Multiplexed drug testing of tumor slices using a microfluidic platform.

Current methods to assess the drug response of individual human cancers are often inaccurate, costly, or slow. Functional approaches that rapidly and directly assess the response of patient cancer tissue to drugs or small molecules offer a promising way to improve drug testing, and have the potential to identify the best therapy for individual patients. We developed a digitally manufactured microfluidic platform for multiplexed drug testing of intact cancer slice cultures, and demonstrate the use of this platform to evaluate drug responses in slice cultures from human glioma xenografts and patient tumor biopsies. This approach retains much of the tissue microenvironment and can provide results rapidly enough, within days of surgery, to guide the choice of effective initial therapies. Our results establish a useful preclinical platform for cancer drug testing and development with the potential to improve cancer personalized medicine.

A microfluidic platform for functional testing of cancer drugs on intact tumor slices.

Present approaches to assess cancer treatments are often inaccurate, costly, and/or cumbersome. Functional testing platforms that use live tumor cells are a promising tool both for drug development and for identifying the optimal therapy for a given patient, i.e. precision oncology. However, current methods that utilize patient-derived cells from dissociated tissue typically lack the microenvironment of the tumor tissue and/or cannot inform on a timescale rapid enough to guide decisions for patient-specific therapy. We have developed a microfluidic platform that allows for multiplexed drug testing of intact tumor slices cultured on a porous membrane. The device is digitally-manufactured in a biocompatible thermoplastic by laser-cutting and solvent bonding. Here we describe the fabrication process in detail, we characterize the fluidic performance of the device, and demonstrate on-device drug-response testing with tumor slices from xenografts and from a patient colorectal tumor.

Neurodevelopmental profile of siblings with Angelman syndrome due to pathogenic UBE3A variants.

BACKGROUND: Angelman syndrome (AS) is a neurodevelopmental disorder caused by a lack of expression of the maternally inherited UBE3A gene on chromosome 15. Individuals with AS due to a UBE3A mutation are more likely to have siblings who also have AS compared with those with AS due to other cytogenetic/molecular mechanisms, but it is unknown whether the developmental outcome of siblings who have AS is similar. METHODS: Through an ongoing AS Natural History Study, we identified seven pairs of siblings with AS due to a UBE3A mutation. We compared the neurodevelopment of the first-born and second-born siblings with AS participants who have a UBE3A mutation and have either typically developing siblings or no siblings. RESULTS: Second-born AS participants due to a UBE3A mutation were more likely to be diagnosed at an earlier age. With the exception of higher expressive language scores among the second-born participants, no other differences were observed in the developmental and adaptive functioning skills across the different groups. CONCLUSIONS: The presence of an older sibling with the same neurodevelopmental disorder is associated with an earlier age of diagnosis and may be associated with an improvement in expressive language skills; the developmental outcome of siblings with AS due to a UBE3A mutation is otherwise comparable.

Multimodel precipitation responses to removal of U.S. sulfur dioxide emissions.

Emissions of aerosols and their precursors are declining due to policies enacted to protect human health, yet we currently lack a full understanding of the magnitude, spatiotemporal pattern, statistical significance, and physical mechanisms of precipitation responses to aerosol reductions. We quantify the global and regional precipitation responses to U.S. SO2 emission reductions using three fully coupled chemistry-climate models: Community Earth System Model version 1, Geophysical Fluid Dynamics Laboratory Coupled Model 3, and Goddard Institute for Space Studies ModelE2. We contrast 200 year (or longer) simulations in which anthropogenic U.S. sulfur dioxide (SO2) emissions are set to zero with present-day control simulations to assess the aerosol, cloud, and precipitation response to U.S. SO2 reductions. In all three models, reductions in aerosol optical depth up to 70% and cloud droplet number column concentration up to 60% occur over the eastern U.S. and extend over the Atlantic Ocean. Precipitation responses occur both locally and remotely, with the models consistently showing an increase in most regions considered. We find a northward shift of the tropical rain belt location of up to 0.35° latitude especially near the Sahel, where the rainy season length and intensity are significantly enhanced in two of the three models. This enhancement is the result of greater warming in the Northern versus Southern Hemispheres, which acts to shift the Intertropical Convergence Zone northward, delivering additional wet season rainfall to the Sahel. Two of our three models thus imply a previously unconsidered benefit of continued U.S. SO2 reductions for Sahel precipitation.

