Elucidation of the K32 Capsular Polysaccharide Structure and Characterization of the KL32 Gene Cluster of Acinetobacter baumannii LUH5549.

Capsular polysaccharide (CPS), isolated from Acinetobacter baumannii LUH5549 carrying the KL32 capsule biosynthesis gene cluster, was studied by sugar analysis, Smith degradation, and one- and two-dimensional 1H and 13C NMR spectroscopy. The K32 CPS was found to be composed of branched pentasaccharide repeats (K units) containing two residues of ß-D-GalpNAc and one residue of ß-D-GlcpA (ß-D-glucuronic acid) in the main chain and one residue each of ß-D-Glcp and α-D-GlcpNAc in the disaccharide side chain. Consistent with the established CPS structure, the KL32 gene cluster includes genes for a UDP-glucose 6-dehydrogenase (Ugd3) responsible for D-GlcA synthesis and four glycosyltransferases that were assigned to specific linkages. Genes encoding an acetyltransferase and an unknown protein product were not involved in CPS biosynthesis. Whilst the KL32 gene cluster has previously been found in the global clone 2 (GC2) lineage, LUH5549 belongs to the sequence type ST354, thus demonstrating horizontal gene transfer between these lineages.

Improving the finite element model accuracy of tissue engineering scaffolds produced by selective laser sintering.

In bone tissue engineering, both geometrical and mechanical properties of a scaffold play a major part in the success of the treatment. The mechanical stresses and strains that act on cells on a scaffold in a physiological environment are a determining factor on the subsequent tissue formation. Computational models are often used to simulate the effect of changes of internal architectures and external loads applied to the scaffold in order to optimise the scaffold geometry for the prospective implantation site. Finite element analysis (FEA) based on computer models of the scaffold is a common technique, but would not take into account actual inaccuracies due to the manufacturing process. Image based FEA using CT scans of fabricated scaffolds can provide a more accurate analysis of the scaffold, and was used in this work in order to accurately simulate and predict the mechanical performance of bone tissue engineering scaffolds, fabricated using selective laser sintering (SLS), with a view to generating a methodology that could be used to optimise scaffold design. The present work revealed that an approach that assumes isotropic properties of SLS fabricated scaffolds will lead to inaccurate predictions of the FE model. However, a dependency of the grey value of the CT scans and the mechanical properties was discovered, which may ultimately lead to accurate FE models without the need of experimental validation.

Reconstructing the fall: individual, behavioural and contextual factors associated with falls in individuals with intellectual disability.

BACKGROUND: Falls are a significant and recurrent problem for individuals with intellectual disability (ID). There has been little exploration of the fall event from the perspective of the individual who falls or their carers. Research has focused predominantly on personal risk factors, leaving the behavioural and contextual analysis of falls much less understood. This study aimed to identify these additional factors as well as briefly explore the fall experience for individuals and their carers. METHOD: A qualitative design was used incorporating fall reconstructions and ethnographic-style interviews conducted in the home setting. Nine people with ID and their carers/family member participated: five pairs were living at home and four were in out-of-family-home settings. Interviews were recorded, transcribed verbatim and major themes identified via thematic analysis. RESULTS: We identified 17 themes that contributed to falls and fell under the three headings of individual, behavioural or contextual factors. Themes include decreased physical capacity, unsafe behaviours, limited hazard awareness and the impact of others in the home on an individual's fall behaviours. Additionally, families and individuals identified a number of consequences and adaptations which they implemented to alleviate possible fall impact. CONCLUSIONS: Qualitative interviews, observational methods and carer assistance are valuable in offering insight into understanding the individual, behavioural and contextual factors associated with falls in people with ID. The fall reconstruction technique may be a useful supplement when evaluating intrinsic risk in programmes designed to reduce falls.

Planning dementia services: new estimates of current and future prevalence rates of dementia for Ireland.

OBJECTIVES: The paper provides new estimates of dementia prevalence at a national and local level in Ireland and new projections of future numbers of people with dementia. METHODS: The prevalence of dementia at a national and local level has been calculated by applying European Collaboration on Dementia (EuroCoDe) prevalence rates to data from the Census of Population 2006. The National Disability Survey has been used to estimate the number of people with Down syndrome and dementia. Projections of future numbers of people with dementia have been calculated by applying EuroCoDe prevalence rates to the most recently available population projections from the Central Statistics Office (CSO). RESULTS: It is estimated that there were 41 740 people with dementia in Ireland in 2006. Estimates show that there are clear regional differences in prevalence of dementia across Ireland, with the largest proportion of people with dementia in the West of Ireland, and the Dublin North Eastern region having the lowest share of dementia. Our best estimate is that there are 700 people with Down syndrome and dementia in Ireland. Applying EuroCoDe prevalence rates to the most recent CSO population projections shows that the prevalence of dementia in Ireland will increase to between 67 493 and 70 000 in 2021 and to between 140 580 and 147 000 in 2041. CONCLUSIONS: Although there are several limitations to these estimates, the data provide timely and useful information for planning effective health and social care services, as well as raising public and professional awareness about dementia at a national level.

