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Biomedical subjects

Paul Webster

Publications and source records attributed to Paul Webster.

At least 37 records · Page 2Linked to original sources
PubMed2005

Bacterial biofilms in surgical specimens of patients with chronic rhinosinusitis.

OBJECTIVES: Biofilms are bacterial pathogens that organize in several chronic and recalcitrant infectious processes. We hypothesize that biofilms play a role in chronic rhinosinusitis (CRS). Our goal is to demonstrate biofilms in mucosal specimens of patients undergoing surgery for CRS. STUDY DESIGN: A prospective study of the presence of biofilms in patients undergoing endoscopic sinus surgery for CRS compared with control patients without CRS. METHODS: There were a total of 30 subjects and 4 controls enrolled. The samples of 24 subjects and 4 controls were cultured and then prepared using standard methods for scanning electron microscopy (SEM). The remaining six subjects' samples were treated using advanced cryofixation methods as preparation to preserve structure for SEM and transmission electron microscopy (TEM). RESULTS: Using strict SEM morphologic criteria, 24 (80%) of the 30 patients were found to have micrographic evidence of biofilms. All controls had healthy appearing cilia and goblet cells without biofilms. The six cryofixation samples showed biofilm structures on SEM micrographs that were correlated with bacterial structures seen at the mucosal surface on the corresponding TEM cross sections. Bacterial cultures were positive on all patients. CONCLUSIONS: Biofilms were demonstrated to be present in patients undergoing surgery for CRS; none of the patients without CRS had any evidence of biofilms. Although SEM is capable of demonstrating the biofilms' three-dimensional structure, glycocalyx, and water channels, it cannot clearly demonstrate the presence of bacteria within the biofilm. We were able to demonstrate evidence of bacteria in the biofilms on the subjects tested using TEM.

Adolescent↗
PubMed2004

Loss of function of KRE5 suppresses temperature sensitivity of mutants lacking mitochondrial anionic lipids.

Disruption of PGS1, which encodes the enzyme that catalyzes the committed step of cardiolipin (CL) synthesis, results in loss of the mitochondrial anionic phospholipids phosphatidylglycerol (PG) and CL. The pgs1Delta mutant exhibits severe growth defects at 37 degrees C. To understand the essential functions of mitochondrial anionic lipids at elevated temperatures, we isolated suppressors of pgs1Delta that grew at 37 degrees C. One of the suppressors has a loss of function mutation in KRE5, which is involved in cell wall biogenesis. The cell wall of pgs1Delta contained markedly reduced beta-1,3-glucan, which was restored in the suppressor. Stabilization of the cell wall with osmotic support alleviated the cell wall defects of pgs1Delta and suppressed the temperature sensitivity of all CL-deficient mutants. Evidence is presented suggesting that the previously reported inability of pgs1Delta to grow in the presence of ethidium bromide was due to defective cell wall integrity, not from "petite lethality." These findings demonstrated that mitochondrial anionic lipids are required for cellular functions that are essential in cell wall biogenesis, the maintenance of cell integrity, and survival at elevated temperature.

Alleles↗
PubMed2004

Antimicrobial activity of innate immune molecules against Streptococcus pneumoniae, Moraxella catarrhalis and nontypeable Haemophilus influenzae.

