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Biomedical subjects

W G Pitt

Publications and source records attributed to W G Pitt.

34 records · Page 2Linked to original sources
PubMed1996

Effect of low-intensity ultrasound upon biofilm structure from confocal scanning laser microscopy observation.

Ultrasonic irradiation at 500 kHz and 10 mW cm-2 of a 24 h old biofilm of P. aeruginosa enhanced the killing of bacteria by gentamicin. To determine whether this bioacoustic effect was caused by ultrasonic-induced changes in the biofilm morphology (biofilm breakup or disruption), the biofilms were examined by confocal scanning laser microscopy (CSLM). Such disruption would be undesirable in the possible ultrasonic treatment of implant infections. The CSLM results showed that the biofilm is a partial monolayer of cells with occasional aggregates of cells, non-cellular materials and extracellular spaces. The aggregates contained large amounts of exopolysaccharide. The structure of biofilm was not changed when the biofilm was exposed to continuous ultrasound at 500 kHz and 10 mW cm-2, the same irradiation parameters that increased cell killing by nearly two orders of magnitude. The observation that low-intensity ultrasound does not disrupt biofilm or disperse the bacteria has significance in the possible use of ultrasound to enhance the action of antibiotics against biofilms.

Acoustics↗
PubMed1994

Ultrasonic enhancement of antibiotic action on gram-negative bacteria.

The effect of gentamicin upon planktonic cultures of Pseudomonas aeruginosa, Escherichia coli, Staphylococcus epidermidis, and Staphylococcus aureus was measured with and without application of 67-kHz ultrasonic stimulation. The ultrasound was applied at levels that had no inhibitory or bactericidal activity against the bacteria. Measurements of the MIC and bactericidal activity of gentamicin against planktonic cultures of P. aeruginosa and E. coli demonstrated that simultaneous application of 67-kHz ultrasound enhanced the effectiveness of the antibiotic. A synergistic effect was observed and bacterial viability was reduced several orders of magnitude when gentamicin concentrations and ultrasonic levels which by themselves did not reduce viability were combined. As the age of the culture increased, the bacteria became more resistant to the effect of the antibiotic alone. Application of ultrasound appeared to reverse this resistance. The ultrasonic treatment-enhanced activity was evident with cultures of P. aeruginosa and E. coli but was not observed with cultures of gram-positive S. epidermidis and S. aureus. These results may have application in the treatment of bacterial biofilm infections on implant devices, which infections are usually more resistant to antibiotic therapy.

Biofilms↗
PubMed1993

Bacterial adhesion to poly(HEMA)-based hydrogels.

The effects of water content and comonomer chemistry upon the adhesion of Pseudomonas aeruginosa to poly(hydroxyethyl methacrylate)-based hydrogels were studied. Hydrogels which varied in swollen water content from 33-69 wt% were polymerized onto glass microscope slides pretreated with a vinyl silane. The hydrogel water content was varied by adding methacrylic acid (1-5 wt%) or N-vinyl pyrrolidone (NVP, 10-25 wt%) or combinations of the two comonomers. The resulting hydrogel surfaces, which were 0.1 mm thick, transparent, and adherent to the glass slide, formed the test surfaces of laminar flow cells (Re = 1.3, wall shear rate = 1.6/s). The bacteria were grown for 8 h in tryptic soy broth (TSB), washed by filtration, and collected on 0.45-microns filters, resuspended in phosphate buffered saline (PBS) at pH = 7.2, and recirculated through the flow cell and across the test surface at 0.85 mL/min for 2 h. Results show that P. aeruginosa adhered less to hydrogels with higher water contents. In the presence of TSB and possible poly(NVP) contamination, the concentration of adherent bacteria was reduced to low and uniform levels independent of the hydrogel chemistry.

Bacterial Adhesion↗
PubMed1993

Air-water interface displaces adsorbed bacteria.

