Context-dependent TGFß family signalling in cell fate regulation.

The transforming growth factor-ß (TGFß) family are a large group of evolutionarily conserved cytokines whose signalling modulates cell fate decision-making across varying cellular contexts at different stages of life. Here we discuss new findings in early embryos that reveal how, in contrast to our original understanding of morphogen interpretation, robust cell fate specification can originate from a noisy combination of signalling inputs and a broad range of signalling levels. We compare this evidence with novel findings on the roles of TGFß family signalling in tissue maintenance and homeostasis during juvenile and adult life, spanning the skeletal, haemopoietic and immune systems. From these comparisons, it emerges that in contrast to robust developing systems, relatively small perturbations in TGFß family signalling have detrimental effects at later stages in life, leading to aberrant cell fate specification and disease, for example in cancer or congenital disorders. Finally, we highlight novel strategies to target and amend dysfunction in signalling and discuss how gleaning knowledge from different fields of biology can help in the development of therapeutics for aberrant TGFß family signalling in disease.

Transition and reversal: long-term experience of the MACE procedure in children.

PURPOSE: Analysis of outcomes and follow-up of children who underwent the Malone antegrade continence enema (MACE) procedure in a UK tertiary paediatric surgery unit. METHODS: Children who underwent a MACE procedure from 1998 to 2020 were identified. Demographic and clinical data were obtained from contemporaneous records. Outcomes were categorised as full (success), partial or failure. RESULTS: Ninety-five children were identified for inclusion (chronic idiopathic constipation (CIC, 59), anorectal malformations (ARM, 23) and Hirschsprung's disease (HD, 13)). Mean age at surgery was 9.4 years (3-19 years) and mean follow-up time was 6 years (0.3-16.8 years). Outcomes were successful in 69% of CIC patients, 78% in ARM and 69% in HD. Twenty (21%) underwent MACE reversal after developing independent continence, with a significant difference between groups (CIC 19%, ARM 9%, HD 54%, p = 0.0047). 50% of patients > 16 years old were transitioned to adult services. CONCLUSION: We report a success rate of 72% for MACE procedures in our unit, with a significant difference in reversal rate between diagnostic groups. Long term, a fifth of patients no longer required their MACE. When these patients reach adolescence, those who require ongoing support outside of the paediatric surgery setting should be safely transitioned to adult services.

Grief and the non-death losses of Covid-19.

Articles in the popular media and testimonies collected in empirical work suggest that many people who have not been bereaved have nevertheless grieved over pandemic-related losses of various kinds. There is a philosophical question about whether any experience of a non-death loss ought to count as grief, hinging upon how the object of grief is construed. However, even if one accepts that certain significant non-death losses are possible targets of grief, many reported cases of putative pandemic-related grief may appear less plausible. For instance, it might be argued that many of these losses are temporary or minor and therefore unlikely to be grieved, and that the associated experiences are phenomenologically dissimilar to grief. In this article, as well as discussing the more general question about the coherence of the idea of non-bereavement grief, we address these obstacles to taking reports of pandemic non-bereavement grief to be literal and true. In particular, we argue that some may have experienced grief over even apparently minor losses during the pandemic. This is generally so, we suggest, only insofar as experiences of such losses form part of an overarching grief process directed at some broader significant loss. Thus, we cast light on both the nature of non-bereavement grief and the kinds of disruption and loss experienced during the pandemic.

A Review of the Use of EQ-5D for Clinical Outcome Assessment in Health Technology Assessment, Regulatory Claims, and Published Literature.

The aim is to identify the extent to which EQ-5D is used as a clinical outcome assessment (COA) endpoint in a non-economic context in health technology assessment (HTA) decisions, regulatory labelling claims and published literature. Drug technology appraisals (TAs) published by HTA agencies in England, France, Germany and the USA between 2019 and 2021 were identified. Product labelling for drugs approved by the European Medicines Agency (EMA) and US Food and Drug Administration (FDA) between 2016 and 2021 were also identified. A systematic literature review (SLR) was also performed. Documents reporting EQ-5D in the context of economic evaluation only were excluded. EQ-5D data were reported for COA in 195 of 1072 (18%) published TAs, with the majority reported for Germany (n = 138). The EQ-5D visual analogue scale (EQ-VAS) was reported most frequently, in 68% of all TAs, and accounted for 100% of Institut für Qualität und Wirtschaftlichkeit im Gesundheitswesen (IQWiG) and 94% of Gemeinsamer Bundesausschuss (G-BA) TAs. In total, 320 drugs were approved or reviewed by the EMA and 735 by the FDA. Of these, 15 reported EQ-5D data from the EMA and 35 from the FDA; however, all EQ-5D data submitted to the FDA were reported in supporting documentation. Reporting of both EQ-5D index and EQ-VAS was most frequent, occurring in 32% of all documents. For the SLR, 329 of 4248 (8%) retrieved records were included. Reporting of both EQ-5D index and EQ-VAS was most frequent, occurring in 36% of studies. Clinical evaluation of recent drug approvals, based on regulatory, HTA and systematic literature reviews, demonstrated limited use of EQ-5D outside the context of economic evaluations. This may be due to the likelihood that the EQ-5D may lack sensitivity to detect improvement in conditions with small expected therapeutic benefit, or because the EQ-5D is not considered an adequate COA tool for clinical evaluation of treatment benefit. EQ-5D, as a COA, was more likely to be used in clinical evaluation of cancer drugs than drugs for treatment in any other disease category. HTA bodies were more likely to use the EQ-5D for COA, especially in Germany.