BACKGROUND: Despite its direct connection to the nasopharynx which harbors otitis media pathogens as part of its normal flora, the middle ear cavity is kept free of these bacteria by as yet unknown mechanisms. Respiratory mucosal epithelia, including those of the middle ear and eustachian tube, secrete antimicrobial effectors including lysozyme, lactoferrin and beta defensins-1 and -2. To elucidate the role of these innate immune molecules in the normal defense and maintenance of sterility of respiratory mucosa such as that of the middle ear, we assessed their effect on the respiratory pathogens nontypeable Haemophilus influenzae (NTHi) 12, Moraxella catarrhalis 035E, and Streptococcus pneumoniae 3, and 6B. METHODS: Two assay methods, the radial assay and the liquid broth assay, were employed for testing the antimicrobial activity of the molecules. This was done in order to minimize the possibility that the observed effects were artifacts of any single assay system employed. Also, transmission electron microscopy (TEM) was employed to evaluate the effect of antimicrobial innate immune molecules on OM pathogens. For the statistical analysis of the data, Student's t-test was performed. RESULTS: Results of the radial diffusion assay showed that beta defensin-2 was active against all four OM pathogens tested, while treatment with beta defensin-1 appeared to only affect M. catarrhalis. The radial assay results also showed that lysozyme was quite effective against S. pneumoniae 3 and 6B and was partially bacteriostatic/bactericidal against M. catarrhalis. Lysozyme however, appeared not to affect the growth of NTHi. Thus, lysozyme seems to have a more pronounced impact on the growth of the Gram-positive S. pneumoniae as compared to that of Gram-negative pathogens. Lactoferrin on the other hand, enhanced the growth of the bacteria tested. The results of the radial assays were confirmed using liquid broth assays for antimicrobial activity, and showed that lysozyme and beta defensin-2 could act synergistically against S. pneumoniae 6B. Moreover, in the liquid broth assay, beta defensin-1 showed a modest inhibitory effect on the growth of S. pneumoniae 6B. As assessed by ultrastructural analysis, lysozyme and beta defensin-2, and to a much lesser extent, beta defensin-1, appeared to be able to cause damage to the bacterial membranes. CONCLUSIONS: Here we report that lysozyme and the beta defensins can inhibit the growth of clinical isolates of otitis media pathogens - namely NTHi strain 12, S. pneumoniae strains 3 and 6B and M. catarrhalis strain 035E - and cause ultrastructural damage to these pathogens. Moreover, we demonstrate that lysozyme and beta defensin-2 can act synergistically against S. pneumoniae. These findings are consistent with the concept that secreted antimicrobial peptides and other components of innate immunity constitute the first line of defense protecting host mucosal surfaces, including the tubotympanal (eustachian tube and middle ear cavity) mucosa, against pathogens.

Anti-Infective Agents↗
PubMed2004

PEGylation significantly affects cellular uptake and intracellular trafficking of non-viral gene delivery particles.

In vitro studies of non-viral gene delivery vectors are typically not performed at physiological conditions, and thus may not provide meaningful results for in vivo investigations. We determine if polycation-plasmid DNA complexes (polyplexes) exploited for in vitro studies behave similarly to variants more applicable to in vivo use by examining their cellular uptake and trafficking. Branched polyethylenimine (25 kDa) or a linear beta-cyclodextrin-containing polymer are each used to formulate polyplexes, which can be PEGylated (PEG: poly(ethylene glycol)) to create particles stable in physiological salt concentrations. Particle size, cellular uptake, intracellular trafficking, and reporter gene expression are reported for polyplexes and for their PEGylated variants. PEGylation confers salt stability to particles but produced a reduction in luciferase expression. Examination of in vitro particle internalization by transmission electron microscopy shows unmodified polyplexes entering cells as large aggregates while PEGylated particles remain small and discrete, both outside and within cells. Unmodified and PEGylated particles enter cells through the endocytic pathway and accumulate in a perinuclear region. Immunolabeling reveals unpackaged exogenous DNA in the cytoplasm and nuclei. It appears all particle types traffic towards the nucleus within vesicles and undergo degradation in vesicles and/or cytoplasm, and eventually some exogenous DNA enters the nucleus, where it is transcribed. In comparing polyplexes and their PEGylated variants, significant differences in particle morphology, cellular uptake, and resultant expression suggest that in vitro studies should be conducted with particles prepared for physiological conditions if the results are to be relevant to in vivo performance.

Animals↗
PubMed2004

Basilar membrane displacement related to endolymphatic sac volume.

OBJECTIVE: To demonstrate that the amount of basilar membrane displacement toward the scala tympani and its attachment to the bony wall of the scala tympani (i.e., interscalar septum) in hydropic temporal bones is related to the intraosseous endolymphatic sac volume. STUDY DESIGN: A retrospective analysis of temporal bones from individuals with the histopathologic diagnosis of "endolymphatic hydrops." METHODS: Fifty-two temporal bones, from 38 patients, with the histopathologic findings of "endolymphatic hydrops" were analyzed microscopically. Data were obtained regarding the displacement of the basilar membrane, endolymphatic sac volume, hair cell loss, strial atrophy, ganglion cell loss, and last measured auditory thresholds. The relationships between these variables were examined statistically. RESULTS: Nineteen of the 52 temporal bones (36.5%) with endolymphatic hydrops showed displacement of the basilar membrane toward the scala tympani in the apical and middle segments of the cochlea. A reduced volume of the endolymphatic sac was significantly related to increased severity of basilar membrane deformation (Rho = -.646; P < or = .001). Multiple regression analysis showed that severity of basilar membrane deformation was the single best predictor of low frequency thresholds while loss of hair cells was the best predictor of pure-tone average threshold. CONCLUSIONS: The displacement of the basilar membrane in the apical and middle segments that may occur with endolymphatic hydrops, to the extent that it impinges on the interscalar septum, is related to a reduction in the intraosseous endolymphatic sac volume.

Aged↗