A pipeline to quantify serum and cerebrospinal fluid microRNAs for diagnosis and detection of relapse in paediatric malignant germ-cell tumours

Background:The current biomarkers alpha-fetoprotein and human chorionic gonadotropin have limited sensitivity and specificity for diagnosing malignant germ-cell tumours (GCTs). MicroRNAs (miRNAs) from the miR-371-373 and miR-302/367 clusters are overexpressed in all malignant GCTs, and some of these miRNAs show elevated serum levels at diagnosis. Here, we developed a robust technical pipeline to quantify these miRNAs in the serum and cerebrospinal fluid (CSF). The pipeline was used in samples from a cohort of exclusively paediatric patients with gonadal and extragonadal malignant GCTs, compared with appropriate tumour and non-tumour control groups.Methods:We developed a method for miRNA quantification that enabled sample adequacy assessment and reliable data normalisation. We performed qRT-PCR profiling for miR-371-373 and miR-302/367 cluster miRNAs in a total of 45 serum and CSF samples, obtained from 25 paediatric patients.Results:The exogenous non-human spike-in cel-miR-39-3p and the endogenous housekeeper miR-30b-5p were optimal for obtaining robust serum and CSF qRT-PCR quantification. A four-serum miRNA panel (miR-371a-3p, miR-372-3p, miR-373-3p and miR-367-3p): (i) showed high sensitivity/specificity for diagnosing paediatric extracranial malignant GCT; (ii) allowed early detection of relapse of a testicular mixed malignant GCT; and (iii) distinguished intracranial malignant GCT from intracranial non-GCT tumours at diagnosis, using CSF and serum samples.Conclusions:The pipeline we have developed is robust, scalable and transferable. It potentially promises to improve clinical management of paediatric (and adult) malignant GCTs

Calculation of health expectancies with administrative data for North Rhine-Westphalia, a Federal State of Germany, 1999–2005

Pinheiro JP, Krämer A. Calculation of health expectancies with administrative data for North Rhine-Westphalia, a Federal State of Germany, 1999-2005. Population Health Metrics. 2009;7(1):4.OBJECTIVES: The main objectives of this study were to prove the feasibility of health expectancy analyses with regional administrative health statistics and to explore the utility of the calculated health expectancies in describing the health state of the population living in North Rhine-Westphalia, a Federal State of Germany. MATERIALS AND METHODS: Administrative population and mortality data as well as health data on disability and long-term care provided by public services were used to calculate: a) the life expectancy and b) the health expectancies Severe-Disability-Free Life Expectancy (SDFLE) and Long-Term-Care-Free Life Expectancy (LTCFLE) from 1999 to 2005. Calculations were done using the Sullivan method. RESULTS: SDFLE at birth was 69.9 years (males 66.2 and females 73.2 years) in 1999 and it increased to 71.7 years (males 68.6 and females 74.7 years) in 2005. The proportion of the SDFLE on the total life expectancy at birth was 89.8% (males 88.6 and females 90.8%) in 1999 and 90.7% (males 89.8 and females 91.4%) in 2005.LTCFLE at birth was 75.3 years (males 73.1 and females 77.5 years) in 1999 and it increased to 76.6 years (males 74.7 and females 78.6 years) in 2005. The proportion of the LTCFLE on the total life expectancy at birth was 96.8% (males 97.8 and females 96.1%) in 1999 and 96.8% (males 97.8 and females 96.2%) in 2005. DISCUSSION AND CONCLUSION: Both health expectancies indicate an improvement in the quantity as well as in the quality of healthy life for the population living in North Rhine Westphalia and therefore suggest a compression of morbidity from 1999 to 2005. The findings however have several limitations in their sensitivity, since we applied dichotomous valuations to the health states. In addition, the results are restricted to comparisons over time because the morbidity concepts do not allow for comparisons with populations other than the German one. Refined calculations with other summary measures of population health and with health data on other morbidity concepts are therefore reasonable

Central defect type partial ACL injury model on goat knees: the effect of infrapatellar fat pad excision

BACKGROUND: The mid-substance central defect injury has been used to investigate the primary healing capacity of the anterior cruciate ligament (ACL) in a goat model. The sagittal plane stability on this model has not been confirmed, and possible effects of fat pad excision on healing have not been evaluated. We hypothesize that excising the fat pad tissue results in poorer ligament healing as assessed histologically and decreased tensile strength of the healing ligament. We further hypothesize that the creation of a central defect does not affect sagittal plane knee stability. METHODS: A mid-substance central defect was created with a 4-mm arthroscopic punch in the ACLs of right knees of all the subjects through a medial mini-arthrotomy. Goats were assigned to groups based on whether the fat pad was preserved (group 1, n = 5) or excised completely (group 2, n = 5). The left knees served as controls in each goat. Histopathology of the defect area along with measurement of type I collagen in one goat from each group were performed at 10th week postoperatively. The remaining knees were evaluated biomechanically at the 12th week, by measuring anterior tibial translation (ATT) of the knee joints at 90° of flexion and testing tensile properties (ultimate tensile load (UTL), ultimate elongation (UE), stiffness (S), failure mode (FM)) of the femur-ACL-tibia complex. RESULTS AND DISCUSSION: Histopathology analysis revealed that the central defect area was fully filled macroscopically and microscopically. However, myxoid degeneration and fibrosis were observed in group 2 and increased collagen type I content was noted in group 2. There were no significant differences within and between groups in terms of ATT values (p = 0.715 and p = 0.149, respectively). There were no significance between or within groups in terms of ultimate tensile load and ultimate elongation; however, group 2 demonstrated greater stiffness than group 1 that was correlated with the fibrotic changes detected microscopically (p = 0.043). CONCLUSIONS: The central defect type injury model was confirmed to be biomechanically stable in a goat model. Resection of the fat pad was noted to negatively affect defect healing and increase ligament stiffness in the central defect injury model

