Virtual voice clinics in the COVID-19 era: have they been helpful?

PURPOSE: In response to the coronavirus pandemic, a tertiary combined Laryngology-Speech Therapy voice clinic was converted to a wholly virtual clinic, with consultations carried out via telephone or video. The aim of our mixed method study was to assess (a) how effective are virtual clinics vs face-to-face clinics in progressing patients' care and (b) what is patient satisfaction with virtual consultation methods. METHODS: Analysis of clinic data from patient databases for both virtual and face-to-face clinics was carried out. A patient satisfaction survey was carried out by 75 of the patients who had attended virtual clinics. RESULTS: There was statistically a significant difference (p value < 0.01) in the proportion of patients prescribed medical therapy, referred for Speech and Language Therapy (SALT) or listed for surgery in the virtual clinic by comparison to the face-to-face clinic. 75 patients completed the questionnaire. 98% of patients were satisfied overall with the virtual method of consultation. 84% believed they would still benefit from face-to-face review. 83% would like the option of a virtual type of clinic in the future. CONCLUSION: Our data clearly demonstrates that face-to-face clinics are superior to virtual clinics, with almost no patients progressed to surgery in virtual consultations. Despite this, virtual methods are still valuable, and many patients have meaningful progression of care. In current circumstances, patients have very high satisfaction with virtual consultations and certain groups have been identified as particularly benefiting. Going forward, an ideal clinic may be a hybrid of face-to-face and virtual appointments as clinically indicated.

Prime editing functionally corrects cystic fibrosis-causing CFTR mutations in human organoids and airway epithelial cells.

Prime editing is a recent, CRISPR-derived genome editing technology capable of introducing precise nucleotide substitutions, insertions, and deletions. Here, we present prime editing approaches to correct L227R- and N1303K-CFTR, two mutations that cause cystic fibrosis and are not eligible for current market-approved modulator therapies. We show that, upon DNA correction of the CFTR gene, the complex glycosylation, localization, and, most importantly, function of the CFTR protein are restored in HEK293T and 16HBE cell lines. These findings were subsequently validated in patient-derived rectal organoids and human nasal epithelial cells. Through analysis of predicted and experimentally identified candidate off-target sites in primary stem cells, we confirm previous reports on the high prime editor (PE) specificity and its potential for a curative CF gene editing therapy. To facilitate future screening of genetic strategies in a translational CF model, a machine learning algorithm was developed for dynamic quantification of CFTR function in organoids (DETECTOR: "detection of targeted editing of CFTR in organoids").

Mitochondrial dynamics and metabolic regulation control T cell fate in the thymus.

Several studies demonstrated that mitochondrial dynamics and metabolic pathways control T cell fate in the periphery. However, little is known about their implication in thymocyte development. Our results showed that thymic progenitors (CD3-CD4-CD8- triple negative, TN), in active division, have essentially a fused mitochondrial morphology and rely on high glycolysis and mitochondrial oxidative phosphorylation (OXPHOS). As TN cells differentiate to double positive (DP, CD4+CD8+) and single positive (SP, CD4+ and CD8+) stages, they became more quiescent, their mitochondria fragment and they downregulate glycolysis and OXPHOS. Accordingly, in vitro inhibition of the mitochondrial fission during progenitor differentiation on OP9-DL4 stroma, affected the TN to DP thymocyte transition by enhancing the percentage of TN and reducing that of DP, leading to a decrease in the total number of thymic cells including SP T cells. We demonstrated that the stage 3 triple negative pre-T (TN3) and the stage 4 triple negative pre-T (TN4) have different metabolic and functional behaviors. While their mitochondrial morphologies are both essentially fused, the LC-MS based analysis of their metabolome showed that they are distinct: TN3 rely more on OXPHOS whereas TN4 are more glycolytic. In line with this, TN4 display an increased Hexokinase II expression in comparison to TN3, associated with high proliferation and glycolysis. The in vivo inhibition of glycolysis using 2-deoxyglucose (2-DG) and the absence of IL-7 signaling, led to a decline in glucose metabolism and mitochondrial membrane potential. In addition, the glucose/IL-7R connection affects the TN3 to TN4 transition (also called ß-selection transition), by enhancing the percentage of TN3, leading to a decrease in the total number of thymocytes. Thus, we identified additional components, essential during ß-selection transition and playing a major role in thymic development.

mTOR inhibition suppresses salinomycin-induced ferroptosis in breast cancer stem cells by ironing out mitochondrial dysfunctions.

