Le rôle du lactate et du N-acétylcystéine intra-tympanique dans la prévention de l’ototoxicité secondaire au cisplatin

Objectifs Aucun agent n’a été approuvé pour prévenir l’ototoxicité secondaire au cisplatin. Nos objectifs consistaient à évaluer la protection auditive offerte par le lactate et le N-acétylcystéine (NAC) intra-tympaniques après injection de cisplatin, ainsi que l’absorption systémique du NAC intra-tympanique. Méthodes Seize cochons d’inde formaient 2 groupes ayant reçu une solution de lactate et de NAC à 20% dans l’oreille testée. L’oreille contro-latérale a reçu une solution saline contrôle. Après 30 minutes, une injection intrapéritonéale de 3 mg/kg de cisplatin a été effectuée et répétée une fois par semaine jusqu’à une dose finale de 24 mg/kg. Les potentiels évoqués auditifs du tronc cérébral (PEATC) ont été mesurés avant les injections, après 9 mg/kg et 24 mg/kg de cisplatin. Les cochlées ont été analysées au microscope électronique à balayage. La diffusion systémique du NAC a été évaluée par chromatographie en phase liquide. Résultats Pour les oreilles contrôles, les seuils auditifs des PEATC ont augmenté uniformément sur toutes les fréquences (28,4 dB en moyenne). Le groupe lactate montrait une augmentation moins importante (17,0 dB). Les basses fréquences étaient nettement moins affectées. Le groupe NAC a subi une augmentation des seuils de 89 dB. La microscopie électronique a démontré une préservation partielle des cellules ciliées externes des cochlées traitées au lactate et une destruction complète de celles traitées au NAC. La chromatographie n’a démontré aucune diffusion de NAC. Conclusions Le lactate offre une protection partielle significative contre l’ototoxicité induite par le cisplatin. Les injections de NAC n’offrent pas de protection lorsque administrées en concentrations élevée. Le NAC intra-tympanique ne se diffuse pas systémiquement.Objectives There is no approved agent to prevent cisplatin-induced ototoxicity. Our objectives are to identify and compare the protective effect of intratympanic injections of lactate or N-acetylcysteine (NAC) in the prevention of cisplatin-induced ototoxicity and to study systemic diffusion of intratympanic NAC. Methods Sixteen guinea pigs, forming two groups, received respectively intratympanic lactate and 20% NAC in one ear. The contra-lateral ears received a control saline solution. After 30 minutes, an intra-peritoneal cisplatin injection of 3 mg/kg was performed and repeated once a week to achieve a final dose of 24mg/kg. Auditory brainstem responses (ABR) were recorded before any injection, after 9mg/kg and after 24mg/kg of cisplatin for the frequencies 2, 4, 6 and 8kHz. Cochleas were analyzed under scanning electron microscope. Systemic diffusion of NAC was studied using high performance liquid chromatography. Results For the control ears, ABR thresholds increased uniformly by an average of 28.4dB. The lactate group showed a lower threshold increase by an average of 17.0dB. The NAC showed an important threshold increase of 89.0dB. Lactate showed a significant hearing protection at 2000Hz (p<0.01). Electron microscopy revealed partial preservation of cochlear outer hair cells stereocilia for the ears treated with lactate and severe disruption for NAC group. No systemic diffusion of NAC was observed with chromatography. Conclusion Lactate offers significant partial protection against cisplatin-induced ototoxicity. NAC does not offer any protection when administered in high concentrations. Intratympanic NAC does not diffuse systemically

Survival Outcomes of Patients with Mycosis Fungoides Involving the External Ear and Ear Canal

OBJECTIVES/HYPOTHESIS: Mycosis Fungoides (MF) is the most common subtype of cutaneous T-cell lymphoma. Disease involvement of specific locations may be more significant than simply the symptoms associated with that site; it is possible that involvement of certain sites could be associated with poor prognosis. We aimed to evaluate the outcomes of patients with MF with documented involvement of the EAC and external ear. STUDY DESIGN: Retrospective analysis. METHODS: We retrospectively reviewed 40 patients with MF that were treated by otologists between 2012 and 2021. RESULTS: We report the largest series of patients with MF involving the external ear and EAC. Of the 40 patients included in this study, 17 presented with Mycosis Fungoides in the otologic region (MFO). Of these 17 MFO patients, 2/17 had involvement of the external ear only, 3/17 of the EAC only, 11/17 of both the external ear and EAC, and 1/17 of the periauricular skin. Of note, 11/14 (79%) patients presenting with EAC disease died compared to11/26 (42%) of patients without involvement. In addition, eight of the 13 (62%) patients with external ear involvement died compared to 14/27 (52%) of patients without involvement. Ear canal involvement was associated with a statistically significant shorter overall survival duration in patients with MF (p = 0.03). Furthermore, disease in the EAC was found to have a hazard ratio value of 2.565 (CI 1.102-5.970). CONCLUSIONS: Involvement of the EAC by MF portends a poor prognosis. This finding highlights the need for a more in-depth otologic evaluation of patients with MF. LEVEL OF EVIDENCE: 4 Laryngoscope, 133:1486-1491, 2023

