide in a cross-link. We show that cross-linked fragments of ordinary cross-linked peptides can be linearized computationally and that a simplified subspectrum can be extracted that is enriched in information on one of the two linked peptides. This allows identifying candidates for this peptide in a simplified database search as we propose in a search strategy here. We conclude that the specific behavior of cross-linked peptides in mass spectrometers can be exploited to relax the requirements on cross-linkers. PMID:26719564

Screening for modulators of cisplatin sensitivity: unbiased screens reveal common themes.

PubMed

Nijwening, Jeroen H; Kuiken, Hendrik J; Beijersbergen, Roderick L

2011-02-01

Cisplatin is a widely used chemotherapeutic agent to treat a variety of solid tumors. The cytotoxic mode of action of cisplatin is mediated by inducing conformational changes in DNA including intra- and inter-strand crosslink adducts. Recognition of these adducts results in the activation of the DNA damage response resulting in cell cycle arrest, repair, and potentially, apoptosis. Despite the clinical efficacy of cisplatin, many tumors are either intrinsically resistant or acquire resistance during treatment. The identification of cisplatin drug response modulators can help us understand these resistance mechanisms, provide biomarkers for treatment strategies, or provide drug targets for combination therapy. Here we discuss functional genetic screens, including one performed by us, set up to identify genes whose inhibition results in increased sensitivity to cisplatin. In summary, the validated genes identified in these screens mainly operate in DNA damage response including nucleotide excision repair, translesion synthesis, and homologous recombination.

Tc-99m Labeled carrier for imaging

DOEpatents

Henze, Eberhard

1984-01-01

Novel radionuclide imaging agents, having particular application for lymphangiography are provided by non-covalently binding Tc-99m to a pharmaceutically acceptable cross-linked polysaccharide. Upon injection of the Tc-99m labeled polysaccharide into the blood stream, optimum contrast can be obtained within one hour.

Novel aminobenzyl and imidobenzyl benzenes

NASA Technical Reports Server (NTRS)

Bell, V. L.; Pratt, J. R.; Stump, B. L.

1976-01-01

Compounds are useful as intermediates for several classes of polymers. Amines can function as cross-linking agents for epoxide and urethane polymers, as well as intermediates for synthesis of thermally-stable addition-type polyimides. Imide derivatives can be obtained by reacting amines with certain monoanhydrides containing olefinic unsaturation.

Biomimetic fiber mesh scaffolds based on gelatin and hydroxyapatite nano-rods: Designing intrinsic skills to attain bone reparation abilities.

PubMed

Sartuqui, Javier; Gravina, A Noel; Rial, Ramón; Benedini, Luciano A; Yahia, L'Hocine; Ruso, Juan M; Messina, Paula V

2016-09-01

Intrinsic material skills have a deep effect on the mechanical and biological performance of bone substitutes, as well as on its associated biodegradation properties. In this work we have manipulated the preparation of collagenous derived fiber mesh frameworks to display a specific composition, morphology, open macroporosity, surface roughness and permeability characteristics. Next, the effect of the induced physicochemical attributes on the scaffold's mechanical behavior, bone bonding potential and biodegradability were evaluated. It was found that the scaffold microstructure, their inherent surface roughness, and the compression strength of the gelatin scaffolds can be modulated by the effect of the cross-linking agent and, essentially, by mimicking the nano-scale size of hydroxyapatite in natural bone. A clear effect of bioactive hydroxyapatite nano-rods on the scaffolds skills can be appreciated and it is greater than the effect of the cross-linking agent, offering a huge perspective for the upcoming progress of bone implant technology. Copyright © 2016 Elsevier B.V. All rights reserved.

Towards producing novel fish gelatin films by combination treatments of ultraviolet radiation and sugars (ribose and lactose) as cross-linking agents.

PubMed

Bhat, Rajeev; Karim, A A

2014-07-01

Developing novel fish gelatin films with better mechanical properties than mammalian gelatin is a challenging but promising endeavor. Studies were undertaken to produce fish gelatin films by combining treatments with different sugars (ribose and lactose) followed 'by' 'and' ultraviolet (UV) radiation, as possible cross-linking agents. Increase in tensile strength and percent elongation at break was recorded, which was more significant in films without sugars that were exposed to UV radiation. Films with added ribose showed decreased solubility after UV treatment and exhibited higher swelling percentage than films with added lactose, which readily dissolved in water. FTIR spectra of all the films showed identical patterns, which indicated no major changes to have occurred in the functional groups as a result of interaction between gelatin, sugars and UV irradiation. The results of this study could be explored for commercial use, depending on industrial needs for either production of edible films or for food packaging purposes.

Occupational asthma and contact dermatitis in a spray painter after introduction of an aziridine cross-linker.

PubMed Central

Leffler, C T; Milton, D K

1999-01-01

A 23-year-old spray painter developed contact dermatitis and respiratory difficulty characterized by small airways obstruction shortly after the polyfunctional aziridine cross-linker CX-100 began to be used in his workplace as a paint activator. The symptoms resolved after he was removed from the workplace and was treated with inhaled and topical steroids. Painters may have an increased risk of asthma due to exposure to a variety of agents, such as isocyanates, alkyd resins, and chromates. This case illustrates the importance of using appropriate work practices and personal protective equipment to minimize exposure. Occupational asthma is diagnosed by a history of work-related symptoms and exposure to known causative agents. The diagnosis is confirmed by serial pulmonary function testing or inhalational challenge testing. The risk of asthma attributable to occupational exposures is probably underappreciated due to underreporting and to inappropriate use of narrow definitions of exposure in epidemiologic studies of attributable risk. Images Figure 1 PMID:10379008

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

Driscoll, Mark S.; Smith, Jennifer L.; Woods, Sean

One of the main problems associated with the use of natural fibers as reinforcing agents in composites is their uptake of moisture. Many natural fibers are lignocellulosic, which causes them to swell and shrink as the amount of available moisture changes. Swelling and shrinking can cause composites to prematurely fail. This paper presents the results of a preliminary study that considers the use of two different low molecular weight monomers, hydroxyethyl methacrylate (HEMA) and hydroxyethyl acrylate (HEA), polymerized by electron beam ionizing radiation, to dimensionally stabilize natural fibers. Eight different treatments consisting of varying amounts of monomer, encapsulating agent, andmore » cross-linkers, were evaluated for their ability to dimensionally stabilize sisal fiber. Results indicate that both polymerized HEA and HEMA can reduce the swelling of sisal fiber. The effectiveness of HEA and HEMA can be further enhanced with the use of a cross-linker (SR 454). The use of hydroxylated monomers to dimensionally stabilize natural fibers may play an important role in reducing delamination and improving fiber-resin adhesion in composites.« less

Regulation of SNM1, an inducible Saccharomyces cerevisiae gene required for repair of DNA cross-links.

PubMed

Wolter, R; Siede, W; Brendel, M

1996-02-05

The interstrand cross-link repair gene SNM1 of Saccharomyces cerevisiae was examined for regulation in response to DNA-damaging agents. Induction of SNM1-lacZ fusions was detected in response to nitrogen mustard, cis-platinum (II) diamine dichloride, UV light, and 8-methoxypsoralen + UVA, but not after heat-shock treatment or incubation with 2-dimethylaminoethylchloride, methylmethane sulfonate or 4-nitroquinoline-N-oxide. The promoter of SNM1 contains a 15 bp motif, which shows homology to the DRE2 box of the RAD2 promoter. Similar motifs have been found in promoter regions of other damage-inducible DNA repair genes. Deletion of this motif results in loss of inducibility of SNM1. Also, a putative negative upstream regulation sequence was found to be responsible for repression of constitutive transcription of SNM1. Surprisingly, no inducibility of SNM1 was found after treatment with DNA-damaging agents in strains without an intact DUN1 gene, while regulation seems unchanged in sad1 mutants.

Bifunctional Crosslinking Agents Enhance Anion Exchange Membrane Efficacy for Vanadium Redox Flow Batteries.

PubMed

Wang, Wenpin; Xu, Min; Wang, Shubo; Xie, Xiaofeng; Lv, Yafei; Ramani, Vijay K

2014-06-02

A series of cross-linked fluorinated poly (aryl ether oxadiazole) membranes (FPAEOM) derivatized with imidazolium groups were prepared. Poly (N-vinylimidazole) (PVI) was used as the bifunctional cross-linking agent to: a) lower vanadium permeability, b) enhance dimensional stability, and c) concomitantly provide added ion exchange capacity in the resultant anion exchange membranes. At a molar ratio of PVI to FPAEOM of 1.5, the resultant membrane (FPAEOM-1.5 PVI) had an ion exchange capacity of 2.2 meq g-1, a vanadium permeability of 6.8×10-7 cm2 min-1, a water uptake of 68 wt.%, and an ionic conductivity of 22.0 mS cm-1, all at 25°C. Single cells prepared with the FPAEOM-1.5 PVI membrane exhibited a higher coulombic efficiency (> 92%) and energy efficiency (> 86%) after 40 test cycles in vanadium redox flow battery. The imidazolium cation showed high chemical stability in highly acidic and oxidizing vanadium solution as opposed to poor stability in alkaline solutions. Based on our DFT studies, this was attributed to the lower HOMO energy (-7.265 eV) of the HSO4- ion (compared to the OH- ion; -5.496 eV) and the larger HOMO-LUMO energy gap (6.394 eV) of dimethylimidazolium bisulfate ([DMIM] [HSO4]) as compared to [DMIM] [OH] (5.387 eV).

mRNA 3' of the A site bound codon is located close to protein S3 on the human 80S ribosome.

PubMed

Molotkov, Maxim V; Graifer, Dmitri M; Popugaeva, Elena A; Bulygin, Konstantin N; Meschaninova, Maria I; Ven'yaminova, Aliya G; Karpova, Galina G

2006-07-01

Ribosomal proteins neighboring the mRNA downstream of the codon bound at the decoding site of human 80S ribosomes were identified using three sets of mRNA analogues that contained a UUU triplet at the 5' terminus and a perfluorophenylazide cross-linker at guanosine, adenosine or uridine residues placed at various locations 3' of this triplet. The positions of modified mRNA nucleotides on the ribosome were governed by tRNA(Phe) cognate to the UUU triplet targeted to the P site. Upon mild UV-irradiation, the mRNA analogues cross-linked preferentially to the 40S subunit, to the proteins and to a lesser extent to the 18S rRNA. Cross-linked nucleotides of 18S rRNA were identified previously. In the present study, it is shown that among the proteins the main target for cross-linking with all the mRNA analogues tested was protein S3 (homologous to prokaryotic S3, S3p); minor cross-linking to protein S2 (S5p) was also detected. Both proteins cross-linked to mRNA analogues in the ternary complexes as well as in the binary complexes (without tRNA). In the ternary complexes protein S15 (S19p) also cross-linked, the yield of the cross-link decreased significantly when the modified nucleotide moved from position +5 to position +12 with respect to the first nucleotide of the P site bound codon. In several ternary complexes minor cross-linking to protein S30 was likewise detected. The results of this study indicate that S3 is a key protein at the mRNA binding site neighboring mRNA downstream of the codon at the decoding site in the human ribosome.

Novel priming and crosslinking systems for use with isocyanatomethacrylate dental adhesives.

PubMed

Chappelow, C C; Power, M D; Bowles, C Q; Miller, R G; Pinzino, C S; Eick, J D

2000-11-01

(a) to design, formulate and evaluate prototype primers and a crosslinking agent for use with isocyanatomethacrylate-based comonomer adhesives and (b) to establish correlations between bond strength and solubility parameter differences between the adhesives and etched dentin, and the permeability coefficients of the adhesives. Equimolar mixtures of 2-isocyanatoethyl methacrylate (IEM) and a methacrylate comonomer were formulated with tri-n-butyl borane oxide (TBBO) as the free radical initiator to have cure times of 6-10 min. Shear bond strengths to dentin were determined for each adhesive mixture (n = 7) using standard testing protocols. Shear bond strengths for the three systems were also determined after application of "reactive primers" to the dentin surface. The "reactive primers" contained 10-20 parts by weight of the respective comonomer mixture and 3.5 parts by weight TBBO in acetone. Solubility parameters difference values (delta delta) and permeability coefficients (P) were approximated for each adhesive system and correlated with shear bond strength values. Additionally, a crosslinking agent was prepared by bulk reaction of an equimolar mixture containing IEM and a methacrylate comonomer. The effects of crosslinker addition on: (a) the setting time of IEM; and (b) the setting times and initiator requirements of selected IEM/comonomer mixtures were determined. Shear bond strength values (MPa): IEM/HEMA 13.6 +/- 2.0 (no primer), 20.1 +/- 2.0 (with primer); IEM/HETMA 9.3 +/- 3.3 (no primer), 20.8 +/- 8.1 (with primer); IEM/AAEMA 13.6 +/- 1.9 (no primer), 17.3 +/- 3.2 (with primer). Also, approximated permeability coefficients showed a significant correlation (r = +0.867, p < 0.001) with shear bond strength values. Crosslinker addition studies with IEM/4-META: (a) at 5-9 mol% reduced the setting time of IEM polymerization by 79%; and (b) at 6 mol% reduced initiator level requirements 60-70% to achieve a comparable setting time, and decreased setting times by ca. 75% for a given initiator level with selected IEM/methacrylate adhesive systems. The shear bond strengths of isocyanatomethacrylate-based dental adhesives can be enhanced by using reactive primers; their setting times and initiator requirements can be improved using a dimethacrylate crosslinker. Approximated permeability coefficients may be useful as indicators of bonding performance for dentin adhesive systems.

Evidence for the Phospholipid Sponge Effect as the Biocidal Mechanism in Surface-Bound Polyquaternary Ammonium Coatings with Variable Cross-Linking Density.

PubMed

Gao, Jing; White, Evan M; Liu, Qiaohong; Locklin, Jason

2017-03-01

Poly quaternary "-oniums" derived from polyethylenimine (PEI), poly(vinyl-N-alkylpyridinium), or chitosan belong to a class of cationic polymers that are efficient antimicrobial agents. When dissolved in solution, the positively charged polycations are able to displace the divalent cations of the cellular phospholipid bilayer and disrupt the ionic cross-links and structural integrity of the membrane. However, when immobilized to a surface where confinement limits diffusion, poly -oniums still show excellent antimicrobial activity, which implies a different biocidal mode of action. Recently, a proposed mechanism, named phospholipid sponge effect, suggested that surface-bound polycationic networks are capable of recruiting negatively charged phospholipids out of the bacterial cell membrane and sequestering them within the polymer matrix.1 However, there has been insufficient evidence to support this hypothesis. In this study, a surface-bound N,N-dodecyl methyl-co-N,N-methylbenzophenone methyl quaternary PEI (DMBQPEI) was prepared to verify the phospholipid sponge effect. By tuning the irradiation time, the cross-linking densities of surface-bound DMBQPEI films were mediated. The modulus of films was measured by PeakForce Quantitative Nanomechanical Mapping (QNM) to indicate the cross-linking density variation with increasing irradiation time. A negative correlation between the film cross-linking density and the absorption of a negatively charged phospholipid (DPhPG) was observed, but no such correlations were observed with a neutral phospholipid (DPhPC), which strongly supported the action of anionic phospholipid suction proposed in the lipid sponge effect. Moreover, the killing efficiency toward S. aureus and E. coli was inversely affected by the cross-linking density of the films, providing evidence for the phospholipid sponge effect. The relationship between killing efficiency and film cross-linking density is discussed.

Low-Melt Polyamic Acid Based Powder Coatings

NASA Technical Reports Server (NTRS)

Jolley, Scott T. (Inventor)

2017-01-01

The present invention is directed to a method for powder coating a metal substrate using a low-melt polyamic acid (PAA) polymer that readily imidizes to polyimides. These low-melt PAAs have been shown to be useful in resins applied as powder coatings to metal surfaces. The resin includes an end-capping material capable of providing crosslinking functionality to at least one end of the low-melt PAA polymer. The end-capping material functions dually as a polymerization chain terminator and crosslinking agent, thus producing resins that have molecular weights low enough to flow well and form good cured films applicable for use in powder coating.

[Fanconi anemia: cellular and molecular features].

PubMed

Macé, G; Briot, D; Guervilly, J-H; Rosselli, F

2007-02-01

Fanconi anemia (FA) is a recessive human cancer prone syndrome featuring bone marrow failure, developmental abnormalities and hypersensitivity to DNA crosslinking agents exposure. 11 among 12 FA gene have been isolated. The biochemical functions of the FANC proteins remain poorly understood. Anyhow, to cope with DNA crosslinks a cell needs a functional FANC pathway. Moreover, the FANC proteins appear to be involved in cell protection against oxidative damage and in the control of TNF-alpha activity. In this review, we describe the current understanding of the FANC pathway and we present how it may be integrated in the complex networks of proteins involved in maintaining the cellular homeostasis.

Proteinaceous Resin and Hydrophilic Encapsulation: A Self-Healing-Related Study

NASA Astrophysics Data System (ADS)

Zheng, Ting

Inspired by living organisms, self-healing materials have been designed as smart materials. Their automatic healing nature is achieved through the use of capsule in which the healing agent is encapsulated. The occurrence of cracks leads to ripping of the capsule, along with crack propagation and release of the healing agent that wets the crack surface to eventually heal (bond) the crack. Such automatic repair of the crack significantly extends the service life of the material. A vast majority of existing self-healing systems have been designed for the epoxy matrix - the most common commercially used thermoset - that possesses low crack resistance. Currently, self-healing systems have not yet been introduced for fully protein-based materials, despite their great potential to replace currently used synthesis precursors for the latter and the eco-friendly nature of self-healing materials. This has been probably due to two major obstacles: poor mechanical properties of the protein-based matrix, and extreme difficulty associated with the encapsulation of hydrophilic healing agents suitable for the protein-based matrix. This study provides possible solutions towards addressing both these obstacles. To improve the inherent mechanical properties of protein-based resin, soy protein isolate (SPI) was chosen as the model in this study. Dialdehyde carboxymethyl cellulose (DCMC) was synthesized and used as the crosslinking agent to modify the SPI film. As-synthesized DCMC - a fully bio-based material - exhibited high mechanical strength, excellent thermal stability, and reduced moisture sensitivity. Good compatibility and effective crosslinking were believed to be the key reasons for such property enhancements. However, these were accompanied by poor crack resistance, where self-healing is a pertinent solution. A novel healing system for the protein matrix was designed in this work via the use of formaldehyde as a healing agent. Subsequently, the well-acknowledged challenge, e.g. hydrophilic agent encapsulation, was addressed through the development of novel polyurethane-Poly(melamine-formaldehyde) (PU-PMF) dual-component capsules. Remarkably, the external PU insulation layer was fabricated through interfacial polymerization based on a water-in-oil-in-oil (W/O/O) emulsion template. Surface tension was identified as the main driving factor for the formation of the external oil phase. The internal PMF layer was observed to strongly influence the internal morphology of the capsule. A protocol was developed, and a typical capsule with dense and neat shell morphology with a shell/capsule diameter (around 3 %) was fabricated. This study provides solutions for the two aforementioned obstacles related to the development of the healing system for the protein-based materials.

Development of biodegradable foamlike materials based on casein and sodium montmorillonite clay

USDA-ARS?s Scientific Manuscript database

Biodegradable foamlike materials based on a naturally occurring polymer (casein protein) and sodium montmorillonite clay (Na+-MMT) were produced through a simple freeze-drying process. By utilizing DL-glyceraldehyde (GC) as a chemical cross-linking agent, the structural integrity of these new aeroge...

Polymerization of Quinone-Crosslinked Marine Bioadhesive Protein

DTIC Science & Technology

1988-10-05

T. (1988) Adhesive protein of ribbed mussels: a natural glue with some features of collagen . To: J. Biol. Chem. Rzepecki, L., Nagafuchi, T. and...protein as inducing agent in the settlement of Mytilus edulis larvae", and "Hemolytic toxins of two marine jellyfish ", respectively. Tatsuhiko

DNA-Protein Cross-Links: Formation, Structural Identities, and Biological Outcomes.

PubMed

Tretyakova, Natalia Y; Groehler, Arnold; Ji, Shaofei

2015-06-16

Noncovalent DNA-protein interactions are at the heart of normal cell function. In eukaryotic cells, genomic DNA is wrapped around histone octamers to allow for chromosomal packaging in the nucleus. Binding of regulatory protein factors to DNA directs replication, controls transcription, and mediates cellular responses to DNA damage. Because of their fundamental significance in all cellular processes involving DNA, dynamic DNA-protein interactions are required for cell survival, and their disruption is likely to have serious biological consequences. DNA-protein cross-links (DPCs) form when cellular proteins become covalently trapped on DNA strands upon exposure to various endogenous, environmental and chemotherapeutic agents. DPCs progressively accumulate in the brain and heart tissues as a result of endogenous exposure to reactive oxygen species and lipid peroxidation products, as well as normal cellular metabolism. A range of structurally diverse DPCs are found following treatment with chemotherapeutic drugs, transition metal ions, and metabolically activated carcinogens. Because of their considerable size and their helix-distorting nature, DPCs interfere with the progression of replication and transcription machineries and hence hamper the faithful expression of genetic information, potentially contributing to mutagenesis and carcinogenesis. Mass spectrometry-based studies have identified hundreds of proteins that can become cross-linked to nuclear DNA in the presence of reactive oxygen species, carcinogen metabolites, and antitumor drugs. While many of these proteins including histones, transcription factors, and repair proteins are known DNA binding partners, other gene products with no documented affinity for DNA also participate in DPC formation. Furthermore, multiple sites within DNA can be targeted for cross-linking including the N7 of guanine, the C-5 methyl group of thymine, and the exocyclic amino groups of guanine, cytosine, and adenine. This structural complexity complicates structural and biological studies of DPC lesions. Two general strategies have been developed for creating DNA strands containing structurally defined, site-specific DPCs. Enzymatic methodologies that trap DNA modifying proteins on their DNA substrate are site specific and efficient, but do not allow for systematic studies of DPC lesion structure on their biological outcomes. Synthetic methodologies for DPC formation are based on solid phase synthesis of oligonucleotide strands containing protein-reactive unnatural DNA bases. The latter approach allows for a wider range of protein substrates to be conjugated to DNA and affords a greater flexibility for the attachment sites within DNA. In this Account, we outline the chemistry of DPC formation in cells, describe our recent efforts to identify the cross-linked proteins by mass spectrometry, and discuss various methodologies for preparing DNA strands containing structurally defined, site specific DPC lesions. Polymerase bypass experiments conducted with model DPCs indicate that the biological outcomes of these bulky lesions are strongly dependent on the peptide/protein size and the exact cross-linking site within DNA. Future studies are needed to elucidate the mechanisms of DPC repair and their biological outcomes in living cells.

DNA-Protein Cross-links: Formation, Structural Identities, and Biological Outcomes

PubMed Central

Tretyakova, Natalia Y.; Groehler, Arnold; Ji, Shaofei

2015-01-01

CONSPECTUS Non-covalent DNA-protein interactions are at the heart of normal cell function. In eukaryotic cells, genomic DNA is wrapped around histone octamers to allow for chromosomal packaging in the nucleus. Binding of regulatory protein factors to DNA directs replication, controls transcription, and mediates cellular responses to DNA damage. Because of their fundamental significance in all cellular processes involving DNA, dynamic DNA-protein interactions are required for cell survival, and their disruption is likely to have serious biological consequences. DNA-protein cross-links (DPCs) form when cellular proteins become covalently trapped on DNA strands upon exposure to various endogenous, environmental and chemotherapeutic agents. DPCs progressively accumulate in the brain and heart tissues as a result of endogenous exposure to reactive oxygen species and lipid peroxidation products, as well as normal cellular metabolism. A range of structurally diverse DPCs are found following treatment with chemotherapeutic drugs, transition metal ions, and metabolically activated carcinogens. Because of their considerable size and their helix-distorting nature, DPCs interfere with the progression of replication and transcription machineries and hence hamper the faithful expression of genetic information, potentially contributing to mutagenesis and carcinogenesis. Mass spectrometry-based studies have identified hundreds of proteins that can become cross-linked to nuclear DNA in the presence of reactive oxygen species, carcinogen metabolites, and antitumor drugs. While many of these proteins including histones, transcription factors, and repair proteins are known DNA binding partners, other gene products with no documented affinity for DNA also participate in DPC formation. Furthermore, multiple sites within DNA can be targeted for cross-linking including the N7 of guanine, the C-5 methyl group of thymine, and the exocyclic amino groups of guanine, cytosine, and adenine. This structural complexity complicates structural and biological studies of DPC lesions. Two general strategies have been developed for creating DNA strands containing structurally defined, site-specific DPCs. Enzymatic methodologies that trap DNA modifying proteins on their DNA substrate are site specific and efficient, but do not allow for systematic studies of DPC lesion structure on their biological outcomes. Synthetic methodologies for DPC formation are based on solid phase synthesis of oligonucleotide strands containing protein-reactive unnatural DNA bases. The latter approach allows for a wider range of protein substrates to be conjugated to DNA and affords a greater flexibility for the attachment sites within DNA. In this Account, we outline the chemistry of DPC formation in cells, describe our recent efforts to identify the cross-linked proteins by mass spectrometry, and discuss various methodologies for preparing DNA strands containing structurally defined, site specific DPC lesions. Polymerase bypass experiments conducted with model DPCs indicate that the biological outcomes of these bulky lesions are strongly dependent on the peptide/protein size and the exact cross-linking site within DNA. Future studies are needed to elucidate the mechanisms of DPC repair and their biological outcomes in living cells. PMID:26032357

Novel actin crosslinker superfamily member identified by a two step degenerate PCR procedure.

PubMed

Byers, T J; Beggs, A H; McNally, E M; Kunkel, L M

1995-07-24

Actin-crosslinking proteins link F-actin into the bundles and networks that constitute the cytoskeleton. Dystrophin, beta-spectrin, alpha-actinin, ABP-120, ABP-280, and fimbrin share homologous actin-binding domains and comprise an actin crosslinker superfamily. We have identified a novel member of this superfamily (ACF7) using a degenerate primer-mediated PCR strategy that was optimized to resolve less-abundant superfamily sequences. The ACF7 gene is on human chromosome 1 and hybridizes to high molecular weight bands on northern blots. Sequence comparisons argue that ACF7 does not fit into one of the existing families, but represents a new class within the superfamily.

Verification, Dosimetry and Biomonitoring of Mustard Gas Exposure via Immunochemical Detection of Mustard Gas Adducts to DNA and Proteins.

DTIC Science & Technology

1990-09-01

90% of all radioactivity. Mustard gas appeared to be a very effective alkylating agent for bases in DNA. Even in blood, with a variety of reactive...stranded than of the single-stranded material is required for effective competition in the ELISA test, probably as a result of interstrand crosslinks...first noticed by their effects on casualties. If a total ban on the use, possession and production of chemical agents will eventually materialize, the

Cross-Linking Interferes With Assessing Sulfur Mustard-Induced DNA Damage in Human Peripheral Blood Lymphocytes Using the Comet Assay

DTIC Science & Technology

2004-01-01

of SM to impede the migration of H,0 2 -damaged mal ian cell lethality with bifunctional alkylating agents . Chemr. Biol. Iriterui. 38:75-86.DNA is an...3100 Ricketts Point Road, Aberdeen Proving Ground, Maryland 21010-5400 N-3 position of adenine, and alkylation leads to depurination of Sulfur...mustard (SM) is a blistering agent that produces DNA DNA strands. Subsequent breakage of phosphodiester bonds at strand breaks. To detect SM-induced DNA

Trifunctional cross-linker for mapping protein-protein interaction networks and comparing protein conformational states

PubMed Central

Tan, Dan; Li, Qiang; Zhang, Mei-Jun; Liu, Chao; Ma, Chengying; Zhang, Pan; Ding, Yue-He; Fan, Sheng-Bo; Tao, Li; Yang, Bing; Li, Xiangke; Ma, Shoucai; Liu, Junjie; Feng, Boya; Liu, Xiaohui; Wang, Hong-Wei; He, Si-Min; Gao, Ning; Ye, Keqiong; Dong, Meng-Qiu; Lei, Xiaoguang

2016-01-01

To improve chemical cross-linking of proteins coupled with mass spectrometry (CXMS), we developed a lysine-targeted enrichable cross-linker containing a biotin tag for affinity purification, a chemical cleavage site to separate cross-linked peptides away from biotin after enrichment, and a spacer arm that can be labeled with stable isotopes for quantitation. By locating the flexible proteins on the surface of 70S ribosome, we show that this trifunctional cross-linker is effective at attaining structural information not easily attainable by crystallography and electron microscopy. From a crude Rrp46 immunoprecipitate, it helped identify two direct binding partners of Rrp46 and 15 protein-protein interactions (PPIs) among the co-immunoprecipitated exosome subunits. Applying it to E. coli and C. elegans lysates, we identified 3130 and 893 inter-linked lysine pairs, representing 677 and 121 PPIs. Using a quantitative CXMS workflow we demonstrate that it can reveal changes in the reactivity of lysine residues due to protein-nucleic acid interaction. DOI: http://dx.doi.org/10.7554/eLife.12509.001 PMID:26952210

Study on structure, mechanical property and cell cytocompatibility of electrospun collagen nanofibers crosslinked by common agents.

