Constitutive internalisation of EP2 differentially regulates G protein signalling.

The prostanoid G protein-coupled receptor (GPCR) EP2 is widely expressed and implicated in endometriosis, osteoporosis, obesity, pre-term labour and cancer. Internalisation and intracellular trafficking are critical for shaping GPCR activity, yet little is known regarding the spatial programming of EP2 signalling and whether this can be exploited pharmacologically. Using three EP2-selective ligands that favour activation of different EP2 pathways, we show that EP2 undergoes limited agonist-driven internalisation but is constitutively internalised via dynamin-dependent, ß-arrestin-independent pathways. EP2 was constitutively trafficked to early and very early endosomes (VEE), which was not altered by ligand activation. APPL1, a key adaptor and regulatory protein of the VEE, did not impact EP2 agonist-mediated cAMP. Internalisation was required for ~70% of the acute butaprost- and AH13205-mediated cAMP signalling, yet PGN9856i, a Gαs-biased agonist, was less dependent on receptor internalisation for its cAMP signalling, particularly in human term pregnant myometrial cells that endogenously express EP2. Inhibition of EP2 internalisation partially reduced calcium signalling activated by butaprost or AH13205 and had no effect on PGE2 secretion. This indicates an agonist-dependent differential spatial requirement for Gαs and Gαq/11 signalling and a role for plasma membrane-initiated Gαq/11-Ca2+-mediated PGE2 secretion. These findings reveal a key role for EP2 constitutive internalisation in its signalling and potential spatial bias in mediating its downstream functions. This, in turn, could highlight important considerations for future selective targeting of EP2 signalling pathways.

Navigating the post-operative nose as a primary care physician.

BACKGROUND: Rhinology procedures represent a wide and diverse category of procedures, which, on the surface, can appear very similar. Thus, it is difficult to navigate postoperative presentations in the general practice setting. OBJECTIVE: This article provides a comprehensive guide to managing acute presentations that might arise in primary care settings following common rhinology surgeries. It outlines common complications, their potential aetiology and first-aid measures that might be employed to temporise patients prior to escalation. It also provides a guide as to potential red flag symptomatology, and when and how to escalate specific presentations. DISCUSSION: Most acute postoperative complications can be effectively managed, or at the very least temporised, in the community setting. When in doubt, discuss with your local otolaryngologist or nearest centre with otolaryngology cover.

Combined upper lid skin crease and endoscopic approach to frontal sinus mucocoeles.

PURPOSE: This case series describes the ophthalmic manifestations of frontal sinus mucoceles and reports the long-term surgical outcomes of a combined endoscopic and upper-lid skin crease drainage approach carried out jointly with otorhinolaryngology. METHODS: We present a retrospective case review of 18 orbits and 15 patients presenting with frontal sinus mucocoeles, all of whom underwent drainage via an adapted anterior orbitotomy approach between January 2015 and July 2023. Data collection included preoperative and postoperative examination findings (visual acuity, extraocular motility, lid retraction, and lagophthalmos), mucocoele recurrence, cosmetic satisfaction, and surgical complications. Patients were followed up for an average of 22 months. RESULTS: All patients underwent successful frontal mucocoele drainage via a modified anterior orbitotomy and simultaneous endonasal approach. At presentation, three (20%) had extraocular restriction leading to diplopia, and six (40%) had proptosis in the eye adjacent to the mucocoele. One patient presented acutely with no light perception in the affected eye due to compressive optic neuropathy. All patients who had reduced extraocular motility before surgery regained full motility post-operatively. Treatment was successful in all cases, and there was no documented mucocoele recurrence during follow-up. Satisfactory aesthetic outcomes were achieved in all cases. Reported complications included temporary forehead numbness and ptosis of the affected eyelid, which resolved without intervention. CONCLUSION: The modified anterior orbitotomy approach to frontal mucocoeles allows optimal frontal sinus access and mucocoele treatment while preserving cosmesis.

A multidisciplinary approach to frontal sinus mucocoeles using an upper lid skin crease incision combined with endoscopic drainage allows full access to the frontal sinus and treatment of the mucocoele while preserving cosmesis.

Post-viral olfactory loss and parosmia.