A preliminary study of phosphodiesterases and adenylyl cyclase signaling pathway on red blood cell deformability of sickle cell patients

Sickle cell disease (SCD) is an inherited hemoglobinopathy characterized by chronic anemia, intravascular hemolysis, and the occurrence of vaso-occlusive crises due to the mechanical obstruction of the microcirculation by poorly deformable red blood cells (RBCs). RBC deformability is a key factor in the pathogenesis of SCD, and is affected by various factors. In this study, we investigated the effects of adenylyl cyclase (AC) signaling pathway modulation and different phosphodiesterase (PDE) modulatory molecules on the deformability and mechanical stress responses of RBC from SCD patients (HbSS genotype) by applying 5 Pa shear stress with an ektacytometer (LORRCA). We evaluated RBC deformability before and after the application of shear stress. AC stimulation with Forskolin had distinct effects on RBC deformability depending on the application of 5 Pa shear stress. RBC deformability was increased by Forskolin before shear stress application but decreased after 5 Pa shear stress. AC inhibition with SQ22536 and protein kinase A (PKA) inhibition with H89 increased RBC deformability before and after the shear stress application. Non-selective PDE inhibition with Pentoxifylline increased RBC deformability. However, modulation of the different PDE types had distinct effects on RBC deformability, with PDE1 inhibition by Vinpocetine increasing deformability while PDE4 inhibition by Rolipram decreased RBC deformability after the shear stress application. The effects of the drugs varied greatly between patients suggesting some could benefit from one drug while others not. Developing drugs targeting the AC signaling pathway could have clinical applications for SCD, but more researches with larger patient cohorts are needed to identify the differences in the responses of sickle RBCs

Distinct Immune Signature Predicts Progression of Vestibular Schwannoma and Unveils a Possible Viral Etiology

BACKGROUND: The management of sub-totally resected sporadic vestibular schwannoma (VS) may include observation, re-resection or irradiation. Identifying the optimal choice can be difficult due to the disease\u27s variable progression rate. We aimed to define an immune signature and associated transcriptomic fingerprint characteristic of rapidly-progressing VS to elucidate the underpinnings of rapidly progressing VS and identify a prognostic model for determining rate of progression. METHODS: We used multiplex immunofluorescence to characterize the immune microenvironment in 17 patients with sporadic VS treated with subtotal surgical resection alone. Transcriptomic analysis revealed differentially-expressed genes and dysregulated pathways when comparing rapidly-progressing VS to slowly or non-progressing VS. RESULTS: Rapidly progressing VS was distinctly enriched in CD4 CONCLUSION: Rapid progression of residual vestibular schwannoma following subtotal surgical resection has an underlying immune etiology that may be virally originating; and despite an abundant adaptive immune response, T-cell immunosenescence may be associated with rapid progression of VS. These findings provide a rationale for clinical trials evaluating immunotherapy in patients with rapidly progressing VS

Oxygen gradient ektacytometry-derived biomarkers are associated with acute complications in sickle cell disease

We investigated the potential of the point of sickling (PoS; the pO2 tension at which red cells start to sickle), determined by oxygen gradient ektacytometry to serve as a biomarker associated with the incidence of acute sickle cell disease-related complications in 177 children and 50 adults. In the pediatric cohort, for every 10 mmHg increase in PoS reflecting a greater likelihood of sickling, the likelihood of an individual experiencing >1 type of acute complication increased; the adjusted odds ratio (aOR) was 1.65. For every 0.1 increase in minimum elongation index (EImin; reflecting improved red blood cell deformability at hypoxia), the aOR was 0.50. In the adult cohort, for every 10 mmHg increase in PoS, we found an aOR of 3.00, although this was not significant after correcting for multiple testing. There was a trend for an association between higher PoS and greater likelihood of vaso-occlusive episodes (VOEs; children aOR, 1.35; adults aOR, 2.22). In children, only EImin was associated with VOEs (aOR, 0.68). When data of both cohorts were pooled, significant associations with PoS and/or EImin were found for all acute complications, independently and when >1 type of acute complication was assessed. These findings indicate that oxygen gradient ektacytometry generates novel biomarkers and provides a rationale for further development of these biomarkers in the assessment of clinical severity, evaluation of novel therapies, and as surrogate clinical trial end points. These biomarkers may be useful in assessing efficacy of novel therapies like pyruvate kinase activators, voxelotor, and L-glutamine