RNA-Seq analysis of a Pax3-expressing myoblast clone in-vitro and effect of culture surface stiffness on differentiation.

Skeletal muscle satellite cells cultured on soft surfaces (12 kPa) show improved differentiation than cells cultured on stiff surfaces (approximately 100 kPa). To better understand the reasons for this, we performed an RNA-Seq analysis for a single satellite cell clone (C1F) derived from the H2kb-tsA58 immortomouse, which differentiates into myotubes under tightly regulated conditions (withdrawal of É£-interferon, 37 °C). The largest change in overall gene expression occurred at day 1, as cells switched from proliferation to differentiation. Surprisingly, further analysis showed that proliferating C1F cells express Pax3 and not Pax7, confirmed by immunostaining, yet their subsequent differentiation into myotubes is normal, and enhanced on softer surfaces, as evidenced by significantly higher expression levels of myogenic regulatory factors, sarcomeric genes, enhanced fusion and improved myofibrillogenesis. Levels of mRNA encoding extracellular matrix structural constituents and related genes were consistently upregulated on hard surfaces, suggesting that a consequence of differentiating satellite cells on hard surfaces is that they attempt to manipulate their niche prior to differentiating. This comprehensive RNA-Seq dataset will be a useful resource for understanding Pax3 expressing cells.

Disruption of the MSL complex inhibits tumour maintenance by exacerbating chromosomal instability.

Rewiring of cellular programmes in malignant cells generates cancer-specific vulnerabilities. Here, using an unbiased screening strategy aimed at identifying non-essential genes required by tumour cells to sustain unlimited proliferative capacity, we identify the male-specific lethal (MSL) acetyltransferase complex as a vulnerability of genetically unstable cancers. We find that disruption of the MSL complex and consequent loss of the associated H4K16ac mark do not substantially alter transcriptional programmes but compromise chromosome integrity and promote chromosomal instability (CIN) that progressively exhausts the proliferative potential of cancer cells through a p53-independent mechanism. This effect is dependent on pre-existing genomic instability, and normal cells are insensitive to MSL disruption. Using cell- and patient-derived xenografts from multiple cancer types, we show that excessive CIN induced by MSL disruption inhibits tumour maintenance. Our findings suggest that targeting MSL may be a valuable means to increase CIN beyond the level tolerated by cancer cells without inducing severe adverse effects in normal tissues.

Prevalence of PD-L1 expression in matched recurrent and/or metastatic sarcoma samples and in a range of selected sarcomas subtypes.

We assessed the frequency of programmed death-ligand 1 (PD-L1) expression by immunohistochemistry (IHC) in a cohort of 522 sarcomas from 457 patients, incuding a subset of 46 patients with 63 matched samples from local recurrence or metastases with primary tumours and/or metachronous metastases. We also investigated the correlation of PD-L1 with the presence and degree of tumour-infiltrating lymphocytes (TILs) in a subset of cases. IHC was performed using the PD-L1 SP263 companion kit (VENTANA) on tissue microarrays from an archival cohort. Evaluation of PD-L1 and TILs was performed on full sections for a subset of 23 cases. Fisher's exact and Mann Whitney test were used to establish significance (P <0.05). PD-L1 positive expression (≥1%) was identified in 31% of undifferentiated pleomorphic sarcomas, 29% of angiosarcomas, 26% of rhabdomyosarcomas, 18% of myxofibrosarcomas, 11% of leiomyosarcomas and 10% of dedifferentiated liposarcomas. Negative expression was present in all atypical lipomatous tumous/well-differentiated lipoasarcomas, myxoid liposarcomas, synovial sarcomas, pleomorphic liposarcomas, and Ewing sarcomas. PD-L1 IHC was concordant in 81% (38 of 47) of matched/paired samples. PD-L1 IHC was discordant in 19% (9 of 47 matched/paired samples), displaying differences in the proportion of cells expressing PD-L1 amongst paired samples with the percentage of PD-L1-positive cells increasing in the metastatic/recurrent site compared to the primary in 6 of 9 cases (67%). Significant correlation between PD-L1 expression and the degree of TILs was exclusively identified in the general cohort of leiomyosarcomas, but not in other sarcoma subtypes or in metastatic/recurrent samples. We conclude that the prevalence of PD-L1 expression in selected sarcomas is variable and likely to be clone dependent. Importantly, we demonstrated that PD-L1 can objectively increase in a small proportion of metastases/recurrent sarcomas, offering the potential of treatment benefit to immune checkpoint inhibitors in this metastatic setting.

