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Query: UNIPROT:P30536 (PBS)
 9,886 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Binding interactions between low molecular weight heparin (LMWH) and heparin-binding peptides (HBP) have been applied as a strategy for the assembly of hydrogels that are capable of sequestering growth factors and delivering them in a controlled manner. In this work, the assembly of four-arm star poly(ethylene glycol) (PEG)-LMWH conjugate with PEG-HBP conjugates has been investigated. The interactions between LMWH and the heparin-binding regions of antithrombin III (ATIII) or the heparin interacting protein (HIP) have been characterized via heparin affinity chromatography and surface plasmon resonance (SPR); results indicate that the two peptides have slightly different affinities for heparin and LMWH, and bind LMWH with micromolar affinity. Solutions of the PEG-LMWH and of mixtures of the PEG-LMWH and PEG-HBP were characterized via both bulk rheology and laser tweezer microrheology. Interestingly, solutions of PEG-LMWH (2.5 wt % in PBS) form hydrogels in the absence of PEG-ATIII or PEG-HIP, with storage moduli, determined via bulk rheological measurements, in excess of the loss moduli over frequencies of 0.1-100 Hz. The addition of PEG-ATIII or PEG-HIP increases the moduli in direct proportion to the number of cross-links introduced. Characterization of the hydrogels via microrheology shows the gel microstructure is composed of polymer-rich fibrillar structures surrounded by polymer-depleted buffer. Potential applications of these hydrogels are discussed.
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PMID:Rheological characterization of polysaccharide-poly(ethylene glycol) star copolymer hydrogels. 1600 30

Lipid-based delivery systems are becoming increasingly popular as carriers of drugs due to their ability to overcome barriers to oral absorption. The purpose of this study was to prepare novel lipid-based formulations of a model drug, piroxicam (PXCM), a poorly water-soluble non-steroidal anti-inflammatory drug (NSAID) using 1,2-dimyristoyl-sn-glycero-3-phosphatidylcholine (DMPC) phospholipid alone, and in combination with polyethylene glycol (PEG 4600). Lipid-based drug delivery systems were prepared using conventional methods of preparation and the following aspects were evaluated (1) in vitro dissolution behavior, (2) absorption via Caco-2 cell monolayers and (3) stability of formulations over a 12-month period. In addition, physical characterization studies using differential scanning calorimetry (DSC) were also performed. Formulations of PXCM were prepared using DMPC in the following combinations (A) 1:1 and (B) 2:1 and a mixture of DMPC and PEG 4600 (C) 2:1:1, respectively. Dissolution studies conducted in phosphate buffered saline (PBS, pH 7.4, 37+/-0.5 degrees C) using the USP type II (paddle) dissolution apparatus showed an increase in dissolution rate and extent of the PXCM from all solid dispersion formulations when compared to the control. As such, the rate of drug release was observed to be fastest with formulation (C) showing the greatest increase of over two-fold compared to the control. Release of PXCM from formulations (A) and (B) was intermediate with the latter showing superior dissolution behavior despite containing lower amounts of the carrier lipid than the former. This observation indicates a possible existence of threshold levels for phospholipids carriers beyond which dissolution could be adversely affected. DSC studies further confirmed the dissolution behavior of these formulations demonstrating different levels of amorphous to crystalline nature. Results of HPLC analysis from Caco-2 cell culture studies showed increase in transport of PXCM from all formulations, with formulation (C) showing the maximum increase followed by formulations (B) and (A), when compared to control. The apparent permeability coefficients (Papp) were calculated to be 7.92x10(-6), 9.48x10(-6), 9.2x10(-6) and 5.6x10(-6)cm/s for formulations (A)-(C) and control, respectively. Overall, permeation appeared to improve for all formulations over the control. Stability studies at various temperatures showed all formulations to have good stability for the first 6 months; then a decline in dissolution rates was observed, especially for PEG-based lipid carrier systems, attributed to the increase in crystalline content of the solid dispersions upon storage.
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PMID:Novel lipid-based formulations enhancing the in vitro dissolution and permeability characteristics of a poorly water-soluble model drug, piroxicam. 1604 87

