Assessment of Allergen-Responsive Regulatory T Cells in Experimental Asthma Induced in Different Mouse Strains.

BACKGROUND: Regulatory T cells (Tregs) are important in regulating responses to innocuous antigens, such as allergens, by controlling the Th2 response, a mechanism that appears to be compromised in atopic asthmatic individuals. Different isogenic mouse strains also have distinct immunological responses and susceptibility to the experimental protocols used to develop lung allergic inflammation. In this work, we investigated the differences in the frequency of Treg cell subtypes among A/J, BALB/c, and C57BL/6, under normal conditions and following induction of allergic asthma with ovalbumin (OVA). METHODS: Subcutaneous sensitization followed by 4 consecutive intranasal OVA challenges induced asthma characteristic changes such as airway hyperreactivity, inflammation, and production of Th2 cytokines (IL-4, IL-13, IL-5, and IL-33) in the lungs of only A/J and BALB/c but not C57BL/6 strain and evaluated by invasive whole-body plethysmography, flow cytometry, and ELISA, respectively. RESULTS: A/J strain naturally showed a higher frequency of CD4+IL-10+ T cells in the lungs of naïve mice compared to the other strains, accompanied by higher frequencies of CD4+IL-4+ T cells. C57BL/6 mice did not develop lung inflammation and presented higher frequency of CD4+CD25+Foxp3+ Treg cells in the bronchoalveolar lavage fluid (BALF) after the allergen challenge. In in vitro settings, allergen-specific stimulation of mediastinal LN (mLN) cells from OVA-challenged animals induced higher frequency of CD4+IL-10+ Treg cells from A/J strain and CD4+CD25+Foxp3+ from C57BL/6. CONCLUSIONS: The observed differences in the frequencies of Treg cell subtypes associated with the susceptibility of the animals to experimental asthma suggest that CD4+CD25+Foxp3+ and IL-10-producing CD4+ Treg cells may play different roles in asthma control. Similar to asthmatic individuals, the lack of an efficient regulatory response and susceptibility to the development of experimental asthma in A/J mice further suggests that this strain could be preferably chosen in experimental models of allergic asthma.

Enhanced influenza A H1N1 T cell epitope recognition and cross-reactivity to protein-O-mannosyltransferase 1 in Pandemrix-associated narcolepsy type 1.

Narcolepsy type 1 (NT1) is a chronic neurological disorder having a strong association with HLA-DQB1*0602, thereby suggesting an immunological origin. Increased risk of NT1 has been reported among children or adolescents vaccinated with AS03 adjuvant-supplemented pandemic H1N1 influenza A vaccine, Pandemrix. Here we show that pediatric Pandemrix-associated NT1 patients have enhanced T-cell immunity against the viral epitopes, neuraminidase 175-189 (NA175-189) and nucleoprotein 214-228 (NP214-228), but also respond to a NA175-189-mimic, brain self-epitope, protein-O-mannosyltransferase 1 (POMT1675-689). A pathogenic role of influenza virus-specific T-cells and T-cell cross-reactivity in NT1 are supported by the up-regulation of IFN-γ, perforin 1 and granzyme B, and by the converging selection of T-cell receptor TRAV10/TRAJ17 and TRAV10/TRAJ24 clonotypes, in response to stimulation either with peptide NA175-189 or POMT1675-689. Moreover, anti-POMT1 serum autoantibodies are increased in Pandemrix-vaccinated children or adolescents. These results thus identify POMT1 as a potential autoantigen recognized by T- and B-cells in NT1.

Is it possible to predict the origin of epithelial cells? - A comparison of secondary transfer of skin epithelial cells versus vaginal mucous membrane cells by direct contact.

