Early mechanisms of neutrophil activation and transmigration in acute lung injury.

Introduction:Neutrophil transmigration is multifactorial and primarily driven by selectins and ß2-integrins (CD11b/CD18), whose expression are dependent on the underlying stimulus. Ventilator-induced lung injury (VILI) results in a predominantly CD18-independent mechanism of neutrophil recruitment, while direct endotoxin-induced lung injury results from a CD18-dependent mechanism. We previously observed that lack of NADPH oxidases DUOX1 and DUOX2 resulted in reduced neutrophil influx in a VILI model of lung injury but had no influence on neutrophil influx after LPS exposure. Based on these observations, we hypothesized that DUOX1/DUOX2 are an important component of CD18-independent mechanisms of neutrophil recruitment in the lung. Methods:We exposed Duoxa -/- (KO) mice and Duoxa +/+ (WT) mice to either an intratracheal exposure of lipopolysaccharide (LPS/endotoxin)-or high tidal volume ventilation and compared expression of neutrophil markers between groups. WT mice (129S6/SvEvTac) were obtained from Taconic Biosciences (One Discovery Drive Suite 304; Rensselaer, NY 1244) and were allowed to acclimatize for one week prior to study enrollment. KO mice were generated as previously described [Grasberger 2012] and bred in-house on a 129S6 background. We provided positive-pressure ventilation at a tidal volume of 10 ml/kg with 2 cmH20 positive end-expiratory pressure (PEEP). Mice were assigned to groups consisting of KO (n = 5) and WT (n = 5) in each group and divided into non-ventilated, positive-pressure ventilation, or LPS IT exposure groups. Positive-pressure ventilation was instituted for 4-h using a FlexiVent (Flexiware 8.1, Scireq, Montreal, QC, Canada). Lipopolysaccharide (Salmonella enterica serotype tryphimurium L6143, Millipore Sigma) was administered via an intratracheal (IT) route at a dose of 0.1 mg/kg. Mice were humanely euthanized at 4-h post-injection consistent with the UC Davis IAUCAC-approved protocol. Results:As previously observed, neutrophilic influx into the airways was significantly impaired in the Duoxa -/- (KO) mice after VILI, but not after LPS exposure. LPS-induced lung injury resulted in upregulation of CD11b+ neutrophils and shedding of CD62L and CD162 regardless of DUOX expression, whereas VILI resulted in upregulation of CD49+ neutrophils in the Duoxa +/+ (WT) mice but not the Duoxa -/- (KO) mice. Conclusion:Our data suggest DUOX is required for CD18-independent mechanisms of neutrophil recruitment in the lung induced by acute lung injury, but not for canonical CD18depedent mechanisms after LPS exposure.

How does small group continuing medical education (CME) impact on practice for rural GPs and their patients, a mixed-methods study.

Studies which report outcomes of continuing medical education (CME) interventions for rural general practitioners (GPs) are limited. This mixed methods study recruited GPs from four CME small group learning (SGL) tutor groups based in different rural locations in the Republic of Ireland. A two-hour teaching module on deprescribing in older patients was devised and implemented. Assessment of educational outcomes was via questionnaires, prescribing audits and qualitative focus groups. All GPs (n = 43) in these CME-SGL groups agreed to participate, 27 of whom (63%) self-identified as being in rural practice. Rural GPs were more likely to be male (56%), in practice for longer (19 years), and attending CME for longer (13 years). The questionnaires indicated learning outcomes were achieved knowledge increased immediately after the education, and was maintained 6 months later. Twenty-four GPs completed audits involving 191 patients. Of these, 152 (79.6%) were de-prescribed medication. In the qualitative focus groups, GPs reported sharing experiences with their peers during CME-SGL helped them to improve patient care and ensured that clinical practice is more consistent across the group. For rural GPs, CME-SGL involving discussion of cases and the practical implementation of guidelines, associated with audit, can lead to changes in patient care.

Does locally delivered small group continuing medical education (CME) meet the learning needs of rural general practitioners?

