Cytokine profiling and transcriptomics in mononuclear cells define immune variants in Meniere Disease.

Meniere Disease (MD) is a chronic inner ear disorder characterized by vertigo attacks, sensorineural hearing loss, tinnitus, and aural fullness. Extensive evidence supporting the inflammatory etiology of MD has been found, therefore, by using transcriptome analysis, we aim to describe the inflammatory variants of MD. We performed Bulk RNAseq on 45 patients with definite MD and 15 healthy controls. MD patients were classified according to their basal levels of IL-1ß into 2 groups: high and low. Differentially expression analysis was performed using the ExpHunter Suite, and cell type proportion was evaluated using the estimation algorithms xCell, ABIS, and CIBERSORTx. MD patients showed 15 differentially expressed genes (DEG) compared to controls. The top DEGs include IGHG1 (p = 1.64 × 10-6) and IGLV3-21 (p = 6.28 × 10-3), supporting a role in the adaptative immune response. Cytokine profiling defines a subgroup of patients with high levels of IL-1ß with up-regulation of IL6 (p = 7.65 × 10-8) and INHBA (p = 3.39 × 10-7) genes. Transcriptomic data from peripheral blood mononuclear cells support a proinflammatory subgroup of MD patients with high levels of IL6 and an increase in naïve B-cells, and memory CD8+ T cells.

Aldosterone inhibits Dot1l expression in guinea pig cochlea.

BACKGROUND: Aldosterone relieves transcriptional repression of epithelial sodium channel (ENaC) by inhibiting Dot1a and Af9 expression and their interaction with ENaC promoter in various tissues. Expressions of ENaC and Af9 in inner ear have been identified. However, it is not known how Dot1l is regulated by aldosterone in inner ear. METHODS: Twenty-eight adult guinea pigs were randomly divided into the control group and treatment group. Aldosterone 1 mg/kg/d was injected intraperitoneally in the treatment group and saline in the control group for 7 days. Animals were killed 1 month later following auditory brainstem response examination. Histomorphology of cochlea was detected with hematoxylin-eosin staining, and Dot1l expression was examined with immunohistochemistry and Western blot. RESULTS: There was no significant difference in ABR thresholds before and after injection of aldosterone or saline in either group. Endolymphatic hydrops was found in 75% of animals in the treatment group. Dot1l was found in both groups in the stria vascularis, Reissner's membrane, spiral limbus, organ of Corti and spiral ligament. Dot1l expression in the treatment group was decreased by aldosterone. CONCLUSIONS: Dot1l in guinea pig cochlea is inhibited by aldosterone with induction of endolymphatic hydrops. Dot1l may be closely related to endolymph regulation by aldosterone and to pathogenesis of Meniere's disease.

A Perspective for Ménière's Disease: In Silico Investigations of Dexamethasone as a Direct Modulator of AQP2.

Ménière's disease is a chronic illness characterized by intermittent episodes of vertigo associated with fluctuating sensorineural hearing loss, tinnitus and aural pressure. This pathology strongly correlates with a dilatation of the fluid compartment of the endolymph, so-called hydrops. Dexamethasone is one of the therapeutic approaches recommended when conventional antivertigo treatments have failed. Several mechanisms of actions have been hypothesized for the mode of action of dexamethasone, such as the anti-inflammatory effect or as a regulator of inner ear water homeostasis. However, none of them have been experimentally confirmed so far. Aquaporins (AQPs) are transmembrane water channels and are hence central in the regulation of transcellular water fluxes. In the present study, we investigated the hypothesis that dexamethasone could impact water fluxes in the inner ear by targeting AQP2. We addressed this question through molecular dynamics simulations approaches and managed to demonstrate a direct interaction between AQP2 and dexamethasone and its significant impact on the channel water permeability. Through compartmentalization of sodium and potassium ions, a significant effect of Na+ upon AQP2 water permeability was highlighted as well. The molecular mechanisms involved in dexamethasone binding and in its regulatory action upon AQP2 function are described.

