The varying effects of a range of preservatives on Myoviridae and Siphoviridae bacteriophages formulated in a semi-solid cream preparation.

Bacteriophages may be formulated into semi-solid bases for therapeutic delivery. This work investigated the effects of a range of preservatives on the viability of Myoviridae and Siphoviridae bacteriophages when these were formulated into a standard semi-solid cream base. The six preservatives tested included: benzoic acid (0·1%), chlorocresol (0·1%), combination hydroxybenzoates (propyl 4-hydroxybenzoates with methyl 4-hydroxybenzoates) (0·1%), methyl 4-hydroxybenzoate (0·08%), 2-phenoxyethanol (1%) and propyl 4-hydroxybenzoate (0·02%). These were each formulated into cetomacrogol cream aqueous to generate six individual semi-solid bases into which Myoviridae and Siphoviridae bacteriophages were added and tested for stability. Optimal bacteriophage stability was seen when the preservative chlorocresol was used. Bacteriophage in the acidic benzoic acid were the least stable, resulting in complete loss of viability after 4-5 weeks. Of the bacteriophages tested, the Myoviridae KOX1 was significantly more stable than the Siphoviridae PAC1 after 91 days in formulations with each of the preservatives. Our results suggest the need for individual testing of specific bacteriophages in pharmaceutical formulations, as their efficacy when exposed to preservatives and excipients in these delivery forms may vary. SIGNIFICANCE AND IMPACT OF THE STUDY: Bacteriophages are being increasingly investigated as alternatives to antibiotics. While bacteriophages can be formulated in diverse ways for therapeutic delivery, there has been scant work on how excipients and preservatives in these formulations affect stability of different bacteriophages. We demonstrate that the nature of preservatives in formulations will affect bacteriophage stability, and that in these formulations, viability of bacteriophage differs according to their morphology. Our work highlights the need for individual testing of specific bacteriophages in pharmaceutical formulations, as efficacy when exposed to preservatives and excipients in these delivery forms may vary.

Preventable mortality in geriatric hip fracture inpatients.

There is a high rate of mortality in elderly patients who sustain a fracture of the hip. We aimed to determine the rate of preventable mortality and errors during the management of these patients. A 12 month prospective study was performed on patients aged > 65 years who had sustained a fracture of the hip. This was conducted at a Level 1 Trauma Centre with no orthogeriatric service. A multidisciplinary review of the medical records by four specialists was performed to analyse errors of management and elements of preventable mortality. During 2011, there were 437 patients aged > 65 years admitted with a fracture of the hip (85 years (66 to 99)) and 20 died while in hospital (86.3 years (67 to 96)). A total of 152 errors were identified in the 80 individual reviews of the 20 deaths. A total of 99 errors (65%) were thought to have at least a moderate effect on death; 45 reviews considering death (57%) were thought to have potentially been preventable. Agreement between the panel of reviewers on the preventability of death was fair. A larger-scale assessment of preventable mortality in elderly patients who sustain a fracture of the hip is required. Multidisciplinary review panels could be considered as part of the quality assurance process in the management of these patients.

Proportional mouse model for aerosol infection by influenza.

AIMS: The aim of this study was to demonstrate a prototype tool for measuring infectivity of an aerosolized human pathogen - influenza A/PR/8/34 (H1N1) virus - using a small-animal model in the Controlled Aerosol Test System (CATS). METHODS AND RESULTS: Intranasal inoculation of nonadapted H1N1 virus into C57BL, BALB/c and CD-1 mice caused infection in all three species. Respiratory exposure of CD-1 mice to the aerosolized virus at graduated doses was accomplished in a modified rodent exposure apparatus. Weight change was recorded for 7 days postexposure, and viral populations in lung tissue homogenates were measured post mortem by DNA amplification (qRT-PCR), direct fluorescence and microscopic evaluation of cytopathic effect. Plots of weight change and of PCR cycle threshold vs delivered dose were linear to threshold doses of ~40 TCID(50) and ~12 TCID(50) , respectively. CONCLUSIONS: MID(50) for inspired H1N1 aerosols in CD-1 mice is between 12 and 40 TCID(50) ; proportionality to dose of weight loss and viral populations makes the CD-1 mouse a useful model for measuring infectivity by inhalation. SIGNIFICANCE AND IMPACT OF THE STUDY: In the CATS, this mouse-virus model provides the first quantitative method to evaluate the ability of respiratory protective technologies to attenuate the infectivity of an inspired pathogenic aerosol.