The emergence of SARS-CoV-2 has brought olfactory dysfunction to the forefront of public awareness, because up to half of infected individuals could develop olfactory dysfunction. Loss of smell-which can be partial or total-in itself is debilitating, but the distortion of sense of smell (parosmia) that can occur as a consequence of a viral upper respiratory tract infection (either alongside a reduction in sense of smell or as a solo symptom) can be very distressing for patients. Incidence of olfactory loss after SARS-CoV-2 infection has been estimated by meta-analysis to be around 50%, with more than one in three who will subsequently report parosmia. While early loss of sense of smell is thought to be due to infection of the supporting cells of the olfactory epithelium, the underlying mechanisms of persistant loss and parosmia remain less clear. Depletion of olfactory sensory neurones, chronic inflammatory infiltrates, and downregulation of receptor expression are thought to contribute. There are few effective therapeutic options, so support and olfactory training are essential. Further research is required before strong recommendations can be made to support treatment with steroids, supplements, or interventions applied topically or injected into the olfactory epithelium in terms of improving recovery of quantitative olfactory function. It is not yet known whether these treatments will also achieve comparable improvements in parosmia. This article aims to contextualise parosmia in the setting of post-viral olfactory dysfunction, explore some of the putative molecular mechanisms, and review some of the treatment options available.

A universal surrogate matrix assay for urea measurement in clinical pharmacokinetic studies of respiratory diseases.

In pharmacokinetic studies for respiratory diseases, urea is a commonly used dilution marker for volume normalization of various biological matrices, owing to the fact that urea diffuses freely throughout the body and is minimally affected by disease states. In this study, we developed a convenient liquid chromatography-tandem mass spectrometry (LC-MS/MS) surrogate matrix assay for accurate urea quantitation in plasma, serum and epithelial lining fluid. Different mass spectrometer platforms and ionization modes were compared in parallel. The LC method and mass spectrometer parameters were comprehensively optimized to reduce interferences, to smooth the baseline and to maximize the signal-to-noise ratio. Saline was selected as the surrogate matrix, and its suitability was confirmed by good parallelism and accurate quality control sample measurements. Reliable and robust assay performance was demonstrated by precision and accuracy, dilution integrity, sensitivity, recovery and stability, all of which met bioanalysis requirements to support clinical studies. The assay performance was also verified and better understood by comparing it with a colorimetric assay and to a surrogate analyte assay. The newly developed surrogate matrix assay has the potential to be further expanded for urea quantitation in numerous physiological matrices.

Functional rewiring of G protein-coupled receptor signaling in human labor.

Current strategies to manage preterm labor center around inhibition of uterine myometrial contractions, yet do not improve neonatal outcomes as they do not address activation of inflammation. Here, we identify that during human labor, activated oxytocin receptor (OTR) reprograms the prostaglandin E2 receptor, EP2, in the pregnant myometrium to suppress relaxatory/Gαs-cAMP signaling and promote pro-labor/inflammatory responses via altered coupling of EP2 from Gαq/11 to Gαi/o. The ability of EP2 to signal via Gαi/o is recapitulated with in vitro OT and only following OTR activation, suggesting direct EP2-OTR crosstalk. Super-resolution imaging with computational modeling reveals OT-dependent reorganization of EP2-OTR complexes to favor conformations for Gαi over Gαs activation. A selective EP2 ligand, PGN9856i, activates the relaxatory/Gαs-cAMP pathway but not the pro-labor/inflammatory responses in term-pregnant myometrium, even following OT. Our study reveals a mechanism, and provides a potential therapeutic solution, whereby EP2-OTR functional associations could be exploited to delay preterm labor.

Immunotherapy as a treatment modality for mucosal melanoma of the head and neck: A systematic review.

