Darwin's finches are a classic example of adaptive radiation, a process by which multiple ecologically distinct species rapidly evolve from a single ancestor. Such evolutionary diversification is typically explained by adaptation to new ecological opportunities. However, the ecological diversification of Darwin's finches following their dispersal to Galápagos was not matched on the same archipelago by other lineages of colonizing land birds, which diversified very little in terms of both species number and morphology. To better understand the causes underlying the extraordinary variation in Darwin's finches, we analyze the evolutionary dynamics of speciation and trait diversification in Thraupidae, including Coerebinae (Darwin's finches and relatives) and, their closely related clade, Sporophilinae. For all traits, we observe an early pulse of speciation and morphological diversification followed by prolonged periods of slower steady‐state rates of change. The primary exception is the apparent recent increase in diversification rate in Darwin's finches coupled with highly variable beak morphology, a potential key factor explaining this adaptive radiation. Our observations illustrate how the exploitation of ecological opportunity by contrasting means can produce clades with similarly high diversification rate yet strikingly different degrees of ecological and morphological differentiation. The ecological diversification of Darwin's finches is unmatched on the same archipelago by any other clade of colonising land birds. For all morphological traits, we observe an early pulse of diversification followed by prolonged periods of slower steady‐state rates of change for the tanager family a substantial increase in speciation rate in Darwin's finches coupled with highly variable beak morphology. Our observations illustrate how the exploitation of ecological opportunity by contrasting means can produce clades high in speciation rates yet strikingly different in their degree of ecological and morphological differentiation. The classic concept of an adaptive radiation involves high levels of ecological diversity evolving rapidly within a single lineage , to the deep, bulky beak of the Large Ground Finch (Geospiza magnirostris)  and the Yellow Warbler (Setophaga petechia aureola) have undergone minimal differentiation compared to their South American mainland or Caribbean counterparts  locate radiating clades within Thraupidae, (b) identify the mode of trait evolution during their evolutionary history, and (c) explore morphological disparity among clades. We predict that shifts in morphological trait evolution will be more evident in Darwin's finches than most of their Caribbean relatives and the rest of Thraupidae. We also expect to find more shifts specifically in the evolution of beak traits, compared to body traits, and for those shifts to delineate distinct subclades within Thraupidae.Following this investigation, we directly compare beak shape diversity in Darwin's finches, mockingbirds, and other Galápagos bird endemics to examine how beak traits have diversified in different lineages colonizing the same environment. Based on observed variation in beak shape across these radiations, we test whether Darwin's finches evolved to occupy a larger area of morphological space compared to other endemic taxa in relation to their respective parent lineages. We also assessed the role of ecological selection by examining whether the extent, kind, and directionality of clade‐specific beak shape variation are reflected in dietary niches.22.1Much of the data relevant to this study was published by Drury et al.  but we iThe clade of Darwin's finches studied here consists of 14 genetically and morphologically distinct species. Recent phylogenomic studies have treated up to four additional Darwin's finch taxa as species . Using the “BAMMtools” package in R  using the “coda” package  and the strength of directed evolution toward the optima (α) can be set to be equal across all regimes but with different phylogenetic state means (“OUM”) or to differ between regimes . To select the optimal model, we used the second order Akaike information criterion (AICc), taking into account sample sizes and a penalty increase for model complexity.Regimes for all branches in the phylogeny were then defined based on those identified from the bayou analyses for each trait. The “OUwie” package fits OU‐based models whereby traits evolve under discrete selective regimes and the models themselves are allow to vary in given parameters  addresses this problem by describing changes in a mean  given changes in covariates  accounting for nonindependence  in the “vegan” package  and a second at the base of the crown Sporophilinae clade  with the Darwin's finch and Sporophilinae clades being two notable exceptions undergoing significantly higher diversification rates. The MAP shifts found in BAMM support this conclusion, with one shift present in the Darwin's finch clade on the branch after the appearance of the Green Warbler‐Finch , and seed finches Oryzoborus (Sporophilinae)  evolving wider beaks than other Coerebinae members. Significant shifts in beak width are also detected in several species of Darwin's finches (Geospiza spp.) and in seed finches (Oryzoborus spp.). At least four shifts in total beak length have occurred during evolution of Thraupidae: Sporophilinae has evolved shorter beaks, while Oryzoborus, Saltator, Geospiza magnirostris, and G. conirostris have all evolved shorter and deeper beaks.Bayou analysis detected three locations on the tree where large beak depth increases, namely in members of ) Figure . Tarsus ) Figure . Larger Geospiza magnirostris and G. conirostris). A more recent phylogeny of Darwin's finches  and the strength of directed evolution toward the optima (α) are allowed to vary. The AICc values for the OUMVA model were much lower than those for other models tested, with values for the simpler OU models  similar to those under BM models  are tightly clustered with other granivores in the PCoA diet space, whereas the 14 Darwin's finch species cover the majority of the diet space occupied by the 349 tanager species representing the entire Thraupidae family Figure . This arThe results of the GEE analysis show there are significant differences in beak morphology among the various dietary guilds Table . For exa44.1Oryzoborus species (Sporophilinae). This implies that selection acts on multiple trait axes, sometimes driving species toward new adaptive optima for those respective traits  and the Vampire Finch , are morphologically similar to their sister species . They lack extreme beak morphologies and fall well within the bounds of the morphospace occupied by Darwin's finches as a group .The high number of shifts in beak evolution detected among the internal branches of Darwin's finches may be biased by incomplete sampling, but this seems unlikely to be the case since the species missing from our phylogenetic analysis, including the Grey Warbler‐Finch , after which the tempo of evolutionary change becomes stable with mostly minor fluctuations in recent times Figure . AlthougWhat factors can help explain the observed patterns of morphological evolution in the tanager family? External factors such as available niches and natural selection are undoubtedly important and provide the usual explanation for early bursts of avian morphological evolution, since these can arise from rapid saturation of ecological niches and consequent narrowing of ecological opportunity , leaf buds, and occasionally scavenging and feeding on blood. Compared with their primarily granivorous ancestors, Darwin's finches occupy an extensive and, importantly, central portion of the beak shape morphospace Figure . This pa4.3Tiaris grassquits, which specialize on grass seeds, colonized the Galápagos Islands and diversified to produce all modern‐day Darwin's finches , Vegetarian Finch (Platyspiza crassirostris), and warbler finches. The range of morphological diversity in Coerebinae therefore results from an interplay between intrinsic and extrinsic factors, with some lineages apparently shaped more by evolvability and others by ecology.Our study explores the patterns of morphological and dietary variation to better understand the relative significance of extrinsic and intrinsic factors in the diversification of Darwin's finches and other similar adaptive radiations. It has long been known that the ancestor of the Coerebinae gave rise to a multitude of morphologically and ecologically distinct forms both on the Caribbean and Galápagos archipelagos, and our analyses confirm that other avian colonists to the Galápagos have not undergone such dramatic modifications. We argue that the ability of Coerebinae to generate multiple new ecomorphs resulted partly from intrinsic factors, perhaps including increased flexibility in their craniofacial developmental program which probably predated their expansion to Galápagos. Nonetheless, environment‐driven factors, such as ecological constraints and purifying selection imposed by available trophic niches, must have played an important role. In particular, a narrow arena of ecological opportunity can help to explain low rates of diversification among ecological specialists in Coerebinae, including Bananaquit (5Our finding that two closely related clades of tanagers (Coerebinae and Sporophilinae) have undergone high rates of net diversification despite markedly different evolutionary histories in contrasting geographical settings suggests that rapid morphological diversification is not necessarily coupled with a particular mode of speciation. The most obvious difference is that Coerebinae speciation is associated with rapid changes in beak morphology linked to partitioning of ecological resources. Members of Sporophilinae have experienced higher net diversification rates with much lower morphological diversification. The Darwin's finches occupy a far larger area of the beak morphospace relative to that of other Galápagos endemic clades evolving novel beak morphologies in addition to forms similar to those of their Caribbean relatives. While ecological and biogeographic factors no doubt play an important role, our findings support the hypothesis that the ancestor of Darwin's finches arrived on the Galápagos Islands already endowed with the genetic propensity to produce the high levels of beak variation needed to explore new dietary niches. A similar combination of increased developmental variability and availability of new ecological opportunities may apply to other adaptive radiations in passerine birds . Future research should focus on the evolution of developmental genetic programs, including that controlling beak morphology, ideally comparing both quickly diversifying and morphologically conservative clades, to determine how changes in genetic mechanisms contribute to avian diversification.The authors declare no competing interests.Ashley Reaney: Conceptualization ; Data curation ; Formal analysis ; Investigation ; Methodology ; Visualization ; Writing‐original draft (lead); Writing‐review & editing . Yanis Bouchenak‐Khelladi: Conceptualization ; Formal analysis ; Methodology ; Validation ; Visualization ; Writing‐review & editing . Joseph Andrew Tobias: Conceptualization ; Data curation ; Investigation ; Methodology ; Validation ; Visualization ; Writing‐review & editing . Arkhat Abzhanov: Conceptualization ; Funding acquisition ; Investigation ; Methodology ; Project administration ; Resources ; Supervision (lead); Validation ; Visualization ; Writing‐original draft ; Writing‐review & editing .App S1Click here for additional data file.Data S2Click here for additional data file.App S3Click here for additional data file.Data S4Click here for additional data file.