The World Health Organisation policy of improving retention of rural health care practitioners recommended that continuing medical education (CME) programmes addressing their needs should be accessible and delivered where they live and work. This cross-sectional study involved a self-administered anonymous questionnaire completed by GPs attending CME small groups (CME-SGL) in Ireland. All GPs attending CME-SGL in a one-month period were invited to complete the questionnaire which examined demographic details, distance to travel to educational meetings/nearest regional hospital, barriers to accessing continuing education, whether CME-SGL met their educational needs, morale and professional isolation. A total of 1,686 responses were collected, of which 332 (19.6%) were from rural GPs. Of these, 289 (87%) reported that their educational needs were fully or mostly met by attending CME-SGL. Compared to urban doctors, rural GPs had further to travel to CME-SGL meetings, were further from the nearest regional hospital, and reported increased barriers to accessing continuing education. Rural GPs reported lower morale and greater levels of professional isolation. Despite considerable barriers to accessing continuing education, rural GPs reported that CME-SGL meets their educational needs. Future research should focus on the potential positive impact this may have on professional isolation and morale.

The introduction of mandatory CPD for newly state registered diagnostic radiographers: An Irish perspective.

INTRODUCTION: Irish diagnostic radiography has undergone significant change with the dual introduction of state registration and mandatory Continuing Professional Development (CPD) in October 2015. We aimed to investigate motivators and barriers around CPD participation, mechanisms of CPD delivery and confidence of radiographers in using e-learning. METHODS: A questionnaire distributed nationally during this period captured Radiographer opinion through the use of closed and open questions. The questionnaire was distributed in hard copy and online formats depending on site preference. RESULTS: 71% of centres participated, rendering 453 responses in total from a possible 1222 respondents employed in those sites at the time of the survey. A varied range of ages and post qualification experience were represented. Respondents indicated use of several CPD options with the majority considering CPD to be important, very important or critical. Social media as a mechanism of CPD delivery was considered acceptable by 48%, while online learning elicited responses ranging from not confident to absolutely confident. Top motivators for CPD activity included interest, developing new knowledge and competency. Principal barriers included funding, time allocation and location issues. CONCLUSION: This study identified Radiographer desire to undertake CPD and the need for developing online CPD offerings. An overall positive perception towards CPD was noted however barriers were identified which require specific redress strategies.

Irish Medical Students Understanding of the Intern Year.

Upon completion of medical school in Ireland, graduates must make the transition to becoming interns. The transition into the intern year may be described as challenging as graduates assume clinical responsibilities. Historically, a survey of interns in 1996 found that 91% felt unprepared for their role. However, recent surveys in 2012 have demonstrated that this is changing with preparedness rates reaching 52%. This can be partially explained by multiple initiatives at the local and national level. Our study aimed evaluate medical student understanding of the intern year and associated factors. An online, cross-sectional survey was sent out to all Irish medical students in 2013 and included questions regarding their understanding of the intern year. Two thousand, two hundred and forty-eight students responded, with 1,224 (55.4%) of students agreeing or strongly agreeing that they had a good understanding of what the intern year entails. This rose to 485 (73.7%) among senior medical students. Of junior medical students, 260 (42.8%) indicated they understood what the intern year, compared to 479 (48.7%) of intermediate medical students. Initiatives to continue improving preparedness for the intern year are essential in ensuring a smooth and less stressful transition into the medical workforce.

The relevance of basic sciences in undergraduate medical education.

BACKGROUND: Evolving and changing undergraduate medical curricula raise concerns that there will no longer be a place for basic sciences. National and international trends show that 5-year programmes with a pre-requisite for school chemistry are growing more prevalent. National reports in Ireland show a decline in the availability of school chemistry and physics. AIM: This observational cohort study considers if the basic sciences of physics, chemistry and biology should be a prerequisite to entering medical school, be part of the core medical curriculum or if they have a place in the practice of medicine. METHODS: Comparisons of means, correlation and linear regression analysis assessed the degree of association between predictors (school and university basic sciences) and outcomes (year and degree GPA) for entrants to a 6-year Irish medical programme between 2006 and 2009 (n = 352). RESULTS: We found no statistically significant difference in medical programme performance between students with/without prior basic science knowledge. The Irish school exit exam and its components were mainly weak predictors of performance (-0.043 ≥ r ≤ 0.396). Success in year one of medicine, which includes a basic science curriculum, was indicative of later success (0.194 ≥ r (2) ≤ 0.534). CONCLUSIONS: University basic sciences were found to be more predictive than school sciences in undergraduate medical performance in our institution. The increasing emphasis of basic sciences in medical practice and the declining availability of school sciences should mandate medical schools in Ireland to consider how removing basic sciences from the curriculum might impact on future applicants.