Continuous-flow multi-pulse electroporation at low DC voltages by microfluidic flipping of the voltage space topology.

Concerns over biosafety, cost, and carrying capacity of viral vectors have accelerated research into physical techniques for gene delivery such as electroporation and mechanoporation. Advances in microfabrication have made it possible to create high electric fields over microscales, resulting in more efficient DNA delivery and higher cell viability. Continuous-flow microfluidic methods are typically more suitable for cellular therapies where a large number of cells need to be transfected under sterile conditions. However, the existing continuous-flow designs used to generate multiple pulses either require expensive peripherals such as high-voltage (>400 V) sources or function generators, or result in reduced cell viability due to the proximity of the cells to the electrodes. In this paper, we report a continuous-flow microfluidic device whose channel geometry reduces instrumentation demands and minimizes cellular toxicity. Our design can generate multiple pulses of high DC electric field strength using significantly lower voltages (15-60 V) than previous designs. The cells flow along a serpentine channel that repeatedly flips the cells between a cathode and an anode at high throughput. The cells must flow through a constriction each time they pass from an anode to a cathode, exposing them to high electric field strength for short durations of time (the "pulse-width"). A conductive biocompatible poly-aniline hydrogel network formed in situ is used to apply the DC voltage without bringing the metal electrodes close to the cells, further sheltering cells from the already low voltage electrodes. The device was used to electroporate multiple cell lines using electric field strengths between 700 and 800 V/cm with transfection efficiencies superior than previous flow-through designs.

Formaldehyde production from isoprene oxidation across NOx regimes.

The chemical link between isoprene and formaldehyde (HCHO) is a strong, non-linear function of NOx (= NO + NO2). This relationship is a linchpin for top-down isoprene emission inventory verification from orbital HCHO column observations. It is also a benchmark for overall photochemical mechanism performance with regard to VOC oxidation. Using a comprehensive suite of airborne in situ observations over the Southeast U.S., we quantify HCHO production across the urban-rural spectrum. Analysis of isoprene and its major first-generation oxidation products allows us to define both a "prompt" yield of HCHO (molecules of HCHO produced per molecule of freshly-emitted isoprene) and the background HCHO mixing ratio (from oxidation of longer-lived hydrocarbons). Over the range of observed NOx values (roughly 0.1 - 2 ppbv), the prompt yield increases by a factor of 3 (from 0.3 to 0.9 ppbv ppbv-1), while background HCHO increases by a factor of 2 (from 1.6 to 3.3 ppbv). We apply the same method to evaluate the performance of both a global chemical transport model (AM3) and a measurement-constrained 0-D steady state box model. Both models reproduce the NOx dependence of the prompt HCHO yield, illustrating that models with updated isoprene oxidation mechanisms can adequately capture the link between HCHO and recent isoprene emissions. On the other hand, both models under-estimate background HCHO mixing ratios, suggesting missing HCHO precursors, inadequate representation of later-generation isoprene degradation and/or under-estimated hydroxyl radical concentrations. Detailed process rates from the box model simulation demonstrate a 3-fold increase in HCHO production across the range of observed NOx values, driven by a 100% increase in OH and a 40% increase in branching of organic peroxy radical reactions to produce HCHO.

Patient safety attitudes of paediatric trainee physicians.

