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PubMed2002

Density and distribution of anionic sites on boar ejaculated and epididymal spermatozoa.

Current knowledge implies that spermatozoa successively acquire negative surface charges as they migrate through the epididymis. Until recently, however, techniques used were not amenable to statistical analysis. In the present study, a novel approach allowing numerical assessment of negative charge labelling was used in order to determine the density and distribution of anionic sites on ejaculated and maturing spermatozoa collected from six regions of the boar epididymis. Labelling was assessed quantitatively for the three morphologically distinct membrane domains on the sperm head. Statistical analysis revealed that labelling density was highest on efferent duct spermatozoa, declined up to the proximal corpus and then increased again. Densities of anionic sites on distal corpus, proximal cauda and ejaculated sperm cells were similar but significantly below the values obtained for efferent duct spermatozoa. All three sperm membrane domains underwent parallel changes. However, the overall density of negative charges on the postacrosomal segment was significantly higher as compared to the acrosomal plasma membrane. These alterations reflect sperm surface modifications through removal and addition of anionic groups. Since charge interactions are considered to play a pivotal role in sperm-egg interactions, these processes should be viewed as an integral part of sperm maturation.

Animals↗

PubMed2004

An air-molding technique for fabricating PDMS planar patch-clamp electrodes.

We present a new technique for fabricating planar patch electrodes in the laboratory. Planar electrodes are micromolded using a micron-sized stream of air to define an aperture in the silicone elastomer, polydimethylsiloxane (PDMS). We have previously demonstrated that planar PDMS electrodes make excellent patch electrodes after surface modification. We demonstrate single-channel measurements of the rSlo channel in Xenopus oocytes and whole-cell measurements in CHO and RBL mammalian cell lines, using planar PDMS electrodes.

Air↗

PubMed2005

Susceptibility of Giardia lamblia to Hovenia dulcis extracts.

Giardia lamblia is the causative agent of giardiasis, a common parasitic infection of the human and animal digestive tract. Although several drugs have been available to treat this infection, they present unpleasant side effects or cytotoxicity. In order to find a more natural treatment for the disease, we analyzed the effects of the methanolic extract and three fractions obtained from Hovenia dulcis Thunb. (Rhamnaceae) leaves on G. lamblia. Comparing all fractions, dichloromethane was more efficient in reducing Giardia growth. The exposition of G. lamblia to this fraction lead to degenerations in the surface, modifications in the cell shape and alterations in the localization of nuclei. Besides that, the adhesion of G. lamblia was also altered. Experiments revealed that the obtained fraction did not present cytotoxic effects in mammalian cells. In summary, dichloromethane fraction has strong antigiardial effects and could become an important new substance for the treatment of giardiasis.

Animals↗

PubMed2006

Bone response to a titanium aluminium nitride coating on metallic implants.

The design, surface characteristics and strength of metallic implants are dependant on their intended use and clinical application. Surface modifications of materials may enable reduction of the time taken for osseointegration and improve the biological response of bio-mechanically favourable metals and alloys. The influence of a titanium aluminium nitride (TAN) coating on the response of bone to commercially pure titanium and austenitic 18/8 stainless steel wire is reported. TAN coated and plain rods of stainless steel and commercially pure titanium were implanted into the mid-shaft of the femur of Wistar rats. The femurs were harvested at four weeks and processed for scanning electron and light microscopy. All implants exhibited a favourable response in bone with no evidence of fibrous encapsulation. There was no significant difference in the amount of new bone formed around the different rods (osseoconduction), however, there was a greater degree of shrinkage separation of bone from the coated rods than from the plain rods (p = 0.017 stainless steel and p = 0.0085 titanium). TAN coating may result in reduced osseointegration between bone and implant.

Aluminum Compounds↗

PubMed2005

Preparation, characterization and application of fluorescent terbium complex-doped zirconia nanoparticles.