Ataxia telangiectasia: more variation at clinical and cellular levels.

Ataxia telangiectasia (A-T) is a rare recessively inherited disorder resulting in a progressive neurological decline. It is caused by biallelic mutation of the ATM gene that encodes a 370 kDa serine/threonine protein kinase responsible for phosphorylating many target proteins. ATM is activated by auto(trans)phosphorylation in response to DNA double strand breaks and leads to the activation of cell cycle checkpoints and either DNA repair or apoptosis as part of the cellular response to DNA damage. The allelic heterogeneity in A-T is striking. While the majority of mutations are truncating, leading to instability and loss of the ATM protein from the allele, a significant proportion of patients carry one of a small number of mutations that are either missense or leaky splice site mutations resulting in retention of some ATM with activity. The allelic heterogeneity in ATM, therefore, results in an equally striking clinical heterogeneity. There is also locus heterogeneity because mutation of the MRE11 gene can cause an obvious A-T like disorder both clinically and also at the cellular level and mutation of the RNF168 gene results in a much milder clinical phenotype, neurologically, with the major clinical feature being an immunological defect.

Medical students' views on selection tools for medical school--a mixed methods study.

It is important to ensure that the tools used in Medical School selection are acceptable to students and applicants. A questionnaire was administered to year 1 medical students in 2010 to determine the suitability of a variety of selection tools and the acceptability of HPAT-Ireland in particular. There were 291 respondents a 77% response rate representing approximately one third of all school leaver entrants that year. While the majority 285 (98%) were in favour of using school leaving examinations there was also support for the use of interviews 215 (74%) and other tools. Three quarters of Irish respondents 159 (76%) agreed that HPAT-Ireland is a fair test overall however section 3 (non-verbal reasoning) appeared less acceptable and relevant than other sections. A little over half had taken a preparatory HPAT-Ireland course 112 (54%). Medical school applicants appear to accept the use of non-traditional tools in the selection process.

Severe reaction to radiotherapy for breast cancer as the presenting feature of ataxia telangiectasia.

BACKGROUND: Severe early and late radiation reaction to radiotherapy is extremely rare in breast cancer patients. Such a reaction prompted an investigation into a 44-year-old mother (patient A-T213). METHODS: A neurological examination was performed and blood lymphocytes and skin fibroblasts were assessed for radiosensitivity chromosomally and by colony-forming assay. The ATM gene was sequenced and ATM mutations modelled by site-directed mutagenesis. The ATM kinase activity was also assessed. RESULTS: Patient A-T213 was normally ambulant with no ataxia and minimal other neurological features. T lymphocytes and skin fibroblasts were unusually radiosensitive, although less sensitive than in classical ataxia telangiectasia (A-T). A lymphoblastoid cell line and skin fibroblasts expressed ATM protein with some retained kinase activity. One missense ATM mutation c.8672G>A (p.Gly2891Asp) and a c.1A>G substitution were identified. In the modelling system, the p.Gly2891Asp mutant protein was expressed and shown to have residual ATM kinase activity. CONCLUSION: Patient A-T213 has a milder form of A-T with biallelic ATM mutations, which may have contributed to breast cancer development, and certainly caused the severe radiation reaction. Ataxia telangiectasia should be investigated as a potential cause of untoward severe early and late radiation reactions in breast cancer patients.

Lymphoid tumours and breast cancer in ataxia telangiectasia; substantial protective effect of residual ATM kinase activity against childhood tumours.

BACKGROUND: Immunodeficiency in ataxia telangiectasia (A-T) is less severe in patients expressing some mutant or normal ATM kinase activity. We, therefore, determined whether expression of residual ATM kinase activity also protected against tumour development in A-T. METHODS: From a total of 296 consecutive genetically confirmed A-T patients from the British Isles and the Netherlands, we identified 66 patients who developed a malignant tumour; 47 lymphoid tumours and 19 non-lymphoid tumours were diagnosed. We determined their ATM mutations, and whether cells from these patients expressed any ATM with residual ATM kinase activity. RESULTS: In childhood, total absence of ATM kinase activity was associated, almost exclusively, with development of lymphoid tumours. There was an overwhelming preponderance of tumours in patients <16 years without kinase activity compared with those with some residual activity, consistent with a substantial protective effect of residual ATM kinase activity against tumour development in childhood. In addition, the presence of eight breast cancers in A-T patients, a 30-fold increased risk, establishes breast cancer as part of the A-T phenotype. CONCLUSION: Overall, a spectrum of tumour types is associated with A-T, consistent with involvement of ATM in different mechanisms of tumour formation. Tumour type was influenced by ATM allelic heterogeneity, residual ATM kinase activity and age.