Ferroptosis constitutes a promising therapeutic strategy against cancer by efficiently targeting the highly tumorigenic and treatment-resistant cancer stem cells (CSCs). We previously showed that the lysosomal iron-targeting drug Salinomycin (Sal) was able to eliminate CSCs by triggering ferroptosis. Here, in a well-established breast CSCs model (human mammary epithelial HMLER CD24low/CD44high), we identified that pharmacological inhibition of the mechanistic target of rapamycin (mTOR), suppresses Sal-induced ferroptosis. Mechanistically, mTOR inhibition modulates iron cellular flux and thereby limits iron-mediated oxidative stress. Furthermore, integration of multi-omics data identified mitochondria as a key target of Sal action, leading to profound functional and structural alteration prevented by mTOR inhibition. On top of that, we found that Sal-induced metabolic plasticity is mainly dependent on the mTOR pathway. Overall, our findings provide experimental evidence for the mechanisms of mTOR as a crucial effector of Sal-induced ferroptosis pointing not only that metabolic reprogramming regulates ferroptosis, but also providing proof-of-concept that careful evaluation of such combination therapy (here mTOR and ferroptosis co-targeting) is required in the development of an effective treatment.

Primary Human Nasal Epithelial Cells: Biobanking in the Context of Precision Medicine.

Human nasal epithelial (HNE) cells are easy to collect by simple, non-invasive nasal brushing. Patient-derived primary HNE cells can be amplified and differentiated into a pseudo-stratified epithelium in air-liquid interface conditions to quantify cyclic AMP-mediated Chloride (Cl-) transport as an index of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) function. If critical steps such as quality of nasal brushing and cell density upon cryopreservation are performed efficiently, HNE cells can be successfully biobanked. Moreover, short-circuit current studies demonstrate that freeze-thawing does not significantly modify HNE cells' electrophysiological properties and response to CFTR modulators. In the culture conditions used in this study, when less than 2 x 106 cells are frozen per cryovial, the failure rate is very high. We recommend freezing at least 3 x 106 cells per cryovial. We show that dual therapies combining a CFTR corrector with a CFTR potentiator have a comparable correction efficacy for CFTR activity in F508del-homozygous HNE cells. Triple therapy VX-445 + VX-661 + VX-770 significantly increased correction of CFTR activity compared to dual therapy VX-809 + VX-770. The measure of CFTR activity in HNE cells is a promising pre-clinical biomarker useful to guide CFTR modulator therapy.

Coblation debulking of a paediatric laryngeal plexiform neurofibroma: a pragmatic response to a rare tumour.

Laryngeal neurofibroma is a rare but important differential diagnosis in a patient presenting with stridor. In paediatric patients, these lesions present a management conundrum: complete surgical resection is the established treatment of choice, but an aggressive approach can be detrimental to developing anatomy. We report the case of a plexiform neurofibroma affecting the right hemilarynx of a 3-year-old boy. Endoscopy revealed a large tumour, involving the right aryepiglottic fold and extending into the piriform sinus, ventricle and the false cord. Given the patient's young age and the challenging tumour location, the lesion was debulked, rather than resected, using coblation (low-temperature plasma radiofrequency ablation). At 30 months follow-up, the neurofibroma has mildly increased in size-in line with expectations that these lesions exhibit slow growth throughout childhood-but there are no significant respiratory symptoms and there is no functional impairment.

Sphenoid mucopyocele causing bilateral sixth nerve palsy.

