Gap junction mediated bioelectric coordination is required for slow muscle development, organization, and function.

Bioelectrical signaling, intercellular communication facilitated by membrane potential and electrochemical coupling, is emerging as a key regulator of animal development. Gap junction (GJ) channels can mediate bioelectric signaling by creating a fast, direct pathway between cells for the movement of ions and other small molecules. In vertebrates, GJ channels are formed by a highly conserved transmembrane protein family called the Connexins. The connexin gene family is large and complex, presenting a challenge in identifying the specific Connexins that create channels within developing and mature tissues. Using the embryonic zebrafish neuromuscular system as a model, we identify a connexin conserved across vertebrate lineages, gjd4, which encodes the Cx46.8 protein, that mediates bioelectric signaling required for appropriate slow muscle development and function. Through a combination of mutant analysis and in vivo imaging we show that gjd4/Cx46.8 creates GJ channels specifically in developing slow muscle cells. Using genetics, pharmacology, and calcium imaging we find that spinal cord generated neural activity is transmitted to developing slow muscle cells and synchronized activity spreads via gjd4/Cx46.8 GJ channels. Finally, we show that bioelectrical signal propagation within the developing neuromuscular system is required for appropriate myofiber organization, and that disruption leads to defects in behavior. Our work reveals the molecular basis for GJ communication among developing muscle cells and reveals how perturbations to bioelectric signaling in the neuromuscular system_may contribute to developmental myopathies. Moreover, this work underscores a critical motif of signal propagation between organ systems and highlights the pivotal role played by GJ communication in coordinating bioelectric signaling during development.

Assessing South African medical interns' experience and confidence in managing obstetric emergencies.

BACKGROUND: Medical doctors in South Africa (SA) are required to complete a 2-year internship at training hospitals, including a 4-month rotation in obstetrics and gynaecology. Following this, doctors are allocated to community service posts, many of which are at district- and primary-level facilities where supervision is limited. Recent triennial Saving Mothers reports identify district hospitals (DHs) as the second leading site for maternal deaths of all causes, the leading site for maternal deaths secondary to obstetric haemorrhage, and the most likely site for the lack of a skilled doctor to be identified as a factor in deaths associated with caesarean delivery. OBJECTIVES: To describe the self-perceived readiness of medical interns completing their training to manage obstetric emergencies, based on the Essential Steps in the Management of Obstetric Emergencies modules in the Health Professions Council of South Africa's internship logbook. METHODS: This cross-sectional descriptive study assessed medical interns in the last 3 months of their training, using a self-administered online questionnaire. Data collection took place between October and December 2019. RESULTS: Cluster sampling of interns at training facilities throughout SA resulted in a total of 182 respondents from 17 hospitals in seven provinces in the country, with an overall response rate of 34.1%. Most interns had experience with and confidence in the management of miscarriage and hypertension in pregnancy. However, gaps in labour ward management, pregnancy-related sepsis and surgical skills were identified. Only 42.3% of respondents were confident in their ability to diagnose obstructed labour, 26.3% had performed an assisted delivery, 39.0% were confident in their knowledge of the indications for and contraindications to assisted deliveries, and 35.7% had been involved in the delivery of a baby with shoulder dystocia. Regarding pregnancy-related sepsis, 54.4% had experience with managing a wound abscess and 29.7% were confident managing puerperal endometritis. While 78.0% felt confident to perform a caesarean section (CS), only 28.6% had performed uterine compression suture for uterine atony at CS. Additionally, there was a statistically significant variation in scores between training hospitals. CONCLUSIONS: An incongruity exists between the shortcomings in DH obstetric services, the prioritisation of placement of community service doctors at primary healthcare facilities and DHs, and the self-perceived readiness of medical interns completing their training to manage obstetric emergencies safely. This situation highlights the importance of clinical support for junior doctors at DHs and standardisation of intern training at accredited facilities across SA.

Targeted candidate gene screens using CRISPR/Cas9 technology.

