Epidemiology of training injuries in amateur taekwondo athletes: a retrospective cohort study

The objectives of this study were to estimate the incidence and describe the pattern and severity of training injuries in taekwondo, and to compare pattern and severity of training injuries with competition injuries. One hundred and fifty-two active Australian amateur taekwondo athletes, aged 12 years or over, completed an online survey comprising questions on training exposure and injury history over the preceding 12 months. The main outcome measures were: overall injury incidence rate per athlete-year; training injury incidence rate per athlete-year, per 1000 athlete-training-sessions, and per 1000 athlete-hours of training; injury severity; and injury proportions by anatomical region and by type of injury. Injury incidence rates were calculated with 95% confidence intervals using standard methods, while injury proportions were compared using Fisher''s exact test. The vast majority (81.5%) of taekwondo injuries in an average athlete-year occurred during training. The training injury incidence rate was estimated to be 1.6 (95% CI: 1.4, 1.9) per athlete-year, 11.8 (95% CI: 10.4, 13.4) per 1000 athlete-training-sessions, and 7.0 (95% CI: 6.1, 7.9) per 1000 athlete-hours of training. Among athletes with five or fewer injuries, the severity and injury pattern of training injuries were, by and large, the same as for competition injuries. Approximately sixty percent (60.3%) of training injuries required treatment by a health professional. Considering the burden of training injuries exceeds that of competition injuries, taekwondo governing bodies and stakeholders are encouraged to devote more efforts towards the identification of risk factors for, and prevention of, training injuries in the sport of taekwondo.     

Complementation of in vitro-assembled spliceosomes

We describe the development and application of a system of in vitro-assembled splicing complexes that can be used for the identification of protein splicing factors which become associated with the spliceosome at the end of the assembly process ("late" splicing components). A splicing reaction performed in the presence of polyvinyl alcohol is interrupted after 15 to 20 minutes, before the appearance of splicing intermediates and products in significant amounts. Following low-speed centrifugation, a pellet is obtained containing splicing complexes that can be solubilized with 0.6 M-KCl. These complexes can be rapidly complemented for splicing in the presence of ATP and Mg2+ with protein factors that are present in HeLa cell nuclear extracts or in chromatographic extract fractions. Biochemical features of the complementation reactions, and conditions for reversible uncoupling of the two splicing steps, are described and discussed. These conditions are used to generate fully assembled spliceosomes in which splicing of the pre-mRNA can occur in the presence of ATP and Mg2+, but in the absence of nuclear extract ("autonomous splicing").     

Spatial distribution and yield of DNA double-strand breaks induced by 3-7 MeV helium ions in human fibroblasts

Accelerated helium ions with mean energies at the target location of 3-7 MeV were used to simulate alpha-particle radiation from radon daughters. The experimental setup and calibration procedure allowed determination of the helium-ion energy distribution and dose in the nuclei of irradiated cells. Using this system, the induction of DNA double-strand breaks and their spatial distributions along DNA were studied in irradiated human fibroblasts. It was found that the apparent number of double-strand breaks as measured by a standard pulsed-field gel assay (FAR assay) decreased with increasing LET in the range 67-120 keV/microm (corresponding to the energy of 7-3 MeV). On the other hand, the generation of small and intermediate-size DNA fragments (0.1-100 kbp) increased with LET, indicating an increased intratrack long-range clustering of breaks. The fragment size distribution was measured in several size classes down to the smallest class of 0.1-2 kbp. When the clustering was taken into account, the actual number of DNA double-strand breaks (separated by at least 0.1 kbp) could be calculated and was found to be in the range 0.010-0.012 breaks/Mbp Gy(-1). This is two- to threefold higher than the apparent yield obtained by the FAR assay. The measured yield of double-strand breaks as a function of LET is compared with theoretical Monte Carlo calculations that simulate the track structure of energy depositions from helium ions as they interact with the 30-nm chromatin fiber. When the calculation is performed to include fragments larger than 0.1 kbp (to correspond to the experimental measurements), there is good agreement between experiment and theory.     