Low-salt diet increases mRNA expression of aldosterone-regulated transporters in the intermediate portion of the endolymphatic sac.

The endolymphatic sac is a small sac-shaped organ at the end of the membranous labyrinth of the inner ear. The endolymphatic sac absorbs the endolymph, in which the ion balance is crucial for inner ear homeostasis. Of the three sections of the endolymphatic sac, the intermediate portion is the center of endolymph absorption, particularly sodium transport, and is thought to be regulated by aldosterone. Disorders of the endolymphatic sac may cause an excess of endolymph (endolymphatic hydrops), a histological observation in Meniere's disease. A low-salt diet is an effective treatment for Meniere's disease, and is based on the assumption that the absorption of endolymph in the endolymphatic sac abates endolymphatic hydrops through a physiological increase in aldosterone level. However, the molecular basis of endolymph absorption in each portion of the endolymphatic sac is largely unknown because of difficulties in gene expression analysis, resulting from its small size and intricate structure. The present study combined reverse transcription-quantitative polymerase chain reaction and laser capture microdissection techniques to analyze the difference of gene expression of the aldosterone-controlled epithelial Na+ channel, thiazide-sensitive Na+-Cl- cotransporter, and Na+, K+-ATPase genes in the three individual portions of the endolymphatic sac in a rat model. A low-salt diet increased the expression of aldosterone-controlled ion transporters, particularly in the intermediate portion of the endolymphatic sac. Our findings will contribute to the understanding of the physiological function of the endolymphatic sac and the pathophysiology of Meniere's disease.

Bidirectional Transport of IgE by CD23 in the Inner Ear of Patients with Meniere's Disease.

Meniere's disease (MD) is a disorder of the inner ear characterized by episodes of spontaneous vertigo, fluctuating hearing loss, and tinnitus. Recent studies have demonstrated that IgE may play a role in the pathogenesis of MD. Patients with MD (n = 103), acoustic neuroma (n = 5), and healthy subjects (n = 72) were recruited into the study. Serum from the participants was analyzed for IgE and type 2-related cytokines. IgE and CD23 expression levels in vestibular end organs of patients, C57BL/6 mice, or mouse HEI-OC1 cells were analyzed. Finally, the role of CD23 in IgE transcytosis was assessed using HEI-OC1 cells. Serum IgE was elevated in patients with MD and positively correlated with clinical symptoms. IL-4, IL-5, IL-10, IL-13, and CD23 levels were increased in patients with MD compared with the control group. In the transcytosis assay, mouse IgE was found to be bidirectionally transported across the HEI-OC1 cell monolayer. Additionally, CD23 downregulation using a small interfering RNA approach significantly reduced the efficiency of IgE transcytosis, suggesting that IgE is transported by CD23. Furthermore, exposure to IL-4 increased CD23 expression and enhanced IgE transcytosis in the HEI-OC1 cells and primary vestibular end organs. Our study indicated that IgE may play a role in the pathophysiology of MD. In addition, CD23-mediated IgE transcytosis in the hair cells may play a critical role in initiating inflammation in the inner ear. Thus, reducing the level of IgE may be a potentially effective approach for MD treatment.

MicroRNA Profiling as a Methodology to Diagnose Ménière's Disease: Potential Application of Machine Learning.