This study continued the development of an isiZulu speech reception threshold (zSRT) test for use with first language, adult speakers of isiZulu.The objective of this study was to determine the convergent and concurrent validity of the zSRT test.One hundred adult isiZulu first-language speakers with normal hearing and 76 first-language, adult isiZulu speakers with conductive or sensorineural hearing losses ranging from mild to severe were assessed on pure tone audiometry and a newly developed isiZulu SRT test. Convergent validity was established through agreement of the zSRT scores with pure tone average (PTA) scores. Concurrent validity was assessed by examining the steepness of the psychometric curve for each word in the zSRT test for each type and degree of hearing loss.Intraclass correlation coefficient analyses showed zSRT scores were in substantial to very high agreement with PTA scores for the normal hearing and hearing loss groups . The mean psychometric slope (%/dB) at 50% correct perception for all words in the zSRT test was 4.92%/dB for the mild conductive hearing loss group, 5.26%/dB for the moderate conductive hearing loss group, 2.85%/dB for the moderately severe sensorineural hearing loss group and 2.47%/dB for the severe sensorineural hearing loss group. These slopes were appropriate for the degree of hearing loss observed in each group.The zSRT test showed convergent and concurrent validity for assessing SRT in first language, adult speakers of isiZulu. This article reports on the convergent and concurrent validity of an isiZulu speech reception threshold (zSRT) test for use with first-language, adult speakers of isiZulu with and without hearing loss.With evidence-based practice now being prioritised for clinical practice, the need for validity evidence of newly developed tests continues to grow amongst clinicians and researchers alike  testing remains a core component of the audiological test battery  and permanent residents of KwaZulu-Natal.For part one of the present study, 100 participants with normal hearing aged 18–60 years (mean age 37 years) were recruited through advertisements and selected based on consecutive sampling , moderate conductive hearing loss , moderately severe sensorineural hearing loss  and severe sensorineural hearing loss .For part two of the present study, 76 adults with hearing loss between the ages of 21 and 59 years (mean age 39 years) were purposely sampled from the clinical databases of two audiology departments in two provincial hospitals and the audiology clinic of a university in EThekwini, Kwa Zulu-Natal, South Africa. The hearing status of each participant was identified from their hospital or clinical database records and confirmed by audiometric assessment at the research facility. These 76 participants were grouped by the degree of hearing loss in their better hearing ear. This created four groups: mild conductive hearing loss . This audiologist was the first-language speaker of isiZulu.All tests were performed in an isolated Industrial Acoustics Company twin audiometric soundproof booth of double wall construction meeting ANSI  standardTesting occurred in two sessions. Participants in part one of the present study only participated in testing session one. Participants in part two of the present study participated in testing sessions one and two.In the first session, all participants completed pure-tone audiometry, tympanometry and SRT testing using the zSRT test. Pure tone audiometry was conducted at octave frequencies from 250 Hz to 8000 Hz using the modified Hughson-Westlake threshold technique . The task was to repeat each word as heard.All 28 words on the zSRT test were first played at 20 dB SL to familiarise each participant with the words, and then in a randomised order at 35–70 dB HL  in 5 dB steps for participants with mild conductive hearing loss, at 40–75 dB HL  in 5 dB steps for participants with moderate conductive hearing loss, 50–85 dB HL  in 5 dB steps for participants with moderately severe sensorineural hearing loss and at 70–95 dB HL  in 5 dB steps for participants with severe sensorineural hearing loss. Participants were given two 5-min rest breaks during this testing to improve co-operation and reduce fatigue. All participant responses were scored by two scorers (the researcher and the isiZulu-speaking audiologists).The PTA  and zSRT scores for both participant groups were confirmed as meeting parametric assumptions by inspecting their histograms, box and whisker plots, and Q-Q plots (data not shown). Agreement between the PTA and zSRT results for each participant group separately, for right and left ears separately, were assessed using intra-class correlation coefficient (ICC) analyses  was calculated. This analysis was not completed for the normal hearing participant group as it had been completed on a separate sample of participants with normal hearing and reported elsewhere  and subsequent annual clearance was obtained during the data collection period. Informed consent was obtained from all participants prior to their participation in the study.For part one of the present study, p < 0.001) difference between PTA and zSRT was observed for each participant group.For part two of the present study, The present study’s results suggest that the zSRT test is a valid measure of SRT in first language, adult speakers of isiZulu with and without hearing loss.The convergent validity of the zSRT test was supported by the very high agreement  observed between participant PTA and zSRT values for participants with and without hearing loss, for both right and left ears. This support is drawn from this study’s use of the zSRT and PTA scores as measures of the theoretical construct of hearing threshold. The degree to which the zSRT and PTA measures of this construct were related suggests high convergent validity. The zSRT results can reasonably be expected to show high levels of agreement with the established PTA results for first-language, adult speakers of isiZulu with hearing loss, when these tests are applied in clinical settings.p < 0.005) higher than the PTA values by an average of 2.9–3.0 dB for participants with normal hearing and by an average of 2–3 dB for participants with hearing loss. Whilst statistically significant, these differences were not considered to be clinically significant given the minimum step size for presentation level used in this study was 5 dB, and differences 6 dB or less between SRT and PTA having been taken to indicate agreement between those two measures for tests in several languages (Carhart, Whilst the zSRT and PTA values were in very high agreement in this study’s participants, mixed model analyses showed the zSRT values to be significantly (Carhart, .The concurrent validity of the zSRT test was supported by the psychometric functions for each word in the zSRT test being consistent with the degrees and types of hearing loss shown by the study’s participants with hearing loss by subgroup, that is, mild conductive, moderate conductive, moderately severe sensorineural and severe sensorineural hearing loss. For increasing degrees of hearing loss, the psychometric functions showed the expected increases in threshold and decreases in slopes (Wilson & Carter, A closer inspection of the psychometric functions of individual words in the isiZulu test across the four hearing loss groups indicates some variability in curves amongst individual words. For example, the psychometric functions for the words /wina/ and /minya/ were consistently very shallow for all degrees and types of hearing loss. This suggests these words could be lowering the concurrent validity of the zSRT test overall and the removal of these words from the zSRT test wordlist could be considered.The present study’s findings further support the potential use of the zSRT test as a valid and reliable measure of SRT in first-language, adult speakers of isiZulu speakers. This zSRT test has now been shown to contain words that are linguistically familiar and homogenous in audibility in isiZulu-speaking adults (Panday et al., The present and previous studies reporting the development of this zSRT test also support the use of rigorous, systematic and multiple methods particularly based on a unified validation framework to accumulate evidence for the validity and reliability of any new test. Previous attempts to use only one or two methods to confirm the validity and/or reliability of new tests should not be considered sufficient for such purposes. It is important to view multiple sources of evidence regarding the validity and reliability on new tests before such tests can be properly considered for clinical use.The present study’s results suggest the zSRT test is a valid measure of SRT in first-language, adult speakers of isiZulu with and without hearing loss, showing both convergent and concurrent validity when used in these populations.This study’s findings are limited by its participants being predominantly recruited from eThekwini and surrounding regions in KwaZulu-Natal, South Africa, although the test words did represent a central dialect of isiZulu. It could also be limited by its use of the SRT method described by Chaiklin and Ventry  with mod


The mainstay of treatment for idiopathic sudden sensorineural hearing loss (SSNHL) includes oral steroids, intratympanic steroid injections or a combination of both. The National Institute for Health and Care Excellence, in their recent hearing loss guidelines, highlighted the paucity of evidence assessing the comparative effectiveness of these treatments; and the National Institute for Health Research (NIHR) Health Technology Assessment Programme has since released a commissioned call for a trial to identify the most effective route of administration of steroids as a first-line treatment for idiopathic SSNHL. For such trials to be run effectively, reliable information is needed on patients with SSNHL: where they present, numbers, demographics, treatment pathways, as well as outcomes. This study will collect these data in a nationwide cohort study of patients presenting with SSNHL across 97 National Health Service (NHS) trusts. The study will be delivered through ear, nose and throat (ENT) trainee networks, the NIHR Clinical Research Network (CRN) Audiology Champions and the NIHR CRN. Importantly, this study will also provide a dataset to develop a prognostic model to predict recovery for patients with idiopathic SSNHL. The study objectives are to: (1) map the patient pathway and identify the characteristics of adult patients presenting to NHS ENT and hearing services with SSNHL, (2) develop a prognostic model to predict recovery for patients with idiopathic SSNHL and (3) establish the impact of idiopathic SSNHL on patients’ quality of life (QoL).Study design: national multicentre prospective cohort study across 97 NHS trusts.Inclusion criteria: adult patients presenting to NHS ENT and hearing services with SSNHL.Outcomes: change in auditory function; change in QoL score.Analysis: multivariable prognostic model, using prespecified candidate predictors. Mean change in QoL scores will be calculated from initial presentation to follow-up.Health Research Authority and NHS Research Ethics Committee approved the study. Publication will be on behalf of study sites and