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Biomedical subjects

M Bradshaw

Publications and source records attributed to M Bradshaw.

At least 19 recordsLinked to original sources
PubMed2000

Cloning, nucleotide sequence, and expression of the gene encoding the bacteriocin boticin B from Clostridium botulinum strain 213B.

Boticin B is a heat-stable bacteriocin produced by Clostridium botulinum strain 213B that has inhibitory activity against various strains of C. botulinum and related clostridia. The gene encoding the bacteriocin was localized to a 3.0-kb HindIII fragment of an 18. 8-kb plasmid, cloned, and sequenced. DNA sequencing revealed the boticin B structural gene, btcB, to be an open reading frame encoding 50 amino acids. A C. botulinum strain 62A transconjugant containing the HindIII fragment inserted into a clostridial shuttle vector expressed boticin B, although at much lower levels than those observed in C. botulinum 213B. To our knowledge, this is the first demonstration and characterization of a bacteriocin from toxigenic group I C. botulinum.

Amino Acid Sequence↗
PubMed2000

Treatment with subantimicrobial dose doxycycline improves the efficacy of scaling and root planing in patients with adult periodontitis.

BACKGROUND: In a previous study, subantimicrobial dose doxycycline (SDD) significantly improved clinical parameters associated with periodontal health in patients with adult periodontitis (AP) when used as an adjunct to a maintenance schedule of supragingival scaling and dental prophylaxis. In this double-blind, placebo-controlled, parallel-group, multicenter study, the efficacy and safety of SDD were evaluated in conjunction with scaling and root planing (SRP) in patients with AP. METHODS: Patients (n = 190) received SRP at the baseline visit and were randomized to receive either SDD 20 mg bid or placebo bid for 9 months. Efficacy parameters included the per-patient mean changes in clinical attachment level (CAL) and probing depth (PD) from baseline, the per-patient percentages of tooth sites with attachment loss (AL) > or = 2 mm and > or = 3 mm from baseline, and the per-patient percentage of tooth sites with bleeding on probing. Prior to analysis, tooth sites were stratified by the degree of disease severity evident at baseline RESULTS: In tooth sites with mild to moderate disease and severe disease (n = 183, intent-to-treat population), improvements in CAL and PD were significantly greater with adjunctive SDD than with adjunctive placebo at 3, 6, and 9 months (all P <0.05). In tooth sites with severe disease, the per-patient percentage of sites with AL > or = 2 mm from baseline to month 9 was significantly lower with adjunctive SDD than with adjunctive placebo (P<0.05). Improvements in clinical outcomes occurred without detrimental shifts in the normal periodontal flora or the acquisition of doxycycline resistance or multiantibiotic resistance. SDD was well tolerated, with a low incidence of discontinuations due to adverse events. CONCLUSIONS: The adjunctive use of SDD with SRP is more effective than SRP alone and may represent a new approach in the long-term management of AP.

Adult↗
PubMed2000

Long-term use of subantimicrobial dose doxycycline does not lead to changes in antimicrobial susceptibility.

BACKGROUND: Adjunctive subantimicrobial dose doxycycline (SDD) with scaling and root planing leads to improved clinical parameters of adult periodontitis, but has raised questions about potential changes in antibiotic susceptibility of the host microflora. Our four studies assessed whether long-term SDD changes antibiotic susceptibility of the oral microflora in adults with periodontitis. METHODS: In studies 1 and 2, adult patients with periodontitis were randomized to receive SDD 10 mg qd, 20 mg qd, 20 mg bid, or placebo. In study 3, patients were randomized to receive SDD 20 mg bid or placebo. No medication was administered in study 4, a follow-up to study 3. Subgingival plaque samples were collected at baseline (all studies) and at 12, 15 to 18, and 24 months (study 1); 12, 18, and 27 months (study 2); 3, 6, and 9 months (study 3); and 3 months post-study 3 (study 4). Antimicrobial susceptibility of isolated bacteria was assessed by: 1) minimum inhibitory concentration (MIC) levels (studies 1 and 2); 2) cross-resistance to non-tetracycline antibiotics (studies 2 and 3); and 3) the proportion of doxycycline-resistant isolates (studies 3 and 4). RESULTS: Organism MIC levels remained constant among all treatment groups at 18 and 24 months compared with baseline (study 1). Observed changes in susceptibility at 12 and 18 months for the 20 mg groups were attributed to the limited number of isolates tested (study 1). There were no statistically significant differences in the proportion of doxycycline-resistant isolates among treatment groups (studies 3 and 4), and no evidence of multi-antibiotic resistance (studies 3 and 4) or cross-resistance (studies 2 and 3) at any timepoint. CONCLUSION: Long-term SDD does not contribute to changes in antibiotic susceptibility.