OBJECTIVE: Diagnosis and treatment of Ménière's disease remains a significant challenge because of our inability to understand what is occurring on a molecular level. MicroRNA (miRNA) perilymph profiling is a safe methodology and may serve as a "liquid biopsy" equivalent. We used machine learning (ML) to evaluate miRNA expression profiles of various inner ear pathologies to predict diagnosis of Ménière's disease. STUDY DESIGN: Prospective cohort study. SETTING: Tertiary academic hospital. SUBJECTS AND METHODS: Perilymph was collected during labyrinthectomy (Ménière's disease, n = 5), stapedotomy (otosclerosis, n = 5), and cochlear implantation (sensorineural hearing loss [SNHL], n = 9). miRNA was isolated and analyzed with the Affymetrix miRNA 4.0 array. Various ML classification models were evaluated with an 80/20 train/test split and cross-validation. Permutation feature importance was performed to understand miRNAs that were critical to the classification models. RESULTS: In terms of miRNA profiles for conductive hearing loss versus Ménière's, 4 models were able to differentiate and identify the 2 disease classes with 100% accuracy. The top-performing models used the same miRNAs in their decision classification model but with different weighted values. All candidate models for SNHL versus Ménière's performed significantly worse, with the best models achieving 66% accuracy. Ménière's models showed unique features distinct from SNHL. CONCLUSIONS: We can use ML to build Ménière's-specific prediction models using miRNA profile alone. However, ML models were less accurate in predicting SNHL from Ménière's, likely from overlap of miRNA biomarkers. The power of this technique is that it identifies biomarkers without knowledge of the pathophysiology, potentially leading to identification of novel biomarkers and diagnostic tests.

A possible mechanism of the formation of endolymphatic hydrops and its associated inner ear disorders.

The pathology of Meniere's disease (MD) is well established to be endolymphatic hydrops. However, the mechanism underlying deafness and vertigo of MD or idiopathic endolymphatic hydrops is still unknown. In order to evaluate the pathogenesis of deafness and vertigo in MD, it seems to be rational to investigate the interrelationship between hydrops and inner ear disorders using animals with experimentally-induced endolymphatic hydrops. In spite of intense efforts by many researchers, the mechanism of vertiginous attack has been unexplained, because animals with experimental hydrops usually did not show vertiginous attack. Recently, there are two reports to succeed to evoke vertiginous attack in animals with experimental hydrops. In the present paper were first surveyed past proposals about underlying mechanism of the development of hydrops and inner ear disorders associated with hydrops, and were discussed the pathogenetic mechanism of vertiginous attack in hydrops. In conclusion, abrupt development of hydrops was thought to play a pivotal role in the onset of vertiginous seizure.

Food-induced stimulation of the antisecretory factor to improve symptoms in Meniere's disease: our results.

PURPOSE: Specially processed cereals (SPC) that increase endogenous antisecretory factor (AF) synthesis have been proposed to improve symptoms of Meniere's disease (MD) with controversial results. The aim of this study was to evaluate the effects of SPC in patients with definite unilateral MD and compare the results to a treatment protocol with intravenous glycerol and dexamethasone. METHODS: Thirteen patients with unilateral MD were treated with SPC and 13 patients were treated with intravenous glycerol and dexamethasone for 12 months. Audio-vestibular evaluation was performed before (T0) and at the end of the treatments (T12). The number of vertigo spells were evaluated before and after therapy and the Efficacy Index (EI) was calculated. Questionnaires for hearing loss (HHIA), tinnitus (THI) and quality of life (TFL) were administered. RESULTS: EI decreased in the SPC group in the second semester compared to the first although not significantly (p = 0.6323). There was a significant reduction for THI score in the SPC group at T12 (p = 0.0325). No significant differences were found between the two groups at T0 (p = 0.4723), while a significant difference was found at T12 (p = 0.0041). Quality of life showed an improvement in daily activities in the SPC group compared to infusion therapy group. CONCLUSION: Our study shows a reduced number of vertigo attacks and a positive effect on the discomfort generated by tinnitus and quality of life in patients with unilateral MD treated with SPC and when compared to patients treated with intravenous glycerol and dexamethasone. No effects on hearing thresholds were noted in both groups.

The evolutionary hypothesis of benign paroxysmal positional vertigo.