OBJECTIVES: To measure the patient safety attitudes of trainee physicians at an academic paediatric hospital. DESIGN: Cross-sectional survey. SETTING: An academic paediatric hospital. PARTICIPANTS: 209 trainee physicians based at the academic paediatric hospital in January 2004. MAIN OUTCOME MEASURES: Patient safety attitudes of trainee physicians measured using the Safety Attitudes Questionnaire (Inpatient Version) and a specific trainee survey. RESULTS: In the Safety Attitudes Questionnaire, responses were most positive in areas associated with independent care: job satisfaction (mean factor score = 77.5) safety climate (76.1), working conditions (75.6), perception of management (70.4) and less positively in areas associated with interdependent care: teamwork climate (64.6) and stress recognition (59.1). In the trainee survey, following a principal component analysis to identify summary factors, responses were most positive in the independent areas of clinical supervision and support (75.0), communication with their immediate senior physician (65.5) and orientation of new personnel (63.9), and less positive in the interdependent areas of handoffs and multiple services, (58.1), role identification during codes (51.0) and support following an adverse event (42.8). The combined independent factor scores were higher than the interdependent (difference = 17.9, 95% CI 16.1 to 19.7, p<0.001). Fellows reported higher independent factor scores than residents (5.5, 95% CI 2.2 to 8.9, p = 0.001), but not for the interdependent scores (-0.5, 95% CI -3.6 to 2.7, p = 0.767). CONCLUSIONS: Trainees appear comfortable with caring independently for patients but less so caring interdependently. With experience, trainee physicians may experience improvement in their ability to act independently but not interdependently. Recently developed patient safety culture instruments may enable additional understanding of what could be implemented to make improvements.

The global atmospheric environment for the next generation.

Air quality, ecosystem exposure to nitrogen deposition, and climate change are intimately coupled problems: we assess changes in the global atmospheric environment between 2000 and 2030 using 26 state-of-the-art global atmospheric chemistry models and three different emissions scenarios. The first (CLE) scenario reflects implementation of current air quality legislation around the world, while the second (MFR) represents a more optimistic case in which all currently feasible technologies are applied to achieve maximum emission reductions. We contrast these scenarios with the more pessimistic IPCC SRES A2 scenario. Ensemble simulations for the year 2000 are consistent among models and show a reasonable agreement with surface ozone, wet deposition, and NO2 satellite observations. Large parts of the world are currently exposed to high ozone concentrations and high deposition of nitrogen to ecosystems. By 2030, global surface ozone is calculated to increase globally by 1.5 +/- 1.2 ppb (CLE) and 4.3 +/- 2.2 ppb (A2), using the ensemble mean model results and associated +/-1 sigma standard deviations. Only the progressive MFR scenario will reduce ozone, by -2.3 +/- 1.1 ppb. Climate change is expected to modify surface ozone by -0.8 +/- 0.6 ppb, with larger decreases over sea than over land. Radiative forcing by ozone increases by 63 +/- 15 and 155 +/- 37 mW m(-2) for CLE and A2, respectively, and decreases by -45 +/- 15 mW m(-2) for MFR. We compute that at present 10.1% of the global natural terrestrial ecosystems are exposed to nitrogen deposition above a critical load of 1 g N m(-2) yr(-1). These percentages increase by 2030 to 15.8% (CLE), 10.5% (MFR), and 25% (A2). This study shows the importance of enforcing current worldwide air quality legislation and the major benefits of going further. Nonattainment of these air quality policy objectives, such as expressed by the SRES-A2 scenario, would further degrade the global atmospheric environment.

Mental health aspects of emergency medical services for children: summary of a consensus conference.

OBJECTIVE: To address the mental health needs of children involved in emergency medical services (EMS). METHODS: A multidisciplinary consensus conference convened to identify mental health needs of children and their families related to pediatric medical emergencies, to examine the impact of psychological aspects of emergencies on recovery and satisfaction with care, and to delineate research questions related to mental health aspects of medical emergencies involving children. RESULTS: The consensus group found that psychological and behavioral factors affect physical as well as emotional recovery after medical emergencies. Children's reactions are critically affected by age and developmental level, characteristics of the emergency medical event, and parent reactions. As frontline health care providers, EMS staff members are in a pivotal position to recognize and effectively manage the mental health needs of patients and their families. CONCLUSIONS: Ecological changes in emergency departments, such as linkages to mental health follow-up services, training of EMS providers and mental health professionals, and focused research that provides an empirical basis for practice, are necessary components for improving current standards of health care.

