Establishment of CD8+ T Cell Thymic Central Tolerance to Tissue-Restricted Antigen Requires PD-1.

Highly self-reactive T cells are censored from the repertoire by both central and peripheral tolerance mechanisms upon receipt of high-affinity TCR signals. Clonal deletion is considered a major driver of central tolerance; however, other mechanisms such as induction of regulatory T cells and functional impairment have been described. An understanding of the interplay between these different central tolerance mechanisms is still lacking. We previously showed that impaired clonal deletion to a model tissue-restricted Ag did not compromise tolerance. In this study, we determined that murine T cells that failed clonal deletion were rendered functionally impaired in the thymus. Programmed cell death protein 1 (PD-1) was induced in the thymus and was required to establish cell-intrinsic tolerance to tissue-restricted Ag in CD8+ thymocytes independently of clonal deletion. In bone marrow chimeras, tolerance was not observed in PD-L1-deficient recipients, but tolerance was largely maintained following adoptive transfer of tolerant thymocytes or T cells to PD-L1-deficient recipients. However, CRISPR-mediated ablation of PD-1 in tolerant T cells resulted in broken tolerance, suggesting different PD-1 signaling requirements for establishing versus maintaining tolerance. Finally, we showed that chronic exposure to high-affinity Ag supported the long-term maintenance of tolerance. Taken together, our study identifies a critical role for PD-1 in establishing central tolerance in autoreactive T cells that escape clonal deletion. It also sheds light on potential mechanisms of action of anti-PD-1 pathway immune checkpoint blockade and the development of immune-related adverse events.

γδ Thymocyte Maturation and Emigration in Adult Mice.

Several unique waves of γδ T cells are generated solely in the fetal/neonatal thymus, whereas additional γδ T cell subsets are generated in adults. One intriguing feature of γδ T cell development is the coordination of differentiation and acquisition of effector function within the fetal thymus; however, it is less clear whether this paradigm holds true in adult animals. In this study, we investigated the relationship between maturation and thymic export of adult-derived γδ thymocytes in mice. In the Rag2pGFP model, immature (CD24+) γδ thymocytes expressed high levels of GFP whereas only a minority of mature (CD24-) γδ thymocytes were GFP+ Similarly, most peripheral GFP+ γδ T cells were immature. Analysis of γδ recent thymic emigrants (RTEs) indicated that most γδ T cell RTEs were CD24+ and GFP+, and adoptive transfer experiments demonstrated that immature γδ thymocytes can mature outside the thymus. Mature γδ T cells largely did not recirculate to the thymus from the periphery; rather, a population of mature γδ thymocytes that produced IFN-γ or IL-17 remained resident in the thymus for at least 60 d. These data support the existence of two populations of γδ T cell RTEs in adult mice: a majority subset that is immature and matures in the periphery after thymic emigration, and a minority subset that completes maturation within the thymus prior to emigration. Additionally, we identified a heterogeneous population of resident γδ thymocytes of unknown functional importance. Collectively, these data shed light on the generation of the γδ T cell compartment in adult mice.

Management of haematoma after thyroid surgery: systematic review and multidisciplinary consensus guidelines from the Difficult Airway Society, the British Association of Endocrine and Thyroid Surgeons and the British Association of Otorhinolaryngology, Head and Neck Surgery.

Haematoma after thyroid surgery can lead to airway obstruction and death. We therefore developed guidelines to improve the safety of peri-operative care of patients undergoing thyroid surgery. We conducted a systematic review to inform recommendations, with expert consensus used in the absence of high-quality evidence, and a Delphi study was used to ratify recommendations. We highlight the importance of multidisciplinary team management and make recommendations in key areas including: monitoring; recognition; post-thyroid surgery emergency box; management of suspected haematoma following thyroid surgery; cognitive aids; post-haematoma evacuation care; day-case thyroid surgery; training; consent and pre-operative communication; postoperative communication; and institutional policies. The guidelines support a multidisciplinary approach to the management of suspected haematoma following thyroid surgery through oxygenation and evaluation; haematoma evacuation; and tracheal intubation. They have been produced with materials to support implementation. While these guidelines are specific to thyroid surgery, the principles may apply to other forms of neck surgery. These guidelines and recommendations provided are the first in this area and it is hoped they will support multidisciplinary team working, improving care and outcomes for patients having thyroid surgery.