PubMed

Luo, Xueshi; Guo, Zhenzhao; He, Ping; Chen, Tian; Li, Lihua; Ding, Shan; Li, Hong

2018-07-01

Collagen electrospun scaffolds properly reproduce the framework of the extracellular matrix (ECM) of tissues that are natural with the fibrous morphology of the protein by coupling large biomimetism of the biological material. However, traditional solvents employed for collagen electrospinning lead to poor mechanical attributes and bad hydro-stability. In this work, by N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride with N-hydroxysulfosuccinimide (EDC-NHS), glutaraldehyde (GTA) and genipin (GP) respectively, electrospun collagen fibers cross-linked, effectively stabilized the fiber morphology over 2months and improved the mechanical properties in both dry and wet state, especially EDC-NHS with large ultimate tensile stress and ε b . The secondary structure of collagen structure still remained and had no obvious difference among various crosslinked samples according to FTIR. On the cell assessment, electrospun collagen fibers crosslinked by EDC-NHS, GTA and GP, were found to support cell adhesion, spreading and proliferation of MC3T3-E1. By contrast, GTA was more effective in preserving explicit fibrous morphology with a relatively lower cell viability both in FBS and BSA soaked mats. Interestingly, GP also had the similar cytocompatibility of MC3T3-E1 as EDC-NHS did. The study proved the feasibility of chemical crosslinker to electrospun collagen for biomedical application. Copyright © 2018. Published by Elsevier B.V.

Inter-crosslinking network gels having both shape memory and high ductility

NASA Astrophysics Data System (ADS)

Amano, Yoshitaka; Hidema, Ruri; Furukawa, Hidemitsu

2012-04-01

Medical treatment for injuries should be easy and quick in many accidents. Plasters or bandages are frequently used to wrap and fix injured parts. If plasters or bandages have additional smart functions, such as cooling, removability and repeatability, they will be much more useful and effective. Here we propose innovative biocompatible materials, that is, nontoxic high-strength shape-memory gels as novel smart medical materials. These smart gels were prepared from two monomers (DMAAm and SA), a polymer (HPC), and an inter-crosslinking agent (Karenz-MOI). In the synthesis of the gels, 1) a shape-memory copolymer network is made from the DMAAm and the SA, and 2) the copolymer and the HPC are crosslinked by the Karenz-MOI. Thus the crosslinking points are connected only between the different polymers. This is our original technique of developing a new network structure of gels, named Inter-Crosslinking Network (ICN). The ICN gels achieve high ductility, going up to 700% strain in tensile tests, while the ICN gels contain about 44% water. Moreover the SA has temperature dependence due to its crystallization properties; thus the ICN gels obtain shape memory properties and are named ICN-SMG. While the Young's modulus of the ICN-SMG is large below their crystallization temperature and the gels behave like plastic materials, the modulus becomes smaller above the temperature and the gels turn back to their original shape.

Effect of UV-ozone treatment on poly(dimethylsiloxane) membranes: surface characterization and gas separation performance.

PubMed

Fu, Ywu-Jang; Qui, Hsuan-zhi; Liao, Kuo-Sung; Lue, Shingjiang Jessie; Hu, Chien-Chieh; Lee, Kueir-Rarn; Lai, Juin-Yih

2010-03-16

A thin SiO(x) selective surface layer was formed on a series of cross-linked poly(dimethylsiloxane) (PDMS) membranes by exposure to ultraviolet light at room temperature in the presence of ozone. The conversion of the cross-linked polysiloxane to SiO(x) was monitored by attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy, X-ray photoelectron spectroscopy (XPS), energy-dispersive X-ray (EDX) microanalysis, contact angle analysis, and atomic force microscopy (AFM). The conversion of the cross-linked polysiloxane to SiO(x) increased with UV-ozone exposure time and cross-linking agent content, and the surface possesses highest conversion. The formation of a SiO(x) layer increased surface roughness, but it decreased water contact angle. Gas permeation measurements on the UV-ozone exposure PDMS membranes documented interesting gas separation properties: the O(2) permeability of the cross-linked PDMS membrane before UV-ozone exposure was 777 barrer, and the O(2)/N(2) selectivity was 1.9; after UV-ozone exposure, the permeability decreased to 127 barrer while the selectivity increased to 5.4. The free volume depth profile of the SiO(x) layer was investigated by novel slow positron beam. The results show that free volume size increased with the depth, yet the degree of siloxane conversion to SiO(x) does not affect the amount of free volume.

Fanconi anemia (FA) binding protein FAAP20 stabilizes FA complementation group A (FANCA) and participates in interstrand cross-link repair.

PubMed

Leung, Justin Wai Chung; Wang, Yucai; Fong, Ka Wing; Huen, Michael Shing Yan; Li, Lei; Chen, Junjie

2012-03-20

The Fanconi anemia (FA) pathway participates in interstrand cross-link (ICL) repair and the maintenance of genomic stability. The FA core complex consists of eight FA proteins and two Fanconi anemia-associated proteins (FAAP24 and FAAP100). The FA core complex has ubiquitin ligase activity responsible for monoubiquitination of the FANCI-FANCD2 (ID) complex, which in turn initiates a cascade of biochemical events that allow processing and removal of cross-linked DNA and thereby promotes cell survival following DNA damage. Here, we report the identification of a unique component of the FA core complex, namely, FAAP20, which contains a RAD18-like ubiquitin-binding zinc-finger domain. Our data suggest that FAAP20 promotes the functional integrity of the FA core complex via its direct interaction with the FA gene product, FANCA. Indeed, somatic knockout cells devoid of FAAP20 displayed the hallmarks of FA cells, including hypersensitivity to DNA cross-linking agents, chromosome aberrations, and reduced FANCD2 monoubiquitination. Taking these data together, our study indicates that FAAP20 is an important player involved in the FA pathway.

Fanconi anemia (FA) binding protein FAAP20 stabilizes FA complementation group A (FANCA) and participates in interstrand cross-link repair

PubMed Central

Leung, Justin Wai Chung; Wang, Yucai; Fong, Ka Wing; Huen, Michael Shing Yan; Li, Lei; Chen, Junjie

2012-01-01

The Fanconi anemia (FA) pathway participates in interstrand cross-link (ICL) repair and the maintenance of genomic stability. The FA core complex consists of eight FA proteins and two Fanconi anemia-associated proteins (FAAP24 and FAAP100). The FA core complex has ubiquitin ligase activity responsible for monoubiquitination of the FANCI-FANCD2 (ID) complex, which in turn initiates a cascade of biochemical events that allow processing and removal of cross-linked DNA and thereby promotes cell survival following DNA damage. Here, we report the identification of a unique component of the FA core complex, namely, FAAP20, which contains a RAD18-like ubiquitin-binding zinc-finger domain. Our data suggest that FAAP20 promotes the functional integrity of the FA core complex via its direct interaction with the FA gene product, FANCA. Indeed, somatic knockout cells devoid of FAAP20 displayed the hallmarks of FA cells, including hypersensitivity to DNA cross-linking agents, chromosome aberrations, and reduced FANCD2 monoubiquitination. Taking these data together, our study indicates that FAAP20 is an important player involved in the FA pathway. PMID:22396592

Ionic Modification Turns Commercial Rubber into a Self-Healing Material.

PubMed

Das, Amit; Sallat, Aladdin; Böhme, Frank; Suckow, Marcus; Basu, Debdipta; Wiessner, Sven; Stöckelhuber, Klaus Werner; Voit, Brigitte; Heinrich, Gert

2015-09-23

Invented by Charles Goodyear, chemical cross-linking of rubbers by sulfur vulcanization is the only method by which modern automobile tires are manufactured. The formation of these cross-linked network structures leads to highly elastic properties, which substantially reduces the viscous properties of these materials. Here, we describe a simple approach to converting commercially available and widely used bromobutyl rubber (BIIR) into a highly elastic material with extraordinary self-healing properties without using conventional cross-linking or vulcanising agents. Transformation of the bromine functionalities of BIIR into ionic imidazolium bromide groups results in the formation of reversible ionic associates that exhibit physical cross-linking ability. The reversibility of the ionic association facilitates the healing processes by temperature- or stress-induced rearrangements, thereby enabling a fully cut sample to retain its original properties after application of the self-healing process. Other mechanical properties, such as the elastic modulus, tensile strength, ductility, and hysteresis loss, were found to be superior to those of conventionally sulfur-cured BIIR. This simple and easy approach to preparing a commercial rubber with self-healing properties offers unique development opportunities in the field of highly engineered materials, such as tires, for which safety, performance, and longer fatigue life are crucial factors.

Room-Temperature Performance of Poly(Ethylene Ether Carbonate)-Based Solid Polymer Electrolytes for All-Solid-State Lithium Batteries.

PubMed

Jung, Yun-Chae; Park, Myung-Soo; Kim, Duck-Hyun; Ue, Makoto; Eftekhari, Ali; Kim, Dong-Won

2017-12-13

Amorphous poly(ethylene ether carbonate) (PEEC), which is a copolymer of ethylene oxide and ethylene carbonate, was synthesized by ring-opening polymerization of ethylene carbonate. This route overcame the common issue of low conductivity of poly(ethylene oxide)(PEO)-based solid polymer electrolytes at low temperatures, and thus the solid polymer electrolyte could be successfully employed at the room temperature. Introducing the ethylene carbonate units into PEEC improved the ionic conductivity, electrochemical stability and lithium transference number compared with PEO. A cross-linked solid polymer electrolyte was synthesized by photo cross-linking reaction using PEEC and tetraethyleneglycol diacrylate as a cross-linking agent, in the form of a flexible thin film. The solid-state Li/LiNi 0.6 Co 0.2 Mn 0.2 O 2 cell assembled with solid polymer electrolyte based on cross-linked PEEC delivered a high initial discharge capacity of 141.4 mAh g -1 and exhibited good capacity retention at room temperature. These results demonstrate the feasibility of using this solid polymer electrolyte in all-solid-state lithium batteries that can operate at ambient temperatures.

Cross-Link Guided Molecular Modeling with ROSETTA

PubMed Central

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

2013-01-01

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

Biocompatibility of Genipin and Glutaraldehyde Cross-Linked Chitosan Materials in the Anterior Chamber of the Eye

PubMed Central

Lai, Jui-Yang

2012-01-01

Chitosan is a naturally occurring cationic polysaccharide and has attracted much attention in the past decade as an important ophthalmic biomaterial. We recently demonstrated that the genipin (GP) cross-linked chitosan is compatible with human retinal pigment epithelial cells. The present work aims to further investigate the in vivo biocompatibility of GP-treated chitosan (GP-chi group) by adopting the anterior chamber of a rabbit eye model. The glutaraldehyde (GTA) cross-linked samples (GTA-chi group) were used for comparison. The 7-mm-diameter membrane implants made from either non-cross-linked chitosan or chemically modified materials with a cross-linking degree of around 80% were inserted in the ocular anterior chamber for 24 weeks and characterized by slit-lamp and specular microscopic examinations, intraocular pressure measurements, and corneal thickness measurements. The interleukin-6 expressions at mRNA level were also detected by quantitative real-time reverse transcription polymerase chain reaction. Results of clinical observations showed that the overall ocular scores in the GTA-chi groups were relatively high. In contrast, the rabbits bearing GP-chi implants in the anterior chamber of the eye exhibited no signs of ocular inflammation. As compared to the non-cross-linked counterparts, the GP-chi samples improved the preservation of corneal endothelial cell density and possessed better anti-inflammatory activities, indicating the benefit action of the GP cross-linker. In summary, the intracameral tissue response to the chemically modified chitosan materials strongly depends on the selection of cross-linking agents. PMID:23109832

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

PubMed

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

2011-06-01

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

Thermo- and electro-dual responsive poly(ionic liquid) electrolyte based smart windows.

PubMed

Chen, Fei; Ren, Yongyuan; Guo, Jiangna; Yan, Feng

2017-01-31

Thermo- and electro-dual responsive poly(ionic liquid) (PIL) based electrolytes were synthesized by co-polymerization of N-isopropylacrylamide (NIPAM) with (or without) 3-butyl-1-vinyl-imidazolium bromide ([BVIm][Br]) using diallyl-viologen (DAV) as both the cross-linking agent and electrochromic material.

Development of the phosphorus and nitrogen containing flame retardant for value added cotton product

USDA-ARS?s Scientific Manuscript database

It is our desire to develop new crosslinking agents for cotton textiles that afford useful flame protection regardless of fabric construction. Herein we present the synthesis and the application of the triazine and piperazine derivatives as flame retardant on cotton. Novel phosphorus-nitrogen contai...

Smart surface coating of drug nanoparticles with cross-linkable polyethylene glycol for bio-responsive and highly efficient drug delivery

NASA Astrophysics Data System (ADS)

Wei, Weijia; Zhang, Xiujuan; Chen, Xianfeng; Zhou, Mengjiao; Xu, Ruirui; Zhang, Xiaohong

2016-04-01

Many drug molecules can be directly used as nanomedicine without the requirement of any inorganic or organic carriers such as silica and liposome nanostructures. This new type of carrier-free drug nanoparticles (NPs) has great potential in clinical treatment because of its ultra-high drug loading capacity and biodegradability. For practical applications, it is essential for such nanomedicine to possess robust stability and minimal premature release of therapeutic molecules during circulation in the blood stream. To meet this requirement, herein, we develop GSH-responsive and crosslinkable amphiphilic polyethylene glycol (PEG) molecules to modify carrier-free drug NPs. These PEG molecules can be cross-linked on the surface of the NPs to endow them with greater stability and the cross-link is sensitive to intracellular environment for bio-responsive drug release. With this elegant design, our experimental results show that the liberation of DOX from DOX-cross-linked PEG NPs is dramatically slower than that from DOX-non-cross-linked PEG NPs, and the DOX release profile can be controlled by tuning the concentration of the reducing agent to break the cross-link between PEG molecules. More importantly, in vivo studies reveal that the DOX-cross-linked PEG NPs exhibit favorable blood circulation half-life (>4 h) and intense accumulation in tumor areas, enabling effective anti-cancer therapy. We expect this work will provide a powerful strategy for stabilizing carrier-free nanomedicines and pave the way to their successful clinical applications in the future.Many drug molecules can be directly used as nanomedicine without the requirement of any inorganic or organic carriers such as silica and liposome nanostructures. This new type of carrier-free drug nanoparticles (NPs) has great potential in clinical treatment because of its ultra-high drug loading capacity and biodegradability. For practical applications, it is essential for such nanomedicine to possess robust stability and minimal premature release of therapeutic molecules during circulation in the blood stream. To meet this requirement, herein, we develop GSH-responsive and crosslinkable amphiphilic polyethylene glycol (PEG) molecules to modify carrier-free drug NPs. These PEG molecules can be cross-linked on the surface of the NPs to endow them with greater stability and the cross-link is sensitive to intracellular environment for bio-responsive drug release. With this elegant design, our experimental results show that the liberation of DOX from DOX-cross-linked PEG NPs is dramatically slower than that from DOX-non-cross-linked PEG NPs, and the DOX release profile can be controlled by tuning the concentration of the reducing agent to break the cross-link between PEG molecules. More importantly, in vivo studies reveal that the DOX-cross-linked PEG NPs exhibit favorable blood circulation half-life (>4 h) and intense accumulation in tumor areas, enabling effective anti-cancer therapy. We expect this work will provide a powerful strategy for stabilizing carrier-free nanomedicines and pave the way to their successful clinical applications in the future. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr09167e

Synthesis of native-like crosslinked duplex RNA and study of its properties.

PubMed

Onizuka, Kazumitsu; Hazemi, Madoka E; Thomas, Justin M; Monteleone, Leanna R; Yamada, Ken; Imoto, Shuhei; Beal, Peter A; Nagatsugi, Fumi

2017-04-01

A variety of enzymes have been found to interact with double-stranded RNA (dsRNA) in order to carry out its functions. We have endeavored to prepare the covalently crosslinked native-like duplex RNA, which could be useful for biochemical studies and RNA nanotechnology. In this study, the interstrand covalently linked duplex RNA was formed by a crosslinking reaction between vinylpurine (VP) and the target cytosine or uracil in RNA. We measured melting temperatures and CD spectra to identify the properties of the VP crosslinked duplex RNA. The crosslinking formation increased the thermodynamic stability without disturbing the natural conformation of dsRNA. In addition, a competitive binding experiment with the duplex RNA binding enzyme, ADAR2, showed the crosslinked dsRNA bound the protein with nearly the same binding affinity as the natural dsRNA, confirming that it has finely preserved the natural traits of duplex RNA. Copyright © 2017. Published by Elsevier Ltd.

Inter-Satellite Communications Considerations and Requirements for Distributed Spacecraft and Formation Flying Systems

NASA Technical Reports Server (NTRS)

Kwadrat, Carl F.; Horne, William D.; Edwards, Bernard L.

2002-01-01

In order to avoid selecting inadequate inter-spacecraft cross-link communications standards for Distributed Spacecraft System (DSS) missions, it is first necessary to identify cross-link communications strategies and requirements common to a cross-section of proposed missions. This paper addresses the cross-link communication strategies and requirements derived from a survey of 39 DSS mission descriptions that are projected for potential launch within the next 20 years. The inter-spacecraft communications strategies presented are derived from the topological and communications constraints from the DSS missions surveyed. Basic functional requirements are derived from an analysis of the fundamental activities that must be undertaken to establish and maintain a cross-link between two DSS spacecraft. Cross-link bandwidth requirements are derived from high-level assessments of mission science objectives and operations concepts. Finally, a preliminary assessment of possible cross-link standards is presented within the context of the basic operational and interoperability requirements.

Combination Platinum-based and DNA Damage Response-targeting Cancer Therapy: Evolution and Future Directions.

PubMed

Basourakos, Spyridon P; Li, Likun; Aparicio, Ana M; Corn, Paul G; Kim, Jeri; Thompson, Timothy C

2017-01-01

Maintenance of genomic stability is a critical determinant of cell survival and is necessary for growth and progression of malignant cells. Interstrand crosslinking (ICL) agents, including platinum-based agents, are first-line chemotherapy treatment for many solid human cancers. In malignant cells, ICL triggers the DNA damage response (DDR). When the damage burden is high and lesions cannot be repaired, malignant cells are unable to divide and ultimately undergo cell death either through mitotic catastrophe or apoptosis. The activities of ICL agents, in particular platinum-based therapies, establish a "molecular landscape," i.e., a pattern of DNA damage that can potentially be further exploited therapeutically with DDR-targeting agents. If the molecular landscape created by platinum-based agents could be better defined at the molecular level, a systematic, mechanistic rationale(s) could be developed for the use of DDR-targeting therapies in combination/maintenance protocols for specific, clinically advanced malignancies. New therapeutic drugs such as poly(ADP-ribose) polymerase (PARP) inhibitors are examples of DDR-targeting therapies that could potentially increase the DNA damage and replication stress imposed by platinum-based agents in tumor cells and provide therapeutic benefit for patients with advanced malignancies. Recent studies have shown that the use of PARP inhibitors together with platinum-based agents is a promising therapy strategy for ovarian cancer patients with "BRCAness", i.e., a phenotypic characteristic of tumors that not only can involve loss-of-function mutations in either BRCA1 or BRCA2, but also encompasses the molecular features of BRCA-mutant tumors. On the basis of these promising results, additional mechanism-based studies focused on the use of various DDR-targeting therapies in combination with platinum-based agents should be considered. This review discusses, in general, (1) ICL agents, primarily platinum-based agents, that establish a molecular landscape that can be further exploited therapeutically; (2) multiple points of potential intervention after ICL agent-induced crosslinking that further predispose to cell death and can be incorporated into a systematic, therapeutic rationale for combination/ maintenance therapy using DDR-targeting agents; and (3) available agents that can be considered for use in combination/maintenance clinical protocols with platinum-based agents for patients with advanced malignancies. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Release of Self-Healing Agents in a Material: What Happens Next?

PubMed

Lee, Min Wook; Yoon, Sam S; Yarin, Alexander L

2017-05-24

A microfluidic chip-like setup consisting of a vascular system of microchannels alternatingly filled with either a resin monomer or a curing agent is used to study the intrinsic physical healing mechanism in self-healing materials. It is observed that, as a prenotched crack propagates across the chip, the resin and curing agent are released from the damaged channels. Subsequently, both the resin and the curing agent wet the surrounding polydimethylsiloxane (PDMS) matrix and spread over the crack banks until the two blobs come in contact, mix, and polymerize through an organometallic cross-linking reaction. Moreover, the polymerized domains form a system of pillars, which span the crack banks on the opposite side. This "stitching" phenomenon prevents further propagation of the crack.

The Fanconi anemia DNA repair pathway: structural and functional insights into a complex disorder.

PubMed

Walden, Helen; Deans, Andrew J

2014-01-01

Mutations in any of at least sixteen FANC genes (FANCA-Q) cause Fanconi anemia, a disorder characterized by sensitivity to DNA interstrand crosslinking agents. The clinical features of cytopenia, developmental defects, and tumor predisposition are similar in each group, suggesting that the gene products participate in a common pathway. The Fanconi anemia DNA repair pathway consists of an anchor complex that recognizes damage caused by interstrand crosslinks, a multisubunit ubiquitin ligase that monoubiquitinates two substrates, and several downstream repair proteins including nucleases and homologous recombination enzymes. We review progress in the use of structural and biochemical approaches to understanding how each FANC protein functions in this pathway.

PEG-based degradable networks for drug delivery applications

NASA Astrophysics Data System (ADS)

Ostroha, Jamie L.

The controlled delivery of therapeutic agents by biodegradable hydrogels has become a popular mechanism for drug administration in recent years. Hydrogels are three-dimensional networks of polymer chains held together by crosslinks. Although the changes which the hydrogel undergoes in solution are important to a wide range of experimental studies, they have not been investigated systematically and the factors which influence the degree of swelling have not been adequately described. Hydrogels made of poly(ethylene glycol) (PEG) will generally resist degradation in aqueous conditions, while a hydrogel made from a copolymer of poly(lactic acid) (PLA) and PEG will degrade via hydrolysis of the lactic acid group. This ability to degrade makes these hydrogels promising candidates for controlled release drug delivery systems. The goal of this research was to characterize the swelling and degradation of both degradable and non-degradable gels and to evaluate the release of different drugs from these hydrogels, where the key variable is the molecular weight of the PEG segment. These hydrogels were formed by the addition and subsequent chemically crosslinking of methacrylate end groups. During crosslinking, both PEG and LA-PEG-LA hydrogels of varied PEG molecular weight were loaded with Vitamin B12, Insulin, Haloperidol, and Dextran. It was shown that increasing PEG molecular weight produces a hydrogel with larger pores, thus increasing water uptake and degradation rate. While many environmental factors do not affect the swelling behavior, they do significantly impact the degradation of the hydrogel, and thus the release of incorporated therapeutic agents.

Formation mechanism of glyoxal-DNA adduct, a DNA cross-link precursor.

PubMed

Vilanova, B; Fernández, D; Casasnovas, R; Pomar, A M; Alvarez-Idaboy, J R; Hernández-Haro, N; Grand, A; Adrover, M; Donoso, J; Frau, J; Muñoz, F; Ortega-Castro, J

2017-05-01

DNA nucleobases undergo non-enzymatic glycation to nucleobase adducts which can play important roles in vivo. In this work, we conducted a comprehensive experimental and theoretical kinetic study of the mechanisms of formation of glyoxal-guanine adducts over a wide pH range in order to elucidate the molecular basis for the glycation process. Also, we performed molecular dynamics simulations to investigate how open or cyclic glyoxal-guanine adducts can cause structural changes in an oligonucleotide model. A thermodynamic study of other glycating agents including methylglyoxal, acrolein, crotonaldehyde, 4-hydroxynonenal and 3-deoxyglucosone revealed that, at neutral pH, cyclic adducts were more stable than open adducts; at basic pH, however, the open adducts of 3-deoxyglucosone, methylglyoxal and glyoxal were more stable than their cyclic counterparts. This result can be ascribed to the ability of the adducts to cross-link DNA. The new insights may contribute to improve our understanding of the connection between glycation and DNA cross-linking. Copyright © 2017 Elsevier B.V. All rights reserved.

Effect of synthesis parameters on polymethacrylic acid xerogel structures and equilibrium swelling

NASA Astrophysics Data System (ADS)

Panić, V.; Jovanović, J.; Adnadjević, B.; Velicković, S.

2009-09-01

Hydrogels based on crosslinked polymethacrylic acid were synthesized via free-radical polymerization in aqueous solution, using N,N'-methylene bisacrylamide as a crosslinking agent and 2,2'-azobis-[2-(2-imidazolin-2-yl)propane] dihydrochloride as an initiator. The influence of the reaction parameters (the neutralization degree of methacrylic acid and the initial monomer concentration) on the equilibrium swelling degree, the swelling kinetic parameters and the basic structural properties of xerogels was investigated. The change of synthesis parameters leads to the change of the basic structural parameters of xerogel, as well as the equilibrium swelling degree and the initial swelling rate of the hydrogels. It is found that there are power form relationships between the equilibrium swelling degree, the initial swelling rate and the structural xerogel’s properties and the change of the neutralization degree of monomer, i.e. the monomer concentration. The examined correlations proved that the crosslinking density is the crucial parameter which determines all the other investigated structural and swelling parameters.

Antioxidant films based on cross-linked methyl cellulose and native Chilean berry for food packaging applications.

PubMed

López de Dicastillo, Carol; Rodríguez, Francisco; Guarda, Abel; Galotto, Maria José

2016-01-20

Development of antioxidant and antimicrobial active food packaging materials based on biodegradable polymer and natural plant extracts has numerous advantages as reduction of synthetic additives into the food, reduction of plastic waste, and food protection against microorganisms and oxidation reactions. In this way, active films based on methylcellulose (MC) and maqui (Aristotelia chilensis) berry fruit extract, as a source of antioxidants agents, were studied. On the other hand, due to the high water affinity of MC, this polymer was firstly cross-linked with glutaraldehyde (GA) at different concentrations. The results showed that the addition of GA decreased water solubility, swelling, water vapor permeability of MC films, and the release of antioxidant substances from the active materials increased with the concentration of GA. Natural extract and active cross-linked films were characterized in order to obtain the optimal formulation with the highest antioxidant activity and the best physical properties for latter active food packaging application. Copyright © 2015 Elsevier Ltd. All rights reserved.

Physicochemical characterization of chitosan/nylon6/polyurethane foam chemically cross-linked ternary blends.

PubMed

Jayakumar, S; Sudha, P N

2013-03-15

Chitosan/nylon6/polyurethane foam (CS/Ny6/PUF) ternary blend was prepared and chemically cross-linked with glutaraldehyde. Structural, thermal and morphological studies were performed for the prepared ternary blends. Characterizations of the ternary blends were investigated by Fourier transform infrared spectroscopy (FTIR), thermo gravimetric analysis (TGA), differential scanning calorimetry (DSC), X-ray diffraction (XRD) and scanning electron microscope (SEM). The FTIR results showed that the strong intermolecular hydrogen bonds took place between CS, Ny6 and PUF. TGA and DSC studies reveal that the thermal stability of the blend is enhanced by glutaraldehyde as crosslinking agent. Results of XRD indicated that the relative crystalline of pure CS film was reduced when the polymeric network was reticulated by glutaraldehyde. Finally, the results of scanning electron microscopy (SEM) indicated that the morphology of the blend is rough and heterogeneous, further it confirms the interaction between the functional groups of the blend components. Copyright © 2012 Elsevier B.V. All rights reserved.

Use of Melt Flow Rate Test in Reliability Study of Thermoplastic Encapsulation Materials in Photovoltaic Modules

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

Moseley, J.; Miller, D.; Shah, Q.-U.-A. S. J.

2011-10-01

Use of thermoplastic materials as encapsulants in photovoltaic (PV) modules presents a potential concern in terms of high temperature creep, which should be evaluated before thermoplastics are qualified for use in the field. Historically, the issue of creep has been avoided by using thermosetting polymers as encapsulants, such as crosslinked ethylene-co-vinyl acetate (EVA). Because they lack crosslinked networks, however, thermoplastics may be subject to phase transitions and visco-elastic flow at the temperatures and mechanical stresses encountered by modules in the field, creating the potential for a number of reliability and safety issues. Thermoplastic materials investigated in this study include PV-grademore » uncured-EVA (without curing agents and therefore not crosslinked); polyvinyl butyral (PVB); thermoplastic polyurethane (TPU); and three polyolefins (PO), which have been proposed for use as PV encapsulation. Two approaches were used to evaluate the performance of these materials as encapsulants: module-level testing and a material-level testing.« less

Influence of Unmodified and β-Glycerophosphate Cross-Linked Chitosan on Anti-Candida Activity of Clotrimazole in Semi-Solid Delivery Systems

PubMed Central

Szymańska, Emilia; Winnicka, Katarzyna; Wieczorek, Piotr; Sacha, Paweł Tomasz; Tryniszewska, Elżbieta Anna

2014-01-01

The combination of an antifungal agent and drug carrier with adjunctive antimicrobial properties represents novel strategy of complex therapy in pharmaceutical technology. The goal of this study was to investigate the unmodified and ion cross-linked chitosan’s influence on anti-Candida activity of clotrimazole used as a model drug in hydrogels. It was particularly crucial to explore whether the chitosans’ structure modification by β-glycerophosphate altered its antifungal properties. Antifungal studies (performed by plate diffusion method according to CLSI reference protocol) revealed that hydrogels obtained with chitosan/β-glycerophosphate displayed lower anti-Candida effect, probably as a result of weakened polycationic properties of chitosan in the presence of ion cross-linker. Designed chitosan hydrogels with clotrimazole were found to be more efficient against tested Candida strains and showed more favorable drug release profile compared to commercially available product. These observations indicate that novel chitosan formulations may be considered as promising semi-solid delivery system of clotrimazole. PMID:25272230

Influence of unmodified and β-glycerophosphate cross-linked chitosan on anti-Candida activity of clotrimazole in semi-solid delivery systems.