In the postgenomic era, the ability to quickly, efficiently, and inexpensively assign function to the zebrafish proteome is critical. Clustered regularly interspaced short palindromic repeats (CRISPRs) have revolutionized the ability to perform reverse genetics because of its simplicity and broad applicability. The CRISPR system is composed of an engineered, gene-specific single guide RNA (sgRNA) and a Cas9 enzyme that causes double-stranded breaks in DNA at the targeted site. This simple, two-part system, when injected into one-cell stage zebrafish embryos, efficiently mutates target loci at a frequency such that injected embryos phenocopy known mutant phenotypes. This property allows for CRISPR-based F0 screening in zebrafish, which provides a means to screen through a large number of candidate genes for their role in a phenotype of interest. While there are important considerations for any successful genetic screen, CRISPR screening has significant benefits over conventional methods and can be accomplished in any lab with modest molecular biology experience.

Incidence of advanced intravenous access in 2 urban EDs.

BACKGROUND: In an emergency department (ED), intravenous (IV) access is frequently accomplished by inspection and palpation of peripheral veins. Failure of these methods indicates severe IV access difficulty and necessitates advanced techniques. Here, we estimate the incidence of advanced IV access in 2 urban EDs with varying resident coverage. METHODS: In this multiple-cohort study, we enrolled data from 2 neighboring urban EDs-a tertiary care ED and a community hospital affiliate. The 2 have similar volumes but the tertiary care ED has more resident coverage (112 vs 20 hours/d). In a prospective data collection (April 2012-2013), we enrolled consecutive patients during hours of scheduled shifts for research assistants. In a retrospective data collection (March 2011-2012), we reviewed charts of a random sample of patients from each ED for similar outcomes. We calculated the incidence of advanced IV access by dividing the number requiring advanced techniques by the number requiring IV access. RESULTS: We determined IV outcomes for 790 patients in the prospective cohort and 669 patients in the retrospective cohort. Between groups, there was no difference in the incidence of advanced IV access in the prospective collection (P = .08) or in the retrospective collection (P = .7). Pooling data from both cohorts and both hospitals, the overall incidence was 3.2 [95% confidence interval, 1.9-5.2] per 100 attempts. CONCLUSION: Advanced IV access is needed in 3.2% of IV attempts in 2 urban EDs with varying levels of resident coverage. We found similar incidence in both EDs.

Condylar fractures of the proximal and middle phalanges.

We describe the treatment of 74 patients with phalangeal condylar fractures. Twelve patients presenting with undisplaced fractures were initially treated nonoperatively; of these, five displaced, requiring fixation. The remaining seven patients, all children, united uneventfully. Sixty-two patients presenting with displaced fractures were treated with internal fixation using a single lag screw through a lateral approach. The patients were treated semi-electively on a day surgery unit. Twenty-seven patients with unicondylar fractures, all operated on within 2 weeks of injury, regained full range of movement. Thirty-eight patients had loss of extension (range 10-35°) with fixed flexion contractures at the proximal interphalangeal and thumb interphalangeal joints and extensor lag at the distal interphalangeal joints (overall mean extension loss 10°). Although fixation was technically easier during the first week, a delay of 2 weeks before fixation made little difference to the outcome. In our experience, fractures can be taken down and fixed internally even 8 weeks after injury. If nonoperative treatment is initially embarked upon, close monitoring is required with weekly radiographs up to 3 weeks, as these fractures will frequently displace.

Gender differences in enrolment in an HIV-treatment programme in rural Lesotho, 2006-2008: a brief report.

The southern African nation of Lesotho has an HIV seroprevalence of approximately 25%. To address the need for HIV care in rural Lesotho, a project called the Rural Health Initative (RHI) was launched in seven clinics in 2006. Data on enrolment were collected retrospectively and analysed for trends in gender enrolment over time. Of 6001 enrolled, 3904 were women (65.1%) and 2097 (34.9%) were men. When analysed by month of enrolment, there was a higher percentage of men enrolled in December compared with the other months of the year (χ(2) = 15.98, P < 0.001). This may be due to the migratory work of the men in the mines of South Africa and suggests a need for targeted interventions to increase male enrolments over the entire calendar year.