Novel zirconia-based fluorescent terbium nanoparticles have been prepared as a fluorescent nanoprobe for time-resolved fluorescence bioassay. The nanoparticles were prepared in a water-in-oil (W/O) microemulsion consisting of a strongly fluorescent Tb(3+) complex, N,N,N(1), N(1)-[2,6-bis(3'-aminomethyl-1'-pyrazolyl)-phenylpyridine]tetrakis(acetate)-Tb(3+)(BPTA-Tb(3+)), Triton X-100, hexanol, and cyclohexane by controlling co-condensation of Zr(OCH(2)CH(3))(4) and ZrOCl(2.) The characterizations by transmission electron microscopy and fluorometric methods indicate that the nanoparticles are uniform in size, 33+/- 4 nm in diameter, and have a fluorescence quantum yield of 8.9% and a long fluorescence lifetime of 2.0 ms. The zirconia-based fluorescent terbium nanoparticles show high stability against basic dissolution in a high pH aqueous buffer compared to the silica-based nanoparticles. A surface modification and bio-conjugation method for the fluorescent nanoparticles was developed, and the nanoparticle-conjugated streptavidin (SA) was used for time-resolved floro-immunoassy (TR-FIA) of human prostate specific antigen (PSA). The result shows that the zirconia-based fluorescent terbium nanoparticles are useful as a fluorescent nanoprobe for time-resolved fluorescence bioassay.

Fluorescence↗

PubMed2005

Controlled release of dexamethasone from microcapsules produced by polyelectrolyte layer-by-layer nanoassembly.

PURPOSE: In an effort to expand the application of core-shell structures fabricated by electrostatic layer-by-layer (LbL) self-assembling for drug delivery, this study reports the controlled release of dexamethasone from microcrystals encapsulated with a polyelectrolyte shell. METHODS: The LbL self-assembly process was used to produce dexamethasone particles encapsulated with up to five double layers formed by alternating the adsorption of positively charged poly(dimethyldiallyl ammonium chloride), negatively charged sodium poly(styrenesulfonate) and depending on the pH positively or negatively charged gelatin A or B onto the surface of the negatively charged dexamethasone particles. The nano-thin shells were characterized by quartz crystal microbalance measurements, microelectrophoresis, microcalorimetry, confocal microscopy, and scanning electron microscopy. In vitro release of dexamethasone from the microcapsules suspended in water or carboxymethylcellulose gels were measured using vertical Franz-type diffusion cells. RESULTS: Sonication of a suspension of negatively charged dexamethasone microcrystals in a solution of PDDA not only reduced aggregation but also reduced the size of the sub-micrometer particles. Assembly of multiple polyelectrolyte layers around these monodispersed cores produced a polyelectrolyte multilayer shell around the drug microcrystals that allowed for controlled release depending on the composition and the number of layers. CONCLUSIONS: Direct surface modification of dexamethasone microcrystals via the LbL process produced monodispersed suspensions with diffusion-controlled sustained drug release via the polyelectrolyte multilayer shell.

Capsules↗

PubMed2006

Ionization states in the microenvironment of solid dosage forms: effect of formulation variables and processing.

PURPOSE: Evaluation of the effect of formulation composition and processing variables on the microenvironment in solid dosage forms, based on ionization of indicator probes. MATERIALS AND METHODS: Sulfonephthalein indicators were intimately mixed with individual excipients, binary excipient mixtures or multi-component blends by the solvent deposition method. Diffuse reflectance visible spectroscopy of these solids provided a measure of indicator ionization extent. Indicator solution studies yielded equations relating solution pH to the ratio of the absorbance signals of the ionized to that of the unionized form, for each indicator. These equations and the spectral data of the indicator-treated solids were used to calculate an acidity function, 'pH(eq)' for the solids. The ionization of incorporated probes was also monitored during various stages of simulated pharmaceutical processing viz. wet and dry mixing. RESULTS: The pH(eq) provided a measure of the physicochemical environment experienced by the probe in the solid. The surface nature of formulation components and their surface area available for interaction influenced the overall properties of the final blend. The extent of probe ionization varied at different stages of a simulated wet mixing-drying process. The pH of the excipient suspension was not a good predictor of the probe ionization in the final dried solid. Indicator ionization is expected to be influenced by the microenvironmental acidity, polarity and ionic strength. Individual excipient properties contributed to the overall microenvironment in powder mixtures even when dry mixed at low water contents. CONCLUSIONS: The environment experienced by a drug in the final solid dosage form will be influenced by the nature of the excipients, the extent of their surfaces available for interaction, surface modification during processing and the amount and nature of solvent used.