Unexpected difficult tracheal intubation secondary to subglottic stenosis leading to emergency front-of-neck airway.

A 26-year-old woman presenting for an elective day case procedure under general anaesthesia had undiagnosed subglottic stenosis leading to a life threatening airway emergency requiring emergency front-of-neck airway. We outline the case and discuss key anaesthetic considerations in subglottic stenosis, including concerning features of a medical/anaesthetic history and the potential for rapid deterioration of a stenotic airway following manipulation. We also consider the effect of anaesthesia on the calibre of subglottic stenosis and the effects of positive pressure ventilation. Subglottic stenosis is a rare condition with congenital, acquired and idiopathic origins; however, iatrogenic trauma is the most common cause. We are aware of a small number of published case reports of previously undiagnosed subglottic stenosis in adults discovered after induction of anaesthesia; situational deterioration to 'cannot intubate, cannot oxygenate' scenarios appear even rarer.

Measuring antimicrobial use on dairy farms: A method comparison cohort study.

Antimicrobial use on UK dairy farms is measured for surveillance purposes, with veterinary sales data as a proxy for use. Two other methods of recording use have been used commonly on-farm: medicine waste bins and farm medicine records. However, none of these methods has been validated to measure antimicrobial use. The objective of this research was to assess agreement between the 3 most common methods for measuring on-farm antimicrobial use with a predetermined reference method on UK dairy farms. Antimicrobial use was measured prospectively on 26 UK dairy farms using medicine waste bins into which participants placed all discarded medicine packaging for a 12-mo period. At the end of 12 mo, farm medicine records and veterinary sales data were obtained retrospectively for participating farms. The reference method used was based on pre- and poststudy inventories combined with veterinary sales data. We investigated the systematic difference between the mean on-farm antimicrobial use measured by each of the 3 methods and a reference method, using one-way repeated-measures ANOVA models. Reliability and clinical relevance of the agreement between each pair of methods was quantified using the concordance correlation coefficient (CCC) and the Bland-Altman method, respectively. When compared with the reference method, veterinary sales data had excellent reliability for injectable antimicrobials and intramammary antimicrobials [95% confidence interval (CI) of CCC > 0.90] and moderate to excellent reliability for other antimicrobials (95% CI of CCC: 0.68-0.97). Medicine waste bins had good to excellent reliability for injectable (95% CI of CCC: 0.84-0.99), and intramammary products (95% CI of CCC: 0.78-0.94) and no agreement for other forms of antimicrobial. Farm medicine records did not agree for any form of antimicrobial when compared with the reference method. The use of veterinary sales data as a proxy for on-farm antimicrobial use in the UK represented excellent statistical reliability and offered clinically good agreement with the reference method when used to measure injectable antimicrobials. This study applies to the UK context and included a relatively small number of farms. However, these results have research and policy implications, both nationally and internationally, and are essential in accurately quantifying agricultural antimicrobial use to inform both animal and human health.

The emergence of magnetic ordering at complex oxide interfaces tuned by defects.

Complex oxides show extreme sensitivity to structural distortions and defects, and the intricate balance of competing interactions which emerge at atomically defined interfaces may give rise to unexpected physics. In the interfaces of non-magnetic complex oxides, one of the most intriguing properties is the emergence of magnetism which is sensitive to chemical defects. Particularly, it is unclear which defects are responsible for the emergent magnetic interfaces. Here, we show direct and clear experimental evidence, supported by theoretical explanation, that the B-site cation stoichiometry is crucial for the creation and control of magnetism at the interface between non-magnetic ABO3-perovskite oxides, LaAlO3 and SrTiO3. We find that consecutive defect formation, driven by atomic charge compensation, establishes the formation of robust perpendicular magnetic moments at the interface. Our observations propose a route to tune these emerging magnetoelectric structures, which are strongly coupled at the polar-nonpolar complex oxide interfaces.

Deciphering the Immunomodulatory Capacity of Oncolytic Vaccinia Virus to Enhance the Immune Response to Breast Cancer.