Relationship of coronary calcium and myocardial perfusion in individuals with chest pain. Assessed by integrated rubidium-82 PET-CT.

AIM: The integrated value of coronary calcium scoring added to myocardial perfusion assessment in hybrid PET-CT imaging remains poorly defined. In the present study, we sought to determine the relationship between calcium burden, other risk factors, and tissue perfusion in a group of patients with chest pain and predominantly intermediate likelihood for coronary artery disease. PATIENTS, METHODS: In 70 patients, coronary calcium scores (CCS) were obtained in addition to rest/dipyridamole stress 82Rb perfusion images using a GE Discovery Rx hybrid PET-CT system. From static perfusion images, summed rest, stress and difference scores (SRS, SSS, SDS) were calculated using a 20-segment model. Absolute CCS was determined according to Agatston and age-, gender-, and ethnicity-matched CCS percentiles were calculated using the MESA database. RESULTS: SSS, SRS and SDS were abnormal (>or=4) in 25 (36%), 17 (24%), and 12 (17%) patients. Mean CCS according to Agatston was 180+/-446(range 0-2122), and CCS percentile was 42+/-43(range 0-99). Absolute CCS correlated mildly but significantly with SSS (r=0.31, p=0.01), while CCS percentile did not (r=0.11, p=0.36). Of 49 patients with normal perfusion, 25 (57%) had CCS=0, and 8 (18%) had a CCS percentile>or=75th. Of 35 patients with a CCS=0, 26 (74%) had normal perfusion. Individuals in whom review of patient records revealed events during follow-up (n=10) had significantly higher SSS than those where no events were recorded (6.0+/-7.2 versus 2.9+/-3.1, p=0.03), and there was a trend towards higher CCS percentiles (62+/-36 versus 35+/-43, p=0.06). CONCLUSION: Coronary calcifications and myocardial tissue perfusion, as interrogated in a single PET-CT imaging session, show only partial agreement in patients with chest pain. Both tests seem to reflect different pathophysiologic components, and may be complementary for definition of individual disease patterns.

Stem cells from fetal membranes - a workshop report.

Stem cells that can be derived from fetal membranes represent an exciting field of research that bears tremendous potential for developmental biology and regenerative medicine. In this report we summarize contributions to a workshop in which newest insights into the characteristics, subtypes and molecular determinants of stem cells from trophoblast and endometrial tissues were presented.

Driver sensitivity to brake pulse duration and magnitude.

Adaptive cruise control (ACC) requires that the driver intervene in situations that exceed the capability of ACC. A brake pulse might provide a particularly compatible means of alerting the driver to situations in which the acceleration authority of the ACC has been exceeded. This study examined the sensitivity of the driver to brake pulses of five different amplitudes (0.01-0.025 g) and five different durations (50-800 ms). Drivers were sensitive to accelerations as low as 0.015 g. Pulse duration interacted with pulse amplitude, such that moderate duration pulses were more detectable than long and short duration pulses at intermediate levels of pulse amplitude. A power function with an exponent of 1.0 accounted for 99% of the variance in drivers' sensitivity to pulse amplitude; however, a power function with an exponent of 0.23 accounted for only 70% of the variance in drivers' sensitivity to pulse duration. These results can help designers create ACC algorithms and develop brake pulse warnings.

Can alcohol-based hand-rub solutions cause you to lose your driver's license? Comparative cutaneous absorption of various alcohols.