collaborators.NCT04108598).ClinicalTrials.gov Registry ( Taking place across 97 National Health Service trusts, the SeaSHeL Study represents the first nationwide prospective cohort study of adults presenting to ear, nose and throat (ENT) and hearing services with sudden sensorineural hearing loss (SSNHL).It is uniquely delivered by National ENT Trainee Research Network, Student and Foundation Doctors in Otolaryngology, National Institute for Health Research (NIHR) Clinical Research Network (CRN) Audiology Champions and the NIHR CRN.We acknowledge that duration between onset of SSNHL and presentation to ENT services as well as follow-up times and treatment strategies will vary across centres and cases, which may impact measured outcomes; however identifying differences in the patient pathway will form an important study finding.Each year approximately 20 people per 100 000 experience a sudden hearing loss in one ear that is located in the inner ear.3 4Idiopathic SSNHL is a serious condition, adversely impacting people’s lives2 8 9Current care pathways for patients suffering from SSNHL appear to vary considerably in terms of the type of service patients first present to, their subsequent referral, length of time between onset of symptoms, presentation and start of treatment, the treatment plan as well as follow-up. The mainstay of treatment options for idiopathic SSNHL include oral steroids, intratympanic steroid injections or a combination of both.11For such a trial to be run effectively, reliable information is needed on patients with SSNHL: where they present, numbers, demographics, treatment pathways, as well as outcomes. This study will collect these data in a nationwide prospective cohort study of adult patients presenting with SSNHL across 97 National Health Service (NHS) trusts. The study will be delivered through National ENT Trainee Research Network (INTEGRATE), Student and Foundation Doctors in Otolaryngology (SFO-UK), NIHR Clinical Research Network (CRN) Audiology Champions, and the NIHR CRN. Importantly, this study will not only provide key data to inform trial design and delivery, but also a unique dataset to develop a prognostic model to predict recovery for patients with idiopathic SSNHL.To map the patient pathway and identify the characteristics of adult patients presenting to NHS ear, nose and throat (ENT) and hearing services with SSNHL.To develop a prognostic model to predict recovery for patients with idiopathic SSNHL.To establish the impact of idiopathic SSNHL on patients’ quality of life (QoL).National multicentre prospective observational cohort study. The study will be reported in accordance with Strengthening the Reporting of Observational Studies in Epidemiology and ‘Transparent Reporting of a multivariable prognostic model for Individual Prognosis or Diagnosis’ reporting guidelines for observational studies.12A multicentre study taking place across 97 NHS trusts providing ENT and hearing services.Adult patients (man or woman) aged over 16 years of age presenting to NHS ENT and hearing services with a history of sudden hearing loss (within a 72-hour window) that is sensorineural in nature.There is a decrease in hearing of ≥30 dB affecting at least three consecutive frequencies , using a premorbid pure tone audiogram as a baseline reference. Or should a baseline pure tone audiogram not be available, the unaffected ear can be used a baseline reference. Or should both ears be affected by hearing loss, a normative reference (ISO7029)Willing and able to provide written informed consent.Patients with mixed or conductive hearing loss (CHL) in ear(s) with the sudden hearing loss. CHL will be defined as a ‘true’ air-bone gap of 15 dB or more in 3 or more contiguous frequencies, between the following specific frequencies: 500 Hz, 1000 Hz, 2000 Hz and 4000 Hz.et al.3 4The minimum sample size for developing a multivariable prognostic model to predict recovery for patients with idiopathic SSNHL was estimated using the method reported by Riley Ninety-seven NHS trusts across the UK have been identified as study sites via INTEGRATE, SFO-UK, NIHR CRN Audiology Champions and the NIHR CRN. An audit carried out in preparation of this study at five NHS trusts, over 6 months, revealed an average presentation rate of one patient with idiopathic SSNHL per trust per month. It appears therefore that our recruitment target over the study duration (32 months) is feasible.Age; gender; social class; presence of vestibular symptoms; precipitating illness; pattern of hearing loss; severity of hearing loss; time between onset of symptoms and treatment; treatment(s) received.Pattern of hearing loss will be defined as: low frequency loss , mid-frequency loss , high-frequency loss  and all frequencies. Severity of hearing loss (on presentation) will be defined as follows: pure tone average (PTA) across six frequencies , classified as: mild (25–40 dB loss), moderate (41–70 dB loss), severe (71–95 dB loss) and profound (>95 dB loss).The change in auditory function in the affected ear from initial presentation to follow-up (at any one time between 6 and 16 weeks from onset of symptoms). Auditory function will be defined as the PTA of air conduction thresholds between the following specific frequencies: 250 Hz, 500 Hz, 1000 Hz, 2000 Hz, 4000 Hz and 8000 Hz. If multiple pure tone audiograms have been carried out between 6 and 16 weeks, the most recent pure tone audiogram will be used for the calculation of auditory function.Change in auditory function classified as:Full recoveryFinal PTA in affected ear within 10 dB (≤10 dB) of PTA of baseline pure tone audiogram.OR should a baseline pure tone audiogram not be available:Final PTA in affected ear within 10 dB of PTA of unaffected ear.OR should both ears be affected with hearing loss:Final PTA in affected ear within 10 dB of a normative reference (ISO7029).Partial to no recoveryFinal PTA in affected ear ≥10 dB of PTA of baseline pure tone audiogram.OR should a baseline pure tone audiogram not be available:Final PTA in affected ear ≥10 dB of PTA of unaffected ear.OR should both ears be affected with hearing loss:Final PTA in affected ear ≥10 dB of PTA of a normative reference (ISO7029).131. Degree of change in auditory functionComplete recoveryFinal PTA in affected ear within 10 dB (≤10 dB) of PTA of baseline pure tone audiogram.OR should a baseline pure tone audiogram not be available:Final PTA in affected ear within 10 dB of PTA of unaffected ear (≤10 dB).OR should both ears be affected with hearing loss:Final PTA in affected ear within 10 dB of a normative reference (ISO7029).13Marked recoveryPTA improvement ≥30 dB .OR should a baseline pure tone audiogram not be available:PTA improvement ≥30 dB .OR should both ears be affected with hearing loss:PTA improvement ≥30 dB ).13Slight recoveryPTA improvement ≥10 dB and <30 dB .OR should a baseline pure tone audiogram not be available:PTA improvement ≥10 dB and <30 dB .OR should both ears be affected with hearing loss:PTA improvement ≥10 dB and <30 dB ).13No improvementPTA improvement <10 dB .OR should a baseline pure tone audiogram not be available:PTA improvement <10 dB .OR should both ears be affected with hearing loss:PTA improvement of <10 dB ).132. Change in QoL score from initial presentation to follow-up at any one time between 6 and 16 weeks following treatment. QoL will be measured with the Hearing Handicap Inventory for Adults (HHIA) (for patients under 65 years of age) or Hearing Handicap Inventory for Elderly (HHIE) (for patients over 65 years of age) and the Health Utility Index Mark 3 (HUI3). QoL data will be collected from a subset of 20 trusts.Out-of-limit thresholds obtained on the pure tone audiogram will be classified as a 96 dB loss at that specific frequency.We will develop a prognostic model to predict recovery for patients with idiopathic SSNHL and assess the impact of idiopathic SSNHL on patients’ QoL. A multivariable prognostic model will be developed using the candidate predictors specified above n=15). Missing outcome data at the end of the study will be imputed using multiple imputation by a chained equations procedure.. Missing16 17The mean change in HHIA, HHIE and HUI3 scores across patients with idiopathic SSNHL will be calculated from initial presentation to follow-up (any one time between 6 and 16 weeks). We will use the non-parametric Wilcoxon and McNemar–Bowker tests with a significance level of 5%. Statistical analysis will be carried out using R V.3.6.2 .A summary of study procedures is outlined in Box 1General information.Patient demographics and socioeconomic status.Medical history and social history.Patient’s first presentation to ear, nose and throat (ENT).Hospital and service patient presenting to.Date of presentation.Referral pathway and treatments received up until first presentation to ENT.Clinical history of sudden sensorineural hearing loss (SSNHL).Examination findings.Audiometry results.Imaging requests/results.Laboratory test requests/results (data on any laboratory test requested for a patient with SSNHL to be collected).Vestibular function tests/results (data on any vestibular function test requested for a patient with SSNHL to be collected).Hearing Handicap Inventory for Adults (HHIA) Questionnaire score OR Hearing Handicap Inventory for Elderly (HHIE) Questionnaire score.*Health Utility Index Mark 3 (HUI3) Questionnaire score.*Treatment plan.Cause of SSNHL identified.Follow-up details.Follow-up (6–16 weeks after onset of symptoms).Date of presentation.Examination findings.Audiometry results.Imaging requests/results.Laboratory test results (data on any laboratory test requested for a patient with SSNHL to be collected).Vestibular function results (data on any vestibular function test requested for a patient with SSNHL to be collected).HHIA Questionnaire score OR HHIE Questionnaire score.*HUI3 Questionnaire score.*Treatments provided since first visit to ENT.Cause of SSNHL identified.Adverse events.*To be collected at quality of life sites only (20 of 97 sites).Patient follow-up data will be collected at 6–16 weeks from onset of symptoms. All data collected will be uploaded onto REDCap, a secure online data collection platform.Three patients suffering from SSNHL were involved in the design of this study from its inception. They were recruited from a database of patients interested in taking part in Patient and Public Involvement activities. They reviewed and gave input on study objectives and design, including reviewing participant information sheets and consent forms to ensure clarity. This group will also be involved in monitoring study progress and in its dissemination including coauthoring a lay summary report.Health Research Authority and NHS Research Ethics Committee has approved the study. IRAS project ID: 258494; REC reference: 19/NW/0556.Risks, Burdens and BenefitsThere are no anticipated risks or direct benefits to participants.Results will be presented at local and inter meetings and published in the scientific and grey literature for the attention of professional and scientific audiences. Publication and presentation of the final results will be on behalf of all study sites and collaborators. A lay summary report will be published for patients and members of the public.Start date: 7 October 2019.Planned end date: 1 May 2022.