Human otoliths, primarily formed from salts of calcium and carbonate, are different from bones of the skeleton, which are composed of calcium phosphate. The echinoderms, which share the earliest common ancestor with us, began to protect the body by making an endoskeleton out of calcium and carbon dioxide dissolved in the sea. In subsequent vertebrates, aerobic respiration supported strong muscle activity, but an occasional shortage of oxygen led to low pH due to the accumulation of lactate produced by anaerobic respiration, increasing the risk of melting bones composed of calcium carbonate. So, all vertebrates used calcium phosphate to increase bone strength, having a stronger ionic bonding than calcium carbonate. But otoliths, which are in the inner ear and thereby not connected to muscles, still use calcium carbonate. Benign paroxysmal positional vertigo (BPPV) is a disorder in which otoliths detached from the utricle enter the semicircular canals and cause a sense of rotation. Otoliths, the calcium carbonate ear bones retaining a long evolutionary history, can be easily broken at low pH. During sleep, shallow breathing produces mild respiratory acidosis and low pH in the blood. Since otoliths are corroded at low pH during nighttime, BPPV occurs frequently in the morning. In addition, diabetes mellitus or gout often decreases pH in the blood and increases the occurrence of BPPV.

Solute carrier family 4 member 1 might participate in the pathogenesis of Meniere's disease in a murine endolymphatic hydrop model.

Background: To date, the pathogenesis of Meniere's disease (MD) remains unclear. Previous research found that the SLC4A1 gene significantly down-regulated. Aims: This study sought to understand the effect of SLC4A1 on the pathogenesis of MD. ELH C57 mice models were induced by intraperitoneal injection of AVP. Material and methods: The mRNA expression levels of SLC4A1, SLC4A10 and SLC26A4 were monitored by real-time quantitative PCR, the protein expression levels of SLC4A1 were monitored by immunoblotting and immunofluorescence before and after the ELH. DIDS is an inhibitor of SLC4A1. The expression levels of SLC4A1 were also monitored in the AVP + DIDS group. Results: We successfully established the model of ELH after applied AVP. The results of HE staining showed displacement of Reissner's membrane with bulge to scala vestibule in ears of the AVP group. Cochlea/ELS SLC4A1 protein and SLC4A1, SLC4A10, SLC26A4 mRNA expressions were reduced significantly in C57 mice of the AVP group. The SLC4A1 protein expression levels and SLC4A1, SLC4A10, SLC26A4 mRNA expression levels declined more obvious in the cochlea and ELS in C57 mice of the AVP + DIDS group. Conclusions and significance: SLC4A1 was a protective factor in the pathogenesis of MD, but the mechanisms were unknown.

Immunolocalization of calcium sensing and transport proteins in the murine endolymphatic sac indicates calciostatic functions within the inner ear.

An exceptionally low calcium (Ca2+) concentration in the inner ear endolymph ([Ca2+]endolymph) is crucial for proper auditory and vestibular function. The endolymphatic sac (ES) is believed to critically contribute to the maintenance of this low [Ca2+]endolymph. Here, we investigated the immunohistochemical localization of proteins that are presumably involved in the sensing and transport of extracellular Ca2+ in the murine ES epithelium. Light microscopic and fluorescence immunolabeling in paraffin-embedded murine ES tissue sections (male C57BL/6 mice, 6-8 weeks old) demonstrated the presence of the calcium-sensing receptor CaSR, transient receptor potential cation channel subtypes TRPV5 and TRPV6, sarco/endoplasmic reticulum Ca2+-ATPases SERCA1 and SERCA2, Na+/Ca2+ exchanger NCX2, and plasma membrane Ca2+ ATPases PMCA1 and PMCA4 in ES epithelial cells. These proteins exhibited (i) membranous (apical or basolateral) or cytoplasmic localization patterns, (ii) a proximal-to-distal labeling gradient within the ES, and (iii) different distribution patterns among ES epithelial cell types (mitochondria-rich cells (MRCs) and ribosome-rich cells (RRCs)). Notably, in the inner ear membranous labyrinth, CaSR was exclusively localized in MRCs, suggesting a unique role of the ES epithelium in CaSR-mediated sensing and control of [Ca2+]endolymph. Structural loss of the distal ES, which is consistently observed in Meniere's disease, may therefore critically disturb [Ca2+]endolymph and contribute to the pathogenesis of Meniere's disease.

Endolymphatic hydrops and ionic transporters: genetic and biohumoral aspects.