Silencing Nociceptor Neurons Reduces Allergic Airway Inflammation

Lung nociceptors initiate cough and bronchoconstriction. To elucidate if these fibers also contribute to allergic airway inflammation, we stimulated lung nociceptors with capsaicin and observed increased neuropeptide release and immune cell infiltration. In contrast, ablating Nav1.8(+) sensory neurons or silencing them with QX-314, a charged sodium channel inhibitor that enters via large-pore ion channels to specifically block nociceptors, substantially reduced ovalbumin- or house-dust-mite-induced airway inflammation and bronchial hyperresponsiveness. We also discovered that IL-5, a cytokine produced by activated immune cells, acts directly on nociceptors to induce the release of vasoactive intestinal peptide (VIP). VIP then stimulates CD4(+) and resident innate lymphoid type 2 cells, creating an inflammatory signaling loop that promotes allergic inflammation. Our results indicate that nociceptors amplify pathological adaptive immune responses and that silencing these neurons with QX-314 interrupts this neuro-immune interplay, revealing a potential new therapeutic strategy for asthma

Effects of Genotypes and Treatment on Oxygenscan Parameters in Sickle Cell Disease

(1) Background: The aim of the present study was to compare oxygen gradient ektacytometry parameters between sickle cell patients of different genotypes (SS, SC, and S/β+) or under different treatments (hydroxyurea or chronic red blood cell exchange). (2) Methods: Oxygen gradient ektacytometry was performed in 167 adults and children at steady state. In addition, five SS patients had oxygenscan measurements at steady state and during an acute complication requiring hospitalization. (3) Results: Red blood cell (RBC) deformability upon deoxygenation (EImin) and in normoxia (EImax) was increased, and the susceptibility of RBC to sickle upon deoxygenation was decreased in SC patients when compared to untreated SS patients older than 5 years old. SS patients under chronic red blood cell exchange had higher EImin and EImax and lower susceptibility of RBC to sickle upon deoxygenation compared to untreated SS patients, SS patients younger than 5 years old, and hydroxyurea-treated SS and SC patients. The susceptibility of RBC to sickle upon deoxygenation was increased in the five SS patients during acute complication compared to steady state, although the difference between steady state and acute complication was variable from one patient to another. (4) Conclusions: The present study demonstrates that oxygen gradient ektacytometry parameters are affected by sickle cell disease (SCD) genotype and treatment

Modulators of red blood cell rheology and eryptosis in sickle cell anemia : the cornerstone between red blood cell microparticles and vascular dysfunction ?