An amelanotic malignant melanoma masquerading as hypergranulation tissue.

We report a case of amelanotic malignant melanoma occurring in the nail sulcus of the hallux, which on initial presentation was mistaken for hypergranulation tissue due to an in-growing toenail. We highlight the importance of this differential diagnosis as such lesions can have serious sequelae.

Finite element predictions compared to experimental results for the effective modulus of bone tissue engineering scaffolds fabricated by selective laser sintering.

A current challenge in bone tissue engineering is to create scaffolds with suitable mechanical properties, high porosity, full interconnectivity and suitable pore size. In this paper, polyamide and polycaprolactone scaffolds were fabricated using a solid free form technique known as selective laser sintering. These scaffolds had fully interconnected pores, minimized strut thickness, and a porosity of approximately 55%. Tensile and compression tests as well as finite element analysis were carried out on these scaffolds. It was found that the values predicted for the effective modulus by the FE model were much higher than the actual values obtained from experimental results. One possible explanation for this discrepancy, viz. the surface roughness of the scaffold and the presence of micropores in the scaffold struts, was investigated with a view to making recommendations on improving FE model configurations for accurate effective property predictions.

The attitudes and practices of general practitioners regarding dementia diagnosis in Ireland.

BACKGROUND: The diagnosis of dementia poses difficulties for general practitioners (GPs) particularly when access to specialist diagnostic services is delayed. Ireland is soon to witness an increase in numbers of people presenting with dementia, yet little is known about the service needs of GPs when attempting to diagnose dementia. OBJECTIVES: To detail the service needs of GPs, especially their training needs, access to diagnostic resources such as CT and MRI brain scans and access to specialist services such as Old Age Psychiatry (OAP), Geriatric Medicine (GM) and Neuropsychology (NP). METHODS: The paper is based on survey data collected from a sample of GPs (n = 300) registered with the Irish College of General Practitioners (ICGP) and on qualitative data collected from a Focus Group (n = 7). RESULTS: GPs were more likely to blame themselves than either the health care system, their patients or family members for the late presentation of dementia in primary care. Stigma was a major obstacle preventing GPs from being more proactive in this area. Rural GPs felt geographically disadvantaged accessing diagnostic services and both rural and urban GPs experienced considerable time delays accessing specialist diagnostic services. CONCLUSIONS: Findings provide compelling evidence that training and access to diagnostic services are only two of several different structural and ideological obstacles that GPs encounter when attempting to diagnose dementia. Future educational supports for GPs need to be developed which concentrate on these areas.

Expression microarray analysis of papillary thyroid carcinoma and benign thyroid tissue: emphasis on the follicular variant and potential markers of malignancy.

The most common sub-variant of papillary thyroid carcinoma (PTC) is the so-called follicular variant (FVPTC), which is a particularly problematic lesion and can be challenging from a diagnostic viewpoint even in resected lesions. Although fine needle aspiration cytology is very useful in the diagnosis of PTC, its accuracy and utility would be greatly facilitated by the development of specific markers for PTC and its common variants. We used the recently developed Applied Biosystems 1700 microarray system to interrogate a series of 11 benign thyroid lesions and conditions and 14 samples of PTC (six with classic morphology and eight with follicular variant morphology). TaqMan(R) reverse transcriptase-polymerase chain reaction was used to validate the expression portfolios of 50 selected transcripts. Our data corroborates potential biomarkers previously identified in the literature, such as LGALS3, S100A11, LYN, BAX, and cluster of differentiation 44 (CD44). However, we have also identified numerous transcripts never previously implicated in thyroid carcinogenesis, and many of which are not represented on other microarray platforms. Diminished expression of metallothioneins featured strongly among these and suggests a possible role for this family as tumour suppressors in PTC. Fifteen transcripts were significantly associated with FVPTC morphology. Surprisingly, these genes were associated with an extremely narrow repertoire of functions, including the major histocompatibility complex and cathepsin families.

Continuous passive motion applied to whole joints stimulates chondrocyte biosynthesis of PRG4.