Actinomyces↗
PubMed1998

Conjugative transfer of the Escherichia coli-Clostridium perfringens shuttle vector pJIR1457 to Clostridium botulinum type A strains.

An RP4-oriT shuttle vector pJIR1457 originally developed for Clostridium perfringens was successfully transferred by conjugation from Escherichia coli to Clostridium botulinum type A strains and to a nontoxigenic C. botulinum type A-transposon Tn916 mutant strain lacking the entire toxin gene cluster. The light chain (LC) of botulinum toxin was highly expressed in the toxin deletion mutant strain from a pJIR1457 construct containing the recombinant botulinal gene for LC. This shuttle vector system will be valuable for genetic analysis of C. botulinum and will enable genetic manipulation and recombinant expression studies of botulinum neurotoxins as pharmaceutical agents.

Botulinum Toxins↗
PubMed1998

An interactive multimedia solution to learning removable partial denture design.

Surveys of the dental profession and dental laboratories show that up to 60% of cases received by laboratories have little or no input from dentists into the design of their patients' removable partial dentures (RPD). Thus, there is a clear mandate for dental schools to teach RPD design in new ways that will give their students confidence and allow their graduates to approach the task readily and easily. This computer-aided learning program addresses that need. In designing it, we have exploited the unique properties of computers as a learning tool. The program uses Quicktime VR to simulate a three-dimensional perspective of diagnostic casts. It is also possible to incorporate animated diagrams to illustrate various points. Control of timed answers promotes the use of inquiry-based learning and allows the program to be interactive and completely problem-oriented. Students can practice at their own pace, on a variety of virtual cases, and thus learn the craft behind the art of designing RPDs. With this foundation, discussions with experts in the field regarding the patients they treat in senior years and as graduates can be conducted at a higher and more productive level. Student evaluation so far has been very encouraging.

Computer-Assisted Instruction↗
PubMed1996

Activation of adenosine A3 receptors on macrophages inhibits tumor necrosis factor-alpha.

Murine macrophage-derived tumor necrosis factor alpha (TNF-alpha) gene expression has been shown to be dramatically induced by bacterial lipopolysaccharide, and to be dependent upon nuclear factor-kappa B (NF-kappa B) binding sites in its promoter for the lipopolysaccharide induction. Murine J774.1 macrophage cells were found to predominantly express the adenosine A3 receptor RNA relative to adenosine A1 receptor or adenosine A2 receptor RNA. Adenosine receptor agonists, in a dose-dependent manner characteristic of the adenosine A3 receptor, blocked the endotoxin induction of the TNF-alpha gene and TNF-alpha protein expression in the J774.1 macrophage cell line. The adenosine A3 receptor antagonist BW-1433 dose-dependently reversed this adenosine inhibitory effect on TNF-alpha gene expression. Thus, the binding of adenosine receptor agonists to the adenosine A3 receptor interrupts the endotoxin CD14 receptor signal transduction pathway and blocks induction of cytokine TNF-alpha, revealing a novel cross-talk between the murine adenosine A3 receptor and the endotoxin CD14 receptor in J774.1 macrophages.