Ménière's disease (MD) is an inner ear disorder, characterized by a burden of symptoms, probably arising from the interplay of genetic and environmental factors. In this brief review, we consider the role of ion channels and transporters in the pathophysiology of MD, focusing on genetic and biohumoral aspects. Pathophysiological mechanisms related to altered concentrations of ions in the endolymph include altered osmotic pressure leading to hydrops and/or immunomodulatory effects of K+ and Endogenous Ouabain (EO) concentrations in the inner ear. Aquaporins 1-5 (AQPs) have been found in the inner ear; AQP2 is the only isoform controlled by a hormone, namely, vasopressin (antidiuretic hormone, ADH). Genetic studies on AQPs have provided inconclusive results. Recently, two genetic polymorphisms have been associated with MD: rs3746951, a missense variant (Gly180Ser) in the Salt-Inducible Kinase-1 (SIK1) gene and rs487119, an intronic variant of gene SLC8A1 coding for a Na+,Ca++ exchanger (NCX-1). EO is a hormone released by the midbrain and adrenal glands. It controls the constitutive capacity of modulating Na+,K+-ATPase activity. Higher plasma levels of EO have been found in MD subjects compared to a control group.

Differential Proinflammatory Signature in Vestibular Migraine and Meniere Disease.

Vestibular Migraine (VM) and Meniere's Disease (MD) are episodic vestibular syndromes defined by a set of associated symptoms such as tinnitus, hearing loss or migraine features during the attacks. Both conditions may show symptom overlap and there is no biological marker to distinguish them. Two subgroups of MD patients have been reported, according to their IL-1ß profile. Therefore, considering the clinical similarity between VM and MD, we aimed to investigate the cytokine profile of MD and VM as a means to distinguish these patients. We have also carried out gene expression microarrays and measured the levels of 14 cytokines and 11 chemokines in 129 MD patients, 82 VM patients, and 66 healthy controls. Gene expression profile in peripheral blood mononuclear cells (PBMC) showed significant differences in MD patients with high and low basal levels of IL- 1ß and VM patients. MD patients with high basal levels of IL- 1ß (MDH) had overall higher levels of cytokines/chemokines when compared to the other subsets. CCL4 levels were significantly different between MDH, MD with low basal levels of IL- 1ß (MDL), VM and controls. Logistic regression identified IL- 1ß, CCL3, CCL22, and CXCL1 levels as capable of differentiating VM patients from MD patients (area under the curve = 0.995), suggesting a high diagnostic value in patients with symptoms overlap.

Proteome of normal human perilymph and perilymph from people with disabling vertigo.

The vast majority of hearing loss, the most common sensory impairment, and vertigo, which commonly causes falls, both reflect underlying dysfunction of inner ear cells. Perilymph sampling can thus provide molecular cues to hearing and balance disorders. While such "liquid biopsy" of the inner ear is not yet in routine clinical practice, previous studies have uncovered alterations in perilymph in patients with certain types of hearing loss. However, the proteome of perilymph from patients with intact hearing has been unknown. Furthermore, no complete characterization of perilymph from patients with vestibular dysfunction has been reported. Here, using liquid-chromatography with tandem mass spectrometry, we analyzed samples of normal perilymph collected from three patients with skull base meningiomas and intact hearing. We identified 228 proteins that were common across the samples, establishing a greatly expanded proteome of the previously inferred normal human perilymph. Further comparison to perilymph obtained from three patients with vestibular dysfunction with drop attacks due to Meniere's disease showed 38 proteins with significantly differential abundance. The abundance of four protein candidates with previously unknown roles in inner ear biology was validated in murine cochleae by immunohistochemistry and in situ hybridization: AACT, HGFAC, EFEMP1, and TGFBI. Together, these results motivate future work in characterizing the normal human perilymph and identifying biomarkers of inner ear disease.

Otopetrin-2 Immunolocalization in the Human Macula Utricle.