We assessed cutaneous ethanol (ETOH) and isopropanol (ISOP) absorption after intensive (30 times per h) use of alcohol-based hand-rub solutions by healthcare workers (HCWs). ETOH was detectable in the breath of 6/20 HCWs (0.001 to 0.0025%) at 1 to 2 min postexposure and in the serum of 2/20 HCWs at 5 to 7 min postexposure. Serum ISOP levels were unrecordable at all time points.

Melanoma cell migration is upregulated by tumour necrosis factor-alpha and suppressed by alpha-melanocyte-stimulating hormone.

We reported recently that the inflammatory cytokine tumour necrosis factor alpha (TNF-alpha) can upregulate integrin expression, cell attachment and invasion of cells through fibronectin in a human melanoma cell line (HBL). Furthermore, the actions of TNF-alpha were suppressed by the addition of an anti-inflammatory peptide alpha-melanocyte-stimulating hormone (alpha-MSH). In the current study, we extend this work investigating to what extent TNF-alpha might stimulate melanoma invasion by promoting cell migration and whether alpha-MSH is also inhibitory. Two human melanoma cell lines were examined in vitro (HBL and C8161) using a scratch migration assay. Analysis using either time-lapse video microscopy or imaging software analysis of migrating 'fronts' of cells revealed that C8161 cells migrated more rapidly than HBL cells. However, when cells were stimulated with TNF-alpha both cell types responded with a significant increase in migration distance over a 16-26 h incubation time. alpha-Melanocyte-stimulating hormone had an inhibitory effect on TNF-alpha-stimulated migration for HBL cells, completely blocking migration at 10(-9) M. In contrast, C8161 cells did not respond to alpha-MSH (as these cells have a loss-of-function melanocortin-1 receptor). However, stable transfection of C8161 cells with the wild-type melanocortin-1 receptor produced cells whose migration was significantly inhibited by alpha-MSH. In addition, the use of a neutralising antibody to the beta(1)-integrin subunit significantly reduced migration in both cell types. This data therefore supports an inflammatory environment promoting melanoma cell migration, and in addition shows that alpha-MSH can inhibit inflammatory stimulated migration. The data also support a fundamental role of the beta(1)-integrin receptor in melanoma cell migration.

Q-VD-OPh, a broad spectrum caspase inhibitor with potent antiapoptotic properties.

In recent years, several inhibitors that prevent caspase activation and apoptosis have emerged. At high doses, however, these inhibitors can have nonspecific effects and/or become cytotoxic. In this study, we determined the effectiveness of broad spectrum caspase inhibitors to prevent apoptosis. A carboxy terminal phenoxy group conjugated to the amino acids valine and aspartate (Q-VD-OPh) potently inhibited apoptosis. Q-VD-OPh was significantly more effective in preventing apoptosis than the widely used inhibitors, ZVAD-fmk and Boc-D-fmk, and was also equally effective in preventing apoptosis mediated by the three major apoptotic pathways, caspase 9/3, caspase 8/10, and caspase 12. In addition to the increased effectiveness, Q-VD-OPh was not toxic to cells even at extremely high concentrations. Our data indicate that the specificity, effectiveness, and reduced toxicity of caspase inhibitors can be significantly enhanced using carboxyterminal o-phenoxy groups and may have important uses in vivo.

Identification of the primary caspase 3 cleavage site in alpha II-spectrin during apoptosis.

Alpha II-spectrin is one of the major proteins responsible for maintaining the cytoskeletal integrity of the cell. The caspase 3-mediated cleavage of alpha II-spectrin during apoptotic cell death may play an important role in altering membrane stability and the formation of apoptotic bodies. In this study, we identified the primary caspase 3 cleavage site in alpha II-spectrin. We found that the transcriptional inhibitor, actinomycin D, induced caspase 3 activation and that caspase 3 activation is coincident with the cleavage of alpha II-spectrin protein at a primary cleavage site. Deletion analysis and site directed mutagenesis identified the primary cleavage site in alpha II spectrin at amino acid 1185 (DETD). The primary caspase 3 cleavage site in alpha II spectrin is conserved in immature and mature B cells. Our results indicate that alpha II-spectrin is initially cleaved at a caspase 3 consensus site and this primary event likely alters the structural conformation of the protein exposing subsequent cleavage sites and altering cytoskeletal integrity. Identification of the primary cleavage site for caspase 3 may help to elucidate the role of alpha II-spectrin in membrane stability and apoptosis as well as provide new insights into alpha II-spectrin autoantibody formation associated with the autoimmune disease, Sjögren's syndrome.