In alleged sexual assault and rape cases, the focus has often been to collect samples from the victim's body, for detection of body fluids or skin cells from the offender. But in many cases intimate body samples from the perpetrator(s) can also be informative. However, in cases where the female victim claims vaginal penetration, the defendant may display an alternative explanation to the DNA findings, i.e. that the victim's skin cells has been secondarily transferred to his penis. We hypothesized that female DNA will be detected in a significantly greater amount on swabs from penis after intercourse than after secondary transfer by skin contact. Fourteen male-female couples were recruited to test the above hypothesis, by collecting penile swabs from 3 specified anatomical locations: Glans, shaft, and the coronal sulcus, after two different situations: Vaginal intercourse and secondary transfer of epithelial cells by skin contact. The results show that penile swabs following intercourse produce significantly higher DNA concentrations than after secondary transfer by skin contact. Our results, indicates which of the anatomical regions is best suited for sampling. The DNA profiling results show a preponderance of female profiles over male profiles following intercourse compared to secondary skin contact. Based on these data, it is possible to make a statistical model to distinguish between samples taken after intercourse and samples taken after secondary transfer by skin contact based on the amount of female DNA and mixture proportion (Mx) between female and male DNA in samples collected from penis swabs.

Contaminating viral sequences in high-throughput sequencing viromics: a linkage study of 700 sequencing libraries.

OBJECTIVES: Sample preparation for high-throughput sequencing (HTS) includes treatment with various laboratory components, potentially carrying viral nucleic acids, the extent of which has not been thoroughly investigated. Our aim was to systematically examine a diverse repertoire of laboratory components used to prepare samples for HTS in order to identify contaminating viral sequences. METHODS: A total of 322 samples of mainly human origin were analysed using eight protocols, applying a wide variety of laboratory components. Several samples (60% of human specimens) were processed using different protocols. In total, 712 sequencing libraries were investigated for viral sequence contamination. RESULTS: Among sequences showing similarity to viruses, 493 were significantly associated with the use of laboratory components. Each of these viral sequences had sporadic appearance, only being identified in a subset of the samples treated with the linked laboratory component, and some were not identified in the non-template control samples. Remarkably, more than 65% of all viral sequences identified were within viral clusters linked to the use of laboratory components. CONCLUSIONS: We show that high prevalence of contaminating viral sequences can be expected in HTS-based virome data and provide an extensive list of novel contaminating viral sequences that can be used for evaluation of viral findings in future virome and metagenome studies. Moreover, we show that detection can be problematic due to stochastic appearance and limited non-template controls. Although the exact origin of these viral sequences requires further research, our results support laboratory-component-linked viral sequence contamination of both biological and synthetic origin.

Optimal sunscreen use, during a sun holiday with a very high ultraviolet index, allows vitamin D synthesis without sunburn.

BACKGROUND: Sunlight contains ultraviolet (UV)A and UVB radiation. UVB is essential for vitamin D synthesis but is the main cause of sunburn and skin cancer. Sunscreen use is advocated to reduce the sun's adverse effects but may compromise vitamin D status. OBJECTIVES: To assess the ability of two intervention sunscreens to inhibit vitamin D synthesis during a week-long sun holiday. METHODS: The impact of sunscreens on vitamin D status was studied during a 1-week sun holiday in Tenerife (28° N). Comparisons were made between two formulations, each with a sun protection factor (SPF) of 15. The UVA-protection factor (PF) was low in one case and high in the other. Healthy Polish volunteers (n = 20 per group) were given the sunscreens and advised on the correct application. Comparisons were also made with discretionary sunscreen use (n = 22) and nonholiday groups (51·8° N, n = 17). Sunscreen use in the intervention groups was measured. Behaviour, UV radiation exposure, clothing cover and sunburn were monitored. Serum 25-hydroxyvitamin D3 [25(OH)D3 ] was assessed by high-performance liquid chromatography-tandem mass spectrometry. RESULTS: Use of intervention sunscreens was the same (P = 0·60), and both equally inhibited sunburn, which was present in the discretionary use group. There was an increase (P < 0·001) in mean ± SD 25(OH)D3 (28·0 ± 16·5 nmol L-1 ) in the discretionary use group. The high and low UVA-PF sunscreen groups showed statistically significant increases (P < 0·001) of 19·0 ± 14·2 and 13·0 ± 11·4 nmol L-1 25(OH)D3 , respectively with P = 0·022 for difference between the intervention sunscreens. The nonholiday group showed a fall (P = 0·08) of 2·5 ± 5·6 nmol L-1 25(OH)D3 . CONCLUSIONS: Sunscreens may be used to prevent sunburn yet allow vitamin D synthesis. A high UVA-PF sunscreen enables significantly higher vitamin D synthesis than a low UVA-PF sunscreen because the former, by default, transmits more UVB than the latter. What's already known about this topic? Action spectra (wavelength dependence) for erythema and the cutaneous formation of vitamin D overlap considerably in the ultraviolet (UV)B region. Theoretically, sunscreens that inhibit erythema should also inhibit vitamin D synthesis. To date, studies on the inhibitory effects of sunscreens on vitamin D synthesis have given conflicting results, possibly, in part, because people typically apply sunscreen suboptimally. Many studies have design flaws. What does this study add? Sunscreens (sun protection factor, SPF 15) applied at sufficient thickness to inhibit sunburn during a week-long holiday with a very high UV index still allow a highly significant improvement of serum 25-hydroxyvitamin D3 concentration. An SPF 15 formulation with high UVA protection enables better vitamin D synthesis than a low UVA protection product. The former allows more UVB transmission.

