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PubMed1984

Characterization of genomic poly(dT-dG).poly(dC-dA) sequences: structure, organization, and conformation.

Hybridization studies suggest the abundant presence of poly(dT-dG).poly(dC-dA) (TG-element), a potential Z-DNA sequence, in eucaryotic genomes. We have isolated and characterized TG-elements from different locations in the human genome: from randomly isolated clones, associated with the actin gene family, and linked to another repeated element. The results indicate that the following features are typical of these TG-elements: the elements consist of 20 to 60 base pairs of (dT-dG)n.(dC-dA)n, the sequences characterized in our study were not flanked by direct or inverted repeats, the sequences are interspersed rather than in satellite blocks, the elements are not usually associated with other repeated elements, and some of the elements are found near coding sequences or in introns. Studies on the conformation of a genomic TG-element in a supercoiled plasmid indicate several distinct properties of the TG-element: it is in the Z-form only at low ionic strength, S1 nuclease recognizes its Z-form with a marked preference for one of the B-Z junctions, and the sensitive region extends for 20 base pairs near the B-Z junction. In contrast to the result with the supercoiled plasmid, S1 nuclease failed to recognize the TG-element in minichromosomes.

Actins↗
PubMed1984

Enhanced gene expression by the poly(dT-dG).poly(dC-dA) sequence.

The sequence poly(dT-dG).poly(dC-dA) (TG-element) is a ubiquitous component of eucaryotic genomes and has the potential to adopt a left-handed DNA conformation (Z-DNA). In this report, we have tested the hypothesis that the TG-element can modulate gene expression. Human genomic DNA fragments (1 to 1.5 kilobases) containing a (dT-dG)n.(dC-dA)n tract (30, 40, or 50 base pairs) or chemically synthesized (dT-dG)n.(dC-dA)n fragments (50 to 130 base pairs) were inserted in the pSV2-cat (simian virus 40 enhancer plus) or pA10-cat (enhancer minus) expression vector plasmid. These constructs were transfected into CV-1 cells or HeLa cells, and their transcription was monitored by assaying chloramphenicol acetyltransferase activity. The results showed that pSV2-cat with the TG-element and pA10-cat with the TG-element synthesized more chloramphenicol acetyltransferase activity (2 to 10 times, depending on the location of the TG-element) than did parental pSV2-cat and pA10-cat DNAs, respectively. Furthermore, the TG-element appeared to have characteristics similar to those of viral enhancers: (i) the TG-element enhanced transcription from a distance, (ii) its closer location to the promoter was more effective, and (iii) its orientation was not crucial. However, its enhancer-like activity was much weaker than that of the simian virus 40 enhancer, and, unlike many viral enhancers, it was equally active in monkey and in human cells. These results suggest that the TG-element may influence the expression of cellular genes.

Acetyltransferases↗
PubMed1998

Cloning of the Nocardia corallina polyhydroxyalkanoate synthase gene and production of poly-(3-hydroxybutyrate-co-3-hydroxyhexanoate) and poly-(3-hydroxyvalerate-co-3-hydroxyheptanoate).

The polyhydroxyalkanoate (PHA) synthase gene (phaCNc) from Nocardia corallina was identified in a lambda library on a 6-kb BamHI fragment. A 2.8-kb XhoII subfragment was found to contain the intact PHA synthase. This 2.8-kb fragment was subjected to DNA sequencing and was found to contain the coding region for the PHA synthase and a small downstream open reading frame of unknown function. On the basis of DNA sequence, phaCNc is closest in homology to the PHA synthases (phaCPaI and phaCPaII) of Pseudomonas aeruginosa (approximately 41% identity and 55% similarity). The 2.8-kb XhoII fragment containing phaCNc was subcloned into broad host range mobilizable plasmids and transferred into Escherichia coli, Klebsiella aerogenes (both containing a plasmid bearing phaA and phaB from Ralstonia eutropha), and PHA-negative strains of R. eutropha and Pseudomonas putida. The recombinant strains were grown on various carbon sources and the resulting polymers were analyzed. In these strains, the PHA synthase from N. corallina was able to mediate the production of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) containing high levels of 3-hydroxyhexanoate when grown on hexanoate and larger even-chain fatty acids and poly(3-hydroxyvalerate-co-3-hydroxyheptanoate) containing high levels of 3-hydroxyheptanoate when grown on heptanoate or larger odd-chain fatty acids.