PubMed

Szymańska, Emilia; Winnicka, Katarzyna; Wieczorek, Piotr; Sacha, Paweł Tomasz; Tryniszewska, Elżbieta Anna

2014-09-30

The combination of an antifungal agent and drug carrier with adjunctive antimicrobial properties represents novel strategy of complex therapy in pharmaceutical technology. The goal of this study was to investigate the unmodified and ion cross-linked chitosan's influence on anti-Candida activity of clotrimazole used as a model drug in hydrogels. It was particularly crucial to explore whether the chitosans' structure modification by β-glycerophosphate altered its antifungal properties. Antifungal studies (performed by plate diffusion method according to CLSI reference protocol) revealed that hydrogels obtained with chitosan/β-glycerophosphate displayed lower anti-Candida effect, probably as a result of weakened polycationic properties of chitosan in the presence of ion cross-linker. Designed chitosan hydrogels with clotrimazole were found to be more efficient against tested Candida strains and showed more favorable drug release profile compared to commercially available product. These observations indicate that novel chitosan formulations may be considered as promising semi-solid delivery system of clotrimazole.

Combination Platinum-based and DNA Damage Response-targeting Cancer Therapy: Evolution and Future Directions

PubMed Central

Basourakos, Spyridon P.; Li, Likun; Aparicio, Ana M.; Corn, Paul G.; Kim, Jeri; Thompson, Timothy C.

2017-01-01

Maintenance of genomic stability is a critical determinant of cell survival and is necessary for growth and progression of malignant cells. Interstrand crosslinking (ICL) agents, including platinum-based agents, are first-line chemotherapy treatment for many solid human cancers. In malignant cells, ICL triggers the DNA damage response (DDR). When the damage burden is high and lesions cannot be repaired, malignant cells are unable to divide and ultimately undergo cell death either through mitotic catastrophe or apoptosis. The activities of ICL agents, in particular platinum-based therapies, establish a “molecular landscape,” i.e., a pattern of DNA damage that can potentially be further exploited therapeutically with DDR-targeting agents. If the molecular landscape created by platinum-based agents could be better defined at the molecular level, a systematic, mechanistic rationale(s) could be developed for the use of DDR-targeting therapies in combination/maintenance protocols for specific, clinically advanced malignancies. New therapeutic drugs such as poly(ADP-ribose) polymerase (PARP) inhibitors are examples of DDR-targeting therapies that could potentially increase the DNA damage and replication stress imposed by platinum-based agents in tumor cells and provide therapeutic benefit for patients with advanced malignancies. Recent studies have shown that the use of PARP inhibitors together with platinum-based agents is a promising therapy strategy for ovarian cancer patients with ”BRCAness”, i.e., a phenotypic characteristic of tumors that not only can involve loss-of-function mutations in either BRCA1 or BRCA2, but also encompasses the molecular features of BRCA-mutant tumors. On the basis of these promising results, additional mechanism-based studies focused on the use of various DDR-targeting therapies in combination with platinum-based agents should be considered. This review discusses, in general, (1) ICL agents, primarily platinum-based agents, that establish a molecular landscape that can be further exploited therapeutically; (2) multiple points of potential intervention after ICL agent–induced crosslinking that further predispose to cell death and can be incorporated into a systematic, therapeutic rationale for combination/maintenance therapy using DDR-targeting agents; and (3) available agents that can be considered for use in combination/maintenance clinical protocols with platinum-based agents for patients with advanced malignancies. PMID:27978798

A novel bio-orthogonal cross-linker for improved protein/protein interaction analysis.

PubMed

Nury, Catherine; Redeker, Virginie; Dautrey, Sébastien; Romieu, Anthony; van der Rest, Guillaume; Renard, Pierre-Yves; Melki, Ronald; Chamot-Rooke, Julia

2015-02-03

The variety of protein cross-linkers developed in recent years illustrates the current requirement for efficient reagents optimized for mass spectrometry (MS) analysis. To date, the most widely used strategy relies on commercial cross-linkers that bear an isotopically labeled tag and N-hydroxysuccinimid-ester (NHS-ester) moieties. Moreover, an enrichment step using liquid chromatography is usually performed after enzymatic digestion of the cross-linked proteins. Unfortunately, this approach suffers from several limitations. First, it requires large amounts of proteins. Second, NHS-ester cross-linkers are poorly efficient because of their fast hydrolysis in water. Finally, data analysis is complicated because of uneven fragmentation of complex isotopic cross-linked peptide mixtures. We therefore synthesized a new type of trifunctional cross-linker to overrule these limitations. This reagent, named NNP9, comprises a rigid core and bears two activated carbamate moieties and an azido group. NNP9 was used to establish intra- and intermolecular cross-links within creatine kinase, then to map the interaction surfaces between α-Synuclein (α-Syn), the aggregation of which leads to Parkinson's disease, and the molecular chaperone Hsc70. We show that NNP9 cross-linking efficiency is significantly higher than that of NHS-ester commercial cross-linkers. The number of cross-linked peptides identified was increased, and a high quality of MS/MS spectra leading to high sequence coverage was observed. Our data demonstrate the potential of NNP9 for an efficient and straightforward characterization of protein-protein interfaces and illustrate the power of using different cross-linkers to map thoroughly the surface interfaces within protein complexes.

In vitro cross-linking of elastin peptides and molecular characterization of the resultant biomaterials.

PubMed

Heinz, Andrea; Ruttkies, Christoph K H; Jahreis, Günther; Schräder, Christoph U; Wichapong, Kanin; Sippl, Wolfgang; Keeley, Fred W; Neubert, Reinhard H H; Schmelzer, Christian E H

2013-04-01

Elastin is a vital protein and the major component of elastic fibers which provides resilience to many vertebrate tissues. Elastin's structure and function are influenced by extensive cross-linking, however, the cross-linking pattern is still unknown. Small peptides containing reactive allysine residues based on sequences of cross-linking domains of human elastin were incubated in vitro to form cross-links characteristic of mature elastin. The resultant insoluble polymeric biomaterials were studied by scanning electron microscopy. Both, the supernatants of the samples and the insoluble polymers, after digestion with pancreatic elastase or trypsin, were furthermore comprehensively characterized on the molecular level using MALDI-TOF/TOF mass spectrometry. MS(2) data was used to develop the software PolyLinX, which is able to sequence not only linear and bifunctionally cross-linked peptides, but for the first time also tri- and tetrafunctionally cross-linked species. Thus, it was possible to identify intra- and intermolecular cross-links including allysine aldols, dehydrolysinonorleucines and dehydromerodesmosines. The formation of the tetrafunctional cross-link desmosine or isodesmosine was unexpected, however, could be confirmed by tandem mass spectrometry and molecular dynamics simulations. The study demonstrated that it is possible to produce biopolymers containing polyfunctional cross-links characteristic of mature elastin from small elastin peptides. MALDI-TOF/TOF mass spectrometry and the newly developed software PolyLinX proved suitable for sequencing of native cross-links in proteolytic digests of elastin-like biomaterials. The study provides important insight into the formation of native elastin cross-links and represents a considerable step towards the characterization of the complex cross-linking pattern of mature elastin. Copyright © 2013 Elsevier B.V. All rights reserved.

A general method for targeted quantitative cross-linking mass spectrometry

USDA-ARS?s Scientific Manuscript database

Chemical cross-linking mass spectrometry (XL-MS) provides protein structural information by identifying covalently linked proximal amino acid residues on protein surfaces. The information gained by this technique is complementary to other structural biology methods such as x-ray crystallography, NM...

Genome-Wide Profiling of RNA–Protein Interactions Using CLIP-Seq

PubMed Central

Stork, Cheryl; Zheng, Sika

2017-01-01

UV crosslinking immunoprecipitation (CLIP) is an increasingly popular technique to study protein–RNA interactions in tissues and cells. Whole cells or tissues are ultraviolet irradiated to generate a covalent bond between RNA and proteins that are in close contact. After partial RNase digestion, antibodies specific to an RNA binding protein (RBP) or a protein–epitope tag is then used to immunoprecipitate the protein–RNA complexes. After stringent washing and gel separation the RBP–RNA complex is excised. The RBP is protease digested to allow purification of the bound RNA. Reverse transcription of the RNA followed by high-throughput sequencing of the cDNA library is now often used to identify protein bound RNA on a genome-wide scale. UV irradiation can result in cDNA truncations and/or mutations at the crosslink sites, which complicates the alignment of the sequencing library to the reference genome and the identification of the crosslinking sites. Meanwhile, one or more amino acids of a crosslinked RBP can remain attached to its bound RNA due to incomplete digestion of the protein. As a result, reverse transcriptase may not read through the crosslink sites, and produce cDNA ending at the crosslinked nucleotide. This is harnessed by one variant of CLIP methods to identify crosslinking sites at a nucleotide resolution. This method, individual nucleotide resolution CLIP (iCLIP) circularizes cDNA to capture the truncated cDNA and also increases the efficiency of ligating sequencing adapters to the library. Here, we describe the detailed procedure of iCLIP. PMID:26965263

RNA–protein binding interface in the telomerase ribonucleoprotein

PubMed Central

Bley, Christopher J.; Qi, Xiaodong; Rand, Dustin P.; Borges, Chad R.; Nelson, Randall W.; Chen, Julian J.-L.

2011-01-01

Telomerase is a specialized reverse transcriptase containing an intrinsic telomerase RNA (TR) which provides the template for telomeric DNA synthesis. Distinct from conventional reverse transcriptases, telomerase has evolved a unique TR-binding domain (TRBD) in the catalytic telomerase reverse transcriptase (TERT) protein, integral for ribonucleoprotein assembly. Two structural elements in the vertebrate TR, the pseudoknot and CR4/5, bind TERT independently and are essential for telomerase enzymatic activity. However, the details of the TR–TERT interaction have remained elusive. In this study, we employed a photoaffinity cross-linking approach to map the CR4/5-TRBD RNA–protein binding interface by identifying RNA and protein residues in close proximity. Photoreactive 5-iodouridines were incorporated into the medaka CR4/5 RNA fragment and UV cross-linked to the medaka TRBD protein fragment. The cross-linking RNA residues were identified by alkaline partial hydrolysis and cross-linked protein residues were identified by mass spectrometry. Three CR4/5 RNA residues (U182, U187, and U205) were found cross-linking to TRBD amino acids Tyr503, Phe355, and Trp477, respectively. This CR4/5 binding pocket is distinct and separate from the previously proposed T pocket in the Tetrahymena TRBD. Based on homologous structural models, our cross-linking data position the essential loop L6.1 adjacent to the TERT C-terminal extension domain. We thus propose that stem-loop 6.1 facilitates proper TERT folding by interacting with both TRBD and C-terminal extension. Revealing the telomerase CR4/5-TRBD binding interface with single-residue resolution provides important insights into telomerase ribonucleoprotein architecture and the function of the essential CR4/5 domain. PMID:22123986

Interaction of collagen with chlorosulphonated paraffin tanning agents: Fourier transform infrared spectroscopic analysis and molecular dynamics simulations.

PubMed

Monti, Susanna; Bramanti, Emilia; Della Porta, Valentina; Onor, Massimo; D'Ulivo, Alessandro; Barone, Vincenzo

2013-09-21

The binding of chlorosulphonated paraffins to collagen triple helices is studied by means of classical molecular dynamics simulations and experimental spectroscopic techniques in order to disclose the principal characteristics of their interaction during the leather fattening process. Indeed, collagen is the main target to develop new leather modifying agents with specific characteristics, and an accurate design of the collagen binders, supported by predictive computational strategies, could be a successful tool to obtain new effective eco-compatible compounds able to impart to the leather the required functionalities and distinctive mechanical properties. Possible effects caused by the tanning agents on the collagen matrix have been identified from both experimental and theoretical points of view. Computational data in agreement with experiment have revealed that chlorosulphonated paraffins can interact favorably with the collagen residues having amine groups in their side chains (Arg, Lys, Asn and Gln) and reduce the tendency of the solvated collagen matrix to swell. However, the interference of chlorosulphonated paraffins with the unfolding process, which is operated mainly by the action of water, can be due both to covalent cross-linking of the collagen chains and intermolecular hydrogen bonding interactions involving also the hydroxyl groups of Hyp, Ser and Thr residues.

Green and Smart: Hydrogels to Facilitate Independent Practical Learning

ERIC Educational Resources Information Center

Hurst, Glenn A.

2017-01-01

A laboratory experiment was developed to enable students to investigate the use of smart hydrogels for potential application in targeted drug delivery. This is challenging for students to explore practically because of the extremely high risks of handling cross-linking agents such as glutaraldehyde. Genipin is a safe and green alternative that has…

Sulfur-Containing Organic-Inorganic Hybrid Gel Compositions and Aerogels

NASA Technical Reports Server (NTRS)

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

2017-01-01

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

Sulfur-Containing Organic-Inorganic Hybrid Gel Compositions and Aerogels

NASA Technical Reports Server (NTRS)

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

2015-01-01

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

21 CFR 172.230 - Microcapsules for flavoring substances.

Code of Federal Regulations, 2013 CFR

2013-04-01

... limitations Succinylated gelatin—Not to exceed 15 percent by combined weight of the microcapsule and flavoring oil. Succinic acid content of the gelatin is 4.5 to 5.5 percent. Arabinogalactan—Complying with § 172... Glutaraldehyde—As cross-linking agent for insolubilizing a coacervate of gum arabic and gelatin. n-Octyl alcohol...

21 CFR 172.230 - Microcapsules for flavoring substances.

Code of Federal Regulations, 2012 CFR

2012-04-01

... limitations Succinylated gelatin—Not to exceed 15 percent by combined weight of the microcapsule and flavoring oil. Succinic acid content of the gelatin is 4.5 to 5.5 percent. Arabinogalactan—Complying with § 172... Glutaraldehyde—As cross-linking agent for insolubilizing a coacervate of gum arabic and gelatin. n-Octyl alcohol...

Strandboard made from soy-based adhesive with high soy content

Treesearch

Zhiyong Cai; James M. Wescott; Jerrold E. Winandy

2005-01-01

A novel green adhesive with high soy content has recently been developed (13) with a process that denatures soy flour, modifies resulting protein with formaldehyde, and uses suitable phenolic crosslinking agents for copolymerization. Compared with mechanical and physical performances of oriented strandboard, the new adhesive showed promise for improving panel...

Polyvinyl pyridine microspheres

NASA Technical Reports Server (NTRS)

Rembaum, Alan (Inventor); Gupta, Amitava (Inventor); Volksen, Willi (Inventor)

1980-01-01

Microspheres are produced by cobalt gamma radiation initiated polymerization of a dilute aqueous vinyl pyridine solution. Addition of cross-linking agent provides higher surface area beads. Addition of monomers such as hydroxyethylmethacrylate acrylamide or methacrylamide increases hydrophilic properties and surface area of the beads. High surface area catalytic supports are formed in the presence of controlled pore glass substrate.

Polyvinyl pyridine microspheres

NASA Technical Reports Server (NTRS)

Rembaum, Alan (Inventor); Gupta, Amitava (Inventor); Volksen, Willi (Inventor)

1979-01-01

Microspheres are produced by cobalt gamma radiation initiated polymerization of a dilute aqueous vinyl pyridine solution. Addition of cross-linking agent provides higher surface area beads. Addition of monomers such as hydroxyethylmethacrylate acrylamide or methacrylamide increases hydrophilic properties and surface area of the beads. High surface area catalytic supports are formed in the presence of controlled pore glass substrate.

30 CFR 56.6000 - Definitions.

Code of Federal Regulations, 2010 CFR

2010-07-01

... Transportation in 49 CFR 173.53, and 173.100. This document is available at any MSHA Metal and Nonmetal Safety..., plus a thickener. Water gel. An explosive material containing substantial portions of water, oxidizers, and fuel, plus a cross-linking agent. [50 FR 4054, Jan. 29, 1985, as amended at 67 FR 38385, June 4...

Hemostatic granules and dressing prepared from formulations of carboxymethyl cellulose, kappa-carrageenan and polyethylene oxide crosslinked by gamma radiation

NASA Astrophysics Data System (ADS)

Barba, Bin Jeremiah D.; Aranilla, Charito T.; Relleve, Lorna S.; Cruz, Veriza Rita C.; Vista, Jeanina Richelle; Abad, Lucille V.

2018-03-01

Uncontrolled hemorrhage remains a persistent problem especially in anatomical areas where compression and tourniquet cannot be applied. Hemostatic agents are materials which can achieve control of bleeding in acute, life-threatening traumatic coagulopathy. In this study, we prepared biocompatible hydrogel-based hemostat crosslinked by ionizing radiation. Granules made from carboxymethyl cellulose and dressing from kappa carrageenan and polyethylene oxide were characterized by FT-IR, SEM, and gel analysis. Gamma radiation with a dose of 25 kGy was used for sterilization process. Stability studies indicate that the products remain effective with a shelf life of up to 18 months based on accelerated aging. Both hemostatic agents were demonstrated to be effective in vitro blood clotting assays showing a low blood clotting index, high platelet adhesion capacity and accelerated clotting time. Hemostat granules and dressing were also used in a femoral artery rat bleeding model where hemorrhage control was achieved in 90 s without compression and resulted in 100% survival rate after a 7 and 14-day observation.

The effect of EGDMA on tensile and thermal properties of irradiated low density polyethylene/sepiolite nanocomposites

NASA Astrophysics Data System (ADS)

Ghazali, Siti Nadia Aini; Mohamad, Zurina; Majid, Rohah A.; Appadu, Sivanesan

2017-07-01

This study presents the influence of ethylene glycol dimethacrylate (EGDMA) as a crosslinking agent through electron beam crosslinking process. Therefore, the effects of EGDMA on irradiated low density polyethylene/sepiolite (LDPE/SEP) nanocomposites on the tensile and thermal properties at 4 part per hundred resin (phr) sepiolite were investigated. The LDPE/SEP nanocomposites were prepared by melt mixing using twin screw extruder at 160 ˚C with a screw speed of 50 rpm. The nanocomposites were then undergone injection moulding process followed by irradiated using 2 MeV electron beam machine at doses ranging from 0 to 200 kGy in the air at ambient temperature. It was found that the tensile strength and Young's modulus were slightly increased with the presence of co-agent. The sample containing 4 phr sepiolite at 200 kGy showed 9% increase in tensile strength when EGDMA was added. However, the result of thermogravimetry analysis (TGA) showed some reduction in thermal stability of nanocomposites on 100 kGy irradiation dose. EGDMA had reduced the optimum irradiation dose without having any adverse effect on tensile and thermal properties.

Sodium alginate adhesives as binders in wood fibers/textile waste fibers biocomposites for building insulation.

PubMed

Lacoste, Clément; El Hage, Roland; Bergeret, Anne; Corn, Stéphane; Lacroix, Patrick

2018-03-15

Alginate derived from seaweed is a natural polysaccharide able to form stable gel through carbohydrate functional groups largely used in the food and pharmaceutical industry. This article deals with the use of sodium alginate as an adhesive binder for wood fibres/textile waste fibres biocomposites. Several aldehyde-based crosslinking agents (glyoxal, glutaraldehyde) were compared for various wood/textile waste ratios (100/0, 50/50, 60/40, 70/30 and 0/100 in weight). The fully biomass derived composites whose properties are herewith described satisfy most of the appropriate requirements for building materials. They are insulating with a thermal conductivity in the range 0.078-0.089 W/m/K for an average density in the range 308-333 kg/m3 according to the biocomposite considered. They are semi-rigid with a maximal mechanical strength of 0.84 MPa under bending and 0.44 MPa under compression for 60/40 w/w wood/textile waste biocomposites with a glutaraldehyde crosslinking agent. Copyright © 2017 Elsevier Ltd. All rights reserved.

Silver nanoparticles-containing dual-function hydrogels based on a guar gum-sodium borohydride system

PubMed Central

Dai, Lei; Nadeau, Ben; An, Xingye; Cheng, Dong; Long, Zhu; Ni, Yonghao

2016-01-01

Dual-function hydrogels, possessing both stimuli-responsive and self-healing properties, have recently attracted attention of both chemists and materials scientists. Here we report a new paradigm using natural polymer (guar gum, GG) and sodium borohydride (NaBH4), for the preparation of silver nanoparticles (AgNPs)-containing smart hydrogels in a simple, fast and economical way. NaBH4 performs as a reducing agent for AgNPs synthesis using silver nitrate (AgNO3) as the precursor. Meanwhile, sodium metaborate (NaBO2) (from NaBH4) behaves as a cross-linking agent between GG molecular chains. The AgNPs/GG hydrogels with excellent viscoelastic properties can be obtained within 3 min at room temperature without the addition of other cross-linkers. The resultant AgNPs/GG hydrogels are flowable and injectable, and they possess excellent pH/thermal responsive properties. Additionally, they exhibit rapid self-healing capacity. This work introduces a facile and scale-up way to prepare a class of hydrogels that can have great potential to biomedical and other industrial applications. PMID:27819289

Experimental scleral cross-linking increases glaucoma damage in a mouse model

PubMed Central

Kimball, Elizabeth C.; Nguyen, Cathy; Steinhart, Matthew R.; Nguyen, Thao D.; Pease, Mary E.; Oglesby, Ericka N.; Oveson, Brian C.; Quigley, Harry A.

2014-01-01

The purpose of this study was to assess the effect of a scleral cross-linking agent on susceptibility to glaucoma damage in a mouse model. CD1 mice underwent 3 subconjunctival injections of 0.5 M glyceraldehyde (GA) in 1 week, then had elevated intraocular pressure (IOP) induced by bead injection. Degree of cross-linking was measured by enzyme-linked immunosorbent assay (ELISA), scleral permeability was measured by fluorescence recovery after photobleaching (FRAP), and the mechanical effects of GA exposure were measured by inflation testing. Control mice had buffer injection or no injection in 2 separate glaucoma experiments. IOP was monitored by Tonolab and retinal ganglion cell (RGC) loss was measured by histological axon counting. To rule out undesirable effects of GA, we performed electroretinography and detailed histology of the retina. GA exposure had no detectable effects on RGC number, retinal structure or function either histologically or electrophysiologically. GA increased cross-linking of sclera by 37% in an ELISA assay, decreased scleral permeability (FRAP, p = 0.001), and produced a steeper pressure—strain behavior by in vitro inflation testing. In two experimental glaucoma experiments, GA-treated eyes had greater RGC axon loss from elevated IOP than either buffer-injected or control eyes, controlling for level of IOP exposure over time (p = 0.01, and 0.049, multivariable regression analyses). This is the first report that experimental alteration of the sclera, by cross-linking, increases susceptibility to RGC damage in mice. PMID:25285424

Radiation synthesis of biocompatible hydrogels of dextran methacrylate

NASA Astrophysics Data System (ADS)

Szafulera, Kamila; Wach, Radosław A.; Olejnik, Alicja K.; Rosiak, Janusz M.; Ulański, Piotr

2018-01-01

The aim of this work was to synthesize biocompatible dextran-based hydrogels through crosslinking initiated by ionizing radiation. A series of derivatives of dextran has been synthesized by coupling of methacrylated glycidyl to the structure of this polysaccharide, yielding dextran methacrylate (Dex-MA) of the degree of methacrylate substitution (DS) up to 1.13 as characterised by FTIR and NMR spectroscopy. Chemically crosslinked hydrogels were formed by electron-beam irradiation of Dex-MA in aqueous solution in the absence of low-molecular-weight additives such as catalysts, monomers or crosslinking agents. Crosslinking of Dex-MA in aqueous solutions of 20 g/l and above was an efficient process, the gels were formed at doses as low as 0.5 kGy (experiments conducted up to 100 kGy) and were characterised by high content of insoluble fraction (70-100%). Due to high crosslinking density the equilibrium degree of swelling of fabricated gels was controlled principally by the initial concentration of Dex-MA solution subjected to irradiation, and it was in the range of 20 to over 100 g of water absorbed by gram of gel. Cytocompatibility of hydrogels was examined using XTT assay through evaluation of the cell viability being in indirect contact with hydrogels. The results indicated that hydrogels of Dex-MA of the average DS below 1 were not cytotoxic. Altogether, our data demonstrate that irradiation of methacrylated dextran in aqueous solution is an efficient method of fabrication of biocompatible hydrogels, which applications in regeneration medicine are anticipated.

Smart surface coating of drug nanoparticles with cross-linkable polyethylene glycol for bio-responsive and highly efficient drug delivery.

PubMed

Wei, Weijia; Zhang, Xiujuan; Chen, Xianfeng; Zhou, Mengjiao; Xu, Ruirui; Zhang, Xiaohong

2016-04-21

Many drug molecules can be directly used as nanomedicine without the requirement of any inorganic or organic carriers such as silica and liposome nanostructures. This new type of carrier-free drug nanoparticles (NPs) has great potential in clinical treatment because of its ultra-high drug loading capacity and biodegradability. For practical applications, it is essential for such nanomedicine to possess robust stability and minimal premature release of therapeutic molecules during circulation in the blood stream. To meet this requirement, herein, we develop GSH-responsive and crosslinkable amphiphilic polyethylene glycol (PEG) molecules to modify carrier-free drug NPs. These PEG molecules can be cross-linked on the surface of the NPs to endow them with greater stability and the cross-link is sensitive to intracellular environment for bio-responsive drug release. With this elegant design, our experimental results show that the liberation of DOX from DOX-cross-linked PEG NPs is dramatically slower than that from DOX-non-cross-linked PEG NPs, and the DOX release profile can be controlled by tuning the concentration of the reducing agent to break the cross-link between PEG molecules. More importantly, in vivo studies reveal that the DOX-cross-linked PEG NPs exhibit favorable blood circulation half-life (>4 h) and intense accumulation in tumor areas, enabling effective anti-cancer therapy. We expect this work will provide a powerful strategy for stabilizing carrier-free nanomedicines and pave the way to their successful clinical applications in the future.

Effects of genipin corneal crosslinking in rabbit corneas.

PubMed

Avila, Marcel Y; Narvaez, Mauricio; Castañeda, Juan P

2016-07-01

To evaluate the effect of genipin, a natural crosslinking agent, in rabbit eyes. Department of Ophthalmology, Universidad Nacional de Colombia Centro de Tecnologia Oftalmica, Bogotá, Colombia. Experimental study. Ex vivo rabbit eyes (16; 8 rabbits) were treated with genipin 1.00%, 0.50%, and 0.25% for 5 minutes with a vacuum device to increase corneal permeability. Penetration was evaluated using Scheimpflug pachymetry (Pentacam). In the in vivo model (20 rabbits; 1 eye treated, 1 eye with vehicle), corneas were crosslinked with genipin as described. Corneal curvature, corneal pachymetry, and intraocular pressure (IOP) assessments as well as slitlamp examinations were performed 0, 7, 30, and 60 days after treatment. In the ex vivo model, Scheimpflug pachymetry showed deep penetration in the rabbit corneas with an increase in corneal density and a dose-dependent relationship. Corneal flattening was observed in treated eyes (mean 4.4 diopters ± 0.5 [SD]) compared with the control eyes. Pachymetry and IOP were stable in all evaluations. No eye showed toxicity in the anterior chamber or in the lens. Corneal crosslinking induced by genipin produced significant flattening of the cornea with no toxicity in rabbit eyes. This crosslinking could be useful in the treatment of corneal ectasia and in the modification of corneal curvature. None of the authors has a financial or proprietary interest in any material or method mentioned. Copyright © 2016 ASCRS and ESCRS. Published by Elsevier Inc. All rights reserved.

Ribose mediated crosslinking of collagen-hydroxyapatite hybrid scaffolds for bone tissue regeneration using biomimetic strategies.

PubMed

Krishnakumar, Gopal Shankar; Gostynska, Natalia; Campodoni, Elisabetta; Dapporto, Massimiliano; Montesi, Monica; Panseri, Silvia; Tampieri, Anna; Kon, Elizaveta; Marcacci, Maurilio; Sprio, Simone; Sandri, Monica

2017-08-01

This study explores for the first time the application of ribose as a highly biocompatible agent for the crosslinking of hybrid mineralized constructs, obtained by bio-inspired mineralization of self-assembling Type I collagen matrix with magnesium-doped-hydroxyapatite nanophase, towards a biomimetic mineralized 3D scaffolds (MgHA/Coll) with excellent compositional and structural mimicry of bone tissue. To this aim, two different crosslinking mechanisms in terms of pre-ribose glycation (before freeze drying) and post-ribose glycation (after freeze drying) were investigated. The obtained results explicate that with controlled freeze-drying, highly anisotropic porous structures with opportune macro-micro porosity are obtained. The physical-chemical features of the scaffolds characterized by XRD, FTIR, ICP and TGA demonstrated structural mimicry analogous to the native bone. The influence of ribose greatly assisted in decreasing solubility and increased enzymatic resistivity of the scaffolds. In addition, enhanced mechanical behaviour in response to compressive forces was achieved. Preliminary cell culture experiments reported good cytocompatibility with extensive cell adhesion, proliferation and colonization. Overall, scaffolds developed by pre-ribose glycation process are preferred, as the related crosslinking technique is more facile and robust to obtain functional scaffolds. As a proof of concept, we have demonstrated that ribose crosslinking is cost-effective, safe and functionally effective. This study also offers new insights and opportunities in developing promising scaffolds for bone tissue engineering. Copyright © 2017 Elsevier B.V. All rights reserved.