Video microscopy was employed to observed the spatial distribution of Staphylococcus epidermidis and Pseudomonas aeruginosa adherent to glass and polymer substrates. During rinsing procedures the bacteria remained in their original positions when the surfaces were rinsed with saline for 3 min followed by ethanol for 3 min before exposure to air. When the surfaces were rinsed with saline only, the air-liquid interface disrupted the spatial distribution of the bacteria, removing and redepositing the bacteria in clumps. A moving air-liquid interface of a gas bubble on substrate also displaced bacteria. Such artefacts produced by air-water interfaces should be avoided during bacterial adhesion experiments.

Adsorption↗
PubMed1993

Fibronectin adsorpton kinetics on phase segregated polyurethaneureas.

The kinetics of fibronectin (FN) adsorption upon three polyurethaneureas (PUU) were measured by Fourier transform infrared spectroscopy. The three polymers had soft blocks composed of polydimethylsiloxane (PDMS), polytetramethyleneoxide (PTMO), and polyethyleneoxide (PEO). On each polymer, the amount of adsorbate increased proportional to the square root of time, but the rates were less than that predicted for purely diffusion controlled adsorption. Adsorption rate constants and sticking coefficients (the fraction of encounters with the surface which result in adsorption) were calculated from the data. The adsorption rate constants are 4.8 x 10(-4), 1.3 x 10(-4), and 2.5 x 10(-5) cm/s on PDMS-PUU, PTMO-PUU and PEO-PUU respectively. The rate constants and sticking coefficients fall within the range of previously reported values for other protein/polymer systems. The sticking coefficients correlate in general with the amount of FN adsorbed at 120 min, and with the extent of conformational change of the adsorbed protein.

Adsorption↗
PubMed1993

Bacterial adhesion to protein-coated hydrogels.

Extended wear soft contact lenses have been implicated in the increased occurrence of corneal bacterial infections. This research investigated the effects of polymer chemistry, water content, and pre-sorbed proteins upon the adherence of Pseudomonas aeruginosa to model hydrogels with chemistries similar to those of extended wear soft contact lenses. The hydrogels were exposed to washed suspensions of P. aeruginosa in a laminar flow cell. Albumin, fibrinogen, desialylated fibrinogen, or mucin were deposited on the hydrogels before exposure to the bacteria. Results showed that with or without protein pre-exposure, bacterial adhesion decreased as water content increased. In the presence of the sorbed protein, the number of adherent bacteria increased by about 45%, and all four proteins caused similar increases in adhesion. Bacterial adhesion was not significantly influenced by the presence of sialic acid residues in the pre-sorbed protein.

Animals↗
PubMed1992

Low fluorescence background electroblotting membrane for DNA sequencing.

A low fluorescence background polypropylene (PP) membrane has been developed for ultimate use as an electroblotting membrane in DNA sequencing based on fluorescence detection. The DNA binding capacity of this membrane is improved by a surface modification using radio frequency plasma discharge (RFPD) in ammonia gas. The RFPD operational parameters are evaluated both in terms of membrane nitrogen content and in terms of the product's capacity for binding radioisotope-labeled DNA fragments. The surface morphologies of the derivatized membranes are examined by scanning electron microscopy; their mechanical and electrical properties, which are important for the subsequent sequencing procedures, are likewise established. Due to the goal of developing a membrane suitable for multiplex processing, in which the electroblotted DNA must withstand dozens of hybridization/stripping cycles, special attention is given the covalent attachment of DNA to the membrane. The modified PP membrane is evaluated in a multiplex sequencing application using radioisotope-labeled DNA probes, and found to yield somewhat better binding of a given amount of electroblotted DNA than the commonly used GeneScreen membrane. A tenfold repetition of the probing indicates little loss of signal; the membrane-bound DNA is stable upon storage and shows no detectable loss in probing efficiency after one month.

Base Sequence↗
PubMed1992

Sticking coefficients of adsorbing proteins.

