Enhanced intracellular Ca2+ concentrations in Escherichia coli and Bacillus subtilis after addition of oligosaccharide elicitors.

Elicitation can lead to overproduction of secondary metabolites in plants and microbes. Potential changes in cytosolic Ca(2+) levels in bacteria were studied in response to elicitation. We report, for the first time, the effect of oligosaccharide elicitors on intracellular Ca(2+) levels. The apoaequorin gene was cloned into Escherichia coli DH5α and Bacillus subtilis 1604 cultures. Addition of elicitors, oligoguluronate and mannan oligosaccharides, to the cultures caused up to 11-fold increase in cytosolic Ca(2+) in E. coli and tenfold increase in B. subtilis. These increases in Ca(2+) levels could therefore contribute to the enhancement of secondary metabolite levels.

What information should we give to our orthognathic patients?

In 2002, whilst the author was an orthodontic registrar, she underwent a bimaxillary osteotomy to correct her class III malocclusion. This therefore gives her a unique perspective into the information she believes prospective patients undergoing similar procedures would find helpful and the choices of format to deliver that advice.

Expression and functional role of negative signalling regulators in tumour development and progression.

Alterations in intracellular signalling pathways such as the mitogen-activated protein kinases (MAPKs) are key common mechanisms of tumour development and progression. As such, there has been intense research into developing drugs that can inhibit or attenuate intracellular signalling. In recent years, there has been increasing recognition that the cell already has innate negative regulatory proteins that achieve this in normal homeostasis. These regulators provide a feedback inhibitory mechanism that controls the intensity and duration of activated signalling by exogenous stimuli. Members of this group include Raf kinase inhibitor protein 1, the MAPK phosphatases, the SPROUTY and SPRED families and similar expression to FGF. A number of studies have now demonstrated significant alterations in expression of negative regulators in malignant tissue in different cancer types. In functional studies, manipulated expression of these regulators has been shown to significantly influence tumour cell behaviour and phenotype. Here, we summarise the evidence for the functional expression of negative signalling regulators in tumour growth and progression and discuss their potential role as cancer biomarkers and targets for novel drug therapy.

Evidence for distinct alterations in the FGF axis in prostate cancer progression to an aggressive clinical phenotype.

Multiple fibroblast growth factor (FGF) axis alterations are known to occur in prostate cancer. Here we simultaneously profiled key components of this axis to determine their relevance in disease progression. An optimized immunohistochemistry protocol was used in expression analysis of FGF2, FGF8, FGFR1, FGFR4, and Sef (similar expression to FGF) in a single TMA of prostate cancer. FGF ligands and receptors were overexpressed in cancers compared to benign samples (p < 0.0001), while Sef expression was reduced (p < 0.0001). There was a positive association between higher grades and increased FGFR4 (p = 0.02), FGF2, and FGF8 (p = 0.002 and p < 0.0001). Sef expression was progressively lower with increasing grade (p = 0.005). Clinical stage was positively associated with FGF2, FGF8, and FGFR4 expression (p = 0.005, 0.03, and 0.012) but not with FGFR1 or Sef expression. Only reduced Sef was associated with bone metastasis (p = 0.02) and was also predictive of subsequent metastasis in initially localized tumours (p = 0.004). Down-regulation of Sef and increased FGFR4 were also the only independent variables associated with disease-specific survival (HR 1.73, p = 0.04 and HR 0.56, p = 0.01). In in vitro studies, silencing Sef enhanced the cell response to FGFs (p < 0.001) and substantially mitigated the effectiveness of an FGFR1 inhibitor. Conversely, increased Sef blocked the response to FGFs and had a comparable suppressive effect to the inhibitor. This study demonstrates that increased FGFR4 and reduced Sef may be critical FGF alterations associated with prostate cancer progression. Sef may also have a role in the tumour response to FGFR inhibition and warrants further investigation in this context.

Elicitation effects of oligosaccharides on the transcriptional level of bacitracin ABC transporter genes in Bacillus licheniformis.

Bacitracin ABC transporter comprises a self-resistance system in Bacillus licheniformis against its own product (bacitracin). Elicitation effects of carbohydrates on transcription level of bcrABC genes encoding bacitracin ABC transporter system were investigated through quantification by real-time PCR. Increases of 68% and 29% in bacitracin A yields after 36 h were achieved in cultures supplemented with oligoguluronate and oligoguluronate/mannan oligosaccharide combination. Cultures with combined elicitors showed 3, 2.6 and 3 transcriptional fold-changes in bcrA, bcrB and bcrC, respectively, compared to control. This could be due to a secondary effect caused by increased bacitracin A yield through elicitation.

Effect of oligosaccharide elicitors on bacitracin A production and evidence of transcriptional level control.