Spatial spread of neural excitation in cochlear implant recipients: comparison of improved ECAP method and psychophysical forward masking.

This study introduces and evaluates a method for measurement of the longitudinal spread of electrically evoked neural excitation in the cochlea, using the Neural Response Telemetry system (NRT) available with the Nucleus((R)) 24 cochlear implant system. The recently released version of the NRT software (version 3.0) enables presentation of the 'masker' and 'probe' on different electrodes. In the present method the probe position was fixed, while the masker position was varied across the electrode array. The amplitude of the response to the partially masked probe provides a measure of the amount of masking, which is dependent on the extent of overlap of the excitation regions of the masker and probe. These measurements were performed in seven subjects implanted with the Nucleus 24 cochlear implant system (four with straight and three with Contour electrode arrays), for basal, middle and apical probe electrodes. Similar excitation profiles were obtained using either the standard NRT subtraction paradigm or an alternative 'Miller' method. The excitation profiles were compared with those obtained from psychophysical forward masking and good agreement was found. The widths of electrically evoked compound action potential (ECAP) and forward masking profiles did not differ significantly. Whereas the width of the ECAP measure was significantly correlated with both the maximum comfortable level and the distance of the electrode band from the modiolus, the width of the forward masking profile was not.

Management of acute displaced midshaft clavicular fractures using Herbert cannulated screw: Technique and results in 114 patients.

PURPOSE: A new and simple operative technique has been developed to provide internal fixation for midshaft clavicle fractures. This involves the use of a large fragment Herbert Screw that is entirely embedded within the bone. Screw fixation is combined with bone grafting from intramedullary reamings of the fracture fragments. The purpose of this report is to assess the outcomes following treatment of midshaft clavicular fracture using this method. MATERIALS AND METHODS: One hundred and fourteen patients with acute displaced midshaft fracture were identified between 2002 and 2007. All patients were followed until fracture union. Patients' medical records were reviewed. Disability of the Arm, Shoulder, and Hand questionnaire (DASH), and American Shoulder and Elbow Surgeons Elbow form (ASES) were posted to all patients. Outcome measures included union rate, time to union, implant removal rate, DASH, and ASES scores. RESULTS: Patients' median age was 29.5 years (interquartile range, 19-44 years). The most common injury mechanism was sports injury (28%). The median time from injury to surgery was 5 days (interquartile range, 2-9 days). Union occurred in an average of 8.8 weeks. Non-union occurred in three cases (2.6%). The re-operation rate for symptomatic hardware prominence screw was 1.7%. The median DASH score was 0.83 and the median ASES was 100 (n = 35). CONCLUSIONS: Intramedullary fixation using cannulated Herbert screw can be used as an effective approach for operative management of midshaft clavicular fractures. Using this method, an appropriate outcome could be achieved and a second intervention for implant removal could be avoided in great majority of cases. LEVEL OF EVIDENCE: Level III.

NMDA receptor antagonists distort visual grouping in rats performing a modified two-choice visual discrimination task.