Sphenoid mucoceles, although rare, should be considered in patients with headache, visual disorders and eye paralysis. Due to close relationships with the orbit and neuromeningeal structures, early recognition is vital. We report the case of a patient who presented with bilateral abducens nerve palsies. At surgery, she was found to have a mucopyocele; this was drained and she required prolonged intravenous antibiotic therapy due to ongoing symptoms and persistent dural enhancement on imaging. A lesion of sufficient size in the clival area has the potential to cause bilateral abducens nerve palsies, though we believe this is the first time it has been described in relation to a sphenoid mucocele. Imaging plays a crucial role in diagnosis, and prompt surgical intervention is essential to avoid serious and permanent complications. The multi-disciplinary team approach is vital-these cases requiring input from ophthalmology, ear nose and throat, microbiology, radiology, neurology and neurosurgery.

Profiling the response to lumacaftor-ivacaftor in children with cystic between fibrosis and new insight from a French-Italian real-life cohort.

INTRODUCTION: Clinical trials for CFTR modulators consider mean changes of clinical status at the cohort level, and thus fail to assess the heterogeneity of the response. We aimed to study the different response profiles to lumacaftor-ivacaftor according to age in children with cystic fibrosis (CF). METHODS: A mathematical framework, including principal component analysis, data clustering, and data completion, was applied to a multicenter cohort of 112 children aged 6-18 years, treated with lumacaftor-ivacaftor. Studied parameters at baseline and 6 months included body mass index (BMI), number of days of antibiotics (ATB), Sweat test (ST), forced expiratory volume in 1 s expressed in percentage predicted (ppFEV1 ), forced vital capacity (ppFVC), and forced expiratory flow at 25%-75% of FVC (ppFEF25-75 ). RESULTS: Change in ppFEV1 was the most significant parameter in characterizing response heterogeneity among the 12-18-year-old patients. Patients with minimal changes in ppFEV1 were further separated by change in BMI and ATB course. In the 6-12-year-old children both BMI and ppFEV1 evolution were the most relevant. ST change was not associated with a clinical response. CONCLUSIONS: Change in ppFEV1 , BMI, and ATB course are the most relevant outcomes to discriminate clinical response profiles in children treated with lumacaftor-ivacaftor. Prepubertal and pubertal children display different response profiles.

Perioperative management of cranial diabetes insipidus in a patient requiring a tracheostomy.

Cranial diabetes insipidus (DI), which can cause life-threatening dehydration, is treated with desmopressin, often intranasally. This is challenging in patients whose nasal airflow is altered, such as those requiring tracheostomy. We report the case of a patient, taking intranasal desmopressin for cranial DI, who underwent partial glossectomy, free-flap reconstruction and tracheostomy. Postoperatively, she could not administer nasal desmopressin due to reduced nasal airflow. She developed uncontrollable thirst, polyuria and hypernatraemia. Symptoms were relieved by switching to an enteric formulation. A literature review showed no cases of patients with DI encountering difficulties following tracheostomy. The Royal Society of Endocrinology recommends perioperative planning for such patients, but gives no specific guidance on medication delivery in the context of altered airway anatomy. Careful perioperative planning is required for head and neck patients with DI, particularly for those undergoing airway alteration that may necessitate a change in the mode of delivery of critical medications.

Primary aspergillosis of the larynx causing acute airway distress.

Laryngeal aspergillosis is most commonly seen as a secondary infection that spreads from the lungs and tracheobronchial tree. Primary invasive aspergillosis of the larynx is rare and most likely seen in an immunocompromised patient. We present a case of a 59-year-old woman who presented with progressive dysphonia and subsequently acute stridor. She is a non-smoker with a recent diagnosis of acute myeloid leukaemia. Fibreoptic nasendoscopy revealed a left sided vocal cord lesion ball valving into the glottic space. Histology taken during emergency debulking confirmed Aspergillus fumigatus and the patient was successfully treated with intravenous antifungals. Although there are cases of primary laryngeal aspergillosis discussed in the literature, to the best of our knowledge this is the first reported case to have caused acute airway distress requiring emergency intervention.

Surgical management of an odontogenic cutaneous fistula.

