Flow-Based CL-SMIA for the Quantification of Protein Biomarkers from Nasal Secretions in Comparison with Sandwich ELISA.

Protein biomarkers in nasal secretions can be used as a measure to differentiate between allergies, airway diseases and infections for non-invasive diagnostics. The point-of-care quantification of biomarker levels using flow-based microarray facilitates precise and rapid diagnosis and displays the potential for targeted and effective treatment. For the first time, we developed a flow-based chemiluminescence sandwich microarray immunoassay (CL-SMIA) for the quantification of nasal interferon-beta (IFN-ß) on the Microarray Chip Reader-Research (MCR-R). Polycarbonate foils are used as a cost-effective surface for immobilizing capture antibodies. By using a commercially available set of anti-human IFN-ß antibodies, the CL-SMIA can be compared directly to an enzyme-linked immunosorbent assay (ELISA) performed in microtiter plates concerning the bioanalytical performance and economic issues. Pre-incubation of the sample with detection antibodies facilitates the lower consumption of detection antibodies, as this allows for a longer interaction time between the antibody and the biomarker. The direct injection of pre-incubated samples into the microarray chips eliminates the adsorption of proteins in the tubing as well as the contamination of the tubing and valves of the MCR-R with clinical samples. The small flow cell allows for a low sample volume of 50 µL. The limit of detection of 4.53 pg mL-1 was slightly increased compared to a sandwich ELISA performed on microtiter plates which were 1.60 pg mL-1. The possibility to perform the CL-SMIA in a multiplexed mode makes it a promising assay for the rapid and cost-effective non-invasive detection of biomarkers in nasal secretions.

Early reduction of SARS-CoV-2-replication in bronchial epithelium by kinin B2 receptor antagonism.

SARS-CoV-2 has evolved to enter the host via the ACE2 receptor which is part of the kinin-kallikrein pathway. This complex pathway is only poorly understood in context of immune regulation but critical to control infection. This study examines SARS-CoV-2-infection and epithelial mechanisms of the kinin-kallikrein-system at the kinin B2 receptor level in SARS-CoV-2-infection that is of direct translational relevance. From acute SARS-CoV-2-positive study participants and -negative controls, transcriptomes of nasal curettages were analyzed. Primary airway epithelial cells (NHBEs) were infected with SARS-CoV-2 and treated with the approved B2R-antagonist icatibant. SARS-CoV-2 RNA RT-qPCR, cytotoxicity assays, plaque assays, and transcriptome analyses were performed. The treatment effect was further studied in a murine airway inflammation model in vivo. Here, we report a broad and strong upregulation of kallikreins and the kinin B2 receptor (B2R) in the nasal mucosa of acutely symptomatic SARS-CoV-2-positive study participants. A B2R-antagonist impeded SARS-CoV-2 replication and spread in NHBEs, as determined in plaque assays on Vero-E6 cells. B2R-antagonism reduced the expression of SARS-CoV-2 entry receptor ACE2, G protein-coupled receptor signaling, and ion transport in vitro and in a murine airway inflammation in vivo model. In summary, this study provides evidence that treatment with B2R-antagonists protects airway epithelial cells from SARS-CoV-2 by inhibiting its replication and spread, through the reduction of ACE2 levels and the interference with several cellular signaling processes. Future clinical studies need to shed light on the airway protection potential of approved B2R-antagonists, like icatibant, in the treatment of early-stage COVID-19. KEY MESSAGES: Induction of kinin B2 receptor in the nose of SARS-CoV-2-positive patients. Treatment with B2R-antagonist protects airway epithelial cells from SARS-CoV-2. B2R-antagonist reduces ACE2 levels in vivo and ex vivo. Protection by B2R-antagonist is mediated by inhibiting viral replication and spread.

Correction to: Clinical manifestations of hydropic ear disease (Menière's).

In the original publication, Fig. 1 was incorrectly published with incomplete legends.