Algorithms↗

PubMed1992

Contribution of complement system on destabilization of liposomes composed of hydrogenated egg phosphatidylcholine in rat fresh plasma.

Large multilamellar vesicles (MLV) composed of hydrogenated egg phosphatidylcholine (HEPC), cholesterol (CH), and dicetyl phosphate (DCP) rapidly release part of an entrapped aqueous marker when incubated with fresh rat plasma and thus have severely limited usefulness as drug carriers. The mechanisms causing the instability of liposomes in plasma were investigated in this study. The leakage of liposomal constituents was completely inhibited by pre-heating at 56 degrees C for 30 min with plasma or by treating with EDTA, K-76COOH, or anti-C3 antiserum but was not inhibited with EGTA/MgCl2. These results indicated that the destabilization of liposomes in fresh rat plasma was induced by activation of the alternative complement pathway (ACP). Furthermore, the complement third component (C3) was detected from the liposomes incubated with fresh plasma by SDS-PAGE followed by Western blotting and immune detection. The C3b deposited on the liposomal surface via ACP was rapidly cleaved to iC3b. The results obtained in the present study suggest a possibility that the liposomes composed of HEPC (without any surface modification) may be effective carriers for macrophages because C3b and its degradative products, iC3b are related to the opsonic function on phagocytosis of foreign particles by macrophages.

Animals↗

PubMed1995

Analysis of metallothionein isoforms by capillary electrophoresis: optimisation of protein separation conditions using micellar electrokinetic capillary chromatography.

Capillary electrophoresis (CE) techniques have been successfully applied to the separation of metallothionein (MT) isoforms and have proved to be rapid, practical and economical. Study of a variety of different electrolytes and capillaries has shown that electrolyte buffer composition and capillary wall surface modifications can have considerable influence on isoform separation and resolution. Ionic surfactants such as sodium dodecyl sulphate (SDS) form micelles at elevated concentrations and the partitioning of molecules between the hydrophobic micelle phase and the aqueous phase and their resulting migration in an electric field is the basis of the technique known as micellar electrokinetic capillary chromatography (MECC). In the present work, we have used sheep and rabbit MT to optimise MECC conditions for analysis of MT isoforms. Capillaries of 57 cm gave much better separations than shorter columns although analysis times were increased to about 12 min. Changing the buffer and SDS concentration or the pH affected the selectivity of isoform separation and up to 5 isoforms in sheep MT and 6 in rabbit MT were completely or partially resolved. Comparing different diameter capillaries we conclude that 25 microns I.D. columns give better separations than 50 or 75 microns I.D. columns although sensitivity is reduced by a factor of about 3 and 5, respectively. Using our MECC conditions, columns coated with C1 or C18 hydrophobic material were not found to be useful in improving MT separation or resolution although further evaluation of these columns is in progress.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1990

Electrochemistry of galvanic couples between carbon and common metallic biomaterials in the presence of crevices.

In vitro experiments were conducted upon some common metallic biomaterials and carbons, both isolated or forming galvanic couples, in a cell specially designed for crevice corrosion studies. The alloys examined were AISI 316L stainless steel, Ti6AI4V and Co-Cr-Mo. The types of carbon were glassy carbon and carbon fibre-reinforced carbon. The surface modifications were evaluated by SEM, AES and ESCA-XPS analyses. AISI 316L stainless steel suffered localized corrosion in open-circuit experiments whilst the other materials remained unattacked. Galvanic currents between metal-carbon couples were measured by zero resistance ammetry. The carbon-metal area ratio was 1:1. The results showed that 316L stainless steel and the Co-Cr-Mo alloy were prone to accelerated corrosion, whilst the Ti6AI4V alloy remained unattacked. The galvanic corrosion currents were also predicted using mixed potential theory from polarization curves obtained for each material. The experimental and theoretical values showed good agreement for the stainless steel and Co-Cr-Mo alloy. Long-term immersion tests with the same couples showed that the only metal not to suffer degradation was the Ti6AI4V alloy.