INTRODUCTION: Mucosal melanoma (MM) is a rare disease, accounting for approximately 1.4% of all melanomas and only 0.03% of all new cancer diagnoses. Traditionally, it has been associated with a poor prognosis, with an overall 5-year survival rate of <25%. Progress in treatment has been hindered by its rarity and lack of evidence. However, studies on the treatment of subcutaneous melanoma with immunotherapy have demonstrated significant improvement in survival rates and have become a core part of oncological strategies. This paper discusses the revision of the evidence for the use of immunotherapy in the head and neck. METHODS: This systematic review was conducted on January 19, 2019. The Medline and Embase databases were searched. In total, 509 articles were collated and screened. Inclusion criteria for the study included treatment-naive cohorts, cohorts with recurrent disease, primary outcomes with overall survival and disease-free survival at 5 years and at the longest follow-up, and studies of adults with MM in whom immunotherapy was reported as a treatment strategy. The exclusion criteria included duplicate papers, anatomical sites other than the head and neck, case reports, and those not published in English. RESULTS: Fifty-two papers out of the 509 collated papers met the inclusion criteria. The results are shown as a comparison of yearly survival rates following different treatment modalities (immunotherapy vs nonimmunotherapy) at 2, 3, and 5 years. It was found that, with immunotherapy, survival rates at all intervals were higher than those without immunotherapy. DISCUSSION: Immunotherapy outcomes in small studies have shown good data for increasing survival rates at yearly intervals in MM of the head and neck. Larger clinical trials are needed to accurately distinguish the efficacy and survival outcomes of immunotherapy when compared with treatment modalities, excluding immunotherapy. However, the ability to perform larger trials is limited by the rarity of MM of the head and neck.

Volumetric absorptive microsampling-LC-MS/MS assays for quantitation of giredestrant in dried human whole blood.

Volumetric absorption microsampling devices offer minimally invasive and user-friendly collection of capillary blood in volumes as low as 10 µl. Herein we describe the assay validation for determination of the selective estrogen receptor degrader giredestrant (GDC-9545) in dried human whole blood collected using the Mitra® and Tasso-M20 devices. Both LC-MS/MS assays met validation acceptance criteria for the linear range 1-1000 ng/ml giredestrant. Mitra and Tasso-M20 samples were stable for 84 and 28 days at ambient conditions, respectively, and for 7-9 days at 40 and -70°C. Blood hematocrit, hyperlipidemia and anticoagulant did not impact quantitation of giredestrant. These validated assays are suitable for the determination of giredestrant in dried blood samples collected using Mitra and Tasso-M20 microsampling devices.

Evolution and genomic organization of the insect sHSP gene cluster and coordinate regulation in phenotypic plasticity.

BACKGROUND: Conserved syntenic gene complexes are rare in Arthropods and likely only retained due to functional constraint. Numerous sHSPs have been identified in the genomes of insects, some of which are located clustered in close proximity. Previous phylogenetic analyses of these clustered sHSP have been limited to a small number of holometabolous insect species and have not determined the pattern of evolution of the clustered sHSP genes (sHSP-C) in insect or Arthropod lineages. RESULTS: Using eight genomes from representative insect orders and three non-insect arthropod genomes we have identified that a syntenic cluster of sHSPs (sHSP-C) is a hallmark of most Arthropod genomes. Using 11 genomes from Hymenopteran species our phylogenetic analyses have refined the evolution of the sHSP-C in Hymenoptera and found that the sHSP-C is order-specific with evidence of birth-and-death evolution in the hymenopteran lineage. Finally we have shown that the honeybee sHSP-C is co-ordinately expressed and is marked by genomic features, including H3K27me3 histone marks consistent with coordinate regulation, during honeybee ovary activation. CONCLUSIONS: The syntenic sHSP-C is present in most insect genomes, and its conserved coordinate expression and regulation implies that it is an integral genomic component of environmental response in arthropods.

British Rhinological Society Consensus Guidance on the use of biological therapies for chronic rhinosinusitis with nasal polyps.

OBJECTIVES: We set out to create Consensus Guidelines, based on current evidence and relative risks of adverse effects and the costs of different treatments, which reflect the views of the British Rhinological Society (BRS) Council on where the use of biologics should be positioned within treatment pathways for CRSwNP, specifically in the setting of the National Health Service (NHS). DESIGN: An expert panel of 16 members was assembled. A review of the literature and evidence synthesis was undertaken and circulated to the panel. We used the RAND/UCLA methodology with a multi-step process to make recommendations on the use of biologics. SETTING: N/A. PARTICIPANTS: N/A. RESULTS: Recommendations were made, based on underlying disease severity, prior treatments and co-morbidities. A group of patients for whom biologics were considered an appropriate treatment option for CRSwNP was defined. CONCLUSIONS: Although biologics are not currently available for the treatment of CRSwNP, the BRS Council have defined a group of patients who have higher rates of "failure" with current treatment pathways, higher resource use and are more likely to suffer with uncontrolled symptoms. We would urge NICE to consider approval of biologics for such indications without applying further restrictions on use.