The objective was to improve the protocol that was used to obtain the first reported piglets from transferred vitrified and warmed zona-intact blastocysts. Blastocysts were collected from superovulated sows and gilts, centrifuged to polarize lipid, vitrified, warmed and cultured for 24h or transferred immediately. Removing the zona pellucida after warming increased the number of cells in the surviving blastocysts (zona-free 60.8+/-4.3, zona-intact 39.1+/-2.8; P<0.05). Thinning the zona pellucida produced similar results to zona removal. Changing the basal medium of the vitrification and warming solutions from modified PBS to phosphate buffered NCSU-23 increased the number of cells (44.7+/-2.2 versus 56.0+/-3.9, respectively; P<0.05). Reducing the plunge temperature of the liquid nitrogen from -196 degrees C to less than -204 degrees C improved the embryo survival rate (61.9% versus 82.9%, respectively; P<0.05). These modifications were incorporated into the vitrification protocol that was used to vitrify and warm 105 blastocysts (that were subsequently transferred into four recipients). Three recipients became pregnant, farrowing three litters (average litter size, 5.3; 18.8% embryo survival in farrowing sows). Changing the warming protocol to using sucrose rather than ethylene glycol resulted in a trend towards improved embryo survival (73.5% versus 91.2%) but this was not statistically significant. Incorporating this modification, 203 blastocysts were vitrified, warmed and transferred into seven recipients. Five became pregnant and 36 fetuses were recovered (average litter size 7.2; 24.8% embryo survival in pregnant sows) at Day 40 of pregnancy. In conclusion, changes made to the vitrification protocol improved pregnancy rate and in vivo embryo survival compared to an earlier study using the original protocol.
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PMID:Changes to porcine blastocyst vitrification methods and improved litter size after transfer. 1605 93

Heparin employed in extracorporeal blood circulation (ECBC) procedures (e.g. open heart operations) often leads to a high incidence of bleeding complications. Protamine employed in heparin neutralization, on the other hand, can cause severe adverse reactions. We previously developed an approach that could prevent both heparin- and protamine-induced toxic side effects concomitantly. This approach consisted of placing a hollow fiber-based bioreactor device containing immobilized protamine (termed a "protamine bioreactor") at the distal end of the ECBC procedure. This protamine bioreactor would remove heparin after heparin served its anticoagulant purpose in the ECBC device, thereby eliminating heparin-induced bleeding risks. In addition, this protamine bioreactor would prevent protamine from entering the patients, thereby aborting any protamine-induced toxic effects. Both in vitro and in vivo studies have successfully demonstrated the feasibility of this approach. Despite promises, early findings also revealed two shortcomings that must be overcome for the protamine bioreactor to be applied clinically. The first drawback was that the cyanate ester linkages, involved in conjugating protamine to the bioreactor device, were unstable and prone to hydrolysis, resulting in the leakage of a significant amount of protamine into circulation during application of the protamine bioreactor. The second deficiency was that the capacity of the protamine bioreactor in heparin removal was rather low, owing to the limited surface area of the hollow fibers for protamine immobilization and subsequently heparin adsorption. In this paper, we present novel strategies to overcome these two limitations. A new conjugation method based on the use of 4-(oxyacetyl)phenoxyacetic acid (OAPA) as the activating reagent was employed to yield stable linkages, via the abundant arginine residues of protamine, onto the hollow fibers. Results showed that while the amount of protamine immobilized on each gram of fibers was relatively comparable between the OAPA and the previous CNBr activation methods (7.45 mg/g versus 7.69 mg/g fibers), there was virtually no detectable leaching of immobilized protamine from the bioreactor by the OAPA method, comparing to 35% leaching of protamine by the previous CNBr method following 72 h of storage of the bioreactor in PBS buffer at 37 degrees C. To improve the capacity and functionality of the protamine bioreactor, two novel approaches were adopted. Long chain and high molecular weight poly-lysine was linked to the hollow fibers, prior to protamine coupling, to create multiple layers of immobilized protamine for subsequent heparin adsorption. In addition, a poly(ethylene glycol) (PEG) chain was inserted between protamine and the hollow fibers to yield a three-dimensional, free dynamic motion for immobilized protamine. Preliminary observations indicated that a four- to five-fold enhancement in heparin adsorption was attained by utilizing each of these new approaches. Aside from their current use, these new strategies can also be employed generically to improve the functionality of any affinity-type bioreactor. Indeed, efforts have been made recently in utilizing these approaches to develop a clinically usable GPIIb/IIIa bioreactor for the treatment of immune thrombocytopenic purpura (ITP)-an autoimmune disease.
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PMID:Strategies for improving the functionality of an affinity bioreactor. 1624 11