Children sustain high levels of skin DNA photodamage, with a modest increase of serum 25-hydroxyvitamin D3 , after a summer holiday in Northern Europe.

BACKGROUND: Childhood solar ultraviolet radiation (UVR) exposure increases the risk of skin cancer in adulthood, which is associated with mutations caused by UVR-induced cyclobutane pyrimidine dimers (CPD). Solar UVR is also the main source of vitamin D, essential for healthy bone development in children. OBJECTIVES: To assess the impact of a 12-day Baltic Sea (54° N) beach holiday on serum 25-hydroxyvitamin D3 [25(OH)D3 ] and CPD in 32 healthy Polish children (skin types I-IV). METHODS: Blood and urine were collected before and after the holiday and assessed for 25(OH)D3 and excreted CPD, respectively, and personal UVR exposure was measured. Diaries were used to record sunbathing, sunburn and sunscreen use. Before- and after-holiday skin redness and pigmentation were measured by reflectance spectroscopy. RESULTS: The average ± SD daily exposure UVR dose was 2·4 ± 1·5 standard erythema doses (SEDs), which is borderline erythemal. The mean concentration of 25(OH)D3 increased (× 1·24 ± 0·19) from 64·7 ± 13·3 to 79·3 ± 18·7 nmol L-1 (P < 0·001). Mean CPD increased 12·6 ± 10·0-fold from 26·9 ± 17·9 to 248·9 ± 113·4 fmol µmol-1 creatinine (P < 0·001). Increased 25(OH)D3 was accompanied by a very much greater increase in DNA damage associated with carcinogenic potential. Overall, skin type had no significant effects on behavioural, clinical or analytical outcomes, but skin types I/II had more CPD (unadjusted P = 0·0496) than skin types III/IV at the end of the holiday. CONCLUSIONS: Careful consideration must be given to the health outcomes of childhood solar exposure, and a much better understanding of the risk-benefit relationships of such exposure is required. Rigorous photoprotection is necessary for children, even in Northern Europe.

Therapeutic administration of bone marrow-derived mesenchymal stromal cells reduces airway inflammation without up-regulating Tregs in experimental asthma.

BACKGROUND: Prophylactic administration of mesenchymal stromal cells (MSCs) derived from adipose (AD-MSC) and bone marrow tissue (BM-MSC) in ovalbumin-induced asthma hinders inflammation in a Treg-dependent manner. It is uncertain whether MSCs act through Tregs when inflammation is already established in asthma induced by a clinically relevant allergen. OBJECTIVE: Evaluate the effect of therapeutic administration of MSCs on inflammation and Treg cells in house dust mite (HDM)-induced asthma. METHODS: BM-MSCs and AD-MSCs were administered intratracheally to C57BL/6 mice 1 day after the last HDM challenge. Lung function, remodelling and parenchymal inflammation were assayed 3 or 7 days after MSCs treatment, through invasive plethysmography and histology, respectively. Bronchoalveolar lavage fluid (BALF) and mediastinal lymph nodes (mLNs) were assessed regarding the inflammatory profile by flow cytometry, ELISA and qRT-PCR. MSCs were studied regarding their potential to induce Treg cells from primed and unprimed lymphocytes in vitro. RESULTS: BM-MSCs, but not AD-MSCs, reduced lung influx of eosinophils and B cells and increased IL-10 levels in HDM-challenged mice. Neither BM-MSCs nor AD-MSCs reduced lung parenchymal inflammation, airway hyperresponsiveness or mucus hypersecretion. BM-MSCs and AD-MSCs did not up-regulate Treg cell counts within the airways and mLNs, but BM-MSCs decreased the pro-inflammatory profile of alveolar macrophages. Co-culture of BM-MSCs and AD-MSCs with allergen-stimulated lymphocytes reduced Treg cell counts in a cell-to-cell contact-independent manner, although co-culture of both MSCs with unprimed lymphocytes up-regulated Treg cell counts. CONCLUSIONS: MSCs therapeutically administered exert anti-inflammatory effects in the airway of HDM-challenged mice, but do not ameliorate lung function or remodelling. Although MSC pre-treatment can increase Treg cell numbers, it is highly unlikely that the MSCs will induce Treg cell expansion when lymphocytes are allergenically primed in an established lung inflammation.