The protein sticking coefficient, phi, the fraction of collisions that result in adsorption, is a function of the molecular interactions between the protein and the surface. A random walk and diffusion-to-capture model was used to describe the kinetics of protein adsorption. The assumption of a constant sticking coefficient leads to a first-order model of the kinetics. A solution of the problem of adsorption from a semi-infinite medium with first-order kinetics at the boundary was obtained by numerical simulation on the computer. The results of the computer simulations match the time dependence observed experimentally. A correlation was developed to estimate phi from experimental data. phi has been found to be in the range 10(-5)-10(-8) for several protein adsorption kinetic studies reported in the literature.

Adsorption↗
PubMed1990

Characterization of hydrophobicity gradients prepared by means of radio frequency plasma discharge.

A hydrophobicity gradient was created by gradually exposing a polydimethylsiloxane film to a radio frequency glow discharge in an oxygen atmosphere. A change in contact angle from 100 to 0 degrees was measured along the gradient surface by means of the Wilhelmy balance technique. The gradient surface was characterized by studying the adsorption from single protein solutions of human albumin, IgG and fibrinogen using total internal reflectance fluorescence (TIRF). Generally, adsorption values similar to monolayer capacities were measured on the hydrophobic side. The adsorption decreased towards the hydrophilic end in correspondence with the change in contact angle. The displacement of proteins with a ethylene oxide-propylene oxide copolymer (EO70PO100EO70) was higher on the hydrophobic side than on the hydrophilic side. Albumin showed an adsorption peak in the wetting transition region, indicating a very strong affinity.

Adsorption↗
PubMed1988

Albumin adsorption on alkyl chain derivatized polyurethanes: I. The effect of C-18 alkylation.

The initial adsorption rate of delipidized Human Serum Albumin (HSA) is increased by addition of C-18 alkyl chains to a polyurethane. The presence of alkyl chains does not appear to influence the total amount of HSA adsorbed after one hour exposure to a 5.0 mg/mL HSA solution. Neither does the desorption following one hour of adsorption appear to be influenced by the presence of alkyl chains. A study of the effects of solution concentration and temperature showed that the initial adsorption rates on both polymers are proportional to the protein concentration raised to the 0.36 power, and that alkylation of the polymer increases the activation energy of the initial adsorption rate above the 14 kJ/mol observed for the underivatized polyurethane. A new technique is presented to quantify the mass of adsorbed protein using Fourier transform infrared spectroscopy and attenuated total reflection optics. This technique uses the absorbance of bulk protein as an internal calibration reference, and appears to be as accurate and perhaps more precise than radiolabeling techniques.

Adsorption↗
PubMed1988

Albumin adsorption on alkyl chain derivatized polyurethanes. II. The effect of alkyl chain length.

Linear alkyl chains containing 2, 10 and 18 carbon atoms were grafted to 10% of the urethane nitrogens in a polyether-polyurethane. The polyurethane was synthesized from methylene bis(p-phenyl isocyanate), 1,4-butanediol, and polytetramethylene oxide of 1000 molecular weight in a molar ratio of 3/2/1. Fourier transform infrared spectroscopy and attenuated total reflectance optics were used to study the adsorption of 5.0 mg/ml human serum albumin (HSA) at 37 degrees C to the derivatized and non-derivatized polymers. Both delipidized HSA and HSA containing 6.5 mol stearic acid per mol of albumin were used to study the effect of chain length upon the initial adsorption rate, the total amount adsorbed in 1 h, and the desorption rate. The initial adsorption rates revealed that non-specific adsorption was similar upon all four polymers. An increase in initial adsorption rate upon the C-18 derivatized polymer was attributed to a specific binding interaction between the HSA and the grafted alkyl chains. The conformational stability of the HSA also affected the adsorption rate. The total amount adsorbed after 1 h decreased as the alkyl chain length increased from 2 to 18 methylene groups. The desorption rate decreased in magnitude as the alkyl chain length increased from C-2 to C-18. These results support a hypothesis that alkyl chain length influences the interaction between albumin and an alkylated polymer system.

Adsorption↗
PubMed1987

Properties of extruded poly(tetramethylene oxide)-polyurethane block copolymers for blood-contacting applications.