Phage anti-immunocomplex assay for clomazone: two-site recognition increasing assay specificity and facilitating adaptation into an on-site format.

The impact of the use of herbicides in agriculture can be minimized by compliance with good management practices that reduce the amount used and their release into the environment. Simple tests that provide real time on-site information about these chemicals are a major aid for these programs. In this work, we show that phage anti-immunocomplex assay (PHAIA), a method that uses phage-borne peptides to detect the formation of antibody-analyte immunocomplexes, is an advantageous technology to produce such field tests. A monoclonal antibody to the herbicide clomazone was raised and used in the development of conventional competitive and noncompetitive PHAIA immunoassays. The sensitivity attained with the PHAIA format was over 10 times higher than that of the competitive format. The cross-reactivity of the two methods was also compared using structurally related compounds, and we observed that the two-site binding of PHAIA "double-checks" the recognition of the analyte, thereby increasing the assay specificity. The positive readout of the noncompetitive PHAIA method allowed adaptation of the assay into a rapid and simple format where as little as 0.4 ng/mL clomazone (more than 10-fold lower than the proposed standard) in water samples from a rice field could be easily detected by simple visual inspection.

Water quality issues and infant diarrhoea in a South American province.

In the province of Salta, in the Northwest region of Argentina, almost two-thirds of the population live in absolute poverty, and diseases associated with poverty are rampant. Almost 12% of the total population of the province are children below 5 years of age; almost half of these infants are living in situations where the basic necessities are not available. Primitive sanitary conditions, including widespread contamination of available water supplies with pathogens, contribute to a major public health problem. Infant mortality was 17% higher for Salta than for Argentina as a whole in 2001. A major cause of death for these children is infectious disease, especially respiratory and intestinal diseases. In Salta, more than half of the total population of infants is affected by diarrhoea annually. The infectious pathogens are diverse: bacteria (predominantly in spring and summer), viruses (especially in the winter) and parasites (endemic in some situations). This paper evaluates current methods used to test for the presence of pathogens in drinking water; discusses why these methods are less than adequate; documents an episode of contamination in a local water supply source; and suggests appropriate methods that can be used to better address this major public health issue effectively.

Are exposure index values consistent in clinical practice? A multi-manufacturer investigation.

The advent of digital radiography poses the risk of unnoticed increases in patient dose. Manufacturers have responded to this by offering an exposure index (EI) value to the clinician. Whilst the EI value is a measure of the air kerma at the detector surface, it has been recommended by international agencies as a method of monitoring radiation dose to the patient. Recent studies by the group have shown that EI values are being used in clinical practice to monitor radiation dose and assess image quality. This study aims to compare the clinical consistency of the EI value in computed radiography (CR) and direct digital radiography (DR) systems. An anthropormorphic phantom was used to simulate four common radiographic examinations: skull, pelvis, chest and hand. These examinations were chosen as they provide contrasting exposure parameters, image detail and radiation dose measurements. Four manufacturers were used for comparison: Agfa Gaevert CR, Carestream CR, Philips Digital Diagnost DR and Siemens DR. For each examination, the phantom was placed in the optimal position and exposure parameters were chosen in accordance with European guidelines and clinical practice. Multiple exposures were taken and the EI recorded. All exposure parameters and clinical conditions remained constant throughout. For both DR systems, the EI values remained consistent throughout. No significant change was noted in any examination. In both CR systems, there were noteworthy fluctuations in the EI values for all examinations. The largest for the Agfa system was a variation of 1.88-2.21 for the skull examination. This represents to the clinician a doubling of detector dose, despite all exposure parameters remaining constant. In the Kodak system, the largest fluctuation was seen for the chest examination where the EI ranged from 2560 to 2660, representing approximately an increase of 30 % in radiation dose, despite consistent parameters. The fluctuations seen with the CR systems are most likely due to image processing delay, replacing of the imaging plate and calibration factors. Fluctuations in EI values may result in confusion to the clinician and unnecessary repeat examinations. The reliability of EI values as a feedback mechanism for CR is also questionable.