Mental health aspects of emergency medical services for children: summary of a consensus conference.

OBJECTIVE: To address the mental health needs of children involved in emergency medical services (EMS). METHODS: A multidisciplinary consensus conference convened to identify mental health needs of children and their families related to pediatric medical emergencies, to examine the impact of psychological aspects of emergencies on recovery and satisfaction with care, and to delineate research questions related to mental health aspects of medical emergencies involving children. RESULTS: The consensus group found that psychological and behavioral factors affect physical as well as emotional recovery after medical emergencies. Children's reactions are critically affected by age and developmental level, characteristics of the emergency medical event, and parent reactions. As frontline health care providers, EMS staff members are in a pivotal position to recognize and effectively manage the mental health needs of patients and their families. CONCLUSIONS: Ecological changes in emergency departments, such as linkages to mental health follow-up services, training of EMS providers and mental health professionals, and focused research that provides an empirical basis for practice, are necessary components for improving current standards of health care.

Genetic tracing reveals a stereotyped sensory map in the olfactory cortex.

The olfactory system translates myriad chemical structures into diverse odour perceptions. To gain insight into how this is accomplished, we prepared mice that coexpressed a transneuronal tracer with only one of about 1,000 different odorant receptors. The tracer travelled from nasal neurons expressing that receptor to the olfactory bulb and then to the olfactory cortex, allowing visualization of cortical neurons that receive input from a particular odorant receptor. These studies revealed a stereotyped sensory map in the olfactory cortex in which signals from a particular receptor are targeted to specific clusters of neurons. Inputs from different receptors overlap spatially and could be combined in single neurons, potentially allowing for an integration of the components of an odorant's combinatorial receptor code. Signals from the same receptor are targeted to multiple olfactory cortical areas, permitting the parallel, and perhaps differential, processing of inputs from a single receptor before delivery to the neocortex and limbic system.

A comparison of ON-PUMP vs OFF-PUMP coronary artery bypass surgery among low, intermediate, and high-risk patients: the Hartford Hospital experience.

BACKGROUND: Off-pump coronary artery bypass (OP-CAB) graft surgery is being used with increasing frequency. This study was designed to compare OP-CAB outcomes with conventional surgical revascularization using cardiopulmonary bypass (CPB) in patients with varying risk categories at a high-volume center. METHODS AND RESULTS: Between 1/1/1999 and 1/31/2001, bypass surgery was performed on 1,312 patients, including 348 OP-CAB cases and 964 CPB cases. Compared to CPB cases, OP-CAB patients were more likely to be female and had a lower incidence of three vessel coronary artery disease, prior percutaneous intervention, and prior bypass surgery. Postoperatively, OP-CAB patients had a lower incidence of renal failure and prolonged ventilatory support, as well as a lower composite endpoint of inhospital mortality, perioperative myocardial infarction, cerebrovascular accident, and/or renal failure. In addition, OP-CAB patients required fewer transfusions and had a shorter total length of hospital stay. In general, morbidity and mortality increased in both OP-CAB and CPB groups with increasing Parsonnet score. CONCLUSIONS: OP-CAB surgery is a safe and effective alternative to conventional coronary artery bypass graft (CABG) surgery, with a lower incidence of major in-hospital adverse clinical events and a decreased requirement for medical resources. Adverse OP-CAB outcomes correlate well with pre-operative Parsonnet Score.

Polio, hepatitis B and AIDS: an integrative theory on a possible vaccine induced pandemic.