Return-to-work intervention during cancer treatment - The providers' experiences.

To explore in-depth understanding of providers' experiences when involved in a return-to-work (RTW) intervention offered during cancer treatment. Semi-structured individual interviews and participant observations at a hospital department and two municipal job centers were carried out, including ten providers (physicians, nurses and social workers). A phenomenological-hermeneutic approach was applied, involving coding, identification of themes and interpretation of findings. Three major themes were identified: Treatment first, Work as an integrated component in cancer rehabilitation, and Challenges in bringing up work issues. Differences in providers' experiences of the RTW intervention offered to cancer patients were found: in the hospital setting RTW was a second priority, whereas in the municipality job centers it was an integrated component. Further studies are needed to investigate how and when occupational rehabilitation services can be implemented across sectors to support cancer patients' RTW. In the future, work issues ought to be systematically presented by providers across sectors as early as possible to support cancer patients' RTW. Cancer patients' individual needs and thoughts about RTW are to be identified by both health care providers during treatment and social workers at the municipality level and shared across sectors.

Evaluation of three-dimensional SonoAVC ultrasound for antral follicle count in infertile women: its agreement with conventional two-dimensional ultrasound and serum levels of anti-Müllerian hormone.

BACKGROUND: Several studies have reported a correlation between antral follicle count by conventional 2D transvaginal sonography and serum anti-Müllerian hormone levels. However, few studies have investigated the effectiveness of 3D SonoAVC transvaginal ultrasound technology, particularly in infertile women. Therefore, this study aims to evaluate the usefulness of three-dimensional (3D) SonoAVC transvaginal ultrasound technology for antral follicle count and its correlation to conventional two-dimensional (2D) transvaginal ultrasound and serum levels of anti-Müllerian hormone in infertile women. METHODS: This cross-sectional study included 42 infertile women with age lower than 40 years that underwent treatment at a private fertility clinic between June and December 2015. Patient data included age, body mass index and cause of infertility. On cycle day 3 the following hormone levels were measured: serum levels of anti-Müllerian hormone, follicle-stimulating hormone, cancer antigen 125, prolactin, thyroid-stimulating hormone and oestradiol; the number of antral follicles was counted as well. The scanning were performed through 2D and 3D technology transvaginal ultrasound. RESULTS: Using a Bland-Altman test we demonstrated that both technologies are quite equivalent. However, antral follicle count is higher using 3D ultrasound technology compared to 2D technology (p < 0.001; Wilcoxon test), this finding is mainly remarkable in ovaries with more than 20 antral follicles. Moreover, the mean time required for manual 2D ultrasound and 3D SonoAVC measurements were 275 ± 109 and 103 ± 57 s, respectively (p < 0.001). Serum AMH concentration correlated to the total number of early antral follicles (correlation coefficients = 0.678 and 0.612; p < 0.001 by 2D ultrasound and 3D SonoAVC, respectively; Spearman's correlation test). CONCLUSIONS: Antral follicle count is better estimated using 3D ultrasound compared to 2D technology. A great advantage of 3D SonoAVC was less time required for an examination and the visual advantage when it need to count more than 20 follicles. TRIAL REGISTRATION: CAAE: 35141114.4.0000.5327 . Registered 10 June 2015.

Focused cardiac ultrasound is feasible in parturients; a prospective observational study.