Acyltransferases↗
PubMed2004

Water-dispersed lamellar phases of symmetric poly(styrene)-block-poly(acrylic acid) diblock copolymers: model systems for flat dense polyelectrolyte brushes.

We investigate the static properties of a water-dispersed lamellar ( L) phase formed in the melt state with a nearly symmetric poly(styrene)-block-poly(acrylic acid) (PS- b-PAA) diblock copolymer. The PAA brush is considered as a model flat polyelectrolyte ( PE) brush of controlled surface density. Thanks to small-angle X-ray scattering, its behavior in water is studied as a function of (i) its ionization, through the pH of the dispersions which is increased by an addition of a known amount of a base, i.e. sodium hydroxyde NaOH, and (ii) in the presence of a monovalent salt, i.e. sodium chloride NaCl, of concentration C(S). At low pH, we find that the brush effectively behaves as a neutral brush. At high pH, the brush is in the so-called "osmotic regime", in which all sodium counterions are trapped within the brush volume and stretch the chains via an osmotic effect. The properties of such a brush in the presence of a monovalent salt, confirm this result, showing a C(S)(-1/3) dependence in the brush height L(O), in agreement with mean-field predictions. The L(O)- C(S) profiles at different ionizations give access to the actual brush internal charge fraction f. The results are found to be in very good quantitative agreement with experimental measures found in the literature, and can be completely and quantitatively described by Oosawa's approach to counterion condensation in a semi-dilute to concentrated solution of charged, rod-like chains.

Acrylic Resins↗
PubMed1995

Inflammation: imaging with methoxy poly(ethylene glycol)-poly-L-lysine-DTPA, a long-circulating graft copolymer.

PURPOSE: To test whether a nontargeted, long-circulating, synthetic polymer accumulates in areas of inflammation, with high capillary permeability and increased regional blood flow. MATERIALS AND METHODS: Methoxy poly(ethylene glycol)-poly-L-lysine (PL)-diethylenetriaminepentaacetic acid (MPEG-PL-DTPA) was labeled with technetium-99m for scintigraphy and with gadolinium for magnetic resonance (MR) imaging. Eleven Escherichia coli-infected rats were injected with 1.0 mCi (37 MBq) of Tc-99m-labeled MPEG-PL-DTPA for scintigraphy. Twelve rats underwent 1.5-T MR imaging after intravenous injection of gadolinium-labeled MPEG-PL-DTPA (35 mumol/kg). RESULTS: Tc-99m-labeled MPEG-PL-DTPA demonstrated nearly eight-fold higher accumulation in infected muscle when compared with normal muscle. Scintigrams and MR images showed areas of inflammation with peak accumulation at 24 hours after injection of Tc-99m- or gadolinium-labeled MPEG-PL-DTPA. CONCLUSION: Nontargeted, long-circulating, copolymers can efficiently accumulate in sites of inflammation and thus represent an alternative to inflammation-specific agents.

Animals↗
PubMed1997

Oral immunization with poly-(D,L-lactide-co-glycolide) and poly-(L-lactic acid) microspheres containing pneumotropic bacterial antigens.