BACKGROUND: In the present study, we investigated the localization of otopetrin-2-a member of the otopetrin family that encodes proton-selective ion channels-in the human macula utricle using immunohistochemistry. METHODS: Macula utricle were acquired at surgery from patients who required transmastoid labyrinthectomy for intractable vertigo due to Meniere's disease (MD; n = 3) and/or vestibular drops attacks (VDA; n = 2) and from temporal bones (n = 2) acquired at autopsy from individuals with no balance disorders. Immunofluorescence staining with otopetrin-2 (rabbit affinity purified polyclonal antibody) and GFAP (mouse monoclonal antibody) to identify vestibular supporting cells was made in formalin fixed cryostat sections or whole microdissected utricle (for flat mount preparations). Secondary antibodies against rabbit and mouse were used for the identification of both proteins. Digital fluorescent images were obtained using a high-resolution laser confocal microscope. RESULTS: Using cryostat sections and flat mount preparations otopetrin-2 immunofluorescence was seen as punctated signal throughout the supporting cells cytoplasm. GFAP immunofluorescence was present in the supporting cell cytoplasm. The distribution of otopetrin-2 was similar in the macula utricle obtained from MD, VDA, or autopsy normative patients. CONCLUSIONS: Otopetrin-2 was localized in supporting cells in a similar fashion that otopetrin-1 previously reported in the mouse macula utricle. The differential expression of otopetrin-2 in the supporting cells of the human macula utricle suggest an important role in the vestibular sensory periphery homeostasis and otolith maintenance.

Histamine H4 receptor gene polymorphisms: a potential contributor to Meniere disease.

BACKGROUND: The immune system is likely involved in the pathophysiology of Meniere's disease (MD). However, its role of patients with MD has not been well studied. Given that histamine H4 receptors are highly expressed in immune system, we tested the hypothesis that histamine H4 receptor gene polymorphisms are a potential contributor to the risk of MD. METHODS: A group of patients was enrolled with a diagnosis of definite MD based on the American Academy of Otolaryngology-Head and Neck Surgery Committee on Hearing and Equilibrium guidelines and a control group of patients without any vestibular disease. We selected one SNP, rs77485247 in HRH4 and conducted an exploratory investigation of its correlations with the symptoms of vertigo and proinflammatory cytokines levels in MD patients. RESULTS: HRH4 rs77485247 polymorphism may be associated with the risk of MD. Furthermore, basal levels of proinflammatory cytokines, such as IL-1ß and TNF-α, in PBMCs are increased in patients with MD compared to control patients. This increased basal level of proinflammatory cytokines is prominent in MD patients with the A allele. CONCLUSIONS: These suggested that HRH4 rs77485247 polymorphism may be an important mediator in regulating proinflammatory cytokines, which are involved in the pathogenesis of MD.

Inner ear pathologies impair sodium-regulated ion transport in Meniere's disease.

Meniere's disease (MD), a syndromal inner ear disease, is commonly associated with a pathological accumulation of endolymphatic fluid in the inner ear, termed "idiopathic" endolymphatic hydrops (iEH). Although numerous precipitating/exacerbating factors have been proposed for MD, its etiology remains elusive. Here, using immunohistochemistry and in situ protein-protein interaction detection assays, we demonstrate mineralocorticoid-controlled sodium transport mechanisms in the epithelium of the extraosseous portion of the endolymphatic sac (eES) in the murine and human inner ears. Histological analysis of the eES in an extensive series of human temporal bones consistently revealed pathological changes in the eES in cases with iEH and a clinical history of MD, but no such changes were found in cases with "secondary" EH due to other otological diseases or in healthy controls. Notably, two etiologically different pathologies-degeneration and developmental hypoplasia-that selectively affect the eES in MD were distinguished. Clinical records from MD cases with degenerative and hypoplastic eES pathology revealed distinct intergroup differences in clinical disease presentation. Overall, we have identified for the first time two inner ear pathologies that are consistently present in MD and can be directly linked to the pathogenesis of EH, and which potentially affect the phenotypical presentation of MD.

Salivary α-amylase levels in vertigo: Can it be an autonomic dysfunction?