The hypothesis that simian virus 40 (SV40) infected polio vaccines may be linked to the evolution of acquired immunodeficiency disorder (AIDS), and certain cancers, has been advanced. Most recently, investigators discussed the likelihood of gene-reshuffling following SV40 infection as a precursor to acquired immune dysfunction. Findings of recent SV40 infections in four children born after 1982 suggest infections were transmitted vertically along gene lines. Earlier observations proved activation of a retrovirus gene by a hepatitis B virus (HBV) protein. This paper proposes a new integrative theory on the origin of AIDS. It advances the possibility of genetic recombinations with oncogene activation by HBV involving simian viruses that likely infected polio vaccinated blood donors to the initial hepatitis B (HB) vaccine trials conducted on gay men in New York City and Ugandan Blacks in the early to mid-1970s. The socio-economic and even military ramifications associated with this politically challenging thesis are discussed.

Detecting suicide risk in a pediatric emergency department: development of a brief screening tool.

OBJECTIVE: To develop a brief screening tool that will allow emergency department (ED) staff to rapidly and accurately detect suicide risk in child and adolescent patients. DESIGN: Cross-sectional survey. Participants. One hundred forty-four children and adolescents, mean age of 13.6 years, presenting to an urban university teaching hospital pediatric ED for primarily psychiatric reasons. Data Collection. As part of a quality improvement initiative, we developed a 14-item screening survey (the Risk of Suicide Questionnaire [RSQ]) that was administered by a triage nurse to all pediatric mental health patients on admission to the ED. All patients were subsequently administered the 30-item Suicide Ideation Questionnaire (SIQ) by a mental health clinician, which served as the criterion standard assessment of suicidality. Other information collected included demographic and clinical characteristics. MAIN OUTCOME MEASURES: Sensitivity, specificity, positive predictive value, negative predictive value (NPV), and area under the receiver operating characteristic curve for responses to individual and combinations of RSQ items, relative to determinations of suicidality by the criterion standard SIQ. RESULTS: Four of the items from the RSQ had a predictive c statistic of 0.87, a sensitivity of 0.98, and a NPV of 0.97. Little improvement in predictive ability was obtained by including other RSQ items (c statistic for the most predictive 4-item model = 0.87; c statistic for the model containing all 14 items = 0.90). Among all possible combinations of 4 RSQ items, the combination of items inquiring about current suicidal behavior, past suicidal ideation, past self-destructive behavior, and current stressors yielded the highest sensitivity (0.98), NPV (0.97), and c statistic (0.87), as assessed by the criterion standard SIQ. CONCLUSIONS: A brief 4-item screening tool can be used by nonmental health clinicians to accurately detect suicidality in children and adolescents who visit an ED. Early and accurate identification of suicidality is a critical first step that could lead to better treatment and improved health outcomes for children and adolescents with mental health concerns.

[Validation of the Inventory for Interpersonal Problems (IIP). Results of a representative study in East and West Germany]. / Zur Validierung des Inventars zur Erfassung Interpersonaler Probleme (IIP). Ergebnisse einer Repräsentativbefragung in Ost- und Westdeutschland.

The Inventory of Interpersonal Problems (IIP) was developed in the USA in the late 1970s and in the mean time has become a well-established instrument applied in numerous clinical studies in German speaking countries under the name "Inventar zur Erfassung interpersonaler Probleme". The test manual was published in German in 1994, but without the corresponding standardisation for a German-speaking population. The present study fills this gap. In a representative study the IIP was completed by 2025 West Germans and 1022 East Germans aged 14-92 years. The representative norms are given and the psychometric dimensions of the questionnaire adjusted for this sample. As a contribution to testing validity the results were also related to the scores of a number of other tests completed by the same sample (Giessen Test--GT; Giessener Beschwerdebogen--GBB; Fragebogen zur Lebenszufriedenheit--FLZ; Angstbewältigungsinventar--ABI and the Fragebogen zum erinnerten elterlichen Erziehungsverhalten [FEE]). The relevant results will be reported.

A genetic approach to trace neural circuits.