Encouraged by recent findings showing the usefulness of nonreplicating antigen delivery systems in the induction ofmucosal immune responses, we investigated microspheres as a means to deliver LW 50020, an immunomodulator consisting of lysates of seven common respiratory pathogens. BALB/c mice were orally immunized with LW 50020 encapsulated into poly-(D,L-lactide-co-glycolide) (PLG) and poly-(L-lactic acid) (PLA) microspheres prepared by either a solvent-evaporation or a solvent-extraction double-emulsion technique. Particle uptake into intestinal Peyer's patches, induction of antibodies in sera and secretion of immunoglobulins by isolated Peyer's patches, lungs and spleen lymphocytes were investigated. Our results revealed size and surface characteristic-dependent uptake of microspheres into Peyer's patches. Microsphere translocation into Peyer's patches was efficient for 0.8-microm microspheres, but poor for 2.0-microm and surface-modified microspheres. We showed an enhanced immune response in the lungs and sera following oral immunization with 0.8-microm PLG solvent-evaporation microspheres. The immunomodulation was statistically significant as compared to free LW 50020. In contrast, oral immunization with other preparations caused reduced or absent modulation of the immune response compared to 0.8-microm microspheres and free antigen. These findings indicate that microspheres displaying small particle sizes, rapid antigen release and high antigen content provide optimal tools to deliver orally applied antigens.

Administration, Oral↗
PubMed2002

Polymeric complexes of lidocaine hydrochloride with poly(acrylic acid) and poly(2-hydroxyethyl vinyl ether).

The specific interactions of local anesthetic lidocaine hydrochloride with poly(acrylic acid) and poly(2-hydroxyethyl vinyl ether), as well as in a triple system composed of the drug and both polymers, have been studied in aqueous solutions by viscometric, turbidimetric, potentiometric, and FTIR spectroscopic methods. The mechanism of the drug binding to the polymers and the structures of the polycomplexes formed are clarified.

Acrylic Resins↗
PubMed2003

Synthesis and characterization of poly(L-lysine)-g-poly(D,L-lactic-co-glycolic acid) biodegradable micelles.

A series of amphipathic graft copolymers composed of poly(L-lysine) (PLL) as the cationic polymer backbone and biodegradable poly(D,L-lactic-co-glycolic acid) (PLGA) as the grafting chains were synthesized and characterized. The terminal group of PLGA was activated and chemically conjugated to the primary epsilon-amine groups of PLL to produce PLL-g-PLGA copolymers. PLL-g-PLGA formed a self-assembling micelle structure in aqueous solution. The micelle size ranged from 69.4 to 149.6 nm in diameter, depending on the grafting percentage of PLGA. Upon increasing the number of PLGA chains grafted onto the PLL backbone, the size of the micelles gradually decreased, at the same time lowering their critical micelle concentration. The micelles were individually separated and had a spherical geometry, as observed by atomic force microscopy (AFM). These PLL-g-PLGA copolymers can be applied as cell adhesive surface coating materials for biodegradable tissue engineering scaffolds and can be used as non-viral DNA carriers for gene therapy.

Biocompatible Materials↗
PubMed2004

The effect of pH on the LCST of poly(N-isopropylacrylamide) and poly(N-isopropylacrylamide-co-acrylic acid).

Poly(N-isopropylacrylamide) (PNIPAAm) and poly(N-isopropylacrylamide-co-acrylic acid) (P(NIPAAm-co-AAc)) hydrogels are synthesized by irradiating the aqueous solutions of NIPAAm and NIPAAm/AAc with 60Co gamma-ray. The effects of pH on the swelling ratio and on the lower critical solution temperature (LCST) are studied by determining the dependence of swelling ratio on temperature in different pH butter solutions. Differential scanning calorimetry (DSC) is applied in determination of the LCST of the hydrogels. Fourier transform infrared (FT-IR) spectrometry is used in the comparison of hydrogels swelled in various pH conditions. As a result, PNIPAAm was found to be a pH-sensitive hydrogel and the LCST of the PNIPAAm and P(NIPAAm-co-AAc) hydrogels are influenced by pH.

Acrylamides↗
PubMed2005

Semi-interpenetrating network of poly(ethylene glycol) and poly(D,L-lactide) for the controlled delivery of protein drugs.