A patient presented to our unit with a long history of a discharging skin infection on his left cheek, which came and went. He had been seen by numerous healthcare practitioners including his general practitioner, general dental practitioner and dermatologist, with no resolution. He was eventually diagnosed with an odontogenic cutaneous fistula (OCF), for which he underwent surgical management. The purpose of the study is to describe the diagnosis and surgical management of an OCF, from initial assessment through to postoperative review and discharge. Following surgical management of the OCF and treatment of the source of infection by dental extraction, the patient is no longer experiencing purulent discharge through his left cheek. The extraoral skin site of drainage at his left cheek has resolved completely, with minimal residual scarring. OCF can be managed by a number of different treatment modalities. The treatment of an OCF by surgical excision is presented.

Unilateral nasal obstruction: a rare presentation of acute myeloid leukaemia.

Myeloid sarcoma, and, with it, Acute Myeloid Leukaemia (AML), is a rare but important differential diagnosis in the consideration of unilateral nasal blockage. These lesions are often misdiagnosed as lymphoma or poorly differentiated carcinoma. We report the case of a patient with unilateral nasal blockage who underwent Endoscopic Sinus Surgery and biopsy. Histology revealed myeloid sarcoma and she was diagnosed with AML. Genetic testing could not be fully undertaken as the biopsy samples were preserved in formalin, which can degrade the quality of the DNA required for the more sensitive fms-like tyrosine kinase 3-internal tandem duplication (FLT3 ITD) test. Given that these levels have a significant impact on treatment decisions, a further biopsy, preserved in saline, was required. This case exemplifies the need for Ear, Nose and Throat clinicians to have a high index of suspicion for this lesion, and a working knowledge of the testing requirements for samples taken.

Cystic fibrosis transmembrane regulator inhibitors CFTR(inh)-172 and GlyH-101 target mitochondrial functions, independently of chloride channel inhibition.

Two highly potent and selective cystic fibrosis (CF) transmembrane regulator (CFTR) inhibitors have been identified by high-throughput screening: the thiazolidinone CFTR(inh)-172 [3-[(3-trifluoromethyl)phenyl]-5-[(4-carboxyphenyl)methylene]- 2-thioxo-4-thiazolidinone] and the glycine hydrazide GlyH-101 [N-(2-naphthalenyl)-((3,5-dibromo-2,4-dihydroxyphenyl)methylene)glycine hydrazide]. Inhibition of the CFTR chloride channel by these compounds has been suggested to be of pharmacological interest in the treatment of secretory diarrheas and polycystic kidney disease. In addition, functional inhibition of CFTR by CFTR(inh)-172 has been proposed to be sufficient to mimic the CF inflammatory profile. In the present study, we investigated the effects of the two compounds on reactive oxygen species (ROS) production and mitochondrial membrane potential in several cell lines: the CFTR-deficient human lung epithelial IB3-1 (expressing the heterozygous F508del/W1282X mutation), the isogenic CFTR-corrected C38, and HeLa and A549 as non-CFTR-expressing controls. Both inhibitors were able to induce a rapid increase in ROS levels and depolarize mitochondria in the four cell types, suggesting that these effects are independent of CFTR inhibition. In HeLa cells, these events were associated with a decrease in the rate of oxygen consumption, with GlyH-101 demonstrating a higher potency than CFTR(inh)-172. The impact of CFTR inhibitors on inflammatory parameters was also tested in HeLa cells. CFTR(inh)-172, but not GlyH-101, induced nuclear translocation of nuclear factor-kappaB (NF-kappaB). CFTR(inh)-172 slightly decreased interleukin-8 secretion, whereas GlyH-101 induced a slight increase. These results support the conclusion that CFTR inhibitors may exert nonspecific effects regarding ROS production, mitochondrial failure, and activation of the NF-kappaB signaling pathway, independently of CFTR inhibition.

Single-session, dual-site robotic surgery for synchronous primary malignancies of the oropharynx and thorax.