Clinical spectrum of ataxia-telangiectasia in adulthood.

OBJECTIVE: To describe the phenotype of adult patients with variant and classic ataxia-telangiectasia (A-T), to raise the degree of clinical suspicion for the diagnosis variant A-T, and to assess a genotype-phenotype relationship for mutations in the ATM gene. METHODS: Retrospective analysis of the clinical characteristics and course of disease in 13 adult patients with variant A-T of 9 families and 6 unrelated adults with classic A-T and mutation analysis of the ATM gene and measurements of ATM protein expression and kinase activity. RESULTS: Patients with variant A-T were only correctly diagnosed in adulthood. They often presented with extrapyramidal symptoms in childhood, whereas cerebellar ataxia appeared later. Four patients with variant A-T developed a malignancy. Patients with classic and variant A-T had elevated serum alpha-fetoprotein levels and chromosome 7/14 rearrangements. The mildest variant A-T phenotype was associated with missense mutations in the ATM gene that resulted in expression of some residual ATM protein with kinase activity. Two splicing mutations, c.331 + 5G>A and c.496 + 5G>A, caused a more severe variant A-T phenotype. The splicing mutation c.331 + 5G>A resulted in less ATM protein and kinase activity than the missense mutations. CONCLUSIONS: Ataxia-telangiectasia (A-T) should be considered in patients with unexplained extrapyramidal symptoms. Early diagnosis is important given the increased risk of malignancies and the higher risk for side effects of subsequent cancer treatment. Measurement of serum alpha-fetoprotein and chromosomal instability precipitates the correct diagnosis. There is a clear genotype-phenotype relation for A-T, since the severity of the phenotype depends on the amount of residual kinase activity as determined by the genotype.

Rheumatoid arthritis: a novel radiographic projection for hand assessment.

Rheumatoid arthritis (RA) is the most common form of inflammatory disease, affecting 1-2% of the population. Posteroanterior (PA) and Brewerton projections are well established in radiographic practice for scoring and monitoring RA, but there is little evidence to demonstrate the diagnostic efficacy of these techniques. This work, by varying the positioning of a cadaveric hand, investigates whether an alternative radiographic projection could yield greater diagnostic information than the traditional techniques. Phase I of the study evaluated moving the hand 15 degrees from the anteroposterior position and then in 5 degrees increments in four directions: medial rotation, lateral rotation, flexion of the wrist and extension of the wrist. Phase II of the study took the optimum projections from Phase I and further manipulated these positions in a direction at right angles to the original position. Images were scored based on joint space visualisation in 29 joints. Results demonstrated that significantly higher diagnostic efficacy was evident with 15 degrees lateral rotation of the hand or 15 degrees flexion at the wrist compared to the Brewerton projection. Either projection is recommended, but on the basis of patient comfort, the latter of these novel positions, now known as the UCD projection, was chosen as the optimum procedure to replace the Brewerton projection. The value of using cadavers for the establishment of optimum radiographic procedures is highlighted.

High-throughput method for ranking the affinity of peptide ligands selected from phage display libraries.

The use of phage display peptide libraries allows rapid isolation of peptide ligands for any target selector molecule. However, due to differences in peptide expression and the heterogeneity of the phage preparations, there is no easy way to compare the binding properties of the selected clones, which operates as a major "bottleneck" of the technology. Here, we present the development of a new type of library that allows rapid comparison of the relative affinity of the selected peptides in a high-throughput screening format. As a model system, a phage display peptide library constructed on a phagemid vector that contains the bacterial alkaline phosphatase gene (BAP) was selected with an antiherbicide antibody. Due to the intrinsic switching capacity of the library, the selected peptides were transferred "en masse" from the phage coat protein to BAP. This was coupled to an optimized affinity ELISA where normalized amounts of the peptide-BAP fusion allow direct comparison of the binding properties of hundreds of peptide ligands. The system was validated by plasmon surface resonance experiments using synthetic peptides, showing that the method discriminates among the affinities of the peptides within 3 orders of magnitude. In addition, the peptide-BAP protein can find direct application as a tracer reagent.