We have prepared a semi-interpenetrating network (IPN) of poly(ethylene glycol) dimethacrylate (PEGDMA) with entrapped poly(D,L-lactide) (PLA) using photochemical techniques. These IPNs were developed for the controlled delivery of protein drugs such as growth factors. The PEG component draws water into the network, forming a hydrogel within the PLA matrix, controlling and facilitating release of the protein drug, while the PLA component both strengthens the PEG hydrogel and enhances the degradation and elimination of the network after the protein drug is released. The rate and extent of swelling and the resultant protein release kinetics could be controlled by varying the PEG/PLA ratio and total PLA content. These IPNs were prepared using a biocompatible benzyl benzoate/benzyl alcohol solvent system that yields a uniform, fine dispersion of the protein throughout the PEG/PLA IPN matrix. IPNs composed of high molecular mass PLA and lower PEG/PLA ratios exhibited lower equilibrium swelling ratios. The release of bovine serum albumin (BSA), a model protein, from these IPNs was characterized by a large initial burst, regardless of the PEG/PLA ratio, due to the entrapment of residual solvent within the network. Microparticles of the PEG/PLA IPNs were also prepared using a modified Prolease strategy. Residual solvent removal was significantly enhanced using this process. The microparticles also exhibited a significant reduction in the initial burst release of protein. Mixtures of different compositions of PEG/PLA microparticles should be useful for the delivery of a variety of protein drugs with different release kinetics from any tissue-engineering matrix.

Acrylates↗
PubMed2005

Poly(ethylene oxide)-b-poly(N-isopropylacrylamide) nanoparticles with cross-linked cores as drug carriers.

Micelle-like nanoparticles that could be used as drug-delivery carriers were developed. The unique feature of these nanoparticles was that the core of poly(ethylene oxide)-b-poly(N-isopropylacrylamide) (PEO-b-PNIPAAm) micelle was lightly cross-linked with a biodegradable cross-linker, N,N-bis(acryloyl)cystamine (BAC). The nanoparticles were characterized by dynamic light scattering and fluorescence measurements. When the BAC content ranged from 0.75 wt% to 0.2 wt% of the mass of NIPAAm, the diameters of the nanoparticles were less than 150 nm. The anti-cancer drug doxorubicin (Dox) and 1,6-diphenyl-1,3,5-hexatriene (DPH) were used as fluorescent probes to study the hydrophobicity of the cores of the nanoparticles; the results showed that the cores of the nanoparticles were hydrophobic enough to sequester Dox and DPH. The nanoparticles with 0.5 wt% BAC stored at room temperature were stable up to 2 weeks, even at dilute concentrations. The degradation of BAC by reducing agent beta-mercaptoethanol was investigated, and the nanoparticles were not detectable 14 days after adding beta-mercaptoethanol.

Acrylic Resins↗
PubMed2005

Fixation of distal femoral osteotomies with self-reinforced poly(L/DL)lactide 70:30 and self-reinforced poly(L/DL)lactide 70: 30/bioactive glass composite rods. an experimental study on rabbits.

Two self-reinforced poly(L/DL)lactide 70:30 or self-reinforced poly (L/DL)lactide 70:30/ bioactive glass (SR-P(L/DL)LA/bioactive glass) composite rods (2 mm x 40 mm) were implanted into the dorsal subcutaneous tissue and osteotomies of the distal femur were fixed with these rods (2 mm x 26 mm) in 36 rabbits. The follow-up times varied from 3 to 100 weeks. After the animals were killed, three-point bending and shear tests and molecular weight measurements were performed for subcutaneously placed rods. Radiological, histological, histomorphometrical, microradiographic and oxytetracycline-fluorescence studies of the osteotomized and intact control femora were performed. After 12 weeks the SR-P(L/DL)LA rods had fragmented into pieces and the mechanical properties could not be measured. The SR-P(L/DL)LA/bioactive glass rods lost their mechanical properties slower, and at 24 weeks the bending strength had decreased by 39% and the shear strength by 50%. After that the mechanical properties of the SR-P(L/DL)LA/bioactive glass rods could not be measured. All osteotomies healed well, and no gross signs of inflammatory reactions were observed. One slight displacement was seen in the three-week follow-up group with SR-P(L/DL)LA rods. Signs of resorption of the implants were seen after 48 weeks in the SR-P(L/DL)LA group and after 24 weeks in the SR-P(L/DL)LA/bioactive glass group. The SR-P(L/DL)LA/bioactive glass rods were almost totally resorbed from the bone at 100 weeks. The present investigation showed that the mechanical strength and fixation properties of the SR-P(L/DL)LA and the SR-P(L/DL)LA/bioactive glass composite rods are suitable for fixation of cancellous bone osteotomies in rabbits.