Animals↗
PubMed1995

Development of bovine microsatellite markers from a microsatellite-enriched library.

A bovine genomic library enriched for DNA fragments bearing poly(dC-dA).poly(dG-dT) sequences was prepared and screened. Twenty-four clones bearing microsatellites were subject to sequence analysis and marker development as appropriate. Three of the 24 clones had microsatellites that were not evaluated for marker development owing to presence of a satellite element in two cases and limited repeat length in the third. Of the remaining 21 clones, all but one yielded polymorphic microsatellites. All but two of the polymorphic markers could be assigned to chromosomal locations by either linkage analysis or on the basis of X-linked inheritance as determined by heterozygosity limited to females. An unexpected clustering of markers was observed as 4 of the 20 polymorphic microsatellites mapped to a region of less than approximately 30 cM on Chromosome (Chr) 19, and four markers displayed X-linked inheritance.

Animals↗
PubMed1995

Polymerase chain reaction based genotyping for characterization of SLA-DQB and SLA-DRB alleles in domestic pigs.

Molecular genotyping of swine major histocompatibility complex SLA-DQB and SLA-DRB genes using polymerase chain reaction (PCR)-based amplification is described. Locus-specific oligonucleotide primers were designed for the analysis of expressed SLA genes by reverse transcription-polymerase chain reaction (RT-PCR). RT-PCR products were sequenced, and the information gained was used to design primers for PCR genotyping of the exon 2 (beta 1) region from genomic DNA templates. A single segregating amplification product was detected for both DQB and DRB in all animals. PCR products were digested with restriction enzymes. Seven SLA-DQB PCR-restriction fragment length polymorphism (RFLP) pattern types were observed for both HaeIII and RsaI that defined 14 SLA-DQB alleles. A total of seven SLA-DRB PCR-RFLP pattern types were defined using MspI (3 RFLP pattern types) and RsaI (6 RFLP pattern types). In order to demonstrate their universal utility, the primers were tested on genomic DNA samples from 10 different swine breeds. No breed-specific alleles were observed. These results show that locus-specific oligonucleotide primers and RFLP analysis provide a simple and rapid method for genotyping expressed SLA-DQB and SLA-DRB from genomic DNA.

Alleles↗
PubMed1995

Specific transcriptional inhibition of bone marrow-derived macrophage tumor necrosis factor-alpha gene expression and protein production using novel enantiomeric carbocyclic nucleoside analogues.

Tumor necrosis factor-alpha (TNF-alpha) is a powerful macrophage-derived proinflammatory cytokine, via both direct effects on host tissues as well as indirectly through the induction of other proinflammatory mediators, including interleukin- (IL) 1 beta and IL-6. Activation of murine bone marrow-derived macrophages (BMDM phi) with lipopolysaccharide (LPS) causes rapid expression of TNF-alpha, which as an autocrine factor enhances BMDM phi function through IL-1 beta and IL-6 production. In this study, we have examined the specific transcriptional inhibition of BMDM phi TNF-alpha using novel enantiomeric carbocyclic nucleoside analogues. BMDM phi were derived in vitro from murine bone marrow progenitors using colony stimulating factor-1 and treated with combinations of LPS (1-100 nG/ml) and the enantiomeric carbocyclic nucleoside (10-100 microM) analogues MDL 201, 112 (9-[(1S,3R)-cis-cyclopentan-3-ol]adenine); MDL 201,451 (9-[1R,3S)-cis-cyclopentan-3-ol]adenine); MDL 201,449 (9-[(1R,3R)-trans-cyclopentan-3-ol]adenine) and MDL 201,484 (9-[(1S,3S)-trans-cyclopentan-3-ol]adenine). Northern blot analysis showed that MDL 201,449 was the most effective agent in vitro at selectively inhibiting TNF-alpha. MDL 201,449 reduced TNF-alpha mRNA levels by nearly 50% for up to 4 hr after the simultaneous addition of LPS and the synthetic agent. In contrast, mRNA and secreted protein levels for IL-1 beta (measured by the D10.S bioassay) and mRNA for TNF-alpha receptor p60 and TNF-alpha receptor p80 were not significantly affected. Carbocyclic nucleoside analogues were effective when added to BMDM phi up-to 2 hr after LPS treatment and at concentrations as low as 10 microM. Regulation of BMDM phi IL-6 by carbocyclic nucleoside analogues in response to LPS appears to be both concentration and time dependent, because IL-6 mRNA and secreted protein levels were inhibited at only high drug concentrations (100 microM) and effective only at longer exposure times (+4 hr of incubation) to LPS. These data support the concept that M phi-derived proinflammatory cytokine gene expression is differentially, rather than coordinately, regulated by selective signal transduction and/or molecular pathways. Enantiomeric carbocyclic nucleoside analogues that specifically inhibit TNF-alpha may have therapeutic potential in inflammatory diseases, such as systemic inflammatory response syndrome, where TNF-alpha has been shown to have an important role in initiating the early stages of disease.