Novel soy protein wound dressings with controlled antibiotic release: mechanical and physical properties.

PubMed

Peles, Zachi; Zilberman, Meital

2012-01-01

Naturally derived materials are becoming widely used in the biomedical field. Soy protein has advantages over various types of natural proteins employed for biomedical applications due to its low price, non-animal origin and relatively long storage time and stability. In the current study soy protein isolate (SPI) was investigated as a matrix for wound dressing applications. The antibiotic drug gentamicin was incorporated into the matrix for local controlled release and, thus, protection against bacterial infection. Homogeneous yellowish films were cast from aqueous solutions. After cross-linking they combined high tensile strength and Young's modulus with the desired ductility. The plasticizer type, cross-linking agent and method of cross-linking were found to strongly affect the tensile properties of the SPI films. Selected SPI films were tested for relevant physical properties and the gentamicin release profile. The cross-linking method affected the degree of water uptake and the weight loss profile. The water vapor transmission rate of the films was in the desired range for wound dressings (∼2300 g m(-2) day(-1)) and was not affected by the cross-linking method. The gentamicin release profile exhibited a moderate burst effect followed by a decreasing release rate which was maintained for at least 4 weeks. Diffusion was the dominant release mechanism of gentamicin from cross-linked SPI films. Appropriate selection of the process parameters yielded SPI wound dressings with the desired mechanical and physical properties and drug release behavior to protect against bacterial infection. These unique structures are thus potentially useful as burn and ulcer dressings. Copyright © 2011 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

In vivo evaluation of folate decorated cross-linked micelles for the delivery of platinum anticancer drugs.

PubMed

Eliezar, Jeaniffer; Scarano, Wei; Boase, Nathan R B; Thurecht, Kristofer J; Stenzel, Martina H

2015-02-09

The biodistribution of micelles with and without folic acid targeting ligands were studied using a block copolymer consisting of acrylic acid (AA) and polyethylene glycol methyl ether acrylate (PEGMEA) blocks. The polymers were prepared using RAFT polymerization in the presence of a folic acid functionalized RAFT agent. Oxoplatin was conjugated onto the acrylic acid block to form amphiphilic polymers which, when diluted in water, formed stable micelles. In order to probe the in vivo stability, a selection of micelles were cross-linked using 1,8-diamino octane. The sizes of the micelles used in this study range between 75 and 200 nm, with both spherical and worm-like conformation. The effects of cross-linking, folate conjugation and different conformation on the biodistribution were studied in female nude mice (BALB/c) following intravenous injection into the tail vein. Using optical imaging to monitor the fluorophore-labeled polymer, the in vivo biodistribution of the micelles was monitored over a 48 h time-course after which the organs were removed and evaluated ex vivo. These experiments showed that both cross-linking and conjugation with folic acid led to increased fluorescence intensities in the organs, especially in the liver and kidneys, while micelles that are not conjugated with folate and not cross-linked are cleared rapidly from the body. Higher accumulation in the spleen, liver, and kidneys was also observed for micelles with worm-like shapes compared to the spherical micelles. While the various factors of cross-linking, micelle shape, and conjugation with folic acid all contribute separately to prolong the circulation time of the micelle, optimization of these parameters for drug delivery devices could potentially overcome adverse effects such as liver and kidney toxicity.

Transparent oxygen and water vapor barriers for flexible electronics using semi-crystalline polymer matrix thin films

NASA Astrophysics Data System (ADS)

Sehgal, Akhil

Electronic components such as organic light emitting diodes (OLED) and photo-voltaics have been of more focus with the advancement of technology. These electronics are susceptible to degradable in the presence of gases such as water vapor and oxygen. Being that these gases are constituents of the atmosphere and can be found in nearly every environment, certain protocols must take place to mitigate the issues that occur. New generation electronics are sensitive to oxidation and corrosion in the presence of extremely low concentrations of moisture and oxygen and therefore the development and improvements of gas barriers are vital for advancements in electronics technology. The improvements of appliances such as flexible solar cells and OLEDs require barriers that need to be flexible in order to achieve high longevity. The area of research has been focused on designing flexible polymer films with composite nanoparticles and cross-linking agents that have low permeability to moisture and oxygen gas. The polymers studied are in the family of methacrylates. Due to the properties of methacrylate polymers, it has been proposed that they are capable of having efficient barrier properties due to their ability to cross link and form crystalline structures with low chain mobility. The change in intensities of the FTIR peaks of different functional groups indicates the cross-linking and crystallinity of the polymer films. The UV-Vis data indicates high transparency of the films. SEM images of the films show continuous and well cured surfaces with minimal deviations, pores and defects. The addition of cross-linking agents and nanoparticles increased polymerization and cross-linking of the methacrylate polymer chains, therefore increasing inter-chain density and long range order. The incorporation of these additives increased the crystallinity of the films and by decreasing the distances and number of voids between polymer chains along with having minimal sorption sites for gases to bond to, the ability of gases such as moisture and oxygen to penetrate through the films has decreased.

Albumin-genipin solder for laser tissue repair.

PubMed

Lauto, A; Foster, L J R; Ferris, L; Avolio, A; Zwaneveld, N; Poole-Warren, L A

2004-01-01

Laser tissue soldering (LTS) is an alternative technique to suturing for tissue repair that avoids foreign body reaction and provides immediate sealing of the wound. One of the major drawbacks of LTS, however, is the weak tensile strength of the solder welds when compared to sutures. In this study, a crosslinking agent of low cytotoxicity was investigated for its ability to enhance the bond strength of albumin solders with sheep intestine. Solder strips were welded onto rectangular sections of sheep small intestine using a diode laser. The laser delivered in continuous mode a power of 170 +/- 10 mW at lambda = 808 nm, through a multimode optical fiber (core size = 200 microm) to achieve a dose of 10.8 +/- 0.5 J/mg. The solder thickness and surface area were kept constant throughout the experiment (thickness = 0.15 +/- 0.01 mm, area = 12 +/- 1.2 mm2). The solder was composed of 62% bovine serum albumin (BSA), 0.38% genipin, 0.25% indocyanin green dye (IG), and water. Tissue welding was also performed with a BSA solder without genipin, as a control group. The repaired tissue was tested for tensile strength by a calibrated tensiometer. Murine fibroblasts were also cultured in extracted media from heat-denatured genipin solder to assess cell growth inhibition in a 48 hours period. The tensile strength of the genipin solder was doubled that of the BSA solder (0.21 +/- 0.04 N and 0.11 +/- 0.04 N, respectively; P = 10(-15) unpaired t-test, N = 30). Media extracted from crosslinked genipin solder showed negligible toxicity to fibroblast cells under the culture conditions examined here. Addition of a chemical crosslinking agent, such as genipin, significantly increased the tensile strength of adhesive-tissue bonds. A proposed mechanism for this enhanced bond strength is the synergistic action of mechanical adhesion with chemical crosslinking by genipin.

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

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

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

2005-03-01

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

Tunable Enzymatic Activity and Enhanced Stability of Cellulase Immobilized in Biohybrid Nanogels.

PubMed

Peng, Huan; Rübsam, Kristin; Jakob, Felix; Schwaneberg, Ulrich; Pich, Andrij

2016-11-14

This paper reports a facile approach for encapsulation of enzymes in nanogels. Our approach is based on the use of reactive copolymers able to get conjugated with enzyme and build 3D colloidal networks or biohybrid nanogels. In a systematic study, we address the following question: how the chemical structure of nanogel network influences the biocatalytic activity of entrapped enzyme? The developed method allows precise control of the enzyme activity and improvement of enzyme resistance against harsh store conditions, chaotropic agents, and organic solvents. The nanogels were constructed via direct chemical cross-linking of water-soluble reactive copolymers poly(N-vinylpyrrolidone-co-N-methacryloxysuccinimide) with proteins such as enhanced green fluorescent protein (EGFP) and cellulase in water-in-oil emulsion. The water-soluble reactive copolymers with controlled amount of reactive succinimide groups and narrow dispersity were synthesized via reversible addition-fragmentation chain transfer (RAFT) polymerization. Poly(ethylene glycol) bis(3-aminopropyl) and branched polyethylenimine were utilized as model cross-linkers to optimize synthesis of nanogels with different architectures in the preliminary experiments. Biofluorescent nanogels with different loading amount of EGFP and varying cross-linking densities were obtained. We demonstrate that the biocatalytic activity of cellulase-conjugated nanogels (CNG) can be elegantly tuned by control of their cross-linking degrees. Circular dichroism (CD) spectra demonstrated that the secondary structures of the immobilized cellulase were changed in the aspect of α-helix contents. The secondary structures of cellulase in highly cross-linked nanogels were strongly altered compared with loosely cross-linked nanogels. The fluorescence resonance energy transfer (FRET) based study further revealed that nanogels with lower cross-linking degree enable higher substrate transport rate, providing easier access to the active site of the enzyme. The biohybrid nanogels demonstrated significantly improved stability in preserving enzymatic activity compared with free cellulase. The functional biohybrid nanogels with tunable enzymatic activity and improved stability are promising candidates for applications in biocatalysis, biomass conversion, or energy utilization fields.

Microencapsulation of citronella oil for mosquito-repellent application: formulation and in vitro permeation studies.

PubMed

Solomon, B; Sahle, F F; Gebre-Mariam, T; Asres, K; Neubert, R H H

2012-01-01

Citronella oil (CO) has been reported to possess a mosquito-repellent action. However, its application in topical preparations is limited due to its rapid volatility. The objective of this study was therefore to reduce the rate of evaporation of the oil via microencapsulation. Microcapsules (MCs) were prepared using gelatin simple coacervation method and sodium sulfate (20%) as a coacervating agent. The MCs were hardened with a cross-linking agent, formaldehyde (37%). The effects of three variables, stirring rate, oil loading and the amount of cross-linking agent, on encapsulation efficiency (EE, %) were studied. Response surface methodology was employed to optimize the EE (%), and a polynomial regression model equation was generated. The effect of the amount of cross-linker was insignificant on EE (%). The response surface plot constructed for the polynomial equation provided an optimum area. The MCs under the optimized conditions provided EE of 60%. The optimized MCs were observed to have a sustained in vitro release profile (70% of the content was released at the 10th hour of the study) with minimum initial burst effect. Topical formulations of the microencapsulated oil and non-microencapsulated oil were prepared with different bases, white petrolatum, wool wax alcohol, hydrophilic ointment (USP) and PEG ointment (USP). In vitro membrane permeation of CO from the ointments was evaluated in Franz diffusion cells using cellulose acetate membrane at 32 °C, with the receptor compartment containing a water-ethanol solution (50:50). The receptor phase samples were analyzed with GC/MS, using citronellal as a reference standard. The results showed that microencapsulation decreased membrane permeation of the CO by at least 50%. The amount of CO permeated was dependent on the type of ointment base used; PEG base exhibited the highest degree of release. Therefore, microencapsulation reduces membrane permeation of CO while maintaining a constant supply of the oil. Copyright © 2011 Elsevier B.V. All rights reserved.

Tungsten Disulfide Nanotubes Reinforced Biodegradable Polymers for Bone Tissue Engineering

PubMed Central

Lalwani, Gaurav; Henslee, Allan M.; Farshid, Behzad; Parmar, Priyanka; Lin, Liangjun; Qin, Yi-Xian; Kasper, F. Kurtis; Mikos, Antonios G.; Sitharaman, Balaji

2013-01-01

In this study, we have investigated the efficacy of inorganic nanotubes as reinforcing agents to improve the mechanical properties of poly(propylene fumarate) (PPF) composites as a function of nanomaterial loading concentration (0.01-0.2 wt%). Tungsten disulfide nanotubes (WSNTs) were used as reinforcing agents in the experimental groups. Single- and multi- walled carbon nanotubes (SWCNTs and MWCNTs) were used as positive controls, and crosslinked PPF composites were used as baseline control. Mechanical testing (compression and three-point bending) shows a significant enhancement (up to 28-190%) in the mechanical properties (compressive modulus, compressive yield strength, flexural modulus, and flexural yield strength) of WSNT reinforced PPF nanocomposites compared to the baseline control. In comparison to positive controls, at various concentrations, significant improvements in the mechanical properties of WSNT nanocomposites were also observed. In general, the inorganic nanotubes (WSNTs) showed a better (up to 127%) or equivalent mechanical reinforcement compared to carbon nanotubes (SWCNTs and MWCNTs). Sol fraction analysis showed significant increases in the crosslinking density of PPF in the presence of WSNTs (0.01-0.2 wt%). Transmission electron microscopy (TEM) analysis on thin sections of crosslinked nanocomposites showed the presence of WSNTs as individual nanotubes in the PPF matrix, whereas SWCNTs and MWCNTs existed as micron sized aggregates. The trend in the surface area of nanostructures obtained by BET surface area analysis was SWCNTs > MWCNTs > WSNTs. The BET surface area analysis, TEM analysis, and sol fraction analysis results taken together suggest that chemical composition (inorganic vs. carbon nanomaterials), presence of functional groups (such as sulfide and oxysulfide), and individual dispersion of the nanomaterials in the polymer matrix (absence of aggregation of the reinforcing agent) are the key parameters affecting the mechanical properties of nanostructure-reinforced PPF composites, and the reason for the observed increases in the mechanical properties compared to the baseline and positive controls. PMID:23727293

In situ diselenide-crosslinked polymeric micelles for ROS-mediated anticancer drug delivery.

PubMed

Deepagan, V G; Kwon, Seunglee; You, Dong Gil; Nguyen, Van Quy; Um, Wooram; Ko, Hyewon; Lee, Hansang; Jo, Dong-Gyu; Kang, Young Mo; Park, Jae Hyung

2016-10-01

Stimuli-responsive micelles have emerged as the drug carrier for cancer therapy since they can exclusively release the drug via their structural changes in response to the specific stimuli of the target site. Herein, we developed the in situ diselenide-crosslinked micelles (DCMs), which are responsive to the abnormal ROS levels of tumoral region, as anticancer drug carriers. The DCMs were spontaneously derived from selenol-bearing triblock copolymers consisting of polyethylene glycol (PEG) and polypeptide derivatives. During micelle formation, doxorubicine (DOX) was effectively encapsulated in the hydrophobic core, and diselenide crosslinks were formed in the shell. The DCMs maintained their structural integrity, at least for 6 days in physiological conditions, even in the presence of destabilizing agents. However, ROS-rich conditions triggered rapid release of DOX from the DOX-encapsulating DCMs (DOX-DCMs) because the hydrophobic diselenide bond was cleaved into hydrophilic selenic acid derivatives. Interestingly, after their systemic administration into the tumor-bearing mice, DOX-DCMs delivered significantly more drug to tumors (1.69-fold and 3.73-fold higher amount compared with their non-crosslinked counterparts and free drug, respectively) and effectively suppressed tumor growth. Overall, our data indicate that DCMs have great potential as drug carriers for anticancer therapy. Copyright © 2016 Elsevier Ltd. All rights reserved.

Eliminating glutaraldehyde from crosslinked collagen films using supercritical CO2.

PubMed

Casali, Dominic M; Yost, Michael J; Matthews, Michael A

2018-01-01

Collagen has received considerable attention as a biomaterial for tissue engineering because of its low immunogenicity, controllable biodegradation, and ability to influence cell growth and proliferation. Frequently, collagen scaffolds require crosslinking to improve mechanical strength, requiring agents like glutaraldehyde that have high residual cytotoxicity. A novel method for extracting residual glutaraldehyde from crosslinked collagen films with supercritical carbon dioxide (CO 2 ) is presented. CO 2 is a nontoxic, nonflammable substance that is relatively inert and can be used to process biomaterials at mild pressures and physiologic temperatures. In this work, it was first determined that type I collagen is chemically compatible with both liquid and supercritical CO 2 . Treated collagen showed minimal changes in physicochemical properties as determined by differential scanning calorimetry, gel electrophoresis, and circular dichroism. CO 2 was subsequently used to extract residual glutaraldehyde from crosslinked collagen films. Glutaraldehyde concentration was reduced by over 95%, from over 20 ppm before treatment to about 1 ppm, in only 1 h. CO 2 treatment caused negligible alteration of thermal stability but did significantly increase film stiffness and tensile strength. However, these changes were minor compared to heat-based removal of glutaraldehyde. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 86-94, 2018. © 2017 Wiley Periodicals, Inc.

Disulfide cross-linked polyurethane micelles as a reduction-triggered drug delivery system for cancer therapy.

PubMed

Yu, Shuangjiang; Ding, Jianxun; He, Chaoliang; Cao, Yue; Xu, Weiguo; Chen, Xuesi

2014-05-01

Nanoscale carriers that stably load drugs in blood circulation and release the payloads in desirable sites in response to a specific trigger are of great interest for smart drug delivery systems. For this purpose, a novel type of disulfide core cross-linked micelles, which are facilely fabricated by cross-linking of poly(ethylene glycol)/polyurethane block copolymers containing cyclic disulfide moieties via a thiol-disulfide exchange reaction, are developed. A broad-spectrum anti-cancer drug, doxorubicin (DOX), is loaded into the micelles as a model drug. The drug release from the core cross-linked polyurethane micelles (CCL-PUMs) loaded with DOX is suppressed in normal phosphate buffer saline (PBS), whereas it is markedly accelerated with addition of an intracellular reducing agent, glutathione (GSH). Notably, although DOX-loaded CCL-PUMs display lower cytotoxicity in vitro compared to either free DOX or DOX-loaded uncross-linked polyurethane micelles, the drug-loaded CCL-PUMs show the highest anti-tumor efficacy with reduced toxicity in vivo. Since enhanced anti-tumor efficacy and reduced toxic side effects are key aspects of efficient cancer therapy, the novel reduction-responsive CCL-PUMs may hold great potential as a bio-triggered drug delivery system for cancer therapy. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Superabsorbent biphasic system based on poly(lactic acid) and poly(acrylic acid)

NASA Astrophysics Data System (ADS)

Sartore, Luciana; Pandini, Stefano; Baldi, Francesco; Bignotti, Fabio

2016-05-01

In this research work, biocomposites based on crosslinked particles of poly(acrylic acid), commonly used as superabsorbent polymer (SAP), and poly-L-lactic acid (PLLA) were developed to elucidate the role of the filler (i.e., polymeric crosslinked particles) on the overall physico-mechanical behavior and to obtain superabsorbent thermoplastic products. Samples prepared by melt-blending of components in different ratios showed a biphasic system with a regular distribution of particles, with diameter ranging from 5 to 10 μm, within the PLLA polymeric matrix. The polymeric biphasic system, coded PLASA i.e. superabsorbent poly(lactic acid), showed excellent swelling properties, demonstrating that cross-linked particles retain their superabsorbent ability, as in their free counterparts, even if distributed in a thermoplastic polymeric matrix. The thermal characteristics of the biocomposites evidence enhanced thermal stability in comparison with neat PLLA and also mechanical properties are markedly modified by addition of crosslinked particles which induce regular stiffening effect. Furthermore, in aqueous environments the particles swell and are leached from PLLA matrix generating very high porosity. These new open-pore PLLA foams, produced in absence of organic solvents and chemical foaming agents, with good physico-mechanical properties appear very promising for several applications, for instance in tissue engineering for scaffold production.

Chitosan-starch beads prepared by ionotropic gelation as potential matrices for controlled release of fertilizers.

PubMed

Perez, Jonas J; Francois, Nora J

2016-09-05

The present study examines the agrochemical application of macrospheres prepared with chitosan and chitosan-starch blends by an easy dripping technique, using a sodium tripolyphosphate aqueous solution as the crosslinking agent. These biopolymers form hydrogels that could be a viable alternative method to obtain controlled-release fertilizers (CRFs). Three different concentrations (ranging from 20 to 100wt/wt% of chitosan) and two crosslinking times (2 or 4h) were used. The resulting polymeric matrices were examined by scanning electron microscopy coupled with energy dispersive X-ray, X-ray diffraction, Fourier transform infrared spectroscopy, solid-state nuclear magnetic resonance, thermogravimetric analysis and differential scanning calorimetry. Ionotropic gelation and neutralization induced the formation of the macrospheres. The crosslinking time and the composition of the polymeric hydrogel controlled the crosslinking degree, the swelling behavior and the fertilizer loading capability. Potassium nitrate-loaded beads were shown to be useful as a controlled-release fertilizer. After 14days of continuous release into distilled water, the cumulative concentration in the release medium reached between 70 and 93% of the initially loaded salt, depending on the matrix used. The prepared beads showed properties that make them suitable for use in the agrochemical industry as CRFs. Copyright © 2016 Elsevier Ltd. All rights reserved.

Enteric-coated epichlorohydrin crosslinked dextran microspheres for site-specific delivery to colon.

PubMed

Rai, Gopal; Yadav, Awesh K; Jain, Narendra K; Agrawal, Govind P

2015-01-01

Enteric-coated epichlorohydrin crosslinked dextran microspheres containing 5-Fluorouracil (5-FU) for colon drug delivery was prepared by emulsification-crosslinking method. The formulation variables studied includes different molecular weights of dextran, volume of crosslinking agent, stirring speed, time and temperature. Dextran microspheres showed mean entrapment efficiencies ranging between 77 and 87% and mean particle size ranging between 10 and 25 µm. About 90% of drug was released from uncoated dextran microspheres within 8 h, suggesting the fast release and indicated the drug loaded in uncoated microspheres, released before they reached colon. Enteric coating (Eudragit-S-100 and Eudragit-L-100) of dextran microspheres was performed by oil-in-oil solvent evaporation method. The release study of 5-FU from coated dextran microspheres was complete retardation in simulated gastric fluid (pH 1.2) and once the coating layer of enteric polymer was dissolved at higher pH (7.4 and 6.8), a controlled release of the drug from the microspheres was observed. Further, the release of drug was found to be higher in the presence of dextranase and rat caecal contents, indicating the susceptibility of dextran microspheres to colonic enzymes. Organ distribution and pharmacokinetic study in albino rats was performed to establish the targeting potential of optimized formulation in the colon.

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

PubMed

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

2004-12-31

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

A PI 4. 6 peroxidase that specifically crosslinks extensin precursors

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

Upham, B.L; Alizadeh, H.; Ryan, K.J.

1991-05-01

The primary cell wall is a microcomposite of cellulose, pectin, hemicellulose and protein. The warp-weft model of the primary cell wall hypothesize that extensin monomers are intermolecularly crosslinked orthogonal to the cellulose microfibril thus mechanically coupling the major load-bearing polymer: cellulose. Media of tomato cell cultures contains heat labile, peroxide dependent crosslinking activity, as determined by the rate of decrease in monomer concentration analyzed via Superose-6. Isoelectric focusing of tomato cell culture media indicated crosslinking was predominantly in the acidic peroxidase fraction (pI4.6). This peroxidase was partially purified by ultracentrifugation, DEAE-Trisacryl and HPLC-DEAE chromatography techniques resulting in a 90 foldmore » purification and 45% yield. A second acidic peroxidase eluted from the HPLC-DEAE column had 25% of the crosslinking activity of the pI 4.6 peroxidase. Purified basic peroxidase had only 0.7% of the activity of the pI 4.6 peroxidase. The specific activity of the pI 4.6 peroxidase was 5,473 mg extensin crosslinked/min/mg peroxidase. The pI 4.6 peroxidase crosslinked the following extensins: tomato I and II, carrot, Ginkgo II and did not crosslink Ginkgo I, Douglas Fir, Maize, Asparagus I and II, and sugarbeet extensins as well as bovine serum albumin. Comparison of motifs common to extensins that are crosslinked by the pI 4.6 peroxidase may help identify the crosslink domain(s) of extension.« less

Cellular and molecular consequences of defective Fanconi anemia proteins in replication-coupled DNA repair: mechanistic insights

PubMed Central

Thompson, Larry H.; Hinz, John M.

2009-01-01

The Fanconi anemia (FA) molecular network consists of 15 “FANC” proteins, of which 13 are associated with mutations in patients with this cancer-prone chromosome instability disorder. Whereas historically the common phenotype associated with FA mutations is marked sensitivity to DNA interstrand crosslinking agents, the literature supports a more global role for FANC proteins in coping with diverse stresses encountered by replicative polymerases. We have attempted to reconcile and integrate numerous observations into a model in which FANC proteins coordinate the following physiological events during DNA crosslink repair: (a) activating a FANCM-ATR-dependent S-phase checkpoint; (b) mediating enzymatic replication-fork breakage and crosslink unhooking; (c) filling the resulting gap by translesion synthesis (TLS) by error-prone polymerase(s); and (d) restoring the resulting one-ended double-strand break by homologous recombination repair (HRR). The FANC core subcomplex (FANCA, B, C, E, F, G, L, FAAP100) promotes TLS for both crosslink and non-crosslink damage such as spontaneous oxidative base damage, UV-C photoproducts, and alkylated bases. TLS likely helps prevent stalled replication forks from breaking, thereby maintaining chromosome continuity. Diverse DNA damages and replication inhibitors result in monoubiquitination of the FANCD2-FANCI complex by the FANCL ubiquitin ligase activity of the core subcomplex upon its recruitment to chromatin by the FANCM-FAAP24 heterodimeric translocase. We speculate that this translocase activity acts as the primary damage sensor and helps remodel blocked replication forks to facilitate checkpoint activation and repair. Monoubiquitination of FANCD2-FANCI is needed for promoting HRR, in which the FANCD1/BRCA2 and FANCN/PALB2 proteins act at an early step. We conclude that the core subcomplex is required for both TLS and HRR occurring separately for non-crosslink damages and for both events during crosslink repair. The FANCJ/BRIP1/BACH1 helicase functions in association with BRCA1 and may remove structural barriers to replication, such as guanine quadruplex structures, and/or assist in crosslink unhooking. PMID:19622404

Application of molecular simulation to investigate chrome(III)-crosslinked collagen problems

NASA Astrophysics Data System (ADS)

Ding, Yun-Qiao; Chen, Cheng-Lung; Gu, Qi-Rui; Liao, Jun-Min; Chuang, Po-Hsiang

2014-04-01

Molecular dynamics simulation with a modified CHARMM (Chemistry at Harvard Macromolecular Mechanics) force field was carried out to investigate the properties of chrome-tanned collagen in comparison with chrome-free collagen under hydrated and dehydrated conditions. An attempt has been made to explain the microcosmic origins of the various properties of the chromium(III)-crosslinked collagen. The present simulation describes the clear crosslinking topology of polychromiums to peptide chains, identifies the linking site and the capacity of the linkage, explains why the efficiency is not 100% in a practical tanning process and provides a new viewpoint on the crosslinking of the polychromium with the side chains of the collagen.

Ara h 2 cross-linking catalyzed by MTGase decreases its allergenicity.

PubMed

Wu, Zhihua; Lian, Jun; Zhao, Ruifang; Li, Kun; Li, Xin; Yang, Anshu; Tong, Ping; Chen, Hongbing

2017-03-22

Peanuts, whose major allergen is Ara h 2, are included among the eight major food allergens. After reduction using dithiothreitol (DTT), cross-linking of Ara h 2 could be catalyzed by microbial transglutaminase (MTGase), a widely used enzyme in the food industry. In this study, Ara h 2 cross-linking was catalyzed by MTGase after it was reduced by DTT. Using mass spectrometry and PLINK software, five cross-linkers were identified, and five linear allergen epitopes were found to be involved in the reactions. The IgE binding capacity of cross-linked Ara h 2 was found to be significantly lower compared to that of native and reduced Ara h 2. After simulated gastric fluid (SGF) digestion, the digested products of the cross-linked Ara h 2, again, had a significantly lower IgE binding capacity compared to untreated and reduced Ara h 2. Furthermore, reduced and cross-linked Ara h 2 (RC-Ara h 2) induced lower sensitization in mice, indicating its lower allergenicity. Reduction and MTGase-catalyzed cross-linking are effective methods to decrease the allergenicity of Ara h 2. The reactions involved linear allergen epitopes destroying the material basis of the allergenicity, and this might develop a new direction for protein desensitization processes.

Permeability Evaluation Through Chitosan Membranes Using Taguchi Design

PubMed Central

Sharma, Vipin; Marwaha, Rakesh Kumar; Dureja, Harish

2010-01-01

In the present study, chitosan membranes capable of imitating permeation characteristics of diclofenac diethylamine across animal skin were prepared using cast drying method. The effect of concentration of chitosan, concentration of cross-linking agent (NaTPP), crosslinking time was studied using Taguchi design. Taguchi design ranked concentration of chitosan as the most important factor influencing the permeation parameters of diclofenac diethylamine. The flux of the diclofenac diethylamine solution through optimized chitosan membrane (T9) was found to be comparable to that obtained across rat skin. The mathematical model developed using multilinear regression analysis can be used to formulate chitosan membranes that can mimic the desired permeation characteristics. The developed chitosan membranes can be utilized as a substitute to animal skin for in vitro permeation studies. PMID:21179329

Permeability evaluation through chitosan membranes using taguchi design.