Animals↗
PubMed2006

Multiphase blends from poly(L-lactide) and poly(methyl mathacrylate).

Melt processing of poly(L-lactide) (PLLA) and poly(methyl methacrylate) (PMMA) was conducted over a targeted range of compositions with PLLAs of 118 and 316 kDa in molecular mass to identify morphologies and the phase relationships in these blends. These blends are of interest for use in biomaterials and the morphologies are critical for tissue-engineering studies where biodegradability, pore connectivity and surface texture control tissue viability and adhesion. Simple extrusion of the two polymers produced multiphase blends with an average domain size near 25 microm. Scanning electron microscopy and dynamic mechanical analysis demonstrated that these blends are immiscible, at least in a metastable sense, and regions of co-continuous structures were identified. Such co-continuous, which occurred generally in accordance with rheology prediction models, exhibit a fine interconnected structure that appears effective for fabricating certain biomaterials. A broad and unexpected transition appears in these blends, as measured by modulated differential scanning calorimetry between 70 and 100 degrees C, which may be the glass transition temperature of an alloy phase. The magnitude of this transition is greatest in the fine-structured co-continuous composition region of blends, suggesting the presence of a complex or other derivative of the two primary phases.

Animals↗
PubMed2006

Self-association and micelle formation of biodegradable poly(ethylene glycol)-poly(L-lactic acid) amphiphilic di-block co-polymers.

Di-block co-polymers of poly(ethylene glycol)-poly(L-lactic acid) (PEG-PLLA) were prepared by ring-opening polymerization, and their self-association and micelle formation were investigated. The block co-polymers have the same block length of the hydrophilic PEG segment (Mn = 2000), but different chain lengths of the hydrophobic PLLA segment (Mn = 700, 1000 and 1300, respectively). The di-block co-polymers synthesized were characterized by GPC, 1H-NMR, TGA and DSC. The critical micelle concentration (CMC) of the PEG-PLLA micelles was determined at various temperatures (5-45 degrees C) using a dye absorption technique involving fluorescence spectrophotometry with pyrene as a probe to monitor the change in the polarity of the microenvironment in the micelle. An increase in the molecular weight of the hydrophobic block decreases the CMC. The partition coefficients of pyrene between the micellar and aqueous phases range from 0.68 x 10(4 )to 1.76 x 10(4), depending on the PLLA content in the block co-polymers. An Arrhenius plot of ln(CMC) versus 1/T exhibited an almost constant CMC at low temperatures (<35 degrees C), followed by an increase of the CMC at higher temperatures (>35 degrees C). The increase of the CMC with temperature indicates the increase of the mobility of the hydrophobic PLLA core. These results were confirmed by 1H-NMR measurements. The micelle size and size distribution were determined by dynamic light scattering (DLS), and were in the range of 78-92 nm. A spherical micelle shape was confirmed by transmission electron microscopy. These results indicate that the CMC and the thermal characteristics of the core-forming segment of the block co-polymer play an important role in the properties of the polymer micelles used for drug delivery.

Biodegradation, Environmental↗
PubMed2006

Poly-alpha,beta-(N-(2-hydroxyethyl)-L-aspartamide)-g-poly(1,3trimethylene carbonate) amphiphilic graft co-polymer as a potential drug carrier.