Tolerability and safety of novel half milliliter formulation of glatiramer acetate for subcutaneous injection: an open-label, multicenter, randomized comparative study

Daily glatiramer acetate (GA) 20 mg/1.0 mL is a first-line treatment for relapsing-remitting multiple sclerosis (RRMS). To reduce the occurrence of injection pain and local injection site reactions (LISRs), a reduced volume formulation of GA was developed. This study compared pain and LISRs after injecting the marketed and the novel formulations. RRMS patients currently injecting GA participated in this multicenter, randomized, crossover comparative study. All patients administered once-daily subcutaneous injections of GA 20 mg/1.0 mL (marketed formulation) or GA 20 mg/0.5 mL (reduced volume formulation) for 14 days. Patients were crossed-over to the alternate treatment for an additional 14 days. Using a Visual Analog Scale (VAS), patients recorded in daily diaries the severity of injection pain immediately and 5 min post-injection, and the presence and severity of LISRs (swelling, redness, itching, lump) within 5 min and 24 h post-injection. VAS pain scores were ranked significantly lower immediately and 5 min after GA 20 mg/0.5 mL injections (p < 0.0001). Although LISRs were rare for both preparations, the severity of reactions ranked significantly lower and fewer symptoms occurred within 5 min and 24 h of using the reduced volume formulation (p < 0.0001). GA injected subcutaneously in a reduced volume formulation is a more tolerable option

Interactions and potential implications of Plasmodium falciparum-hookworm coinfection in different age groups in south-central Côte d'Ivoire

BACKGROUND: Given the widespread distribution of Plasmodium and helminth infections, and similarities of ecological requirements for disease transmission, coinfection is a common phenomenon in sub-Saharan Africa and elsewhere in the tropics. Interactions of Plasmodium falciparum and soil-transmitted helminths, including immunological responses and clinical outcomes of the host, need further scientific inquiry. Understanding the complex interactions between these parasitic infections is of public health relevance considering that control measures targeting malaria and helminthiases are going to scale.METHODOLOGY: A cross-sectional survey was carried out in April 2010 in infants, young school-aged children, and young non-pregnant women in south-central Côte d'Ivoire. Stool, urine, and blood samples were collected and subjected to standardized, quality-controlled methods. Soil-transmitted helminth infections were identified and quantified in stool. Finger-prick blood samples were used to determine Plasmodium spp. infection, parasitemia, and hemoglobin concentrations. Iron, vitamin A, riboflavin, and inflammation status were measured in venous blood samples.PRINCIPAL FINDINGS: Multivariate regression analysis revealed specific association between infection and demographic, socioeconomic, host inflammatory and nutritional factors. Non-pregnant women infected with P. falciparum had significantly lower odds of hookworm infection, whilst a significant positive association was found between both parasitic infections in 6- to 8-year-old children. Coinfected children had lower odds of anemia and iron deficiency than their counterparts infected with P. falciparum alone.CONCLUSIONS/SIGNIFICANCE: Our findings suggest that interaction between P. falciparum and light-intensity hookworm infections vary with age and, in school-aged children, may benefit the host through preventing iron deficiency anemia. This observation warrants additional investigation to elucidate the mechanisms and consequences of coinfections, as this information could have important implications when implementing integrated control measures against malaria and helminthiases

Sedation AND Weaning In Children (SANDWICH): protocol for a cluster randomised stepped wedge trial.

Introduction: Weaning from ventilation is a complex process involving several stages that include recognition of patient readiness to begin the weaning process; steps to reduce ventilation while optimising sedation in order not to induce distress; and removing the endotracheal tube. Delay at any stage can prolong the duration of mechanical ventilation. We developed a multi-component intervention targeted at helping clinicians to safely expedite this process and minimise the harms associated with unnecessary mechanical ventilation. Methods and analysis: This is a 20-month cluster-randomised stepped wedge clinical and cost-effectiveness trial with an internal pilot and a process evaluation. It is being conducted in 18 paediatric intensive care units in the UK to evaluate a protocol-based intervention for reducing the duration of invasive mechanical ventilation. Following an initial eight-week baseline data collection period in all sites, one site will be randomly chosen to transition to the intervention every four weeks and will start an eight-week training period after which it will continue the intervention for the remaining duration of the study. We aim to recruit approximately 10,000 patients. The primary analysis will compare data from before the training (control) with that from after the training (intervention) in each site. Full details of the analyses will be in the statistical analysis plan. Ethics and dissemination: This Protocol was reviewed and approved by NRES Committee East Midlands - Nottingham 1 Research Ethics Committee (reference: 17/EM/0301). All sites started patient recruitment on 5 February 2018 before randomisation in April 2018. Results will be disseminated in 2020. The results will be presented at national and international conferences and published in peer reviewed medical journals

The present and future of serum diagnostic tests for testicular germ cell tumours.