BACKGROUND: In pregnancy, the major causes of morbidity and mortality include diseases that affect circulation profoundly and restoration of normal circulation is a key objective. Focused cardiac ultrasound provides information on the individual determinants of circulation and has a significant impact on patient management. This study aimed to determine the feasibility of focused cardiac ultrasound in term parturients. METHODS: Forty parturients underwent focused cardiac ultrasound. Parasternal long-axis, parasternal short-axis, 4-chamber and subcostal views were performed in the left-lateral position and the supine position by a novice operator. All images were graded on the following scale: 1 = no image; 2 = poor and insufficient image; 3 = sufficient image quality; 4 = good image quality; 5 = perfect image quality. A cut-off grade of three signified images sufficient for clinical decision-making. RESULTS: Images sufficient for clinical decision-making were obtained in all patients when including all imaging views. 37/40 (93 %) patients presented images of grade ≥ 4, whereas 10/40 (25%) patients had images of grade 5. Image grades were higher in the left lateral position (all P < 0.001). Parturients with a body mass index ≤ 30 kg/m2 did not present different image grades than parturients with a higher body mass index (P = 0.753). Subcostal views could not be obtained. CONCLUSION: Focused cardiac ultrasound is feasible in the obstetric population and images of sufficient quality for clinical decision-making were obtained in all parturients by an operator with limited experience.

Ultrasound-guidance outperforms the palpation technique for peripheral venous catheterisation in anaesthetised toddlers: a randomised study.

BACKGROUND: Traditional palpation technique for peripheral venous catheter (PVC) placement can be challenging. We aimed to compare the Dynamic Needle Tip Positioning (DNTP) ultrasound technique with the palpation technique for PVC placement in anaesthetised toddlers undergoing elective, low-risk procedures. METHODS: The study was a randomised, controlled, crossover study. Five operators applied both techniques for PVC on 50 children < 4 years. The primary endpoint was first attempt success rate. The secondary endpoints were: (1) overall success rate, (2) number of skin perforations, (3) number of needle redirections, (4) number of catheters used, (5) total time, (6) needle manipulation time, (7) distance to the nearest flexion crease, (8) anatomical region of catheterisation, (9) size of the catheters and (10) the effects of visibility, and palpability of the veins. RESULTS: The first attempt success rate and the overall success rate were significantly higher in the DNTP group (42/50 vs. 30/50, P = 0.029), (50/50 vs. 42/50, P = 0.008). Furthermore, a lower number of skin perforations (60 vs. 84, P = 0.013) and needle redirections (14 vs. 131, P < 0.001) and increased distance to the nearest flexion crease (P < 0.001) were seen in the DNTP group. There were no significant differences in number of catheters used (60 vs. 75 P = 0.050) or total time (P = 0.073), however, the needle manipulation time was longer in the DNTP group (P = 0.011). The success rate decreased with less visible and palpable veins in the palpation group (p = 0.006). CONCLUSION: Ultrasound-guidance outperforms the traditional palpation technique for peripheral venous catheterisation in anaesthetised toddlers undergoing elective, low-risk procedures.

The tyrosine kinase inhibitor dasatinib reduces lung inflammation and remodelling in experimental allergic asthma.

BACKGROUND AND PURPOSE: Asthma is characterized by chronic lung inflammation and airway hyperresponsiveness. Despite recent advances in understanding of its pathophysiology, asthma remains a major public health problem, and new therapeutic strategies are urgently needed. In this context, we sought to ascertain whether treatment with the TK inhibitor dasatinib might repair inflammatory and remodelling processes, thus improving lung function, in a murine model of asthma. EXPERIMENTAL APPROACH: Animals were sensitized and subsequently challenged, with ovalbumin (OVA) or saline. Twenty-four hours after the last challenge, animals were treated with dasatinib, dexamethasone, or saline, every 12 h for 7 consecutive days. Twenty-four hours after the last treatment, the animals were killed, and data were collected. Lung structure and remodelling were evaluated by morphometric analysis, immunohistochemistry, and transmission electron microscopy of lung sections. Inflammation was assessed by cytometric analysis and ELISA, and lung function was evaluated by invasive whole-body plethysmography. KEY RESULTS: In OVA mice, dasatinib, and dexamethasone led to significant reductions in airway hyperresponsiveness. Dasatinib was also able to attenuate alveolar collapse, contraction index, and collagen fibre deposition, as well as increasing elastic fibre content, in OVA mice. Concerning the inflammatory process, dasatinib reduced inflammatory cell influx to the airway and lung-draining mediastinal lymph nodes, without inducing the thymic atrophy promoted by dexamethasone. CONCLUSIONS AND IMPLICATIONS: In this model of allergic asthma, dasatinib effectively blunted the inflammatory and remodelling processes in asthmatic lungs, enhancing airway repair and thus improving lung mechanics.