PubMed

Sharma, Vipin; Marwaha, Rakesh Kumar; Dureja, Harish

2010-01-01

In the present study, chitosan membranes capable of imitating permeation characteristics of diclofenac diethylamine across animal skin were prepared using cast drying method. The effect of concentration of chitosan, concentration of cross-linking agent (NaTPP), crosslinking time was studied using Taguchi design. Taguchi design ranked concentration of chitosan as the most important factor influencing the permeation parameters of diclofenac diethylamine. The flux of the diclofenac diethylamine solution through optimized chitosan membrane (T9) was found to be comparable to that obtained across rat skin. The mathematical model developed using multilinear regression analysis can be used to formulate chitosan membranes that can mimic the desired permeation characteristics. The developed chitosan membranes can be utilized as a substitute to animal skin for in vitro permeation studies.

Hierarchically assembled theranostic nanostructures for siRNA delivery and imaging applications.

PubMed

Shrestha, Ritu; Elsabahy, Mahmoud; Luehmann, Hannah; Samarajeewa, Sandani; Florez-Malaver, Stephanie; Lee, Nam S; Welch, Michael J; Liu, Yongjian; Wooley, Karen L

2012-10-24

Dual functional hierarchically assembled nanostructures, with two unique functions of carrying therapeutic cargo electrostatically and maintaining radiolabeled imaging agents covalently within separate component building blocks, have been developed via the supramolecular assembly of several spherical cationic shell cross-linked nanoparticles clustered around a central anionic shell cross-linked cylinder. The shells of the cationic nanoparticles and the hydrophobic core domain of the anionic central cylindrical nanostructure of the assemblies were utilized to complex negatively charged nucleic acids (siRNA) and to undergo radiolabeling, respectively, for potential theranostic applications. The assemblies exhibited exceptional cell transfection and radiolabeling efficiencies, providing an overall advantage over the individual components, which could each facilitate only one or the other of the functions.

Mechanism of cell death resulting from DNA interstrand cross-linking in mammalian cells

PubMed Central

Osawa, T; Davies, D; Hartley, J A

2011-01-01

DNA interstrand cross-links (ICLs) are critical cytotoxic lesions produced by cancer chemotherapeutic agents such as the nitrogen mustards and platinum drugs; however, the exact mechanism of ICL-induced cell death is unclear. Here, we show a novel mechanism of p53-independent apoptotic cell death involving prolonged cell-cycle (G2) arrest, ICL repair involving HR, transient mitosis, incomplete cytokinesis, and gross chromosomal abnormalities resulting from ICLs in mammalian cells. This characteristic ‘giant' cell death, observed by using time-lapse video microscopy, was reduced in ICL repair ERCC1- and XRCC3-deficient cells. Collectively, the results illustrate the coordination of ICL-induced cellular responses, including cell-cycle arrest, DNA damage repair, and cell death. PMID:21814285

Crosslinking hydroxylated reduced graphene oxide with RAFT-CTA: A nano-initiator for preparation of well-defined amino acid-based polymer nanohybrids.

PubMed

Namvari, Mina; Biswas, Chandra S; Wang, Qiao; Liang, Wenlang; Stadler, Florian J

2017-10-15

Here, we demonstrate a novel reversible addition-fragmentation chain transfer agent (RAFT-CTA)-modified reduced graphene oxide nanosheets (CTA-rGONSs) by crosslinking rGONSs with a RAFT-CTA via esterification reaction. These nano CTA-rGONSs were used to polymerize a hydrophobic amino acid-based methacrylamide (N-acryloyl-l-phenylalanine methyl ester) monomer with different monomer/initiator ratios. Thermogravimetric analysis showed that the polymer-graphene composites were thermally more stable than GO itself. M n of the polymers increased with increasing monomer/initiator ratio, while the polydispersity index decreased, indicating controlled polymerization. The composites were stable in DMF even after two months. Copyright © 2017 Elsevier Inc. All rights reserved.

Reconfigurable Polymer Networks for Improved Treatment of Intracranial Aneurysms

NASA Astrophysics Data System (ADS)

Ninh, Chi Suze Q.

Endovascular embolization of intracranial aneurysms is a minimally invasive treatment in which an implanted material forms a clot to isolate the weakened vessel. Current strategy suffers from long-term potential failure modes. These potential failure modes include (1) enzymatic degradation of the fibrin clot that leads to compaction of the embolic agent, (2) incomplete filling of the aneurysm sac by embolic agent, and (3) challenging geometry of wide neck aneurysms. In the case of wide neck aneurysms, usually an assisting metal stent is used to help open the artery. However, metal stents with much higher modulus in comparison to the soft blood vessel can cause biocompatibilities issues in the long term such as infection and scarring. Motivated to solve these challenges associated with endovascular embolization, strategies to synthesize and engineer reconfigurable and biodegradable polymers as alternative therapies are evaluated in this thesis. (1) Reconfiguration of fibrin gel's modulus was achieved through crosslinking with genipin released from a biodegradable polymer matrix. (2) Reconfigurability can also be achieved by transforming triblock co-polymer hydrogel into photoresponsive material through incorporation of melanin nanoparticles as efficient photosensitizers. (3) Finally, reconfigurability can be conferred on biodegradable polyester networks via Diels-Alder coupling of furan pendant groups and dimaleimide crosslinking agent. Taken all together, this thesis describes strategies to transform a broad class of polymer networks into reconfigurable materials for improved treatment of intracranial aneurysms as well as for other biomedical applications.

Electrochemical Sensor Based on Rh(III) Ion-Imprinted Polymer as a New Modifying Agent for Rhodium Determination.

PubMed

Bai, Huiping; Xiong, Caiyun; Wang, Chunqiong; Liu, Peng; Dong, Su; Cao, Qiue

2018-05-01

A rhodium (III) ion carbon paste electrode (CPE) based on an ion imprinted polymer (IIP) as a new modifying agent has been prepared and studied. Rh(III) ion imprinted polymer was synthesized by copolymerization of acrylamide-Rh(III) complex and ethylene glycol dimethacrylate according to the precipitation polymerization. Acrylamide acted as both functional monomer and complexing agent to create selective coordination sites in a cross-linked polymer. The ion imprinted carbon paste electrode (IIP-CPE) was prepared by mixing rhodium IIP-nanoparticles and graphite powder in n-eicosane as an adhesive and then embedding them in a Teflon tube. Amperometric i-t curve method was applied as the determination technique. Several parameters, including the functional monomer, molar ratio of template, monomer and cross-linking agent, the amounts of IIP, the applied potential, the buffer solution and pH have been studied. According to the results, IIP-CPE showed a considerably higher response in comparison with the electrode embedded with non-imprinted polymer (NIP), indicating the formation of suitable recognition sites in the IIP structure during the polymerization stage. The introduced electrode showed a linear range of 1.00×10-8~3.0×10-5 mol·L-1 and detection limit of 6.0 nmol L-1 (S/N = 3). The IIP-CPE was successfully applied for the trace rhodium determination in catalyst and plant samples with RSD of less than 3.3% (n = 5) and recoveries in the range of 95.5~102.5%.

Protein specific polymeric immunomicrospheres

NASA Technical Reports Server (NTRS)

Yen, Shiao-Ping S. (Inventor); Dreyer, William J. (Inventor); Rembaum, Alan (Inventor)

1980-01-01

Small, round, bio-compatible microspheres capable of covalently bonding proteins and having a uniform diameter below about 3500 A are prepared by substantially instantaneously initiating polymerization of an aqueous emulsion containing no more than 35% total monomer including an acrylic monomer substituted with a covalently bondable group such as hydroxyl, amino or carboxyl and a minor amount of a cross-linking agent.

Biotin-transfer from a trifunctional crosslinker for identification of cell surface receptors of soluble protein ligands

PubMed Central

Tremblay, Tammy-Lynn; Hill, Jennifer J.

2017-01-01

Here we describe a novel crosslinker and its application as a biotin-transfer reagent to identify cell surface receptors of soluble protein ligands on live cells. This crosslinker contains three functional groups: an aldehyde-reactive aminooxy group, a sulfhydryl, and a biotin (ASB). It is readily synthesized via a 3-step addition reaction using standard solid-phase peptide synthesis methods and commercially available intermediates, allowing access to laboratories without specialized synthetic chemistry capabilities. For the biotin-transfer method, ASB is linked to a protein ligand through the sulfhydryl group in a two-step process that allows the introduction of a disulfide bond between the ligand and the crosslinker. Incubation of the labelled ligand with oxidized live cells leads to the formation of crosslinks with aldehyde-containing glycans on the cell surface receptor. Subsequent reduction of the disulfide bond results in biotin transfer from the ligand to the cell surface receptor. Protein biotinylation that is mediated by ligand binding to its receptor is differentiated from background biotinylation events by comparison with a similarly labelled control protein using comparative proteomic mass spectrometry to quantify streptavidin-bound proteins. Using this method, we successfully identified the cell surface receptors of a peptide hormone, a monoclonal antibody, and a single-domain antibody-Fc fusion construct. PMID:28422167

Global Membrane Protein Interactome Analysis using In vivo Crosslinking and Mass Spectrometry-based Protein Correlation Profiling*

PubMed Central

Larance, Mark; Kirkwood, Kathryn J.; Tinti, Michele; Brenes Murillo, Alejandro; Ferguson, Michael A. J.; Lamond, Angus I.

2016-01-01

We present a methodology using in vivo crosslinking combined with HPLC-MS for the global analysis of endogenous protein complexes by protein correlation profiling. Formaldehyde crosslinked protein complexes were extracted with high yield using denaturing buffers that maintained complex solubility during chromatographic separation. We show this efficiently detects both integral membrane and membrane-associated protein complexes,in addition to soluble complexes, allowing identification and analysis of complexes not accessible in native extracts. We compare the protein complexes detected by HPLC-MS protein correlation profiling in both native and formaldehyde crosslinked U2OS cell extracts. These proteome-wide data sets of both in vivo crosslinked and native protein complexes from U2OS cells are freely available via a searchable online database (www.peptracker.com/epd). Raw data are also available via ProteomeXchange (identifier PXD003754). PMID:27114452

A Crosslinker Based on a Tethered Electrophile for Mapping Kinase-Substrate Networks

PubMed Central

Riel-Mehan, Megan M; Shokat, Kevan M

2014-01-01

SUMMARY Despite the continuing progress made towards mapping kinase signaling networks, there are still many phosphorylation events for which the responsible kinase has not yet been identified. We are interested in addressing this problem through forming covalent crosslinks between a peptide substrate and the corresponding phosphorylating kinase. Previously we reported a dialdehyde-based kinase binding probe capable of such a reaction with a peptide containing a cysteine substituted for the phosphorylatable ser/thr/tyr residue. Here, we examine the yield of a previously reported dialdehyde-based probe, and report that the dialdehyde based probes possesses a significant limitation in terms of crosslinked kinase-substrate product yield. To address this limitation, we develop a crosslinking scheme based on a kinase activity-based probe, and this new cross-linker provides an increase in efficiency and substrate specificity, including in the context of cell lysate. PMID:24746561

Bionic multisilicon copolymers used as novel cross-linking agents for preparing anion exchange hybrid membranes.

PubMed

Wu, Yonghui; Luo, Jingyi; Wu, Cuiming; Xu, Tongwen; Fu, Yanxun

2011-05-26

Bionic multisilicon copolymers have long-main chains and many branched chains, and contain multifunctional groups of -N(+)(CH(3))(3)Cl(-) and -Si(OCH(3))(3), which are similar to the stem, branch, fruit, and acetabula of a vine from bionic aspect, respectively. They have high flexibility, charge density, and cross-linking ability and thus can be used as novel cross-linking agents for preparing anion-exchange hybrid membranes. High content of -Si(OCH(3))(3) groups (68-78%) is suitable to enhance membrane stabilities. The membranes are stable in 65 °C water up to 120 h and can keep integrity in 2 mol/L NaOH for 192 h. High content of -N(+)(CH(3))(3)Cl(-) groups (42-55%) is suitable to enhance membrane electrical properties. The membranes have low membrane resistance (R(m), 0.59-0.94 Ω cm(2)) and high diffusion dialysis performance. The acid (H(+)) dialysis coefficients (U(H)) are in the range of 0.007-0.075 m h(-1) at room temperature and 0.015-0.115 m h(-1) at 40 °C. The separation factor (S(H/Fe)) can reach up to 43 at room temerature and 49 at 40 °C. All of the membranes are highly homogeneous, mechanically stable (21-31 MPa, 25-147%), and thermally stable (227-275 °C for halide form membranes, and 157-172 °C for OH(-) form membranes). Hence, the investigation of multisilicon copolymers will give rise to a new developing field in material and membrane sciences.

Chemical crosslinking and mass spectrometry studies of the structure and dynamics of membrane proteins and receptors.

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

Haskins, William E.; Leavell, Michael D.; Lane, Pamela

2005-03-01

Membrane proteins make up a diverse and important subset of proteins for which structural information is limited. In this study, chemical cross-linking and mass spectrometry were used to explore the structure of the G-protein-coupled photoreceptor bovine rhodopsin in the dark-state conformation. All experiments were performed in rod outer segment membranes using amino acid 'handles' in the native protein sequence and thus minimizing perturbations to the native protein structure. Cysteine and lysine residues were covalently cross-linked using commercially available reagents with a range of linker arm lengths. Following chemical digestion of cross-linked protein, cross-linked peptides were identified by accurate mass measurementmore » using liquid chromatography-fourier transform mass spectrometry and an automated data analysis pipeline. Assignments were confirmed and, if necessary, resolved, by tandem MS. The relative reactivity of lysine residues participating in cross-links was evaluated by labeling with NHS-esters. A distinct pattern of cross-link formation within the C-terminal domain, and between loop I and the C-terminal domain, emerged. Theoretical distances based on cross-linking were compared to inter-atomic distances determined from the energy-minimized X-ray crystal structure and Monte Carlo conformational search procedures. In general, the observed cross-links can be explained by re-positioning participating side-chains without significantly altering backbone structure. One exception, between C3 16 and K325, requires backbone motion to bring the reactive atoms into sufficient proximity for cross-linking. Evidence from other studies suggests that residues around K325 for a region of high backbone mobility. These findings show that cross-linking studies can provide insight into the structural dynamics of membrane proteins in their native environment.« less

Cross-linking of initiation factor IF3 to Escherichia coli 30S ribosomal subunit by trans-diamminedichloroplatinum(II): characterization of two cross-linking sites in 16S rRNA; a possible way of functioning for IF3.

PubMed Central

Ehresmann, C; Moine, H; Mougel, M; Dondon, J; Grunberg-Manago, M; Ebel, J P; Ehresmann, B

1986-01-01

The initiation factor IF3 is platinated with trans-diamminedichloroplatinum(II) and cross-linked to Escherichia coli 30S ribosomal subunit. Two cross-linking sites are unambiguously identified on the 16S rRNA: a major one, in the region 819-859 in the central domain, and a minor one, in the region 1506-1529 in the 3'-terminal domain. Specific features of these sequences together with their particular location within the 30S subunit lead us to postulate a role for IF3, that conciliates topographical and functional observations made so far. Images PMID:2425339

Optimization of biodegradable sponges as controlled release drug matrices. I. Effect of moisture level on chitosan sponge mechanical properties.

PubMed

Foda, Nagwa H; El-laithy, Hanan M; Tadros, Mina I

2004-04-01

Cross-linked chitosan sponges as controlled release drug carrier systems were developed. Tramadol hydrochloride, a centrally acting analgesic, was used as a model drug. The sponges were prepared by freeze-drying 1.25% and 2.5% (w/w) high and low M.wt. chitosan solutions, respectively, using glutaraldehyde as a cross-linking agent. The hardness of the prepared sponges was a function of glutaraldehyde concentration and volume where the optimum concentration that offered accepted sponge consistency was 5%. Below or above 5%, very soft or very hard and brittle sponges were obtained, respectively. The determined drug content in the prepared sponges was uniform and did not deviate markedly from the calculated amount. Scanning electron microscopy (SEM) was used to characterize the internal structures of the sponges. The SEM photos revealed that cross-linked high M.wt. chitosan sponges have larger size surface pores that form connections (channels) with the interior of the sponge than cross-linked low M.wt. ones. Moreover, crystals of the incorporated Tramadol hydrochloride were detected on the lamellae and within pores in both chitosan sponges. Differences in pore size and dissolution medium uptake capacity were crucial factors for the more delayed drug release from cross-linked low M.wt. chitosan sponges over high M.wt. ones at pH 7.4. Kinetic analysis of the release data using linear regression followed the Higuchi diffusion model over 12 hours. Setting storage conditions at room temperature under 80-92% relative humidity resulted in soft, elastic, and compressible sponges.

Immobilization of a phosphonated analog of matrix phosphoproteins within cross-linked collagen as a templating mechanism for biomimetic mineralization

PubMed Central

Gu, Li-sha; Kim, Young Kyung; Liu, Yan; Takahashi, Kei; Arun, Senthil; Wimmer, Courtney E.; Osorio, Raquel; Ling, Jun-qi; Looney, Stephen W.; Pashley, David H.; Tay, Franklin R.

2010-01-01

Immobilization of phosphoproteins on a collagen matrix is important for induction of intrafibrillar apatite mineralization. Unlike phosphate esters, polyphosphonic acid has no reactive sites for covalent binding to collagen amine groups. Binding of polyvinylphosphonic acid (PVPA), a biomimetic templating analog of matrix phosphoproteins, to collagen was found to be electrostatic in nature. Thus, an alternative retention mechanism was designed for immobilization of PVPA to collagen by cross-linking the latter with carbodiimide (EDC). This mechanism is based on the principle of size exclusion entrapment of PVPA molecules within the internal water compartments of collagen. By cross-linking collagen with EDC, a zero-length cross-linking agent, the sieving property of collagen is increased, enabling the PVPA to be immobilized within the collagen. Absence of covalent cross-linking between PVPA and collagen was confirmed by FT-IR spectroscopy. Based on these results, a concentration range for immobilized PVPA to template intrafibrillar apatite deposition was established and validated using a single-layer reconstituted type I collagen mineralization model. In the presence of a polyacrylic acid-containing mineralization medium, optimal intrafibrillar mineralization of the EDC-cross-linked collagen was achieved using 500 and 1,000 μg/mL PVPA. The mineralized fibrils exhibited a hierarchical order of intrafibrillar mineral infiltration, as manifested by the appearance of electron-dense periodicity within unstained fibrils. Understanding the basic processes in intrafibrillar mineralization of reconstituted collagen creates opportunities for the design of tissue engineering materials for hard tissue repair and regeneration. PMID:20688200

Isozyme-specific comprehensive characterization of transglutaminase-crosslinked substrates in kidney fibrosis.

PubMed

Tatsukawa, Hideki; Otsu, Risa; Tani, Yuji; Wakita, Ryosuke; Hitomi, Kiyotaka

2018-05-09

Chronic kidney disease is characterized by prolonged decline in renal function, excessive accumulation of ECM, and progressive tissue fibrosis. Transglutaminase (TG) is a crosslinking enzyme that catalyzes the formation of covalent bonds between glutamine and lysine residues, and is involved in the induction of renal fibrosis via the stabilization of ECM and the activation of TGF-β1. Despite the accumulating evidences indicating that TG2 is a key enzyme in fibrosis, genetic knockout of TG2 reduced by only 50% the elevated protein crosslinking and fibrous protein in renal fibrosis model, whereas treatment with TG inhibitor almost completely reduced these levels. Here, we also clarified the distributions of TG isozymes and their in situ activities and identified the isozyme-specific crosslinked substrates for both TG1 and TG2 in fibrotic kidney. We found that TG1 activity was markedly enhanced in renal tubular epithelium and interstitial areas, whereas TG2 activity increased only in the extracellular space. In total, 47 and 67 possible candidates were identified as TG1 and TG2 substrates, respectively, only in fibrotic kidney. Among them, several possible substrates related to renal disease and fibrosis were identified. These findings provide novel insights into the mechanisms of renal fibrosis through the targeting of isozyme-specific TG substrates.

CLMSVault: A Software Suite for Protein Cross-Linking Mass-Spectrometry Data Analysis and Visualization.

PubMed

Courcelles, Mathieu; Coulombe-Huntington, Jasmin; Cossette, Émilie; Gingras, Anne-Claude; Thibault, Pierre; Tyers, Mike

2017-07-07

Protein cross-linking mass spectrometry (CL-MS) enables the sensitive detection of protein interactions and the inference of protein complex topology. The detection of chemical cross-links between protein residues can identify intra- and interprotein contact sites or provide physical constraints for molecular modeling of protein structure. Recent innovations in cross-linker design, sample preparation, mass spectrometry, and software tools have significantly improved CL-MS approaches. Although a number of algorithms now exist for the identification of cross-linked peptides from mass spectral data, a dearth of user-friendly analysis tools represent a practical bottleneck to the broad adoption of the approach. To facilitate the analysis of CL-MS data, we developed CLMSVault, a software suite designed to leverage existing CL-MS algorithms and provide intuitive and flexible tools for cross-platform data interpretation. CLMSVault stores and combines complementary information obtained from different cross-linkers and search algorithms. CLMSVault provides filtering, comparison, and visualization tools to support CL-MS analyses and includes a workflow for label-free quantification of cross-linked peptides. An embedded 3D viewer enables the visualization of quantitative data and the mapping of cross-linked sites onto PDB structural models. We demonstrate the application of CLMSVault for the analysis of a noncovalent Cdc34-ubiquitin protein complex cross-linked under different conditions. CLMSVault is open-source software (available at https://gitlab.com/courcelm/clmsvault.git ), and a live demo is available at http://democlmsvault.tyerslab.com/ .

New Elastomeric Materials Based on Natural Rubber Obtained by Electron Beam Irradiation for Food and Pharmaceutical Use

PubMed Central

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

2016-01-01

The efficiency of polyfunctional monomers as cross-linking co-agents on the chemical properties of natural rubber vulcanized by electron beam irradiation was studied. The following polyfunctional monomers were used: trimethylolpropane-trimethacrylate, zinc-diacrylate, ethylene glycol dimethacrylate, triallylcyanurate and triallylisocyanurate. The electron beam treatment was done using irradiation doses in the range of 75 kGy–300 kGy. The gel fraction, crosslink density and effects of different aqueous solutions, by absorption tests, have been investigated as a function of polyfunctional monomers type and absorbed dose. The samples gel fraction and crosslink density were determined on the basis of equilibrium solvent-swelling measurements by applying the modified Flory–Rehner equation for tetra functional networks. The absorption tests were done in accordance with the SR ISI 1817:2015 using distilled water, acetic acid (10%), sodium hydroxide (1%), ethylic alcohol (96%), physiological serum (sodium chloride 0.9%) and glucose (glucose monohydrate 10%). The samples structure and morphology were investigated by Fourier Transform Infrared Spectroscopy and Scanning Electron Microscopy techniques. PMID:28774150

Recent Developments in Hardware-in-the-Loop Formation Navigation and Control

NASA Technical Reports Server (NTRS)

Mitchell, Jason W.; Luquette, Richard J.

2005-01-01

The Formation Flying Test-Bed (FFTB) at NASA Goddard Space Flight Center (GSFC) provides a hardware-in-the-loop test environment for formation navigation and control. The facility is evolving as a modular, hybrid, dynamic simulation facility for end-tc-end guidance, navigation, and control (GN&C) design and analysis of formation flying spacecraft. The core capabilities of the FFTB, as a platform for testing critical hardware and software algorithms in-the-loop, are reviewed with a focus on many recent improvements. Two significant upgrades to the FFTB are a message-oriented middleware (MOM) architecture, and a software crosslink for inter-spacecraft ranging. The MOM architecture provides a common messaging bus for software agents, easing integration, arid supporting the GSFC Mission Services Evolution Center (GMSEC) architecture via software bridge. Additionally, the FFTB s hardware capabilities are expanding. Recently, two Low-Power Transceivers (LPTs) with ranging capability have been introduced into the FFTB. The LPT crosslinks will be connected to a modified Crosslink Channel Simulator (CCS), which applies realistic space-environment effects to the Radio Frequency (RF) signals produced by the LPTs.

Inactivation of human DGAT2 by oxidative stress on cysteine residues

PubMed Central

Choi, Kwangman; Kwon, Eun Bin; Kang, Mingu; Kim, Dong-eun; Jeong, Hyejeong; Kim, Janghwan; Kim, Jong Heon; Kim, Mun Ock; Han, Sang-Bae

2017-01-01

Diacylglycerol acyltransferases (DGATs) have a crucial role in the biosynthesis of triacylglycerol (TG), the major storage form of metabolic energy in eukaryotic organisms. Even though DGAT2, one of two distinct DGATs, has a vital role in TG biosynthesis, little is known about the regulation of DGAT2 activity. In this study, we examined the role of cysteine and its oxidation in the enzymatic activity of human DGAT2 in vitro. Human DGAT2 activity was considerably inhibited not only by thiol-modifying reagents (NEM and IA) but also by ROS-related chemicals (H2O2 and β-lapachone), while human DGAT1 and GPAT1 were little affected. Particularly, ROS-related chemicals concomitantly induced intermolecular disulfide crosslinking of human DGAT2. Both the oxidative inactivation and disulfide crosslinking were almost completely reversed by the treatment with DTT, a disulfide-reducing agent. These results clearly demonstrated the significant role of ROS-induced intermolecular crosslinking in the inactivation of human DGAT2 and also suggested DGAT2 as a redox-sensitive regulator in TG biosynthesis. PMID:28700690

New Elastomeric Materials Based on Natural Rubber Obtained by Electron Beam Irradiation for Food and Pharmaceutical Use.

PubMed

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

2016-12-21

The efficiency of polyfunctional monomers as cross-linking co-agents on the chemical properties of natural rubber vulcanized by electron beam irradiation was studied. The following polyfunctional monomers were used: trimethylolpropane-trimethacrylate, zinc-diacrylate, ethylene glycol dimethacrylate, triallylcyanurate and triallylisocyanurate. The electron beam treatment was done using irradiation doses in the range of 75 kGy-300 kGy. The gel fraction, crosslink density and effects of different aqueous solutions, by absorption tests, have been investigated as a function of polyfunctional monomers type and absorbed dose. The samples gel fraction and crosslink density were determined on the basis of equilibrium solvent-swelling measurements by applying the modified Flory-Rehner equation for tetra functional networks. The absorption tests were done in accordance with the SR ISI 1817:2015 using distilled water, acetic acid (10%), sodium hydroxide (1%), ethylic alcohol (96%), physiological serum (sodium chloride 0.9%) and glucose (glucose monohydrate 10%). The samples structure and morphology were investigated by Fourier Transform Infrared Spectroscopy and Scanning Electron Microscopy techniques.

A novel fish collagen scaffold as dural substitute.

PubMed

Li, Qing; Mu, Lanlan; Zhang, Fenghua; Sun, Yue; Chen, Quan; Xie, Cuicui; Wang, Hongmei

2017-11-01

The novel fish collagen scaffolds were prepared by lyophilization. The collagen sponges and chitosan were chemically cross-linked with the 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) as a cross-linking agent by pressing in one special mould. The collagen scaffolds were analyzed by scanning electron microscopy (SEM) and mechanical property, and the in vitro collagenase degradation was tested. The results revealed that the scaffold has a suitable porosity, elasticity and prevent fluid leakage, suggesting potential applications in the tissue-engineered. In vitro collagenase degradation demonstrated that the collagen cross-linking with EDC by pressing played an important role in their resistance to biodegradation. Moreover, the scaffold proved excellent biocompatibility for the activity and proliferation of mouse embryonic fibroblasts cells (MEFs) in vitro. The rabbit dural defect model demonstrated that the scaffolds could prevent brain tissue adhesion, which reduce the opportunity of inflammation, facilitate the growth of fibroblasts and enhance the tissue regeneration and healing. The novel fish collagen scaffold as dural substitute, demonstrate a capability for using in the field of tissue engineering. Copyright © 2017. Published by Elsevier B.V.

The Aggregation Paths and Products of Aβ42 Dimers Are Distinct from Those of the Aβ42 Monomer.

PubMed

O'Malley, Tiernan T; Witbold, William M; Linse, Sara; Walsh, Dominic M

2016-11-08

Extracts of Alzheimer's disease (AD) brain that contain what appear to be sodium dodecyl sulfate-stable amyloid β-protein (Aβ) dimers potently block LTP and impair memory consolidation. Brain-derived dimers can be physically separated the Aβ monomer, consist primarily of Aβ42, and resist denaturation by chaotropic agents. In nature, covalently cross-linked Aβ dimers could be generated in two ways: by the formation of a dityrosine (DiY) or an isopeptide ε-(γ-glutamyl)-lysine (Q-K) bond. We enzymatically cross-linked recombinant Aβ42 monomer to produce DiY and Q-K dimers and then used a range of biophysical methods to study their aggregation. Both Q-K and DiY dimers aggregate to form soluble assemblies distinct from the fibrillar aggregates formed by the Aβ monomer. The results suggest that the cross-links disfavor fibril formation from Aβ dimers, thereby enhancing the concentration of soluble aggregates akin to those in aqueous extracts of AD brain. Thus, it seems that Aβ dimers may play an important role in determining the formation of soluble rather than insoluble aggregates.