Robust and Comprehensive Targeted Metabolomics Method for Quantification of 50 Different Primary, Secondary, and Sulfated Bile Acids in Multiple Biological Species (Human, Monkey, Rabbit, Dog, and Rat) and Matrices (Plasma and Urine) Using Liquid Chromatography High Resolution Mass Spectrometry (LC-HRMS) Analysis.

Bile acids (BAs) are biomolecules synthesized in the liver from cholesterol and are constituents of bile. The in-vivo BA pool includes more than 50 known diverse BAs which are unconjugated, amino acid conjugated, sulfated, and glucuronidated metabolites. Hemostasis of bile acids is known to be highly regulated and an interplay between liver metabolism, gut microbiome function, intestinal absorption, and enterohepatic recirculation. Interruption of BA homeostasis has been attributed to several metabolic diseases and drug induced liver injury (DILI), and their use as potential biomarkers is increasingly becoming important. Speciated quantitative and comprehensive profiling of BAs in various biomatrices from humans and preclinical animal species are important to understand their significance and biological function. Consequently, a versatile one single bioanalytical method for BAs is required to accommodate quantitation in a broad range of biomatrices from human and preclinical animal species. Here we report a versatile, comprehensive, and high throughput liquid chromatography-high resolution mass spectrometry (LC-HRMS) targeted metabolomics method for quantitative analysis of 50 different BAs in multiple matrices including human serum, plasma, and urine and plasma and urine of preclinical animal species (rat, rabbit, dog, and monkey). The method has been sufficiently qualified for accuracy, precision, robustness, and ruggedness and addresses the issue of nonspecific binding of bile acids to plastic for urine samples. Application of this method includes comparison for BA analysis between matched plasma and serum samples, human and animal species differences in BA pools, data analysis, and visualization of complex BA data using BA indices or ratios to understand BA biology, metabolism, and transport.

Tackling nasal symptoms in athletes: Moving towards personalized medicine.

Adequate nasal breathing is indispensable for athletes, and nasal symptoms have been shown to interfere with their subjective feeling of comfortable breathing and quality of life. Nasal symptoms are caused by either structural abnormalities or mucosal pathology. Structural pathologies are managed differently from mucosal disease, and therefore, adequate diagnosis is of utmost importance in athletes in order to choose the correct treatment option for the individual. Literature suggests that nasal symptoms are more prevalent in athletes compared to the general population and certain sports environments might even trigger the development of symptoms. Given the high demands of respiratory function in athletes, insight into triggering factors is of high importance for disease prevention. Also, it has been suggested that athletes are more neglectful to their symptoms and hence remain undertreated, meaning that special attention should be paid to education of athletes and their caregivers. This review aims at giving an overview of nasal physiology in exercise as well as the possible types of nasal pathology. Additionally, diagnostic and treatment options are discussed and we focus on unmet needs for the management and prevention of these symptoms in athletes within the concept of precision medicine.

Tracheostomy care and decannulation during the COVID-19 pandemic. A multidisciplinary clinical practice guideline.

PURPOSE: Traditional critical care dogma regarding the benefits of early tracheostomy during invasive ventilation has had to be revisited due to the risk of COVID-19 to patients and healthcare staff. Standard practises that have evolved to minimise the risks associated with tracheostomy must be comprehensively reviewed in light of the numerous potential episodes for aerosol generating procedures. We meet the urgent need for safe practise standards by presenting the experience of two major London teaching hospitals, and synthesise our findings into an evidence-based guideline for multidisciplinary care of the tracheostomy patient. METHODS: This is a narrative review presenting the extensive experience of over 120 patients with tracheostomy, with a pragmatic analysis of currently available evidence for safe tracheostomy care in COVID-19 patients. RESULTS: Tracheostomy care involves many potentially aerosol generating procedures which may pose a risk of viral transmission to staff and patients. We make a series of recommendations to ameliorate this risk through infection control strategies, equipment modification, and individualised decannulation protocols. In addition, we discuss the multidisciplinary collaboration that is absolutely fundamental to safe and effective practise. CONCLUSION: COVID-19 requires a radical rethink of many tenets of tracheostomy care, and controversy continues to exist regarding the optimal techniques to minimise risk to patients and healthcare workers. Safe practise requires a coordinated multidisciplinary team approach to infection control, weaning and decannulation, with integrated processes for continuous prospective data collection and audit.