UNLABELLED: Continuous passive motion (CPM) is currently a part of patient rehabilitation regimens after a variety of orthopedic surgical procedures. While CPM can enhance the joint healing process, the direct effects of CPM on cartilage metabolism remain unknown. Recent in vivo and in vitro observations suggest that mechanical stimuli can regulate articular cartilage metabolism of proteoglycan 4 (PRG4), a putative lubricating and chondroprotective molecule found in synovial fluid and at the articular cartilage surface. OBJECTIVES: (1) Determine the topographical variation in intrinsic cartilage PRG4 secretion. (2) Apply a CPM device to whole joints in bioreactors and assess effects of CPM on PRG4 biosynthesis. METHODS: A bioreactor was developed to apply CPM to bovine stifle joints in vitro. Effects of 24h of CPM on PRG4 biosynthesis were determined. RESULTS: PRG4 secretion rate varied markedly over the joint surface. Rehabilitative joint motion applied in the form of CPM regulated PRG4 biosynthesis, in a manner dependent on the duty cycle of cartilage sliding against opposing tissues. Specifically, in certain regions of the femoral condyle that were continuously or intermittently sliding against meniscus and tibial cartilage during CPM, chondrocyte PRG4 synthesis was higher with CPM than without. CONCLUSIONS: Rehabilitative joint motion, applied in the form of CPM, stimulates chondrocyte PRG4 metabolism. The stimulation of PRG4 synthesis is one mechanism by which CPM may benefit cartilage and joint health in post-operative rehabilitation.

TRAIL sensitisation by arsenic trioxide is caspase-8 dependent and involves modulation of death receptor components and Akt.

The majority of leukaemic cells are resistant to apoptosis induced by tumour necrosis factor-related apoptosis-inducing ligand (TRAIL). Here, we show that sublethal concentrations of arsenic trioxide (ATO) specifically enhanced TRAIL-induced apoptosis in leukaemic but not in other tumour cell lines. The combination of ATO and TRAIL synergistically enhanced cleavage of caspase-8, which was blocked by the caspase inhibitor IETD.fmk as well as in cells deficient for caspase-8, suggesting a requirement for the death-inducing signalling complex. Arsenic trioxide led to increased cell surface expression of DR5 (death receptor 5), inhibition of the serine/threonine kinase Akt and downregulation of the short isoform of FLIP (FLICE-inhibitory protein, FLIPS). Inhibition of the phosphatidylinositol 3 kinase (PI3K) was equally efficient in sensitising leukaemic cells to TRAIL with similar effects on DR5 and FLIPS expression, suggesting that ATO may in part act through inhibition of the PI3K/Akt signalling pathway. These results indicate that the enhancement in TRAIL-mediated apoptosis induced by ATO is due to alteration in the levels of multiple components and regulators of the death receptor-mediated pathway. These findings offer a promising and novel strategy involving a combination of TRAIL and ATO, or more specific Akt inhibitors in the treatment of various haematopoietic malignancies.

ret/PTC and BRAF act as distinct molecular, time-dependant triggers in a sporadic Irish cohort of papillary thyroid carcinoma.

The purpose of this study was to assess BRAF mutation rates in various thyroid tissues and to investigate if concomitant mutations with ret/PTC activation occurred in inflammatory and neoplastic lesions. To this end, we developed a novel Taqman based screening assay for the common T1799A BRAF mutation. Heterozygous T1799A mutations were detected in 13 of 34 (44%) papillary thyroid carcinomas (PTCs) tested. No such mutations were detected in the other tissue types tested. Concomitant presence of both oncogenes was reported in 5 of the 34 PTCs. A significant temporal trend was observed, with ret/PTC chimera detected for the most part before 1997 and BRAF mutations being more prevalent after 1997. The results suggest that some environmental/etiological agent(s) may have influenced the pathobiology of thyroid tumor development, among the population examined, over time.

Threshold 2: the reliability, validity and sensitivity to change of the Threshold Assessment Grid (TAG).

OBJECTIVE: This study investigated the psychometric properties of the Threshold Assessment Grid (TAG), a new assessment of the severity of mental health problems. METHOD: A total of 605 patients were recruited from 10 mental health adult and elderly services in London, UK. TAG ratings and other standardized definitions of severe mental illness were completed by referrers. TAG, Global Assessment of Functioning (GAF), Camberwell Assessment of Need Short Appraisal Schedule (CANSAS) and Health of the Nation Outcome Scale (HoNOS) ratings were completed by mental health service staff. Construct validation on extreme groups was investigated. RESULTS: Construct and concurrent validity were good. Referrer TAG scores predicted mental health team view of referral suitability, but not whether assessments were offered. Test-retest reliability was good, interrater reliability ranged from good to poor in different domains (but adequate for total TAG score), internal consistency was appropriate. Sensitivity to change requires further investigation. CONCLUSION: The TAG can be recommended for use by all agencies when making referrals to mental health services.

