The physiological and biochemical basis of potency thresholds modeled using human estrogen receptor alpha: implications for identifying endocrine disruptors.

The endocrine system functions by interactions between ligands and receptors. Ligands exhibit potency for binding to and interacting with receptors. Potency is the product of affinity and efficacy. Potency and physiological concentration determine the ability of a ligand to produce physiological effects. The kinetic behavior of ligand-receptor interactions conforms to the laws of mass action. The laws of mass action define the relationship between the affinity of a ligand and the fraction of cognate receptors that it occupies at any physiological concentration. We previously identified the minimum ligand potency required to produce clinically observable estrogenic agonist effects via the human estrogen receptor-alpha (ERα). By examining data on botanical estrogens and dietary supplements, we demonstrated that ERα ligands with potency lower than one one-thousandth that of the primary endogenous hormone 17ß-estradiol (E2) do not produce clinically observable estrogenic effects. This allowed us to propose a Human-Relevant Potency Threshold (HRPT) for ERα ligands of 1 × 10-4 relative to E2. Here, we test the hypothesis that the HRPT for ERα arises from the receptor occupancy by the normal metabolic milieu of endogenous ERα ligands. The metabolic milieu comprises precursors to hormones, metabolites of hormones, and other normal products of metabolism. We have calculated fractional receptor occupancies for ERα ligands with potencies below and above the previously established HRPT when normal circulating levels of some endogenous ERα ligands and E2 were also present. Fractional receptor occupancy calculations showed that individual ERα ligands with potencies more than tenfold higher than the HRPT can compete for occupancy at ERα against individual components of the endogenous metabolic milieu and against mixtures of those components at concentrations found naturally in human blood. Ligands with potencies less than tenfold higher than the HRPT were unable to compete successfully for ERα. These results show that the HRPT for ERα agonism (10-4 relative to E2) proposed previously is quite conservative and should be considered strong evidence against the potential for disruption of the estrogenic pathway. For chemicals with potency 10-3 of E2, the potential for estrogenic endocrine disruption must be considered equivocal and subject to the presence of corroborative evidence. Most importantly, this work demonstrates that the endogenous metabolic milieu is responsible for the observed ERα agonist HRPT, that this HRPT applies also to ERα antagonists, and it provides a compelling mechanistic explanation for the HRPT that is grounded in basic principles of molecular kinetics using well characterized properties and concentrations of endogenous components of normal metabolism.

The quiet crossing of ocean tipping points.

Anthropogenic climate change profoundly alters the ocean's environmental conditions, which, in turn, impact marine ecosystems. Some of these changes are happening fast and may be difficult to reverse. The identification and monitoring of such changes, which also includes tipping points, is an ongoing and emerging research effort. Prevention of negative impacts requires mitigation efforts based on feasible research-based pathways. Climate-induced tipping points are traditionally associated with singular catastrophic events (relative to natural variations) of dramatic negative impact. High-probability high-impact ocean tipping points due to warming, ocean acidification, and deoxygenation may be more fragmented both regionally and in time but add up to global dimensions. These tipping points in combination with gradual changes need to be addressed as seriously as singular catastrophic events in order to prevent the cumulative and often compounding negative societal and Earth system impacts.

Comparable Outcomes Between Autologous Chondrocyte Implantation and Osteochondral Allograft Transplantation in the Setting of Patellar Realignment.

PURPOSE: To determine clinical and functional outcomes in patients treated with autologous chondrocyte implantation (ACI) or osteochondral allograft (OCA) transplantation for chondral defects secondary to patellar instability with concomitant medial patellofemoral ligament (MPFL) reconstruction and tibial tubercle osteotomy (TTO) for patellar realignment. METHODS: A retrospective review identified patients who underwent ACI or OCA transplantation with concomitant MPFL reconstruction and TTO. Patients were excluded if they did not have concomitant MPFL reconstruction and TTO, had the presence of other intra-articular pathologies, or failed to complete postoperative subjective outcome evaluations at a minimum of 2 years following surgery. Subjective outcome measures included the Knee injury and Osteoarthritis Outcome Score for Joint Replacement, International Knee Documentation Committee evaluation, and 12-item Short Form Health Survey physical scores, collected a minimum of 2 years after surgery. Defect location, size, complications, and rate of subsequent surgery were determined. RESULTS: Eighteen total patients were included in this study. The ACI cohort included 11 patients with 13 total defects that were treated with ACI. The OCA cohort included 7 patients with 10 total defects that were treated with OCA. This was due to a number of patients in either group having multiple cartilage defects. Twenty-three total chondral defects were compared to analyze clinical and functional outcomes following surgical correction (ACI: n = 13, OCA: n = 10). Five defects were noted on the femoral condyle and 18 on the patellar facets/central ridge. Defects were comparable between groups, including size measured during index arthroscopy (ACI = 3.34 cm2 [95% CI, 2.3-4.4 cm2] vs OCA = 4.03 cm2 [95% CI, 3.1-5.0 cm2]; P = .351), Outerbridge classification (ACI = 54.8% grade 4 vs OCA = 60.0% grade 4; P ≥ .999), and Area Measurement and Depth Underlying Structures score (ACI = 47.1 vs OCA = 58.6; P = .298). Postoperative outcomes were comparable, including revision rate (ACI = 15.4% vs OCA = 10.0%; P ≥ .999) and 2-year International Knee Documentation Committee scores (ACI = 74.2 [95% CI, 65.2-83.2] vs OCA = 51.2 [95% CI, 30.3-72.1]; P = .077). ACI did have significantly higher 2-year Knee injury and Osteoarthritis Outcome Score for Joint Replacement (85.1 [95% CI, 76.9-93.3] vs 63.7 [95% CI, 49.1-78.3]; P = .031) and 12-item Short Form Health Survey scores (54.1 [95% CI, 52.0-56.2] vs 42.6 [95% CI, 35.8-49.4]; P = .007) compared to OCA. CONCLUSIONS: ACI or OCA transplantation for chondral defects with concomitant MPFL reconstruction and TTO can be safely performed in an outpatient setting with functional and clinical outcomes being comparable. LEVEL OF EVIDENCE: Level III, retrospective case series study.