Alloys↗

PubMed1991

Effect of plasma glow, glutaraldehyde and carbodiimide treatments on the enzymic degradation of poly (L-lactic acid) and poly (gamma-benzyl-L-glutamate) films.

The hydrolytic and enzymic degradation of poly(L-lactic acid) (PLA) and poly(gamma-benzyl L-glutamate) (PBGA) films, together with a series of surface treatments, were studied, as a function of exposure time. The degradation of these polymers was monitored by weight loss, contact angle, pH changes and tensile strength studies. Glutaraldehyde treatment retained the maximum strength of PLA in buffer, followed by carbodiimide, compared with control films. On the other hand, plasma glow reversed the effect. The ability of alpha-chymotrypsin, carboxypeptidase, ficin, esterase, bromelain and leucine aminopeptidase to modulate the degradation of PLA and PBGA was also investigated. Addition of these enzymes to the polymer-buffer system reduced the tensile strength of these polymers variably. Among the six enzymes studied, leucine aminopeptidase showed the highest enzymic effect on the degradation of the glutaraldehyde-treated and bare PLA or bare PBGA films. However, glutaraldehyde-cross-linked PLA demonstrated maximum stability in buffers or in all other enzyme systems studied compared with bare PLA. It is conceivable that surface treatments on these polymers might have altered their physical and chemical configuration and the subsequent degradation properties. Surface modifications may provide new ways of controlling the biodegradation of polymers for a variety of biomedical applications.

Biocompatible Materials↗

PubMed1992

Sterically stabilized liposomes.

Many recent reports have demonstrated that rapid uptake of liposomes in vivo by cells of the mononuclear phagocytic system (MPS), which has restricted their therapeutic utility, can be overcome by incorporation of lipids derivatized with the hydrophilic polymer polyethylene glycol (PEG). The structure-function relationship of PEG-derivatized phosphatidylethanolamine (PEG-PE) has been examined by measurement of blood lifetime and tissue distribution in both mice and rats. The results are reviewed and contrasted with those from liposomes without PEG-PE or other surface modifications. With a PEG molecular weight in the range of 1000 to 5000, prolonged circulation and reduced MPS uptake is achieved. After 24 h, up to 35% of the injected dose remains in the blood and less than 10% is taken up by the two major organs of the MPS, liver and spleen, compared with 1% and up to 50%, respectively, for liposomes without PEG-PE. Other important advantages of PEG-PE have been identified: prolonged circulation is independent of liposome cholesterol content, degree of hydrocarbon chain saturation in either the PC or the PE lipid anchor, lipid dose, or addition of most other negatively charged lipids. This versatility in lipid composition and dose is important for controlling drug release in a liposome-based therapeutic agent. Steric stabilization has been proposed as a theoretical basis for the results and some initial results testing this hypothesis have been reported. A description of a theoretical model is presented here and evaluated with the data available. The results are compared with other particulate drug carriers and the range of potential applications are considered.

Animals↗

PubMed1996

Highly efficient protein separations in high-performance capillary electrophoresis, using hydrophilic coatings on polysiloxane-bonded columns.

Three methods for preparing hydrophilic coatings on polysiloxane bonded CElect H-type capillary electrophoresis columns have been shown. The polyalkylsiloxane-bonded phase is the first coating layer on the capillary surface, and nonionic surfactant, hydrophilic polymer, or polymer surfactant, adsorbed onto this first later through hydrophobic interactions, forms the second coating layer. The resultant capillary surfaces are inert, hydrophilic, and suitable for highly efficient protein separations. The effectiveness and applicability of these capillary surface modification methods were tested for the separations of a variety of proteins over a wide range of buffer pH values under different capillary electrophoretic operations modes.

Buffers↗

PubMed1990

Ultrastructural observations on regenerating hair cells in the chick basilar papilla.