The role of oligosaccharides as biotic elicitors for the enhanced production of antibiotics and enzymes has been established in recent years. These findings could have significant promise for the pharmaceutical and biotechnology industries. Before the elicitors could be used at large-scale, reproducible production and high levels of enhancement are essential. In order to ensure the robustness of elicitation and exploit the process an understanding of this phenomenon at the molecular level is required. Bacitracin produced by Bacillus licheniformis is widely used as an animal feed additive and antibiotic against Gram-positive bacteria. In this work we report, the effect of single and multiple additions of different elicitors to B. licheniformis cultures for the production of bacitracin A. The results obtained showed that single addition of oligoguluronate to the cultures enhanced the levels of bacitracin A by 23.3%. Moreover, multiple elicitor addition of oligoguluronate and mannan oligosaccharide further enhanced the production of bacitracin A by 13.2% and 36.5% compared to single elicitor addition and control cultures, respectively. In addition, for the first time, it was found that elicitor supplementation of cultures increased the transcription level of the bacitracin biosynthetic genes bacABC, indicating transcriptional level control of the elicitor.

Management of postorthodontic demineralized white lesions with microabrasion: a quantitative assessment.

INTRODUCTION: The aim of this study was to quantify changes in postorthodontic demineralized enamel lesion surface areas after microabrasion. METHOD: The study group comprised 8 orthodontic patients (mean age,17.3 years) with multiple decalcified enamel lesions after fixed orthodontic therapy. Two demineralized areas were randomly selected for interventive treatment in each patient. Microabrasion was undertaken on these lesions by using a well-accepted 18% hydrochloric acid and pumice technique. Standardized intraoral images were taken of the lesions before and immediately after microabrasion. Image-processing software was used to quantify (mm2) the visible areas of the demineralized lesions before and after microabrasion. The total labial surface area of each tooth was also determined, and the area affected by demineralization was expressed as a percentage of total tooth surface. Images were reanalyzed a month later to determine the repeatability of the method. RESULTS: Microabrasion significantly reduced visible enamel demineralization (P < .001, paired t test). The mean reduction in lesion size after treatment was 83% (SD, 8.2%; range, 61%-92%). The quantification methodology was found to be highly repeatable with an intraclass correlation coefficient of 0.98. CONCLUSIONS: Microabrasion is an effective treatment approach for the cosmetic improvement of long-standing postorthodontic demineralized enamel lesions.

The diary of an orthognathic patient aged 30 3/4.

This article reports on the experiences of an orthodontist who has actually undergone combined orthodontic and orthognathic treatment. The aim is to give the reader an insight into not only what we, the orthodontists, fail to tell our orthognathic patients, but also what they fail to tell us.

Image analysis of oronasal fistulas in cleft palate patients acquired with an intraoral camera.

The aim of this study was to examine the clinical technique of using an intraoral camera to monitor the size of residual oronasal fistulas in cleft lip-cleft palate patients, to assess its repeatability on study casts and patients, and to compare its use with other methods. Seventeen plaster study casts of cleft palate patients with oronasal fistulas obtained from a 5-year series of 160 patients were used. For the clinical study, 13 patients presenting in a clinic prospectively over a 1-year period were imaged twice by the camera. The area of each fistula on each study cast was measured in the laboratory first using a previously described graph paper and caliper technique and second with the intraoral camera. Images were imported into a computer and subjected to image enhancement and area measurement. The camera was calibrated by imaging a standard periodontal probe within the fistula area. The measurements were repeated using a double-blind technique on randomly renumbered casts to assess the repeatability of measurement of the methods. The clinical images were randomly and blindly numbered and subjected to image enhancement and processing in the same way as for the study casts. Area measurements were computed. Statistical analysis of repeatability of measurement using a paired sample t test showed no significant difference between measurements, indicating a lack of systematic error. An intraclass correlation coefficient of 0.97 for the graph paper and 0.84 for the camera method showed acceptable random error between the repeated records for each of the two methods. The graph paper method remained slightly more repeatable. The mean fistula area of the study casts between each method was not statistically different when compared with a paired samples t test (p = 0.08). The methods were compared using the limits of agreement technique, which showed clinically acceptable repeatability. The clinical study of repeated measures showed no systematic differences when subjected to a t test (p = 0.109) and little random error with an intraclass correlation coefficient of 0.98. The fistula size seen in the clinical study ranged from 18.54 to 271.55 mm. Direct measurements subsequently taken on 13 patients in the clinic without study models showed a wide variation in the size of residual fistulas presenting in a multidisciplinary clinic. It was concluded that an intraoral camera method could be used in place of the previous graph paper method and could be developed for clinical and scientific purposes. This technique may offer advantages over the graph paper method, as it facilitates easy visualization of oronasal fistulas and objective fistulas size determination and permits easy storage of data in clinical records.