A method to establish the relationship between chronological age and stage of union from radiographic assessment of epiphyseal fusion at the knee: an Irish population study.

Characteristic changes during epiphyseal union provide a skeletal age, which when compared with age-based standards provides an estimation of chronological age. Currently there are no data on epiphyseal union for the purposes of age estimation specific to an Irish population. This cross-sectional study aims to investigate the relationship between stage of epiphyseal union at the knee joint and chronological age in a modern Irish population. A novel radiographic method that sub-divides the continuum of development into five specific stages of union is presented. Anteroposterior and lateral knee radiographs of 148 males and 86 females, aged 9-19 years, were examined. Fusion was scored as Stage 0, non-union; Stage 1, beginning union; Stage 2, active union; Stage 3, recent union; or Stage 4, complete union. Stage of epiphyseal union is correlated with chronological age in both males and females. Mean age gradually increases with each stage of union and also varies between male and female subjects. A statistically significant difference in mean age was recorded between stages when compared to the previous stage, for the three epiphyses. Irish children are comparable to those from previously published studies with epiphyseal union in females occurring earlier than males. A significant difference was noted between the mean age of union for males and females for each of Stages 1 and 2 for the femur and Stages 0, 1, 2 and 3 for the tibia and the fibula. The results also suggest that the stages of union occur at earlier ages in this Irish population. Implementation of standardized methodology is necessary to investigate if this is due to a secular or population variation in maturation or to a methodology which clearly identifies five stages of union.

Poster - Thurs Eve-05: An assessment of PDDs and outputs predicted by a Monte Carlo-based treatment planning system for electron beams.

Monte Carlo simulation is currently considered to be the most accurate method of calculating dose distributions for electron beam therapy, and commercial treatment planning software using simplified macro Monte Carlo is available for electron treatment planning. In this work, Eclipse V8.1.18 is being investigated in preparation for the clinical use of CT-based electron treatment planning. Water tank measurements of percentage depth doses (PDDs) and absolute outputs at depth of maximum dose (Zmax ) under different geometric conditions are compared to the results calculated by Eclipse. The measurements are carried out for a range of electron energies (6, 9, 12, and 16 MeV) for the standard open field (10×10 cm2 ) and for circular cutouts (2, 3, and 6 cm diameters) at SSD of 100 cm. In addition, extended SSDs (105 and 110 cm) and oblique beam incident (gantry 345 degree) for the open field and 3 cm diameter cutout are measured and compared to Eclipse. For PDDs, the results predicted by Eclipse are generally acceptable, falling mostly within 5% of those measured in water. For output, the results predicted by Eclipse are similar, falling mostly within 3% of those measured in water. We observed the greatest differences between Eclipse and measurements near the water surface and in high dose gradients for PDDs. A similar observation is noted for a small field in the case of outputs.

Polyclonal antibody-based noncompetitive immunoassay for small analytes developed with short peptide loops isolated from phage libraries.

To date, there are a few technologies for the development of noncompetitive immunoassays for small molecules, the most common of which relies on the use of anti-immunocomplex antibodies. This approach is laborious, case specific, and relies upon monoclonal antibody technology for its implementation. We recently demonstrated that, in the case of monoclonal antibody-based immunoassays, short peptide loops isolated from phage display libraries can be used as substitutes of the anti-immunocomplex antibodies for noncompetitive immunodetection of small molecules. The aim of this work was to demonstrate that such phage ligands can be isolated even when the selector antibodies are polyclonal in nature. Using phenoxybenzoic acid (PBA), a major pyrethroid metabolite, as a model system, we isolated the CFNGKDWLYC peptide after panning a cyclic peptide library on the PBA/anti-PBA immunocomplex. The sensitivity of the noncompetitive enzyme-linked immunosorbent assay (ELISA) setup with this peptide was 5-fold (heterologous) or 400-fold (homologous) higher than that of the competitive assay setup with the same antibody. Phage anti-immunocomplex assay (PHAIA) was also easily adapted into a rapid and highly sensitive dipstick assay. The method not only provides a positive readout but also constitutes a major shortcut in the development of sensitive polyclonal-based assays, avoiding the need of synthesizing heterologous competing haptens.