The current study investigated role of telocytes (TCs) in angiogenesis during embryonic development of quail using immunohistochemistry (IHC), transmission electron microscopy (TEM), and scanning electron microscopy (SEM). The angiogenic apparatus consisted of TCs, endothelial cells, and macrophages. TCs were identified morphologically by their telopodes and podoms using TEM and SEM and immunohistochemically using CD34, and vascular endothelial growth factor (VEGF). TCs also expressed CD68. TCs formed a three-dimensional network and established direct contact with blood vessels, sprouting endothelial cells, and active macrophages, while exerting their effect through paracrine signaling. VEGF was also expressed by endothelial cells and macrophages. Matrix metalloproteinase–9 (MMP-9) was expressed by TCs, endothelial cells, and macrophages. In conclusion, the expression of VEGF by TCs, endothelial cells, and macrophages is required for the proliferation and migration of endothelial cells and vascular growth. The expression of MMP-9 by TCs, endothelial cells, and macrophages is essential for the degradation of extracellular matrix (ECM) components during neoangiogenesis. Macrophages may facilitate phagocytosis and elimination of the degraded ECM components. TCs consist of a cell body and telopodes (TPs), which contain focal dilatations and thin podomeres. TPs are the unique features that distinguish TCs by electron microscopy2. The cell body contains mitochondria, small Golgi apparatus, and smooth and rough endoplasmic reticulum4. Podoms contain endoplasmic reticulum, mitochondria, caveolae, and secretory vesicles4.Telocytes (TCs) are a unique type of interstitial cells. They play a critical role during tissue development and hemostasis5.Angiogenesis is a physiological process through which blood vessels grow. During embryogenesis, two mechanisms of angiogenesis occur: sprouting angiogenesis and intussusceptive angiogenesis. The latter occurs during splitting of the blood vessels in current vascular tissue. Sprouting angiogenesis is a well-known type in which endothelial sprouting occurs in response an angiogenic stimulus . Sprouting is involved in the angiogenesisof tissues devoid of blood vessels. Endothelial sprouting requires sequential steps, including enzymatic degradation of capillary basement membrane, endothelial cell (EC) proliferation, directed migration of ECs, tubulogenesis, vessel fusion, vessel pruning, and pericyte stabilization7. They express VEGF, which facilitates angiogenesis2. The current study aims to investigate the role of TCs in angiogenesis during embryonic development of quail bird using immunohistochemistry (IHC), transmission electron microcopy (TEM), and scanning electron microscopy (SEM).The role of TCs angiogenesis is described in many tissues and organs during development and tissue repairCoturnix japonica) eggs were used in the current study. The source of the eggs was the Farm of Department of Histology, Faculty of Veterinary Medicine, South Valley University, Qena, Egypt. The egg incubation protocol was carried out according to Soliman8. Eggs were incubated at 37.5 °C at a relative humidity 65%. Quail embryos were collected from days 5 and 8 (five for each day). The eggs were kept at − 20 °C for 4 h and were opened from the wide end, with embryos carefully excised from the egg shells. The embryos were placed on a Petri dish and fixed after washing with saline 0.9% NaCl. Sample collection was conducting in accordance with the guidelines of the Institutional Ethical Committee of Veterinary Medicine, South Valley University, Egypt, and following the Egyptian animal laws.Fertile quail .ve Table  for 24 h12 and the solution of the Ultra Vision Detection System . Procedures were performed according to the manufacturer’s instructions15. The Richert Leica RM 2125 microtome was used to cut 5-µm paraffin sections. The sections were dewaxed by xylene, hydrated by ascending grades of alcohol, and washed by phosphate-buffered saline   three times for 5 min. Blocking of the endogenous peroxidase was carried out using drops of 3% hydrogen peroxide in methanol for 20 min at room temperature. The section was thoroughly rinsed using running tap water for 10 min. Antigen retrieval was performed by applying 10-mm sodium citrate buffer . The neck skin was carefully excised and trimmed to a small-sized length of 2.0 to 3.0 mm. Karnovsky fixative (Table 21 and Periodic acid–Schiff (PAS)22. Staining of semithin sections was performed after dissolving the resin using a saturated alcoholic solution of sodium hydroxide. The stained sections were examined by a Leitz Dialux 20 microscope and a Canon digital camera (Canon PowerShot A95).Resin-embedding technique was performed using Karnovsky’s fixativeUltra-thin sections (60 nm) of resin-embedded samples at embryonic days 5 and 8 were cut using the Reichert ultra-microtome. The sections were stained using uranyl acetate followed by lead citrate for 15 min for each stain. The stained grids were examined using a JEOL100CX II transmission electron microscope at the central laboratory of South Valley University, Egypt.The samples were fixed in Karnovsky’s fixative and washed. Sample washing was performed using the Na–phosphate buffer (pH 7.3) for four times at 15 min. Samples were postfixed in 1% osmic acid in 0.1 M Na–phosphate buffer for an additional 2 h at room temperature and washed in Na–phosphate buffer. The samples were dehydrated by ascending grades of alcohol  for 30 min at each concentration and 100% for 2 days with many changes. Samples were treated with isoamyl acetate for 2 days and dried using the critical-point drying method with the Critical Point Drying Procedure Polaron E3000 CPD apparatus (Germany). Sample coating with gold was performed using the JEOL 1100 E-ion sputtering device (Japan) and examined with a JEOL SEM (JSM 5500 LV) at 10 kV, at the central laboratory of South Valley University, Egypt.31.To distinguish different types of cells and structures, coloring of TEM and SEM images was performed using the Photo Filter 6.3.2 program. The black and white image was colored using adjust then color balance to change the color to a desirable colors and degrees. Coloring transfer was performed by the stamp tool located at the right panel. Changing color was performed again using adjust then color balance to color another cell or structure. The methods used were previously described by many authors32. Arrival guidelines the study was carried out in compliance with the ARRIVE guidelines33.The National Ethics Committee of South Valley University and veterinary authorities in South Valley University Province, Egypt, approved the method of this study. ‘All procedures were performed in accordance with the relevant guidelines and regulationsTCs were recognized around blood capillaries Fig. A, large By SEM, TCs were distinguished by TPs that formed a 3D network around blood vessels Fig. A,B,D–F a34, proliferation, and inhibition of differentiation of stem or progenitor cells35.The current study investigated the role of TCs during the early stages of angiogenesis in embryonic quail. Using TEM, SEM, and IHC, typical TCs were identified forming a 3D network in the neck skin. TCs had strong immunoaffinity for CD34 CD34 is a member protein of the transmembrane sialomucin protein family and is used for the identification of hematopoietic stem cells and non-hematopoietic progenitors, including vascular endothelial progenitors and embryonic fibroblasts, multipotent mesenchymal stromal cells, interstitial dendritic cells, and epithelial progenitors. Functional implications of CD34 are linked to cell adhesion36. The findings of the current study suggest the role of TCs in angiogenesis, which is supported by previous studies. They secrete VEGF and endothelial growth factor, which regulate angiogenesis and the proliferation of ECs6. The angiogenic role of TCs is also noted during myocardial infarction. TCs produce angiogenic microRNAs, such as let‐7e, 10a, 21, 27b, 100, 126‐3p, 130a, 143, 155, and 50337.Perivascular TCs formed a heterocontact with the blood vessels and release of secretory vesicles for paracrine singling. TCs established contact with sprouting ECs, which indicated neovascularization. They also expressed VEGF, which is essential for the development and growth of the circulatory system and for the regulation of proliferation and migration of vascular ECs38. The relation between TCs and macrophages may reveal that TCs involved in the phagocytic activities via indirect pathway. The effect of TCs on the activation of macrophages has been studied using an in vitro coculture of peritoneal macrophages (pMACs). The authors showed that communication between pMACs and TCs occurs through heterocellular junctions and the paracrine mode, and they suggest that activation of pMACs occurs via the mitochondrial signaling pathway39.TCs communicated through direct contact and paracrine signaling with active macrophages. Avian macrophage identified by their rounded profile and contained large phagosomes with materials of different stages of digestion40. Based on the biochemical properties, MMP subtypes are categorized as collagenases, gelatinases, stromelysins, and membrane-type MMPs (MT-MMPs)41. Gelatinases are gelatinase A (MMP-2) and gelatinase B (MMP-9)42. MMP-9 plays a critical role in angiogenesis, immune cell migration, activation of cytokines and chemokines, and progression and metastasis of cancer cells45. MMP-9 degrades collagen types IV, V, XIk´, XIVl´, elastin, aggrecan, link protein, decorinr, lamininn, entactin, SPARCq, myelin basic proteinm, ∞2Mn, ∞1Pli, IL-1βj, and proTNF-∞k47. In the current study, TCs exhibited proteolytic activities that had a strong immunoaffinity for MMP-9. MMP-9 plays an essential role in the steps of angiogenesis and degrades the basement membrane of capillaries. MMP-9 also promotes EC migration43. Vascular pruning requires post hypoxia activation of MMP-948. CD68 is a sialomucin belongs to class D scavenger receptor. CD68 protein is common in late endosomes and lysosomes. Thus, CD68 protein is found in the granules of macrophages, and other cells of the mononuclear phagocyte system such as Kupffer in the liver, microglial cells in the brain, Hofbauer cell in the placenta, osteoclast in bone49. CD68 is also expressed by other immune cells including dendritic cells, neutrophils, basophils, and mast cells, activated T-cells and some proportion of mature B-cells as well as epithelium of renal tubules50. Increased CD68 expression in macrophage is associated with high vascularity51. In the current study, TCs expressed CD68 as well as macrophages. Stromal cells/telocytes express CD68 in the human adult trigeminal ganglion52. Cd68 is one of the complement Receptors that expressed by antigen presenting cells. Complement Receptors are implicated in in cell migration and phagocytosis and immune regulation. Complement regulatory proteins that expressed by antigen presenting cells have an essential role in limiting cell activation. Complement receptors cooperate with different receptors to regulate myeloid cell responses53.MMP, or matrixins, are a type of endopeptidases. They are specialized to degrade the extracellular matrix components and other proteins and contribute to tissue remodeling54. Pericyte has an essential role during angiogenesis. Development of the endothelial tube is associated with pericytes, that the endothelial sprouting act as a migration sign. pericytes are derived from the differentiation of resident mesenchymal precursors or