Stimuli sensitive super-macroporous cryogels based on photo-crosslinked 2-hydroxyethylcellulose and chitosan.

PubMed

Stoyneva, Veselina; Momekova, Denitsa; Kostova, Bistra; Petrov, Petar

2014-01-01

Original pH sensitive cryogels, based on two biodegradable natural polymers chitosan (CS) and 2-hydroxyethylcellulose (HEC), were obtained via cryogenic treatment of semi-dilute aqueous solutions and UV induced crosslinking in frozen state. H₂O₂ and N,N'-methylenebisacrylamide (BisAAm) were used as photoinitiator and crosslinking agent, respectively. BisAAm facilitated the formation of polymer co-network and increased both the gel fraction yield and mechanical strength of cryogels. The influence of chitosan content on the physico-mechanical properties of HEC-CS cryogels was investigated. In general, the increase of CS fraction in the polymer co-network increased the degree of swelling and enhanced significantly the storage modulus of materials. All HEC-CS cryogels obtained were opalescent sponge-like materials, which quickly release/uptake water due to their open porous structure. The incorporation of CS provided pH dependent swelling and good bioadhesive properties of cryogels. HEC-CS cryogels were further exploited as drug delivery systems of the highly water soluble drug metronidazole belonging to BCS Class l. Copyright © 2013 Elsevier Ltd. All rights reserved.

One-pot synthesis of redox-responsive polymers-coated mesoporous silica nanoparticles and their controlled drug release.

PubMed

Sun, Jiao-Tong; Piao, Ji-Gang; Wang, Long-Hai; Javed, Mohsin; Hong, Chun-Yan; Pan, Cai-Yuan

2013-09-01

A versatile one-pot strategy for the preparation of reversibly cross-linked polymer-coated mesoporous silica nanoparticles (MSNs) via surface reversible addition-fragmentation chain transfer (RAFT) polymerization is presented for the first time in this paper. The less reactive monomer oligo(ethylene glycol) acrylate (OEGA) and the more reactive cross-linker N,N'-cystaminebismethacrylamide (CBMA) are chosen to be copolymerized on the external surfaces of RAFT agent-functionalized MSNs to form the cross-linked polymer shells. Owing to the reversible cleavage and restoration of disulfide bonds via reduction/oxidation reactions, the polymer shells can control the on/off switching of the nanopores and regulate the drug loading and release. The redox-responsive release of doxorubicin (DOX) from this drug carrier is realized. The protein adsorption, in vitro cytotoxicity assays, and endocytosis studies demonstrate that this biocompatible vehicle is a potential candidate for delivering drugs. It is expected that this versatile grafting strategy may help fabricate satisfying MSN-based drug delivery systems for clinical application. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Immobilization of pectinase onto chitosan magnetic nanoparticles by macromolecular cross-linker.

PubMed

Sojitra, Uttam V; Nadar, Shamraja S; Rathod, Virendra K

2017-02-10

Pectinase was immobilized onto chitosan magnetic nanoparticles (CMNPs) by dextran polyaldehyde as a macromolecular cross-linking agent. The parameters like cross-linking concentration, time and CMNPs to enzyme ratio were optimized. Further, prepared magnetic pectinase nanobiocatalyst was characterized by FT-IR and XRD. The thermal kinetic studies for immobilized pectinase showed two folds improved thermal stability in the range of 55-75°C as compared to free form. The V max and K m values of immobilized pectinase were found to be nearly equal to native form which indicated that conformational flexibility of pectinase was retained even after immobilization. The residual activity of immobilized pectinase was 85% after seven successive cycles of reuse, while it retained upto 89% residual activity on storage of fifteen days which exhibited excellent stability and durability. The conformational changes in pectinase after immobilization were evaluated by FT-IR spectroscopy data analysis tools. Finally, magnetic pectinase nanobiocatalyst was employed for apple juice clarification which showed turbidity reduction upto 74% after 150min treatment. Copyright © 2016 Elsevier Ltd. All rights reserved.

Meniscal material properties are minimally affected by matrix stabilization using glutaraldehyde and glycation with ribose.

PubMed

Hunter, Shawn A; Noyes, Frank R; Haridas, Balakrishna; Levy, Martin S; Butler, David L

2005-05-01

Knee meniscus replacement holds promise, but current allografts are susceptible to biodegradation. Matrix stabilization with glutaraldehyde, a crosslinking agent used clinically to fabricate cardiovascular bioprostheses, or with glycation, a process of crosslinking collagen with sugars such as ribose, is a potential means of rendering tissue resistant to such degradation. However, stabilization should not significantly alter meniscal material properties, which could disturb normal function in the knee. Our objective was to evaluate the effects of glutaraldehyde- and glycation-induced matrix stabilization on the material properties of porcine meniscus. Normal untreated meniscus specimens were tested in confined compression at one of three applied stresses (0.069, 0.208, 0.347 MPa), subjected to either a glutaraldehyde or glycation stabilization treatment, and then re-tested to measure changes in tissue aggregate modulus, permeability, and compressive strain at equilibrium. Changes in these properties significantly increased with glutaraldehyde concentration and exposure time to ribose. One glutaraldehyde and three glycation treatments did not alter aggregate modulus or compressive strain at equilibrium compared to controls (p > 0.10). However, all treatments increased permeability by at least 108% compared to controls (p < 0.001). This study reveals a dose-dependent relationship between meniscal material properties and certain stabilization conditions and identifies treatments that minimally affect these properties. Further research is necessary to determine whether these treatments prevent enzymatic degradation before and after surgical implantation in the knee.

Optimization of protein cross-linking in bicomponent electrospun scaffolds for therapeutic use

NASA Astrophysics Data System (ADS)

Papa, Antonio; Guarino, Vincenzo; Cirillo, Valentina; Oliviero, Olimpia; Ambrosio, Luigi

2015-12-01

Bio-instructive electrospun scaffolds based on the combination of synthetic polymers, such as PCL or PLLA, and natural polymers (e.g., collagen) have been extensively investigated as temporary extracellular matrix (ECM) analogues able to support cell proliferation and stem cell differentiation for the regeneration of several tissues. The growing use of natural polymers as carrier of bioactive molecules is introducing new ideas for the design of polymeric drug delivery systems based on electrospun fibers with improved bioavailability, therapeutic efficacy and programmed drug release. In particular, the release mechanism is driven by the use of water soluble proteins (i.e., collagen, gelatin) which fully degrade in in vitro microenvironment, thus delivering the active principles. However, these protein are generally rapidly digested by enzymes (i.e., collagenase) produced by many different cell types, both in vivo and in vitro with significant drawbacks in tissue engineering and controlled drug delivery. Here, we aim at investigating different chemical strategies to improve the in vitro stability and mechanical strength of scaffolds against enzymatic degradation, by modifying the biodegradation rates of proteins embedded in bicomponent fibers. By comparing scaffolds treated by different cross-linking agents (i.e., GC, EDC, BDDGE), we have provided an extensive morphological/chemical/physical characterization via SEM and TGA to identify the best conditions to control drug release via protein degradation from bicomponent fibers without compromising in vitro cell response.

Optimization of protein cross-linking in bicomponent electrospun scaffolds for therapeutic use

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

Papa, Antonio; IMAST SCaRL, Piazza Bovio 22, 80133 Naples; Guarino, Vincenzo, E-mail: vincenzo.guarino@cnr.it

Bio-instructive electrospun scaffolds based on the combination of synthetic polymers, such as PCL or PLLA, and natural polymers (e.g., collagen) have been extensively investigated as temporary extracellular matrix (ECM) analogues able to support cell proliferation and stem cell differentiation for the regeneration of several tissues. The growing use of natural polymers as carrier of bioactive molecules is introducing new ideas for the design of polymeric drug delivery systems based on electrospun fibers with improved bioavailability, therapeutic efficacy and programmed drug release. In particular, the release mechanism is driven by the use of water soluble proteins (i.e., collagen, gelatin) which fullymore » degrade in in vitro microenvironment, thus delivering the active principles. However, these protein are generally rapidly digested by enzymes (i.e., collagenase) produced by many different cell types, both in vivo and in vitro with significant drawbacks in tissue engineering and controlled drug delivery. Here, we aim at investigating different chemical strategies to improve the in vitro stability and mechanical strength of scaffolds against enzymatic degradation, by modifying the biodegradation rates of proteins embedded in bicomponent fibers. By comparing scaffolds treated by different cross-linking agents (i.e., GC, EDC, BDDGE), we have provided an extensive morphological/chemical/physical characterization via SEM and TGA to identify the best conditions to control drug release via protein degradation from bicomponent fibers without compromising in vitro cell response.« less

Isotope-labeled cross-linkers and Fourier transform ion cyclotron resonance mass spectrometry for structural analysis of a protein/peptide complex.

PubMed

Ihling, Christian; Schmidt, Andreas; Kalkhof, Stefan; Schulz, Daniela M; Stingl, Christoph; Mechtler, Karl; Haack, Michael; Beck-Sickinger, Annette G; Cooper, Dermot M F; Sinz, Andrea

2006-08-01

For structural studies of proteins and their complexes, chemical cross-linking combined with mass spectrometry presents a promising strategy to obtain structural data of protein interfaces from low quantities of proteins within a short time. We explore the use of isotope-labeled cross-linkers in combination with Fourier transform ion cyclotron resonance (FTICR) mass spectrometry for a more efficient identification of cross-linker containing species. For our studies, we chose the calcium-independent complex between calmodulin and a 25-amino acid peptide from the C-terminal region of adenylyl cyclase 8 containing an "IQ-like motif." Cross-linking reactions between calmodulin and the peptide were performed in the absence of calcium using the amine-reactive, isotope-labeled (d0 and d4) cross-linkers BS3 (bis[sulfosuccinimidyl]suberate) and BS2G (bis[sulfosuccinimidyl]glutarate). Tryptic in-gel digestion of excised gel bands from covalently cross-linked complexes resulted in complicated peptide mixtures, which were analyzed by nano-HPLC/nano-ESI-FTICR mass spectrometry. In cases where more than one reactive functional group, e.g., amine groups of lysine residues, is present in a sequence stretch, MS/MS analysis is a prerequisite for unambiguously identifying the modified residues. MS/MS experiments revealed two lysine residues in the central alpha-helix of calmodulin as well as three lysine residues both in the C-terminal and N-terminal lobes of calmodulin to be cross-linked with one single lysine residue of the adenylyl cyclase 8 peptide. Further cross-linking studies will have to be conducted to propose a structural model for the calmodulin/peptide complex, which is formed in the absence of calcium. The combination of using isotope-labeled cross-linkers, determining the accurate mass of intact cross-linked products, and verifying the amino acid sequences of cross-linked species by MS/MS presents a convenient approach that offers the perspective to obtain structural data of protein assemblies within a few days.

Preparation of small bio-compatible microspheres

NASA Technical Reports Server (NTRS)

Rembaum, Alan (Inventor); Yen, Shiao-Ping S. (Inventor); Dreyer, William J. (Inventor)

1979-01-01

Small, round, bio-compatible microspheres capable of covalently bonding proteins and having a uniform diameter below about 3500 A are prepared by substantially instantaneously initiating polymerization of an aqueous emulsion containing no more than 35% total monomer including an acrylic monomer substituted with a covalently bondable group such a hydroxyl, amino or carboxyl and a minor amount of a cross-linking agent.

Smart Polymer Surfaces

DTIC Science & Technology

2009-02-27

films: Inhibition of dewetting in thin polymer films”, Carroll, Gregory T., Sojka, Melissa E., Lei, Xuegong, Turro, Nicholas J., Koberstein, Jeffrey T...at Sandia was that the polymer films, designed to have specific interactions with particular warfare agents, would dewet the surface of a surface...crosslinking or dewetting . Patterned dewetting constitutes a completely new way of generating micro thin film structures that might be useful in

21 CFR 176.180 - Components of paper and paperboard in contact with dry food.

Code of Federal Regulations, 2014 CFR

2014-04-01

.... Tetraethylenepentamine Polymerization cross-linking agent. α-[p-(1,1,3,3-Tetramethylbutyl)phenyl]-omega hydroxypoly... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Components of paper and paperboard in contact with... preservative in coatings and sizings. 1,2-Benzisothiazolin-3-one (CAS Registry No. 2634-33-5) For use only as a...

Detection of DNA damage induced in vivo by a cross-linking agent with a circular channel crucible oscillating viscometer.

PubMed

Balbi, C; Abelmoschi, M L; Roner, R; Giaretti, W; Parodi, S; Santi, L

1985-11-01

DNA damage induced in vivo by the cross-linking agent mitomycin C (MMC) was investigated with a new oscillating crucible viscometer. Viscosity was measured by lysing rat liver nuclei in an alkaline lysing solution (pH 12.5; 25 degrees C). In control samples the viscosity increased very slowly with time, reaching a plateau only after 10-12 h. The process was accelerated and the maximum viscosity was decreased by alkaline single-stranded breaks arising from methylation and subsequent depurination of DNA in vitro with dimethylsulphate (DMS). MMC, when given alone, had no evident effect on the time needed for reaching plateau viscosity but it induced a small increase in maximum viscosity. When MMC was given in association with DMS, the time of disentanglement remained unchanged (accelerated) but maximum viscosity was increased in a dose dependent way. We conclude that these data clearly confirm that the slow steady increase of the viscosity of control DNA with time reflects mainly the process of unwinding of the two strands. The speed of this process seems to depend only from the number of unwinding points in DNA (breaks).

Phosphorylation of psyllium seed polysaccharide and its characterization.

PubMed

Rao, Monica R P; Warrier, Deepa U; Gaikwad, Snehal R; Shevate, Prachi M

2016-04-01

Psyllium is widely used as a medicinally active natural polysaccharide for treating conditions like constipation, diarrhea, and irritable bowel syndrome, inflammatory bowel disease, ulcerative colitis and colon cancer. Studies have been performed to characterize and modify the polysaccharide obtained from psyllium seed husk and to evaluate its use as a pharmaceutical excipient, but no studies have been performed to evaluate the properties of the polysaccharide present in psyllium seeds. The present study focuses on phosphorylation of psyllium seed polysaccharide (PPS) using sodium tri-meta phosphate as the cross-linking agent. The modified phosphorylated psyllium seed polysaccharide was then evaluated for physicochemical properties, rheological properties, spectral analysis, thermal analysis, crosslinking density and acute oral toxicity studies. The modified polysaccharide (PhPPS) has a high swelling index due to which it can be categorized as a hydrogel. The percent increase in swelling of PhPPS as compared to PPS was found to be 90.26%. The PPS & PhPPS mucilages of all strengths were found to have shear thinning properties. These findings are suggestive of the potential use of PhPPS as gelling & suspending agent. PhPPS was found to have a mucoadhesive property which was comparable with carbopol. Copyright © 2015 Elsevier B.V. All rights reserved.

Altered Cross-Linking of HSP27 by Zerumbone as a Novel Strategy for Overcoming HSP27-Mediated Radioresistance

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

Choi, Seo-Hyun; School of Life Sciences and Biotechnology, Korea University, Seoul; Lee, Yoon-Jin

Purpose: HSP27 or HSP25 negatively regulates apoptosis pathways after radiation or chemotherapeutic agents. Abrogation of HSP27 function may be a candidate target for overcoming radio- and chemoresistance. Methods and Materials: Zerumbone (ZER), a cytotoxic component isolated from Zingiber zerumbet smith. Clonogenic survival assay and flow cytometry after Annexin V staining were performed to determine in vitro sensitization effects of ZER with ionizing radiation. A nude mouse xenografting system was also applied to detect in vivo radiosensitizing effects of ZER. Results: ZER produced cross-linking of HSP27, which was dependent on inhibition of the monomeric form of HSP27. ZER was directly insertedmore » between the disulfide bond in the HSP27 dimer and modified normal HSP27 dimerization. Pretreatment with ZER before radiation inhibited the binding affinity between HSP27 and apoptotic molecules, such as cytochrome c and PKC{delta}, and induced sensitization in vitro and in an in vivo xenografted nude mouse system. Structural analogs lacking only the carbonyl group in ZER, such as {alpha}-humulene (HUM) and 8-hydroxy-humulen (8-OH-HUM), did not affect normal cross-linking of HSP27 and did not induce radiosensitization. Conclusions: We suggest that altered cross-linking of HSP27 by ZER is a good strategy for abolishing HSP27-mediated resistance.« less

Altered cross-linking of HSP27 by zerumbone as a novel strategy for overcoming HSP27-mediated radioresistance.

PubMed

Choi, Seo-Hyun; Lee, Yoon-Jin; Seo, Woo Duck; Lee, Hae-June; Nam, Joo-Won; Lee, Yoo Jin; Kim, Joon; Seo, Eun-Kyoung; Lee, Yun-Sil

2011-03-15

HSP27 or HSP25 negatively regulates apoptosis pathways after radiation or chemotherapeutic agents. Abrogation of HSP27 function may be a candidate target for overcoming radio- and chemoresistance. Zerumbone (ZER), a cytotoxic component isolated from Zingiber zerumbet smith. Clonogenic survival assay and flow cytometry after Annexin V staining were performed to determine in vitro sensitization effects of ZER with ionizing radiation. A nude mouse xenografting system was also applied to detect in vivo radiosensitizing effects of ZER. ZER produced cross-linking of HSP27, which was dependent on inhibition of the monomeric form of HSP27. ZER was directly inserted between the disulfide bond in the HSP27 dimer and modified normal HSP27 dimerization. Pretreatment with ZER before radiation inhibited the binding affinity between HSP27 and apoptotic molecules, such as cytochrome c and PKCδ, and induced sensitization in vitro and in an in vivo xenografted nude mouse system. Structural analogs lacking only the carbonyl group in ZER, such as α-humulene (HUM) and 8-hydroxy-humulen (8-OH-HUM), did not affect normal cross-linking of HSP27 and did not induce radiosensitization. We suggest that altered cross-linking of HSP27 by ZER is a good strategy for abolishing HSP27-mediated resistance. Copyright © 2011 Elsevier Inc. All rights reserved.

Synthesis of controlled polymeric cross-linked coatings via iniferter polymerisation in the presence of tetraethyl thiuram disulphide chain terminator.

PubMed

Bossi, A; Whitcombe, M J; Uludag, Y; Fowler, S; Chianella, I; Subrahmanyam, S; Sanchez, I; Piletsky, S A

2010-05-15

A "grafting from" approach has been used for controlled deposition of cross-linked polymers by living radical polymerisation. Borosilicate glass was modified with N,N-diethylaminodithiocarbamoylpropyl(trimethoxy)silane, in order to confine the iniferter reactive groups solely at its surface, then placed in solution with monomers and cross-linker. The polymerisation was initiated by UV irradiation. Formation of the cross-linked polymers was studied in terms of time course of the reaction, type of monomers incorporated and influence of oxygen. Grafted surfaces were characterised by AFM, FT-IR, ellipsometry and contact angle measurements. The ability to control the grafted layer improved dramatically when the chain terminator agent, N,N-N',N'-tetraethyl thiuram disulphide (TED) was added. Upon irradiation TED increases the concentration of passive capping radicals and decreases the possibility of recombination of active macro-radicals, thus prolonging their lifetime. In the absence of TED the thickness of produced coatings was below 10 nm. TED added at different concentrations assisted in the formation of grafted layers of 10-130 nm thickness. Iniferter chemistry in the presence of TED can be used for growing nanometre-scale polymer layers on solid supports. It constitutes a robust general platform for controlled grafting and offer a general solution to address the needs of surface derivatisation in sensors technology. 2010 Elsevier B.V. All rights reserved.

Biocompatibility of NGF-grafted GTG membranes for peripheral nerve repair using cultured Schwann cells.

PubMed

Chen, Pei-Ru; Chen, Ming-Hong; Sun, Jui-Sheng; Chen, Mei-Hsiu; Tsai, Chien-Chen; Lin, Feng-Huei

2004-11-01

We previously developed a biodegradable composite with potentially good biocompatibility composed by tricalcium phosphate and gluataraldehyde cross-linking gelatin (GTG) with good mechanical property feasible for surgical manipulation. The purpose of this study was to evaluate the feasibility of immobilizing nerve growth factor (NGF) onto the composite (GTG) with carbodiimide (GEN composite). Cultured Schwann cells were seeded onto the GTG and GEN composites. For comparison, GTG membrane soaked in NGF solution without carbodiimide (GN composite) as cross-linking agent was also used to culture Schwann cells. Cell morphology was observed by a scanning electron microscope. Cell survival, cytotoxicity and cellular metabolism on the NGF-grafted GTG membrane were assessed quantitatively in terms of cell protein content, leakage of cytosolic lactate dehydrogenase (LDH) activity and by the well-established MTT assay, respectively. The result of LDH study did not show significant difference among GTG, NGF-modified GTG and control group. This indicated that GTG composite, whether cross-linking with NGF or not, has little cytotoxic effect. Comparing the protein content and MTT assay among GEN, GN composite and control group, the data confirmed more attachment of Schwann cells on GEN composite. Although GTG cross-linking with NGF did not promote Schwann cell proliferation, the techniques we used in this study provided a method to fabricate a novel biomaterial incorporation of Schwann cells and covalently immobilized NGF.

Photoinitiated chemical vapor deposition of cytocompatible poly(2-hydroxyethyl methacrylate) films.

PubMed

McMahon, Brian J; Pfluger, Courtney A; Sun, Bing; Ziemer, Katherine S; Burkey, Daniel D; Carrier, Rebecca L

2014-07-01

Poly(2-hydroxyethyl methacrylate) (pHEMA) is a widely utilized biomaterial due to lack of toxicity and suitable mechanical properties; conformal thin pHEMA films produced via chemical vapor deposition (CVD) would thus have broad biomedical applications. Thin films of pHEMA were deposited using photoinitiated CVD (piCVD). Incorporation of ethylene glycol diacrylate (EGDA) into the pHEMA polymer film as a crosslinker, confirmed via Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy, resulted in varied swelling and degradation behavior. 2-Hydroxyethyl methacrylate-only films showed significant thickness loss (up to 40%), possibly due to extraction of low-molecular-weight species or erosion, after 24 h in aqueous solution, whereas films crosslinked with EGDA (9.25-12.4%) were stable for up to 21 days. These results differ significantly from those obtained with plasma-polymerized pHEMA, which degraded steadily over a 21-day period, even with crosslinking. This suggests that the piCVD films differ structurally from those fabricated via plasma polymerization (plasma-enhanced CVD). piCVD pHEMA coatings proved to be good cell culture materials, with Caco-2 cell attachment and viability comparable to results obtained on tissue-culture polystyrene. Thus, thin film CVD pHEMA offers the advantage of enabling conformal coating of a cell culture substrate with tunable properties depending on method of preparation and incorporation of crosslinking agents. © 2013 Wiley Periodicals, Inc.

Epoxy Monomers Cured by High Cellulosic Nanocrystal Loading.

PubMed

Khelifa, Farid; Habibi, Youssef; Bonnaud, Leila; Dubois, Philippe

2016-04-27

The present study focuses on the use of cellulose nanocrystals (CNC) as the main constituent of a nanocomposite material and takes advantage of hydroxyl groups, characteristic of the CNC chemical structure, to thermally cross-link an epoxy resin. An original and simple approach is proposed, based on the collective sticking of CNC building blocks with the help of a DGEBA/TGPAP-based epoxy resin. Scientific findings suggest that hydroxyl groups act as a toxic-free cross-linking agent of the resin. The enhanced protection against water degradation as compared to neat CNC film and the improvement of mechanical properties of the synthesized films are attributed to a good compatibility between the CNC and the resin. Moreover, the preservation of CNC optical properties at high concentrations opens the way to applying these materials in photonic devices.

Dextran/Albumin hydrogel sealant for Dacron(R) vascular prosthesis.

PubMed

Lisman, Anna; Butruk, Beata; Wasiak, Iga; Ciach, Tomasz

2014-05-01

In this paper, the authors describe a novel type of hydrogel coating prepared from the copolymer of human serum albumin and oxidized dextran. The material was designed as a hydrogel sealant for polyester (Dacron®)-based vascular grafts. Dextran was chosen as a coating material due to its anti-thrombogenic properties. Prepared hydrogels were compared with similar, already known biomaterial made from gelatine with the same cross-linking agent. Obtained hydrogels, prepared from various ratios of oxidized dextran/albumin or oxidized dextran/gelatine, showed different cross-linking densities, which caused differences in swelling, degradation rate and mechanical properties. Permeability tests confirmed the complete tightness of the hydrogel-modified prosthesis. Results showed that application of the hydrogel coating provided leakage-free prosthesis and eliminated the need of pre-clotting.

Enzymic cross-linkage of monomeric extensin precursors in vitro. [Lycopersicon esculentum

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

Everdeen, D.S.; Kiefer, S.; Willard, J.J.

Rapidly growing tomato (Lycopersicon esculentum) cell suspension cultures contain transiently high levels of cell surface, salt-elutable, monomeric precursors to the covalently cross-linked extensin network of the primary cell wall. Thus, the authors purified a highly soluble monomeric extensin substrate from rapidly growing cells, and devised a soluble in vitro cross-linking assay based on Superose-6 fast protein liquid chromatography separation, which resolved extensin monomers from the newly formed oligomers within 25 minutes. Salt elution of slowly growing (early stationary phase) cells yielded little or no extensin monomers but did give a highly active enzymic preparation that specifically cross-linked extensin monomers inmore » the presence of hydrogen peroxide, judging from: (a) a decrease in the extensin monomer peak on fast protein liquid chromatography gel filtration, (b) appearance of oligomeric peaks, and (c) direct electron microscopical observation of the cross-linked oligomers. The cross-linking reaction had a broad pH optimum between 5.5 and 6.5. An approach to substrate saturation of the enzyme required extensin monomer concentrations of 20 to 40 milligrams per milliliter. Preincubation with catalase completely inhibited the cross-linking reaction, which was highly dependent on hydrogen peroxide and optimal at 15 to 50 micromolar. They therefore identified the cross-linking activity as extensin peroxidase.« less

Genome-wide Mapping of Cellular Protein–RNA Interactions Enabled by Chemical Crosslinking

PubMed Central

Li, Xiaoyu; Song, Jinghui; Yi, Chengqi

2014-01-01

RNA–protein interactions influence many biological processes. Identifying the binding sites of RNA-binding proteins (RBPs) remains one of the most fundamental and important challenges to the studies of such interactions. Capturing RNA and RBPs via chemical crosslinking allows stringent purification procedures that significantly remove the non-specific RNA and protein interactions. Two major types of chemical crosslinking strategies have been developed to date, i.e., UV-enabled crosslinking and enzymatic mechanism-based covalent capture. In this review, we compare such strategies and their current applications, with an emphasis on the technologies themselves rather than the biology that has been revealed. We hope such methods could benefit broader audience and also urge for the development of new methods to study RNA−RBP interactions. PMID:24747191

Preparation and Characterization of Chitosan-coated Fe3O4 Nanoparticles using Ex-Situ Co-Precipitation Method and Tripolyphosphate/Sulphate as Dual Crosslinkers

NASA Astrophysics Data System (ADS)

Wulandari, Ika O.; Mardila, Vita T.; Santjojo, D. J. Djoko H.; Sabarudin, Akhmad

2018-01-01

The unique properties of nanomaterial provide great opportunities to develop in several fields. Several types of nanoparticles have been proven beneficial for biomedical and therapeutic agent development. Particularly for clinical use, nanoparticles must be biocompatible and non-toxic. Iron oxide nanoparticles consist of either magnetite (Fe3O4) or maghemite (γ-Fe2O3) was eligible to use for in vivo application including targeting drug delivery. Due to their distinct properties, these nanoparticles could be directed to the specific target under external magnetic field. However, nanoparticles have a tendency to form agglomeration. Therefore, surface modification was required to reduce the agglomeration. In this study, nanoparticles of Fe3O4 were produced and coated by biomaterial (chitosan) using ex-situ co-precipitation method. Nanoparticles of Fe3O4 were synthesized by adding ammonia water into iron ferric and ferrous solution. Synthesis process of Fe3O4 was conducted prior to adding chitosan. Chitosan was then cross-linked by a combination of tripolyphosphate/sulphate. The different composition ratio and crosslinking time provide the different physical and magnetic characteristics of nanoparticles. Particle and crystallite size was determined by using Scanning Electron Microscopy (SEM) and X-Ray Diffraction (XRD) respectively, whereas magnetic characteristic was determined by Electron Spin Resonance (ESR). The results showed that the ratio enhancement between chitosan: Fe3O4 increase the particle size, while decreased the crystallite size. Morphology and particle size were influenced by the ratio of crosslinkers. It was found that the higher tripolyphosphate content was contributed to the small size and more spherical morphology. In addition, the influence of crosslinking time toward crystallite size was determined by altering stirring time. The longer duration of crosslinking time, provide the larger crystallite size of chitosan-Fe3O4. There was an interesting correlation between particle size and amount of Fe3O4 phase with ESR curve.