Vaccinia virus (VACV) is a double-stranded DNA virus that devotes a large portion of its 200 kbp genome to suppressing and manipulating the immune response of its host. Here, we investigated how targeted removal of immunomodulatory genes from the VACV genome impacted immune cells in the tumor microenvironment with the intention of improving the therapeutic efficacy of VACV in breast cancer. We performed a head-to-head comparison of six mutant oncolytic VACVs, each harboring deletions in genes that modulate different cellular pathways, such as nucleotide metabolism, apoptosis, inflammation, and chemokine and interferon signaling. We found that even minor changes to the VACV genome can impact the immune cell compartment in the tumor microenvironment. Viral genome modifications had the capacity to alter lymphocytic and myeloid cell compositions in tumors and spleens, PD-1 expression, and the percentages of virus-targeted and tumor-targeted CD8+ T cells. We observed that while some gene deletions improved responses in the nonimmunogenic 4T1 tumor model, very little therapeutic improvement was seen in the immunogenic HER2/neu TuBo model with the various genome modifications. We observed that the most promising candidate genes for deletion were those that interfere with interferon signaling. Collectively, this research helped focus attention on the pathways that modulate the immune response in the context of VACV oncolytic virotherapy. They also suggest that the greatest benefits to be obtained with these treatments may not always be seen in "hot tumors."

Analysis of a National Programme for Selective Internal Radiation Therapy for Colorectal Cancer Liver Metastases.

AIMS: Patients with chemotherapy-refractory colorectal cancer liver metastases have limited therapeutic options. Selective internal radiation therapy (SIRT) delivers yttrium 90 microspheres as a minimally invasive procedure. This prospective, single-arm, observational, service-evaluation study was part of National Health Service England Commissioning through Evaluation. METHODS: Patients eligible for treatment had histologically confirmed carcinoma with liver-only/liver-dominant metastases with clinical progression during or following oxaliplatin-based and irinotecan-based chemotherapy. All patients received SIRT plus standard of care. The primary outcome was overall survival; secondary outcomes included safety, progression-free survival (PFS) and liver-specific PFS (LPFS). RESULTS: Between December 2013 and March 2017, 399 patients were treated in 10 centres with a median follow-up of 14.3 months (95% confidence interval 9.2-19.4). The median overall survival was 7.6 months (95% confidence interval 6.9-8.3). The median PFS and LPFS were 3.0 months (95% confidence interval 2.8-3.1) and 3.7 months (95% confidence interval 3.2-4.3), respectively. During the follow-up period, 143 patients experienced an adverse event and 8% of the events were grade 3. CONCLUSION: Survival estimates from this pragmatic study show clinical outcomes attainable in the National Health Service comparable with previously published data. This study shows the value of a registry-based commissioning model to aid national commissioning decisions for highly specialist cancer treatments.

Clinical and Psychosocial Factors Influencing Retinal Screening Uptake Among Young Adults with Type 2 Diabetes.

PURPOSE OF REVIEW: Young adults with type 2 diabetes (T2D, 18-39 years) experience early-onset and rapid progression of diabetic retinopathy (DR), the leading cause of vision loss for working age adults. Despite this, uptake of retinal screening, the crucial first step in preventing vision loss from DR, is low. The aim of this review is to summarize the clinical and psychosocial factors affecting uptake of retinal screening. RECENT FINDINGS: Barriers include lack of diabetes-related symptoms, low personal DR risk perception, high rates of depression and diabetes-related distress, fatalism about inevitability of complications, time and financial constraints, disengagement with existing diabetes self-management services, and perceived stigma due to having a condition associated with older adults. Young adults with T2D are an under-researched population who face an accumulation of barriers to retinal screening. Tailored interventions that address the needs, characteristics, and priorities of young adults with T2D are warranted.

Nutrient-extraction blender preparation reduces postprandial glucose responses from fruit juice consumption.