migration from the adventitia of the adjacent vessel55. This process is known as pericyte-driven angiogenic process in which endothelial cells are preceded and guided by migrating pericytes56. pericytes may inhibit endothelial growth and migration58. Previous studies revealed a remarkable correlation between pericyte contribution and microvessel stabilization60. pericyte investment is linked to maintenance of capillary integrity in vivo61.Formation of vascular basal lamina and investment of the pericytes occurs at the end of the proliferative stage and the onset of the mature or quiescent stage of capillary activity62. endothelial cell-pericyte interactions stimulate upregulation of basal lamina-encoding genes and proteins, including fibronectin laminin, and integrins63. On the other hand, pericytes invasion may establish tubes and enhance the subsequent penetration of endothelial cells64. NG2 expressing pericyte progenitors located in close proximity to blood vessels located closed to blood vessels65. Thus, the reciprocal interaction between endothelial cells and pericytes has a fundamental role in angiogenic process.In the current study, TCs formed 3D network at the site of neovascularization. Nehls and his colleagues noticed that the initial endothelial sprouting may not depend on pericyte. Despite of pericyte is one of the initiative types of cells that invade the nascent vascular tissue and distribute at the tip of the growing endothelial sproutsIn conclusion, TCs facilitate angiogenesis. They express VEGF, which promotes endothelial proliferation and migration. They also release MMP-9, which plays a critical role in the degradation of the capillary basement membrane, EC migration, and vascular pruning.Supplementary Information


Captioning is the process of transcribing speech and acoustical information into text to help deaf and hard of hearing people accessing to the auditory track of audiovisual media. In addition to the verbal transcription, it includes information such as sound effects, speaker identification, or music tagging. However, it just takes into account a limited spectrum of the whole acoustic information available in the soundtrack, and hence, an important amount of emotional information is lost when attending just to the normative compliant captions. In this article, it is shown, by means of behavioral and EEG measurements, how emotional information related to sounds and music used by the creator in the audiovisual work is perceived differently by normal hearing group and hearing disabled group when applying standard captioning. Audio and captions activate similar processing areas, respectively, in each group, although not with the same intensity. Moreover, captions require higher activation of voluntary attentional circuits, as well as language-related areas. Captions transcribing musical information increase attentional activity, instead of emotional processing. It is widely accepted that music produces emotional responses, being one of its defining features , (Phone). In the case of music, information of the type of music, sensation transmitted, and identification of the piece  must be included, e.g., (Rock music), (Horror music), . This verbal representation does not include the emotional information of sounds and music  measurements before emotional motor response by means of EEG. The focus of the present study is to examine what happens just before a motor response (associated to emotion detection) while watching videos with audio or captions in two groups of participants: normal hearing and deaf or c subjects.The decision of using the ERPs prior to motor response is based on the following results: the emotional and cognitive networks involved in decision making can be tracked by ERPs , 24.1% with a high school degree, 27.6% with a college degree, and 48.3% with post-graduate studies.In one group, 16 participants with self-reported normal hearing were recruited, eight females and eight males, aged between 20 and 60 , 38.5% with a high school degree, 38.5% with a college degree, and 23.1% with post-graduate studies.In the other group, 13 participants with self-reported hearing loss were recruited, seven females and six males, aged between 20 and 60  EEG recordings, obtained using a custom-designed electrode Neuroscan cap and an ATI EEG system (Advantek SRL). Impedances were kept under 5 kΩ. Additional channels were included to monitor eye movement . The reference electrodes were placed on the mastoids, and the ground electrode was placed on the forehead. Data were processed to an average reference following acquisition with a band-pass filter of 0.05–30 Hz and a sample rate of 512 Hz. An artifact rejection criterion of 100 μV was used to exclude eye blinks. Individual subject averages were visually inspected to insure that clean recordings were obtained. Eye and muscle movement artifacts were identified off-line on a trial-by-trial basis through visual inspection, and they were removed prior to data averaging and ERP analysis. Noisy channels were sparingly replaced with linear interpolations from clean channels (around 6 ± 3.5 channels per record and subject). From the remaining artifact-free trials, averages were computed for each participant and each condition. The analysis epochs for ERPs were 500 ms before motor response. EEG analysis was carried out on frequent (non-target) trials to avoid contamination by motor-related neural activity associated with making a response. ERPs obtained were averaged separately for each condition and each subject. A Bayesian Model Averaging (BMA) analysis over all electrodes was performed by opening a time window of −20 to +20 ms around the highest negative amplitude peak measured in Cz electrode.Participants were cited in individual sessions. They were first asked to fill in a survey including questions about their age, gender, education level, type, and degree of hearing loss and hearing aids if applicable.Then, they were asked to sit in an armchair facing a 17″ screen with speakers placed at 1.5 m in front of the participant. They were asked to remove their hearing aids, but to keep their glasses on if needed. The 128-EEG cap was fixed to their head. A press button was placed under their left hand. Participants were explained that they were going to watch a video and were asked to press the button grabbed in their left hand whenever they felt any emotion while watching the video. Lights in the room were turned off, and the corresponding video was launched. Normal hearing participants watched the video with soundtrack (audio and captioned sequences), while participants with hearing loss watched the video without soundtrack (muted and captioned sequences). The press button in their left hand was connected with one of the computers, and each pressure was transmitted and registered as a mark in the EEG track.We registered the total number of times each participant pressed the button with their left hand indicating they were feeling an emotion. The scores were registered for three conditions: Audio (pressures occurring during audio fragments display), Caption (pressures occurring during captioned fragments display), and Mute (pressures occurring during muted fragment display).Nonparametric Mann–Whitney tests were used to compare the number of button presses in the different conditions and groups. The Mann–Whitney statistic was selected as the Shapiro–Wilk test rejected normality in some conditions, and the size of the samples was not sufficiently large (less than 20) to assume the normal distribution in the rest of the conditions. The results are shown in p = 0.00528) in the scores for Audio (19.87 ± 9.63) and Caption (9.81 ± 9.69) conditions, suggesting that video auditory stimuli produced a number of emotional reactions twice as much as the captions stimuli did. A second Mann–Whitney test was conducted to compare the number of button presses in Caption and Mute conditions in the hearing loss group. There was no significant difference (p = 0.52218) in the scores for the Mute (7 ± 4.1) and Caption (7.15 ± 4.86) conditions, suggesting that captions do not produce additional emotional reactions to the visual stimuli.A Mann–Whitney test was conducted to compare the number of button presses in Caption and Audio conditions in the normal hearing group. The standardized results show a significant difference (p = 0.79486) in the scores for the hearing group (9.81 ± 9.69) and the hearing loss group (7.15 ± 4.86), suggesting that the emotional reaction to captions is similar in both groups.Finally, a Mann–Whitney test was conducted to compare the number of button presses in Caption condition in the normal hearing group and in the hearing loss group. There was no significant difference  shows high activation in the left temporal pole (TP) lobe for Audio and Mute conditions in the hearing group and the hearing loss group, respectively. The difference between both groups concerns the magnitude obtained with the Hotelling’s T2 test that shows  greater activation of these cerebral areas in the hearing loss group  compared with the hearing group (≅500). In addition, high activation appears in the frontal inferior lobe only in the hearing loss group. For the Caption condition, activation is found in the right temporal lobe in both groups, with greater activation in the hearing loss group  compared with the hearing group .Regarding NS300 maps , 3, NS30SD of 1.27) shows a significantly greater activation in the hearing loss group. Activation in the right parietal lobe for the Audio condition appears only in the hearing group. For the caption stimuli, greater activation is found in the hearing group  compared with the hearing loss group (≅800).Regarding NS100 maps , 3, actiIn the hearing group, the average NS100 latency measured was 165.85 ms (±6.14) for the Audio condition and 161.91 ms (±8.22) for the Caption condition. In the case of NS300, the results were 377.79 ms (±50.57) for Audio and 370.08 ms (±43.99) for Caption. No significant differences were found in the NS100 or NS300 latencies between the Audio and Caption conditions in this group.In the hearing loss group, the average NS100 wave latency for the Mute condition was 116.15 ms (±19.30) and 125.84 ms (±18.17) for the Caption condition. Regarding the NS300 wave, the latency was 338.46 ms (±7.77) for the Mute and 334.15 ms (±6.15) for the Caption. No significant differences were found in NS100 or NS300 latencies between the Mute and Caption conditions in this group.p = 0.0001) was found between the hearing group (165.85 ± 6.14) and the hearing loss group (125.85 ± 18.17). For NS300, a significant difference (p = 0.00078) was likewise found between the hearing group (377.79 ± 50.57) and the hearing loss group (334.15 ± 6.15). These results suggest that the reaction time is lower in the hearing loss group for the Caption condition.Significant differences between groups where found in the Caption condition. An important result from our study is the existence of two differentiated negative ERPs, labeled NS100 and NS300, regarding the latency and cerebral activation produced prior to the motor response (button press), without significant differences between both groups. The negative ERPs close to the motor response present in anticipatory processes reflect the emotional and cognitive processing of stimuli and neurophysiological processes related to decision making  areas, meaning that images activate the same areas, and probably the same emotional and cognitive processes. The lower and middle temporal areas are associated with visual and auditory processing, object and face recognition, and word meaning , but again, the hearing loss group activated these areas with higher intensity. The shorter reaction time in the hearing loss group can be related to this higher activation.As for the laterality of the processes, we found highest left hemisphere activity for NS300 in both groups for Audio and Mute conditions, which seems to be related to attentional positive emotional processes. Other scientific studies demonstrated that the valence of emotions is represented bilaterally in our brain, emerging differentially positive in the left hemisphere and negative in the right hemisphere  and 300 ms (NS300) prior to emotional response onset. These waves were present with different intensities or in different areas in the normal hearing and hearing loss groups, demonstrating that:−Both groups mobilized temporal perception and processing areas.