Alcohol use and the management of multidrug-resistant tuberculosis in Tomsk, Russian Federation.

SETTING: Alcohol use increases the risk of multidrug-resistant tuberculosis (MDR-TB) and poses challenges for successful MDR-TB treatment, including the potential for additional adverse events. AIM: To investigate the association between alcohol consumption during MDR-TB treatment and adverse events and treatment outcomes in a cohort of patients in Tomsk, Russia. DESIGN: From 2000 to 2004, retrospective data were collected on 407 MDR-TB patients in Tomsk. Factors associated with treatment outcomes were assessed using logistic regression. RESULTS: Of the 407 patients, 253 (62.2%) consumed alcohol during treatment ('drinkers'), and 367 (90.2%) had at least one documented adverse advent. No significant differences were noted in frequency of adverse events in drinkers vs. non-drinkers. Drinkers had less favourable treatment outcomes (OR 0.28, 95%CI 0.18-0.45). Among drinkers, favourable treatment outcome was associated with adherence to at least 80% of prescribed doses (OR 2.89, 95%CI 1.30-6.43) and the occurrence of an adverse event requiring treatment interruption (OR 2.49, 95%CI 1.11-5.59). CONCLUSIONS: Alcohol use did not appear to increase the risk of adverse events during MDR-TB treatment; however, alcohol consumption was associated with poor outcome. Our findings suggest that individuals who drink alcohol should receive aggressive attention to optimise treatment adherence and manage adverse events.

Reliability of a common solution-based taste perception test: implications for validity and a briefer test.

The aim of this study was to assess test-retest reliability of a common method for quantifying taste perception and its association with gustatory responses and individual risk for obesity and related health conditions. Forty-six healthy adults rated 20 mixtures comprised of 5 dairy beverages varied in fat content and mixed with sugar concentrations of 0%, 5%, 10%, and 20%, following existing procedures. Individuals rated the sweetness, creaminess, and pleasantness of each mixture during two taste testing sessions occurring 7±2 days apart. Test-retest correlations were of the expected magnitudes (r≥.50) only for the pleasantness ratings of mixtures with higher sugar concentrations. Correlations for sweetness and creaminess taste perception ratings were low, indicating that such ratings may not be reliable over approximately one week, and challenging the validity of such ratings for measuring trait taste perception. A shortened version of the test may be warranted.

Continuous intravenous infusion of ketamine for maintenance sedation.

Ketamine HCl is a rapidly acting general anesthetic with sedative and analgesic properties that has been reported to have favorable effects on the cardiovascular and pulmonary systems. The goal of this review is to determine the hemodynamic and pulmonary effects of continuous intravenous (IV) ketamine infusion in mechanically ventilated patients, and to determine whether sufficient evidence exists to support its use as an agent for maintenance anesthesia. PubMed/Medline, EMBASE, and Index Medicus databases as well as relevant bibliographies were searched. Studies were independently evaluated for inclusion and exclusion criteria, as well as study parameters, by two evaluators. Any discrepancy was resolved by a third evaluator. Twenty studies (281 patients) met the inclusion criteria for this review including 11 prospective studies (250 patients). Data suggests that ketamine decreases airway resistance, improves dynamic compliance, and preserves functional residual capacity, minute ventilation and tidal volume, while retaining protective pharyngeal and laryngeal reflexes. In patients with refractory bronchospasm, continuous infusion of intravenous ketamine decreases audible wheeze, bronchodilator requirements, and hypercarbia. It also improves respiratory rate and oxygenation, and does not promote respiratory depression. Additionally, ketamine does not result in significant perturbations in blood pressure, heart rate, or vascular resistance. Ketamine may be a safe and effective tool for maintenance sedation of mechanically ventilated patients, however a large prospective clinical trial is warranted.