Autonomous splicing and complementation of in vivo-assembled spliceosomes

We have used an in vivo system generating assayable amounts of a specific pre-mRNA to study the relationship between splicing and an operationally defined nuclear matrix preparation (NM). When NM is prepared by extraction of DNase I-treated nuclei with an approximately physiological concentration of KCl (0.1 M), a portion of NM-associated precursor can be spliced in vitro in the presence of ATP and Mg2+ and in the absence of splicing extract ("autonomous splicing"). We propose that the autonomous reaction, which does not exhibit a temporal lag and is half-complete in 5 min, occurs in fully assembled, matrix-bound ribonucleoprotein complexes (in vivo spliceosomes). Extraction of the NM with concentrations of KCl greater than 0.4 M eliminates autonomous splicing but leaves behind preassembled complexes that can be complemented for splicing with HeLa cell nuclear extract. The splicing complementing factor, representing one or more activities present in the nuclear extract and also in the cytoplasmic S100 fraction, is relatively heat resistant, devoid of an RNA component, and does not bind to DEAE-Sepharose in 0.1 M KCl. It exists in the nucleus in two forms; bound to autonomous spliceosomes and free in the nucleoplasm. Biochemical features of the complementation reaction, and conditions for reversible uncoupling of the two splicing steps are described and discussed.     

Variations in Multiple Birth Rates and Impact on Perinatal Outcomes in Europe

Objective

Infants from multiple pregnancies have higher rates of preterm birth, stillbirth and neonatal death and differences in multiple birth rates (MBR) exist between countries. We aimed to describe differences in MBR in Europe and to investigate the impact of these differences on adverse perinatal outcomes at a population level.

Methods

We used national aggregate birth data on multiple pregnancies, maternal age, gestational age (GA), stillbirth and neonatal death collected in the Euro-Peristat project (29 countries in 2010, N = 5 074 643 births). We also used European Society of Human Reproduction and Embryology (ESHRE) data on assisted conception and single embryo transfer (SET). The impact of MBR on outcomes was studied using meta-analysis techniques with random-effects models to derive pooled risk ratios (pRR) overall and for four groups of country defined by their MBR. We computed population attributable risks (PAR) for these groups.

Results

In 2010, the average MBR was 16.8 per 1000 women giving birth, ranging from 9.1 (Romania) to 26.5 (Cyprus). Compared to singletons, multiples had a nine-fold increased risk (pRR 9.4, 95% Cl 9.1–9.8) of preterm birth (<37 weeks GA), an almost 12-fold increased risk (pRR 11.7, 95% CI 11.0–12.4) of very preterm birth (<32 weeks GA). Pooled RR were 2.4 (95% Cl 1.5–3.6) for fetal mortality at or after 28 weeks GA and 7.0 (95% Cl 6.1–8.0) for neonatal mortality. PAR of neonatal death and very preterm birth were higher in countries with high MBR compared to low MBR (17.1% (95% CI 13.8–20.2) versus 9.8% (95% Cl 9.6–11.0) for neonatal death and 29.6% (96% CI 28.5–30.6) versus 17.5% (95% CI 15.7–18.3) for very preterm births, respectively).

Conclusions

Wide variations in MBR and their impact on population outcomes imply that efforts by countries to reduce MBR could improve perinatal outcomes, enabling better long-term child health.     