Blood transcriptomic signature in type-2 biomarker-low severe asthma and asthma control.

BACKGROUND: Patients with type-2 (T2) cytokine-low severe asthma often have persistent symptoms despite suppression of T2 inflammation with corticosteroids. OBJECTIVES: We sought to analyze whole blood transcriptome from 738 samples in T2-biomarker-high/-low patients with severe asthma to relate transcriptomic signatures to T2 biomarkers and asthma symptom scores. METHODS: Bulk RNA-seq data were generated for blood samples (baseline, week 24, week 48) from 301 participants recruited to a randomized clinical trial of corticosteroid optimization in severe asthma. Unsupervised clustering, differential gene expression analysis, and pathway analysis were performed. Patients were grouped by T2-biomarker status and symptoms. Associations between clinical characteristics and differentially expressed genes (DEGs) associated with biomarker and symptom levels were investigated. RESULTS: Unsupervised clustering identified 2 clusters; cluster 2 patients were blood eosinophil-low/symptom-high and more likely to be receiving oral corticosteroids (OCSs). Differential gene expression analysis of these clusters, with and without stratification for OCSs, identified 2960 and 4162 DEGs, respectively. Six hundred twenty-seven of 2960 genes remained after adjusting for OCSs by subtracting OCS signature genes. Pathway analysis identified dolichyl-diphosphooligosaccharide biosynthesis and assembly of RNA polymerase I complex as significantly enriched pathways. No stable DEGs were associated with high symptoms in T2-biomarker-low patients, but numerous associated with elevated T2 biomarkers, including 15 that were upregulated at all time points irrespective of symptom level. CONCLUSIONS: OCSs have a considerable effect on whole blood transcriptome. Differential gene expression analysis demonstrates a clear T2-biomarker transcriptomic signature, but no signature was found in association with T2-biomarker-low patients, including those with a high symptom burden.

Contrasting principal stratum and hypothetical strategy estimands in multi-period crossover trials with incomplete data.

Complete case analyses of complete crossover designs provide an opportunity to make comparisons based on patients who can tolerate all treatments. It is argued that this provides a means of estimating a principal stratum strategy estimand, something which is difficult to do in parallel group trials. While some trial users will consider this a relevant aim, others may be interested in hypothetical strategy estimands, that is, the effect that would be found if all patients completed the trial. Whether these estimands differ importantly is a question of interest to the different users of the trial results. This paper derives the difference between principal stratum strategy and hypothetical strategy estimands, where the former is estimated by a complete-case analysis of the crossover design, and a model for the dropout process is assumed. Complete crossover designs, that is, those where all treatments appear in all sequences, and which compare t treatments over p periods with respect to a continuous outcome are considered. Numerical results are presented for Williams designs with four and six periods. Results from a trial of obstructive sleep apnoea-hypopnoea (TOMADO) are also used for illustration. The results demonstrate that the percentage difference between the estimands is modest, exceeding 5% only when the trial has been severely affected by dropouts or if the within-subject correlation is low.

Pre-conditioning of gingival epithelial cells with sub-apoptotic concentrations of curcumin prevents pro-inflammatory cytokine release.