Controlled trial of active tuberculosis case finding in a Brazilian favela.

SETTING: A large, impoverished squatters' settlement (favela), Rio de Janeiro, Brazil. OBJECTIVE: To assess the community impact of active case finding for tuberculosis (TB) compared to an enhanced case-finding strategy. DESIGN: A pair-matched, cluster-randomized trial comparing household symptom screening and spot sputum collection (Arm 1) vs. distribution of an educational pamphlet (Arm 2) was performed in a large Brazilian favela. We compared TB case-notification rates, time from symptom onset to treatment start and treatment completion proportions between arms. Fourteen neighborhoods (estimated population 58,587) were pair-matched by prior TB case rates and randomly allocated to one of two interventions. TB was diagnosed using acid-fast bacilli smears. New TB cases were interviewed and clinic records were reviewed. RESULTS: A total of 193 TB cases were identified in the 14 study neighborhoods (incidence proportion 329 per 100,000 population). The case identification rate in Arm 1 was 934/100,000 person-years (py) vs. 604/100,000 py in Arm 2 (RR 1.55, 95%CI 1.10-1.99). No significant differences were found in time from cough onset to treatment start or proportion completing treatment. CONCLUSIONS: A door-to-door case-finding campaign was more effective (while ongoing) at detecting prevalent cases and influencing people to come for care than leafleting, but no differences were seen in time to treatment start or treatment completion.

The effect of weaner diet protein content and diet quality on the long-term performance of pigs to slaughter.

Short and long-term effects of manipulating dietary CP content and diet quality in weaner diets on health and performance of pigs were investigated in a 2 x 2 factorial combination of CP inclusion (high-CP, 230 g of CP/kg vs. low-CP, 170 g of CP/kg) and diet quality (high-quality, cooked cereals, and animal protein vs. low-quality, raw cereals, and plant protein). Diets were fed ad libitum for 14 d postweaning to pigs weaned at 29.4+/-3.1 d of age and 9.9+/-1.0 kg of BW. From d 14 to slaughter at 104+/-3 kg, all pigs were fed the same series of standard commercial diets. There were 15 replicates per treatment in the weaner phase (<30 kg) and 5 replicates per treatment in the grower-finisher phase (>30 kg). High-quality diets promoted gut health as indicated by improved fecal lactobacilli to coliform ratio (P=0.002) and decreased fecal enterotoxigenic Escherichia coli counts on d 11 postweaning (P=0.028), reducing the risk of postweaning diarrhea and improving pig health from weaning to the end of the weaner phase. Reducing CP content had no effect on gut health. High-CP (P=0.053) and high-quality (P=0.025) diets independently increased ADG during the first 14 d postweaning compared with low-CP and low-quality diets, respectively. There were no interactions between dietary CP content and quality on any of the response criteria investigated. Despite differences in the immediate postweaning period, there was no effect of manipulating diet quality or CP content for 2 wk postweaning on lifetime performance with pigs reaching slaughter weight in 128+/-7 d. These results indicate that high-quality diets may protect pig gut health during the immediate postweaning period. However, it may be possible to use less expensive, decreased quality weaner diets without any adverse effects on long-term performance when weaning older, heavier pigs and where health status, environmental control, and stock management are all maintained to a high standard.

The pathway study: results of a pilot feasibility study in patients suspected of having lung carcinoma investigated in a conventional chest clinic setting compared to a centralised two-stop pathway.