Transient outward current inhibition by propafenone and 5-hydroxypropafenone in cultured neonatal rat ventricular myocytes.

The antiarrhythmic agent propafenone and its primary electropharmacologically active metabolite, 5-hydroxypropafenone, are known inhibitors of cardiac myocyte repolarizing currents. We recently documented potent propafenone inhibition of the transient outward potassium current (Ito) in human atrial myocytes from patients in the newborn and infant age range. In the current study we characterized ventricular Ito inhibition by propafenone and 5-hydroxypropafenone in neonatal myocytes enzymatically isolated from 2-day-old Sprague-Dawley rat pups. Using the whole-cell patch-clamp technique in ventricular myocytes kept in primary culture for 1-4 days, we observed comparably potent Ito inhibition by both agents, yielding 50% maximal inhibitory concentration (IC50) values of 2.1 +/- 0.5 and 1.5 +/- 0.2 microM for propafenone and 5-hydroxypropafenone, respectively. Ito blockade by both of these agents was time, concentration, and voltage dependent, but use independent. There was no drug effect on steady-state voltage dependence of Ito inactivation, or on the time course of Ito recovery from inactivation. These findings are consistent with an open channel-blocking mechanism as suggested by other models. We conclude that both propafenone and 5-hydroxypropafenone are potent Ito inhibitors in neonatal rat ventricular myocytes, with potencies exceeding those demonstrated for propafenone in adult rat ventricular myocytes or in human atrial myocytes from patients of all ages.

The Mycobacterium tuberculosis cmaA2 gene encodes a mycolic acid trans-cyclopropane synthetase.

Infection with Mycobacterium tuberculosis remains a major global health emergency. Although detailed understanding of the molecular events of M. tuberculosis pathogenesis is still limited, recent genetic analyses have implicated specific lipids of the cell envelope as important effectors in M. tuberculosis pathogenesis. We have shown that pcaA, a novel member of a family of M. tuberculosis S-adenosyl methionine (SAM)-dependent methyl transferases, is required for alpha-mycolic acid cyclopropanation and lethal chronic persistent M. tuberculosis infection. To examine the apparent redundancy between pcaA and cmaA2, another cyclopropane synthetase of M. tuberculosis thought to be involved in alpha-mycolate synthesis, we have disrupted the cmaA2 gene in virulent M. tuberculosis by specialized transduction. Inactivation of cmaA2 causes accumulation of unsaturated derivatives of both the methoxy- and ketomycolates. Analysis by proton NMR indicates that the mycolic acids of the cmaA2 mutant lack trans-cyclopropane rings but are otherwise intact with respect to cyclopropane and methyl branch content. Thus, cmaA2 is required for the synthesis of the trans cyclopropane rings of both the methoxymycolates and ketomycolates. These results define cmaA2 as a trans-cyclopropane synthetase and expand our knowledge of the substrate specificity of a large family of highly homologous mycolic acid methyl transferases recently shown to be critical to M. tuberculosis pathogenesis.

Threshold 3: the feasibility of the Threshold Assessment Grid (TAG) for routine assessment of the severity of mental health problems.

BACKGROUND: Evidence-based practice requires the development of measures which are suitable for everyday clinical use ('feasible'). There is no consensus as to how to establish feasibility. METHOD: The feasibility of a new assessment - the Threshold Assessment Grid (TAG) - for use when making referrals to mental health services was tested by training mental health teams in using the TAG and other standardised assessments, asking referrers to ten mental health services in London also to complete a TAG, surveying TAG users, and evaluating a feedback meeting at which TAG data were presented. RESULTS: One hundred and one (61%) mental health staff received training, and 445 (74%) referrers of 600 patients completed TAGs. Sixty-five (65%) questionnaires from TAG users were completed, and 24 (80%) people attending feedback meetings evaluated the TAG. These allowed the extent to which the TAG is brief, simple, relevant, acceptable, available and valuable to be investigated. CONCLUSION: The TAG exhibited good feasibility when used by mental health staff, and moderate feasibility when used by referrers. This approach can be used to investigate the feasibility of other standardised assessments.

Stories of relative privilege: power and social change in feminist community psychology.