Cell proliferation on PVA/sodium alginate and PVA/poly(γ-glutamic acid) electrospun fiber.

PubMed

Yang, Jen Ming; Yang, Jhe Hao; Tsou, Shu Chun; Ding, Chian Hua; Hsu, Chih Chin; Yang, Kai Chiang; Yang, Chun Chen; Chen, Ko Shao; Chen, Szi Wen; Wang, Jong Shyan

2016-09-01

To overcome the obstacles of easy dissolution of PVA nanofibers without crosslinking treatment and the poor electrospinnability of the PVA cross-linked nanofibers via electrospinning process, the PVA based electrospun hydrogel nanofibers are prepared with post-crosslinking method. To expect the electrospun hydrogel fibers might be a promising scaffold for cell culture and tissue engineering applications, the evaluation of cell proliferation on the post-crosslinking electrospun fibers is conducted in this study. At beginning, poly(vinyl alcohol) (PVA), PVA/sodium alginate (PVASA) and PVA/poly(γ-glutamic acid) (PVAPGA) electrospun fibers were prepared by electrospinning method. The electrospun PVA, PVASA and PVAPGA nanofibers were treated with post-cross-linking method with glutaraldehyde (Glu) as crosslinking agent. These electrospun fibers were characterized with thermogravimetry analysis (TGA) and their morphologies were observed with a scanning electron microscope (SEM). To support the evaluation and explanation of cell growth on the fiber, the study of 3T3 mouse fibroblast cell growth on the surface of pure PVA, SA, and PGA thin films is conducted. The proliferation of 3T3 on the electrospun fiber surface of PVA, PVASA, and PVAPGA was evaluated by seeding 3T3 fibroblast cells on these crosslinked electrospun fibers. The cell viability on electrospun fibers was conducted with water-soluble tetrazolium salt-1 assay (Cell Proliferation Reagent WST-1). The morphology of the cells on the fibers was also observed with SEM. The results of WST-1 assay revealed that 3T3 cells cultured on different electrospun fibers had similar viability, and the cell viability increased with time for all electrospun fibers. From the morphology of the cells on electrospun fibers, it is found that 3T3 cells attached on all electrospun fiber after 1day seeded. Cell-cell communication was noticed on day 3 for all electrospun fibers. Extracellular matrix (ECM) productions were found and cell-ECM adhesion was shown on day 7. The cell number was also increased on all of the crosslinked electrospun fibers. It seems that the PVA based electrospun hydrogel nanofibers prepared with post-crosslinking method can be used as scaffold for tissue engineering. Copyright © 2016 Elsevier B.V. All rights reserved.

Novel Hydrogels from Renewable Resources

NASA Astrophysics Data System (ADS)

Karaaslan, Muzafer Ahmet

2011-12-01

The cell wall of most plant biomass from forest and agricultural resources consists of three major polymers, cellulose, hemicellulose and lignin. Of these, hemicelluloses have gained increasing attention as sustainable raw materials. In the first part of this study, novel pH-sensitive semi-IPN hydrogels based on hemicelluloses and chitosan were prepared using glutaraldehyde as the crosslinking agent. The hemicellulose isolated from aspen was analyzed for sugar content by HPLC, and its molecular weight distribution was determined by high performance size exclusion chromatography. Results revealed that hemicellulose had a broad molecular weight distribution with a fair amount of polymeric units, together with xylose, arabinose and glucose. The effect of hemicellulose content on mechanical properties and swelling behavior of hydrogels were investigated. The semi-IPNs hydrogel structure was confirmed by FT-IR, X-ray study and ninhydrin assay method. X-ray analysis showed that higher hemicellulose contents yielded higher crystallinity. Mechanical properties were mainly dependent on the crosslink density and average molecular weight between crosslinks. Swelling ratios increased with increasing hemicellulose content and were high at low pH values due to repulsion between similarly charged groups. In vitro release study of a model drug showed that these semi-IPN hydrogels could be used for controlled drug delivery into gastric fluid. The aim of the second part of this study was to control the crosslink density and the mechanical properties of hemicellulose/chitosan semi-IPN hydrogels by changing the crosslinking sequence. It has been hypothesized that by performing the crosslinking step before introducing hemicellulose, covalent crosslinking of chitosan would not be hindered and therefore more and/or shorter crosslinks could be formed. Furthermore, additional secondary interactions and crystalline domains introduced through hemicellulose could be favorable in terms of mechanical stability of semi-IPN hydrogels. In this last section of this study, the natural affinity of hemicellulose to cellulose was utilized to coat cellulose whiskers with chemically modified hemicellulose isolated from wood fiber. Surface modified cellulose nanowhiskers were used to prepare nanocomposite hydrogels using free radical polymerization of 2-hydroxyethyl methacrylate (HEMA), a biocompatible monomer. The effect of morphology and concentration of the incorporated nanocrystals on the hydrogel network was related to the mechanical properties, viscoelastic behavior and swelling of the hydrogels.

A multicomponent complex is required for the AAUAAA-dependent cross-linking of a 64-kilodalton protein to polyadenylation substrates.

PubMed Central

Wilusz, J; Shenk, T; Takagaki, Y; Manley, J L

1990-01-01

A 64-kilodalton (kDa) polypeptide that is cross-linked by UV light specifically to polyadenylation substrate RNAs containing a functional AAUAAA element has been identified previously. Fractionated HeLa nuclear components that can be combined to regenerate efficient and accurate polyadenylation in vitro have now been screened for the presence of the 64-kDa protein. None of the individual components contained an activity which could generate the 64-kDa species upon UV cross-linking in the presence of substrate RNA. It was necessary to mix two components, cleavage stimulation factor and specificity factor, to reconstitute 64-kDa protein-RNA cross-linking. The addition of cleavage factors to this mixture very efficiently reconstituted the AAUAAA-specific 64-kDa protein-RNA interaction. The 64-kDa protein, therefore, is present in highly purified, reconstituted polyadenylation reactions. However, it is necessary to form a multicomponent complex to efficiently cross-link the protein to a substrate RNA. Images PMID:2304466

Shear bond strength of composite to deep dentin after treatment with two different collagen cross-linking agents at varying time intervals.

PubMed

Srinivasulu, S; Vidhya, S; Sujatha, M; Mahalaxmi, S

2012-01-01

This in vitro study evaluated the shear bond strength of composite resin to deep dentin using a total etch adhesive after treatment with two collagen cross-linking agents at varying time intervals. Thirty freshly extracted human maxillary central incisors were sectioned longitudinally into equal mesial and distal halves (n=60). The proximal deep dentin was exposed, maintaining a remaining dentin thickness (RDT) of approximately 1 mm. The specimens were randomly divided into three groups based on the surface treatment of dentin prior to bonding as follows: group I (n=12, control): no prior dentin surface treatment; group II (n=24): dentin surface pretreated with 10% sodium ascorbate; and group III (n=24): dentin surface pretreated with 6.5% proanthocyanidin. Groups II and III were further subdivided into two subgroups of 12 specimens each, based on the pretreatment time of five minutes (subgroup A) and 10 minutes (subgroup B). Shear bond strength of the specimens was tested with a universal testing machine, and the data were statistically analyzed. Significantly higher shear bond strength to deep dentin was observed in teeth treated with 10% sodium ascorbate (group II) and 6.5% proanthocyanidin (group III) compared to the control group (group I). Among the collagen cross-linkers used, specimens treated with proanthocyanidin showed significantly higher shear bond strength values than those treated with sodium ascorbate. No significant difference was observed between the five-minute and 10-minute pretreatment times in groups II and III. It can be concluded that dentin surface pretreatment with both 10% sodium ascorbate and 6.5% proanthocyanidin resulted in significant improvement in bond strength of resin composite to deep dentin.

FANCD2 monoubiquitination and activation by hexavalent chromium [Cr(VI)] exposure

PubMed Central

Vilcheck, Susan K.; Ceryak, Susan; O’Brien, Travis J.; Patierno, Steven R.

2007-01-01

Fanconi anemia (FA) is a rare autosomal recessive disorder characterized by congenital abnormalities, progressive bone marrow failure, and cancer susceptibility. FA cells are hypersensitive to DNA crosslinking agents. FA is a genetically heterogeneous disease with at least 11 complementation groups. The eight cloned FA proteins interact in a common pathway with established DNA-damage-response proteins, including BRCA1 and ATM. Six FA proteins (A, C, E, F, G, and L) regulate the monoubiquitination of FANCD2 after DNA damage by crosslinking agents, which targets FANCD2 to BRCA1 nuclear foci containing BRCA2 (FANCD1) and RAD51. Some forms of hexavalent chromium [Cr(VI)] are implicated as respiratory carcinogens and induce several types of DNA lesions, including DNA interstrand crosslinks. We have shown that FA-A fibroblasts are hypersensitive to both Cr(VI)-induced apoptosis and clonogenic lethality. Here we show that Cr(VI) treatment induced monoubiquitination of FANCD2 in normal human fibroblasts, providing the first molecular evidence of Cr(VI)-induced activation of the FA pathway. FA-A fibroblasts demonstrated no FANCD2 monoubiquitination, in keeping with the requirement of FA-A for this modification. We also found that Cr(VI) treatment induced significantly more S-phase-dependent DNA double strand breaks (DSBs), as measured by γ-H2AX expression, in FA-A fibroblasts compared to normal cells. However, and notably, DSBs were repaired equally in both normal and FA-A fibroblasts during recovery from Cr(VI) treatment. While previous research on FA has defined the genetic causes of this disease, it is critical in terms of individual risk assessment to address how cells from FA patients respond to genotoxic insult. PMID:16893675

Preparation of three-dimensional macroporous chitosan-gelatin B microspheres and HepG2-cell culture.

PubMed

Huang, Fang; Cui, Long; Peng, Cheng-Hong; Wu, Xu-Bo; Han, Bao-San; Dong, Ya-Dong

2016-12-01

Chitosan-gelatin B microspheres with an open, interconnected, highly macroporous (100-200 µm) structure were prepared via a three-step protocol combining freeze-drying with an electrostatic and ionic cross-linking method. Saturated tripolyphosphate ethanol solution (85% ethanol) was chosen as the crosslinking agent to prevent destruction of the porous structure and to improve the biostability of the chitosan-gelatin B microspheres, with N-(3-dimethylaminopropyl)-N'-ethyl-carbodiimide/N-hydroxysuccinimide as a second crosslinking agent to react with gelatin A and fixed chitosan-gelatin B microspheres to attain improved biocompatibility. Water absorption of the three-dimensional macroporous chitosan-gelatin B microspheres (3D-P-CGMs) was 12.84, with a porosity of 85.45%. In vitro lysozyme degradation after 1, 3, 5, 7, 10, 14, and 21 days showed improved biodegradation in the 3D-P-CGMs. The morphology of human hepatoma cell lines (HepG2 cells) cultured on the 3D-P-CGMs was spherical, unlike that of cells cultured under traditional two-dimensional conditions. Scanning electron microscopy and paraffin sections were used to confirm the porous structure of the 3D-P-CGMs. HepG2 cells were able to migrate inside through the pore. Cell proliferation and levels of albumin and lactate dehydrogenase suggested that the 3D-P-CGMs could provide a larger specific surface area and an appropriate microenvironment for cell growth and survival. Hence, the 3D-P-CGMs are eminently suitable as macroporous scaffolds for cell cultures in tissue engineering and cell carrier studies. Copyright © 2014 John Wiley & Sons, Ltd. Copyright © 2014 John Wiley & Sons, Ltd.

D-Optimal mixture experimental design for stealth biodegradable crosslinked docetaxel-loaded poly-ε-caprolactone nanoparticles manufactured by dispersion polymerization.

PubMed

Ogunwuyi, O; Adesina, S; Akala, E O

2015-03-01

We report here our efforts on the development of stealth biodegradable crosslinked poly-ε-caprolactone nanoparticles by free radical dispersion polymerization suitable for the delivery of bioactive agents. The uniqueness of the dispersion polymerization technique is that it is surfactant free, thereby obviating the problems known to be associated with the use of surfactants in the fabrication of nanoparticles for biomedical applications. Aided by a statistical software for experimental design and analysis, we used D-optimal mixture statistical experimental design to generate thirty batches of nanoparticles prepared by varying the proportion of the components (poly-ε-caprolactone macromonomer, crosslinker, initiators and stabilizer) in acetone/water system. Morphology of the nanoparticles was examined using scanning electron microscopy (SEM). Particle size and zeta potential were measured by dynamic light scattering (DLS). Scheffe polynomial models were generated to predict particle size (nm) and particle surface zeta potential (mV) as functions of the proportion of the components. Solutions were returned from simultaneous optimization of the response variables for component combinations to (a) minimize nanoparticle size (small nanoparticles are internalized into disease organs easily, avoid reticuloendothelial clearance and lung filtration) and (b) maximization of the negative zeta potential values, as it is known that, following injection into the blood stream, nanoparticles with a positive zeta potential pose a threat of causing transient embolism and rapid clearance compared to negatively charged particles. In vitro availability isotherms show that the nanoparticles sustained the release of docetaxel for 72 to 120 hours depending on the formulation. The data show that nanotechnology platforms for controlled delivery of bioactive agents can be developed based on the nanoparticles.

Two-Dimensional Nanostructure- Reinforced Biodegradable Polymeric Nanocomposites for Bone Tissue Engineering

PubMed Central

Lalwani, Gaurav; Henslee, Allan M.; Farshid, Behzad; Lin, Liangjun; Kasper, F. Kurtis; Qin, Yi-Xian; Mikos, Antonios G.; Sitharaman, Balaji

2013-01-01

This study investigates the efficacy of two dimensional (2D) carbon and inorganic nanostructures as reinforcing agents of crosslinked composites of the biodegradable and biocompatible polymer polypropylene fumarate (PPF) as a function of nanostructure concentration. PPF composites were reinforced using various 2D nanostructures: single- and multi-walled graphene oxide nanoribbons (SWGONRs, MWGONRs), graphene oxide nanoplatelets (GONPs), and molybdenum di-sulfite nanoplatelets (MSNPs) at 0.01–0.2 weight% concentrations. Cross-linked PPF was used as the baseline control, and PPF composites reinforced with single- or multi-walled carbon nanotubes (SWCNT, MWCNT) were used as positive controls. Compression and flexural testing show a significant enhancement (i.e., compressive modulus = 35–108%, compressive yield strength = 26–93%, flexural modulus = 15–53%, and flexural yield strength = 101–262% greater than the baseline control) in the mechanical properties of the 2D-reinforced PPF nanocomposites. MSNPs nanocomposites consistently showed the highest values among the experimental or control groups in all the mechanical measurements. In general, the inorganic nanoparticle MSNPs showed a better or equivalent mechanical reinforcement compared to carbon nanomaterials, and 2-D nanostructures (GONP, MSNP) are better reinforcing agents compared to 1-D nanostructures (e.g. SWCNTs). The results also indicate that the extent of mechanical reinforcement is closely dependent on the nanostructure morphology and follows the trend nanoplatelets > nanoribbons > nanotubes. Transmission electron microscopy of the cross-linked nanocomposites indicates good dispersion of nanomaterials in the polymer matrix without the use of a surfactant. The sol-fraction analysis showed significant changes in the polymer cross-linking in the presence of MSNP (0.01–0.2 wt %) and higher loading concentrations of GONP and MWGONR (0.1–0.2 wt%). The analysis of surface area and aspect ratio of the nanostructures taken together with the above results indicates differences in nanostructure architecture (2D vs. 1D nanostructures), as well as the chemical compositions (inorganic vs. carbon nanostructures), number of functional groups, and structural defects for the 2D nanostructures maybe key properties that affect the mechanical properties of 2D nanostructure-reinforced PPF nanocomposites, and the reason for the enhanced mechanical properties compared to the controls. PMID:23405887

Evidence for the Location of the Allosteric Activation Switch in the Multisubunit Phosphorylase Kinase Complex from Mass Spectrometric Identification of Chemically Crosslinked Peptides*

PubMed Central

Nadeau, Owen W.; Anderson, David W.; Yang, Qing; Artigues, Antonio; Paschall, Justin E.; Wyckoff, Gerald J.; McClintock, Jennifer L.; Carlson, Gerald M.

2007-01-01

Phosphorylase kinase (PhK), an (αβγδ)4 complex, regulates glycogenolysis. Its activity, catalyzed by the γ subunit, is tightly controlled by phosphorylation and activators acting through allosteric sites on its regulatory α, β and δ subunits. Activation of the catalytic γ subunit in the PhK complex by phosphorylation is known to be predominantly mediated by the regulatory β subunit, which undergoes a conformational change that is structurally linked with the γ subunit and that is characterized by the ability to form β-β dimers using a short chemical crosslinker. To determine potential regions of interaction of the β and γ subunits, we have used chemical crosslinking and 2-hybrid screening. The β and γ subunits were chemically crosslinked to each other in phosphorylated PhK, and crosslinked peptides were identified in digests of the kinase by Fourier transform mass spectrometry in combination with a search engine developed ‘in house’ that generates a hypothetical list of crosslinked peptides. Such a conjugate between β and γ was identified, verified by MS/MS and shown to correspond to crosslinking between K303 in the C-terminal regulatory domain of γ (γCRD) and R18 in the N-terminal regulatory region of β (β1-31), which contains the phosphorylatable serines 11 and 26. A synthetic peptide corresponding to residues 1-22 of β inhibited the crosslinking between β and γ in the complex, and was itself crosslinked to K303 of γ. Through the use of 2-hybrid screening, the β1-31 region was also shown to control β subunit self-interactions, which were favored by truncation of this region or by mutation of the phosphorylatable serines 11 and 26, thus providing structural evidence for a phosphorylation-dependent subunit communication network in the PhK complex involving at least these two regulatory regions of the β and γ subunits. The sum of our results considered together with previous findings implicates the γCRD as being an allosteric activation switch in PhK that interacts with all three of the enzyme’s regulatory subunits and is proximal to the active site cleft. PMID:17123541

Neighborhood of 16S rRNA nucleotides U788/U789 in the 30S ribosomal subunit determined by site-directed crosslinking.

PubMed

Mundus, D; Wollenzien, P

1998-11-01

Site-specific photo crosslinking has been used to investigate the RNA neighborhood of 16S rRNA positions U788/ U789 in Escherichia coli 30S subunits. For these studies, site-specific psoralen (SSP) which contains a sulfhydryl group on a 17 A side chain was first added to nucleotides U788/U789 using a complementary guide DNA by annealing and phototransfer. Modified RNA was purified from the DNA and unmodified RNA. For some experiments, the SSP, which normally crosslinks at an 8 A distance, was derivitized with azidophenacylbromide (APAB) resulting in the photoreactive azido moiety at a maximum of 25 A from the 4' position on psoralen (SSP25APA). 16S rRNA containing SSP, SSP25APA or control 16S rRNA were reconstituted and 30S particles were isolated. The reconstituted subunits containing SSP or SSP25APA had normal protein composition, were active in tRNA binding and had the usual pattern of chemical reactivity except for increased kethoxal reactivity at G791 and modest changes in four other regions. Irradiation of the derivatized 30S subunits in activation buffer produced several intramolecular RNA crosslinks that were visualized and separated by gel electrophoresis and characterized by primer extension. Four major crosslink sites made by the SSP reagent were identified at positions U561/U562, U920/U921, C866 and U723; a fifth major crosslink at G693 was identified when the SSP25APA reagent was used. A number of additional crosslinks of lower frequency were seen, particularly with the APA reagent. These data indicate a central location close to the decoding region and central pseudoknot for nucleotides U788/U789 in the activated 30S subunit.

Tissue Engineering of Ureteral Grafts: Preparation of Biocompatible Crosslinked Ureteral Scaffolds of Porcine Origin

PubMed Central

Koch, Holger; Hammer, Niels; Ossmann, Susann; Schierle, Katrin; Sack, Ulrich; Hofmann, Jörg; Wecks, Mike; Boldt, Andreas

2015-01-01

The surgical reconstruction of ureteric defects is often associated with post-operative complications and requires additional medical care. Decellularized ureters originating from porcine donors could represent an alternative therapy. Our aim was to investigate the possibility of manufacturing decellularized ureters, the characteristics of the extracellular matrix (ECM) and the biocompatibility of these grafts in vitro/in vivo after treatment with different crosslinking agents. To achieve these goals, native ureters were obtained from pigs and were decellularized. The success of decellularization and the ECM composition were characterized by (immuno)histological staining methods and a DNA-assay. In vitro: scaffolds were crosslinked either with carbodiimide (CDI), genipin (GP), glutaraldehyde, left chemically untreated or were lyophilized. Scaffolds in each group were reseeded with Caco2, LS48, 3T3 cells, or native rat smooth muscle cells (SMC). After 2 weeks, the number of ingrown cells was quantified. In vivo: crosslinked scaffolds were implanted subcutaneously into rats and the type of infiltrating cells were determined after 1, 9, and 30 days. After decellularization, scaffold morphology and composition of ECM were maintained, all cellular components were removed, DNA destroyed and strongly reduced. In vitro: GP and CDI scaffolds revealed a higher number of ingrown 3T3 and SMC cells as compared to untreated scaffolds. In vivo: at day 30, implants were predominantly infiltrated by fibroblasts and M2 anti-inflammatory macrophages. A maximum of MMP3 was observed in the CDI group at day 30. TIMP1 was below the detection limit. In this study, we demonstrated the potential of decellularization to create biocompatible porcine ureteric grafts, whereas a CDI-crosslink may facilitate the remodeling process. The use of decellularized ureteric grafts may represent a novel therapeutic method in reconstruction of ureteric defects. PMID:26157796

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

PubMed

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

2014-09-02

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

In Vitro Cytocompatibility of One- and Two-Dimensional Nanostructure-Reinforced Biodegradable Polymeric Nanocomposites

PubMed Central

Farshid, Behzad; Lalwani, Gaurav; Sitharaman, Balaji

2015-01-01

This study investigates the in vitro cytocompatibility of one- and two-dimensional (1-D and 2-D) carbon and inorganic nanomaterial reinforced polymeric nanocomposites fabricated using biodegradable polymer poly (propylene fumarate), crosslinking agent N-vinyl pyrrolidone (NVP) and following nanomaterials: single- and multi- walled carbon nanotubes, single- and multi- walled graphene oxide nanoribbons, graphene oxide nanoplatelets, molybdenum disulfide nanoplatelets, or tungsten disulfide nanotubes dispersed between 0.02–0.2 wt% concentrations in the polymer. The extraction media of unreacted components, crosslinked nanocomposites and their degradation products between 1X-100X dilutions were examined for effects on viability and attachment employing two cell lines: NIH3T3 fibroblasts and MC3T3 pre-osteoblasts. The extraction media of unreacted PPF/NVP elicited acute dose-dependent cytotoxicity attributed to leaching of unreacted components into cell culture media. However, extraction media of crosslinked nanocomposites showed no dose dependent adverse effects. Further, all crosslinked nanocomposites showed high viability (78–100%), high cellular attachment (40–55%), and spreading that was confirmed by confocal and scanning electron microscopy. Degradation products of nanocomposites showed a mild dose-dependent cytotoxicity possibly due to acidic degradation components of PPF. In general, compared to PPF control, none of the nanocomposites showed significant differences in cellular response to the unreacted components, crosslinked nanocomposites and their degradation products. The initial minor cytotoxic response and lower cell attachment numbers were observed only for a few nanocomposite groups; these effects were absent at later time points for all PPF nanocomposites. The favorable cytocompatibility results for all the nanocomposites opens avenues for in vivo safety and efficacy studies for bone tissue engineering applications. PMID:25367032

Phosphodiester-mediated reaction of cisplatin with guanine in oligodeoxyribonucleotides.

PubMed

Campbell, Meghan A; Miller, Paul S

2008-12-02

The cancer chemotherapeutic agent cis-diamminedichloroplatinum(II) or cisplatin reacts primarily with guanines in DNA to form 1,2-Pt-GG and 1,3-Pt-GNG intrastrand cross-links and, to a lesser extent, G-G interstrand cross-links. Recent NMR evidence has suggested that cisplatin can also form a coordination complex with the phosphodiester internucleotide linkage of DNA. We have examined the effects of the phosphodiester backbone on the reactions of cisplatin with oligodeoxyribonucleotides that lack or contain a GTG sequence. Cisplatin forms a stable adduct with TpT that can be isolated by reversed phase HPLC. The cis-Pt-TpT adduct contains a single Pt, as determined by atomic absorption spectroscopy (AAS) and by electrospray ionization mass spectrometry (ESI-MS), and is resistant to digestion by snake venom phosphodiesterase. Treatment of the adduct with sodium cyanide regenerates TpT. Similar adduct formation was observed when T(pT)(8) was treated with cisplatin, but not when the phosphodiester linkages of T(pT)(8) were replaced with methylphosphonate groups. These results suggest that the platinum may be coordinated with the oxygens of the thymine and possibly with those of the phosphodiester group. As expected, reaction of a 9-mer containing a GTG sequence with cisplatin yielded an adduct that contained a 1,3-Pt-GTG intrastrand cross-link. However, we found that the number and placement of phosphodiesters surrounding a GTG sequence significantly affected intrastrand cross-link formation. Increasing the number of negatively charged phosphodiesters in the oligonucleotide increased the amount of GTG platination. Surrounding the GTG sequence with nonionic methylphosphonate linkages inhibited or eliminated cross-link formation. These observations suggest that interactions between cisplatin and the negatively charged phosphodiester backbone may play an important role in facilitating platination of guanine nucleotides in DNA.

Development of horseradish peroxidase-based cross-linked enzyme aggregates and their environmental exploitation for bioremediation purposes.

PubMed

Bilal, Muhammad; Iqbal, Hafiz M N; Hu, Hongbo; Wang, Wei; Zhang, Xuehong

2017-03-01

In the present study, horseradish peroxidase (HRP), in-house isolated crude cocktail enzyme, from Armoracia rusticana was cross-linked using a new type of cross-linking agent, i.e., ethylene glycol-bis [succinic acid N-hydroxysuccinimide, (EG-NHS)], which is mild in nature as compared to the glutaraldehyde (GA). The HRP-immobilized cross-linked enzyme aggregates (HRP-CLEAs) were developed using a wider range of EG-NHS and notably no adverse effect was observed. In a comparative evaluation, in the case of EG-NHS, a high-level stability in the residual activity was recorded, whereas a sharp decrease was observed in the case of glutaraldehyde. Following initial cross-linker evaluation, the HRP-CLEAs were tested to investigate their bio-catalytic efficacy for bioremediation purposes using a newly developed packed bed reactor system (PBRS). A maximal of 94.26% degradation of textile-based methyl orange dye was recorded within the shortest time frame, following 91.73% degradation of basic red 9, 84.35% degradation of indigo, 81.47% degradation of Rhodamin B, and 73.6% degradation of Rhodamine 6G, respectively, under the same working environment. Notably, the HRP-CLEAs retained almost 60% of its original activity after methyl orange dye degradation in seven consecutive cycles using PBRS. Furthermore, after HRP-CLEAs-mediated treatment in the PBRS, a significant toxicity reduction in the dye samples was recorded as compared to their pristine counterparts. In conclusion, the results suggest that the newly developed HRP-CLEAs have a great potential for industrial exploitation, to tackle numerous industrial dye-based emergent pollutants. Copyright © 2016 Elsevier Ltd. All rights reserved.

DNA Damage Induced by Alkylating Agents and Repair Pathways

PubMed Central

Kondo, Natsuko; Takahashi, Akihisa; Ono, Koji; Ohnishi, Takeo

2010-01-01

The cytotoxic effects of alkylating agents are strongly attenuated by cellular DNA repair processes, necessitating a clear understanding of the repair mechanisms. Simple methylating agents form adducts at N- and O-atoms. N-methylations are removed by base excision repair, AlkB homologues, or nucleotide excision repair (NER). O6-methylguanine (MeG), which can eventually become cytotoxic and mutagenic, is repaired by O6-methylguanine-DNA methyltransferase, and O6MeG:T mispairs are recognized by the mismatch repair system (MMR). MMR cannot repair the O6MeG/T mispairs, which eventually lead to double-strand breaks. Bifunctional alkylating agents form interstrand cross-links (ICLs) which are more complex and highly cytotoxic. ICLs are repaired by complex of NER factors (e.g., endnuclease xeroderma pigmentosum complementation group F-excision repair cross-complementing rodent repair deficiency complementation group 1), Fanconi anemia repair, and homologous recombination. A detailed understanding of how cells cope with DNA damage caused by alkylating agents is therefore potentially useful in clinical medicine. PMID:21113301

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

Walker, E.M. Jr.; Gale, G.R.