Asphyxia causes ultrasonographic D-shaping of the left ventricle--an experimental porcine study.

INTRODUCTION: In critical care, early diagnosis and correct treatment are of the utmost importance. Focused ultrasonography has gained acceptance as a pivotal tool for this by elucidating the underlying pathology. For example, massive pulmonary embolism is characterised by right ventricular dilatation. However, theoretically these characteristics might also be generated by asphyxia and the consequent hypoxia. We aimed to evaluate the ultrasonographic characteristics of asphyxia in a porcine model. METHODS: Nineteen (13 intervention, 6 control) piglets were subjected to asphyxia until cardiac arrest, by disconnecting the ventilator tube. Ultrasonographic short-axis cine loops of the left ventricle were obtained every 30 seconds. The left ventricular (LV) eccentricity index (index of LV D-shaping) was quantified along with LV end-diastolic/end-systolic areas. Invasive pressures were measured throughout. RESULTS: The LV eccentricity index increased from 1.14 (1.10-1.31) to 1.86 (1.48-2.38), (P = 0.002), after 1.5 min, receded thereafter to baseline levels followed by a second increase after 5.5 min. LV end-diastolic area decreased from 11.6 cm(2) (11.1-13.2) to 6.3 cm(2) (3.3 -11.0) after 2.0 min (P = 0.009). Subsequently, values returned to the baseline level. DISCUSSION: The early and transient acute dilatation of the RV, coinciding with D-shaping of the LV and decrease in LV end-diastolic area seen in our study represent a combination of ultrasonographic characteristics normally attributed to pulmonary embolism. Early changes in ventricular chamber sizes and shape with septal flattening related to asphyxia can occur, but appear to be transient and disappear as circulatory collapse progresses, in an animal model. Despite this, asphyxia may represent a cause of ultrasonographic misinterpretation.

Teenagers and young adults with cancer in Europe: from national programmes to a European integrated coordinated project.

Over 14 000 patients aged 15-24 are estimated to be diagnosed with cancer in the European Union (EU) each year. Teenagers and young adults (TYA) often fall down gaps between children's and adults cancer services. The specific challenges of providing optimal care to them are described, but we present a summary of recent progress. Progress to overcome these challenges is happening at different rates across Europe. We summarise the European national projects in this field but more recently we have seen the beginnings of European coordination. Within the EU 7th Funding Programme (FP7) European Network for Cancer Research in Children and Adolescents programme (ENCCA), a specific European Network for Teenagers and Young Adults with Cancer has held a series of scientific meetings, including professionals, patients and caregivers. This group has proposed unanswered research questions and agreed key features of a high-quality service that can improve outcomes for TYA with cancer, including the primacy of collaboration between adult and paediatric services to eliminate the gap in the management of TYA with cancer.

Forensic ancestry analysis with two capillary electrophoresis ancestry informative marker (AIM) panels: Results of a collaborative EDNAP exercise.