UNLABELLED: The best chance of cure in non-small cell lung cancer (NSCLC) is surgical resection, but UK rates of 8% compare poorly to 25% in the USA and Europe. Delays in diagnosis in the current UK system may be one reason for such discrepancy. To address this problem we set up a rapid diagnostic system and compared it to the conventional method of investigations in a pilot randomised trial. METHODS: Eighty-eight patients were prospectively enrolled from three District General Hospitals and randomised to either investigation locally or to the rapid system at The Royal Marsden Hospital. The pilot end-points were feasibility and audit of radical treatment rates to enable estimates for patient numbers for the full study. RESULTS: Forty-five and 43 patients were in the central and conventional arms, respectively (65% male, median age 69 years). There was a 4-week improvement in time to first treatment in those in the central arm (P=0.0025) with 13/30 (43%) and 9/27 (33%) patients having radical treatment in the central and conventional arms, respectively. Patients in the conventional arm felt the diagnostic process was too slow (P=0.02) while those in the central arm seemed to have a better care experience (P=0.01). There were significantly less visits to the general practitioner (GP) in the central arm (P=0.02). CONCLUSIONS: This pilot study demonstrates that the full study is feasible but would require the commitment and involvement of a large number of patients and physicians. The results show several advantages to investigations and diagnosis in the central arm, particularly in time to treatment initiation, patient satisfaction and rate of radical treatments. The improved rate of radical treatment could lead to an improved survival rate.

Radiation biodosimetry: applications for spaceflight.

The multiparametric dosimetry system that we are developing for medical radiological defense applications could be adapted for spaceflight environments. The system complements the internationally accepted personnel dosimeters and cytogenetic analysis of chromosome aberrations, considered the best means of documenting radiation doses for health records. Our system consists of a portable hematology analyzer, molecular biodosimetry using nucleic acid and antigen-based diagnostic equipment, and a dose assessment management software application. A dry-capillary tube reagent-based centrifuge blood cell counter (QBC Autoread Plus, Becton [correction of Beckon] Dickinson Bioscience) measures peripheral blood lymphocytes and monocytes, which could determine radiation dose based on the kinetics of blood cell depletion. Molecular biomarkers for ionizing radiation exposure (gene expression changes, blood proteins) can be measured in real time using such diagnostic detection technologies as miniaturized nucleic acid sequences and antigen-based biosensors, but they require validation of dose-dependent targets and development of optimized protocols and analysis systems. The Biodosimetry Assessment Tool, a software application, calculates radiation dose based on a patient's physical signs and symptoms and blood cell count analysis. It also annotates location of personnel dosimeters, displays a summary of a patient's dosimetric information to healthcare professionals, and archives the data for further use. These radiation assessment diagnostic technologies can have dual-use applications supporting general medical-related care.

Clonal relationships in a shelter-associated outbreak of drug-resistant tuberculosis: 1983-1997.

SETTING: An outbreak of tuberculosis caused by Mycobacterium tuberculosis resistant to isoniazid and streptomycin (HS-resistant) was documented in Boston's homeless population in 1984. Isolate relatedness was confirmed at the time by phage typing. In the late 1990s, cases of HS-resistant tuberculosis in the homeless were also documented, confirmed by RFLP typing using IS6110. None of the phage typed isolates from the 1980s were viable for performing RFLP analysis. We attempted to determine, using mixed-linker PCR (M-L PCR) finger-printing, whether or not these cases were all due to the same strain of M. tuberculosis. DESIGN: Isolates from 10 HS-resistant patients-four non-viable isolates from the 1980s and six viable isolates from 1996-1997-were sent to the Centers for Disease Control and Prevention for M-L PCR fingerprinting. These results were combined with record reviews of older cases and an ongoing epidemiologic investigation. RESULTS: Eight of 10 of the isolates were clonal, and the other two were strongly suspected matches. Epidemiologic investigation determined that transmission continued to occur after the initial outbreak in 1984-1985, and that a streptomycin-monoresistant variant of the strain was also circulating. CONCLUSION: M-L PCR fingerprinting combined with epidemiology was able to document links between cases across 15 years.

Observation of radiation-specific damage in human cells exposed to depleted uranium: dicentric frequency and neoplastic transformation as endpoints.