We aim to demonstrate possible autonomic dysfunction based on salivary α-amylase measurements during and after the vertigo attacks associated with Ménière disease (MD) and benign paroxysmal positional vertigo (BPPV). Patients admitted to the emergency room with a diagnosis of vertigo attacks caused by either MD (n = 15) or BPPV (n = 9) constituted the study groups. The control group (n = 10) consisted of volunteer patients admitted to the emergency department with minor soft-tissue trauma. The first saliva samples were obtained immediately during the attacks and the second and third samples were obtained on the third and fifteenth days of the attack, respectively. In the controls, the first sample was obtained after admission to the hospital and the second sample was obtained on the third day. Salivary α-amylase levels were evaluated. The difference between salivary α-amylase levels in patients with MD and BPPV was not significant. The amylase value measured early after the BPPV attack was significantly lower than that of the controls (p = 0.008). Although not significant, an undulating pattern of salivary α-amylase levels was observed with both diseases. An autonomic imbalance could be partly demonstrated by salivary α-amylase measurement early after the attack in patients with BPPV. Therefore, amylase may be a promising marker that is worth further investigation.

Proinflammatory cytokines and response to molds in mononuclear cells of patients with Meniere disease.

Epidemiological studies have found a higher prevalence of allergic symptoms and positive prick tests in patients with Meniere's disease (MD); however the effect of allergenic extracts in MD has not been established. Thus, this study aims to determine the effect of Aspergillus and Penicillium stimulation in cytokine release and gene expression profile in MD. Patients with MD showed higher basal levels of IL-1ß, IL-1RA, IL-6 and TNF-α when compared to healthy controls. We observed that IL-1ß levels had a bimodal distribution suggesting two different subgroups of patients, with low and high basal levels of cytokines. Gene expression profile in peripheral blood mononuclear cells (PBMC) showed significant differences in patients with high and low basal levels of IL-1ß. We found that both mold extracts triggered a significant release of TNF-α in MD patients, which were not found in controls. Moreover, after mold stimulation, MD patients showed a different gene expression profile in PBMC, according to the basal levels of IL-1ß. The results indicate that a subset of MD patients have higher basal levels of proinflammatory cytokines and the exposure to Aspergillus and Penicillium extracts may trigger additional TNF-α release and contribute to exacerbate inflammation.

Ménière's Disease Pathophysiology: Endolymphatic Sac Immunohistochemical Study of Aquaporin-2, V2R Vasopressin Receptor, NKCC2, and TRPV4.

Objectives Endolymphatic sac (ELS) pathophysiology in Ménière's disease (MD) remains poorly understood. We identified from the literature a group of proteins expressed on the ELS and involved in endolymph volume regulation: aquaporin-2 (AQP2), vasopressin receptor V2R, sodium potassium chloride cotransporter 2 (NKCC2), and transient receptor potential cation channel V4 (TRPV4). Our objective was to determine whether their ELS expression was altered in MD, to better understand the pathophysiology of endolymphatic hydrops. Study Design Prospective case-control study. Setting Tertiary care center. Subjects Twenty-four patients with definite MD undergoing endolymphatic duct blockage surgery were recruited, as well as 23 controls with no history of MD undergoing surgery for vestibular schwannoma (VS). Methods ELS biopsies and blood samples for plasma arginine vasopressin (AVP) were obtained. Immunohistochemistry for AQP2, V2R, NKCC2, and TRPV4 was performed. Slides were scanned digitally for highly sensitive pixel density analysis by specialized software (VIS; Visiopharm). Results Global scores generated by the software represent total and relative protein expression density of 3 staining intensity levels, exclusively on ELS epithelium. AQP2 expression density was significantly elevated in MD compared to VS ( P = .003). There was no significant difference in plasma AVP, V2R, NKCC2, and TRPV4 expression. Conclusion This original study evaluates simultaneous in situ expression of AQP2, V2R, NKCC2, and TRPV4 on the human ELS in MD, with a control group. Our results show only AQP2 upregulation on the ELS of patients with MD. We suggest a constitutively increased expression of AQP2 in MD, independent of its regulatory axis (AVP-V2R). Acquired regulator sequence mutations could support this model.