There is increasing interest in forensic ancestry tests, which are part of a growing number of DNA analyses that can enhance routine profiling by obtaining additional genetic information about unidentified DNA donors. Nearly all ancestry tests use single nucleotide polymorphisms (SNPs), but these currently rely on SNaPshot single base extension chemistry that can fail to detect mixed DNA. Insertion-deletion polymorphism (Indel) tests have been developed using dye-labeled primers that allow direct capillary electrophoresis detection of PCR products (PCR-to-CE). PCR-to-CE maintains the direct relationship between input DNA and signal strength as each marker is detected with a single dye, so mixed DNA is more reliably detected. We report the results of a collaborative inter-laboratory exercise of 19 participants (15 from the EDNAP European DNA Profiling group) that assessed a 34-plex SNP test using SNaPshot and a 46-plex Indel test using PCR-to-CE. Laboratories were asked to type five samples with different ancestries and detect an additional mixed DNA sample. Statistical inference of ancestry was made by participants using the Snipper online Bayes analysis portal plus an optional PCA module that analyzes the genotype data alongside calculation of Bayes likelihood ratios. Exercise results indicated consistent genotyping performance from both tests, reaching a particularly high level of reliability for the Indel test. SNP genotyping gave 93.5% concordance (compared to the organizing laboratory's data) that rose to 97.3% excluding one laboratory with a large number of miscalled genotypes. Indel genotyping gave a higher concordance rate of 99.8% and a reduced no-call rate compared to SNP analysis. All participants detected the mixture from their Indel peak height data and successfully assigned the correct ancestry to the other samples using Snipper, with the exception of one laboratory with SNP miscalls that incorrectly assigned ancestry of two samples and did not obtain informative likelihood ratios for a third. Therefore, successful ancestry assignments were achieved by participants in 92 of 95 Snipper analyses. This exercise demonstrates that ancestry inference tests based on binary marker sets can be readily adopted by laboratories that already have well-established CE regimes in place. The Indel test proved to be easy to use and allowed all exercise participants to detect the DNA mixture as well as achieving complete and concordant profiles in nearly all cases. Lastly, two participants successfully ran parallel next-generation sequencing analyses (each using different systems) and achieved high levels of genotyping concordance using the exercise PCR primer mixes unmodified.

Visualization and prediction of porosity in roller compacted ribbons with near-infrared chemical imaging (NIR-CI).

The porosity of roller compacted ribbon is recognized as an important critical quality attribute which has a huge impact on the final product quality. The purpose of this study was to investigate the use of near-infrared chemical imaging (NIR-CI) for porosity estimation of ribbons produced at different roll pressures. Two off-line methods were utilized as reference methods. The relatively fast method (oil absorption) was comparable with the more time-consuming mercury intrusion method (R(2)=0.98). Therefore, the oil method was selected as the reference off line method. It was confirmed by both reference methods that ribbons compressed at a higher pressure resulted in a lower mean porosity. Using NIR-CI in combination with multivariate data analysis it was possible to visualize and predict the porosity distribution of the ribbons. This approach is considered important for process monitoring and control of continuously operating roller compaction line.

RNA/DNA co-analysis from human skin and contact traces--results of a sixth collaborative EDNAP exercise.

The European DNA profiling group (EDNAP) organized a sixth collaborative exercise on RNA/DNA co-analysis for body fluid/tissue identification and STR profiling. The task was to identify skin samples/contact traces using specific RNA biomarkers and test three housekeeping genes for their suitability as reference genes. Eight stains, a skin RNA dilution series and, optionally, bona fide or mock casework samples of human or non-human origin were analyzed by 22 participating laboratories using RNA extraction or RNA/DNA co-extraction methods. Two sets of previously described skin-specific markers were used: skin1 pentaplex (LCE1C, LCE1D, LCE2D, IL1F7 and CCL27) and skin2 triplex (LOR, KRT9 and CDSN) in conjunction with a housekeeping gene, HKG, triplex (B2M, UBC and UCE). The laboratories used different chemistries and instrumentation. All laboratories were able to successfully isolate and detect mRNA in contact traces (e.g., human skin, palm-, hand- and fingerprints, clothing, car interiors, computer accessories and electronic devices). The simultaneous extraction of RNA and DNA provides an opportunity for positive identification of the tissue source of origin by mRNA profiling as well as a simultaneous identification of the body fluid donor by STR profiling. The skin markers LCE1C and LOR and the housekeeping gene marker B2M were detected in the majority of contact traces. Detection of the other markers was inconsistent, possibly due to the low amounts and/or poor quality of the genetic material present in shed skin cells. The results of this and the previous collaborative RNA exercises support RNA profiling as a reliable body fluid/tissue identification method that can easily be combined with current STR typing technology.

Effects of levosimendan in patients with left ventricular hypertrophy undergoing aortic valve replacement.