This experiment was designed to investigate cellular and subcellular maturational changes in regenerated immature sensory cells and support cells of the chick basilar papilla following gentamycin treatment. Scanning and transmission electron microscopy were used. The experimental animals received one subcutaneous injection of gentamycin sulfate daily (50 mg/kg) for five or 10 days. The animals receiving five days of injection were sacrificed the following day. The remaining animals were allowed to survive either seven or 28 days before sacrifice and preparation for electron microscopy. The initial lesion consisted of total degeneration of hair cells within 500 microns of the proximal tip providing the opportunity to study a 'pure' population of regenerating sensory cells. Sensory cell regeneration could be identified by one day after terminating gentamycin treatment. Early in development sensory cell precursors were morphologically very similar to supporting cells. A density gradient, based on cytoplasmic staining characteristics, was established which increased from cells displaying low density at the base of the supporting cell layer to high density cells at the luminal surface. These changes in density were equated to increase in number of and types of cytoplasmic organelles. In contrast to the empty appearing cytoplasm of the support cell, the cytoplasm of the hair cell precursor contained numerous mitochondria, clusters of ribosomes, and vesicles. As the cell approached the surface, mitochondria became more numerous as did smooth and coarse endoplasmic reticulum and Golgi apparatus. This gradient suggested that determination of the cellular phenotype occurred at the level of the basal membrane followed by migration to the surface, during which time differentiation was characterized by an increase in number and complexity of cellular organelles. Luminal surface modifications occurred as soon as the cell erupted. The development of stereocilia, rootlet, cuticular plate and cellular polarization followed the normal embryogenetic pattern. At 28 days, stereocilia organization was still incomplete as was the orientation of the bundle. To the extent that proper orientation of hair cells or bundles is necessary for normal transduction, mature function at 28 days would not be anticipated. Innervation of the presumptive hair cell precursors could be observed one day after treatment, early in the course of hair cell differentiation. Synaptogenesis followed the normal embryogenetic sequence; however, afferent and efferent nerve terminals remained immature appearing at 28 days. This observation may have physiological implications manifested by delay of hearing

Afferent Pathways↗

PubMed2004

Artificial surface-induced cytokine synthesis: effect of heparin coating and complement inhibition.

BACKGROUND: Contact between blood and artificial surfaces induces an inflammatory response including activation of leukocytes and platelets, as well as complement and other plasma cascade systems. In the present study we investigated the roles of complement and surface modification in polyvinylchloride-induced cytokine production. METHODS: Human whole blood was incubated in rotating loops of polyvinylchloride or heparin-coated polyvinylchloride tubing for 4 hours. Plasma concentrations of the cytokines tumor necrosis factor alpha, interleukin (IL) 1 beta, IL-6, IL-8, IL-10, and monocyte chemoattractant protein 1 (MCP-1) were quantified. RESULTS: Polyvinylchloride induced a substantial increase in IL-8 and MCP-1, which was abolished by cycloheximide, indicating that they were synthesized during incubation. Interleukin 8 synthesis was completely complement-dependent since it was abolished by neutralizing antibodies to factor D and complement factor 5, as well as by a complement factor 5a receptor antagonist. Monocyte chemoattractant protein 1 synthesis was reduced by approximately half the amount by the complement inhibitors. Heparin-coated polyvinylchloride efficiently prevented synthesis of both IL-8 and MCP-1. Addition of recombinant human complement factor 5a to blood incubated in heparin-coated polyvinylchloride restored IL-8 and MCP-1 production completely and partly, respectively. In contrast to IL-8 and MCP-1, tumor necrosis factor alpha, IL-1 beta, interleukin 6 and IL-10 increased only marginally. A minor but significant increase in IL-1 beta was complement-dependent, whereas a similar increase in IL-10 was completely prevented by heparin-coated polyvinylchloride. No significant changes were observed for tumor necrosis factor alpha and IL-6. CONCLUSIONS: Polyvinylchloride induced a marked increase in IL-8 and MCP-1, in contrast to a marginal increase in tumor necrosis factor alpha, IL-1 beta, IL-6, and IL-10. The increase in IL-8 and MCP-1 was prevented by heparin-coated polyvinylchloride. Interleukin 8 production was totally complement-dependent and mediated by complement factor 5a.

Anticoagulants↗

PubMed2005

Differential effect of heparin coating and complement inhibition on artificial surface-induced eicosanoid production.