Single-session, dual-site robotic surgery for synchronous malignancies is not widely reported. To our knowledge, there are no previous reports of transoral robotic surgery (TORS) and robot-assisted thoracic surgery (RATS) in a single sitting. A 49-year-old male presented with a neck lump. Biopsies and imaging proved synchronous primaries of the tonsil and lung. The morbidity of primary dual-site chemoradiotherapy, or open surgery, presented a management challenge. We therefore opted for sequential robotic resections, in a single sitting. The patient was discharged on Day 6 post-operatively and was able to start radiotherapy less than 3 weeks post-operatively. Undertaking TORS and RATS in a single sitting is feasible and safe. This approach allowed fast-tracking of adjuvant oncological therapy, arguably conveying the greatest chance of cure. We detail the rationale and utility of this novel approach and describe the surgical and anaesthetic challenges of two teams undertaking sequential robotic procedures in a single sitting.

Dibromidodimethyl-dipyridine-platinum(IV).

In the title complex, [PtBr(2)(CH(3))(2)(C(5)H(5)N)(2)], the Pt(IV) metal centre lies on a twofold rotation axis and adopts a slightly distorted octa-hedral coordination geometry. The structure displays weak intra-molecular C-H⋯Br hydrogen-bonding inter-actions.

Treatment of trivalvular rheumatic heart disease: why it matters where we live.

A 42-year-old woman who had recently emigrated to the UK from Ethiopia presented to her general practitioner with increasing breathlessness and palpitations. Transoesophageal echocardiogram revealed rheumatic heart disease affecting three valves, and severe aortic stenosis. ECG demonstrated atrial fibrillation. She was treated operatively with mitral and aortic valve replacement and tricuspid valve repair and put on lifelong warfarin. Four months later she remains well, with substantially increased exercise tolerance. This case report summarises treatment options for severe rheumatic heart disease and discusses differences in prevention and management between patients presenting in the UK and in the developing world.

Long-term CFTR inhibition modulates 15d-prostaglandin J2 in human pulmonary cells.

Prostaglandins, the products of arachidonic acid release and oxidation by phospholipase A(2) and cyclooxygenases (COX) 1 and 2 respectively, are known as important inflammation mediators. However, their diversity in structure, properties and cell specificity make their physiological function difficult to define. In the lung, the prostaglandin D(2) (PGD(2)) metabolite 15d-PGJ(2) is known to modulate the properties of a large number of intracellular compounds, leading to both pro- and anti-inflammatory effects. In the lung, the serous sub-mucosal glands, that strongly express CFTR (cystic fibrosis transmembrane conductance regulator), play an important role in the defence against inflammation, and their derivatives Calu-3 cells are largely used in in vitro experiments. The present study was undertaken to determine whether the PGD synthase-PGD(2)-15d-PGJ(2) pathway is active in Calu-3 cells, and whether its activity requires a functional CFTR. Both cellular and released PGD(2) and 15d-PGJ(2) were measured in cells treated with CFTR inhibitors and stimulated or not with inflammatory IL-1ß. Pretreatment with either CFTR(inh172) or GlyH101 inhibitors decreased the basal cell content of both prostaglandins, and so did acute stimulation with IL-1ß, but the latter was dramatically reversed in CFTR(inh172)-treated cells. CFTR(inh172) also altered the release of inflammation mediators PGE(2) and IL-8, and this effect was blunted by exogenous 15d-PGJ(2). CFTR(inh172)-induced modulation of 15d-PGJ(2) cellular content was not detected in CFTR-silenced Calu-3 cells, but it was reproduced in pulmonary CFBE41o-cells, which express F508del-CFTR. These results show that cellular 15d-PGJ(2) production, which controls PGE(2) and IL-8 release, is disturbed by CFTR dysfunction. In Calu-3 cells, 15d-PGJ(2) production resulted from COX-2-regulated COX-1 activation, while CFTR(inh172)-induced alteration of 15d-PGJ(2) synthesis involved both decreased expression of PGD synthase and disturbed relationships between both COXs. CFTR-mediated regulation of PGD synthase-PGD(2)-15d-PGJ(2) pathway and cellular 15d-PGJ(2) effects may involve a large number of molecular reactive pathways. Their exploration should help understand the development of CF inflammation and might bring new perspectives in its treatment.