Although whole-fruit consumption is regarded as protective against type 2 diabetes (T2DM), conventionally prepared fruit juice is associated with increased T2DM risk, and current public health advice recommends its restriction. 'Nutrient extractor' style blenders are increasing in popularity worldwide as an alternative means of juicing fruit, but little is known about their effect on postprandial glucose levels. The current study investigated the effect of nutrient extraction on postprandial blood glucose response and glycemic index (GI) compared with a glucose control for both mixed fruit and a high GI fruit (mango). Remarkably, consumption of nutrient-extracted mixed fruit resulted in a significant lowering of the GI (32.7±8.5) compared with whole mixed fruit (66.2±8.2, P<0.05). For the high GI mango, there were no differences between nutrient-extracted and whole fruit, indicating that even for a high GI fruit the effect of nutrient extraction does not increase GI compared with the whole fruit. These findings suggest that, in contrast to conventionally prepared fruit juice, fruit juice prepared by nutrient extraction in some cases elicits a more favorable postprandial glycemic response than whole fruit and even for high GI fruits do not worsen the response. The mechanism responsible for this effect is currently unclear. However, these results suggest that fruit homogenized by nutrient extraction should be considered as a potential dietetic strategy for glycemic control.

Vision Amniotic Leak Detector (ALD) to Eliminate Amniotic Fluid Leakage as a Cause of Vaginal Wetness in Pregnancy: A NICE Medical Technology Guidance.

In prelabour rupture of membranes (PROM) or preterm PROM the amniotic membranes rupture prior to labour. Where this is not overt a speculum examination is undertaken to confirm diagnosis. The Vision Amniotic Leak Detector (ALD) is a panty liner that can diagnose amniotic fluid as a cause of vaginal wetness. It was evaluated by the UK National Institute for Health and Care Excellence (NICE) as part of the Medical Technologies Evaluation Programme. The sponsor (CommonSense Ltd) identified five studies, of which three were deemed within scope by the External Assessment Centre (EAC). Two of these three used an inappropriate comparator. The EAC recalculated the diagnostic accuracy of Vision ALD using speculum examination as the comparator: sensitivity of 97% (95% CI 93-99%), negative predictive value of 96% (95% CI 92-98%). A negative result would therefore allow patients to be discharged with confidence. In the sponsor's cost-consequence model only patients with a positive Vision ALD result would have a speculum examination, producing a cost saving of around £10 per patient. The EAC felt that some costs were unjustified and the model did not include infection outcomes or use in a community setting. The EAC revised the sponsor's model and found the results were most sensitive to clinician costs. Vision ALD was associated with savings of around £15-£25 per patient when administration in lower-cost community healthcare avoided a referral to a higher-cost secondary-care centre. NICE published guidance MTG15 in July 2013 recommending that the case for adopting Vision ALD was supported by the evidence.

The ScanTrainer obstetrics and gynaecology ultrasound virtual reality training simulator: A cost model to determine the cost viability of replacing clinical training with simulation training.

The aim of this study was to produce an economic cost model comparing the use of the Medaphor ScanTrainer virtual reality training simulator for obstetrics and gynaecology ultrasound to achieve basic competence, with the traditional training method. A literature search and survey of expert opinion were used to identify resources used in training. An executable model was produced in Excel. The model showed a cost saving for a clinic using the ScanTrainer of £7114 per annum. The uncertainties of the model were explored and it was found to be robust. Threshold values for the key drivers of the model were identified. Using the ScanTrainer is cost saving for clinics with at least two trainees per year to train, if it would take at least six lists to train them using the traditional training method and if a traditional training list has at least two fewer patients than a standard list.

Spontaneous neuronal activity predicts intersubject variations in executive control of attention.

Executive control of attention regulates our thoughts, emotion and behavior. Individual differences in executive control are associated with task-related differences in brain activity. But it is unknown whether attentional differences depend on endogenous (resting state) brain activity and to what extent regional fluctuations and functional connectivity contribute to individual variations in executive control processing. Here, we explored the potential contribution of intrinsic brain activity to executive control by using resting-state functional magnetic resonance imaging (fMRI). Using the amplitude of low-frequency fluctuations (ALFF) as an index of spontaneous brain activity, we found that ALFF in the right precuneus (PCUN) and the medial part of left superior frontal gyrus (msFC) was significantly correlated with the efficiency of executive control processing. Crucially, the strengths of functional connectivity between the right PCUN/left msFC and distributed brain regions, including the left fusiform gyrus, right inferior frontal gyrus, left superior frontal gyrus and right precentral gyrus, were correlated with individual differences in executive performance. Together, the ALFF and functional connectivity accounted for 67% of the variability in behavioral performance. Moreover, the strength of functional connectivity between specific regions could predict more individual variability in executive control performance than regionally specific fluctuations. In conclusion, our findings suggest that spontaneous brain activity may reflect or underpin executive control of attention. It will provide new insights into the origins of inter-individual variability in human executive control processing.