High-Frequency Horizontal Semicircular Canal Function in Certain Menière's Disease.

OBJECTIVES: To characterize video head impulse testing (vHIT) in definitive Menière's disease and to investigate the relationship between vHIT results and other audiovestibular function tests. DESIGN: Fifty-four patients with certain Menière's disease, that is, patients with clinically definite Menière's disease and endolymphatic hydrops visualized by locally enhanced inner ear MR imaging, were recruited for this study. All patients underwent vHIT. The vestibular-ocular reflex gain at 60 ms and refixation saccades were the outcome parameters measured. Saccades were characterized by determining the saccade frequency, their mean latency, and their mean velocity. RESULTS: Seven of 54 patients had bilateral normal vHITs. Clearly pathologically vHITs with decreased gain and refixation saccades were observed in further seven patients. The majority of patients exhibited vHITs with refixation saccades but normal gain. Saccades mostly occurred bilaterally. There was no correlation between vHIT gain or saccades and caloric irrigation, cervical vestibular evoked myogenic potential or audiometry for Menière's ears. Furthermore, vHIT gain or saccades correlated neither with age nor with the duration of the disease. CONCLUSIONS: Pathological low vHIT gain values are rarely observed in patients with certain Menière's disease, while refixation saccades occur very frequently. In the majority of patients, saccades occur bilaterally. In Menière's disease, in contrast to vestibular neuritis, there is no compensatory decrease of saccade latency over time.

Clinical manifestations of hydropic ear disease (Menière's).

INTRODUCTION: Hydropic ear disease, initially described by and named after Prosper Menière, is one of the most frequent vertigo disorders and one of the most frequent inner ear disorders. It is the syndrome of endolymphatic hydrops which until 2007 could be diagnostically confirmed only by post-mortem histology. In the past, various attempts to formulate clinical diagnostic criteria have been undertaken but were hampered by the inability to ascertain the diagnosis in living patients. With the milestone achievement of endolymphatic hydrops imaging, today the pathology can be ascertained. In this study, we have performed a detailed analysis of the clinical features of hydropic ear disease for the first time by examining a large cohort of patients with morphologically confirmed endolymphatic hydrops using a detailed physician-administered neurotologic face-to-face interview. RESULTS: During a hydropic vertigo attack, the patients report nausea, vomiting, sweating, urge to defecate, urge to urinate, phosphenes, headache, photophobia, phonophobia and even transient loss of consciousness. A third of the patients does not experience auditory symptoms during the vertigo attacks. Vertigo attacks last less than 20 min in more than one-fourth of the patients. Audiometric hearing loss has its greatest diagnostic value at the frequencies of 1 kHz and below. Cochleovestibular symptom onset simultaneity is associated with a high frequency of drop-attacks. Migraine and autoimmune disorders are not associated with hydropic ear disease. CONCLUSION: This study marks the beginning of the clinical characterization of hydropic ear disease. The findings have important implications for the future formulation of clinical diagnostic criteria.

Relationship Between the Extent of Endolymphatic Hydrops and the Severity and Fluctuation of Audiovestibular Symptoms in Patients With Menière's Disease and MRI Evidence of Hydrops.

OBJECTIVE: To characterize the short-, middle-, and long-term occurrence of vertigo attacks in a large population of Menière's disease (MD) and to investigate the relationship between the extent of endolymphatic hydrops (ELH) and the severity of audiovestibular symptoms. STUDY DESIGN: Prospective observational study. METHODS: One hundred ninety-two patients with clinically definite MD participated in this study. The degree of ELH was visualized by locally enhanced inner ear magnetic resonance imaging. The occurrence and intensity of vertigo attacks, hearing loss, tinnitus, and aural fullness were documented in patient diaries. RESULTS: There was no significant correlation between the extent of cochlear or vestibular hydrops and the number of definite vertigo days, neither with regard to a short-term nor with regard to a middle-term time period. There was also no correlation between the extent of ELH and the intensity or activity of the coexisting aural symptoms hearing loss, tinnitus, and aural fullness. The duration of the disease significantly correlated with the extent of both cochlear and vestibular hydrops, but not with the number of definite vertigo days. CONCLUSION: The ELH was progressive in the long-term course of the disease in this large population of definite MD patients, but short-term and middle-term fluctuations of the symptom severity did not involve measurable variations of the ELH. Furthermore, the symptom severity did not decrease with increasing disease duration.