La drépanocytose est la première maladie génétique en France et dans le monde. Cette hémoglobinopathie mène à la production d’une hémoglobine mutée, appelée hémoglobine S (HbS), qui a la particularité de polymériser en condition désoxygénée, ce qui mène à la falciformation des globules rouges (GRs). Les GRs des patients drépanocytaires (SS) sont plus fragiles et moins déformables que des globules rouges d’individus non malades. Les patients souffrent ainsi d’une anémie hémolytique chronique, de crises vaso-occlusives douloureuses répétées et de nombreuses autres complications aiguës et chroniques. Cependant, ce mécanisme physiopathologique ne permet pas d’expliquer le caractère extrêmement polymorphe de l’expression clinique de la maladie. D’une maladie considérée à l’origine comme uniquement érythrocytaire, la drépanocytose est ainsi aujourd’hui également étudiée sous le prisme d’une pathologie vasculaire, où anomalies du globule rouge et dysfonction vasculaire seraient étroitement liées. Le stress oxydant et les facteurs pro-inflammatoires majorés ainsi que la biodisponibilité du monoxyde d’azote abaissée participeraient fortement à la physiopathologie de la maladie et notamment à la survenue d’une dysfonction vasculaire chronique. Nous avons montré au cours de cette thèse que le plasma des patients SS ainsi que des molécules plasmatiques inflammatoires augmentées dans la maladie exerçaient un effet délétère sur la rhéologie de leurs GRs, rhéologie qui participe à la survenue des complications cliniques. Nous avons également identifié que le stress oxydant majoré et la biodisponibilité du NO abaissée chez les malades altéraient la rhéologie des GRs. Ces résultats pourraient donc expliquer le bénéfice clinique de l’utilisation de la plasmaphérèse dans des rares cas de complications sévères résistantes à l’érythraphérèse. Nous avons également mis en évidence que le stress oxydant et le NO modulaient le phénomène d’éryptose ou mort du globule rouge chez les patients. Ce mécanisme s’accompagnerait de l’émission de microparticules de GRs (GR-MPs), qui se trouvent en concentration majorée chez les patients et de façon plus importante lors des crises. Nos études ont permis de mettre en évidence un lien entre ces GRs-MPs et la dysfonction macrovasculaire des patients. Nous avons également montré que des GR-MPs directement isolées chez les patients exerçaient un rôle délétère in vitro sur des cellules endothéliales de la macrocirculation. Celles-ci favorisaient l’expression de molécules d’adhérence et la production de cytokines pro-inflammatoires impliquées dans la dysfonction vasculaire et dans les complications de la maladie. Nos résultats ont révélé que ces propriétés toxiques des GR-MPs pourraient être due à l’activation du récepteur TLR4 sur l’endothélium. Des thérapies ciblées sur le stress oxydant, le NO ou encore TLR4 pourraient permette ainsi de limiter ces phénomènes et donc la survenue de complications. Le traitement de référence est actuellement l’hydroxyurée. Le mécanisme d’action principal de cette molécule repose sur l’augmentation de la production d’hémoglobine fœtale et donc la diminution de la concentration en HbS. Cette thèse a permis de mettre en évidence que l’hydroxyurée augmentait également la biodisponibilité du NO chez les patients, en lien probable avec un effet donneur de NO. Cet effet permettrait d’améliorer la déformabilité des GRs au travers d’une s-nitrosylation de la β-spectrine mais également d’abaisser le stress oxydant érythrocytaire. Ce traitement pourrait donc potentiellement réduire les phénomènes d’éryptose et d’émission de GR-MPs par ses propriétés sur le métabolisme du NO. Enfin, l’hydroxyurée menait également à inhiber la voie de production du NO intra-érythrocytaire. Les mécanismes à l’origine de cette inhibition ainsi que ses conséquences sur la production endogène de NO par les GRs restent cependant à être explorés.Sickle cell disease is the first genetic disorder in France and worldwide. This hemoglobinopathy leads to the production of a mutated hemoglobin called hemoglobin S (HbS) which has the property to polymerize in deoxygenated condition, leading to red blood cell (RBC) sickling. Sickle cell anemia (SCA) patient RBCs are more fragile and less deformable than RBC from healthy individuals. Patients suffer from hemolytic anemia, repeated painful vaso-occlusive crisis and other acute and chronical complications. However, this physiopathological mechanism cannot explain the clinical expression variability of the disease. Therefore, SCA is now considered as both as an erythrocyte disease and a vascular disease where RBC alterations and vessels dysfunction are closely linked. Oxidative stress, pro-inflammatory factors and decreased nitric oxide (NO) bioavailability could contribute to the physiopathology of the disease and to a chronic vascular dysfunction. We showed that plasma from SCA patients and inflammatory plasma factors that are majored in patient blood had deleterious effects on SCA RBC rheology, rheology that is known to participate to clinical complications. We also identified that majored oxidative stress and decreased NO altered RBC deformability. Those results could explain the clinical benefits of plasmapheresis which is used in severe complications that are resistant to blood transfusion. We also observed that oxidative stress and NO modulated eryptosis i.e.: the red blood cell death. This mechanism could be accompanied by red blood cell microparticles (RBC-MPs) release. The concentration of RBC-MPs is majored in patient blood at steady state and even more during crisis. Our study revealed a link between SCA patients RBC-MPs concentration and macrovascular dysfunction. We also showed that RBC-MPs directly isolated from patient blood had deleterious effect in vitro on macrovascular endothelial cells. Indeed, they promoted the expression of adhesion molecules and the production of inflammatory cytokines that are involved in vascular dysfunction and clinical complications. Our results also showed that those toxic properties could be attributed to their ability to activate TLR4 on endothelial cells. Therefore, therapies targeting oxidative stress, NO or TLR4 could limit those mechanisms and the onset of complications. Currently, hydroxyurea is the reference medication in SCA. Hydroxyurea increased the production of fetal hemoglobin and so decreased the concentration of HbS. This thesis also showed that this drug could increase NO bioavailability in patients, potentially in link with its NO donor property. This effect could improve SCA RBC deformability by promoting the S-Nitrosylation of the β-spectrine protein and could also decrease RBC oxidative stress. Therefore, this treatment could potentially decrease eryptosis and RBC-MPs released by its effects on NO metabolism. Finally, we showed that hydroxyurea therapy inhibited RBC NO production pathway. The mechanisms at the origin of this inhibition and the consequences on the endogenous NO production still need to be explore