A biodegradable amphiphilic graft polymer was successfully synthesized by grafting hydrophobic poly(1,3-trimethylene carbonate) (PTMC) sequences onto a hydrophilic poly-alpha,beta-(N-(2-hydroxyethyl)-L-aspartamide) (PHEA) backbone. The graft polymer, PHEA-g-PTMC, was synthesized by ring-opening polymerization initiated by the macroinitiator PHEA bearing hydroxyl groups without adding any catalyst. The graft polymer was characterized by Fourier transform infrared spectroscopy, 1H-nuclear magnetic resonance spectroscopy, combined size-exclusion chromatography and multiangle laser light scattering analysis. Two drugs with distinct water solubility, prednisone acetate and tegafur, were encapsulated in the PHEA-g-PTMC nanoparticles. The in vitro release of two drugs from PHEA-g-PTMC nanoparticle drug-delivery systems was investigated.

Biocompatible Materials↗
PubMed1997

Immobilization of functionalized alkyl-poly(ethylene oxide) surfactants on poly(ethylene) surfaces by means of an argon plasma treatment.

Alkyl-poly(ethylene oxide) (PEO) surfactants containing a terminal hydroxyl, sulfate, or carboxylate group were grafted at the surface of poly(ethylene) (PE) samples to improve their blood compatibility. Grafting was achieved by immobilizing PEO surfactants on PE using an argon plasma treatment. The sulfate group containing PEO surfactant was synthesized by sulfating polyoxyethylene(20)stearylether (Brij78; B) with chlorosulfonic acid. A carboxylate-terminated surfactant was synthesized by a substitution reaction of the sodium alkoxide form of B with sodium iodoacetate. XPS analysis of the modified PE samples showed that at short plasma treatment times of up to 5 s the structure of the immobilized surfactants is largely retained. When plasma treatment times longer than 30 s were applied, the PEO chains of the surfactants were degraded. The wettability of the modified PE samples was improved compared to the unmodified PE samples. The wettability of the modified samples did not change when they were stored in air at room temperature for at least 12 weeks.

Alkylation↗
PubMedDate not supplied

Characterizing poly(epsilon-caprolactone)-b-chitooligosaccharide-b-poly(ethylene glycol) (PCP) copolymer micelles for doxorubicin (DOX) delivery: effects of crosslinked of amine groups.

New amine-groups containing tri-block copolymers and micelles that consisting of poly(epsilon-caprolactone)-b-chitooligosaccharide-b-poly(ethylene glycol) (PCL-b-COS-b-PEG, PCP), were synthesized, characterized, and evaluated for delivering doxorubicin (DOX) with or without crosslinked amine groups by genipin. The characteristics of the PCP copolymers of Fourier-transform infrared spectrometry (FT-IR) verify the amine and ester groups of the COS and the PCL of the copolymers, respectively. 1H nuclear magnetic resonance (1H NMR) spectra verify the structures of the PCP copolymers consisting two PCL and PEG polymers reacted onto the COS block. In addition, gel permeation chromatography (GPC) determines the number average molecular weight of the tri-block copolymers (Mn) of approximately 11340 Da/mole. The PCP copolymers can self-assemble to form polymeric micelles at the critical micelle concentration (CMC) of 1.0 microM as determined by the UV-VIS absorption spectra. The mean diameter of the PCP micelles is 90 nm, as determined using a dynamic light-scattering (DLS) analyzer. Moreover, the zeta potentials of PCP micelles change from neutral to cationic state when pH of suspension mediums varied from 7.4 to 3.0. For evaluating delivery characteristics of hydrophobic DOX, it was loaded into PCP micelles with or without crosslinked by genipin. The burst release and release period of DOX for the crosslinked micelles are significantly reduced (P < 0.003, n = 3, for pH = 7.4) and sustained (e.g., 8 days), respectively, than those non-crosslinked ones (e.g., 4 days). In conclusion, new tri-block amine groups containing PCP copolymers are synthesized that can self-assemble as PCP micelles. After post-crosslinked amine groups of DOX loaded the micelles, they can effectively reduce the burst release and sustain the release of DOX at different pH dissolution mediums. Further applications of PCP copolymers and micelles for drug delivery can be explored in future.

Amines↗