BACKGROUND: Left ventricular hypertrophy is associated with adverse outcomes, including death, during cardiac surgery. This may be facilitated by an increased oxygen demand and diastolic dysfunction. Levosimendan augments haemodynamics without further oxygen consumption and improves echocardiographic indices of diastolic dysfunction. This study aimed to describe the haemodynamic effects of short-term pre- and intra-operative levosimendan infusion including advanced echocardiographic measures of diastolic and systolic heart function. METHODS: The study was randomised, double-blinded and placebo-controlled performed at a single-centre university hospital. Patients with left ventricular hypertrophy and ejection fraction > 45% scheduled for single procedure aortic valve replacement were included and randomised to infusion of either levosimendan 0.1 µg/kg/min or placebo from 4 h before anaesthesia to the end of surgery. Outcome measures were echocardiographic indices of left ventricular diastolic function: E/e' (primary endpoint), e', e'/a' and indices of systolic function: longitudinal strain, ejection fraction and s'. Patients were followed until 6 months after surgery. In addition, invasive haemodynamic measures were obtained perioperatively. RESULTS: The trial was prematurely terminated due to an overall high incidence of post-operative atrial fibrillation (15/20, P = 0.002) after inclusion of 20 patients. The relative decrease in perioperative cardiac index was lower (P = 0.016) in the levosimendan group. There was no difference in E/e', and similar results were found for all measures of systolic function. CONCLUSION: Short-term levosimendan caused a transient relative increase in cardiac index, but no effect was seen on the first post-operative day and up to 6 months post-operatively with indices of systolic and diastolic heart function.

Glucocorticoids decrease Treg cell numbers in lungs of allergic mice.

Glucocorticoids have been the hallmark anti-inflammatory drug used to treat asthma. It has been shown that glucocorticoids ameliorate asthma by increasing numbers and activity of Tregs, in contrast recent data show that glucocorticoid might have an opposite effect on Treg cells from normal mice. Since Tregs are target cells that act on the resolution of asthma, the aim of this study was to elucidate the effect of glucocorticoid treatment on lung Tregs in mouse models of asthma. Allergen challenged mice were treated with either oral dexamethasone or nebulized budesonide. Broncoalveolar lavage and airway hyperresponsiveness were evaluated after allergenic challenge. Lung, thymic and lymph node cells were phenotyped on Treg through flow cytometry. Lung cytokine secretion was detected by ELISA. Although dexamethasone inhibited airway inflammation and hyperresponsiveness, improving resolution, we have found that both dexamethasone and budesonide induce a reduction of Treg numbers on lungs and lymphoid organs of allergen challenged mice. The reduction of lung Treg levels was independent of mice strain or type of allergen challenge. Our study also indicates that both glucocorticoids do not increase Treg activity through production of IL-10. Glucocorticoid systemic or localized treatment induced thymic atrophy. Taken together, our results demonstrate that glucocorticoids decrease Treg numbers and activity in different asthma mouse models, probably by reducing thymic production of T cells. Therefore, it is possible that glucocorticoids do not have beneficial effects on lung populations of Treg cells from asthmatic patients.

Winning or losing does matter: Acute cardiac admissions in New Zealand during Rugby World Cup tournaments.

AIM: The relationship between watching major sporting events and cardiac hospital admissions is contentious. This study is the first to investigate cardiovascular admissions during Rugby World Cup (RWC) tournaments. METHODS AND RESULTS: New Zealand (NZ) public hospital admissions data were analysed for cardiovascular events during the 1999, 2003, 2007 and 2011 RWC tournaments. The exposure period was the day of the NZ All Blacks last match and 2 days post-match. The control period was equivalent days in October or November for 3 years prior to or after each RWC. The NZ 2011 semi-final win and 2000 Olympics opening ceremony were also analysed. There were 281 ± 14 and 3313 ± 379 cardiac admissions in the exposure and control periods. The semi-final loss in 2003 was associated with a 50% (p < 0.01) increase in pooled heart failure admissions and a 20% (p < 0.05) increase in pooled acute coronary syndromes admissions. Increases in heart failure were specific to women with a two-fold increase on match day and 2-days post (p < 0.01). There was no increase in male heart failure admissions but arrhythmias increased 2.6 times (p < 0.01) 1-day after losing the 2003 semi-final. In contrast, admissions were typically lower after the 2011 semi-final win and Olympics opening ceremony. CONCLUSION: This is the first study to find a relationship between hospital admissions for heart failure in women and a major sporting event. Preventive health measures should be considered in fans with cardiovascular disease or at high risk of cardiovascular events during sporting events. Winning or losing does matter.