MTS-conjugated-antiactive caspase 3 antibodies inhibit actinomycin D-induced apoptosis.

Caspase 3 is critically involved in the pathway of apoptosis. We have conjugated a MTS-transport-peptide to monoclonal and polyclonal anti-caspase-3 antibodies to suppress Actinomycin D-induced apoptosis in human lymphoma T cells. The advantage of using trans-membrane antibodies compared to conventional apoptosis inhibitors is their specific target recognition in the living cell and their lower toxicity compared to conventional apoptosis inhibitors. We could show that a MTS-transport-peptide modified monoclonal anti-caspase-3 antibody reduces Actinomycin D induced apoptosis, as shown by DNA ladder electrophoresis and cell death ELISA. These results indicate that antibodies have a therapeutic potential to inhibit apoptosis in a variety of diseases.

A study of transcallosal inhibition in schizophrenia using transcranial magnetic stimulation.

A considerable body of imaging research has demonstrated morphological changes in the corpus callosum (CC) of patients with schizophrenia. Transcranial magnetic stimulation (TMS) allows the possibility for the in vivo investigation of a variety of aspects of brain function including the spread of information across the CC. We aimed to investigate whether patients with schizophrenia demonstrate abnormalities of transcallosal inhibition (TCI), a TMS parameter measured with both single and paired pulse experiments. 25 patients with DSM-IV schizophrenia and 20 normal volunteers participated in the study. Electromyographic (EMG) recordings from the bilateral abductor pollicis brevis (APB) muscle were made during focal TMS stimulation to the motor cortex. Experimental paradigms were utilised to measure both the timing and degree of the effect of TCI. The patient group demonstrated a reduction in the degree of TCI at rest and during a sustained muscle contraction. TCI commenced at the same time in the patient and the control group but was of prolonged duration in the patient group although the length of TCI correlated with medication dose. Patients with schizophrenia demonstrate a reduction in the degree of TCI that appeared independent of medication dose. The latency of TCI is not altered in the patient group suggesting that cortical inhibitory mechanisms, rather than corpus callosal ones, are likely to be the cause of these TCI alterations.

The application of transcranial magnetic stimulation in psychiatry and neurosciences research.

OBJECTIVE: Over recent years transcranial magnetic stimulation (TMS) has become widely applied in the study of neuropsychiatric disorders. The aim of this article is to review the application of TMS as an investigative tool and as a potential therapeutic modality in psychiatric disorders. METHOD: A comprehensive literature review. RESULTS: When applied as an investigative tool, TMS provides innovative ways to directly study the excitability of the cortex, cortical regional connectivity, the plasticity of brain responses and cognitive functioning in illness and disease states. A number of studies suggest the potential of treatment with TMS in disease states, especially in patients with depression, although difficulties exist with the interpretation of the published literature. CONCLUSION: TMS has a considerable role in neuropsychiatric research. It appears to have considerable potential as a therapeutic tool in depression, and perhaps a role in several other disorders, although widespread application requires larger trials and establishment of sustained response.

The role of religion in predicting adolescent alcohol use and problem drinking.