DNA markers and crossbreeding scheme as means to select sires for heterosis in egg production of chickens

Genotypes for 24 microsatellite markers, dispersed across the chicken genome, were used to predict progeny performance and heterosis for egg production (number and mass) in 'layers' (egg-type chickens). These markers were used to evaluate genetic distance between each of 39 sires sampled from two-layer male-lines; Rhode Island Red (RIR) and White egg Leghorn (Leghorn), and a DNA pool of 30 randomly sampled females from a Brown-egg female line (Silver). Each sire was analysed for egg production across months in the laying period and cumulatively in each of three subperiods; onset (2 month), mid (9 month) and late (1 month). The average Reynolds' genetic distance between Leghorn sires and the Silver female line (theta;=0.6) was significantly higher than that between RIR sires and the Silver female line (theta;=0.5). Neither performance nor heterosis values in the RIR sire's daughters were associated with genetic distance values between sires and the Silver female line. On the other hand, performance as well as heterosis values of Leghorn's daughters were positively associated with genetic distance. This association was particularly evident in the mid-subperiod. If 25% of the most genetically distant Leghorn sires from the Silver female line had been selected in a single generation on the basis of DNA markers information only, average egg production of the crossbred daughters would have been improved by about nine eggs (3%). In principle, further improvement is possible if selection to increase genetic distance between the parental lines is carried on.     

cis-Acting elements within CFTR 5'-flanking DNA are not sufficient to decrease gene expression in response to phorbol ester

The cystic fibrosis transmembrane conductance regulator gene (CFTR) is regulated in a tissue-specific and developmental fashion. Although it has been known for some time that phorbol esters decrease CFTR expression in cell lines that have high CFTR mRNA levels, the cis-acting elements that control this down-regulation remain ill-defined. The role of cis-acting elements within the CFTR minimal promoter in modulating responses to phorbol 12-myristate 13-acetate (PMA) and forskolin was assessed using luciferase reporter gene (luc)-containing plasmids transfected into Calu-3 and HT-29 cells. PMA treatment had no effect on luciferase activity in Calu-3 cells transiently transfected with plasmids containing luc driven by up to 2.3 kb of CFTR 5'-flanking DNA. PMA increased luciferase activity in transfected HT-29 cells. A more extensive region of DNA was evaluated using a yeast artificial chromosome (YAC) containing luc driven by approximately 335 of CFTR 5'-flanking DNA (y5'luc) stably introduced into HT-29 cells. Clonal cell lines containing y5'luc were created and assessed for luciferase activity at baseline and in response to forskolin and PMA. There was a wide range of baseline luciferase activities among the clones (42-1038 units/microg protein) that was not entirely due to the number of luc copies present within the cells. Treatment with both PMA and forskolin led to increased luciferase activity in six randomly selected clonal cell lines. As expected, endogenous CFTR expression increased in response to forskolin and decreased in response to PMA. These studies demonstrate that luc-containing YAC vectors can be used to study CFTR expression in human cells. In addition, these data suggest that important regulatory elements responsible for decreased CFTR expression in response to PMA are not located upstream of CFTR in the approximately 335 kb 5'-flanking sequence included in this YAC construct.     

Single-step detection of mutant huntingtin in animal and human tissues: A bioassay for Huntington’s disease

The genetic mutation causing Huntington’s disease is a polyglutamine expansion in the huntingtin protein where more than 37 glutamines cause disease by formation of toxic intracellular fragments, aggregates, and cell death. Despite a clear pathogenic role for mutant huntingtin, understanding huntingtin expression during the presymptomatic phase of the disease or during disease progression has remained obscure. Central to clarifying the role in the pathomechanism of disease is the ability to easily and accurately measure mutant huntingtin in accessible human tissue samples as well as cell and animal models. Here we describe a highly sensitive time-resolved Förster resonance energy transfer (FRET) assay for quantification of soluble mutant huntingtin in brain, plasma, and cerebrospinal fluid. Surprisingly, in mice, soluble huntingtin levels decrease during disease progression, inversely correlating with brain aggregate load. Mutant huntingtin is easily detected in human brain and blood-derived fractions, providing a utility to assess mutant huntingtin expression during disease course as well as a pharmacodynamic marker for disease-modifying therapeutics targeting expression, cleavage, or degradation of mutant huntingtin. The design of the homogeneous one-step method for huntingtin detection is such that it can be easily applied to measure other proteins of interest.