BACKGROUND AND OBJECTIVE: Plaque-induced gingival inflammation (gingivitis) is ubiquitous in humans. The epithelial barrier reacts to the presence of oral bacteria and induces inflammatory cascades. The objective of this study was to investigate the mechanism by which the small molecule micronutrient curcumin could decrease inflammatory response in vitro to oral bacterium heat-killed Fusobacterium nucleatum as curcumin could be a useful compound for combatting gingivitis already consumed by humans. METHODS: H400 oral epithelial cell line was pre-conditioned with curcumin and the production of cytokines was measured by enzyme-linked immunosorbent assay (ELISA) and translocation of transcription factors was used to monitor inflammatory responses. Haem oxygenase (HO-1) expression and molecules that HO-1 releases were evaluated for their potential to reduce the quantity of cytokine production. Immunofluorescence microscopy and Western blotting were used to evaluate changes in transcription factor and enzyme location. RESULTS: Pre-conditioning of H400 cells with a sub-apoptotic concentration of curcumin (20 µM) attenuated secretion of Granulocyte-Macrophage - Colony-Stimulating Factor (GM-CSF) and reduced NFkB nuclear translocation. This pre-conditioning caused an increase in nuclear Nrf2; an initial drop (at 8 h) followed by an adaptive increase (at 24 h) in glutathione; and an increase in haem oxygenase (HO-1) expression. Inhibition of HO-1 by SnPPIX prevented the curcumin-induced attenuation of GM-CSF production. HO-1 catalyses the breakdown of haem to carbon monoxide, free iron and biliverdin: the HO-1/CO anti-inflammatory pathway. Elevations in carbon monoxide, achieved using carbon monoxide releasing molecule-2 (CORM2) treatment alone abrogated F. nucleatum-induced cytokine production. Biliverdin is converted to bilirubin by biliverdin reductase (BVR). This pleiotropic protein was found to increase in cell membrane expression upon curcumin treatment. CONCLUSION: Curcumin decreased inflammatory cytokine production induced by Fusobacterium nucleatum in H400 oral epithelial cells. The mechanism of action appears to be driven by the increase of haem oxygenase and the production of carbon monoxide.

The Infant KIdney Dialysis and Utrafiltration (I-KID) Study: A Stepped-Wedge Cluster-Randomized Study in Infants, Comparing Peritoneal Dialysis, Continuous Venovenous Hemofiltration, and Newcastle Infant Dialysis Ultrafiltration System, a Novel Infant Hemodialysis Device.

OBJECTIVES: Renal replacement therapy (RRT) options are limited for small babies because of lack of available technology. We investigated the precision of ultrafiltration, biochemical clearances, clinical efficacy, outcomes, and safety profile for a novel non-Conformité Européenne-marked hemodialysis device for babies under 8 kg, the Newcastle Infant Dialysis Ultrafiltration System (NIDUS), compared with the current options of peritoneal dialysis (PD) or continuous venovenous hemofiltration (CVVH). DESIGN: Nonblinded cluster-randomized cross-sectional stepped-wedge design with four periods, three sequences, and two clusters per sequence. SETTING: Clusters were six U.K. PICUs. PATIENTS: Babies less than 8 kg requiring RRT for fluid overload or biochemical disturbance. INTERVENTIONS: In controls, RRT was delivered by PD or CVVH, and in interventions, NIDUS was used. The primary outcome was precision of ultrafiltration compared with prescription; secondary outcomes included biochemical clearances. MEASUREMENTS AND MAIN RESULTS: At closure, 97 participants were recruited from the six PICUs (62 control and 35 intervention). The primary outcome, obtained from 62 control and 21 intervention patients, showed that ultrafiltration with NIDUS was closer to that prescribed than with control: sd controls, 18.75, intervention, 2.95 (mL/hr); adjusted ratio, 0.13; 95% CI, 0.03-0.71; p = 0.018. Creatinine clearance was smallest and least variable for PD (mean, sd ) = (0.08, 0.03) mL/min/kg, larger for NIDUS (0.46, 0.30), and largest for CVVH (1.20, 0.72). Adverse events were reported in all groups. In this critically ill population with multiple organ failure, mortality was lowest for PD and highest for CVVH, with NIDUS in between. CONCLUSIONS: NIDUS delivers accurate, controllable fluid removal and adequate clearances, indicating that it has important potential alongside other modalities for infant RRT.

Exacerbation Profile and Risk Factors in a Type-2-Low Enriched Severe Asthma Cohort: A Clinical Trial to Assess Asthma Exacerbation Phenotypes.