RATIONALE: Visual perception is impaired during pathological psychosis, which can be mimicked by NMDA receptor antagonists. However, the underlying mechanisms are poorly understood, partly due to limits of current rodent models for visual integration. OBJECTIVES: The objectives of the study are (1) to develop a rodent task that can differentiate between effects on perception and nonspecific effects on task performance and (2) to test whether NMDA receptor antagonists affect visual perception in rats. METHODS: We used an adaptation of Glass patterns to assess visual grouping in rats using a two-choice visual discrimination task in an infrared touch screen conditioning chamber. After rats learned to discriminate between a radial and a concentric bipole pattern, the ability to discriminate between these patterns was tested at various levels of distortion and a psychometric function was fit to obtain the maximum task performance and signal level needed for half-maximum performance. RESULTS: NMDA receptor antagonists ketamine and phencyclidine at low doses increased the signal quality needed to discriminate between the visual patterns, without affecting the ability to discriminate between undistorted images. At higher doses, the ability to perform the task even with undistorted images was impaired, which was associated with stereotypic behaviour and increased impulsivity. CONCLUSIONS: The Glass pattern-based visual grouping task is able to differentiate the effect of psychotomimetic NMDA receptor antagonists on visual perception from the effects on motor and memory functions. The half-maximum performance signal level allows quantification of cognitive psychosis in rodents, which can be translated to human psychometric functions and can be used in the development of more effective treatments.

Delayed surgical treatment of displaced midshaft clavicle fracture using Herbert cannulated screw with intramedullary bone graft.

There is an increasing trend towards operative treatment for displaced midshaft clavicle fractures. This retrospective study was performed to assess the outcome of delayed fixation of displaced midshaft clavicle fractures and test the null hypothesis that there is no difference in results between early and delayed surgical treatment for displaced midshaft clavicle fractures. Using the hospital database, two groups of patients who were surgically treated using Herbert cannulated screw for displaced midshaft clavicle fractures were identified. There were 114 cases in acute (median time to operation of 5 days) and 21 cases in delayed groups (median time to operation 10.5 weeks). Thirty-five cases were available for follow-up in the acute and 16 cases in the delayed series. The primary outcome was assessed by the Disabilities of the Arm, Shoulder and Hand (DASH) score, American Shoulder and Elbow Surgeons score (ASES) and Constant-Murley score. Union occurred in 14 cases in the delayed group. The median DASH, mean ASES and Constant-Murley scores were 4.1, 97 and 88.7, respectively. Compared with the delayed group, the early group had a higher union rate (P = 0.033), trend towards lower DASH score (P = 0.051), and higher ASES score (P = 0.047). The delayed group had significantly more problems with prominent, symptomatic screws that required removal (P = 0.002). There were no significant differences in union time and complication rate. Delayed fixation of displaced midshaft fractures using the Herbert cannulated bone screw and bone graft is effective and provides a good functional outcome that only slightly reduced from that recorded for early fixation.

Spatial spread of neural excitation: comparison of compound action potential and forward-masking data in cochlear implant recipients.

The feasibility of using the electrically evoked compound action potential (ECAP), measured with the NRT system (Neural Response Telemetry) and the NRT software (version 2.04), to quantify the longitudinal spread of neural excitation was examined in four subjects fitted with the Nucleus C124M cochlear implant. The ECAP and psychophysical forward-masking profiles were measured using stimulation on each of three electrodes, in basal, middle and apical positions. Spatial spread profiles derived from the ECAP measure produced broader functions than those derived from the psychophysical measure. These results, together with investigation of the change of ECAP spread pattern with stimulation current, suggest that functions derived from this 'simple' method were more influenced by the spread of electric field from excited neuron to electrode array than by breadth of the neural excitation pattern. The recently released NRT version 3.0 permits the masker and probe pulses to be delivered to separate electrodes, thus removing a fundamental limitation of version 2.04. Results from a pilot study, in which this capability was exploited, suggest that the spatial functions from this 'advanced' method may provide a better correlation with results from psychophysical forward masking.

Application of loudness models to sound processing for cochlear implants.

A new paradigm for processing sound signals for multiple-electrode cochlear implants is introduced, and results are presented from an initial psychophysical evaluation of its effect on the perceived loudness of complex sounds. A real-time processing scheme based on this paradigm, called SpeL, has been developed primarily to improve control of loudness for implant users. SpeL differs from previous schemes in several ways. Most importantly, it incorporates a published numerical model which predicts the loudness perceived by implant users for complex patterns of pulsatile electric stimulation as a function of the pulses' physical parameters. This model is controlled by the output of a corresponding model that estimates the loudness perceived by normally hearing listeners for complex sounds. The latter model produces an estimate of the specific loudness arising from an acoustic signal. In SpeL, the specific loudness function, which describes the contribution to total loudness of each of a number of frequency bands (or cochlear positions), is converted to a pattern of electric stimulation on an appropriate set of electrodes. By application of the loudness model for electric stimulation, this pattern is designed to produce a specific loudness function for the implant user which approximates that produced by the normal-hearing model for the same input signal. The results of loudness magnitude estimation experiments with five users of the SpeL scheme confirmed that the psychophysical functions relating overall loudness perceived to input sound level for five complex acoustic signals were, on average, very similar to those for normal hearing.