BACKGROUND: Contact between blood and artificial surfaces induces an inflammatory response including activation of leukocytes and platelets, as well as complement and other plasma cascade systems. In the present study we investigated the roles of complement and surface modification in polyvinyl chloride-induced synthesis of eicosanoids (arachidonic acid metabolites). METHODS: Human whole blood was incubated in rotating loops of polyvinyl chloride or heparin-coated polyvinyl chloride tubing for 4 hours. Plasma concentrations of the eicosanoids leukotriene B4, prostaglandin E2 and thromboxane B2 were quantified. RESULTS: Polyvinyl chloride induced a substantial increase in leukotriene B4, prostaglandin E2, and thromboxane B2. Inhibition of complement activation by the complement factor 3 binding peptide compstatin or blockade of the complement factor 5a receptor with a specific antagonist significantly and specifically inhibited the synthesis of leukotriene B4, whereas thromboxane B2 and prostaglandin E2 synthesis were apparently complement independent. The increase in all three mediators was significantly reduced by the heparin coating. Indomethacin abolished the increase of the cyclooxygenase products prostaglandin E2 and thromboxane B2, but had no effect on the increase of the lipoxygenase product leukotriene B4, consistent with the specificity of indomethacin for the cyclooxygenase and confirming the specificity of complement inhibition. CONCLUSIONS: Polyvinyl chloride-induced increase in all three eicosanoids was attenuated by heparin coating, whereas complement inhibition selectively reduced the synthesis of leukotriene B4.

Complement Inactivator Proteins↗

PubMed2004

Incorporation of biodegradable nanoparticles into human airway epithelium cells-in vitro study of the suitability as a vehicle for drug or gene delivery in pulmonary diseases.

PURPOSE: Nanoparticles are able to enhance drug or DNA stability for purposes of optimised deposition to targeted tissues. Surface modifications can mediate drug targeting. The suitability of nanoparticles synthesised out of porcine gelatin, human serum albumin, and polyalkylcyanoacrylate as drug and gene carriers for pulmonary application was investigated in vitro on primary airway epithelium cells and the cell line 16HBE14o-. METHODS: The uptake of nanoparticles into these cells was examined by confocal laser scan microscopy (CLSM) and flow cytometry (FACS). Further the cytotoxicity of nanoparticles was evaluated by an LDH-release-test and the inflammatory potential of the nanoparticles was assessed by measuring IL-8 release. RESULTS: CLSM and FACS experiments showed that the nanoparticles were incorporated into bronchial epithelial cells provoking little or no cytotoxicity and no inflammation as measured by IL-8 release. CONCLUSIONS: Based on their low cytotoxicity and the missing inflammatory potential in combination with an efficient uptake in human bronchial epithelial cells, protein-based nanoparticles are suitable drug and gene carriers for pulmonary application.

Biodegradation, Environmental↗

PubMed2005

Tethering poly(ethylene glycol)s to improve the surface biocompatibility of poly(acrylonitrile-co-maleic acid) asymmetric membranes.

To improve the surface biocompatibility, asymmetric membranes fabricated from poly(acrylonitrile-co-maleic acid)s (PANCMAs) synthesized by water-phase precipitation copolymerization were tethered (or immobilized) with poly(ethylene glycol)s (PEGs) by esterification reaction. Chemical changes on the membrane surface were characterized by Fourier transform infrared spectroscopy and elemental analysis to confirm the immobilization of PEG onto the PANCMA membranes. The hydrophilicity and blood compatibility of the PEG-tethered PANCMA membrane were investigated by water contact angle, water absorption, protein adsorption, plasma platelets adhesion and cell adhesion measurements, and the results were compared with the corresponding PANCMA membranes. It was found that, after the tethering of PEG, the hydrophilicity of the membrane can be improved significantly, and the protein adsorption, platelets adhesion and macrophage attachment on the membrane surface are obviously suppressed. Furthermore, not only the content of maleic acid in PANCMA, which influences the tethering density of PEG, but also the molecular weight of PEG has great effect on the surface modification of PANCMA membranes for biocompatibility.

Acrylic Resins↗