The bulk and surface properties and blood compatibility of a series of polyurethanes based on methylene bis(p-phenyl isocyanate), 1,4-butanediol, and poly(tetramethylene oxide) of molecular weight 1000 were studied. The hard-to-soft segment ratio of these multiphase polymers was varied, and the effect of substituting a poly(dimethylsiloxane)-containing polyol in place of 5% of the polyether soft segment was studied. Bulk properties such as tensile strength and modulus increased with hard segment content, as did surface wettability and ESCA nitrogen content. However, blood compatibility measured by a canine ex vivo blood-contacting experiment was not found to vary with hard/soft segment ratio. The addition of the silicone-containing polyol did not significantly lower the surface wettability, although ESCA-measured silicon content increased and physical properties were unfavourably affected by the incorporation of this co-soft segment. Incorporation of the siloxane-containing component resulted in increased platelet adhesion and fibrinogen deposition at most blood contact times in comparison with the other polyurethanes.

Animals↗
PubMed1986

FTIR-ATR studies of the effect of shear rate upon albumin adsorption onto polyurethaneurea.

The effect of wall shear rate upon albumin adsorption onto a polyurethaneurea, Biomer, was studied using FTIR-ATR spectroscopy. In these experiments, nondiffusion controlled adsorption was observed from 10 mg/ml solutions at known wall shear rates. Results show that increasing wall shear rate during adsorption does not significantly affect the adsorption kinetics, but does decrease the rate of protein desorption. These results suggest that albumin adsorbed at higher shear rates is more tightly bound to the surface. Comparison of the data to simple mathematical models suggests that more than one layer of protein adsorbs in 2 h.

Adsorption↗
PubMedDate not supplied

The effect of ultrasonic frequency upon enhanced killing of P. aeruginosa biofilms.

It is widely recognized that the bacteria sequestered in a biofilm on a medical implant are much more resistant to antibiotics than their planktonic counterparts. Recent studies have shown that application of antibiotic along with low power ultrasound significantly increases the killing of planktonic bacteria by the antibiotic. Herein is reported a similar application of antibiotic and ultrasound to sessile bacteria in biofilms of Pseudomonas aeruginosa on a polyethylene substrate. Biofilm viability was measured after exposure to 12 micrograms/ml gentamicin sulfate and 10 mW/cm2 ultrasound at frequencies of 70 kHz, 500 kHz, 2.25 MHz, and 10 MHz. The results indicate that a significantly greater fraction of the bacteria was killed by gentamicin when they were subjected to ultrasound. However, ultrasound by itself did not have any deleterious effect on the biofilm viability. In addition, lower-frequency insonation is significantly more effective than higher frequency in reducing bacterial viability within the biofilm. The possible mechanisms of synergistic action are discussed.

Anti-Bacterial Agents↗
PubMedDate not supplied

Markers of thromboembolization in a bovine ex vivo left ventricular assist device model.

The production of blood microemboli (BME) was studied using an ex vivo exteriorized left ventricular assist device (LVAD) model in calves. Each of eight calves received a series of three LVADs, each operating for 24 hr. Blood microemboli were measured directly by a laser (624 nm and 828 nm) light scattering microemboli detection (LSMD) system through the LVAD outflow cannula and by constant pressure filtration (CPF) of blood samples from the LVAD outflow cannula. Hematologic parameters were also measured. After LVAD removal, perivalvular thrombi were evaluated using polar coordinate mapping. The average LSMD and CPF results correlated. For example, in one series of three calves, one ventricle exhibited significantly greater thrombogenesis than did the other ventricles, as indicated by both the LSMD and CPF results. In a series of five calves, one calf developed an abnormally high activated thromboplastin time (APTT), even in the absence of heparin. For two of the three ventricles tested in that calf, microemboli concentration (CPF), Factor XII activity, level of fibrin degradation products (FDP), and accumulated thrombus were significantly lower than for the other calves. The whole blood viscosity (WBV, at 230 s-1) in this calf also decreased to lower values than were seen with the other calves.

Animals↗