Depleted uranium (DU) is a dense heavy metal used primarily in military applications. Published data from our laboratory have demonstrated that DU exposure in vitro to immortalised human osteoblast cells (HOS) is both neoplastically transforming and genotoxic. DU possesses both a radiological (alpha-particle) and chemical (metal) component. Since DU has a low specific activity in comparison to natural uranium, it is not considered to be a significant radiological hazard. The potential contribution of radiation to DU-induced biological effects is unknown and the involvement of radiation in DU-induced biological effects could have significant implications for current risk estimates for internalised DU exposure. Two approaches were used to address this question. The frequency of dicentrics was measured in HOS cells following DU exposure in vitro. Data demonstrated that DU exposure (50 microM, 24 h) induced a significant elevation in dicentric frequency in vitro in contrast to incubation with the heavy metals, nickel and tungsten which did not increase dicentric frequency above background levels. Using the same concentration (50 microM) of three uranyl nitrate compounds that have different uranium isotopic concentrations and therefore, different specific activities, the effect on neoplastic transformation in vitro was examined. HOS cells were exposed to one of three-uranyl nitrate compounds (238U-uranyl nitrate, specific activity 0.33 microCi.g-1; DU-uranyl nitrate, specific activity 0.44 microCi.g-1; and 235U-uranyl nitrate, specific activity 2.2 microCi.g-1) delivered at a concentration of 50 microM for 24 h. Results showed, at equal uranium concentration, there was a specific activity dependent increase in neoplastic transformation frequency. Taken together these data suggest that radiation can play a role in DU-induced biological effects in vitro.

Proto-oncogene expression: a predictive assay for radiation biodosimetry applications.

Using a model system of in vitro human peripheral blood lymphocytes, the effect of low-dose (0.25 to 1.50 Gy) 250-kVp X ray radiation (1 Gy.min-1) on the expression of several proto-oncogenes was examined (c-Haras, c-src, c-met, c-jun, c-fos, and c-myc) and beta-actin from 0.25 to 17 h post-radiation. RNA was extracted from cells harvested at various times after exposure and examined for levels of particular mRNAs by northern blot hybridisation. A progressive time- and dose-dependent increase in mRNA levels was observed for c-Haras mRNA, while the other proto-oncogenes (c-src, c-met, c-fos, c-jun and c-myc) examined were variable during the same time period. beta-actin levels were initially decreased but at 17 h post-radiation had returned to control levels. A comparison of the rate of c-Haras transcription at 5 and 17 h post-irradiation revealed that c-Haras transcription was higher at 5 h than at 17 h. These findings suggest that the level of specific proto-oncogene expression, particularly c-Haras, may be useful early diagnostic molecular biomarkers for biodosimetry applications. The use of real-time PCR technologies to quantify gene expression changes will also be discussed.

Biological effects of embedded depleted uranium (DU): summary of armed forces radiobiology research institute research.

The Persian Gulf War resulted in injuries of US Coalition personnel by fragments of depleted uranium (DU). Fragments not immediately threatening the health of the individuals were allowed to remain in place, based on long-standing treatment protocols designed for other kinds of metal shrapnel injuries. However, questions were soon raised as to whether this approach is appropriate for a metal with the unique radiological and toxicological properties of DU. The Armed Forces Radiobiology Research Institute (AFRRI) is investigating health effects of embedded fragments of DU to determine whether current surgical fragment removal policies remain appropriate for this metal. These studies employ rodents implanted with DU pellets as well as cultured human cells exposed to DU compounds. Results indicate uranium from implanted DU fragments distributed to tissues far-removed from implantation sites, including bone, kidney, muscle, and liver. Despite levels of uranium in the kidney that were nephrotoxic after acute exposure, no histological or functional kidney toxicity was observed. However, results suggest the need for further studies of long-term health impact, since DU was found to be mutagenic, and it transformed human osteoblast cells to a tumorigenic phenotype. It also altered neurophysiological parameters in rat hippocampus, crossed the placental barrier, and entered fetal tissue. This report summarizes AFRRI's depleted uranium research to date.