−The deaf group mobilized these areas with much higher intensity and adding voluntary cerebral resources  when watching muted videos without captions.−Hearing people mobilized these resources with moderate intensity levels and activated perception integration areas  when watching the same videos with audio.−The presence of captions increased and focused the activation of visual and word processing areas in both groups.On the one hand, these results indicate that when a subject with hearing loss is watching a video without captions, a higher voluntary attentional effort is needed prior to motor response compared with the normal hearing group. According to previous works, this greater energy is related to higher brain resource consumption as a consequence of hearing loss. If we add captions to the video, this attention effort increases and focuses on visual and word processing. Other studies with deaf and hard of hearing participants showed that captions cause a shift in attention from video information to captioned information (which results in an increased level of information assimilated from captions to the detriment of the information assimilated from the video sources), especially in participants with profound and severe hearing loss who rely on captions as the principal source of context information.On the other hand, these results show that auditory stimuli produce significantly more number of emotional reactions than captions. Studies on the emotional response to music showed that less than 2 s of music can produce basic emotional states as happiness, sadness, or fear, and that the emotional response depends on musical parameters such as mode, tempo, register, dynamics, articulation, or timbre. These parameters do not have a literal translation, and thus, captions transcribing information associated with non-verbal sounds generate additional intensity of attentional activity, instead of emotional response.Finally, we found that, when watching captioned videos, all the cerebral activations are produced in the right hemisphere, while when they are not present, both hemispheres interact. The explanation of this fact remains open for further research since the generally assumed approach, establishing that the right hemisphere is the one in charge of processing negative emotions, and the left hemisphere positive emotions, does not bring light to these findings.With this study, we want to contribute from a scientific basis to the enrichment of the deaf and hard of hearing people audio-visual experience. Our main conclusion is that if we want deaf people to feel the emotion produced by sounds in a similar manner as hearing people do, we need to provide other non-verbal representations of the sound, exploring other stimuli, rather than literal captions, triggering more direct emotional reactions. There is increasing research on the correspondences between the sense of hearing and the sense of touch and, thus, the potential of vibrotactile technologies to produce musical experience (Vieillard et al., The datasets generated for this study are available on request to the corresponding author.The studies involving human participants were reviewed and approved by Carlos III University of Madrid Ethics Committee. The patients/participants provided their written informed consent to participate in this study.TO and PR contributed to all aspects of the work. BR and JS-P contributed to the conception and design of the study. ML contributed to the data analysis, discussion, and article writing. All authors contributed to manuscript revision, read and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Chorioangioma is the most common type of placental tumor. A primigravida woman was noted on routine scan at 21 weeks of gestation to have a placental mass measuring 1.8 x 2.2cm. A detailed ultrasound scan revealed a well circumscribed, hypoechoic lesion protruding into the amniotic cavity; hence a preliminary diagnosis of placental chorioangioma was made and close prenatal surveillance was scheduled. At 34 weeks of gestation, the mass was measuring 6.27 x 5.38cm. The patient experienced reduced fetal movements at 37 weeks gestation. A small-for-gestational age but normal female neonate was delivered by caesarean section. Histopathological analysis of the placenta confirmed the diagnosis. According to our case, a giant placental chorioangioma may have a favorable outcome without any medical intervention. Non-trophoblastic tumors are the most common benign neoplasms of the placenta. They include chorioangioma, teratoma, leiomyoma and hepatocellular adenoma. Chorioangioma is the most common subtype . Most coA 29-year-old Caucasian primigravida woman with unremarkable medical history was noted on routine scan at 21 weeks of gestation to have a placental mass measuring 1.8 x 2.2 cm . A detaiChorioangioma is the most frequent non-trophoblastic benign tumor of the placenta, with an estimated incidence of 0.6% . MaternaDuring histopathology examination, these tumors demonstrate angiomatous and cellular matrix with degenerative changes of calcification. In larger tumors, hyaline and myxoid degeneration is noticed . Due to In summary, diagnosis of placental chorioangioma by ultrasonography is only suggestive and the final diagnosis is established upon histological examination of the tumor. Our case proves that placental chorioangioma can be asymptomatic even when the size is greater than 4 cm. Moreover, MRI was not used antenatally to observe the characteristics of the placental tumor. Hence, transabdominal ultrasound is considered an acceptable method for regular surveillance of the mass during pregnancy. Our patient had a favorable outcome without any medical intervention and delivery was performed full-term. The neonate was not admitted to the neonatal unit, which demonstrates that fetal anemia was not developed despite the large size of the chorioangioma. Early prenatal diagnosis, close monitoring and appropriate intervention are considered critical in order to reduce fetal morbidity and mortality.


The present study was to investigate the dynamics of endolymphatic hydrops (EH) and hearing function, and explore whether the hearing loss is caused by EH alone and whether the hearing function can be modulated by changes in the EH. The extent of EH visualized by gadolinium (Gd)-enhanced inner ear magnetic resonance imaging, hearing thresholds and the summating potential/action potential ratio (-SP/AP ratio) of electrocochleography (ECochG) were recorded prior to and following surgery in 22 patients with intractable Meniere’s disease (MD) who underwent endolymphatic duct blockage (EDB). The difference value of the hearing threshold and -SP/AP ratio was significantly positively correlated with the difference value of the endolymph to vestibule-volume ratio (EVVR) and grading of cochlear hydrops between prior to and following surgery. Among patients with a decreased EVVR, the average hearing threshold and -SP/AP ratio was significantly decreased following surgery, as compared to that prior to surgery. Six out of seven patients with a hearing improvement (≥10-dB decline) and 4/5 patients with a negative conversion of EcochG showed downgrading of their hydrops in the cochlea and/or vestibule. By contrast, among patients with an increased EVVR, the average hearing threshold and -SP/AP ratio tended to increase following EDB, as compared with that prior to surgery. One out of two patients with a hearing deterioration (≥10-dB elevation) showed upgrading of her hydrops in both cochlea and vestibule. The present results showed the downgrading of cochlear and/or vestibular hydrops accompanied by the downregulation of the hearing threshold and -SP/AP ratio of EcochG, as well as the upgrading of cochlear and/or vestibular hydrops that tended to upregulate the hearing threshold and -SP/AP ratio of EcochG; this suggested that hearing loss is likely to be caused by hydrops and that the impaired hearing function be modulated by changes in the hydrops. Meniere’s disease (MD) is a well-recognized but inadequately understood condition associated with the classic triad of episodic vertigo, low-tone fluctuating sensorineural hearing loss and tinnitus. Numerous studies have confirmed that virtually all cases of ‘‘true” MD present with endolymphatic hydrops (EH) in the inner ear –4. HowevIn the present study, the dynamics of EH were evaluated using gadolinium (Gd)-enhanced inner ear MRI, changes in hearing thresholds and summating potential/action potential ratio (-SP/AP ratio) of electrocochleography (ECochG) in a group of patients who had undergone endolymphatic duct blockage (EDB) for the treatment of MD; the aim was to clarify whether or not the change of hydrops is likely to modulate the hearing function and -SP/AP ratio, which would verify a cause-effect relationship between hydrops and auditory physiology.This study was conducted at a large tertiary referral center. Between January 2018 and December 2018, a total of 2,160 patients with vertigo or dizziness were referred to our otology-led vertigo clinic. According to the diagnostic criteria for MD jointly formulated by the Classification Committee of the Bárány Society [The present study was approved by the Medical Ethics Committee of the Second Xiangya Hospital (certificate number: S452). All participants provided their written informed consent in accordance with the Declaration of Helsinki. Authors had access to information that could identify individual participants during or after data collection.On the day of the examination, that is, immediately before contrast medium application and 1 day before magnetic resonance imaging (MRI), patients underwent pure tone audiometry (PTA) and extratympanic ECochG either prior to or following surgery. Following the confirmation of both cochlear and vestibular hydrops by Gd-enhanced MRI and once informed consent was obtained, the patients underwent surgery. From February 10 to March 30, 2020, a total of 22 patients were recalled for repeat PTA, ECochG and MRI scans examinations. A total of 4 patients were excluded from the study, due to lack of follow-up Gd-MRI scans, while 22 subjects  met the inclusion criteria.Hearing function was measured by a pure-tone audiometer and was evaluated based on the four-tone average air-conductive hearing threshold formulated by (a+b+c+d)/4; a, b, c and d represent hearing levels at 0.25, 0.5, 1.0 and 2.0 kHz, respectively. Changes in the hearing levels were defined as worse (≥10-dB elevation), better (≥10-dB decline) or same (-10 dB< x <10 dB).The silver-ball electrode for EcochG was placed on the upper wall of the deepest part of the ear canal near the eardrum, and extratympanic ECochG was recorded bilaterally by averaging 1,000 sweeps after alternating click stimuli (rate: 15/sec) with levels of 100 dB nHL. This test was considered positive at an -SP/AP of >0.40 at our hospital. For further analysis reasons, cases where EcochG traces were unobtainable were classed as negative.