Rationale: The past 25 years have seen huge progress in understanding of the pathobiology of type-2 (T2) asthma, identification of measurable biomarkers, and the emergence of novel monoclonal antibody treatments. Although present in a minority of patients with severe asthma, very little is known about the mechanisms underlying T2-low asthma, making it a significant unmet need in asthma research. Objectives: The objective of this study was to explore the differences between study exacerbators and nonexacerbators, to describe physiological changes at exacerbation in those who are T2HIGH and T2LOW at the time of exacerbation, and to evaluate the stability of inflammatory phenotypes when stable and at exacerbation. Methods: Exacerbation assessment was a prespecified secondary analysis of data from a 48-week, multicenter, randomized controlled clinical study comparing the use of biomarkers and symptoms to adjust steroid treatment in a T2-low severe asthma-enriched cohort. Participants were phenotyped as T2LOW (fractional exhaled nitric oxide ⩽ 20 ppb and blood eosinophil count ⩽ 150 cells/µl) or T2HIGH (fractional exhaled nitric oxide > 20 or blood eosinophil count > 150) at study enrollment and at each exacerbation. Here, we report the findings of the exacerbation analyses, including comparison of exacerbators and nonexacerbators, the physiological changes at exacerbation in those who had evidence of T2 biology at exacerbation versus those that did not, and the stability of inflammatory phenotypes when stable and at exacerbation. Measurements and Main Results: Of the 301 participants, 60.8% (183) had one or more self-reported exacerbations (total of 390). Exacerbators were more likely to be female, have a higher body mass index, and have more exacerbations requiring oral corticosteroid and unscheduled primary care attendances for exacerbations. At enrollment, 23.6% (71) were T2LOW and 76.4% (230) T2HIGH. The T2LOW group had more asthma primary care attendances, were more likely to have a previous admission to HDU (high dependency unit)/ICU and to be receiving maintenance oral corticosteroids. At exacerbation, the T2LOW events were indistinguishable from T2HIGH exacerbations in terms of lung function (mean fall in T2LOW FEV1, 200 [400] ml vs. T2HIGH 200 [300] ml; P = 0.93) and symptom increase (ACQ5: T2LOW, 1.4 [0.8] vs. T2HIGH, 1.3 [0.8]; P = 0.72), with no increase in T2 biomarkers from stable to exacerbation state in the T2LOW exacerbations. The inflammatory phenotype within individual patients was dynamic; inflammatory phenotype at study entry did not have a significant association with exacerbation phenotype. Conclusions: Asthma exacerbations demonstrating a T2LOW phenotype were physiologically and symptomatically similar to T2HIGH exacerbations. T2LOW asthma was an unstable phenotype, suggesting that exacerbation phenotyping should occur at the time of exacerbation. The clinically significant exacerbations in participants without evidence of T2 biology at the time of exacerbation highlight the unmet and pressing need to further understand the mechanisms at play in non-T2 asthma. Clinical trial registered with www.clinicaltrials.gov (NCT02717689).

Oxidized Zirconium Versus Cobalt Chromium for Primary TKA: No Difference in Midterm Revision Rates From the American Joint Replacement Registry.

BACKGROUND: Cobalt chromium (CoCr) is the most commonly used material in TKA; however, the use of oxidized zirconium (OxZr) implants has increased. The advantages to this material demonstrated in basic science studies have not been borne out in clinical studies to date. QUESTION/PURPOSE: In the setting of the American Joint Replacement Registry (AJRR), how do revision rates differ between CoCr and OxZr after primary TKA? METHODS: The AJRR was accessed for all primary TKAs performed between 2012 and 2020 for osteoarthritis, resulting in 441,605 procedures (68,506 with OxZr and 373,099 with CoCr). The AJRR is the largest joint replacement registry worldwide and collects procedure-specific details, making it ideal for large-scale comparisons of implant materials in the United States. Competing risk survival analyses were used to evaluate the all-cause revision rates of primary TKAs, comparing CoCr and OxZr implants. Data from the Centers for Medicare and Medicaid Services claims from 2012 to 2017 were also cross-referenced to capture additional revisions from other institutions. Revision rates were tabulated and subclassified by indication. Multivariate Cox regression was used to account for confounding variables such as age, gender, region, and hospital size. RESULTS: After controlling for confounding variables, there were no differences between the OxZr and CoCr groups in terms of the rate of all-cause revision at a mean follow-up of 46 ± 23 months and 44 ± 24 months for CoCr and OxZr implants, respectively (hazard ratio 1.055 [95% confidence interval 0.979 to 1.137]; p = 0.16) The univariate analysis demonstrated increased rates of revisions for pain and instability in the OxZr group (p = 0.003 and p < 0.001, respectively). CONCLUSION: These findings suggest there is no difference in all-cause revision between OxZr and CoCr implants in the short-term to mid-term. However, further long-term in vivo studies are needed to monitor the safety and all-cause revision rate of OxZr implants compared with those of CoCr implants. OxZr implants may be favorable in patients who have sensitivity to metal. Despite similar short-term to mid-term all-cause revision rates to CoCr implants, because of the limitations of this study, definitive recommendations for or against the use of OxZr cannot be made. LEVEL OF EVIDENCE: Level III, therapeutic study.

Factors affecting adherence with treatment advice in a clinical trial of patients with severe asthma.