OBJECTIVE: There are racial differences in adolescents' propensity to consume alcohol--with white adolescents tending to consume more alcohol than black adolescents--but there is no clear explanation for why such differences exist. The purpose of this study was to examine the relationship between religiosity, a cultural factor that is not well understood currently, and racial differences in adolescent alcohol use. METHOD: Participants were white and black ninth-grade adolescents (N = 899; 54% female, 57.5% white) involved in a 3-year longitudinal study of ways to reduce alcohol use and sexual risk-taking behavior among adolescents in Ohio and Kentucky. RESULTS: Findings indicate that religiosity is differentially associated with alcohol use and problem drinking for white and black adolescents. Religious service attendance was the most significant predictor of alcohol use for black adolescents, whereas religious fundamentalism was most important for white adolescents. In contrast, frequency of prayer was the significant predictor of problem drinking for black adolescents, whereas the level of importance placed on religion was the significant predictor for white adolescents. Important gender differences also emerged in both prediction models and are discussed. CONCLUSIONS: Since there is great heterogeneity among adolescents (in terms of race and gender) in their alcohol use and misuse, the "one-size-fits-all" approach to alcohol treatment and prevention is likely inappropriate. Moreover, conceptualizations of alcohol use and misuse, and its prevention and treatment, should include the consideration of such key cultural factors as religiosity.

Characterization of a TGFbeta-responsive human trophoblast-derived cell line.

The placenta is formed by developing trophoblast cells to facilitate fluid, gas and nutrient exchange with the mother. Inappropriate trophoblast responsiveness can lead to life threatening complications during pregnancy including intrauterine growth retardation, pre-eclampsia, spontaneous abortion and malignancy that could lead to fetal loss. Transforming growth factor beta (TGFbeta) is a multifunctional cytokine required for embryonic development and is an important regulator of human trophoblast function. Although TGFbeta is critical for placental and embryonic development, there are currently no established TGFbeta-responsive human trophoblast-derived cell lines available to study the mechanisms by which TGFbeta regulates trophoblast function. Our studies have examined the transformed human trophoblast-derived cell line, ED27, to determine if it is responsive to TGFbeta. Our data indicate that TGFbeta dose responsively and reversibly inhibits cell growth in ED27 cells and induces classic TGFbeta response genes, fibronectin and plasminogen activator inhibitor 1 (PAI-1). TGFbeta also induces an inhibitor of trophoblast invasion, tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) in ED27 cells. Our studies have identified a human trophoblast-derived cell line that parallels isolated primary human trophoblasts in their responses to TGFbeta. This cell line may provide us with the opportunity to determine TGFbeta-mediated responses on human trophoblast functions not previously possible.

Speech-based interaction with in-vehicle computers: the effect of speech-based e-mail on drivers' attention to the roadway.

As computer applications for cars emerge, a speech-based interface offers an appealing alternative to the visually demanding direct manipulation interface. However, speech-based systems may pose cognitive demands that could undermine driving safety. This study used a car-following task to evaluate how a speech-based e-mail system affects drivers' response to the periodic braking of a lead vehicle. The study included 24 drivers between the ages of 18 and 24 years. A baseline condition with no e-mail system was compared with a simple and a complex e-mail system in both simple and complex driving environments. The results show a 30% (310 ms) increase in reaction time when the speech-based system is used. Subjective workload ratings and probe questions also indicate that speech-based interaction introduces a significant cognitive load, which was highest for the complex e-mail system. These data show that a speech-based interface is not a panacea that eliminates the potential distraction of in-vehicle computers. Actual or potential applications of this research include design of in-vehicle information systems and evaluation of their contributions to driver distraction.

A randomized double-blind, placebo-controlled trial of the EMLA patch for the reduction of pain associated with intramuscular injection in four to six-year-old children.

UNLABELLED: The effectiveness of a eutectic mixture lidocaine-prilocaine topical anaesthetic cream (EMLA) patch compared with a placebo patch in the reduction of pain associated with intramuscular immunization was evaluated. As part of the study, 161 children (aged 4-6-y) undergoing routine diphtheria, pertussis, tetanus and polio (DPTP) immunization in five urban and five rural private office settings were randomly assigned to an EMLA patch (n = 83) or a placebo patch control group (n = 78). Pain measurements included: child's self-report on a Faces Pain Scale; facial action on the Child Facial Coding System; the Children's Hospital of Eastern Ontario Pain Scale and parent and technician ratings on a Visual Analogue Scale. Parents also rated their own and their child's immunization-related anxiety on a Visual Analogue Scale. The EMLA patch group had significantly less pain on all four pain measures compared with the placebo group. Of the children in the placebo group, 43% had clinically significant pain, compared with 17% of children in the EMLA patch group. No severe adverse symptoms occurred as a result of either EMLA or placebo patch application. CONCLUSION: The EMLA patch reduced immunization pain in 4 to 6-y-old children during needle injection.