®, Bayer AG) 4 h before the MRI scan (IV-Gd), as well as intratympanic administration of 8-fold-diluted Gd (IT-Gd) in both ears or the ear undergoing surgery for the follow-up Gd-MRI, if patients were reluctant to receive intratympanic injections in their healthy ear, 24 h prior to the MRI scan. All scans were performed on a 3T MRI scanner  using a 12-channel head coil. Three-dimensional real inversion recovery  sequence MRI images were collected, as previously described [MRI was performed using a single-dose (0.2 ml/kg) intravenous administration of gadopentetate dimeglumine . The reduction of vestibular hydrops was defined as an EVVR decrease of >10 points (%). No change in the vestibular hydrops was defined as an EVVR change of ≤10 points (%). The complete reversal of vestibular hydrops was defined as an EVVR of <30 points (%).The surgical procedure used for EDB was similar to that described previously . The tecFor statistical analysis, we used paired Student’s t-tests for two-group comparisons and Pearson correlations for evaluating the relationship. p values lower than .05 were considered significant. All data were statistically treated with SPSS version 26.0 .According to MRI scans, the grade of the cochlear EH in 22 patients was subjectively rated as follows: Mild EH (I) in 13 ears and significant EH (II) in 9 ears prior to surgery, and no hydrops (N) in 2 ears, mild EH (I) in 11 ears and significant EH (II) in 9 ears following surgery. A total of 4 patients showed downgrading of cochlear hydrops and 1 patient showed upgrading .Based on the EVVR results, the vestibular EH in 22 patients was classified as follows: Mild EH (I) in 7 ears and significant EH (II) in 15 ears prior to surgery, and no hydrops (N) in 4 ears, mild EH (I) in 7 ears and significant EH (II) in 11 ears following surgery. A total of 6 patients showed downgrading of vestibular hydrops and 2 patients showed upgrading . The aveFigs As shown in However, the difference value of the -SP/AP ratio had a significant positive correlation with the difference value of the EVVR   and gradIn vivo visualization of hydrops with Gd-MRI is no longer limited to only showing evidence of the hydrops for the diagnosis of MD. Changes in EH may be used to objectively evaluate the effects of various treatments for patients with MD ,20. HighAlthough the pathology of MD is well established to be EH. However, the mechanism underlying deafness of MD or idiopathic endolymphatic hydrops is still unknown. The most popular assumption is that the low-tone hearing loss is caused by the displacement of the basilar membrane (BM) from its resting position and the uncoupling of the outer hair cell stereocilia from the tectorial membrane due to hydrops –31; thisBased on a mechanical theory about hearing loss in MD, the low-tone sensorineural hearing loss in MD is caused by the mechanical impact of high endolymphatic pressure , which dRecently, Ito et al reported a positive correlation between the control of vertigo and the decrease in the volume of EH, and no correlation between changes in the hearing function and the volume of EH following sac drainage surgery ,47. HigaThe limitation of the present study is the small number of patients. Although fluctuation in the composition and volume of the endolymph is considered to contribute to the fluctuating nature of the symptoms experienced by sufferers of Meniere’s. On average, MD stabilizes with no further vestibular attacks by about 8 years after the onset of symptoms; however, hearing loss typically becomes permanent as MD progresses . NeverthIn conclusion, in the present study, it was first reported that the dynamic change of EH visualized by Gd-MRI was likely to modulate hearing function in 22 MD patients following EDB, highlighting the possibility that the downgrading of cochlear and/or vestibular hydrops could downregulate the hearing threshold and -SP/AP ratio, whereas the upgrading of cochlear and/or vestibular hydrops tended to upregulate the hearing threshold and -SP/AP ratio, suggesting that the hearing loss in MD is likely to be caused by hydrops and this disordered hearing function can be modulated by changes in the hydrops.S1 File(DOCX)Click here for additional data file.


Several factors may play a role in the ability of patients with Alzheimer’s disease to perform activities of daily living (ADL). The aim of this study was to examine the impact of different aspects of physical performance and cognitive functions on ADL in patients suffering from mild-to-moderate Alzheimer’s disease.We conducted secondary analyses on cross-sectional baseline data from the randomized controlled multicentre study “Preserving quality of life, physical health and functional ability in Alzheimer’s Disease: The effect of physical exercise” (ADEX). In total, 185 AD patients (76 women and 109 men), with a mean age on 70,4 years, were included. Data from physical performance tests , Sit to Stand test (STS)) and cognitive tests , Symbol Digit Modalities Test (SDMT), Stroop Color and Word test (Stroop)) were used. Their associations with ADL, measured on the ADCS-ADL scale was assessed in multivariable regression analyses.r = 0.33, MMSE: r = 0.42) and instrumental ADL , but not with basic ADL. Adjusting for age and sex, the associations between SDMT and MMSE to total ADL and instrumental ADL persisted. No significant associations were found between Astrand, TUG, STS or Stroop and total ADL, basic ADL or instrumental ADL.SDMT and MMSE had significant, moderate correlations with total ADL (SDMT: Total ADL and instrumental ADL are associated with cognitive functions, including executive function. No significant association between examined physical performance parameters and ADL functions was observed, and consequently does not support an impact of physical function on ADL functions in patients with mild-to-moderate Alzheimer’s disease and relatively well-preserved physical function. Strategies aimed to improve cognition may be better suited to improve ADL function in patients with mild-to-moderate Alzheimer’s disease.NCT01681602. Registered 10 September 2012, retrospectively registered. Alzheimer’s disease (AD) is a neurodegenerative disease with early and prominent memory impairment, and decline in the ability to independently carry out activities of daily living (ADL) –3. ADL fSeveral patient-related factors such as cognitive function, neuropsychiatric symptoms and disease duration may have deleterious impact on ADL functions in AD –8, patieWith regards to the relationship between cognition and ADL , 2, 18, Other interventions have also shown to be able to improve ADL in patients with MCI and dementia. For example, a combination of cognitive-physical training has shown beneficial in both patients with dementia and MCI . The appPreservation of executive function, rather than memory, has been linked to slower decline in ADL in AD patients  and furtA better understanding of the individual contributions of different cognitive functions and the impact of physical performance would potentially enable targeted interventions aimed at improving ADL functions. As stated above many studies have investigated the relationship between physical performance, cognition and ADL, in isolation. However, how all three variables are related still remains unclear. An exception is a rather recent cross-sectional study in which it was reported that the relationship between aerobic fitness and ADL function in AD patients was mediated by global cognition .The objectives of the present study were to examine the possible association between physical performance, cognition, and ADL functions. We hypothesised that physical performance and cognition, respectively, were associated with ADL functions. Further we hypothesised that IADL is more cognitively taxing than BADL, and therefore less dependent of physical function and the converse regarding IADL. Further, we wished to assess the impact of physical performance and cognitive functions on ADL, and to examine the possible mediating role of cognitive function with regards to the relationship between physical performance and ADL function.disease: The effect of physical exercise study). The ADEX study was a multicentre single-blinded study, conducted between January 2012 and June 2014, investigating the impact of a 16-week aerobic exercise intervention on symptoms of AD [This is a secondary analysis of cross-sectional data from a previously conducted randomized controlled trial using baseline data from patients enrolled in the ADEX study , McKhann et al., 1984. All participants were diagnosed by a physician who was a specialist in either geriatric medicine, psychiatry or neurology. As a part of the diagnostic process, all subjects underwent as a minimum formal cognitive testing, ADL assessment, blood samples and structural brain scans (CT or MRI). Patients were recruited from and followed in 8 memory clinics in Denmark. Details on objectives and design including inclusion and exclusion criteria have previously been described in detail [A total of 200 community-dwelling patients with mild-to-moderate AD were included in the ADEX study, all meeting criteria for probable AD according to the Alzheimer’s criteria by the n detail . For theCognitive assessments were performed in a quiet environment without interruptions. The Mini-Mental State Examination (MMSE) was used to assess global cognitive function . Process2max (ml/kg/min)) based on workload and average heart rate during the last minute of the test, corrected for age and body weight [To assess aerobic fitness, the 6-min Astrand Cycle Ergometer test  was used. The estimated maximum oxygen uptake  scale . The ADCp < 0.05.Unadjusted associations between total ADL, BADL and IADL, and measures of physical performance  and cognitive function  were assessed by Pearson’s correlation coefficient. Using multivariable linear regression models the beforementioned relationships were examined (adjusted for age and sex) by the corresponding β-coefficients where the measures of physical performance and cognitive function where entered individually. In order to assess a potential mediating effect of cognition on the association between physical performance and ADL, the regressions for the measures of physical performance were additionally adjusted for the measures of cognitive functions . A small number of patients had missing data for Stroop, SDMT and Astrand, which were handled with listwise deletion. Statistical analyses were performed using SAS v9.4 and R v3.5.1. Statistical significance was accepted at Baseline characteristics for all 185 participants included in the study are presented in Table r = 0.33, P < 0.05) and IADL , but not with BADL. Moreover, MMSE was positively and significantly correlated with total ADL  and IADL , but not with BADL. None of the physical parameters nor Stroop were correlated with ADL.Pearson correlation coefficients (PCC) are presented in Table p = 0.0019) and IADL . Similarly, MMSE was associated with total ADL  and IADL . Concerning BADL, no significant associations with physical nor cognitive parameters were found. No significant associations were found between Astrand, TUG, STS or Stroop and ADL, BADL or IADL.The corresponding