BACKGROUND: Understanding why patients with severe asthma do not follow healthcare provider (HCP) advice to adjust treatment is critical to achieving personalised disease management. METHODS: We reviewed patient choice to follow HCP advice to adjust asthma treatment in a UK-based randomised, controlled, single-blind (study participant), multicentre, parallel group 48-week clinical study comparing biomarker-directed treatment adjustment with standard care in severe asthma. RESULTS: Of 1572 treatment advisories (291 participants), instructions were followed in 1377 cases (87.6%). Patients were more likely to follow advice to remain on treatment (96.7%) than to either reduce (70.3%) or increase (67.1%) their treatment, with 64% of patients following all treatment advice. Multivariate analysis associated belonging to an ethnic minority group (OR 3.10, 95% CI 1.68-5.73) and prior study medication changes (two or more changes: OR 2.77, 95% CI 1.51-5.10) with failure to follow treatment advice. In contrast, emergency room attendance in the prior year (OR 0.54, 95% CI 0.32-0.92) was associated with following treatment advice. The largest effect was seen with transition onto or off oral corticosteroids (OR 29.28, 95% CI 16.07-53.36) when compared with those requested to maintain treatment. Centre was also an important determinant regarding the likelihood of patients to follow treatment advice. CONCLUSIONS: Belonging to an ethnic minority group and multiple prior treatment adjustments were associated with not following HCP treatment advice. Patients also responded differently to HCP advice across UK specialist centres. These findings have implications for the generalisability of models of care in severe asthma and require further focused studies.

Relationship between inflammatory status and microbial composition in severe asthma and during exacerbation.

BACKGROUND: In T2-mediated severe asthma, biologic therapies, such as mepolizumab, are increasingly used to control disease. Current biomarkers can indicate adequate suppression of T2 inflammation, but it is unclear whether they provide information about airway microbial composition. We investigated the relationships between current T2 biomarkers and microbial profiles, characteristics associated with a ProteobacteriaHIGH microbial profile and the effects of mepolizumab on airway ecology. METHODS: Microbiota sequencing was performed on sputum samples obtained at stable and exacerbation state from 140 subjects with severe asthma participating in two clinical trials. Inflammatory subgroups were compared on the basis of biomarkers, including FeNO and sputum and blood eosinophils. ProteobacteriaHIGH subjects were identified by Proteobacteria to Firmicutes ratio ≥0.485. Where paired sputum from stable visits was available, we compared microbial composition at baseline and following ≥12 weeks of mepolizumab. RESULTS: Microbial composition was not related to inflammatory subgroup based on sputum or blood eosinophils. FeNO ≥50 ppb when stable and at exacerbation indicated a group with less dispersed microbial profiles characterised by high alpha-diversity and low Proteobacteria. ProteobacteriaHIGH subjects were neutrophilic and had a longer time from asthma diagnosis than ProteobacteriaLOW subjects. In those studied, mepolizumab did not alter airway bacterial load or lead to increased Proteobacteria. CONCLUSION: High FeNO could indicate a subgroup of severe asthma less likely to benefit from antimicrobial strategies at exacerbation or in the context of poor control. Where FeNO is <50 ppb, biomarkers of microbial composition are required to identify those likely to respond to microbiome-directed strategies. We found no evidence that mepolizumab alters airway microbial composition.

Radiomics and radiogenomics in gliomas: a contemporary update.

The natural history and treatment landscape of primary brain tumours are complicated by the varied tumour behaviour of primary or secondary gliomas (high-grade transformation of low-grade lesions), as well as the dilemmas with identification of radiation necrosis, tumour progression, and pseudoprogression on MRI. Radiomics and radiogenomics promise to offer precise diagnosis, predict prognosis, and assess tumour response to modern chemotherapy/immunotherapy and radiation therapy. This is achieved by a triumvirate of morphological, textural, and functional signatures, derived from a high-throughput extraction of quantitative voxel-level MR image metrics. However, the lack of standardisation of acquisition parameters and inconsistent methodology between working groups have made validations unreliable, hence multi-centre studies involving heterogenous study populations are warranted. We elucidate novel radiomic and radiogenomic workflow concepts and state-of-the-art descriptors in sub-visual MR image processing, with relevant literature on applications of such machine learning techniques in glioma management.

Reproducibility of 19 F-MR ventilation imaging in healthy volunteers.