Mammalian nervous system function involves billions of neurons which are interconnected in a multitude of neural circuits. Here we describe a genetic approach to chart neural circuits. By using an olfactory-specific promoter, we selectively expressed barley lectin in sensory neurons in the olfactory epithelium and vomeronasal organ of transgenic mice. The lectin was transported through the axons of those neurons to the olfactory bulb, transferred to the bulb neurons with which they synapse, and transported through the axons of bulb neurons to the olfactory cortex. The lectin also was retrogradely transported from the bulb to neuromodulatory brain areas. No evidence could be obtained for adverse effects of the lectin on odorant receptor gene expression, sensory axon targeting in the bulb, or the generation or transmission of signals by olfactory sensory neurons. Transneuronal transfer was detected prenatally in the odor-sensing pathway, but only postnatally in the pheromone-sensing pathway, suggesting that odors, but not pheromones, may be sensed in utero. Our studies demonstrate that a plant lectin can serve as a transneuronal tracer when its expression is genetically targeted to a subset of neurons. This technology can potentially be applied to a variety of vertebrate and invertebrate neural systems and may be particularly valuable for mapping connections formed by small subsets of neurons and for studying the development of connectivity as it occurs in utero.

Characterization of topotecan-mediated redistribution of DNA topoisomerase I by digital imaging microscopy.

Topographical image measures have been used to characterize the subnuclear distribution of DNA topoisomerase I in human tumor cell lines. This topographical analysis allowed a mathematical description of staining patterns to be produced that did not depend on subjective grading. The redistribution of topoisomerase I in response to increasing concentrations of topotecan was then monitored by this method. The cell lines were stained for topoisomerase I by indirect immunofluorescence methods. Digital imaging microscopy and image analysis were used to extract the nucleus from each cell, and nine parameters describing the topography of the distribution of topoisomerase I within the nucleus were computed for each. Use of multivariate analysis of variance enabled this nine-parameter set to be reduced to a single canonical variable, representing 60-90% of the observed internuclear variance. Plotting the canonical variable vs drug concentration resulted in dose-response curves that could be fitted well by a simple Emax model. From these curve fits, EC50 and Emax values for drug-induced redistribution of topoisomerase I were determined. Our results indicate that neither the maximum extent of topoisomerase I redistribution (Emax) nor the EC50 for drug-induced redistribution correlated well with the growth inhibition produced by continuous exposure to topotecan in these cell lines. However, the EC50 determined for the 1-h high-concentration exposure did reflect the growth inhibition produced in cells exposed to the drug for 1 h. The methodology described may also be generally applied to any antigen of interest.

Patterns of interpersonal problems associated with binge eating disorder.

OBJECTIVE: To determine if interpersonal problems reported by individuals with binge eating disorder (BED) are distinct from psychiatric norms, and whether specific types of interpersonal problems are predictive of BED treatment outcome. METHOD: Subjects were 88 females with BED who completed a treatment trial for binge eating and weight loss. Pretreatment data on the Inventory of Interpersonal Problems (IIP) was compared (1) to that of psychiatric and nonpsychiatric norms and (2) for individuals with good versus poor outcome for eating disorder symptoms and weight loss. RESULTS: BED patients reported less distress for problems with being too vindictive (hostile dominance) than psychiatric norms. Patients with good eating disorder outcome reported less distress for problems with social avoidance; patients with good weight loss outcome reported greater distress over problems with vindictiveness. DISCUSSION: BED treatment may be enhanced by an initial focus on specific interpersonal difficulties.

Factors affecting utilization of treatment services by sexually abused girls.

This study describes the naturalistic therapy experiences of a sample of sexually abused girls and the relationship of these experiences to demographic factors, abuse experiences, psychopathology, and family functioning. The sample consisted of 81 sexually abused girls, aged 6 to 16, participating in a longitudinal study of the effects of sexual abuse. Results indicated strong effects for abuse experiences and child psychopathology on the total amount of therapy received. Patterns of treatment utilization were associated with ethnic minority status, but these differences are confounded by differing abuse experiences for racial groups in the sample. Other patterns of treatment utilization are discussed, as well as issues for further research and implications for treatment providers.