Cisplatin, an agent widely used in the chemotherapy of a variety of human malignancies, is often dose-limited owing to its nephrotoxicity. Some of the approaches under consideration, regarding the reduction of cisplatin nephrotoxicity, include the use of hydration and osmotic diuresis, pharmacological diuretics, chelating agents or agents which otherwise react with cisplatin or reverse cisplatin-induced deoxyribonucleic acid cross-links, and antioxidants to destroy free radicals, especially superoxide radicals, produced by cisplatin. The effects of each of these and other interventions on cisplatin-induced nephrotoxicity are delineated, along with their proposed mechanisms and effects on therapeutic efficacy. The current status of development ofmore » organoplatinum analogs yielding congeners with less nephrotoxicity and greater efficacy is discussed briefly. Finally, a possible role of endogenous and/or exogenous prostaglandins in protecting against or reversing heavy metal nephrotoxicity is suggested.« less

Ionic Liquid Epoxy Resin Monomers

NASA Technical Reports Server (NTRS)

Paley, Mark S. (Inventor)

2013-01-01

Ionic liquid epoxide monomers capable of reacting with cross-linking agents to form polymers with high tensile and adhesive strengths. Ionic liquid epoxide monomers comprising at least one bis(glycidyl) N-substituted nitrogen heterocyclic cation are made from nitrogen heterocycles corresponding to the bis(glycidyl) N-substituted nitrogen heterocyclic cations by a method involving a non-nucleophilic anion, an alkali metal cation, epichlorohydrin, and a strong base.

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.

Polymer Claw: Instant Underwater Adhesive

DTIC Science & Technology

2012-03-23

technology is the use of pressure sensitive microcapsules , which release reactive amine crosslinkers into an adhesive putty when pressed against the...PROIECT GOALS AND OBIECTIVES 2 2 KEY ACCOMPLISHMENTS 2 3.1 KICKOFF MEETING 3 3.2 AMINE MICROENCAPSULATION 3 3.3 CAUSTIC CLEANING AGENT 5 3.4...caustic, and the abrasive brush. We successfully synthesized amine-filled microcapsules and a dry mixture of caustic ingredients that only activate when

Pelletized ponderosa pine bark for adsorption of toxic heavy metals from water

Treesearch

Miyoung Oh; Mandla A. Tshabalala

2007-01-01

Bark flour from ponderosa pine (Pinus ponderosa) was consolidated into pellets using citric acid as cross-linking agent. The pellets were evaluated for removal of toxic heavy metals from synthetic aqueous solutions. When soaked in water, pellets did not leach tannins, and they showed high adsorption capacity for Cu(ll), Zn(ll), Cd(ll). and Ni(ll) under both equilibrium...

Post-Fusion Membrane Reorganization.

DTIC Science & Technology

1993-01-27

diphosphoglycerate , and NEM (a crosslinking agent), and ethanol treatments all had reproducible and very specific effects on the kinetic phases and the fusion product...actually, at the ultrastructure level , a double membrane multiply perforated with fusion sites (or pores). Also, because the heat treatment was within...relationships. Moreover. 2.3- Diphosphoglycerate (2-3-DPG). a naturally occuring metabolite which is known to have a regulatory role in spectrin-cytoskeletal

pH responsive N-succinyl chitosan/Poly (acrylamide-co-acrylic acid) hydrogels and in vitro release of 5-fluorouracil.

PubMed

Bashir, Shahid; Teo, Yin Yin; Naeem, Sumaira; Ramesh, S; Ramesh, K

2017-01-01

There has been significant progress in the last few decades in addressing the biomedical applications of polymer hydrogels. Particularly, stimuli responsive hydrogels have been inspected as elegant drug delivery systems capable to deliver at the appropriate site of action within the specific time. The present work describes the synthesis of pH responsive semi-interpenetrating network (semi-IPN) hydrogels of N-succinyl-chitosan (NSC) via Schiff base mechanism using glutaraldehyde as a crosslinking agent and Poly (acrylamide-co-acrylic acid)(Poly (AAm-co-AA)) was embedded within the N-succinyl chitosan network. The physico-chemical interactions were characterized by Fourier transform infrared (FTIR), X-ray diffraction (XRD), thermogravimetric analysis (TGA), and field emission scanning electron microscope (FESEM). The synthesized hydrogels constitute porous structure. The swelling ability was analyzed in physiological mediums of pH 7.4 and pH 1.2 at 37°C. Swelling properties of formulations with various amounts of NSC/ Poly (AAm-co-AA) and crosslinking agent at pH 7.4 and pH 1.2 were investigated. Hydrogels showed higher swelling ratios at pH 7.4 while lower at pH 1.2. Swelling kinetics and diffusion parameters were also determined. Drug loading, encapsulation efficiency, and in vitro release of 5-fluorouracil (5-FU) from the synthesized hydrogels were observed. In vitro release profile revealed the significant influence of pH, amount of NSC, Poly (AAm-co-AA), and crosslinking agent on the release of 5-FU. Accordingly, rapid and large release of drug was observed at pH 7.4 than at pH 1.2. The maximum encapsulation efficiency and release of 5-FU from SP2 were found to be 72.45% and 85.99%, respectively. Kinetics of drug release suggested controlled release mechanism of 5-FU is according to trend of non-Fickian. From the above results, it can be concluded that the synthesized hydrogels have capability to adapt their potential exploitation as targeted oral drug delivery carriers.

pH responsive N-succinyl chitosan/Poly (acrylamide-co-acrylic acid) hydrogels and in vitro release of 5-fluorouracil

PubMed Central

Bashir, Shahid; Teo, Yin Yin; Naeem, Sumaira; Ramesh, S.; Ramesh, K.

2017-01-01

There has been significant progress in the last few decades in addressing the biomedical applications of polymer hydrogels. Particularly, stimuli responsive hydrogels have been inspected as elegant drug delivery systems capable to deliver at the appropriate site of action within the specific time. The present work describes the synthesis of pH responsive semi-interpenetrating network (semi-IPN) hydrogels of N-succinyl-chitosan (NSC) via Schiff base mechanism using glutaraldehyde as a crosslinking agent and Poly (acrylamide-co-acrylic acid)(Poly (AAm-co-AA)) was embedded within the N-succinyl chitosan network. The physico-chemical interactions were characterized by Fourier transform infrared (FTIR), X-ray diffraction (XRD), thermogravimetric analysis (TGA), and field emission scanning electron microscope (FESEM). The synthesized hydrogels constitute porous structure. The swelling ability was analyzed in physiological mediums of pH 7.4 and pH 1.2 at 37°C. Swelling properties of formulations with various amounts of NSC/ Poly (AAm-co-AA) and crosslinking agent at pH 7.4 and pH 1.2 were investigated. Hydrogels showed higher swelling ratios at pH 7.4 while lower at pH 1.2. Swelling kinetics and diffusion parameters were also determined. Drug loading, encapsulation efficiency, and in vitro release of 5-fluorouracil (5-FU) from the synthesized hydrogels were observed. In vitro release profile revealed the significant influence of pH, amount of NSC, Poly (AAm-co-AA), and crosslinking agent on the release of 5-FU. Accordingly, rapid and large release of drug was observed at pH 7.4 than at pH 1.2. The maximum encapsulation efficiency and release of 5-FU from SP2 were found to be 72.45% and 85.99%, respectively. Kinetics of drug release suggested controlled release mechanism of 5-FU is according to trend of non-Fickian. From the above results, it can be concluded that the synthesized hydrogels have capability to adapt their potential exploitation as targeted oral drug delivery carriers. PMID:28678803

Sarcosine influences apoptosis and growth of prostate cells via cell-type specific regulation of distinct sets of genes.

PubMed

Rodrigo, Miguel A Merlos; Strmiska, Vladislav; Horackova, Eva; Buchtelova, Hana; Michalek, Petr; Stiborova, Marie; Eckschlager, Tomas; Adam, Vojtech; Heger, Zbynek

2018-02-01

Sarcosine is a widely discussed oncometabolite of prostate cells. Although several reports described connections between sarcosine and various phenotypic changes of prostate cancer (PCa) cells, there is still a lack of insights on the complex phenomena of its effects on gene expression patterns, particularly in non-malignant and non-metastatic cells. To shed more light on this phenomenon, we performed parallel microarray profiling of RNA isolated from non-malignant (PNT1A), malignant (22Rv1), and metastatic (PC-3) prostate cell lines treated with sarcosine. Microarray results were experimentally verified using semi-quantitative-RT-PCR, clonogenic assay, through testing of the susceptibility of cells pre-incubated with sarcosine to anticancer agents with different modes of actions (inhibitors of topoisomerase II, DNA cross-linking agent, antimicrotubule agent and inhibitor of histone deacetylases) and by evaluation of activation of executioner caspases 3/7. We identified that irrespective of the cell type, sarcosine stimulates up-regulation of distinct sets of genes involved in cell cycle and mitosis, while down-regulates expression of genes driving apoptosis. Moreover, it was found that in all cell types, sarcosine had pronounced stimulatory effects on clonogenicity. Except of an inhibitor of histone deacetylase valproic acid, efficiency of all agents was significantly (P < 0.05) decreased in sarcosine pre-incubated cells. Our comparative study brings evidence that sarcosine affects not only metastatic PCa cells, but also their malignant and non-malignant counterparts and induces very similar changes in cells behavior, but via distinct cell-type specific targets. © 2017 Wiley Periodicals, Inc.

Alkaline Comet Assay for Assessing DNA Damage in Individual Cells.

PubMed

Pu, Xinzhu; Wang, Zemin; Klaunig, James E

2015-08-06

Single-cell gel electrophoresis, commonly called a comet assay, is a simple and sensitive method for assessing DNA damage at the single-cell level. It is an important technique in genetic toxicological studies. The comet assay performed under alkaline conditions (pH >13) is considered the optimal version for identifying agents with genotoxic activity. The alkaline comet assay is capable of detecting DNA double-strand breaks, single-strand breaks, alkali-labile sites, DNA-DNA/DNA-protein cross-linking, and incomplete excision repair sites. The inclusion of digestion of lesion-specific DNA repair enzymes in the procedure allows the detection of various DNA base alterations, such as oxidative base damage. This unit describes alkaline comet assay procedures for assessing DNA strand breaks and oxidative base alterations. These methods can be applied in a variety of cells from in vitro and in vivo experiments, as well as human studies. Copyright © 2015 John Wiley & Sons, Inc.

Multimodal and self-healable interfaces enable strong and tough graphene-derived materials

NASA Astrophysics Data System (ADS)

Liu, Yilun; Xu, Zhiping

2014-10-01

Recent studies have shown that graphene-derived materials not only feature outstanding multifunctional properties, but also act as model materials to implant nanoscale structural engineering insights into their macroscopic performance optimization. In this work, we explore strengthening and toughening strategies of this class of materials by introducing multimodal crosslinks, including long, strong and short, self-healable ones. We identify two failure modes by fracturing functionalized graphene sheets or their crosslinks, and the role of brick-and-mortar hierarchy in mechanical enhancement. Theoretical analysis and atomistic simulation results show that multimodal crosslinks synergistically transfer tensile load to enhance the strength, whereas reversible rupture and formation of healable crosslinks improve the toughness. These findings lay the ground for future development of high-performance paper-, fiber- or film-like macroscopic materials from low-dimensional structures with engineerable interfaces.

Preparation and characterization of (St-DVB-MAA) ion exchange resins

NASA Astrophysics Data System (ADS)

Jiang, Shanquan; Sun, Xiangwei; Ling, Lixing; Wang, Shumin; Wu, Wufeng; Cheng, Shihong; Hu, Yue; Zhong, Chunyan

2017-08-01

In this paper, used polyvinyl alcohol as dispersing agent, Benzoyl peroxide as initiator of polymerization, Divinyl benzene as cross-linking agent, Styrene and 2-Methylpropenoic acid as monomer, ion exchange resin (copolymer of St-DVB-MAA)were prepared by suspension polymerization on 80°C. The structures, components and properties of the prepared composite micro gels were characterized by Fourier transform infrared spectroscopy (FT-IR), thermogravimetric analysis (TGA). The experiment of ion exchange was conducted by resin to deal with copper ions in the solution. The result showed that performance of the ion exchange capacity was excellent, which impacted by pH.

Stoichiometry of the large conductance bacterial mechanosensitive channel of E. coli. A biochemical study

NASA Technical Reports Server (NTRS)

Sukharev, S. I.; Schroeder, M. J.; McCaslin, D. R.

1999-01-01

MscL, a 15 kDa transmembrane protein, is the only component involved in the formation of a 3 nS channel in the inner membrane of Escherichia coli that opens in response to mechanical or osmotic stress. While previous data had suggested that the functional MscL complex might be a hexamer, a recent crystallographic study of the MscL homologue from M. tuberculosis reveals a pentameric structure. The present work further examines the stoichiometry of the E. coli MscL using a variety of biochemical approaches. Detergent-purified 6His-MscL in solution and MscL in the membrane could be chemically crosslinked with the products displaying ladderlike patterns on SDS gels. Three crosslinking agents (EDC, DMS, and DMA) used at saturating concentrations invariably generated pentamers as the largest product. DSS produced additional bands corresponding to larger complexes although the pentamer band appeared to be the predominant product at high levels of crosslinker. It is not clear whether these extra bands reflect a difference in the crosslinking chemistry of DSS or whether its spacer arm is the longest of those used, or a combination of both facts. For the detergent-solubilized 6His-MscL both sedimentation equilibrium and gel chromatography showed the presence of multiple species. Thus the longer spacer arm could permit both intra- and intercomplex linkages. Nonetheless, the patterns obtained with all agents are consistent with and strongly suggest a pentameric organization for the MscL channel. Expression of MscL as genetically engineered double or triple subunit tandems yields low numbers of functional channels as compared to expressed monomers. The double-tandem assemblies must have an even number of subunits and crosslinking in the membrane confirmed hexamerization. Gel chromatography clearly demonstrated that the channels formed from the double tandems were larger than those formed from WT MscL, consistent with the native channel being pentameric. The observation that both double and triple tandems form channels of normal conductance implies that the pentameric assembly is to some degree independent of the number of subunit repeats in the polypeptide precursor. The channel is thus a pentameric core with the 'extra' subunits left out of the functional complex. From sedimentation equilibrium and size-exclusion chromatography, we also conclude that MscL complexes are not in a dynamic equilibrium with monomers, but are pre-assembled; and thus, their gating properties must result from changes in the conformation of the entire complex induced by the mechanical stress.

Stoichiometry of the large conductance bacterial mechanosensitive channel of E. coli. A biochemical study.

PubMed

Sukharev, S I; Schroeder, M J; McCaslin, D R

1999-10-01

MscL, a 15 kDa transmembrane protein, is the only component involved in the formation of a 3 nS channel in the inner membrane of Escherichia coli that opens in response to mechanical or osmotic stress. While previous data had suggested that the functional MscL complex might be a hexamer, a recent crystallographic study of the MscL homologue from M. tuberculosis reveals a pentameric structure. The present work further examines the stoichiometry of the E. coli MscL using a variety of biochemical approaches. Detergent-purified 6His-MscL in solution and MscL in the membrane could be chemically crosslinked with the products displaying ladderlike patterns on SDS gels. Three crosslinking agents (EDC, DMS, and DMA) used at saturating concentrations invariably generated pentamers as the largest product. DSS produced additional bands corresponding to larger complexes although the pentamer band appeared to be the predominant product at high levels of crosslinker. It is not clear whether these extra bands reflect a difference in the crosslinking chemistry of DSS or whether its spacer arm is the longest of those used, or a combination of both facts. For the detergent-solubilized 6His-MscL both sedimentation equilibrium and gel chromatography showed the presence of multiple species. Thus the longer spacer arm could permit both intra- and intercomplex linkages. Nonetheless, the patterns obtained with all agents are consistent with and strongly suggest a pentameric organization for the MscL channel. Expression of MscL as genetically engineered double or triple subunit tandems yields low numbers of functional channels as compared to expressed monomers. The double-tandem assemblies must have an even number of subunits and crosslinking in the membrane confirmed hexamerization. Gel chromatography clearly demonstrated that the channels formed from the double tandems were larger than those formed from WT MscL, consistent with the native channel being pentameric. The observation that both double and triple tandems form channels of normal conductance implies that the pentameric assembly is to some degree independent of the number of subunit repeats in the polypeptide precursor. The channel is thus a pentameric core with the 'extra' subunits left out of the functional complex. From sedimentation equilibrium and size-exclusion chromatography, we also conclude that MscL complexes are not in a dynamic equilibrium with monomers, but are pre-assembled; and thus, their gating properties must result from changes in the conformation of the entire complex induced by the mechanical stress.

Genipin cross-linked type II collagen/chondroitin sulfate composite hydrogel-like cell delivery system induces differentiation of adipose-derived stem cells and regenerates degenerated nucleus pulposus.

PubMed

Zhou, Xiaopeng; Wang, Jingkai; Fang, Weijing; Tao, Yiqing; Zhao, Tengfei; Xia, Kaishun; Liang, Chengzhen; Hua, Jianming; Li, Fangcai; Chen, Qixin

2018-04-15

Nucleus pulposus (NP) degeneration is usually the origin of intervertebral disc degeneration and consequent lower back pain. Although adipose-derived stem cell (ADSC)-based therapy is regarded to be promising for the treatment of degenerated NP, there is a lack of viable cell carriers to transplant ADSCs into the NP while maintaining cell function. In this study, we developed a type II collagen/chondroitin sulfate (CS) composite hydrogel-like ADSC (CCSA) delivery system with genipin as the cross-linking agent. The induction effect of the scaffold on ADSC differentiation was studied in vitro, and a rat coccygeal vertebrae degeneration model was used to investigate the regenerative effect of the CCSA system on the degenerated NP in vivo. The results showed that the CCSA delivery system cross-linked with 0.02% genipin was biocompatible and promoted the expressions of NP-specific genes. After the injection of the CCSA system, the disc height, water content, extracellular matrix synthesis, and structure of the degenerated NP were partly restored. Our CCSA delivery system uses minimally invasive approaches to promote the regeneration of degenerated NP and provides an exciting new avenue for the treatment of degenerative disc disease. Nucleus pulposus (NP) degeneration is usually the origin of intervertebral disc degeneration and consequent lower back pain. Stem cell-based tissue engineering is a promising method in NP regeneration, but there is a lack of viable cell carriers to transplant ADSCs into the NP while maintaining cell function. In this study, we developed a type II collagen/chondroitin sulfate (CS) composite hydrogel-like ADSC (CCSA) delivery system with genipin as the cross-linking agent. Although several research groups have studied the fabrication of injectable hydrogel with biological matrix, our study differs from other works. We chose type II collagen and CS, the two primary native components in the NP, as the main materials and combined them according to the natural ratio of collagen and sGAG in the NP. The delivery system is preloaded with ADSCs and can be injected into the NP with a needle, followed by in situ gelation. Genipin is used as a cross-linker to improve the bio-stability of the scaffold, with low cytotoxicity. We investigated the stimulatory effects of our scaffold on the differentiation of ADSCs in vitro and the regenerative effect of the CCSA delivery system on degenerated NP in vivo. Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

A novel smart injectable hydrogel prepared by microbial transglutaminase and human-like collagen: Its characterization and biocompatibility.

PubMed

Zhao, Leilei; Li, Xian; Zhao, Jiaqi; Ma, Saijian; Ma, Xiaoxuan; Fan, Daidi; Zhu, Chenhui; Liu, Yannan

2016-11-01

Various tissue scaffold materials are increasingly used to repair skin defects by cross-linking because of the ability to fill and implant in any form via operation. However, crosslinker residues cannot be easily removed from scaffold materials prepared by chemical crosslinking methods, limiting their use for skin tissue engineering. Here, microbial transglutaminase (MTGase), a nontoxic crosslinker with high specific activity and reaction rate under mild conditions, was employed crosslinks in human-like collagen (HLC) to yield novel smart MTGase crosslinked with human-like collagen (MTGH) hydrogels, which are sensitive to temperature and/or enzymes. Various ratios of MTGase/HLC were performed, and their physicochemical properties were characterized, including the swelling ratio, the elastic modulus, the morphology and the porosity. The degradation behavior and mechanism of MTGase in concentration-dependent manner involved in formation hydrogels were identifying in vitro. The cell attachment in vitro and biocompatibility in vivo were also investigated. The results demonstrated that the use of different concentrations of MTGase to crosslink HLC produced products with different degradation times and biocompatibilities. The 50U/g MTGase-prepared MTGH hydrogels had a higher density of crosslinks, which made them more resistant to degradation by collagenase I and collagenase II. However, 40U/g MTGase-prepared MTGH hydrogels were more suitable for cell attachment. In addition, compared with the Collagen Implant I® (SUM) used in animal experiments, the 40U/g MTGase-prepared MTGH hydrogels had a lower toxicity and better biocompatibility. Therefore, 40U/g MTGase crosslinked with HLC should be used to prepare MTGH hydrogels for potential application as soft materials for skin tissue engineering. Copyright © 2016 Elsevier B.V. All rights reserved.

Chemical cross-linking of the urease complex from Helicobacter pylori and analysis by Fourier transform ion cyclotron resonance mass spectrometry and molecular modeling

NASA Astrophysics Data System (ADS)

Carlsohn, Elisabet; Ångström, Jonas; Emmett, Mark R.; Marshall, Alan G.; Nilsson, Carol L.

2004-05-01

Chemical cross-linking of proteins is a well-established method for structural mapping of small protein complexes. When combined with mass spectrometry, cross-linking can reveal protein topology and identify contact sites between the peptide surfaces. When applied to surface-exposed proteins from pathogenic organisms, the method can reveal structural details that are useful in vaccine design. In order to investigate the possibilities of applying cross-linking on larger protein complexes, we selected the urease enzyme from Helicobacter pylori as a model. This membrane-associated protein complex consists of two subunits: [alpha] (26.5 kDa) and [beta] (61.7 kDa). Three ([alpha][beta]) heterodimers form a trimeric ([alpha][beta])3 assembly which further associates into a unique dodecameric 1.1 MDa complex composed of four ([alpha][beta])3 units. Cross-linked peptides from trypsin-digested urease complex were analyzed by Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) and molecular modeling. Two potential cross-linked peptides (present in the cross-linked sample but undetectable in [alpha], [beta], and native complex) were assigned. Molecular modeling of urease [alpha][beta] complex and trimeric urease units ([alpha][beta])3 revealed a linkage site between the [alpha]-subunit and the [beta]-subunit, and an internal cross-linkage in the [beta]-subunit.

Mapping of contact sites in complex formation between light-activated rhodopsin and transducin by covalent crosslinking: Use of a chemically preactivated reagent

PubMed Central

Itoh, Yoshiki; Cai, Kewen; Khorana, H. Gobind

2001-01-01

Contact sites in interaction between light-activated rhodopsin and transducin (T) have been investigated by using a chemically preactivated crosslinking reagent, N-succinimidyl 3-(2-pyridyldithio)propionate. The 3 propionyl-N-succinimidyl group in the reagent was attached by a disulfide exchange reaction to rhodopsin mutants containing single reactive cysteine groups in the cytoplasmic loops. Complex formation between the derivatized rhodopsin mutants and T was carried out by illumination at λ > 495 nm. Subsequent increase in pH (from 6 to 7.5 or higher) of the complex resulted in crosslinking of rhodopsin to the Tα subunit. Crosslinking to Tα was demonstrated for the rhodopsin mutants K141C, S240C, and K248C, and the crosslinked sites in Tα were identified for the rhodopsin mutant S240C. The peptides carrying the crosslinking moiety were isolated from the trypsin-digested peptide mixture, and their identification was carried out by matrix-assisted laser desorption ionization–time of flight mass spectrometry. The main site of crosslinking is within the peptide sequence, Leu-19–Arg-28 at the N-terminal region of Tα. The total results show that both the N and the C termini of Tα are in close vicinity to the third cytoplasmic loop of rhodopsin in the complex between rhodopsin and T. PMID:11320238

The effect of riboflavin/UVA cross-linking on anti-degeneration and promoting angiogenic capability of decellularized liver matrix.

PubMed

Xiang, Junxi; Liu, Peng; Zheng, Xinglong; Dong, Dinghui; Fan, Shujuan; Dong, Jian; Zhang, Xufeng; Liu, Xuemin; Wang, Bo; Lv, Yi

2017-10-01

Weak mechanical property and unstable degradation rate limited the application of decellularized liver matrix in tissue engineering. The aim of this study was to explore a new method for improving the mechanical properties, anti-degeneration and angiogenic capability of decellularized liver matrix. This was achieved by a novel approach using riboflavin/ultraviolet A treatment to induce collagen cross-linking of decellularized matrix. Histological staining and scanning electron microscope showed that the diameter of cross-linked fibers significantly increased compared with the control group. The average peak load and Young's modulus of decellularized matrix were obviously improved after cross-linking. Then we implanted the modified matrix into the rat hepatic injury model to test the anti-degeneration and angiogenic capability of riboflavin/UVA cross-linked decellularized liver scaffolds in vivo. The results indicated that cross-linked scaffolds degrade more slowly than those in the control group. In the experiment group, average microvessel density in the implanted matrix was higher than that in the control group since the first week after implantation. In conclusion, we initiated the method to improve the biomechanical properties of decellularized liver scaffolds by riboflavin/UVA cross-linking, and more importantly, its improvement on anti-degeneration and angiogenesis was identified. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2662-2669, 2017. © 2017 Wiley Periodicals, Inc.

Reflective article having a sacrificial cathodic layer

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

Kabagambe, Benjamin; Buchanan, Michael J.; Scott, Matthew S.

The present invention relates to reflective articles, such as solar mirrors, that include a sacrificial cathodic layer. The reflective article, more particularly includes a substrate, such as glass, having a multi-layered coating thereon that includes a lead-free sacrificial cathodic layer. The sacrificial cathodic layer includes at least one transition metal, such as a particulate transition metal, which can be in the form of flakes (e.g., zinc flakes). The sacrificial cathodic layer can include an inorganic matrix formed from one or more organo-titanates. Alternatively, the sacrificial cathodic layer can include an organic polymer matrix (e.g., a crosslinked organic polymer matrix formedmore » from an organic polymer and an aminoplast crosslinking agent). The reflective article also includes an outer organic polymer coating, that can be electrodeposited over the sacrificial cathodic layer.« less

Crystallization behaviour of nanostructured hybrid SiO2-TiO2 gel glasses to nanocomposites.

PubMed

Tsvetelina, Gerganova; Yordanka, Ivanova; Yuliya, Vueva; Miranda, Salvado Isabel M; Helena, Fernandes Maria

2010-04-01

The crystallization behaviour of hybrid SiO2-TiO2 nanocomposites derived from titanosiloxanes by sol-gel method has been investigated depending on the type of siloxane precursor and the pirolysis temperature. The resulting hybrid titanosiloxanes, crosslinked with trimethylsilil isocyanate (nitrogen-modified) or methyltrietoxisilane (carbon-modified), were pirolyzed in an inert atmosphere in the temperature range between 600 to 1100 degrees C in order to form C-(N)-Si-O-TiO2 nanocomposites. By means of XRD, FTIR, 29Si NMR, SEM, TEM and AFM investigations have been established that the transformation of the nanostructured SiO2-TiO2 hybrid materials into nanocomposites as well as the crystalline size depend on the titanium content and the type of cross-linking agents used in the synthesizes.

Solving the problem of building models of crosslinked polymers: an example focussing on validation of the properties of crosslinked epoxy resins.

PubMed

Hall, Stephen A; Howlin, Brendan J; Hamerton, Ian; Baidak, Alex; Billaud, Claude; Ward, Steven

2012-01-01

The construction of molecular models of crosslinked polymers is an area of some difficulty and considerable interest. We report here a new method of constructing these models and validate the method by modelling three epoxy systems based on the epoxy monomers bisphenol F diglycidyl ether (BFDGE) and triglycidyl-p-amino phenol (TGAP) with the curing agent diamino diphenyl sulphone (DDS). The main emphasis of the work concerns the improvement of the techniques for the molecular simulation of these epoxies and specific attention is paid towards model construction techniques, including automated model building and prediction of glass transition temperatures (T(g)). Typical models comprise some 4200-4600 atoms (ca. 120-130 monomers). In a parallel empirical study, these systems have been cast, cured and analysed by dynamic mechanical thermal analysis (DMTA) to measure T(g). Results for the three epoxy systems yield good agreement with experimental T(g) ranges of 200-220°C, 270-285°C and 285-290°C with corresponding simulated ranges of 210-230°C, 250-300°C, and 250-300°C respectively.

Solving the Problem of Building Models of Crosslinked Polymers: An Example Focussing on Validation of the Properties of Crosslinked Epoxy Resins

PubMed Central

Hall, Stephen A.; Howlin, Brendan J; Hamerton, Ian; Baidak, Alex; Billaud, Claude; Ward, Steven

2012-01-01

The construction of molecular models of crosslinked polymers is an area of some difficulty and considerable interest. We report here a new method of constructing these models and validate the method by modelling three epoxy systems based on the epoxy monomers bisphenol F diglycidyl ether (BFDGE) and triglycidyl-p-amino phenol (TGAP) with the curing agent diamino diphenyl sulphone (DDS). The main emphasis of the work concerns the improvement of the techniques for the molecular simulation of these epoxies and specific attention is paid towards model construction techniques, including automated model building and prediction of glass transition temperatures (Tg). Typical models comprise some 4200–4600 atoms (ca. 120–130 monomers). In a parallel empirical study, these systems have been cast, cured and analysed by dynamic mechanical thermal analysis (DMTA) to measure Tg. Results for the three epoxy systems yield good agreement with experimental Tg ranges of 200–220°C, 270–285°C and 285–290°C with corresponding simulated ranges of 210–230°C, 250–300°C, and 250–300°C respectively. PMID:22916182

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

NASA Astrophysics Data System (ADS)

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

2018-03-01

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