Feedback of personal retinal images appears to have a motivational impact in people with non-proliferative diabetic retinopathy and suboptimal HbA1c: findings of a pilot study.

AIM: To conduct a pilot study to explore the potential impact of visual feedback of personal retinal images on diabetes outcomes. METHODS: Twenty-five participants with non-proliferative diabetic retinopathy and suboptimal HbA(1c) (> 53 mmol/mol; > 7%) were randomized to receive visual feedback of their own retinal images or to a control group. At baseline and 3-month follow-up, HbA(1c), standard measures of beliefs, diabetes-related distress and self-care activities were assessed. RESULTS: In unadjusted models, relative to controls, the intervention group showed significantly greater improvement in HbA(1c) at 3-month follow-up (-0.6% vs. +0.3%, P < 0.01), as well as enhanced motivation to improve blood glucose management (P < 0.05). CONCLUSIONS: This small pilot study provides preliminary evidence that visual feedback of personal retinal images may offer a practical educational strategy for clinicians in eye care services to improve diabetes outcomes in non-target compliant patients. A fully powered randomized controlled trial is required to confirm these findings and determine the optimal use of feedback to produce sustained effects.

The effect of monoalkyl phosphates and fluoride on dissolution of hydroxyapatite, and interactions with saliva.

The aims were to investigate the effect of monoalkyl phosphates (MAPs) and fluoride on dissolution rate of native and saliva-coated hydroxyapatite (HA). Fluoride at 300 mg/l (as NaF) inhibited dissolution of native HA by 12%, while potassium and sodium dodecyl phosphates (PDP, SDP), at 0.1% or higher, inhibited dissolution by 26-34%. MAPs, but not fluoride, also showed persistence of action. MAPs at 0.5% and fluoride at 300 mg/l were then tested separately against HA pre-treated with human saliva for 2 or 18 h. Agents were applied with brushing to half the specimens, and without brushing to the other half. In control (water-treated) specimens, pre-treatment of HA with human saliva reduced dissolution rate on average by 41% (2 h) and 63% (18 h). Brushing did not have a statistically significant effect on dissolution rate of saliva-coated specimens. In brushed specimens, fluoride significantly increased the inhibition due to 2- or 18-hour saliva pre-treatment. It is hypothesised that brushing partially removes the salivary film and allows KOH-soluble calcium fluoride formation at the surfaces of HA particles. Inhibition was reduced by PDP in 2-hour/non-brushed specimens and in 18-hour/brushed specimens. PDP did not affect dissolution rates in the remaining groups and SDP did not affect dissolution rate in any group. Possible reasons for these variable results are discussed. The experiments show that pre-treatment with saliva can significantly modify results of tests on potential anti-erosive agents and it is recommended that saliva pre-treatment should be a routine part of testing such agents.

Protein engineering and preclinical development of a GM-CSF receptor antibody for the treatment of rheumatoid arthritis.

BACKGROUND AND PURPOSE: For antibody therapies against receptor targets, in vivo outcomes can be difficult to predict because of target-mediated clearance or antigen 'sink' effects. The purpose of this work was to engineer an antibody to the GM-CSF receptor α (GM-CSFRα) with pharmacological properties optimized for chronic, s.c. treatment of rheumatoid arthritis (RA) patients. EXPERIMENTAL APPROACH: We used an in silico model of receptor occupancy to guide the target affinity and a combinatorial phage display approach for affinity maturation. Mechanism of action and internalization assays were performed on the optimized antibody in vitro before refining the modelling predictions of the eventual dosing in man. Finally, in vivo pharmacology studies in cynomolgus monkeys were carried out to inform the predictions and support future clinical development. KEY RESULTS: Antibody potency was improved 8600-fold, and the target affinity was reached. The refined model predicted pharmacodynamic effects at doses as low as 1 mg kg(-1) and a study in cynomolgus monkeys confirmed in vivo efficacy at 1 mg kg(-1) dosing. CONCLUSIONS AND IMPLICATIONS: This rational approach to antibody drug discovery enabled the isolation of a potent molecule compatible with chronic, s.c. self-administration by RA patients. We believe this general approach enables the development of optimal biopharmaceuticals.