Adenine↗
PubMed1995

Interactive computer teaching/learning in the dental curriculum: partial denture design.

Interactive multimedia is an emerging industry with a broad domain. It operates from a base of computer technology to enable the production, distribution and sharing of ideas. The use of multimedia in the educational setting is in keeping with the concept of learning, rather than teaching and, with this in mind, a pilot program in interactive computer learning has been developed which is geared specifically to the requirements of junior students. The design sequence and philosophy used in this program follow the teaching at Sydney University Dental School. Its purpose is to facilitate students in mastering the technical aspects of denture framework design, leading them through a set sequence of steps from saddle identification to finish. Future plans, using more sophisticated equipment, are to develop more advanced programs for senior (fourth year) students, including applications for treatment planning and also the rudiments of osseointegrated implant work.

Australia↗
PubMed1993

The insulin receptor substrate (IRS-1) is a PEST protein that is susceptible to calpain degradation in vitro.

The insulin receptor substrate 1 (IRS-1) contains at least 11 sequence motifs that are rich in proline (P), glutamic acid (E), serine (S), and threonine (T), i.e., PEST regions. Proteins with PEST regions turn over rapidly. IRS-1 is degraded rapidly in vivo upon exposure of 3T3-L1 adipocytes to insulin. The intracellular, calcium-dependent, neutral proteases known as calpains are one possible mechanism by which IRS-1 may be degraded. To begin to investigate this possibility, purified exogenous calpain was shown to degrade IRS-1 in cell-free extracts from basal and insulin-treated cells and rat recombinant IRS-1 in vitro. Only two proteolytic fragments could be detected. One had a mol wt of approximately 79 kDa, arising from the C-terminus end, and the second had a mol wt of approximately 90 kDa arising from near the N-terminus, possibly a product of the same cleavage event, since the mol wt of IRS-1 from insulin-treated cells was approximately 170 kDa. These results suggest that IRS-1 may serve as a substrate for calpain in vivo, accounting for its rapid degradation.

3T3 Cells↗
PubMed1979

A comparison of amphotericin B alone and combined with flucytosine in the treatment of cryptoccal meningitis.

We compared amphotericin B therapy for cryptococcal meningitis with a newer regimen containing both amphotericin B and flucytosine. In 50 patients with 51 courses of therapy adherent to the protocol, 27 courses were with amphotericin B and 24 with the combination. Even though the combination regimen was given for only six weeks and amphotericin B for 10 weeks, the combination cured or improved more patients (16 vs 11), produced fewer failures or relapses (three vs. 11), more rapid sterilization of the cerebrospinal fluid (P less than 0.001) and less nephrotoxicity (P less than 0.05) than did amphotericin B alone. The number of deaths was the same (five) with each regimen. Adverse reactions to flucytosine occurred in 11 of 34 patients but were not life threatening. We conclude that combined flucytosine-amphoericin B therapy is the regimen of choice in cryptococcal meningitis.

Adolescent↗