PURPOSE: To assess the reproducibility of percentage ventilated lung volume (%VV) measurements in healthy volunteers acquired by fluorine (19 F)-MRI of inhaled perfluoropropane, implemented at two research sites. METHODS: In this prospective, ethically approved study, 40 healthy participants were recruited (May 2018-June 2019) to one of two research sites. Participants underwent a single MRI scan session on a 3T scanner, involving periodic inhalation of a 79% perfluoropropane/21% oxygen gas mixture. Each gas inhalation session lasted about 30 seconds, consisting of three deep breaths of gas followed by a breath-hold. Four 19 F-MR ventilation images were acquired per participant, each separated by approximately 6 minutes. The value of %VV was determined by registering separately acquired 1 H images to ventilation images before semi-automated image segmentation, performed independently by two observers. Reproducibility of %VV measurements was assessed by components of variance, intraclass correlation coefficients, coefficients of variation (CoV), and the Dice similarity coefficient. RESULTS: The MRI scans were well tolerated throughout, with no adverse events. There was a high degree of consistency in %VV measurements for each participant (CoVobserver1 = 0.43%; CoVobserver2 = 0.63%), with overall precision of %VV measurements determined to be within ± 1.7% (95% confidence interval). Interobserver agreement in %VV measurements revealed a high mean Dice similarity coefficient (SD) of 0.97 (0.02), with only minor discrepancies between observers. CONCLUSION: We demonstrate good reproducibility of %VV measurements in a group of healthy participants using 19 F-MRI of inhaled perfluoropropane. Our methods have been successfully implemented across two different study sites, supporting the feasibility of performing larger multicenter clinical studies.

A mechanistic evaluation of the potential for octamethylcyclotetrasiloxane to produce effects via endocrine modes of action.

This review is a hypothesis driven, mechanistic evaluation of the potential for octamethylcyclotetrasiloxane (D4) to produce any effects via endocrine modes of action. D4 is a volatile, lipophilic liquid used in the production of high molecular weight dimethylsiloxane polymers. These are used in a variety of industrial, medical, cleaning, and personal care products, and they may contain low levels of residual D4. Low concentrations of D4 are found in the environment and there is potential for low level human exposure. All of the measured environmental and workplace levels of D4 fall below no observed effect levels (NOEL). Most of the effects of high dose D4 involve the female reproductive system. In the mature intact female rat following chronic high dose exposure, D4 may cause inhibition of mating and ovulation, decreased live litter sizes, small increases in the estrogen to progesterone ratio primarily through decreases in progesterone, and increases in uterine hyperplasia. When endogenous estrogens are very low, high dose D4 causes increases in some uterine parameters. To assess whether these high dose effects can be attributed to an endocrine mode of action, endpoints are ranked for relevance and strength, consistent with published concepts. When sufficient information is available the level of activity of D4 for producing the observed effect is compared with that of potent endocrines. The conclusions reached are that all of the effects of D4 fall well short of any established criteria for D4 to be capable of producing any adverse effect via an endocrine mode of action.

Implementation of Universal Infant Free School Meals: a pilot study in NE England exploring the impact on Key Stage 1 pupil's dietary intake.

OBJECTIVE: To consider the principal effect of an interaction between year (pre- and post-Universal Infant Free School Meals (UIFSM)) and school on pupil's dietary intakes. DESIGN: A repeated cross-sectional survey using dietary data from 2008 to 2009 (pre-) and 2017 to 2018 (post-UIFSM). SETTING: Two primary schools, NE England. PARTICIPANTS: Pupils aged 4-7 years (2008-2009 n 121; 2017-2018 n 87). RESULTS: At lunchtime, there was a statistically significant decrease in pupils non-milk extrinsic sugars intake (%E NMEs) pre- to post-UIFSM (mean change -4·6 %; 95 % CI -6·3, -2·9); this was reflected in total diet (-3·8 %; -5·2, -2·7 %). A year and school interaction was found for mean Ca intakes: post-UIFSM pupils in School 2 had a similar mean intake as pre; in School 1 intakes had increased (difference of difference: -120 mg; 95 % CI -179, -62); no reflection in total diet. Post-UIFSM mean portions of yogurt decreased in School 2 and remained similar in School 1 (-0·25; -0·46, -0·04); this was similar for 'cake/pudding' and fruit. CONCLUSIONS: Within the limitations, these findings highlight positives and limitations following UIFSM implementation and demonstrate the role of school-level food practices on pupil's choices. To facilitate maximum potential of UIFSM, national levers, such as discussions on updating school food standards, including sugars, could consider removing the daily 'pudding' option and advocate 'fruit only' options 1 d/week, as some schools do currently. Small school-level changes could maximise positive health impacts by decreasing NMEs intake. A more robust evaluation is imperative to consider dietary impacts, equitability and wider effects on schools and families.

Implementation of the 'TAKE STOCK' Hot Debrief Tool in the ED: a quality improvement project.