The Effect of Elevated Intracranial Pressure on Frequency Tuning of Air-Conducted Ocular Vestibular Myogenic Potentials in Ménière's Disease Patients.

OBJECTIVE: To investigate the effect of increased intracranial pressure on ocular vestibular-evoked myogenic potential (oVEMP) amplitudes and frequency tuning in patients with Ménière's disease (MD) to elucidate whether oVEMPS recorded under such conditions could provide a simple and accurate diagnostic test for MD. SETTING: Tertiary referral center for otology and neurotology. PATIENTS: Ten patients with certain unilateral MD (mean age = 48.2 yr, range 25-75 yr, 6 males and 4 females) as confirmed by a locally enhanced inner ear MRI (LEIM) were enrolled in this study. DESIGN: Air-conducted tone-burst oVEMP amplitudes were measured in response to 500 Hz and 1000 Hz in the horizontal plane (0 degree), a 20-degree head-down position. RESULTS: Tilting the patients from the horizontal position to the 20-degree head-down position led to a large reduction in oVEMP amplitudes to the 500 Hz tone burst (3.02 µV vs 1.17 µV, p = 0.005) and to a smaller one in the 1000 Hz tone burst (2.28 µV vs 1.78 µV, p = 0.013) in the Ménière's ear. Accordingly, the 500/1000 Hz frequency-tuning ratio was significantly decreased in the Ménière's ear as a result of this manoeuver (1.36 vs 0.75, p = 0.005). CONCLUSION: oVEMP amplitudes and frequency tuning in MD patients show a similar behavior to that found in healthy control subjects. oVEMP testing of putative MD patients in the tilted position is therefore unlikely to be diagnostically useful.

Utilisation of multi-frequency VEMPs improves diagnostic accuracy for Meniere's disease.

To determine whether vestibular evoked myogenic potential (VEMP) measurements that combine the VEMP 500/1000 Hz frequency tuning ratio and the inter-aural asymmetry ratio can reliably detect unilateral Meniere's disease ears as compared to healthy controls. Forty-two consecutive patients with certain unilateral Meniere's disease (as confirmed using a locally enhanced inner ear MRI (LEIM)) were assessed. Cervical vestibular evoked myogenic potentials (cVEMP) and ocular vestibular evoked myogenic potentials (oVEMP) were recorded at 500 and 1000 Hz. The VEMP amplitudes, asymmetry ratios, and the 500/1000 Hz amplitude ratios were compared with those of 21 age-matched healthy controls. A multi-frequency VEMPs score that combined: (1) the cVEMP 500/1000 Hz amplitude ratio, (2) the oVEMP 500/1000 Hz amplitude ratio, (3) the 500 Hz cVEMP asymmetry ratio, (4) the 1000 Hz cVEMP asymmetry ratio, produced a ROC curve with an area under the curve (AUC) of 0.814. The inclusion of audiology data further improved the result to 0.906. This score can be used to discriminate with a good degree of clinical accuracy between Meniere's ears (unilateral) and those of healthy controls. Multi-frequency VEMP analysis offers a simple, cost-effective solution to the diagnostic difficulties presented by Meniere's disease.

Effect of Spatial Orientation of the Horizontal Semicircular Canal on the Vestibulo-Ocular Reflex.