Chondroitin sulfate (CS) is a potential candidate for colon-specific drug carriers. However, the readily water-soluble nature limits its application as a solid-state drug-delivery vehicle. In this study, the CS formation of a polyelectrolyte complex (PEC) with Ca2+ (CS-Ca) was adapted to retain CS in a solid form for use in a drug-delivery system. Pre-treated CS with poly(ethylene glycol) diglycidyl ether (EX-810) followed by complexation with Ca2+ was also tested (CS-Ca-EX). Diclofenac sodium was used as a drug probe to evaluate the performance of the drug-release behavior of the complexes. The amount of diclofenac sodium released was higher in simulated intestinal fluid (SIF) than in simulated gastric fluid (SGF) due to the anionic groups on CS or the higher solubility of drug itself in PBS. The release profile of diclofenac sodium from CS-Ca-EX was most notably sustained when compared to other groups. Enzymatic degradation by chondroitinase ABC of CS, CS-Ca and CS-Ca-EX exhibited a similar degradation mechanism and GPC revealed the dissolution rate of CS from the three matrix types was, in decreasing order: CS, CS-Ca, CS-Ca-EX. The synergy of the anti-inflammatory activity of diclofenac sodium in CS-based complexes was evaluated using the carrageenan-induced edema rat test. The percentage of swelling was lower for all experimental groups as compared to the control, untreated group. The anti-inflammatory activity of diclofenac in the CS matrix gradually increased up to 9 h but CS-Ca or CS-Ca-EX matrices showed less potency than the CS matrix in reducing inflammation.
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PMID:Oral sustained delivery of diclofenac sodium using calcium chondroitin sulfate matrix. 1626 56

PEGylated polyamidoamine (PAMAM) dendrimers as drug carriers have been a topic of interest because of their biomedically favorable features, including minimal toxicity, reduced immunogenicity, and excellent solubility in aqueous and most organic solutions. A PEG shell on dendrimer surface may provide steric hindrance, known as stealth properties of PEG, to stabilize drug molecules to be delivered. In this article, the effects of PEG and coupling sequence of drug, PEG, and dendrimer in modulating the stability of delivered drug molecules were evaluated. N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide was chosen as a model peptide. Dendritic peptides, that is, peptide-dendrimer, peptide-PAMAM-PEG, and peptide-PEG-dendrimer, were constructed based on Starbursttrade mark G3.0 PAMAM dendrimer and characterized by (1)H-NMR spectroscopy. Hydrolysis of dendritic peptides was catalyzed by alpha-chymotrypsin in pH 7.4 PBS buffer containing 5% DMF (v/v) at room temperature. The enzymatic stability of dendritic peptides was peptide-PAMAM-PEG > peptide-PAMAM > free peptide > peptide-PEG-PAMAM. The ratio of PEG/peptide could be reduced for increasing peptide loading while maintaining the delivered peptides' relatively high enzymatic stability. The quantitative analysis of dendritic peptide/enzyme interactions provided the understandings of the molecular structure/stability relationships of dendrimer/drug for the design of an optimal PEGylated dendrimer-based drug-delivery system.
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PMID:In vitro enzymatic stability of dendritic peptides. 1627 Mar 46

The anthelmintic properties of tanniferous plants and of their secondary metabolites represent one possible alternative to chemotherapy that is currently being explored as a means of achieving sustainable control of gastrointestinal nematodes in ruminants. Previous in vivo and in vitro results suggest that tanniferous plants can have direct anti-parasitic effect against different stages of nematodes. However, the mode of action of the bioactive plant compounds remains obscure. The objectives of the current study were (1) to examine the hypothesis that extracts of tanniferous plants might interfere with the exsheathment of third-stage infective larvae (L3); (2) to assess the role of tannins in the process by examining the consequence of adding an inhibitor of tannins (polyethylene glycol: PEG) to extracts. The effects of 4 tanniferous plant extracts on exsheathment have been examined on L3 of Haemonchus contortus and Trichostrongylus colubriformis. Artificial exsheathment was induced in vitro by adding hypochloride solution to larval suspension. The evolution of exsheathment with time was measured by repeated observations at 10-min interval for 60 min. The selected plants were: genista (Sarothamnus scoparius), heather (Erica erigena), pine tree (Pinus sylvestris), and chestnut tree (Castanea sativa), with tannin contents ranging from 1.5 to 24.7% of DM. Extracts of a non-tanniferous plant (rye grass, tannin content: 0.3% of DM) were included in the assay as negative controls. The extracts were tested at the concentration of 600 microg/ml and the effects were compared to the rate of exsheathment of control larvae in PBS. No statistical differences in the pattern of exsheathment was observed after addition of rye grass or genista extracts for both nematode species and with heather extracts for T. colubriformis. In contrast, pine tree extracts on larvae of both species and heather extracts with H. contortus induced a significant delay in exsheathment. Last, contact with chest nut extracts led to a total inhibition of the process for both nematodes. These results suggest that extracts of tanniferous plants might affect a key process in the very early stages of larval invasion of the host. In most cases, the addition of PEG led to a total or partial restoration towards control values. This suggests that tannins are largely involved in the inhibitory process. However, other secondary metabolites may also interfere with the process that would help to explain some of the differences in response observed between the two nematode species.
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PMID:Effects of four tanniferous plant extracts on the in vitro exsheathment of third-stage larvae of parasitic nematodes. 1638 90