Testicular germ cell tumours (GCTs) are the most common malignancy occurring in young adult men and the incidence of these tumours is increasing. Current research priorities in this field include improving overall survival for patients classified as being 'poor-risk' and reducing late effects of treatment for patients classified as 'good-risk'. Testicular GCTs are broadly classified into seminomas and nonseminomatous GCTs (NSGCTs). The conventional serum protein tumour markers α-fetoprotein (AFP), human chorionic gonadotrophin (hCG) and lactate dehydrogenase (LDH) show some utility in the management of testicular malignant GCT. However, AFP and hCG display limited sensitivity and specificity, being indicative of yolk sac tumour (AFP) and choriocarcinoma or syncytiotrophoblast (hCG) subtypes. Furthermore, LDH is a very nonspecific biomarker. Consequently, seminomas and NSGCTs comprising a pure embryonal carcinoma subtype are generally negative for these conventional markers. As a result, novel universal biomarkers for testicular malignant GCTs are required. MicroRNAs are short, non-protein-coding RNAs that show much general promise as biomarkers. MicroRNAs from two 'clusters', miR-371-373 and miR-302-367, are overexpressed in all malignant GCTs, regardless of age (adult or paediatric), site (gonadal or extragonadal) and subtype (seminomas, yolk sac tumours or embryonal carcinomas). A panel of four circulating microRNAs from these two clusters (miR-371a-3p, miR-372-3p, miR-373-3p and miR-367-3p) is highly sensitive and specific for the diagnosis of malignant GCT, including seminoma and embryonal carcinoma. In the future, circulating microRNAs might be useful in diagnosis, disease monitoring and prognostication of malignant testicular GCTs, which might also reduce reliance on serial CT scanning. For translation into clinical practice, important practical considerations now need addressing.The authors would like to acknowledge grant funding from CwCUK/GOSHCC (M.J.M. N.C. grant W1058), SPARKS (M.J.M. N.C. grant 11CAM01), CRUK (N.C. grant A13080) MRC (M.J.M. grant MC_EX_G0800464) and National Health Service funding to the Royal Marsden/Institute of Cancer Research National Institute for Health Research Biomedical Research Centre for Cancer (R.A.H.). The authors also thank the Max Williamson Fund, the Josh Carrick Foundation and The Perse Preparatory School, Cambridge for support.This is the author accepted manuscript. The final version is available fromNature Publishing Group via https://doi.org/10.1038/nrurol.2016.17

Smc5/6 coordinates formation and resolution of joint molecules with chromosome morphology to ensure meiotic divisions

During meiosis, Structural Maintenance of Chromosome (SMC) complexes underpin two fundamental features of meiosis: homologous recombination and chromosome segregation. While meiotic functions of the cohesin and condensin complexes have been delineated, the role of the third SMC complex, Smc5/6, remains enigmatic. Here we identify specific, essential meiotic functions for the Smc5/6 complex in homologous recombination and the regulation of cohesin. We show that Smc5/6 is enriched at centromeres and cohesin-association sites where it regulates sister-chromatid cohesion and the timely removal of cohesin from chromosomal arms, respectively. Smc5/6 also localizes to recombination hotspots, where it promotes normal formation and resolution of a subset of joint-molecule intermediates. In this regard, Smc5/6 functions independently of the major crossover pathway defined by the MutLγ complex. Furthermore, we show that Smc5/6 is required for stable chromosomal localization of the XPF-family endonuclease, Mus81-Mms4Eme1. Our data suggest that the Smc5/6 complex is required for specific recombination and chromosomal processes throughout meiosis and that in its absence, attempts at cell division with unresolved joint molecules and residual cohesin lead to severe recombination-induced meiotic catastroph

Measuring the Meltdown: Drivers of Global Amphibian Extinction and Decline

Habitat loss, climate change, over-exploitation, disease and other factors have been hypothesised in the global decline of amphibian biodiversity. However, the relative importance of and synergies among different drivers are still poorly understood. We present the largest global analysis of roughly 45% of known amphibians (2,583 species) to quantify the influences of life history, climate, human density and habitat loss on declines and extinction risk. Multi-model Bayesian inference reveals that large amphibian species with small geographic range and pronounced seasonality in temperature and precipitation are most likely to be Red-Listed by IUCN. Elevated habitat loss and human densities are also correlated with high threat risk. Range size, habitat loss and more extreme seasonality in precipitation contributed to decline risk in the 2,454 species that declined between 1980 and 2004, compared to species that were stable (n = 1,545) or had increased (n = 28). These empirical results show that amphibian species with restricted ranges should be urgently targeted for conservation