β-coefficients from multivariable linear regression models, adjusted for age and sex, are presented in Table Since no association between physical performance parameters and ADL functions were found, a mediating effect of cognition was not excepted. However, the regressions for physical performance parameters adjusted for the measures of cognitive functions are presented in Table In the present study, we add to the limited research on how physical performance and cognitive subdomains impact on ADL in patients with mild-to-moderate AD.We found that cognition and ADL were significantly and positively correlated. Specifically, with regards to cognitive domains we found that processing speed and attention correlated significantly with IADL but not BADL. The findings support our hypothesis that cognitive function is of importance for ADL functions and especially for more complex ADL functions captured by IADLs . MeasureIn general, a significant moderate correlation was observed between general cognitive function and the instrumental subscale of ADL, while no correlation was found to basic ADL, suggesting that IADL is more reliant on cognition. When examining the relationship in more detail, processing speed and attention were found to be associated with total ADL and IADL, but not BADL, mirroring the findings for general cognitive function. As peviously stated, the findings may indicate that those complex activities comprising IADL are most reliant on cognition. Supporting this finding, IADL has been found to be more strongly associated with executive function than BADL in patients with mild-to-moderate AD , 62. MorAnother important finding is the lack of significant association between ADL functions and physical performance i.e. aerobic fitness, measure of mobility, and strength and endurance of lower extremities, which is not consistent with the findings of a recent meta-analysis establishing an association between TUG and STS and worsening in ADL in populations of older adults . Our resOur findings add to the relatively limited research on the role of aerobic and physical performance on ADL in patients with AD. The findings provide important input for future design of interventions aimed at improving ADL. Our findings of an association between cognitive function, and specifically a measure of procession speed and mental speed, indicate that interventions in patients with mild to moderate AD focusing on improving these cognitive functions may be more beneficial for improving ADL. The fact that we did not find an association between ADL and aerobic performance would seem to indicate that in mild to moderate AD, interventions to improve ADL functions may be less effective, if targeting aerobic function. It may be speculated, based on our findings that in more advanced patients, interventions improving aerobic and physical performance may improve ADL through an effect on BADL. In order to further investigate the impact of physical performance on ADL, especially considering recent research regarding effects of cognitive-motor interventions on ADL , 39, it We acknowledge that this study has some limitations. The study is a cross-sectional study excluding an analysis of a causal relationship. Analyses of intervention is of interest in order to investigate how physical performance, cognition and ADL benefit from physical exercise , 45. HowIn conclusion, this study found a significant and positive association between cognition, including processing speed and attention and total ADL and IADL functions in patients with mild-to-moderate AD. No association was found between cognition and BADL, suggesting that BADL is less cognitively demanding and less reliant on cognitive function. Although our findings do not support physical function as being important for performance of ADL functions in patients with mild-to-moderate AD and with relatively well-preserved physical function, it cannot be interpreted as if such an association does not exist in persons with more severe dementia or more severe ADL impairment. Our study could imply that interventions targeting cognition may also have a positive effect on instrumental ADL in patients with mild-to-moderate AD.


The purpose of this study is to investigate Black-White differences in hearing problems among older adults living in the United States. Secondary data analyses were conducted using the 2017 American Community Survey (ACS) with a replication analysis in the 2016 ACS. The ACS is an annual nationally representative survey of Americans living in community settings and institutions. The sample size of older Americans (age 65+) in 2017 was 467,789 Non-Hispanic Whites (NHW) and 45,105 Non-Hispanic Blacks (NHB). In the 2016 ACS, there were 459,692 NHW and 45,990 NHB respondents aged 65+. Measures of hearing problems, age, race/ethnicity, education level and household income were based on self-report. Data were weighted to adjust for non-response and differential selection probabilities. The prevalence of hearing problems was markedly higher among older NHW (15.4% in both waves) in comparison to NHB . In the 2017 ACS, the age-sex adjusted odds of hearing loss were 69% higher for NHW compared to NHB, which increased to 91% higher odds when household income and education level were taken into account . Further analyses by 10 year age cohorts indicated comparable findings (fully adjusted ORs range from 1.89 to 1.98). Findings from the 2016 ACS were very similar . NHW have a much higher prevalence and almost double the odds of hearing loss compared NHB. Future research should investigate if melanin plays an otoprotective role through enhancing the antioxidant capability of cochlea.


Brain-derived nerve growth factor (BDNF) plays an important role in cochlear development so it is plausible that it could restore hearing loss if delivered directly into the cochlea. We wished to confirm our previous report that a single intracochlear injection of brain-derived nerve growth factor (BDNF) was beneficial for hearing in guinea pigs. We wished to assess the reproducibility of our results and assess possible improved methods with a view to developing a clinical treatment for sensorineural hearing loss.CDDP was used to create partial hearing loss in 25 guinea pigs. After 30 days the animals underwent ABR testing and unilateral BDNF injection through the round window in one ear and saline injection into the other ear. After allowing possible effects to stabilize, thirty days later, ABR threshold testing was repeated to assess change in threshold.Final ABR thresholds were 60–70 dB and were about 11 dB better in the ears treated with BDNF.Our original finding that Intracochlear BDNF can improve hearing in guinea pigs was confirmed, but the improvement demonstrated by the methods in this paper is too small for clinical application. Brain-derived nerve growth factor (BDNF) is important for hearing. BDNF is known to play a significant role in cochlear development , but itsGene therapy is one consideration. Ongoing research involving promising, molecular biological approaches to hearing loss may provide treatment someday but knowledge of gene control and methods of administration of genetic modifiers is lacking. Could simple injection of BDNF to the cochlea achieve positive results? The answer to this question is unclear but it seems important to address a possible simple solution for hearing loss. After all, the research findings that relate to BDNF and hearing are based on the expression of BDNF, not control of its activity. The possibility that direct injection of BDNF into the cochlea deserves study as a possible clinical intervention for hearing loss.There is uncertainty regarding BDNF research. Much of the BDNF research focuses on hair cell survival or in vitro findings, not actual hearing. BDNF may result in morphologic improvements but not hearing in pigeons . For cliWe considered some improvements on our initial design. In our previous study  we used Our lab has previously explored the possibility that simply injecting the BDNF into the cochlea is associated with better hearing. If such a simple treatment were successful, the application could help millions. The purpose of this study was to assess the reproducibility and possibly confirm our previous findings. In our previous study we reported that a single intracochlear injection of BDNF resulted in auditory brainstem response (ABR) thresholds across several frequencies that were from 2 to 27 dB better than saline-injected ears . We foun2+) activity. BDNF acts on the TrkB receptor. Mice that lack either BDNF or the TrkB receptor have no hearing [Neurotrophins such as BDNF act by signaling through tyrosine kinases, reducing the formation of oxygen free radicals and up-regulation of apoptotic genes and mitigation of intracellular calcium .ABR thresholds were entered into a database and Analysis of Variance for Repeated Measures was performed using SPSS v24 software and a significance level of p = 0.254) for ABR thresholds by day. ANOVA results indicates that differences across frequencies were not statistically significant (p = 0.088) so all frequencies were included together in further analysis.Figure p = 0.034). Across all frequencies final, rounded ABR thresholds in BDNF-treated ears on day 60 were 60+/− 23 dB (mean +/− 95%CI) in BDNF-treated ears and 71 +/− 20 dB in the saline-treated ears for a mean difference of 11 dB.Threshold differences were not significantly different between the treated and non-treated ears at days 0 or 30, but at day 60  the traces diverged significantly. Figure CDDP induced a mean increase in ABR threshold on both sides of 9 dB across all frequencies. Intracochlear injection increased the thresholds further by an average of 16 dB .Several findings deserve discussion.First, our results indicate that intracochlear BDNF is associated with lower thresholds of about 11 dB. The results of this study are consistent with some of our previous work, indicating a small difference in threshold in ears treated with BDNF . We wereSecond, the 11 dB improvement in hearing did not overcome the loss induced by the intracochlear injection, highlighting the importance of controls in research. The effect of the injection procedure is evident as the increase in threshold between 30 and 60 days in both the BDNF- and the saline-treated ears. This finding would seem to be important for any agent injected into the cochlea for the purpose of improving hearing.Third, it is odd that the ABR thresholds did not differ by frequency as one might expect from the human literature. In this project as well as our previous work has also found that, in rodents, CDDP affects all thresholds across the range of frequencies tested. Differences between human and rodent hearing should be considered if this research is to be applied to humans. For example, rodents have excellent hearing in higher frequency ranges than humans, perhaps up to 40 kHz. We have sampled some of both ranges. Anesthetic time limits the feasibility of testing at very many frequencies.Fourth, the CDDP did not create as much hearing loss as it has in past work in our lab . TypicalAlthough it CDDP is commonly thought to affect higher frequencies more than lower ones this was not reflected in our data. ABR thresholds at day 30 were similar across frequencies. We have found this in other studies as well .Intracochlear BDNF injection can benefit hearing in guinea pigs but does not overcome the hearing loss induced by the injection itself. The methods as used in this paper are inadequate for clinical application.