Thermosensitive poly(organophosphazenes) bearing hydrophobic isoleucine ethyl esters group and hydrophilic alpha-amino-omega-methoxy-poly(ethylene glycol) of the molecular weight 550 along with hydrolysis-sensitive glycyl lactate ethyl esters have been synthesized for sustained delivery of anticancer drug. The aqueous solution of poly(organophosphazenes) containing doxorubicin, that represents chemotherapeutic agent for cancer treatment, was transformed into hydrogel with good gel strength at body temperature via hydrophobic interactions. Solubility of hydrophobic doxorubicin in the aqueous poly(organophosphazene) solution was dramatically improved as compared with that in PBS (0.01 M, pH 7.4). The hydrogel property of poly(organophosphazenes) was affected on incorporation of doxorubicin, resulting in increase of gelation temperature and decrease of gel strength. The release of loaded doxorubicin from the polymer hydrogel was significantly sustained over 20 days and the effect of gel strength, polymer concentration and drug concentration on the release rate were observed. The release of doxorubicin from the polymer gels was effectively controlled by the gel strength. As a result of investigating anticancer efficacy using cancer cell line of mouse lymphoblast of P388D1, the efficacy of doxorubicin was observed to be constant over a prolonged period of times for more than 30 days, indicating that the release of doxorubicin was sustained for a long time without any initial burst release, and that the delivery system was an excellent candidate for locally injectable gel-depot system.
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PMID:Controlled release of doxorubicin from thermosensitive poly(organophosphazene) hydrogels. 1667 86

The influence of equilibration time before vitrification on the viability of vitrified morula- to blastocyst-stage bovine embryos and in vivo viability of vitrified embryos following transfer to recipients were investigated. In experiment 1, the embryos were exposed to an equilibration solution (50% VSED) containing 12.5% v/v ethylene glycol and 12.5% v/v dimethyl sulfoxide in modified Dulbecco's phosphate buffered saline with 4 mg/ml BSA (m-PBS) for 1, 2 and 5 minutes at room temperature (22 to 24 degrees C). The embryos were then placed in 15mul vitrification solution (VSED) consisting of 25% v/v ethylene glycol and 25% v/v dimethyl sulfoxide in m-PBS and were loaded into 0.25 ml plastic straws at room temperature. After 30 seconds, the straws were placed in liquid nitrogen (LN(2)) vapor for 2 minutes, plunged and stored in LN(2). To thaw, the straws were warmed in water at 20 degrees C for 15 seconds and the contents of the straws were expelled into a plastic dish. The embryos were diluted in 0.5 M sucrose + m-PBS for 5 minutes and were cultured in TCM-199 supplemented with bovine oviductal epithelial tissue. Viability of the embryos was assessed by the forming or reforming of the blastocoele after 24 hours of culture. High in vitro survival rates (73 approximately 90%) of vitrified embryos were obtained after 1 and 2 minute equilibrations, but was reduced (P<0.05) after 5 minute equilibration. In Experiment 2, morula- to blastocyst-stage embryos were vitrified after 1 minute equilibration in 50% VSED and 30 seconds of exposure to VSED. The vitrified-warmed embryos were transferred to recipient heifers at 7 days after estrus (1 embryo per recipient). Five (38%) of 13 (40%) of 10 recipients that had received blastocysts were diagnosed as pregnant using ultrasonography 60 days following transfer.
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PMID:Vitrification of bovine embryos in a mixture of ethylene glycol and dimethyl sulfoxide. 1672 27

The aim of the present study was to compare the survival rates of goat morulae and blastocysts after different freezing procedures. The viability of frozen-thawed embryos was assessed both in vivo and in vitro. Two cryoprotectants, ethylene glycol and glycerol, were used and three cryoprotectant removal procedures were compared: progressive dilution in 1.0, 0.5, 0.3 and 0 M of cryoprotectant in PBS; a similar progressive dilution with cryoprotectant in PBS plus 0.25 M of sucrose; or one-step transfer in PBS containing 0.25 M of sucrose. In vitro development of frozen-thawed blastocysts was always higher than that of frozen morulae irrespective of the cryoprotectant (52 129 = 40.3% vs 23 161 = 14.3% ; P< 0.001). In vivo, however, frozen-thawed morulae developed equally as well as blastocysts after an identical freezing-thawing protocol. Development both in vivo and in vitro showed ethylene glycol to be a better cryoprotectant than glycerol for goat embryos at both developmental stages (23 vs 0%, 45 vs 35% in vitro; 34.5 vs 21%, 35 vs 23% in vivo for morulae and blastocysts, respectively).
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PMID:In vivo and in vitro survival of goat embryos after freezing with ethylene glycol or glycerol. 1672 58


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