OBJECTIVE: To determine if an alignment of the horizontal semi-circular canal (hSCC) with the plane of rotation would enhance the vestibular-ocular reflex (VOR) gain result as it has been previously suggested. STUDY DESIGN: Comparative study of a physiological vestibular function test in healthy subjects. SETTING: Tertiary referral center for otology and neurotology. PATIENTS: Twenty two healthy volunteers were recruited for this study. Their mean age was 25.6 years and the sex distribution was 14:8 (M:F). None of the subjects had a history of audiovestibular disorders. INTERVENTION: The video Head Impulse Test (v-HIT) was performed with the hSCC in the conventional position (head upright, horizontal gaze) and also with the hSCC in-line with the earth horizontal. MAIN OUTCOME MEASURES: depending on the alignment of the hSCC with the plane of head rotation. RESULTS: There was no significant difference between the results, either for the VOR gain at 60 ms, or the regression slope gain, when the two alternative head positions were compared. CONCLUSIONS: The data acquired in this study show that the VOR as measured by the v-HIT is not enhanced by aligning the plane of the hSCC with the plane of rotation during the testing procedure. Hence, we recommend that the positioning of the patient, with the head upright and a horizontal gaze direction should be routinely used in the clinical evaluation of the angular VOR by v-HIT.

The Effect of Increasing Intracranial Pressure on Ocular Vestibular-Evoked Myogenic Potential Frequency Tuning.

OBJECTIVE: Ocular vestibular-evoked myogenic potentials (oVEMPs) represent extraocular muscle activity in response to vestibular stimulation. The authors sought to investigate whether posture-induced increase of the intracranial pressure (ICP) modulated oVEMP frequency tuning, that is, the amplitude ratio between 500-Hz and 1000-Hz stimuli. DESIGN: Ten healthy subjects were enrolled in this study. The subjects were positioned in the horizontal plane (0 degree) and in a 30-degree head-downwards position to elevate the ICP. In both positions, oVEMPs were recorded using 500-Hz and 1000-Hz air-conducted tone bursts. RESULTS: When tilting the subject from the horizontal plane to the 30-degree head-down position, oVEMP amplitudes in response to 500-Hz tone bursts distinctly decreased (3.40 µV versus 2.06 µV; p < 0.001), whereas amplitudes to 1000 Hz were only slightly diminished (2.74 µV versus 2.48 µV; p = 0.251). Correspondingly, the 500/1000-Hz amplitude ratio significantly decreased when tilting the subjects from 0- to 30-degree inclination (1.59 versus 1.05; p = 0.029). Latencies were not modulated by head-down position. CONCLUSIONS: Increasing ICP systematically alters oVEMPs in terms of absolute amplitudes and frequency tuning characteristics. oVEMPs are therefore in principle suited for noninvasive ICP monitoring.

Enhancing the reproducibility of ocular vestibular evoked myogenic potentials by use of a visual target originating from a head-mounted laser.

Ocular vestibular evoked myogenic potentials (oVEMPs) represent extraocular muscle activity in response to vestibular stimulation. oVEMP amplitudes are known to increase with increasing upward gaze angle, while the patient fixates a visual target. We investigated two different methods of presenting a visual target during oVEMP recordings. 57 healthy subjects were enrolled in this study. oVEMPs were elicited by 500 Hz air-conducted tone bursts while the subjects were looking upward at a marking which was either fixed on the wall or originated from a head-mounted laser attached to a headband, in either case corresponding to a 35° upward gaze angle. oVEMP amplitudes and latencies did not differ between the subjects looking at the fixed marking and the ones looking at the laser marking. The intra-individual standard deviation of amplitudes obtained by two separate measurements for each subject, however, as a measure of test-retest reliability, was significantly smaller for the laser headband group (0.60) in comparison to the group looking at the fixed marking (0.96; p = 0.007). The intraclass correlation coefficient revealed better test-retest reliability for oVEMP amplitudes when using the laser headband (0.957) than using the fixed marking (0.908). Hence, the use of a visual target originating from a headband enhances the reproducibility of oVEMPs. This might be due to the fact that the laser headband ensures a constant gaze angle and rules out the influence of small involuntary head movements on the gaze angle.