Deletion of UCP2 in iNOS deficient mice reduces the severity of the disease during experimental autoimmune encephalomyelitis.

Uncoupling protein 2 is a member of the mitochondrial anion carrier family that is widely expressed in neurons and the immune cells of humans. Deletion of Ucp2 gene in mice pre-activates the immune system leading to higher resistance toward infection and to an increased susceptibility to develop chronic inflammatory diseases as previously exemplified with the Experimental Autoimmune Encephalomyelitis (EAE), a mouse model for multiple sclerosis. Given that oxidative stress is enhanced in Ucp2-/- mice and that nitric oxide (NO) also plays a critical function in redox balance and in chronic inflammation, we generated mice deficient for both Ucp2 and iNos genes and submitted them to EAE. Mice lacking iNos gene exhibited the highest clinical score (3.4+/-0.5 p<0.05). Surprisingly, mice deficient for both genes developed milder disease with reduced immune cell infiltration, cytokines and ROS production as compared to iNos-/- mice.

Resveratrol rescues cAMP-dependent anionic transport in the cystic fibrosis pancreatic cell line CFPAC1.

BACKGROUND AND PURPOSE: The cystic fibrosis transmembrane conductance regulator (CFTR) is a cAMP-dependent chloride channel in the plasma membrane of epithelia whose mutation is the cause of the genetic disease cystic fibrosis (CF). The most frequent CFTR mutation is deletion of Phe(508) and this mutant protein (delF508CFTR) does not readily translocate to the plasma membrane and is rapidly degraded within the cell. We hypothesized that treating epithelial cells with resveratrol, a natural polyphenolic, phyto-ooestrogenic compound from grapes, could modulate both the expression and localization of CFTR. EXPERIMENTAL APPROACH: Cells endogenously expressing CFTR (MDCK1 and CAPAN1 cells) or delF508CFTR (CFPAC1 and airway epithelial cells, deriving from human bronchial biopsies) were treated with resveratrol for 2 or 18 h. The effect of this treatment on CFTR and delF508CFTR expression and localization was evaluated using RT-PCR, Western blot and immunocytochemistry. Halide efflux was measured with a fluorescent dye and with halide-sensitive electrodes. Production of interleukin-8 by these cells was assayed by ELISA. KEY RESULTS: Resveratrol treatment increased CFTR expression or maturation in immunoblotting experiments in MDCK1 cells or in CFPAC1 cells. Indirect immunofluorescence experiments showed a shift of delF508CFTR localization towards the (peri)-membrane area in CFPAC1 cells and in human airway epithelial cells. A cAMP-dependent increase in membrane permeability to halide was detected in resveratrol-treated CFPAC1 cells, and was inhibited by a selective inhibitor of CFTR. CONCLUSION AND IMPLICATIONS: These results show that resveratrol modulated CFTR expression and localization and could rescue cAMP-dependent chloride transport in delF508CFTR cells.

Glutathione S-transferases related to P. aeruginosa lung infection in cystic fibrosis children: preliminary study.

OBJECTIVES: In cystic fibrosis (CF) children, we investigated the predictive impact of glutathione S-transferases (GST) activity and genotypes P1, M1 and T1, and antioxidant levels on stage-severity of Pseudomonas aeruginosa lung infection. METHODS: GST activity was determined in whole blood by spectrophotometry, and GST genotypes by multiplex PCR RFLP for 36 CF and 9 control children. Levels of glutathione in erythrocyte and vitamins A, E and C in plasma were measured by HPLC. RESULTS: No difference in GST activity and no relationship between GST activity and antioxidant levels were observed in CF children as compared to controls. However, GST activity was lower in CF children with severe clinical status and infection, and the frequency of GSTP1 wild type genotype AA, prevalent in uninfected CF children (75%), decreased in infected ones (33%). CONCLUSION: GST activity and genotype could play an important role in modulating P. aeruginosa lung infection in CF patients.