Stories about community work in New Zealand and Scotland are presented to describe and reflect on issues central to feminist community psychology. Organizing a lesbian festival, Ingrid Huygens describes feminist processes used to equalize resources across Maori (indigenous) and Pakeha (white) groups. Heather Hamerton presents her experiences as a researcher using collective memory work to reflect on adolescent experiences related to gender, ethnicity, and class. Sharon Cahill chronicles dilemmas and insights from focus groups about anger with women living in public housing in Scotland. Each story chronicles experiences related to oppression and privilege, and describes the author's emotions and reflections. Individually and collectively, the stories illustrate the potential offered by narrative methods and participatory processes for challenging inequalities and encouraging social justice.

Apparent symptom overreporting in combat veterans evaluated for PTSD.

Psychometric studies have consistently shown that combat veterans evaluated for posttraumatic stress disorder (PTSD) appear to overreport psychopathology as exhibited by (a) extreme and diffuse levels of psychopathology across instruments measuring different domains of mental illness, and (b) extreme elevations on the validity scale of the MMPI-MMPI-2, in a "fake-bad" direction. The phenomenon of this ubiquitous presentational style is not well understood at present. In this review we describe and delineate the assessment problem posed by this apparent symptom overreporting, and we review the literature regarding several potential explanatory factors. Finally, we address conceptual and practical issues relevant to reaching a better understanding of the phenomenon, and ultimately the clinical syndrome of combat-related PTSD, in both research and clinical settings.

Predictors of torsades de pointes in rabbit ventricles perfused with sedating and nonsedating histamine H1-receptor antagonists.

Several nonsedating histamine H1-receptor antagonists are associated with torsades de pointes ventricular tachycardia. The objectives of this study were to: (i) compare electrocardiographic, monophasic action potential, and arrhythmogenic effects of sedating and nonsedating H1-receptor antagonists, and (ii) identify correlates of drug-induced torsades de pointes in an isolated ventricle model. Isolated, electrically paced (1-3 Hz) rabbit ventricles were Langendorff-perfused with either drug-free Tyrode's solution or one of the following: (i) the sedating H1-receptor antagonist hydroxyzine (0.1-30 microM), (ii) cetirizine, a nonsedating metabolite of hydroxyzine (1-300 microM), and (iii) the nonsedating, putatively arrhythmogenic H1-receptor antagonist astemizole (0.1-30 microM). Volume conducted electrocardiographic signals and monophasic action potentials from the periapical left ventricular endocardium and epicardium were recorded. There were no apparent changes in control (n = 15) or hydroxyzine-perfused (n = 7) hearts. Cetirizine (n = 13) produced a mild biphasic electrocardiographic QT interval prolongation and was associated with early afterdepolarizations, but not with torsades de pointes. Astemizole (n = 11) lengthened QT intervals, and at high concentration (30 microM) induced torsades de pointes in 10 of 11 hearts (P < 0.001 vs. all other groups). These findings are consistent with previously reported repolarizing current inhibition by cetirizine, but may additionally indicate "compensatory" inhibition of inward currents at higher concentrations. By contrast, astemizole-induced changes are consistent with unopposed repolarizing current inhibition.

The 2.0 A structure of malarial purine phosphoribosyltransferase in complex with a transition-state analogue inhibitor.

Malaria is a leading cause of worldwide mortality from infectious disease. Plasmodium falciparum proliferation in human erythrocytes requires purine salvage by hypoxanthine-guanine-xanthine phosphoribosyltransferase (HGXPRTase). The enzyme is a target for the development of novel antimalarials. Design and synthesis of transition-state analogue inhibitors permitted cocrystallization with the malarial enzyme and refinement of the complex to 2.0 A resolution. Catalytic site contacts in the malarial enzyme are similar to those of human hypoxanthine-guanine phosphoribosyltransferase (HGPRTase) despite distinct substrate specificity. The crystal structure of malarial HGXPRTase with bound inhibitor, pyrophosphate, and two Mg(2+) ions reveals features unique to the transition-state analogue complex. Substrate-assisted catalysis occurs by ribooxocarbenium stabilization from the O5' lone pair and a pyrophosphate oxygen. A dissociative reaction coordinate path is implicated in which the primary reaction coordinate motion is the ribosyl C1' in motion between relatively immobile purine base and (Mg)(2)-pyrophosphate. Several short hydrogen bonds form in the complex of the enzyme and inhibitor. The proton NMR spectrum of the transition-state analogue complex of malarial HGXPRTase contains two downfield signals at 14.3 and 15.3 ppm. Despite the structural similarity to the human enzyme, the NMR spectra of the complexes reveal differences in hydrogen bonding between the transition-state analogue complexes of the human and malarial HG(X)PRTases. The X-ray crystal structures and NMR spectra reveal chemical and structural features that suggest a strategy for the design of malaria-specific transition-state inhibitors.