The Alaska Native diabetes program.

OBJECTIVES: To provide optimum health care to indigenous people with diabetes, to prevent diabetes, and to monitor the epidemiology of diabetes and selected complications. The purposes of this paper are to describe the program and to present data that highlights the major problems and successes. STUDY DESIGN: Descriptive epidemiology report of diabetes and population service program based on yearly chart review data. METHODS: Almost half of Alaska Natives with diabetes have no direct access to physicians or hospitals. Health care delivery is now managed by the tribes themselves. Program emphases include maintenance of a population-based registry, formal training for village health aides, physical activity programs, patient education, primary prevention activities and adherence to standards of care to prevent complications. A centralized registry is maintained to assure that epidemiological data is available and patients are not lost to follow-up. Each year a random sample of charts at each major facility is audited against nationally standardized care guidelines. RESULTS: The prevalence of diabetes among Alaska Natives increased 80% over the 13 years from 1985 to 1998 (15.7/1000 to 28.3/1000, age adjusted to U.S. 1980 population). For the years 1986-1998 the incidence rates of lower extremity amputation and end stage renal disease were 6.1/1000 and 2.0/1000 respectively. The level of care provided to Alaska Native patients is comparable to that provided to the general diabetic patient population seen in Alaskan urban clinics. CONCLUSIONS: In spite of logistic challenges, care provided to Alaska Native people with diabetes compares favorably to that provided in other settings. Incidence rates of lower extremity amputation and end stage renal disease also remain comparable to or lower than those in other U.S. populations. Many aspects of our system could be extended to other chronic disease programs serving isolated indigenous populations. Primary prevention of diabetes remains a major challenge as life styles change.

'VIT1', a novel gene associated with vitiligo.

To define genes associated with the pigmentary disorder vitiligo, gene expression was compared in non-lesional melanocytes cultured from three vitiligo patients and from three control melanocyte cultures by differential display. A basic local alignment search tool search did not reveal homology of six differentially expressed cDNA fragments to previously identified expressed sequence tags; thus, one was used to screen a melanocyte cDNA library. The underlying VIT1 gene maps to chromosome 2p16. The 3' portion of the VIT1 message is complementary to the 3' end of hMSH6 mRNA, enabling the formation of RNA-RNA hybrids, which may interfere with G/T mismatch repair function. Moreover, the aligned cDNA sequence revealed an open reading frame identical to a hypothetical protein expressed in brain, with a similarity to Drosophila calmodulin, and containing a zinc-finger motif partially identical to N-recognin. Expression of ORF mRNA was confirmed for multiple skin cell types, suggesting its importance for skin physiology.

Human performance models and rear-end collision avoidance algorithms.

Collision warning systems offer a promising approach to mitigate rear-end collisions, but substantial uncertainty exists regarding the joint performance of the driver and the collision warning algorithms. A simple deterministic model of driver performance was used to examine kinematics-based and perceptual-based rear-end collision avoidance algorithms over a range of collision situations, algorithm parameters, and assumptions regarding driver performance. The results show that the assumptions concerning driver reaction times have important consequences for algorithm performance, with underestimates dramatically undermining the safety benefit of the warning. Additionally, under some circumstances, when drivers rely on the warning algorithms, larger headways can result in more severe collisions. This reflects the nonlinear interaction among the collision situation, the algorithm, and driver response that should not be attributed to the complexities of driver behavior but to the kinematics of the situation. Comparisons made with experimental data demonstrate that a simple human performance model can capture important elements of system performance and complement expensive human-in-the-loop experiments. Actual or potential applications of this research include selection of an appropriate algorithm, more accurate specification of algorithm parameters, and guidance for future experiments.