Hot debriefing (HoD) describes a structured team-based discussion which may be initiated following a significant event. Benefits may include improved teamwork, staff well-being and identification of learning opportunities. Existing literature indicates that while staff value HoD following significant events, it is infrequently undertaken in practice. Internationally, several frameworks for HoD have been developed, although none are widely adopted for use in the ED. A quality improvement project was conducted to introduce HoD into a single UK ED in North West England, between January and March 2019. Following stakeholder consultation, the 9-item 'TAKE STOCK' tool was developed. Implementation of the tool increased the number of HoD (0-2.2 HoD episodes/week). Findings from the first plan-do-study-act (PDSA) cycle are presented, which revealed the key strengths and limitations of this model. Staff perceptions of the tool were evaluated using a self-administered short questionnaire designed by the authors. Satisfaction with TAKE STOCK was assessed using 10-point numerical scales. Across respondents (n-15), average satisfaction scores exceeded 9 out of 10 concerning patient care, staff self-care, decision-making, education, teamwork and identification of equipment issues. Implementation of HoD into the ED is feasible and viewed as beneficial by staff. Implementation toolkits for TAKE STOCK have been requested by 42 additional UK hospitals and ambulance trusts, demonstrating significant interest in its use. Research is now required to formally validate HoD frameworks for use in the ED, and assess whether HoD results in sustained improvements to staff and patient outcomes.

Adjuvant radiotherapy versus early salvage radiotherapy following radical prostatectomy (TROG 08.03/ANZUP RAVES): a randomised, controlled, phase 3, non-inferiority trial.

BACKGROUND: Adjuvant radiotherapy has been shown to halve the risk of biochemical progression for patients with high-risk disease after radical prostatectomy. Early salvage radiotherapy could result in similar biochemical control with lower treatment toxicity. We aimed to compare biochemical progression between patients given adjuvant radiotherapy and those given salvage radiotherapy. METHODS: We did a phase 3, randomised, controlled, non-inferiority trial across 32 oncology centres in Australia and New Zealand. Eligible patients were aged at least 18 years and had undergone a radical prostatectomy for adenocarcinoma of the prostate with pathological staging showing high-risk features defined as positive surgical margins, extraprostatic extension, or seminal vesicle invasion; had an Eastern Cooperative Oncology Group performance status of 0-1, and had a postoperative prostate-specific antigen (PSA) concentration of 0·10 ng/mL or less. Patients were randomly assigned (1:1) using a minimisation technique via an internet-based, independently generated allocation to either adjuvant radiotherapy within 6 months of radical prostatectomy or early salvage radiotherapy triggered by a PSA of 0·20 ng/mL or more. Allocation sequence was concealed from investigators and patients, but treatment assignment for individual randomisations was not masked. Patients were stratified by radiotherapy centre, preoperative PSA, Gleason score, surgical margin status, and seminal vesicle invasion status. Radiotherapy in both groups was 64 Gy in 32 fractions to the prostate bed without androgen deprivation therapy with real-time review of plan quality on all cases before treatment. The primary endpoint was freedom from biochemical progression. Salvage radiotherapy would be deemed non-inferior to adjuvant radiotherapy if freedom from biochemical progression at 5 years was within 10% of that for adjuvant radiotherapy with a hazard ratio (HR) for salvage radiotherapy versus adjuvant radiotherapy of 1·48. The primary analysis was done on an intention-to-treat basis. This study is registered with ClinicalTrials.gov, NCT00860652. FINDINGS: Between March 27, 2009, and Dec 31, 2015, 333 patients were randomly assigned (166 to adjuvant radiotherapy; 167 to salvage radiotherapy). Median follow-up was 6·1 years (IQR 4·3-7·5). An independent data monitoring committee recommended premature closure of enrolment because of unexpectedly low event rates. 84 (50%) patients in the salvage radiotherapy group had radiotherapy triggered by a PSA of 0·20 ng/mL or more. 5-year freedom from biochemical progression was 86% (95% CI 81-92) in the adjuvant radiotherapy group versus 87% (82-93) in the salvage radiotherapy group (stratified HR 1·12, 95% CI 0·65-1·90; pnon-inferiority=0·15). The grade 2 or worse genitourinary toxicity rate was lower in the salvage radiotherapy group (90 [54%] of 167) than in the adjuvant radiotherapy group (116 [70%] of 166). The grade 2 or worse gastrointestinal toxicity rate was similar between the salvage radiotherapy group (16 [10%]) and the adjuvant radiotherapy group (24 [14%]). INTERPRETATION: Salvage radiotherapy did not meet trial specified criteria for non-inferiority. However, these data support the use of salvage radiotherapy as it results in similar biochemical control to adjuvant radiotherapy, spares around half of men from pelvic radiation, and is associated with significantly lower genitourinary toxicity. FUNDING: New Zealand Health Research Council, Australian National Health Medical Research Council, Cancer Council Victoria, Cancer Council NSW, Auckland Hospital Charitable Trust, Trans-Tasman Radiation Oncology Group Seed Funding, Cancer Research Trust New Zealand, Royal Australian and New Zealand College of Radiologists, Cancer Institute NSW, Prostate Cancer Foundation Australia, and Cancer Australia.

A randomized, placebo-controlled trial evaluating effects of lebrikizumab on airway eosinophilic inflammation and remodelling in uncontrolled asthma (CLAVIER).