Posture-induced changes of ocular vestibular evoked myogenic potentials suggest a modulation by intracranial pressure.

Ocular vestibular evoked myogenic potentials (oVEMPs) represent extraocular muscle activity in response to vestibular stimulation. We sought to investigate whether oVEMPs are modulated by increasing intracranial pressure (ICP). Air-conducted oVEMPs were elicited in 20 healthy subjects lying supine on a tilt table. In order to elevate the ICP, the table was stepwise tilted from the horizontal plane to a 30° declination, corresponding to a 0°, 10°, 20° and 30° head-down position. At each inclination angle, oVEMP recording was performed in two head positions: (1) the head in line with the body and (2) the head positioned horizontally with the body tilted. When tilting both the body and head, oVEMP amplitudes gradually declined from 4.59 µV at 0° to 2.24 µV at 30° head-down position, revealing a highly significant reduction in amplitudes for all tilt angles when compared to the baseline value (p < 0.001). In parallel, the response prevalence decreased and latencies prolonged. Similar effects were observed when the body was tilted but the head positioned horizontally, even though the decrease in oVEMP amplitudes was less pronounced. A gravitoinertial force effect upon the otolith organs could thereby be excluded as a possible confounder. Hence, oVEMPs were most likely modulated by increasing ICP. In the range of the horizontal plane to a 30° head-down tilt, there was a linear correlation between oVEMP amplitudes and the inclination angle. oVEMPs might in principle be suited for non-invasive ICP monitoring.

Longitudinal assessment of endolymphatic hydrops with contrast-enhanced magnetic resonance imaging of the labyrinth.

OBJECTIVE: To visualize the progression of endolymphatic hydrops in a patient with Ménière's disease. PATIENT AND METHODS: A 53-year-old patient with definite Ménière's disease underwent locally enhanced inner ear magnetic resonance imaging (LEIM) for visualizing endolymphatic hydrops. Two years later, a second magnetic resonance imaging was performed for follow-up. RESULTS: The first magnetic resonance image confirmed the presence of endolymphatic hydrops and a treatment with betahistine was begun. Two years later, no more vertigo attacks occurred. The audiometry showed a deterioration of the hearing loss on both sides, whereas the caloric irrigation revealed normal results on both occasions. In comparison to the first examination, the second LEIM showed an augmented endolymphatic hydrops in both the cochlear and the vestibular compartments. CONCLUSION: For the first time, this case of Ménière's disease shows an enlargement of endolymphatic hydrops in 1 individual in the course of 2 years. A further development of the LEIM technique is desirable to enable the volumetric quantification of endolymphatic hydrops and to monitor the effect of therapies on the course of Ménière's disease.

Ocular vestibular evoked myogenic potential frequency tuning in certain Menière's disease.

Ocular vestibular evoked myogenic potentials (oVEMP) represent extraocular muscle activity in response to vestibular stimulation. To specify the value of oVEMP in the diagnostics of Menière's disease, the amplitude ratio between 500 and 1000 Hz stimuli was investigated. Thirty-nine patients with certain Menière's disease, i.e. definite Menière's disease with visualization of endolymphatic hydrops by magnetic resonance imaging and 19 age-matched healthy controls were enrolled in this study. oVEMP were recorded using 500 and 1000 Hz air-conducted tone bursts. For Menière's ears, the 500/1000 Hz amplitude ratio (mean ratio = 1.20) was significantly smaller when compared to unaffected ears of Menière's patients (mean ratio = 1.80; p = 0.008) or healthy controls (mean ratio = 1.81; p = 0.011). The amplitude ratio was neither correlated with the degree of endolymphatic hydrops nor with the duration of disease. While an older age was associated with a diminished amplitude ratio in healthy controls, there was no correlation between the amplitude ratio and age in Menière's ears. Hence, the calculation of the oVEMP 500/1000 Hz amplitude ratio may be a valuable diagnostic tool for Menière's disease.