Royal Society of Medicine Surgical Innovation Day Adrian Tanner Prize Winner 2019: Multidisciplinary care and surgical innovation for the benefit of the patient with head and neck cancer of unknown primary.

Head and neck carcinoma of unknown primary accounts for approximately 1-5% of all head and neck cancers and presents a genuine diagnostic and therapeutic dilemma. Despite advanced investigations, the primary tumour location remains unknown in up to 40% of these cases. Transoral robotic surgery presents a viable diagnostic and therapeutic option in these patients. This surgical innovation alongside advances in the understanding of head and neck cancer biology means that a multidisciplinary approach in the management of these complex patients is of utmost importance to ensure optimal therapeutic outcomes.

Defining appropriateness criteria for endoscopic sinus surgery in the management of adult dental implant patients with incidental maxillary sinus findings on conebeam computed tomography.

OBJECTIVES: Conebeam computed tomography (CBCT) imaging is commonly requested by dental implant surgeons, preoperatively, for patients being considered for dental implants. Incidental maxillary sinus findings often result in otolaryngology (ENT) referral for further assessment. CBCT findings include transient and benign mucosal changes that may not require any intervention and therefore unnecessarily delay implant surgery. We aim to define appropriateness criteria for ESS in the management of adult dental implant patients with incidental maxillary sinus findings on CBCT and provide guidance to both dental implant and ENT surgeons. DESIGN: The RAND/UCLA appropriateness methodology was used to develop and define the appropriateness criteria. SETTING: A virtual panel of 13 international experts in ESS. PARTICIPANTS: The expert panel completed two rounds of a modified Delphi ranking process for nine clinical scenarios, considering various factors affecting decision-making processes. MAIN OUTCOME MEASURES: To define appropriateness criteria for ESS in adult dental implant patients who have incidental maxillary sinus findings on CBCT. RESULTS: Patients with clinical symptoms and endoscopic findings of chronic rhinosinusitis together with an obstructed ostiomeatal complex (OMC) and concentric mucosal thickening of the ipsilateral maxillary sinus or pansinusitis were deemed appropriate candidates for ESS prior to their dental implant. ESS was not appropriate in asymptomatic patients with a patent OMC and mucosal thickening isolated to floor of the ipsilateral maxillary sinus. For uncertain scenarios, further discussion between dental implant and ENT surgeon should be considered. CONCLUSIONS: This study has developed and reported a list of appropriateness criteria to offer ESS in adult dental implant patients with incidental maxillary sinus findings on CBCT.

Recommendation of a practical guideline for safe tracheostomy during the COVID-19 pandemic.

PURPOSE: The COVID-19 pandemic is placing unprecedented demand upon critical care services for invasive mechanical ventilation. There is current uncertainty regarding the role of tracheostomy for weaning ventilated patients with COVID-19 pneumonia. This is due to a number of factors including prognosis, optimal healthcare resource utilisation, and safety of healthcare workers when performing such a high-risk aerosol-generating procedure. METHODS: Literature review and proposed practical guideline based on the experience of a tertiary healthcare institution with 195 critical care admissions for COVID-19 up until 4th April 2020. RESULTS: A synthesis of the current international literature and reported experience is presented with respect to prognosis, viral load and staff safety, thus leading to a pragmatic recommendation that tracheostomy is not performed until at least 14 days after endotracheal intubation in COVID-19 patients. Practical steps to minimise aerosol generation in percutaneous tracheostomy are outlined and we describe the process and framework for setting up a dedicated tracheostomy team. CONCLUSION: In selected COVID-19 patients, there is a role for tracheostomy to aid in weaning and optimise healthcare resource utilisation. Both percutaneous and open techniques can be performed safely with careful modifications to technique and appropriate enhanced personal protective equipment. ORL-HNS surgeons can play a valuable role in forming tracheostomy teams to support critical care teams during this global pandemic.