BACKGROUND: The anti-interleukin 13 (IL-13) monoclonal antibody lebrikizumab improves lung function in patients with moderate-to-severe uncontrolled asthma, but its effects on airway inflammation and remodelling are unknown. CLAVIER was designed to assess lebrikizumab's effect on eosinophilic inflammation and remodelling. OBJECTIVE: To report safety and efficacy results from enrolled participants with available data from CLAVIER. METHODS: We performed bronchoscopy on patients with uncontrolled asthma before and after 12 weeks of randomized double-blinded treatment with lebrikizumab (n = 31) or placebo (n = 33). The pre-specified primary end-point was relative change in airway subepithelial eosinophils per mm2 of basement membrane (cells/mm2 ). Pre-specified secondary and exploratory outcomes included change in IL-13-associated biomarkers and measures of airway remodelling. RESULTS: There was a baseline imbalance in tissue eosinophils and high variability between treatment groups. There was no discernible change in adjusted mean subepithelial eosinophils/mm2 in response to lebrikizumab (95% CI, -82.5%, 97.5%). As previously observed, FEV1 increased after lebrikizumab treatment. Moreover, subepithelial collagen thickness decreased 21.5% after lebrikizumab treatment (95% CI, -32.9%, -10.2%), and fractional exhaled nitric oxide, CCL26 and SERPINB2 mRNA expression in bronchial tissues also reduced. Lebrikizumab was well tolerated, with a safety profile consistent with other lebrikizumab asthma studies. CONCLUSIONS & CLINICAL RELEVANCE: We did not observe reduced tissue eosinophil numbers in association with lebrikizumab treatment. However, in pre-specified exploratory analyses, lebrikizumab treatment was associated with reduced degree of subepithelial fibrosis, a feature of airway remodelling, as well as improved lung function and reduced key pharmacodynamic biomarkers in bronchial tissues. These results reinforce the importance of IL-13 in airway pathobiology and suggest that neutralization of IL-13 may reduce asthmatic airway remodelling. CLINICAL TRIAL REGISTRATION: NCT02099656.

Perineural invasion by prostate adenocarcinoma in needle biopsies predicts bone metastasis: Ten year data from the TROG 03.04 RADAR Trial.

AIMS: Perineural invasion (PNI) by prostatic adenocarcinoma is debated as a prognostic parameter. This study investigates the prognostic predictive value of PNI in a series of patients with locally advanced prostate cancer treated with radiotherapy and androgen deprivation using 10 years outcome data from the TROG 03.04 RADAR trial. METHODS: Diagnostic prostate biopsies from 976 patients were reviewed and the presence of PNI noted. Patients were followed for 10 years according to the trial protocol or until death. The primary endpoint for the study was time to bone metastasis. Secondary endpoints included time to soft tissue metastasis, transition to castration resistance, prostate cancer-specific mortality and all-cause mortality. RESULTS: PNI was detected in 449 cases (46%), with 234 cases (24%) having PNI in more than one core. The presence of PNI was significantly associated with higher ISUP grade, clinical T staging category, National Comprehensive Cancer Network risk group, and percent positive biopsy cores. The cumulative probability of bone metastases according to PNI status was significant over the 10 years follow-up interval of the study (log-rank test P < 0.0001). PNI was associated with all endpoints on univariable analysis. After adjusting for baseline clinicopathological and treatment factors, bone metastasis was the only endpoint in which PNI retained its prognostic significance (hazard ratio 1.42, 95% confidence interval 1.05-1.92, P = 0.021). CONCLUSIONS: The association between PNI and the development of bone metastases supports the inclusion of this parameter as a component of the routine histology report. Further this association suggests that evaluation of PNI may assist in selecting those patients who should be monitored more closely during follow-up.

Highly efficient stepped wedge designs for clusters of unequal size.

The stepped wedge design (SWD) is a form of cluster randomized trial, usually comparing two treatments, which is divided into time periods and sequences, with clusters allocated to sequences. Typically all sequences start with the standard treatment and end with the new treatment, with the change happening at different times in the different sequences. The clusters will usually differ in size but this is overlooked in much of the existing literature. This paper considers the case when clusters have different sizes and determines how efficient designs can be found. The approach uses an approximation to the variance of the treatment effect, which is expressed in terms of the proportions of clusters and of individuals allocated to each sequence of the design. The roles of these sets of proportions in determining an efficient design are discussed and illustrated using two SWDs, one in the treatment of sexually transmitted diseases and one in renal replacement therapy. Cluster-balanced designs, which allocate equal numbers of clusters to each sequence, are shown to have excellent statistical and practical properties; suggestions are made about the practical application of the results for these designs. The paper concentrates on the cross-sectional case, where subjects are measured once, but it is briefly indicated how the methods can be extended to the closed-cohort design.